Obonyo, 2, Mark Odhiambo MSC Spplied Epidemiology, 2018

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SERO-PREVALENCE AND FACTORS ASSOCIATED

WITH BRUCELLOSIS IN GOATS AND SHEEP AND


ASSESSMENT OF PASTORALISTS, KNOWLEDGE
ATTITUDE AND PRACTICES TOWARDS BRUCELLOSIS
IN GARISSA COUNTY

MARK ODHIAMBO OBONYO

MASTER OF SCIENCE

(Applied Epidemiology)

JOMO KENYATTA UNIVERSITY OF

AGRICULTURE AND TECHNOLOGY.

2018
Sero-prevalence and Factors Associated with Brucellosis in Goats and
Sheep and Assessment of Pastoralists, Knowledge Attitude and
Practices towards Brucellosis in Garissa County

Mark Odhiambo Obonyo

A thesis submitted in partial fulfilment for the Degree of Master of


Science in Applied Epidemiology in the Jomo Kenyatta University of
Agriculture and Technology.

2018
DECLARATION

This thesis is my original work and has not been presented for a degree in any other
university

Signature: .......................................................... Date: ...............................................

Mark Odhiambo Obonyo

This proposal has been submitted for examination with our approval as the university
supervisors

Signature: .......................................................... Date: ...............................................

Prof. Gideon Kikuvi, PhD

JKUAT, Kenya

Signature: .......................................................... Date: ...............................................

Dr. Kariuki Njenga, PhD

KEMRI, Kenya

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DEDICATION

This work is dedicated to my wife Norah Mbithe, sons Israel and Ethan, my mother
Dorcas, sisters Beatrice and Mercy and my brother Kennedy for their support, prayers,
encouragement, and motivation to see this thesis become a reality.

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ACKNOWLEDGEMENTS

First I would like to thank the almighty God for giving me strength and energy to
accomplish this work. Secondly, I would wish to express my gratitude to my supervisors
Prof. Gideon Kikuvi and Dr. Kariuki Njenga for their guidance and assistance during the
research work.

I would like to also extend my sincere gratitude to Kenya Field Epidemiology and
Laboratory Program (K-FELTP) for financial support to carry out the study. I would
also wish to extend my sincere gratitude to the K-FELTP faculty members for their
untiring support to ensure that I accomplish this work.

I also wish to acknowledge all the staff of the department of veterinary services and
livestock production of Garissa County and staff at the Regional veterinary
investigations Laboratory (RVIL) in Garissa and Central Veterinary Laboratory (CVL)
in Kabete Nairobi for their testing and supply of reagents for sample testing.

I would also like to cooperation and efforts towards the success of this study especially
in connection to sample acknowledge the study participants and the community leaders
for support provided during the study.

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TABLE OF CONTENTS

DECLARATION .............................................................................................................II

DEDICATION ............................................................................................................... III

ACKNOWLEDGEMENTS .......................................................................................... IV

TABLE OF CONTENTS ................................................................................................ V

LIST OF TABLES .......................................................................................................... X

LIST OF FIGURES ..................................................................................................... XII

LIST OF APPENDICES ........................................................................................... XIII

LIST OF EQUATIONS ............................................................................................. XIV

LIST OF ACRONYMS AND ABBREVIATIONS ................................................... XV

ABSTRACT ............................................................................................................... XVII

CHAPTER ONE ............................................................................................................ 19

INTRODUCTION .......................................................................................................... 19

1.1 Background information......................................................................................... 19

1.2 Statement of the Problem ....................................................................................... 21

1.3 Justification of the Study ........................................................................................ 23

1.4 Research questions ................................................................................................. 24

1.5 Objectives ............................................................................................................... 25

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1.5.1 Broad objective ................................................................................................ 25

1.5.2 Specific objectives ........................................................................................... 26

CHAPTER TWO ........................................................................................................... 27

LITERATURE REVIEW.............................................................................................. 27

2.1 Etiology of Brucellosis ........................................................................................... 27

2.2 Morphology of Brucella spps ................................................................................. 27

2.3 Taxonomy of Brucella spps .................................................................................... 28

2.4 History and importance of Brucella melitensis ...................................................... 30

2.5 Brucella melitensis infection in sheep and goats .................................................... 30

2.6 Pathological lesions in animals .............................................................................. 31

2.7 Brucella melitensis infection in humans ................................................................ 32

2.8 Diagnosis of Brucellosis in sheep and goats .......................................................... 34

2.8.1 Clinical signs of Brucellosis in sheep and goats .............................................. 34

2.8.2 Staining and microscopy of Brucella organisms ............................................. 35

2.8.3 Serology of Brucellosis .................................................................................... 35

2.8.4 Culture of Brucella organisms ......................................................................... 36

2.8.5 Biochemical tests for Brucella organisms ....................................................... 38

2.8.6 Susceptibility to phages by Brucella organisms .............................................. 38

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2.8.7 Other methods for detection of Brucella organisms ........................................ 39

2.8.8 Sero-prevalence of Brucellosis in small ruminants ......................................... 39

2.8.9 Risk factors for small ruminant Brucellosis .................................................... 41

2.8.10 Knowledge attitude and practices towards Brucellosis ................................. 43

2.8.11 Conceptual framework ................................................................................... 44

CHAPTER THREE ....................................................................................................... 45

MATERIALS AND METHODS .................................................................................. 45

3.1 Study area ........................................................................................................... 45

3.2 Study design ........................................................................................................... 46

3.3 Study Population .................................................................................................... 46

3.4 Sample size determination for sheep and Goats ................................................. 47

3.5 Sampling design for sheep and Goats .................................................................... 50

3.6 Blood sample collection ......................................................................................... 52

3.7 Serological testing .................................................................................................. 53

3.8 Data Management ................................................................................................... 56

3.8.1 Questionnaire survey ....................................................................................... 56

3.8.2 Data analysis .................................................................................................... 57

3.9 Ethical Approvals and Considerations ................................................................... 57

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CHAPTER FOUR .......................................................................................................... 58

RESULTS ....................................................................................................................... 58

4.1 Sero-prevalence of Brucellosis at Individual and Herd Level ............................... 58

4.1.1 Herd characteristics.......................................................................................... 58

4.1.2 Brucella Sero-prevalence at individual animal level by animal species .......... 58

4.1.3 Brucella Sero-prevalence at individual animal level by sub-location sampled


.................................................................................................................................. 59

4.1.4 Adjusted Individual animal level Sero-Prevalence .......................................... 60

4.1.5 Brucella Sero-prevalence at Herd Level .......................................................... 61

4.2 Factors Associated with Brucellosis Herd Sero-positivity ..................................... 62

4.2.1 Bivariate Analysis for Herd level Factors Associated with Brucellosis in sheep
and goats ................................................................................................................... 62

4.2.2 Multivariable analysis to determine independent factors associated with


Brucella herd sero-positivity..................................................................................... 65

4.3 Determination of Knowledge, Attitude and Practices towards Brucellosis among


pastoralists in Garissa County ...................................................................................... 66

4.3.1 Demographic characteristics of study participants .......................................... 66

4.3.2 Knowledge of pastoralists toward Brucellosis................................................. 67

4.3.3 Respondents’ attitude towards Brucellosis ...................................................... 72

4.3.4 Respondents’ practices towards Brucellosis .................................................... 74

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4.3.5 Respondents’ requirement of more Information on Brucellosis ...................... 75

CHAPTER FIVE ............................................................................................................ 76

DISCUSSIONS, CONCLUSSION AND RECOMMEDATIONS ............................. 76

5.1 Discussion .............................................................................................................. 76

5.1.1 Highlight of major findings of the study ......................................................... 76

5.1.2 Sero-prevalence of Brucellosis in sheep and goats .......................................... 77

5.1.3 Herd-level factors associated with Brucellosis sero-positivity in sheep and


goats .......................................................................................................................... 78

5.1.4 Knowledge Attitude and Practices towards Brucellosis .................................. 81

5.2 Limitations of the study .......................................................................................... 85

5.3 Conclusions and Recommendations ....................................................................... 86

5.3.1 Conclusions...................................................................................................... 86

5.3.2 Recommendations ............................................................................................ 86

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LIST OF TABLES

Table 3.1: Sheep and goats population, Garissa County ................................................. 47

Table 3.2: List of sub-locations in Garissa and Balambala Township and the Sub-
locations randomly selected for Sampling highlighted in yellow ................... 50

Table 3.3: Distribution of number herds sampled per sub-location ................................ 52

Table 4.1: Distribution of individual animal prevalence of Brucella antibodies in sheep


and goats by sub-location, Garissa 2013 (n=2400) ......................................... 60

Table 4.2: Comparison of Number of Herds with Positive Reactors with Number of
animals testing Positive (n=62) ....................................................................... 61

Table 4.3: Comparison of factors associated with Brucella Sero-positivity among


positive and negative herds, Garissa 2013....................................................... 64

Table 4.4: Multivariable Logistic Regression Analysis of the Variables associated with
Herd-level Sero-positivity for Brucella spps in Sheep and Goats, Garissa
County, 2013 ................................................................................................... 65

Table 4.5: Distribution of Socio-demographic characteristics of study participants,


Garissa County-2013 (n=120) ......................................................................... 66

Table 4.6: Distribution of responses of participants’ on animal species affected by


Brucellosis and signs and symptoms of Brucellosis in animals (n=95) .......... 68

Table 4.7: Responses of respondents’ on signs and symptoms of Brucellosis in humans,


Garissa County, 2013 (n=95) .......................................................................... 72

Table 4.8: Distribution of respondents’ responses on attitude and perceptions towards


Brucellosis, Garissa County, 2013 (n=95) ...................................................... 73

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Table 4.9: Distribution of respondents’ responses on practices towards Brucellosis,
Garissa County, 2013 (n=95) .......................................................................... 74

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LIST OF FIGURES

Figure 2.1: Conceptual Framework ................................................................................ 44

Figure 3.1: Map of Kenya showing Garissa County highlighted in yellow color and
study area highlighted in red color ............................................................. 46

Figure 4.1: Distribution of Prevalence of Brucella antibodies by test type and species,
Garissa County, 2013 (n=2,400) ................................................................. 59

Figure 4.2: Distribution of study participants’ responses on causes of Brucellosis,


Garissa County, 2013 (n=95) ...................................................................... 67

Figure 4.3: Distribution of respondents’ responses on mode of transmission of


Brucellosis to humans, Garissa County, 2013 (n=27) ................................ 70

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LIST OF APPENDICES

Appendix 1: Questionnaire in English .......................................................................... 108

Appendix 2: Consent form in English .......................................................................... 126

Appendix 3: Translated Questionnaire in Somali Language ........................................ 130

Appendix 4: Translated consent form in Somali Language .......................................... 147

Appendix 5: Manuscript on Sero-prevalence and herd level factors associated with


Brucellosis in sheep and goats in Garissa County, 2013. ....................... 152

Appendix 6: Manuscript on Brucellosis KAP survey ................................................... 178

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LIST OF EQUATIONS

Equation 1: Formula for Sample Size Calculations....................................................... 47

Equation 2: Equation for determination of true sero-prevalence .................................. 56

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LIST OF ACRONYMS AND ABBREVIATIONS

CDC Centers for Disease Control and Prevention

CFSPH Center for Food security and Public Health

CFT Complement Fixation Test

CI Confidence Interval

CO2 Carbon dioxide

DNA Deoxyribonucleic Acid

ELISA Enzyme Linked Immuno-Sorbent Assay

FAO Food and Agriculture Organization

H2 S Hydrogen sulphide

ICSP International Committee on Systematics of Prokaryotes

OIE World Organization for Animal Health

OR Odds Ratio

PCR Polymerase Chain Reaction

RBPT Rose Bengal Plate Test

RVIL Regional Veterinary Investigations Laboratory

SLPS Smooth Lipopolysaccharide

SPP Species

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WHO World Health Organization

ZDU Zoonotic Disease Unit, Kenya

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ABSTRACT

Brucellosis, a zoonosis of major public health importance, is endemic in livestock in


Kenya. Unfortunately, reliable data on Brucellosis in Kenya is scarce and data on sero-
prevalence and risk factors associated with small ruminant Brucellosis in Garissa County
is unknown. This was a cross-sectional study carried out to determine the sero-
prevalence of Brucellosis and identify herd-level factors associated sero-prevalence in
small ruminants in Garissa County of North Eastern Kenya. The study also assessed the
pastoralists’ knowledge, attitude and practices towards Brucellosis. A total of 2,400 sera
from 120 flocks were collected from sheep and goats which were randomly selected
using a multi-stage sampling technique and data on potential herd-level factors were
collected from the pastoralists’ ≥15 years using a pre-tested structured questionnaire.
The sera were analyzed using Rose Bengal Plate Test (RBPT) and sero-positive reactors
confirmed by Complement Fixation Test (CFT) using serial interpretation. A sample
was considered to be positive when both tests results were positive and a herd was
considered positive when a single animal within the herd tested positive on both tests.
Multivariable logistic regression was used to investigate for independent factors
associated with flock Brucellosis sero-positivity in small ruminants. The overall sero-
prevalence of Brucellosis at individual animal-level was 20.0% (95% CI: 18.2% to
22.0%); in goats 24.3% (95% CI: 21.8% to 27.1%) and sheep 12.5% (95% CI: 10.2% to
15.2). Overall true herd-level sero-prevalence was 65.8% (95% CI: 54.3% to 77.2%).
Seeking veterinary services [aOR=0.30 (95% CI: 0.12 to 0.76], introduction of new
animals into the flock [aOR=8.0 (95% CI: 3.09 to 20.70] and experiencing abortions in
the flock [aOR=3.43 (95% CI: 1.33 to 8.88] were independently associated with
Brucellosis herd sero-prevalence in small ruminants. A total of 120 pastoralists were
interviewed of which 95 (79%) had heard of Brucellosis and 17(18%) mentioned
bacteria/germ as cause. Forty-four (46%) would do nothing if they had aborting animal
in their herd, 91 (96%) consumed raw milk in the past year and 72 (76%) assisted an
animal during parturition process and none used glove. The study highlights
considerable high sero-prevalence of Brucellosis and factors that contributes for its

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transmission in small ruminants in Garissa County. This poses potential public health
threat associated with zoonotic transmission. The study also highlights that though the
community has some knowledge on Brucellosis, attitudes and practices are poor.
Enhanced public health education by the County government is recommended for
effective prevention and control of Brucellosis in animals and humans in the area. Need
to conduct animal-human linked study in the area for holistic understanding of
epidemiology of the disease in the area.

xviii
CHAPTER ONE

INTRODUCTION

1.1 Background information

Brucellosis is one of the world’s most widespread zoonotic diseases caused by species of
the genus Brucella (Moreno et al., 2002). Brucellosis in sheep and goats occurs
worldwide. The Mediterranean countries of Europe, northern and eastern Africa, Near
East countries, India, Central Asia, Mexico and Central and South America are
especially affected. While B. melitensis has never been detected in some countries, there
are no reliable reports that it has ever been eradicated from small ruminants (Omer et al.,
2000; Radostits et al., 2007; Taleski et al., 2002).

Brucellosis is a herd or flock problem. It is spread within the herd primarily by ingestion
of contaminated material. Venereal infections can also occur, but this is mainly seen
with B. suis infections. Congenital (in utero) or perinatal infections may also occur, with
the ensuing development of latent infections. Spread between herds usually occurs by
the introduction of asymptomatic chronically-infected animals. Brucellosis is a disease
of the sexually mature animals with predilection for placentas, fetal fluids and testes of
male animals. The disease in animals is transmitted by direct or indirect contact with
infected or contaminated materials. Both wild and domestic animals are susceptible to
infection with Brucella and may serve as carriers for other domestic animals. Initial
infection in the reservoir species is often followed by abortion and subsequent delayed
or permanent infertility. Infected animals shed the organisms in uterine discharges
following abortion and subsequent parturition, and also in the colostrum and milk
(Radostits et al., 2007). Risk factors that have been identified to enhance the
transmission of Brucellosis between and within herds include history of abortions within
a herd, mixing of different animals during grazing and in watering places, introduction
of new non-quarantined animals whose Brucellosis status is not known into the herd as

19
replacement stock or for breeding purposes and mixing with wild animals in watering
and grazing places (Coelho et al., 2008; Bamaiyi et al., 2014; Sharifi et al., 2014; Muma
et al., 2007a; Al-Majali et al., 2007).

Transmission of infection to humans occurs through breaks in the skin, following direct
contact with tissues, blood, urine, vaginal discharges, aborted fetuses or placentas. Food-
borne infection occurs following ingestion of raw milk and other dairy products, but
rarely from eating raw meat from infected animals. Occupational airborne infection in
laboratories and abattoirs has also been documented. Accidental inoculation of live
vaccines (such as B. abortus Strain 19 and B. melitensis Rev.1) can also occur, resulting
in human infections. There are also case reports of venereal and congenital infection in
humans (Al-Majali, 2005; Queipo-Ortuno et al., 1997).

Human infections are characterized by a variable incubation period (from several days
up to several months), and clinical signs and symptoms of continued, intermittent or
irregular fever of variable duration, with headaches, weakness, profuse sweating, chills,
depression and weight loss. Localized suppurative infections may also occur. The course
of the disease can be variable especially in persons either not or inadequately treated
(Al-Majali, 2005).

Diagnosis of clinical Brucellosis in humans and animals is initially made by use of


appropriate serological or other immunological tests, and confirmed by bacteriological
isolation and identification of the agent (OIE, 2009a).

Methods of prevention include health education to reduce occupational and food-borne


risks, including pasteurization of all dairy products. However, education campaigns have
never resulted in fully eliminating the risks of infection, and the ultimate prevention of
human infection remains elimination of the infection among animals. This can be
achieved by a combination of test and isolation/slaughter of positive animals,
vaccination of susceptible animals and control of animal movements (Corbel, 1997).

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Various countries all over the world have implemented different control policies using
combinations of these control measures, with reported success in some countries and
failure in others (Cutler et al., 2005; Blasco, 1997).

1.2 Statement of the Problem

Brucellosis infection in sheep and goats causes heavy economic losses in animal
production resulting from clinical disease, abortion, neonatal losses, reduced fertility,
decreased milk production, slaughtering of the infected animals, cost of veterinary care,
and replacement animals. In addition, the disease is an impediment to free animal
movement and it is also a limiting factor for international trade of animals and their
products (Radostits et al., 2007). The disease in humans is of great public health
implication due to losses from high medical expenses, productivity losses and high costs
of management of sequelae (Al-Majali, 2005).

In humans, Brucellosis, especially caused by Brucella melitensis, remains one of the


most common zoonotic diseases worldwide with more than 500,000 human cases
reported annually. The true incidence of human Brucellosis worldwide is unknown and
the estimated burden of the disease varies widely, from <0.03 to >160 per 100,000
population (Pappas et al., 2006; Taleski et al., 2002). The bacterial pathogen is classified
by the US Centers for Disease Control and Prevention as a category (B) pathogen that
has potential for development as a biological weapon with a potential of aerosol
transmission (Seleem et al., 2009).

In Kenya, there is no systematic control program for Brucellosis in livestock. Control


program is based on individual farmer initiative and is based on testing ruminants more
than 6 months of age with slaughter of serologically positives, and voluntary vaccination
of calves using Brucella abortus S19 vaccine and Brucella melitensis Rev 1 vaccine for
lambs and kids (McDermott and Arimi, 2002).

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Brucellosis remains endemic in livestock and humans in several parts of Kenya and a
hospital based study done in Garissa County among febrile patients, found a Brucellosis
sero-prevalence of 31.8% and the true prevalence of 15.4% by PCR (Kiambi, 2014).
Given that infected animals are the source of human infection, the increasing incidence
of human Brucellosis probably reflects a similar trend in domestic animals. Animal
Brucellosis has been reported every year particularly from the arid and semi-arid
pastoral areas of the country (Kenya DVS Annual reports, 1999-2010). A study of
Brucellosis in goats herds done in Kenya in 1976 in Garissa found a herd prevalence of
6.8% (McDermott and Arimi, 2002). In other parts of the world, a cross-sectional study
to determine sero-prevalence of Brucella melitensis and to identify risk factors
associated with goat sero-positivity in Michoacán, Mexico, blood samples were
collected from 5,114 animals from 79 herds. Sera were tested for antibodies against B.
melitensis using the Rose Bengal plate test (RBPT) and the complement-fixation test
(CFT). A total of 55 herds out of the 79 herds tested had at least one seropositive animal.
The animal-level true sero-prevalence was 9.8% (Solorio-Rivera et al., 2007). In a study
done in Spain using RBPT and CFT where 56 herds (35 ovine and 21 caprine) were
sampled. Sixteen (29%) flocks (3 caprine and 13 ovine) were Brucellosis sero-positive.
Overall, 0.7% of sheep and 0.1% of goats were sero-positive (Reviriego et al., 2000). In
Africa, a study done in goats in Eastern and Western Uganda using Brucellosis card test
(CT) and Tube Agglutination Test (TAT) in parallel, found an individual animal-level
sero-prevalence of 4% and a herd-level sero-prevalence of 13% with CT; and a 13%
animal-level sero-prevalence and 43% herd-level sero-prevalence with TAT
(Kabagambe et al., 2001). In Eriteria, a study using Rose Bengal Plate Test (RBPT)
detected an animal-level sero-prevalence of 14.3% and a herd-level sero-prevalence of
56. 3% (Omer et al., 2000).

In order for a Brucellosis control programme to be efficient, it is important to understand


community local knowledge, attitudes and practices relating to Brucellosis to improve
information delivery and initiate relevant control measures through disease education

22
among community participants (Smits, 2012). However in Kenya, there is scarcity of
information on local community KAP on Brucellosis. A KAP study of Brucellosis
conducted in Kenya among people with high level of contact with livestock found that
the disease awareness and knowledge of the transmission routes were poor (Kangethe et
al., 2007).

1.3 Justification of the Study

Livestock plays a crucial role in the livelihood of the majority of residents of Garissa
county who are predominantly pastoralists keeping mainly sheep, goats, cattle and
camels. The pastoral production system is characterized by pastoralists keeping
relatively large herds that graze freely in vast communal lands with watering points.
Livestock owners practice a free grazing system, using communal grazing grounds and
watering points where cattle and small ruminants graze separately. The majority of herd
owners are semi-sedentary with only a few still practicing nomadism. Animals are
usually kept in the animal shade (boma) at night and young stock of usually less than
two months of age share the house with humans. Older cattle are kept in a separate shade
with sheep and goats. Livestock owners also keep relatively large herds and flocks for
meat and milk for their families, as a source of savings, and to meet some cultural and
social values such as dowry, celebrations and gifts and for social prestige. It is has been
estimated that more than 95% of the household income is derived from livestock and
livestock products (Garissa, 2013). However this dependence on livestock makes people
vulnerable to zoonotic diseases. Some of the socio cultural practices such as
consumption of raw milk make residents of Garissa at greater risk of infection with
Brucellosis.

Although there are no published studies that incriminate Brucella spps as cause of
abortions in goats and sheep in Garissa, Brucellosis has been suspected on basis of
clinical grounds by local veterinary officers working in Garissa County. Unfortunately,

23
reliable data on Brucellosis in Kenya is scarce and data on sero-prevalence and risk
factors associated with sheep and goat Brucellosis in Garissa County is not known.

Therefore a better understanding of the sero-prevalence of Brucellosis and associated


herd-level risk factors for exposure across different geographical areas in Kenya is
important in generating evidence based information that may help towards formulating
control strategies especially in livestock and this will impact in reducing the incidence of
infection in both livestock and humans. Control of Brucellosis in Garissa County is
important in safeguarding the livelihoods of majority of people in Garissa County who
largely depends on livestock keeping, enhancing food security in the region and in long
term promoting good health and wellbeing of people living in Garissa which directly
linked to the sustainable development goal 3 (SDG3).

Improvement of knowledge, attitudes and practices among pastoralists could have a


significant impact on the reduction of many zoonotic infections including Brucellosis
(Kangethe et al., 2007).

1.4 Research questions

1. What is the individual animal-level and herd-level sero-prevalence of Brucellosis


in goats and sheep in Garissa County?
2. What are the herd-level factors associated with Brucellosis sero-prevalence in
Garissa County?
3. What are the farmer’s knowledge, attitude and practices regarding Brucellosis in
Garissa County?

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1.5 Objectives
1.5.1 Broad objective

To evaluate individual animal-level and herd-level sero-prevalence, determine herd-level


factors associated with Brucellosis in goats and sheep and assess farmer’s knowledge,
attitude and practices regarding Brucellosis in Garissa County.

25
1.5.2 Specific objectives

1. To determine individual animal-level and herd-level sero-prevalence of


Brucellosis in goats and sheep in Garissa County.
2. To determine herd-level factors associated with Brucellosis sero-prevalence in
goats and sheep in Garissa County.
3. To evaluate pastoralists knowledge, attitude and practices regarding Brucellosis
in Garissa County.

26
CHAPTER TWO

LITERATURE REVIEW

2.1 Etiology of Brucellosis

Ten Brucella spps are currently recognized, seven of them that affect terrestrial animals
are: B. abortus, B. melitensis, B. suis, B. ovis, B. canis, B. neotomae, and B. microti
(Scholz et al., 2008) and two that affect marine mammals are: B. ceti and B.
pinnipedialis (Foster et al., 2007) and B. inopinata isolated from a breast implant wound
of a woman with clinical signs of Brucellosis (Scholz et al., 2010). Recently, a novel
Brucella spps has been isolated from postpartum uteruses of pregnant baboons with
history of stillbirth (Schlabritz-Loutsevitch, 2009).The newly isolated Brucella spps has
not yet been classified nor included in the above mentioned list. The first three species
are called classical Brucella and within these species, seven biovars are recognized for
B. abortus, three for B. melitensis and five for B. suis. The remaining species have not
been differentiated into biovars.

Brucellosis in sheep and goats is mainly caused by Brucella melitensis, containing three
biovars (biovars 1, 2 and 3). All three biovars cause disease in small ruminants, but their
geographic distribution varies. Brucella abortus and Brucella suis infections also occur
occasionally in small ruminants, but clinical disease seems to be rare (CSFPH, 2009). B.
ovis infections occur in sheep in sub-Saharan Africa and have been associated with
epididymitis, orchitis and infertility in rams (McDermott and Arimi, 2002).

2.2 Morphology of Brucella spps

Brucella spps are gram-negative cocco-bacilli or short rods 0.6 to 1.5 μm long by 0.5 to
0.7 μm in width, arranged singly and less frequently in pairs or small groups. They are

27
non-spore-forming and non-capsulated. Although they are described as non-motile, they
carry all the genes except the chemotactic system, necessary to assemble a functional
flagellum (Fretin et al., 2005). Brucella usually does not show bipolar staining. They are
not truly acid-fast but resist discoloration by weak acids, thus stain red by the Stamp's
modification of Ziehl-Nielsen method, which is sometimes used for the microscopic
diagnosis of Brucellosis from smears of solid or liquid specimens. The morphology of
Brucella spps is fairly constant except in old cultures, where pleomorphic forms may be
evident (Godfroid et al., 2005; OIE, 2009a).

On suitable solid media Brucella colonies are visible after 2 days incubation. After 4
days incubation, Brucella colonies are round, 1-2 mm in diameter, with smooth (S)
margins, translucent and a pale honey color when plates are viewed in the daylight
through a transparent medium. When viewed from above, colonies appear convex and
pearly white. Later, colonies become larger and slightly darker. Smooth Brucella
cultures, especially B. mellitensis cultures, have a tendency to undergo variation during
growth, especially with subcultures, and dissociate to rough (R) forms, and sometimes
mucoïd (M) forms. Colonies are then much less transparent with a more granular, dull
surface (R) or a sticky glutinous texture (M), and range in color from matt white to
brown in reflected or transmitted light. Intermediate (I) forms between S, R and M forms
may occur in cultures undergoing dissociation to the non-smooth state. Changes in the
colonial morphology are generally associated with changes in virulence, serological
properties and phage sensitivity. (Alton et al., 1988; Quinn et al., 1999; OIE, 2009a;
Godfroid et al., 2010).

2.3 Taxonomy of Brucella spps

Due to high degree of DNA homology in DNA-DNA hybridization assays (>90%


identity), including the recently recognized marine mammal strains , it was proposed
that the genus Brucella should be a monospecific genus, with B. melitensis as the sole
species and the other species should be considered as biovars (Verger et al., 1985, 1987,
28
1998, 2000; Xavier, 2009). Conversely, several molecular genotyping methods have
been developed and applied to characterize Brucella spps, indicating that significant
DNA polymorphisms occur between species, which favor the current multi-species
classification of Brucella (Halling, 2005). Importantly, comparison of genome
sequences of B. suis and B. melitensis demonstrated that exist clusters of genes that are
unique in both species (designated genetic islands). It is reasonable to hypothesize that
these unique genes may contribute to the differences in host specificity between
Brucella spps (Tsolis, 2002). Furthermore, recent studies based on comparative whole
genome analysis of several Brucella spps indicate that there is limited divergence with a
large number of pseudo genes. Interestingly, these genomic analyses do not clearly
explain the host preferences of Brucella spps (Wattam, 2009; Foster, 2009). One of
these studies indicates that at the B. ovis is the basal lineage to the rest of the Brucella
spps and that apparently most Brucella spps diverged from their common B. ovis
ancestor in the past 86,000 to 296,000 years (Foster, 2009).

The International Committee on Systematics of Prokaryotes (ICSP), Subcommittee on


the taxonomy of Brucella recommended a taxonomic classification that includes
different species within the genus, either classical or new, which are still considered as
individual species. Therefore, the genus currently group ten species, namely B.
melitensis, B. abortus, B. suis, B. neotomae, B. ovis, B. canis, B. ceti, B. pinnipedialis, B.
microti and B. innopinata (ICSP, 2010).

B. ovis and B. canis have only been encountered in the rough form whereas the rest of
the species occurs in smooth form. Three biovars are recognized for B. melitensis (1-3),
seven for Brucella abortus (1-6 and 9), and five for Brucella suis (1-5) (Godfroid et al.,
2010).

29
2.4 History and importance of Brucella melitensis

Sheep and goats Brucellosis (excluding Brucella ovis infection which is rarely
pathogenic for humans) is a zoonotic infection with important effects on both public
health and animal health and production and is widespread in many areas of the world,
particularly in some Mediterranean and Middle Eastern countries. Brucella mellitensis,
the main etiologic agent of Brucellosis in small ruminants, was the first species in the
genus Brucella described. It was first isolated by Bruce in 1887 from the spleens of
soldiers dying of Mediterranean fever on the island of Malta. Bruce called it
Micrococcus melitensis. The origin of the disease remained a mystery for nearly 20
years until it was discovered that goats were the source of infection for human
population (Nicoletti, 2002).

2.5 Brucella melitensis infection in sheep and goats

The major route of infection is through the mucous membranes of the oropharynx and
upper respiratory tract or the conjunctiva following contact with contaminated pastures.
Other potential routes of infection are through the mucous membranes of the male or
female genital tract. After gaining entrance to the body, the organisms encounter the
cellular defenses of the host, but generally succeed in arriving via the lymph channels at
the nearest lymph node. The fate of invading bacteria is mainly determined by the
cellular defenses of the host, chiefly macrophages and T lymphocytes, though specific
antibody undoubtedly plays a part. The outcome depends on the ruminant species
infected, age, immune status of the host, pregnancy status, and the virulence and number
of the invading Brucella. When the bacteria prevail over the body defenses, a bacteremia
is generally established. This bacteremia is detectable after 10 to 20 days and persists
from 30 days to more than two months. If the animal is pregnant, bacteremia often leads
to the invasion of the uterus. At the same time, infection becomes established in various
lymph nodes and organs, often in the udder and sometimes in the spleen. During this
first stage of infection, the major clinical sign is abortion but other signs due to a
30
localization of Brucella may be observed i.e. orchitis, epididymitis, hygroma, arthritis,
metritis, subclinical mastitis, occasionally retained placenta. However, numerous
animals develop self-limiting infections or they become asymptomatic latent carriers.
Abortion generally does not occur if the female becomes infected at the third trimester
of pregnancy (Bishop et al., 1994; Radostits et al., 2007; CSFPH, 2009)

The second stage is characterized by either elimination of Brucella or, more frequently,
by a persistent infection of mammary glands and supramammary and genital lymph
nodes with constant or intermittent shedding of the organisms in the milk and genital
secretions (Radostits et al., 2007; Benkirane. A, 2006)

Animals generally abort once during the second trimester, but re-invasion of the uterus
occurs in subsequent pregnancies with shedding in fluids and membranes. The
pregnancy can also continue to full-term. The proportion of newly infected females that
abort varies with the circumstances. The percentage of infected females lambing/kidding
in a flock may reach 40% (Radostits et al., 2007). Females that are born into an infected
environment and subsequently infected generally abort less frequently as compared to
those born in uninfected environment but later get infected. This explains the high level
of abortions in newly infected flocks and their relatively low frequency in flocks where
infection is enzootic (Radostits et al., 2007). Greatly reduced milk yield follows
abortion, and infection of the udder following a normal birth also leads to a considerable
reduction in yield. In spite of this, clinical signs of mastitis are seldom detectable in
naturally infected goats (Alton 1990; Radostits et al., 2007; Godfroid et al., 2005).

2.6 Pathological lesions in animals

Brucella-infected animals generally develop granulomatous inflammatory lesions which


frequently are found in lymphoid tissues and organs such as reproductive organs, udder
and supramammary lymph nodes and sometimes joints and synovial membranes. The
lesions when present are not pathognomonic. The following could be observed:

31
necrotizing placentitis, palpable testicular alterations, necrotizing orchitis and
epididymitis with subsequent granulomatous, necrotizing seminal vesiculitis and
prostatitis. Acute mastitis with palpable nodules and the production of clotted and
watery milk may occur. Some aborted fetuses may have an excess of blood-stained
fluids in the body cavities, with enlarged spleen and liver. Others appear normal.
Infected fetal membranes show changes affecting part or all of the membrane. The
necrotic cotyledons lose their blood-red appearance becoming edematous and dull-grey
in color (Radostits et al., 2007; Godfroid et al., 2004).

2.7 Brucella melitensis infection in humans

Human Brucellosis is widely distributed all over the world, with regions of high
endemicity such as Mediterranean, Middle East, Latin America, parts of Asia and Africa
(Corbel, 1997). The incubation period of Brucellosis normally is 1–3 weeks, but it can
be several months before showing signs of infection. B. melitensis is associated with
acute infection whereas the infections with other species are usually sub-acute and
prolonged (Mantur et al., 2007). B. melitensis is the most virulent Brucella for humans
with a few organisms (10 to 100) being sufficient to cause a debilitating infection
(Fugier et al., 2007).

The World Health Organization (WHO) laboratory biosafety manual classifies Brucella
(and particularly B. melitensis) in risk group III (OIE, 2009b). Humans acquire
Brucellosis mainly through ingestion of contaminated milk and unpasteurized dairy
products. Contact of mucosa and skin abrasions with fluids and tissues from aborted
fetuses of infected animals are also important sources of Brucella transmission (Fugier
et al., 2007). Furthermore, people may be infected by inhalation of contaminated dust or
aerosols. Thus, Brucella is one of the most common laboratory acquired pathogens
worldwide and is included in the potential biological weapon list (Xavier, 2009).

32
Humans are accidentally infected and almost always are dead-end hosts of Brucella
infections. The disease is primarily an occupational risk in exposed professions, i.e.
veterinarians, farmers, laboratory technicians, abattoir workers, and others who work
with animals and their products. People living near infected premises may also contract
infection. The primary source is the animal and infection is contracted either by direct or
indirect contact through the skin or mucous membranes or ingestion of contaminated
products, especially fresh dairy products. The maximum danger is therefore during the
lambing or kidding period. Dairy products are the main source of infection for people
who do not have direct contact with animals. However much of the milk which is
consumed is nowadays rendered safe by pasteurization or boiling, but cheese made from
sheep and goat milk is preferably prepared from untreated milk may come from Brucella
infected animals. During the course of cheese manufacture, any Brucella present in the
milk become trapped in the clot and thus concentrated in the cheese, although bacteria
may subsequently be inactivated by manufacturing or ripening processes. The
prevalence of human Brucellosis acquired from dairy products is seasonal, reaching a
peak soon after kidding and lambing. Abattoir workers handling infected animals are
also at risk, especially from the contents of uteri and udders. The handling of raw wool
has been identified as a potential source of infection of workers involved. B. melitensis
is also easily acquired by laboratory infection (Kangethe et al., 2007; OIE, 2009a; OIE,
2009b).

Human infections with B. melitensis have variable clinical manifestations and can
become life threatening (Colmenero, 2002). Although the majority of patients present
with general symptoms, such as fever, malaise, sweats and lymphadenopathy and/or
hepatosplenomegaly, a more severe form of the disease can be accompanied with osteo-
articular signs (spondylitis, arthritis and osteomyelitis) or genitourinary tract changes
(orchitis, epididymitis, glomerulonephritis and kidney abscesses) (Colmenero, 2002).

Human Brucellosis is also known for complications and involvement of internal organs
and its symptoms can be very diverse depending on the site of infection and include
33
encephalitis, meningitis, spondylitis, arthritis, endocarditis, orchitis, and prostatitis
(Acha and Szyfres, 2003). Spontaneous abortions, mostly in the first and second
trimesters of pregnancy, are seen in pregnant women infected with Brucella (Khan et al.,
2001). Although rare complication, Brucella endocarditis (<2% of cases) is most
commonly associated with B. melitensis infection and is the most severe complication. It
accounts for at least 80% of deaths due to Brucellosis (Reguera et al., 2003). Lack of
appropriate therapy during the acute phase may result in localization of Brucella in
various tissues and organs and lead to sub-acute or chronic disease that is very hard to
treat (Young, 1995). Nervous, genitourinary, hepatosplenic and cardiovascular
complications been observed. Brucellosis is termed chronic when it includes one or
more of the signs described above and persists or recurs over a period of six months or
more. Brucella dermatitis traditionally known as “allergy" to Brucella has also been
associated with B. melitensis. Symptoms and signs of Brucellosis usually referred to as
fever of unknown origin can be confused with other diseases including enteric fever,
malaria, rheumatic fever, tuberculosis, cholecystitis, thrombophlebitis, fungal infection,
autoimmune disease and tumors (Mantur et al., 2007). Live animal vaccines B.
melitensis Rev. 1 and B. abortus strain 19 are known to cause disease in humans. The
course of the disease with vaccine strains is usually shorter and more benign (Acha and
Szyfres, 2003). Direct person-to-person spread of Brucellosis is extremely rare. Mothers
who are breast-feeding may transmit the infection to their infants and sexual
transmission has also been reported (Carrera et al., 2006; Kato et al., 2007).

2.8 Diagnosis of Brucellosis in sheep and goats


2.8.1 Clinical signs of Brucellosis in sheep and goats

Brucellosis should be considered in flocks and herds with history of abortions and
stillbirths occurring without concurrent illness. Other diseases causing abortion in small
ruminants, particularly chlamydiosis and coxiellosis, should be considered. B. ovis can
also cause epididymitis and orchitis in rams (CSFPH, 2009).

34
2.8.2 Staining and microscopy of Brucella organisms

Microscopic examination of smears stained with the Stamp's modification of the Ziehl-
Neelsen method can be useful for a presumptive diagnosis, particularly if the direct
examination is supported by serology. Brucella spps are not truly acid-fast, but they are
resistant to decolorization by weak acids, and stain red against a blue background.
Brucella organisms are coccobacilli or short rods, usually arranged singly but sometimes
in pairs or small groups. This test is not definitive. Other organisms that cause abortions
such as Chlamydophila abortus and Coxiella burnetii can resemble Brucella. B. ovis,
which causes epididymitis and orchitis in rams, can also be confused with B. melitensis.
Immuno-staining is sometimes used to identify Brucella in smears (Alton et al., 1988).

2.8.3 Serology of Brucellosis

Serology can be used for a presumptive diagnosis of Brucellosis, or to screen flocks.


Serological tests are not completely specific and cannot always distinguish reactions due
to B. melitensis from cross-reactions to other bacteria, particularly Yersinia
enterocolitica O: 9. The most commonly used serological tests in small ruminants are
the buffered Brucella antigen tests, the card agglutination test (CAT) and rose bengal
plate agglutination tests(RBPT) and the complement fixation test (CFT). Indirect or
competitive enzyme-linked immunosorbent assays (ELISAs) are also used. A brucellin
allergic skin test is sometimes used to test unvaccinated sheep and goats for B.
melitensis. This test is performed by injecting the allergen into the lower eyelid resulting
into delayed type hypersensitivity characterized by a local swelling and induration (OIE,
2009a, b; CSFPH, 2007).

2.8.3.1 Antigenic cross reactions of Brucella organisms

All smooth Brucella strains show complete cross-reaction with each other in
agglutination tests with unabsorbed polyclonal antisera, a cross-reaction which does not

35
extend to non-smooth variants. Cross-reactions between non smooth strains can be
demonstrated by agglutination tests with unabsorbed anti-R sera. Lipopolysaccharide
(LPS) comprises the major surface antigens of the corresponding colonial phase
involved in agglutination. The S-LPS molecules carry the A and M antigens, which have
different quantitative distribution among the smooth Brucella strains. This is of value in
differentiating biovars of the major species using absorbed monospecific A and M
antisera. Serological cross-reactions have been reported between smooth Brucella and
various other Gram negative bacteria, e.g. Escherichia coli O: 116 and O: 157,
Salmonella group N (O: 30) of Kaufmann-White, Pseudomonas multophilia, Vibrio
cholera and especially Yersinia enterocolitica O: 9. These organisms can induce
significant levels of antibodies which cross react with S-LPS Brucella antigens in
diagnostic tests (Nielsen et al., 2006; Munoz et al., 2005; CSFPH, 2007).

2.8.4 Culture of Brucella organisms

A definitive diagnosis can be made if B. melitensis is cultured from an animal. Brucella


spps can be isolated on a variety of plain media, or selective media. Vaginal swabs and
milk samples are the best samples to isolate B. melitensis from live sheep and goats. B.
melitensis can also be cultured from aborted fetuses (stomach contents, spleen and lung)
or the placenta. The spleen, mammary and genital lymph nodes, udder and late pregnant
or early post-parturient uterus are the most reliable samples to collect at necropsy. This
organism can also be cultured from semen, the testis or epididymis, and arthritis or
hygroma fluids.

Brucella spps are aerobic, but some strains require an atmosphere containing 5-10%
carbon dioxide (CO2) added for growth, especially on primary isolation e.g. B. abortus
wild type (biovars 1-4). Others, like B. abortus wild type (biovars 5, 6, 9), B. abortus
S19 vaccine strain, B. melitensis, and B. suis, do not require CO2 for growth. The
optimum pH for growth varies from 6.6 to 7.4, and culture media should be adequately
buffered near pH 6.8 for optimum growth. The optimum growth temperature is 36-38°C,
36
but most strains can grow between 20°C and 40°C. Brucella requires biotin, thiamin and
nicotinamide. The growth is improved by serum or blood, but haemin (V-factor) and
nicotinamide-adenine dinucleotide (X-factor) are not required. The growth of most
Brucella strains is inhibited on media containing bile salts, tellurite or selenite. Growth
is usually poor in liquid media unless culture is vigorously agitated. Growth in static
liquid media favors dissociation of smooth-phase cultures to non-smooth forms.
Continuous and vigorous aeration will prevent this, provided a neutral pH is maintained.
In semi solid media, CO2-independent Brucella strains produce a uniform turbidity from
the surface down to a depth of a few millimeters, while cultures of CO2-requiring strains
produce a disk of growth a few millimeters below the surface of the medium (Godfroid
et al., 2010; OIE, 2009a, b, CSFPH, 2007).

Since B. melitensis does not require serum or CO2 for growth and can be isolated on
ordinary solid media under aerobic conditions at 37°C, however, the use of nonselective
media cannot be recommended because of the risk of overgrowing contaminants usually
present in field samples. Selective media are needed for isolation purposes. The Farrell’s
selective medium, developed for the isolation of B. abortus from milk (Farrell, 1974), is
also recommended for the isolation of B. melitensis (Alton et al., 1988). However,
nalidixic acid and bacitracin, at the concentration used in this medium, may have
inhibitory effects on some B. melitensis strains (Marín et al., 1996b). Thus, its sensitivity
for the isolation of B. melitensis from naturally infected sheep is sometimes lower than
that obtained with the less selective Thayer-Martin’s modified medium (Marín et al.,
1996a). The sensitivity of bacteriological diagnosis is significantly increased by the
simultaneous use of both the Farrell’s and the modified Thayer-Martin’s media (Marín
et al., 1996b). Non selective, biphasic medium, known as Castaneda’s medium, is
recommended for the isolation of Brucella from blood and other body fluids or milk,
where enrichment culture is usually advised. Castaneda’s medium is used because
Brucella tends to dissociate in broth medium, and this interferes with bio-typing by
conventional bacteriological techniques. B. melitensis can be identified to the species

37
and biovars level by phage typing and cultural, biochemical and serological
characteristics. Genetic techniques can also be used for bio-typing. Biotyping of
Brucella spps is performed using different tests, the most important being agglutination
tests with antibodies against rough or smooth LPS, i.e. against the A or M epitopes of
‘O’ chain polysaccharides (O-LPS); lysis by phages, dependence on CO2 for growth,
measured usually in primary cultures; production of H2S; growth in the presence of
basal fuchsine or thionine; and the crystal violet or acriflavine tests (Alton et al., 1988).
The vaccine strain (B. melitensis strain Rev.1) can be distinguished from field strains by
its growth characteristics and sensitivity to antibiotics and other additives (Godfroid et
al., 2010; OIE, 2009a, b).

2.8.5 Biochemical tests for Brucella organisms

The metabolism of Brucella is oxidative and Brucella cultures show no ability to acidify
carbohydrate media in conventional tests. The Brucella spps are catalase positive and
usually oxidase positive, and they reduce nitrate to nitrite (except B. ovis and some B.
canis strains). The production of H2S from sulphur containing amino-acids also varies.
B. melitensis does not produce H2S. Urease activity varies from fast to very slow. Indole
is not produced from tryptophane and acetyl-methyl-carbinol is not produced from
glucose (Alton et al., 1988; Quinn et al., 1999; Corbel et al., 2005; Godfroid et al.,
2010).

2.8.6 Susceptibility to phages by Brucella organisms

Over 40 Brucella phages have been reported to be lytic for Brucella members. All
phages are specific for the genus Brucella, and are not known to be active against any
other bacteria that have been tested. Thus, lysis by Brucella phages is a useful test to
confirm the identity of Brucella spps and for speciation within the genus. The Brucella
phages currently used for

38
Brucella typing are: Tbilisi (Tb), Weybridge (Wb), Izatnagar1 (Iz1) and R/C.

The three former phages are used for differentiation of smooth Brucella spps. R/C is
lytic for B. ovis and B. canis (OIE, 2009a).

2.8.7 Other methods for detection of Brucella organisms

Animal inoculation is uncommonly used for isolation, but occasionally necessary when
other techniques fail. Guinea pigs or mice can be used (CSFPH, 2009).

Several PCR based methods have also been developed. The best validated methods are
based on the detection of specific sequences of Brucella spps such as the 16S-23S genes,
the IS711 insertion sequence or the bcsp31 gene encoding a 31-kDa protein (Ouahrani-
Bettache et al., 1996; Baddour and Al-Khalifa, 2008).

2.8.8 Sero-prevalence of Brucellosis in small ruminants

In a study conducted in Jordan on ovine and caprine Brucellosis (Brucella melitensis) in


aborted animals in Jordanian sheep and goat flocks, 255 biological samples were
collected from188 animals (81 sheep and 107 goats) during the lambing season from
September 2009 to April 2010 from the Mafraq region of Jordan. Sampled animals
belonged to 93 sheep and goat flocks that had abortion cases in the region. One hundred
and seven (41.9%) biological samples were positive for the omp2 primers that were able
to identify all Brucella species in the collected samples which were obtained from 86
aborted animals (86/188 = 45.7%). These positive samples were obtained from 28 sheep
and 33 goats. The prevalence rate of B. melitensis was 27.1% (51/188) among aborted
animals (Samadi et al., 2010).

A survey to estimate the seroprevalence of ovine and caprine brucellosis was conducted
in the region of Trás-os-Montes e Alto Douro, Northeast of Portugal. In total, 278,097
small ruminants and 5,466 flocks from 13 Livestock Farmers Organizations (OPP’s)

39
were analyzed. A total of 487 (8.9%) flocks had one or more serologically positive
animals with values ranging between 8.2% and 9.7%. The individual seroprevalence was
0.44%. There were significant differences in seroprevalence rates among herd sizes,
species and constitution of herd, production’s type and OPP (Coelho et al., 2013).

In Iran, a cross-sectional study conducted to estimate seroprevalence and to identify


flock-level factors associated with seropositivity to Brucellosis in small ruminants in
Kerman province, southeastern Iran. In October-November 2011, serum samples were
randomly collected from 1767 sheep and 1233 goats, older than 18 months, from 300
flocks. The sera were initially screened for the presence of anti-Brucella antibodies using
the Rose-Bengal test; those found to be positive were then examined by Wright and 2-
mercaptoethanol Brucella agglutination tests. A total of 63 (21%) flocks had at least one
seropositive animal (Sharifi et al., 2014).

In a cross-sectional study done in Ethiopia in in South Omo Zone of Southern Ethiopia


in goats, out of 384 tested animals; 20 (5.2%) of them were found positive for RBPT. Up
on further testing the RBPT positive sera by CFT, 16 (4.2%) were found seropositive.
Thus, overall number of seropositive goats in selected areas of South Omo Zone was 16
(4.2%) after RBPT and CFT. This study also found that Seroprevalence of caprine
brucellosis was significantly associated with abortion rate in the animals (Ashagrie et al.,
2011). In another cross-sectional study done in Somali Regional State, Eastern Ethiopia,
sera samples were screened by Rose Bengal Plate Test (RBPT), and all samples tested
positive by the RBPT were subjected to Complement Fixation Test (CFT) for
confirmation and a total of 730 sera samples (421 of sheep and 309 of goats) were tested
of which 12 serum samples that were positive by RBPT, 11 were positive by CFT
(Bekele et al., 2011). Yet in another cross sectional study carried out from October 2008
to April 2009 to determine the sero-prevalence of brucellosis in small ruminants in and
around Bahir Dar, northwest Ethiopia, a total of 500 serum samples (270 from sheep and
230 from goats) were collected from extensive management system with no history of
vaccination. All serum samples were initially screened by Rose-Bengal-Plate Test
40
(RBPT) and positive reactors to RBPT (n=6) were further tested by complement fixation
test (CFT) for confirmation of which 2 (0.4%) turned positive (Ferede et al., 2011). In
another study conducted in Ethiopia in Dire Dawa region Eastern Ethiopia in small
ruminants, out of 384 sheep and goats sera tested using Rose Bengal plate test (RBPT)
and complement fixation test (CFT), 36 (9.38%) reacted positively using RBPT. Of
these reactant sera, 35 also tested positive using CFT, giving an overall prevalence of
small ruminant brucellosis of 9.11% (Negash et al., 2012). In another sero-prevalence
study on small ruminant Brucellosis in selected districts of Afar and Somali pastoral
areas of Eastern Ethiopia, Sera from 2000 sheep and goats were tested by Rose Bengal
Plate test (RBPT) and Indirect Enzyme Linked Immuno – Sorbent Assay (I-ELISA). Out
of the 2000 sera tested 1.9 % (n= 38) were positive to RBPT and 9.7 % (n=193) were
positive to I – ELISA. Brucella antibodies were more prevalent in goats (13.2%) as
compared to sheep (5.6%). This difference was statistically significant (X2 = 32.5; P <
0.001) (Teshale et al., 2006).

In a study conducted in Niger to determine sero-prevalence and potential risk factors for
Brucella

Spps infection in traditional cattle, sheep and goats reared in urban, peri-urban and rural
areas, 5,192 animals from 681 herds were included in the study. Serum samples and
hygroma fluids were collected and tested for Brucellosis antibodies using indirect
ELISA. The true adjusted herd-level sero-prevalence of Brucellosis ranged between
11.2% and 17.2% and the true adjusted animal-level sero-prevalence was 1.3%
(Boukary et al., 2013).

2.8.9 Risk factors for small ruminant Brucellosis

In a cross-sectional study conducted in Iran to determine risk factors of small ruminant


brucellosis in Southeast Iran in 2012, after adjustment for the sampling fraction, a
multivariable multi-level logistic model was used to detect the potential risk factors of

41
the infection. The final model identified presence of purchased animals (OR=8.39; 95%
CI: 1.10-64.90) as an independent factor associated with Brucellosis sero-prevalence
(Sharifi et al., 2012; Sharifi et al., 2012).

In a case control study conducted in Mexico to determine risk factors for brucellosis
seropositivity of goat herds in the Mexicali valley of Baja California, Mexico, an
important risk factor for goat herd brucellosis seropositivity in the Mexicali Valley was
importation of goats from another state in Mexico. This was explained to be attributed to
transportation of goats from higher-prevalence regions of the country to Mexicali which
is a low-prevalence region (Mikolon et al., 1998).

In a cross-sectional study conducted in Spain to determine risk factors for Brucellosis


sero-prevalence of sheep and goat flocks in Spain, Eleven risk factors were studied at the
group-level by logistic regression using flock brucellosis-status as outcome, and by
linear regression using percentage of brucellosis-seropositivity as outcome. Both final
models contained the same variables: contact with sheep and grazing in communal
pastures as risk factors, and frequency of disinfecting practices as a protective factor
(Reviriego et al., 2000).

In a study conducted in Niger to determine sero-prevalence and potential risk factors for
Brucella

Spps infection in traditional cattle, sheep and goats reared in urban, peri-urban and rural
areas, (Boukary et al., 2013). At herd level, the risk of transmission was increased by
transhumance (OR of 5.4; 95% CI: 2.84–10.41), the occurrence of abortions (OR of 3.0;
95% CI: 1.40–6.41), and for herds having more than 50 animals (OR of 11.0; 95% CI:
3.75–32.46) (Boukary et al., 2013).

42
2.8.10 Knowledge attitude and practices towards Brucellosis

In Africa, a KAP study on Brucellosis conducted in Egypt showed a relative high


general knowledge of Brucellosis but still a high-risk behavior among livestock owners
which the authors concluded might contribute to a high sero-prevalence of brucellosis
among livestock in the area (Holt et al., 2013). Another study done in Uganda found that
respondents had moderate overall knowledge of human and animal Brucellosis and agro-
pastoralists were found to have better knowledge compared to the pure pastoralists
(Kansiime et al., 2014).

43
2.8.11 Conceptual framework

Husbandry Practices Community Knowledge attitude


and practices
 Herd size
 Herd Immunity  Education level
 Management /production system  Awareness of brucellosis
(nomadic pastoralism vs non-  Awareness of cause of Brucellosis
nomadic)  Awareness of modes transmission
 Uncontrolled livestock movement to humans and animals
 Herding of different species  Awareness on signs and symptoms
together in humans and animals
 Use of common pastures and  Attitudes and practices towards
water sources Brucellosis e.g. consumption of
 Purchase of infected animals from unpasteurized milk and milk
the market for replacement or products and assisting animals
upgrading during parturition without
 Coming into contact with wildlife protective gear (gloves)
 Seeking veterinary services
 Presence of aborting animals in
the herd

Infection of sheep and goats with Brucellosis

Infection of humans with Brucellosis

Figure 2.1: Conceptual Framework

44
CHAPTER THREE

MATERIALS AND METHODS

3.1 Study area

Garissa County is one of the three counties of the original North Eastern Province. The
County is further divided into seven sub-counties namely: Garissa Township,
Balambala, Lagdera, Dadaab, Fafi, Ijara and Hulugho. It boarders Isiolo county to the
Northwest, Wajir county to the North and Republic of Somalia to the East, Tana river
county to the west and Lamu county to the south. The county covers an area of 44,175
square kilometers. Garissa County is low lying with altitudes ranging between 70 m and
400 m above sea level. The River Tana, which runs along the western boundary of the
County, is the only permanent river. Though it is not confined within the county’s
boundaries, the river has tremendous influence over the climate, settlement patterns, and
economic activities within the County, as it forms the single most important source of
water for the fast growing Garissa Town and the surrounding areas. The county is
generally semi-arid and receives annual rainfall of about 40 mm.

The soils range from the sandstone, dark clays in some patches, to alluvial soils along
the river Tana basin. Garissa County’s economy is mainly pastoral. The main economic
activity is keeping of cattle, camels, goats and sheep. Increased livestock population has
led to overgrazing and competition over watering grounds. The county receives rain in
two seasons, these are the long rains season between March and April and the short rain
season between October and December. The rainfall is unreliable with some torrential
rains which in many cases are detrimental to vegetation growth. The temperatures in the
county are high ranging from 20oC to 38oC. The natives of Garissa County are mainly
Somali’s whose main activity is pastoralism, and their staple food is mainly meat and
milk (Arid lands development project, Kenya 2011). The study was focused in Garissa
Township and Balambala sub-counties highlighted (Figure 3.1).

45
Figure 3.1: Map of Kenya showing Garissa County highlighted in yellow color and
study area highlighted in red color

3.2 Study design

This was a cross-sectional study done in Garissa County, Balambala and Garissa
Township sub-counties.

3.3 Study Population

Goat and sheep populations in the County comprised the study population. According to
2009 Kenya national housing and population census, the sheep and goat population in
the county was estimated at 3, 315,061 sheep and goats with 2, 090,613 goats and
1,224,448 sheep. Garissa (constituting Balambala and Garissa Township) had estimated
46
goat population of 1,000,856 and 312,601 sheep and a total of 1,313,457 (Table 3.1).
The two sub-counties were chosen due to the large population of sheep and goats
compared to other sub-counties. The pastoralists’ within Balambala and Garissa sub-
counties also constituted the study population

Table 3.1: Sheep and goats population, Garissa County

Sub-county Sheep Goats Total


Garissa 312,601 1,000,856 1,313,457
Lagdera 489,282 561,883 1,051,165
Fafi 98,889 179,226 278,115
Ijara 323,676 348,648 672,324
Total 1,224, 448 2,090,613 3,315,061

Source: Kenya National Bureau of Statistics 2009 population and housing census
report

3.4 Sample size determination for sheep and Goats

Herds were regarded as the primary sampling units. A herd was defined as any cluster
or aggregate of animals, e.g. flock or group of animals under the same management
system but not necessarily owned by a single individual that are able to frequently mix.
Animals in a herd share common risk factors for disease, so the distribution of disease
within the herd is assumed to be relatively homogenous. The total number of herds to be
sampled was calculated using the formula for simple random sampling as suggested by
(Thrushfield et al., 2013).

Equation 1: Formula for Sample Size Calculations

47
48
Where:

n= minimum number of herds or flocks to be sampled;

1.96= z-score at 95% confidence interval

d = desired absolute precision assumed at 10%;

p = expected herd or flock prevalence assumed at 16%

Based on a Brucellosis survey done in Kajiado, which has similar ecosystem as Garissa
by the Kenya Zoonotic disease unit (ZDU), sero-prevalence of Brucellosis in goats was
16% and 11% in sheep (Osoro et al., 2015). The sero-prevalence in goats was used to
calculate the sample size. Based on the above parameters, a minimum herd sample size
of 52 herds was determined. The resulting number of herds needed for sampling was
multiplied by a design effect to consider the multistage level clustering of the sampling
design that was employed. When no estimate of intra-cluster correlation coefficient is
available, a qualitative assessment of intra-cluster correlation coefficient might be used
(Ariwan and Frerichs, 1996). Suggested qualitative levels of ‘‘low,’’ ‘‘medium,’’ and
‘‘high’’ intra-cluster correlations are associated with values of the design effect: 2, 4,
and 7, respectively were proposed. The design effect is the variance of an estimated
proportion obtained by a cluster sample divided by the variance for a simple random
sample (Dargatz and Hill, 1996). For this study, a low design effect of 2 was assumed
due to high homogeneity and low heterogeneity within and between herds. The adjusted
minimum sample size after multiplying with the design effect was 104 herds. In
addition, 10% contingency was applied which gave rise to 114 minimum number of
herds.

49
3.5 Sampling design for sheep and Goats

This was undertaken using a multiple stage sampling technique. The first stage was
random selection of sub-locations in Garissa County to be sampled. As at the time of the
study, sub-location were the lowest government administrative unit headed by assistant
chief. Due to resource limitation a total of about a third of the sub-locations (12) was
randomly selected from list of 33 sub-locations of Garissa County (Table 3.2).

Table 3.2: List of sub-locations in Garissa and Balambala Township and the Sub-
locations randomly selected for Sampling highlighted in yellow

No Sub-County Name of sub-locations for sampling


1 Garissa Township sub-county Township
2 Galbet
3 Medina
4 Malika
5 Waberi
6 Iftin
7 Korakora
8 Edet
9 Boralgi
10 Jarirot
11 Sankuri
12 Balich
13 Bololoweyn
14 Shidey
15 Raya
16 Atheley
17 Shabaha
18 Shimbirey
19 Abdisamit
20 Balambala sub-county Saka
21 Saka junction
22 Daley
23 Balambala
24 Dujis
25 Jarajara
26 Kasha
27 Ashadin
28 Kuno

50
29 Danyere
30 Ohio
31 Kone
32 Sikley
33 Libahoi
34 Madey
35 Urgad
36 Dogob

Due to lack of comprehensive information on the number of sheep and goats herds (n) in
the study area, a base line study was conducted. During the baseline study, the selected
sub-locations were visited and with assistance of the local administration (assistant chief
and community elders), local veterinary staff and community animal health workers
(CAHWs), a comprehensive list of households and approximate number of animals
owned by each household was generated. A final list of herds in a particular sub-location
was generated putting into consideration the definition of herd as earlier outlined.
Household(s) whose animals met criteria for a herd were merged and listed as a single
herd under the name of the eldest member representing the merged households or as was
agreeable among the merged households. This final list of households and number of
animals owned by each household acted as a proxy to the number of herds of goats and
sheep in each sub-location and constituted the sampling frame from which actual
sampling was conducted. The second step was to identify the individual herds to sample
in each sub-location. The number of herds to sample per sub-location depended on the
number of herds in each sub-location (Sampling proportion to size). The third stage was
a systematic random sampling (SRS) to select individual animals in the herd for sample
collection. All eligible animals for sampling were placed within an enclosure with one
gate. The first animal per species was randomly selected and subsequent animals were
selected when animals of particular species were allowed to pass through the gate;
factoring in the kth interval (sampling interval) determined based on the number of
animals to be sampled within that species. This was repeated until the number of animals
to be sampled for that species was achieved. Due to logistical reasons, sampling of
individual animals within a herd was done proportional to size of the herd such that for

51
small herds (< 10 animals), all the animals in the herd were sampled, for medium herds
with sizes (>10 but <50), all the animals to a maximum of 20 animals were sampled and
for large herds, (>50 animals), a maximum of 20 animals were sampled. In this set up,
sheep and goats are raised together as one herd therefore the above sample size applied
to both. In any particular herd, sampling of goats and sheep was done proportional to
their size within the herd. To facilitate sampling of the herds, list of selected herds for a
particular sub-location were sent in advance to a community animal health worker
(CAHW) within the sub-location to mobilize the pastoralists whose herds were selected
for sampling. A total of 120 herds were sampled during the study and the distribution of
the number of herds sampled per sub-location is as shown in the Table 3.3 below.

Table 3.3: Distribution of number herds sampled per sub-location

Sub-location Total herds listed Number of herds sampled


Kuno 50 14
Madey 32 9
Shimbirey 42 12
Sikley 36 10
Balambala 54 15
Medina 30 8
Kasha 28 8
Dogob 18 5
Boralgi 38 11
Galbet 22 6
Ohio 44 12
Abdisamit 32 9
Total 426 120

3.6 Blood sample collection

Sheep and goats selected for sample collection were individually restrained and 9ml
blood collected in plain red topped vacuum plastic tubes (Vacutainer®) from the jugular

52
vein. To allow clot separation, all blood samples were left to stand for approximately 15
to 30 minutes in a slanting manner at ambient temperature to separate serum from clot.
Serum was collected from the Vacutainer using a disposable plastic Pasteur pipette,
dispensed to an Eppendorf /cryovial tube and stored in a cool-box containing ice in the
field. Eppendorf/cryovial tubes were then stored in the freezer at -20°C until used for
serological testing.

3.7 Serological testing

The test procedures was done at the regional veterinary investigations laboratory (RVIL)
in Garissa using test protocols as outlined by OIE (OIE, 2009a) and the manufacturer’s
specifications for the tests. Screening for presence of antibodies in collected sera
specimens was done using Rose Bengal Plate test (RBPT) and complement fixation test
(CFT). The sensitivity and specificity of RBPT were 89% and 97%, respectively and
that of CFT was 88% and 100%. The CFT and RBPT test antigens (Brucella arbortus
strain 99), control sera and other reagents were obtained from Atlas medical, William
James House, Cowley Rd. Cambridge Cb4, 4WX and sensitized sheep red blood cells
(SRBC) were obtained from the central veterinary laboratory (national veterinary
reference laboratory in Kenya). The sera specimens were tested serially first using RBPT
then CFT for those that tested positive on RBPT. An animal was considered positive if
the serum specimen tested positive on both RBPT and CFT whereas a herd was
considered positive if at least a single serum specimen from an animal within the herd
tested positive on both RBPT and CFT. For RBPT, Rose Bengal test antigen was
prepared from killed standard strain of B. abortus strain 99 and stained with Rose Bengal
dye, in an acidic buffer pH 3.65. Serum samples and the antigen were left at room
temperature for an hour before the test commenced. The bottle containing the antigen
was well shaken so that the suspension was homogeneous. Thereafter 30µl of the sample
was added after swirling for a minute onto a white tile and same volume of antigen
alongside the antigen spots using a micro-pippete. The sera and antigen were then
thoroughly mixed using a wooden splint, using one wooden splint for each test, until a
53
circular zone of approximately 2 cm was formed. The white tile was then rocked on both
clockwise and anti-clockwise for four minutes (timing was done using a laboratory
buzzer). Agglutination (due to antigen and antibody complex formation) was thereafter
observed in a well-lit place to avoid false positive reading due to formation of fibrin.
Magnifying glass was used to examine those tests that were suspected to have micro-
agglutination. A positive test results was any visible agglutination observed and negative
results was absence of any visible agglutination. Control serum was tested every day to
provide minimal agglutination before the actual testing began to verify the sensitivity of
the test conditions. For CFT, there was dilution of the test serum and appropriate
working standards with equal volume of veronal buffered saline in small tubes which
were then incubated at 58°C for 50 minutes to inactivate the native complement.
Thereafter, 25µl of diluted test serum was dispensed on a round bottom 96 well micro-
titre on the first and second rows of the well. Then 25µl of the veronal buffered saline
was added to all wells except those on the first row. Serial doubling dilution was applied
by transferring 25µl of serum from third row onwards and discarded 25µl of the mixture
in last row. This serial dilution was done four times. This was followed by dispensing
25µl of antigen to each well except in the first row followed by 25µl of complement to
each well. Control wells having diluent only; control wells having complement and
diluent; and control wells having diluent, complement and antigen each at 75µl volume
in each control wells were then set up. Control serum that results into a minimum
positive reaction for each set of tests to ascertain the sensitivity of test conditions was
tested each day. This was followed by incubation of the plates at 37°C for 30 minutes
and thereafter addition of 25μl of sensitized sheep red blood cells (SRBC) to each well.
This was followed by re-incubation of the plates at 37°C for 30 minutes. Thereafter the
plates were centrifuged at 100rpm for 10 minutes to allow the SRBC that did not
undergo hemolysis to settle. The degree of hemolysis was compared with standard
corresponding to 0, 25, 50, 75 and 100% hemolysis. Absence of complementary activity
was also checked for the serum in the first row. Sera samples having SRBCs
sedimentation at a dilution ≥ 1:5 were considered to be positive for Brucellosis. Using
54
the diagnostic sensitivity and specificity of RBPT and CFT, combined sensitivity and
specificity for the RBPT and CFT using a serial interpretation was calculated to be 78%
and 100% respectively (Ausvet animal health services, 2009). This combined sensitivity
and specificity together with the resulting apparent prevalence (AP) were factored in a
formula suggested by Rogan and Gladen (Rogan and Gladen, 1978) to obtain the true
prevalence (TP) at both the individual animal level and herd level.

55
Equation 2: Equation for determination of true sero-prevalence

3.8 Data Management


3.8.1 Questionnaire survey

A pre-tested structured questionnaire was used to collect information about potential


factors associated with Brucellosis herd sero-positivity and knowledge, attitude and
practices of pastoralists towards Brucellosis. The potential factors included herd size,
stocking density, management and breeding practices, purchasing of breeding stock in
the past one year; keeping other animals with sheep and goats and history of abortion in
animals in past one year. Some of the questions to assess knowledge attitude and
practices towards Brucellosis included what is Brucellosis, causes in both humans and
livestock, transmission, symptoms and signs in animals and humans, occurrence of
disease in the farmer/ farmer’s family/ neighbor, attitude towards handling infected
animals, drinking raw milk and consumption of unprocessed milk products and practices
like assisting animals during parturition or during abortion and methods of disposal of
aborted fetuses and placenta. The questionnaire was administered immediately after
bleeding the animals and services of a trained Somali speaking translator was used for
ease of administration. The translator was trained on questionnaire administration prior
to embarking on data collection to minimize on time taken to administer the
questionnaires. The pre-testing of the questionnaire was done on five herds to check for
consistency and any ambiguity. Subsequently, revisions were made on the questionnaire
based on pre-testing findings.

56
3.8.2 Data analysis

Data was entered, cleaned and analyzed using Epi Info 7 (CDC, Atlanta, GA, USA) and
Ms. Excel 2013 (Microsoft, Seattle, WA, USA). Univariate analysis was performed
where proportions’ was calculated for categorical variables and means and medians for
continuous variables. Bivariate analysis was carried out to evaluate the association
between herd Brucella sero-positivity and the potential factors. Odds ratios and 95%
confidence interval (CI) was calculated and factors with p-value of ≤ 0.05 were
considered statistically significant.

For selection of independent variables for inclusion into the initial multiple logistic
regression model, the entry criterion was p-value ≤ 0.20. The model was developed by
stepwise forward selection approach, dropping the least significant independent variable
until all the remaining predictor variables were significant (p-value ≤ 0.05). All
biologically and statistically plausible two-way interactions between variables remaining
in the final model was tested and retained if significant.

3.9 Ethical Approvals and Considerations

Protocol approval was sought and obtained from Board of post graduate studies of Jomo
Kenyatta University of Agriculture and Technology (JKUAT) and ethical clearance was
sought and obtained from Kenyatta National Hospital/University of Nairobi-Ethics and
Research Committee (KNH-UoN ERC). The aim and procedures of the study was
explained to participants’ who were required to give written consent prior to their
voluntary participation in the study. Blood samples were collected from animals of
consenting individuals and were only used to test for antibodies against Brucellosis.
Confidentiality of laboratory information was observed and maintained.

57
CHAPTER FOUR

RESULTS

4.1 Sero-prevalence of Brucellosis at Individual and Herd Level


4.1.1 Herd characteristics

A total of 2,400 animals composed of 979 (41%) sheep and 1471 (59%) goats were
sampled. These animals were sampled from 120 herds from 12 out of the 36 sub-
locations in Garissa Township and Balambala sub-counties. The 120 herds had a total of
12,945 animals of which 5,263 were sheep and 7,682 were goats. The median herd size
was 112 animals (range: 58 to 140 animals).

4.1.2 Brucella Sero-prevalence at individual animal level by animal species

Of the 2,400 sera samples examined, 18.2% (95% CI: 16.7% to 19.8%) were positive for
Brucella antibodies on RBPT. Of the samples positive on RBPT, 14.1%; (95% CI:
12.1% to 16.4%) were from sheep and 20.3% (95% CI: 18.4% to 22.5%) were from
goats. Of the samples testing positive on RBPT, 16.4% (95% CI: 15.0% to 18.0%) tested
positive on CFT of which 10.6% (95% CI: 8.8% to 12.7%) were sheep and 19.7% (95%
CI: 17.8% to 21.8%) were goats (Figure 4.1).

58
Figure 4.1: Distribution of Prevalence of Brucella antibodies by test type and
species, Garissa County, 2013 (n=2,400)

4.1.3 Brucella Sero-prevalence at individual animal level by sub-location sampled

Based on CFT, highest sero-prevalence of 37.8% (95% CI: 30.92% to 45.03%) was
recorded in Abdisimit sub-location followed by 25.4% (95% CI: 20.21% to 31.21%) in
Ohio and 22.5% (95% CI: 17.9% to 27.67%) in Kuno; whereas the lowest sero-
prevalence of 8.5% (95% CI: 5.2% to 13%) was recorded in Sikley and 2% (95% CI:
0.82% to 4.11%) in Balambala (Table 4.1).

59
Table 4.1: Distribution of individual animal prevalence of Brucella antibodies in
sheep and goats by sub-location, Garissa 2013 (n=2400)

Sub-location Total number CFT Positive 95% CI


sampled
n (%) (Lower Limit to Upper Limit)
Abdisimit 180 68 (37.8) 30.9 – 45.0
Ohio 240 61 (25.4) 20.2 – 31.2
Kuno 280 63 (22.5) 17.9 – 27.7
Galbet 140 28 (20) 14.0 – 27.2
Madey 180 32 (17.8) 12.7 – 23.9
Shimbirey 240 39 (16.3) 12.0 – 21.3
Boralgi 220 33 (15.0) 10.7 – 20.2
Dogob 100 15 (15.0) 9.0 – 23.0
Kasha 160 17 (10.6) 6.5 – 16.1
Medina 160 15 (9.4) 5.6 – 14.7
Sikley 200 17 (8.5) 5.2 – 13.0
Balambala 300 6 (2) 0.8 – 4.1
Total 2400 394 (16.4) 15.0 – 18.0

4.1.4 Adjusted Individual animal level Sero-Prevalence

Considering samples that were positive by both the RBPT and CFT, and when adjusted
to the two tests sensitivities and specificities, the overall true sero-prevalence of
Brucellosis at individual animal level in sheep and goats was 20.0% (95% CI: 18.2% to
22.0%); and in goats it was 24.3% (95% CI: 21.8% to 27.1%) and sheep 12.5% (95%
CI: 10.2% to 15.2%).

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4.1.5 Brucella Sero-prevalence at Herd Level

At the herd level, of the 120 herds sampled, the overall apparent herd prevalence was
51.7% 95% CI: 42.8% to 60.4%). The overall true herd sero-prevalence of Brucellosis
was 65.8% (95% CI: 54.3% to 77.2%); in goats it was 24.4% (95% CI: 18.2% to 22.0%)
and in sheep it was 12.5% (95% CI: 10.2% to 15.2%). Among the herds with positive
animals, the median number of animals testing positive was 8 animals (range: 3 to 15
animals). A total of 14 herds had four animals testing positive, 13 herds had five animals
testing positive and 14 herds had 10 animals testing positive (Table 4.2).

Table 4.2: Comparison of Number of Herds with Positive Reactors with Number of
animals testing Positive (n=62)

Number of animals testing positive Number of herds with positive reactors


Less than 3 0
3 9
4 14
5 13
6 2
7 1
8 5
9 1
10 14
11 1
12 1
13 0
14 0
15 1
Above 15 0
Total 62

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4.2 Factors Associated with Brucellosis Herd Sero-positivity
4.2.1 Bivariate Analysis for Herd level Factors Associated with Brucellosis in sheep
and goats

In determining both the risk factors and protective factors associated with Brucella herd
sero-positivity, several factors were examined at the bivariate analysis (Table 4.3).
Since Garissa County is in a pastoral set up where herds mix especially during grazing
and at watering points, 98 (82%) of the herds had come into contact with other herds of
sheep and goats. Herds that came into contact with other herds had almost 4 times higher
odds of having positive reactors to for Brucella compared to those herds that did not
come into contact with other herds and this was statistically significant (OR= 3.6; p-
value = 0.02). In contrast, coming into contact with wildlife or wildlife abortus was not
associated with herd Brucella sero-positivity.

In overall, abortion in sheep and goats herds was reported in 84 (70%) of the herds in
past one year. Of the herds that tested sero-positive, 52 (84%) had experienced abortion
in the past year compared to 32 (55%) in sero-negative herds. A herd that experienced
abortions within the past year were more than 4 times increased odds to have sero-
positive animals compared to herds that had not experienced abortion, an association
that was statistically significant (OR = 4.2; p-value = 0.001). However, experiencing
abortion in cattle herds (OR= 0.3; p-value = 0.52) was not associated with herd Brucella
sero-positivity in sheep and goats.

When comparing herds that introduced a new animal into the herd in the past year to
those herds that did not, herds that introduced new animals into the herds were more
than eight times increased chances of having sero-positive reactors for Brucellosis
compared to those that did not (OR =8.1; p-value <0.001).

62
The chances of a herd having seropositive reactors was almost three times higher among
those herds that lend male animals to other herds for breeding purposes as compared to
those herds that did not lend their male animals to other herds for breeding. This was
also statistically significant (OR= 2.8; p-value = 0.04).

Herds whose owners sought veterinary services in the past year compared to those that
whose owners did not seek veterinary services in the past year, had 60% reduced
chances of having sero-positive reactors to Brucella and this was statistically significant
(OR= 0.4; p-value = 0.02). Herd size (OR= 1.6; p-value = 0.33) was not statistically
associated with Brucella herd sero-positivity in sheep and goats.

All the respondents’ did not have calving pens for the herds, none of the animals
introduced into the herd over past year were quarantined before introduction, all the
respondents’ did not vaccinate their animals against Brucellosis.

63
Table 4.3: Comparison of factors associated with Brucella Sero-positivity among
positive and negative herds, Garissa 2013

(Positive herds n=62; Negative herds n=58)

Variable Herds Herds POR* (95% CI) P-value


Positive Negative
n (%) n (%)
Contact other sheep and goats herds in past year
Yes 56 (90) 42 (72) 3.6 (1.3 - 9.9) 0.02
No 6 (10) 16 (28) Reference
Contact wildlife in past year
Yes 23 (37) 17 (29) 1.4 (0.7 - 3.1) 0.48
No 39 (63) 41 (71) Reference
Contact wildlife abortus in past year
Yes 10 (44) 6 (35) 1.4 (0.4 - 5.1) 0.85
No 13 (56) 11 (65) Reference
Type of wildlife contact in past year
Ruminant 15 (65) 8 (47) 2.1 (0.6 – 7.6) 0.41
Non-ruminant 8 (35) 9 (53) Reference
Abortion in sheep and goat herd in past year
Yes 52 (84) 32 (55) 4.2 (1.8 - 9.9) 0.001
No 10 (16) 26 (45) Reference
Abortion in cattle herd in past year
Yes 1 (50) 4 (80) 0.3 (0.01 - 8.6) 0.52
No 1 (50) 1 (20) Reference
Introduction of new animals into sheep and goat herd in past year
Yes 37 (60) 9 (16) 8.1 (3.4 – 19.3) <0.001
No 25 (40) 49 (84) Reference
Lend male animals to other sheep and goat herds for breeding in past year
Yes 21 (34) 9 (16) 2.8 (1.2 - 6.8) 0.04
No 41 (66) 49 (84) Reference
Seek veterinary services in past year
Yes 15 (24) 27 (47) 0.4 (0.2 - 0.8) 0.02
No 47 (47) 31 (53) Reference
Herd size
<100 21 (34) 14 (24) 1.6 (0.7 – 3.6) 0.33
64
*POR=Prevalence odds ratio
≥100 41 (64) 44 (76) Reference

4.2.2 Multivariable analysis to determine independent factors associated with


Brucella herd sero-positivity

The multivariable logistic regression model revealed that introduction of new animals
into the herd [adjusted odd ratio (aOR) = 8.0; p-value <0.0001), experiencing abortion in
sheep and goats herd in past year (aOR = 3.4; 95%; p-value=0.01) and seeking of
veterinary services (aOR = 0.3; 95% CI: 0.1 to 0.8) were the independent factors
associated with sheep and goats herds sero-positivity to Brucella antigens, with
introduction of new animals into the herd and experiencing abortion in sheep and goats
herds in past year being risk factors and seeking veterinary services/advice in past year
being protective factors (Table 4.4).

Table 4.4: Multivariable Logistic Regression Analysis of the Variables associated


with Herd-level Sero-positivity for Brucella spps in Sheep and Goats, Garissa
County, 2013

(Positive herds n=62; Negative herds n=58)

Variable Odds Z-
95% CI Coefficient S.E. P-Value
Characteristics Ratio** Statistic
Introduction of new
animals into sheep and
8.0 3.1- 20.7 2.08 0.486 4.28 <0.0001
goats herd in past year
(Yes/No)
Experience abortion in
sheep and goats herds 3.4 1.3 – 8.9 1.23 0.485 2.54 0.01
in past year (Yes/No)
Seeking veterinary
advice/services in past 0.3 0.1 – 0.8 -1.20 0.471 -2.55 0.01
year (Yes/No)

65
**aOR=adjusted odds ratio

4.3 Determination of Knowledge, Attitude and Practices towards Brucellosis among


pastoralists in Garissa County
4.3.1 Demographic characteristics of study participants

A total of 120 participants were interviewed to assess their knowledge, attitude and
practices towards Brucellosis. The median age of the study participants was 16 years
(Range: 15 – 70 years), with 102 (85%) aged below 35 years. There were 90 (75%)
males; and 92 (77%) of all study participants had no formal education. In regard to
primary role of the study participants in management of the herd, 58 (48%) were herd
owners, 38 (32%) were herders and 24 (20%) were mostly involved in milking animals.
Eighty-three (69%) of the participants were married and 37 (31%) were single (Table
4.5).

Table 4.5: Distribution of Socio-demographic characteristics of study participants,


Garissa County-2013 (n=120)

Variable Frequency n (%)


Age group
15-24 53 (44)
25-34 49 (41)
35-44 14 (12)
>45 4 (3)
Gender
Male 90 (75)
Female 30 (25)
Education level
No formal education 92 (77)
At least primary education 28 (23)
Primary role in Herd
Herd owner 58 (48)
Herding 38 (32)
Milking 24 (20)
Marital status
Married 83 (69)
66
Single 37 (31)

4.3.2 Knowledge of pastoralists toward Brucellosis

4.3.2.1 Awareness and cause of Brucellosis in animals and Humans

Among the study participants, 95 (79%) had heard of Brucellosis and the local name for
Brucellosis in Somali was “Diis”. Among those who had heard of Brucellosis, 17 (18%)
mentioned germs/bacteria as the cause of Brucellosis in both animals and humans
(Figure 4.3). Of the source of information about Brucellosis, 44 (46%) heard of it
through community health workers, 19 (20%) from a family member, 19 (20%) religious
leaders, eight (8%) from veterinary staff and five (5%) from local FM stations.

Figure 4.2: Distribution of study participants’ responses on causes of Brucellosis,


Garissa County, 2013 (n=95)

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4.3.2.2 Knowledge of respondents’about the animals affected and signs/symptoms
of Brucellosis in animals

In regard to animal species affected by Brucellosis, 62 (65%) mentioned goats, 47 (49%)


camels, 45 (47%) cattle and 32 (34%) sheep. Fifty-six (59%) of respondents mentioned
abortion as most common sign, 21 (22%) mentioned retained placenta, 20 (21%)
swollen joints or hygroma and 11 (12%) mentioned mastitis or swollen udder and teats
(Table 4.6).

Table 4.6: Distribution of responses of participants’ on animal species affected by


Brucellosis and signs and symptoms of Brucellosis in animals (n=95)

Frequency

Variable n (%)
Animal species affected
Goats 62 (65)
Camels 47 (49)
Cattle 45 (47)
Sheep 32 (34)
Signs and symptoms of Brucellosis**
Abortions 56 (59)
Retained Placenta 21 (22)
Swollen joints 20 (21)
Mastitis 11 (12)

** Respondents were allowed to choose multiple responses

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4.3.2.3 Knowledge of respondents on modes of transmission of Brucellosis to
humans

A summary of information about the knowledge of respondents about mode of


transmission of Brucellosis to humans is in Figure 4.3. Concerning Brucellosis being
zoonotic disease, only 27 (28%) of the respondents knew of this. Among the respondents
who knew that Brucellosis is a zoonotic disease, 22 (82%) mentioned drinking raw milk
as most common mode of transmission of Brucellosis from animals to humans, followed
by eating milk products from raw milk mentioned by 11 (41%) respondents.

69
Figure 4.3: Distribution of respondents’ responses on mode of transmission of
Brucellosis to humans, Garissa County, 2013 (n=27)

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4.3.2.4 Knowledge of respondents on signs/symptoms of Brucellosis in humans

Summary of information on the knowledge of respondents’ on signs and symptoms of


Brucellosis in humans is as shown in Table 4.7. Forty-six (48%) of the respondents had
a family member diagnosed with Brucellosis in the past, 43 (45%) had seen somebody
who is not a family member with Brucellosis and 38 (40%) respondents had been
diagnosed with Brucellosis in the past. Concerning signs and symptoms, 71 (75%) of
respondents mentioned fever, 45 (47%) chills, 37 (47%) night sweats, 14 (15%) painful
scrotum in men and 7 (8%) spontaneous abortion in women.

Variable Frequency n (%)


Brucellosis occurrence
Family member diagnosed with Brucellosis in the past 46 (48)
Seen person not family diagnosed with Brucellosis 43 (45)
Self was diagnosed with Brucellosis in the past 38 (40)
Signs and symptoms
Fever 71 (75)
Loss of appetite 56 (59)
Joint pain 48 (51)
Chills 45 (47)
Headache 38 (40)
Night sweats 37 (39)
Back pain 33 (35)
Fatigue 29 (31)
Malaise 29 (31)
Vomiting 14 (15)
Painful scrotum in men 14 (15)
Diarrhea 12 (13)
Blurred vision 9 (10)
Miscarriage in women 7 (8)
Nausea 5 (5)

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Table 4.7: Responses of respondents’ on signs and symptoms of Brucellosis in
humans, Garissa County, 2013 (n=95)

4.3.3 Respondents’ attitude towards Brucellosis

A summary of information about the attitude of study participants about Brucellosis is


summarized in Table 4.8. A total of 63 (67%) of the respondents thought that
Brucellosis is a serious disease in animals whereas 61 (64%) thought that Brucellosis is
a serious disease in human. Only 13 (14%) thought that Brucellosis can be prevented in
animals of which six (46%) mentioned vaccination. In regard to attitude and perception
of respondents when they suspect they have Brucellosis, 33 (35%) thought that
Brucellosis in human can be treated/cured of which 14 (42%) mentioned visiting health
facility. When encountered with an aborting animal in the herd, 83 (87%) of the
respondents would do nothing.

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Table 4.8: Distribution of respondents’ responses on attitude and perceptions
towards Brucellosis, Garissa County, 2013 (n=95)

Attitudes and perceptions Frequency n (%)


Attitude and Perception on Brucellosis seriousness
Serious Disease in Animals 64 (67)
Serious Disease in Humans 61 (64)
Attitude towards Brucellosis prevention in animals
Brucellosis can be prevented in animals 13 (14)
Prevention by vaccination 6 (46)
Prevention by contacting veterinary office 4 (31)
Prevention by isolation of sick and aborting animals 3 (23)
Attitude when suspecting human Brucellosis
Brucellosis can be cured in humans 33 (35)
Visit health facility 14 (42)
Praying 8 (24)
Consuming herbal medicine 6 (18)
Visit local chemist and purchase medicine 5 (15)
Attitude towards aborting animals
Do nothing 44 (46)
Treat aborting animals with antibiotics 16 (17)
Sell 11 (12)
Inform veterinary office 10 (11)
Isolate 8 (8)
Slaughter 5 (5)

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4.3.4 Respondents’ practices towards Brucellosis

Respondents’ responses regarding practices towards Brucellosis are as shown in Table


4.9. A total of 91 (96%) of the respondents consumed raw milk in the past year and 34
(36%) assisted an animal during parturition process, abortion or removal of retained
placenta and none used gloves during the process. All (100%) disposed fetal material by
dumping.

Table 4.9: Distribution of respondents’ responses on practices towards Brucellosis,


Garissa County, 2013 (n=95)

Practices of respondents Frequency n (%)

Consumption of raw milk 91 (96)

Participation in slaughtering/butchering an animal 72 (76)

Assisted an animal during birthing/abortion/removal of 34 (36)


retained placenta
Used gloves when assisting during abortions and 0 (0)
parturitions
Disposal of fetal materials

Dumping 34 (100)

Burning 5 (15)

Burying 0 (0)

Consumption of raw milk products 13 (14)

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4.3.5 Respondents’ requirement of more Information on Brucellosis

A total of 92 (97%) of the respondents believed that they were not sufficiently informed
about Brucellosis and required more information. The most favored mode of receiving
information on Brucellosis was through the local FM radio stations mentioned by 36
(39%) of respondents, 23 (25%) favored religious leaders, 18 (20%) local community
meetings (barazas) and 15 (16%) community health workers/community animal health
workers.

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CHAPTER FIVE

DISCUSSIONS, CONCLUSSION AND RECOMMEDATIONS

5.1 Discussion
5.1.1 Highlight of major findings of the study

This study reports the serological analysis of exposure to Brucella spps in sheep and
goats and determination of factors associated with sero-positivity from a population
based sample of sheep and goats herds in Garissa district (currently Garissa Township
and Balambala sub-counties). The study estimated the true sero-prevalence of
Brucellosis in sheep and goats both at the individual animal and herd level incorporating
the sensitivity and specificity of the diagnostic tests used and the uncertainties in these
tests. The result of this study does confirm that there is very high level of probability that
the sheep and goats herds in Garissa could have been exposed to Brucella spps hence
enhanced chances of zoonotic transmission to humans and exposure to other livestock
species like cattle and camels which are also susceptible to Brucellosis and are also
reared in this setup. The study also highlights the need for adjusting for the sensitivity
and specificity of serological tests used in order to make reliable interpretation of
serological surveys, since lack of adjustment based on sensitivity and specificity could
lead to unbiased estimates of sero-prevalence resulting into significantly different
estimates. Seroprevalence of Brucellosis in sheep and goats in this set up varied by
abortion status of the herd and introduction of new animals into the herd which were risk
factors and seeking veterinary care which was protective factor. The study also
examined the farmers’ knowledge, attitudes and practices towards Brucellosis and
highlights the knowledge gaps that exist among pastoralists in the study area towards
Brucellosis as well as the poor attitude and practices that put them at risk of getting
infected with Brucellosis.

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5.1.2 Sero-prevalence of Brucellosis in sheep and goats

Adjusted Brucellosis sero-prevalence in the sheep and goats herds in Garissa County
appears to be high at 20.0% individual animal level and 65.8% at herd-level. This
prevalence varied between the sheep and goats. Reports from literature suggest a very
variable small ruminant Brucellosis sero-prevalence at individual and herd-level in
Kenya across various African countries. Estimates include animal-level sero-prevalence
of between 0.4% and 9.7%, and 15% to 45% herd-level sero-prevalence in Ethiopia
(Megersa et al., 2011; Mohammed et al., 2015; Tsehay et al., 2014; Tsegay et al., 2015;
Dabassa et al., 2013; Yesuf et al., 2012; Deddefo et al., 2015). In Niger, between 0.4%
and 3.6% individual animal-level and 17.8% herd-level sero-prevalence have been found
(Boukary et al., 2013); In Egypt between 0.44% and 12.1% individual animal-level
(Hegazy et al., 2011a and Hegagazy et al., 2011b); In Nigeria individual animal level of
3.3% in sheep and 4.5% in goats (Brisibe et al., 1996) have been reported; In Sudan
between 0.9% and 22% individual animal-level (Gumaa et al., 2014; Abdallah et al.,
2015; Omer et al., 2007) and in Uganda, 4% individual animal-level and 13% herd-level
sero-prevalence in goats have been reported (Kabagambe et al., 2001). In Kenya, studies
done in Kiambu, Kajiado and Marsabit Counties estimated individual animal-level sero-
prevalence ranging from 1.3% to 16.1% and herd-level sero-prevalence ranging from
5.6% to 68% (Osoro et al., 2015). Another study done in Baringo County estimated an
individual animal sero-prevalence of 13.04% in goats and 8.23% in sheep (Kosgei et al.,
2015). These variations in estimates of sero-prevalence of Brucellosis in small ruminants
may be largely attributed to the varied animal husbandry and production systems,
various agro-ecological zones in which the studies were carried out, the sampling
methodology employed and diagnostic tests. These factors have been shown to
contribute to variation in the obtained sero-prevalence of Brucellosis infection in
livestock among different researchers (Mangen et al., 2002; McDermott and Arimi,
2002; Nielsen, 2002; Díaz-Aparicio et al., 2002). Another important issue is the
difference in sensitivity and specificity of serological tests used for screening. This

77
factor contributes to the variability of results among researchers (Nielsen, 2002). The
RBPT used for screening individual animals in this study is a cheap, rapid and highly
sensitive test (OIE 2009a). However, its specificity is low because the smooth
lipopolysaccharides of the Brucella antigen can cross-react with antibodies from closely
related Gram-negative bacteria such as Yersinia enterocolitica, Escherichia coli,
Salmonella spps and Sternotrophomonas maltophilia as well as antibodies produced by
B. abortus S19 vaccine (Nielsen, 2002). The study also found varied differences in sero-
prevalence of Brucellosis between sheep and goats. This could be attributed to the
greater susceptibility of goats to Brucella infection and also excreting the organism for
longer periods compared to the sheep (Tsehay, et al., 2014).

5.1.3 Herd-level factors associated with Brucellosis sero-positivity in sheep and


goats

The introduction of new animals into the sheep and goats herd from unscreened herds
was a major risk factor observed in this study. These animals are usually introduced into
the herd as replacement stocks by the pastoralists. This result is in agreement with
several authors who found out that the introduction of animals from non-free Brucellosis
herds or from herds of which their Brucellosis status was unknown was a major factor
associated with Brucellosis both in sheep and goat herds and in cattle (Kabagambe et al.,
2001, Refai, 2002; Coelho et al., 2008; Bamaiyi et al., 2014; Sharifi et al., 2012; Sharifi
et al., 2014). Other studies suggest that the introduction of infected animals can lead to
an increase in the individual level prevalence due to the fact that the longer they are in
contact with rest of the flock, the higher the risk of spread would be (Corbel, 2006;
Rahman et al., 2013). Practices that involve movement of animals between herds have
also been found likely to be risky. Evidence exists that one of the main reasons for the
inefficiency of most Brucellosis control campaign is the lack of control in the movement
of animals and one suggested key prevention measure is to avoid introduction of
infected animals by maintaining a completely closed herd or by carefully screening

78
purchased animals before introducing them into the herd, a practice that is very rare in
pastoral communities (Kabagambe et al., 2001, Refai, 2002; Nielsen et al., 1990).

Presence of female animals which had aborted in the herd was found to be significantly
associated with herd sero-positivity. Abortion in livestock represents the major
complaint attributed to Brucella infections (Kabagambe et al., 2001; Muma et al.,
2007a; Muma et al., 2007b, Musa et al., 2008; Schelling et al., 2003). Females infected
with Brucellosis are known to excrete high concentrations of the organism in their milk,
placental membranes and aborted fetuses. Such females have been reported to continue
shedding the organisms for several months (Radostits et al., 2007). This results into
environmental contamination and as a result there is enhanced high risk of transmission
of the pathogen between animals of the same herd as well as other herds during free
mixing in grazing and watering places. Zoonotic transmission to humans through direct
contact with contaminated material such as fetal membranes and aborted fetuses is also
enhanced. This is supported by the fact that majority of the respondents’ interviewed
reported assisting animals during abortions and birthing processes and handling fetal
materials without any protective clothing putting them at risk of coming down with the
disease should such materials be contaminated with Brucella organisms. Similarly the
owners reported consuming unpasteurized milk and milk products and just dumping
aborted fetuses and fetal membranes, practices which clearly shows their ignorance of
the transmision of the disease (Obonyo & Gufu; 2015).

Lending male animals to other herds for breeding purposes and seeking veterinary
services in the past year were also factors that were significantly associated with
Brucellosis herd sero-positivity with the former being a risk factor and the latter a
protective factor. Lending of male animals for breeding has been reported by other
authors to be a risk factor associated with Brucella sero-positivity in animals (Al-Majali
et al., 2007; Reviriego et al., 2000). Although venereal route is not considered an
important route for Brucellosis transmission in small ruminants under natural conditions,

79
practices that involve movement of animals between flocks are considered risky due to
potential of mechanical transmission (Radostits et al., 2007; Benkirane, 2006).

80
Though not significant in the final multivariable model, contact between herds was
found as a significant risk factor at the bivariate analysis. This could be attributed to the
nomadic pastoral lifestyle of the community where there is frequent migration of
pastoral herds. Considering the contagious nature of Brucella spps, sharing common
grazing land and drinking water places facilitate transmission of Brucellosis as well as
other diseases between potentially infected herds and clean herds (Megersa et al., 2011;
Schelling et al., 2003; Smits, 2014).

5.1.4 Knowledge Attitude and Practices towards Brucellosis

The KAPs survey showed that Brucellosis is known by the general community in the
present study area, since more than three quarters of the study respondents had heard of
Brucellosis. This is similar to findings of previous studies done in Uganda among
pastoral communities living along Lake Mburo; in Egypt among cattle and Buffalo
farmers in a village in Nile Delta region and among small ruminant farmers in the peri-
urban areas of Dushanbe Tajikistan in which 99.3%, 83.2% and 57% of the respondents’
had heard of Brucellosis (Kansiime et al., 2014; Holt et al., 2013; Grahn, 2013).

However, the awareness of Brucellosis among study participants in Uganda and Egypt
were higher compared to our study but that in Tajikistan was lower. In contrast to this
finding, a study done among small-scale dairy farmers in an urban and peri-urban area of
Tajikistan and another one done among urban and peri-urban dairy and non-dairy
farming households in Kenya found that most respondents had not heard of Brucellosis.
In the Kenyan study done in Kiambu County, 30% of dairy respondents and 22% of
non-dairy respondents knew of the existence of Brucellosis whereas in Tajikistan 85%
of the respondents had never heard of Brucellosis (Lindahl et al., 2015; Kange’the et al.,
2007). Perhaps an explanation as to why the pastoral communities are more aware of
Brucellosis compared to farmers in urban or peri-urban areas could be due to their close
proximity and interaction with animals resulting into in-built indigenous knowledge over
years which is subsequently passed down from one generation to the next. Despite a
81
higher proportion of the study participants had heard about Brucellosis, majority had
little or no knowledge about the cause of the disease. Less than a fifth of the participants
correctly mentioned germ/bacteria as cause of Brucellosis. Poor knowledge regarding
etiology of Brucellosis could negatively impact on respondents’ preventive and control
methods of Brucellosis in both humans and animals due to misconception on the cause
hence need to enhance public health education in this set up.

The main sources of information on Brucellosis in this study area was community health
workers (CHWs) followed by family members. Contrary to this finding, the study in
Uganda and the two studies in Tajikistan found main source of information to be from
friends/relatives (Kansiime et al., 2014; Grahn, 2013; Lindahl et al., 2015). Few
participants in the current study mentioned mass media (radio/TV) as a source of
information about Brucellosis, which was similar to the studies in Uganda and
Tajikistan. These findings implies the powerful role the community health volunteers
play in terms of relaying important health messages to nomadic pastoralists in this area
who in most circumstances have challenges in accessing basic health care services.
Deliberate moves should therefore be undertaken to incorporate the two in all aspects of
health care education for the pastoralists.

Based on results of this study, the respondents’ had basic knowledge about the animal
species affected and signs/symptoms of Brucellosis in animals. In this regard, about two
thirds mentioned goats, close to a half sheep and cattle, and majority were not aware that
camels could be affected. This findings contrast with the findings of studies in Tajikistan
(Lindahl et al., 2015) where 82% of respondents knew that cattle, sheep and goats could
be affected and the study in Egypt (Holt et al., 2011) in which 98.1% mentioned cattle,
86% sheep and 85% goats. However, the current study was fairly in agreement with
another study in Tajikistan (Grahn, 2013) in which two thirds mentioned that all animals
could be affected. With regards to clinical signs of Brucellosis in animals, more than
half of the respondents mentioned abortion as the major clinical sign. This finding was
in agreement with findings of a study done among pastoralists in Kaduna state in Nigeria
82
and the study in Egypt in which 94.4% and 59.5% of respondents mentioned abortion as
the major clinical sign (Holt et al., 2011; Buhari et al., 2015). However the current study
finding was different from that done in Tajikistan where only 11% of respondents’
mentioned abortion as a clinical signs of Brucellosis in animals (Grahn, 2013).
Knowledge of the animal species affected and signs/symptoms of Brucellosis in animals
are crucial because it positively impacts on farmers’ practices towards prevention and
control measures of Brucellosis in both animals and humans.

In this study, majority of the study participants did not know that Brucellosis is a
zoonotic disease, findings which were similar to those in previous studies conducted in
Ghana and Nigeria which found very low awareness of zoonotic nature of Brucellosis
(Buhari et al., 2015; Addo et al; 2011). Among those who were aware of the zoonotic
nature of Brucellosis, consumption of raw milk and raw milk products was the most
cited mode of transmission of Brucellosis from animals to humans. The respondents’
response regarding consumption of milk as a mode of transmission was comparable to
findings in Egypt and Uganda (Kansiime et al., 2014; Holt et al., 2011). However in the
current study, the respondents’ had low awareness on other modes of transmission such
as handling of aborted fetuses and fetal membranes, consumption of raw or undercooked
meat, assisting animals during parturition and slaughtering animals; most of which have
been identified in many studies as major risk factors for transmission of Brucellosis from
animals to humans (Kozukeev et al., 2003; Earhart et al., 2009). Such low knowledge on
mode of transmission of Brucellosis from animals to humans have been documented
elsewhere (Grahn, 2013; Buhari et al., 2015; Addo et al., 2011).

In the current study, the majority of the study participants identified fever, joint pains
and muscle pains in that order as the major signs and symptoms of Brucellosis. This was
consistent with the findings of a previous study in Kyrgyzstan where fever and joint pain
(locally known as “Tajik”) were mentioned as main signs and symptoms of Brucellosis
in humans (Grahn, 2013) as well as a study in Nigeria where all respondents knew signs
and symptoms of Brucellosis in humans (Adesokan et al., 2013). However, the finding
83
of the current study is different from the results of previous studies conducted in other
parts of Nigeria and in Ghana (Buhari et al., 2015; Addo et al., 2011) where almost all
participants were not aware of signs and symptoms of Brucellosis in humans. The
respondents’ basic knowledge about the signs and symptoms of Brucellosis in humans
could have significant impact if the community knowledge is enhanced thus reducing
diagnosis and treatment delay which in the long run will prevent sequelae and prolonged
human suffering.

The present study showed that a considerable proportion of the study respondents
perceived that Brucellosis was a serious disease in both animals and humans. However,
despite this high perception of risk, most respondents’ had poor attitudes towards
prevention of Brucellosis in animals and treatment of Brucellosis in suspected humans.
Regarding respondents’ opinion on actions most would take when confronted with an
aborting animal in their herd, majority would do nothing about it whereas others would
attempt treating the animal with antibiotics or sell the animal. Very few mentioned
isolation of the animal or seeking veterinary services. Failure to isolate suspected
animals has been cited as one of the major risk factors for transmission of Brucellosis
within and between herds as susceptible animals can become infected through contact
with infected animals aborted tissues or consumption of pasture or water contaminated
with aborted materials (Laing and Wagner, 1988). Frequent migration of pastoralists
with their animals increases the chances of different herds coming into contact with
other potentially infected herds thus spreading diseases (Megersa et al., 2011; Boukary
et al., 2013). This is more important when considering the high levels of infectiousness
of Brucellosis making practices such as sharing grazing land and drinking water points
by pastoral communities a major transmission pathway of Brucellosis between different
herds (Makita et al., 2011; Mekonnen et al., 2010).

The study participants engaged in risky practices that could expose them to infection
with Brucellosis. Nearly all respondents consumed raw milk, about three quarter assisted
animals during abortions or parturition and handled aborted materials/fetal membranes
84
and a third participated in slaughtering or butchering an animal. Of those who assisted
aborting animals, three quarter dumped the aborted materials and none used any
protective clothing. Such risky practices have been shown to be important risk factors
for Brucellosis transmission to human (Kozukeev et al., 2003; Earhart et al 2009; Sofian
et al., 2008; Kiambi, 2014; Regassa et al., 2009). Female animals infected with Brucella
spps excrete high concentrations of the organism in their milk, placental membranes and
aborted fetuses (Radostits et al., 2007; Laing and Wagner, 1988). Goats have also been
shown to have prolonged secretion of Brucella organisms in milk compared to sheep
(Poester & Santos, 2013). Furthermore, Brucella spps have been shown to survive in
aborted fetuses, manure and water for periods of up to 150 to 240 days (Saegerman et
al., 2010). Therefore, there is a high risk of transmission of the pathogen between
animals and from animals to humans through direct contact with contaminated materials
such as fetal membranes, aborted fetuses, manure and other animal products.
Introduction of new animals into the herd without quarantine and borrowing or lending
breeding males to other farmers or even taking a female to be served at a neighbor’s
farm have been identified as major risk factors for transmission of Brucellosis within
and between herds as shown in studies in several countries (Boukary et al., 2013;
Kabagambe et al., 2001; Muma et al., 2007b, Al-Majali et al., 2009; Chand & Chhabra,
2013, Chiebao et al., 2013; Patel et al., 2014).

5.2 Limitations of the study

1. Study focused on small ruminants’ Brucellosis only and left out other livestock
species: cattle and camels deemed to be susceptible to Brucellosis. The study
also did not study Brucellosis in humans to correlate findings in the animals.
2. The present study did not attempt culture of the Brucella organisms and therefore
was not able to identify the various species and biovars of Brucella orgaisms
circulating in the sheep and goats
3. The present was also not able to evaluate the socio-economic impact of the
disease in humans and in limiting livestock production in the study area.
85
5.3 Conclusions and Recommendations
5.3.1 Conclusions

1. The present study confirms a considerably high prevalence of Brucellosis both at


individual animal and herd level in sheep and goats. Therefore there is enhanced
potential for zoonotic transmission and exposure to other susceptible livestock
species that are reared in the region.
2. The introduction of new non-quarantined animals with unknown Brucellosis
status into the herd, occurrence of abortions within the herd and lending of male
animals for breeding purposes were found to be the major risk factors for the
spread of the disease within and between herds.
3. In addition, the study showed that the community had some knowledge towards
Brucellosis but attitude and practices were poor. At present, there is no officially
coordinated program for control of Brucellosis in Kenya. Understanding of the
knowledge, perceptions and practices have been defined as important pillars
regarding the feasibility and the acceptability of potential measures that might be
instituted. Enhanced public health education on the cause, symptoms and mode
of transmission of Brucellosis would be important towards the prevention and
control of Brucellosis in the present study area. This can be achieved by targeted
messages in local FM radios and integrating the community health volunteers in
control and prevention efforts.

5.3.2 Recommendations

1. Necessary measures should be taken by the national and County governments


involved in health of livestock and humans in the region to prevent the
transmission of Brucellosis to the human population and as well as transmission
of Brucellosis to other livestock species. There is therefore need for enhanced
public health education as a prevention and control strategy for Brucellosis in
both humans and animals in this region.
86
2. Due to nomadic pastoral lifestyle of the community, control of Brucellosis in
livestock is quite challenging not only because of the number and complexity of
risk factors involved, but also because the risk factors that are tightly linked and
often inherent to the livestock production practices. The above factors when
combined with the close interaction between the people and their livestock
coupled with the socio-cultural lifestyle and poor knowledge attitude and
practices poses a serious public health concern and has been known to enhance
transmission of Brucellosis to the human population. These reasons make control
of Brucellosis both in livestock and humans challenging. At present, there is no
officially coordinated program for control of Brucellosis in Kenya. However the
greater horn of Africa where Kenya belongs has made greater strides towards
controlling of the disease. The African Union Inter-African Bureau on Animal
Resources (AU-IBAR) came up with a strategy dubbed “Standard Methods and
Procedures (SMPs) for control of Brucellosis in the Greater Horn of Africa”
which outlines minimum standards, methods and procedures on various subject
areas of surveillance, laboratory procedures and disease control that must be met
for harmonized regional control of the disease (AU-IBAR, 2014). If fully
adopted, these SMPs will go a long way towards control of livestock Brucellosis
in the greater horn of Africa. However for the full potential of the SMPs to be
realized, there is need to evaluate attitudes of communities involved and their
roles need to be clearly defined in the various control strategies especially
regarding the feasibility and the acceptability of potential measures. Measures
including selective vaccination programs combined with the slaughtering of
known infected animals (test and slaughter) as well as testing animals newly
introduced into the herd can be considered. This suggested control measures can
only be effective with involvement and buy –in of the community including
integrating the CAHWs in the control strategies.

87
3. Other studies to be conducted in the region should include other species of
livestock which are also susceptible to Brucellosis as well as humans in a linked
animal-human study; attempt culture of the Brucella organisms to identify the
various species and biovars of Brucella spps circulating in both livestock and
human in this region and evaluate the socio-economic impact of the disease in
humans and iit effect on limiting livestock production.
4. However for effective control of Brucellosis in the present study area, an
integrated approach should be promoted that takes into account the relationship
between humans, animals and environment in the context of ‘‘One health
approach’’.

88
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107
APPENDICES

Appendix 1: Questionnaire in English

The questionnaire was administered to the respondents with assistance of an


interpreter who speaks the local Somali language. The interpreter was trained
prior to the beginning of the investigation.

Questionnaire No: ………………

Title: Sero-prevalence and Factors associated with Brucellosis in Goats and Sheep,
Garissa County, 2013

Name of the farmer: .............................................................................................................

Name of interviewer: ...........................................................................................................

Date of interview: ................................................................................................................

Time of interview: ...............................................................................................................

Socio demographic data of respondent

Age in years:
.........................................................................................................................

Gender:
.................................................................................................................................

County:
..................................................................................................................................

108
Division:
................................................................................................................................

Location:
................................................................................................................................

Sub-location:
.........................................................................................................................

Village:
.................................................................................................................................

GPS point:
Latitude.................................Longitude...................................Elevation.............................
..

Highest level of formal attained education:

0=No formal education

1 = Primary

2 = Secondary

3 = College/graduate

5 = Other

Primary occupation:

1 = Employed full time on farm

2 =Employed part time on farm

109
3 =Self-employed off farm

4 =Employed off farm - agriculture

5 = Employed off farm

6= Other

Primary role:

1=Milking

2=Slaughtering

3 =Butchering

4= Cleaning barns

5= assisting in animal delivery

6= herding/

7=Feeding animals

8=Other

Laboratory information

What is the number of sheep and goats in the herd:


……………Sheep…………………Goats

How many sheep and goats are sampled:


………………Sheep………………………….Goats

110
How many are positive on RBPT:
………………………Sheep…………………………Goats

From those positive on RBPT, how many are positive on CFT:


…………Sheep………..Goats

Overall herd status:

Positive

Negative

Factors associated with Brucellosis in sheep and goats herds

1. Do you own other animals apart from sheep and goats? (If No skip to question 4)

1=Yes

2=No

2. If yes above, which ones and how many are they?

Cattle………………..

Camels……………….

Donkey………………

Poultry……………….

Others (specify)……………………………………….

3. How do you raise your animals?

111
1=Raise them together

2=Raise them separately

4. Do you own all the sheep and goats in the herds? (If Yes, skip to question 6)

1=Yes

2=No

5. If No above, who else owns the animals in this herd?

1=Neighbor

2=Friend

3=Family members

6. Do you have a calving pen for your herd?

1=Yes

2=No

7. Did your herd come into contact with other herds during grazing or watering in
year?

1=Yes

2=No

8. Did your animals come into contact with wild animals in the past year?

1=Yes

112
2=No

9. If yes, which wild animals?

• Zebra

• Buffalo

• Antelopes

• Waterbuck

• Warthogs

• Others (specify):
………………………………………………………………………….

10. Have you ever found evidence of wildlife abortion in your farm or where you graze
or water your animals?

1=Yes

2=No

11. Did you experience abortions or still births in your sheep and goats herd in the
past year? ( If No skip to question 12)

1=Yes

2=No

If yes above, what was the number of animals aborting?


113
Sheep……………................................

Goats……………................................

12. Did you experience abortions in your other animals herd in the past year? (Skip
if answer to question 1 was No and go to question 13)

1=Yes

2=No

If yes, to above question which other animal(s) aborted and how many were
they?

Cattle ……………………………..

Camels …………………………….

Others……………………………...

13. Did you introduce any new animal(s) into your sheep and goat herd in the past
year through buying, dowry, compensations etc? (If No, skip to question 15)

1=Yes

2=No

If yes above, How many were they?

Sheep …………………………..

Goats …………………………..

14. Was the animal(s) tested for diseases and quarantined before introduction?
114
1=Yes

2=No

15. Did you lend your male animals for breeding to other herds in past year?

1=Yes

2=No

16. Did you vaccinate your animals against infectious diseases in past year? (If No,
skip to question 18)

1=Yes

2=No

17. If yes above, was Brucellosis one of the diseases you vaccinated against?

1=Yes

2=No

18. Did you get veterinary advice on management of your sheep and goats in past
year?

1=Yes

2=No

Farmers Knowledge Attitude and Practices regarding Brucellosis

115
19. Have you heard of the disease called Brucellosis? (If No, questioning ends here)

1=Yes

2=No

20. If yes above, what do you think is the cause of this disease in animals?

1= Food,

2=Water,

3=Wild animals,

4=Sexual

5=Witchcraft

6= Hereditary

7=Don’t know

8=Others (specify) …………………………………………………….

21. In your opinion, which animals are affected by Brucellosis? (Check all that are
mentioned.)

1=Goats

2=Sheep

3=Cattle

4=Camels

116
5=Others (specify)
………………………………………………………………………

22. In your opinion, what are the common signs and symptoms of Brucellosis in
sheep and goats? (Check all that are mentioned.)

1=Abortions

2=Retained placenta

3=Swollen testes (orchitis)

4=Infertility

5=Metritis (Pus from Vulvular area)

6=Arthritis/Hygroma (Swollen joint)

7=Mastitis

8=Others (specify):
…………………………………………………………………….

23. Do you know whether animals can transmit Brucellosis to humans? (If No/don’t
know, skip to question 22)

1=Yes

2=No /don’t know

24. If yes above, how is Brucellosis transmitted from animals to humans? (Check all
that are mentioned.)

117
1=Helping animals to deliver/abort by bear hands

2=Handling fetal tissues/aborted fetuses with bear hands

3=Drinking raw milk

4=Consuming products processed from raw milk

5=Slaughtering animals

6=Milking animals

7=Eating uncooked meat

8=Living with animals

9=Others (specify):
………………………………………………………………………

25. Have you ever been diagnosed with Brucellosis in the past?

1=Yes

2=No /don’t know

26. Have any of your family members been diagnosed with Brucellosis in the past?

1=Yes

2=No /don’t know

118
27. Have you seen any other person with Brucellosis in the past apart from your
family member?

1=Yes

2=No/don’t know

28. In your opinion, what are the signs/symptoms of Brucellosis in humans? (Check
all that are mentioned.)
 Fever  Night sweating
 Headache  Diarrhea
 Chills  Nausea
 Fatigue  Vomiting
 Malaise  Blurred vision
 Loss of appetite  Spontaneous abortion in women
 Joint pain  Painful scrotum in men
 Back pain

29. Where did you first learn about Brucellosis? (Check all that are mentioned.)

1=Newspapers and magazines

2=Radio

3=TV

4=Veterinary officials

5=Brochures, posters and other printed materials

6=Community Animal Health workers

7=Family, friends, neighbors and colleagues

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8=Religious leaders

9=Teachers

10=When I was diagnosed with the disease

11=Other (please explain):


…………………………………………………………..

30. In your opinion, how serious a disease is Brucellosis in animals in your area?
(Check one.)

1=Very serious

2=Somewhat serious

3=Not very serious

31. In your opinion, how serious a disease is Brucellosis in humans in your area?
(Check one.)

1=Very serious

2=somewhat serious

3=Not very serious

32. Have you ever heard how you can prevent Brucellosis in animals?

Yes

No/Don’t know

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33. How do you prevent animals from getting Brucellosis? (Please check all that are
mentioned.)

Vaccinations

1=Isolation of sick or aborting animals from the rest of the herd

2=Reporting to veterinary authority

3=Don’t know

4=Other (please explain)


………….………………………..............................................

34. Can Brucellosis be cured in humans? (If No, skip to question 36)

1=Yes

2=No/don’t know

35. If yes above, how can someone with Brucellosis be cured? (Check all that are
mentioned.)

1=Herbal remedies

2=Home rest without medicine

3=Praying

4=Specific drugs given by health facility

5=Do not know

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6=Others (specify):
………………………………………………………………………

36. What actions would you take if you were found out or suspected that your
animals have Brucellosis? (Check all that are mentioned.)

1=Slaughter and consume the meat

2=Sell animal to a neighbor

3=Sell animal to a butcher

4=Sell animal in a market

5=Call a veterinarian

6=Do nothing

7=Others (specify)
…………………………………………………………………….

37. What action(s) would you take as regarding aborting animals in your herd?
(Check all that are mentioned.)

1=Call a veterinarian

2=Self treat them

3=Separate them from the rest of the herd

4=Sell them

5=Slaughter them

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6=Do nothing

7=Others
8=(specify)…………………………………………………………………………
….

38. What would you do if you thought you had symptoms of Brucellosis? (Check all
that apply.)

1=Go to health facility

2=Go to pharmacy

3=Got to traditional healer

4=Go to be prayed for

5=Other (specify):
…………………………………………………………………………

39. Have you assisted sheep and goats or any of your animals during
parturition/abortion/removal of retained placenta in the past year? (If No/don’t
know skip to question 42)

1=Yes

2=No/don’t know

40. Did you use any protective gloves or masks when assisting with the parturition or
abortion of animals or whilst handling placentas and aborted fetuses?

1=Yes

123
2=No/don’t know

41. How did you dispose of the aborted fetuses and placentas?

1=Burning

2=Dumping

3=Burying

42. Did you participate in slaughtering/butchering of animals in past year?

1=Yes

2=No/don’t know

43. Did you participate in processing of raw milk products in past year?

1=Yes

2=No/don’t know

44. Did you consume raw milk or milk products made of raw milk in past year?

1=Yes

2=No/don’t know

45. Do you feel well informed about Brucellosis?

1=Yes

2=No

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46. Do you wish you could get more information about Brucellosis?

1=Yes

2=No

47. What are the sources of information that you think can most effectively reach
people like you with information on Brucellosis? (Please choose the three most
effective sources.)

1=Newspapers and magazines

2=Radio

3=TV

4=Billboards

5=Brochures, posters and other printed materials

6=Health workers

7=Family, friends, neighbors and colleagues

8=Religious leaders

8=Teachers

9=Other (please explain):


…………………………………………………………………

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Appendix 2: Consent form in English

Title of study:

Sero-prevalence and associated Factors for Brucellosis in Goats and Sheep, Garissa
County

Introduction:

My name is Mark Odhiambo Obonyo. I am trying to learn more about Brucellosis.


Brucellosis is a zoonotic disease that is of public health importance. It is transmitted
from animals to humans when people get exposed to infected livestock and their
products which may act as a source of infection.

Purpose of study:

Due to the great public health importance of this disease, I am requesting for your
participation in this study whose main objective is to find out how many of your animals
and others in this village and other villages in Garissa County are exposed to this serious
disease, what are the factors associated with transmission or acquisition of this disease
by your animals and also gauge your knowledge, attitude and practice. This is important
for the relevant authorities to find ways of dealing with this disease in this area. You are
being asked to join this study because your herd was picked by chance among other
herds in this area.

Expectations of the study:

If you agree to participate in the study, I wish to test some of your animals to determine
whether they could have been exposed to Brucellosis. If you agree to take part in the
study, a trained laboratory technologist will withdraw 5-10mls of blood (that can fill a
big spoon) from the jugular vein selected animals using a syringe and needle. The blood

126
specimen was transported to Garissa regional veterinary investigation laboratory where I
was test for Brucellosis. I shall then ask you some questions which are written on a
paper on animal husbandry and your knowledge, attitude and practices regarding
Brucellosis. The test results shall be availed as soon as possible to County veterinary
officer of Garissa who shall forward them to you and advice on any necessary control
measures if need be.

Risks:

There are no envisaged risks from participating in this study. However minor bruising
and bleeding may be noticed on the selected sheep and goats during withdrawal of blood
sample.

Benefits:

The results of this study was communicated and disseminated to the people concerned
for them to take action on the recommendations that was come out from the study
results. This was include necessary control measures if need be.

Confidentiality:

Any information obtained from you was kept confidential and used solely for purposes
of this research only. The results of this research may be published in scientific journals
or presented at medical or veterinary meetings, but your identity will not be disclosed.

Compensation:

If you accept to take part in this study, there was no payment for participation.

Alternatives:

127
You have a choice to agree or not to agree to participate in this study. If you agree to
participate in study you are allowed to withdraw from the study at any time if you so
wish without any consequences whatsoever.

Approval of the study:

This study was approved by:

The Kenyatta National Hospital/University of Nairobi-Ethics and Research Committee


(KNH/UON-ERC)

P O BOX 20723-00202, Nairobi, Kenya

Email: uonknh_erc@uonbi.ac.ke

Tel: (254-020) 2726300 ext 44355 for UoN or 726300-9 for KNH

And

Board of Post graduate studies

Jomo Kenyatta University of Agriculture and Technology

P.O. Box 62,000, Juja, Kenya

In case of any further questions or concerns, you can address them to the directors of the
above institutions.

Consent:

I have been fully informed about the study, the risks and benefits of it. I had the
opportunity to ask questions which were satisfactorily answered. I therefore consent to
voluntarily participate in the study.

128
Name of participant………………………………………………………………………

Signature/ thumb print of participant……………………………………………………..

Date………………………………………………………………………………………..

Name of researcher/research assistant……………………………………………………..

Signature……………………………………………….. Date
…………………………….

129
Appendix 3: Translated Questionnaire in Somali Language

LIFAAQA 1: SU’AALO

Cinwaanka : Sero-prevalence and iyo xaqiiqooyinka la xiriira cudurka Diis ee k u


dhaca ariga iyo idaha , degmada Gaarisa ,2013

Magaca wareysi
qaadaha.................................................................................................................

Taariikhda
wareysiga......................................................................................................................

Waqtiga.................................................................................................................................

GPS
point......................................................................................................................................

Qayaas ahaan Inta u dhaxeysa GPS iyo goobta


xoolaha(KMs).....................................................

Warbixinta Sheybaarka Meeqey dhan yihiin xoolaha ari iyo idaba


............................................

Ari iyo ido meeqaa ayaa sample laga qaaday


?............................................................................

Meeqaa RBPT positive noqotay


?.................................................................................................

RBPT positive kuwa ah meeqaa sii ah CFT positive ?


................................................................

130
Muxuu yahay herd-level sero-positivity xoolaha
?.......................................................................

Meeqaa loo badalay herd-level sero-


positivity.............................................................................

XAQIIQOOYIN LA XIRII CUDURKA DIIS EE KU DHACA XOOLAHA SIDA


ARIGA IYO IDAHA

1. Maleedahay xoolo kale markii laga reebo idaha iyo ariga?(hadii jaawantu tahay
maya suaasha 4 ugudub)

Haa……

Maya…………..

Hadii jaawabta suaasha kore haa tahay , meeqaad leedahay waana


xayawaankee/xoolohee?

Lo’o…………….

Geel…………….

Dameero…………………

Dooro………………

Kuwakale(fadlan sheeg)……………….

2. Xoolaha side baad udhaqataa?

Halmeel hal xero……………..

131
Kala xero………….

3. Adi miyaa wada leh ariga iyo idaha meesha joogo?(hadii ee jaabtu tahay haa
suaasha 6-aad ugub

Haa………

Maya………

4. Hadii jaawabtu tahay maya ,yaa kale oo kulaleh xoolaha?

……………………………………………………………………………………………

……………………………………………………………………………………………

……………………………………………………………………………………………

5. Xoolaha maladaaqeen xoolo kale 6-dii bilood ee lasoo dhaafay?

Haa…………………….

Maya…………………

6. Xoolaha mala kulmeen xoolo kale markii ee biyaha cabaayeen?

Haa,………………

Maya……………

7. Xoolaha malakulmeen xayawaanka duurka 6-dii bilood ee soo dhaafay?

Haa ……………………….

Maya………………

132
8. Xoolahaga (idaha iyo ariga) dilan maku aragtay sanadkii lasoo dhaafay?

(Hadii jaawabtu tahay maya ugudub saasha 11)

Haa…………………….

Maya……………

9. Xoolaha aad ku aragtay dilan waa meeqaa?

Idaha……………………………………………………………………

Ariga……………………………

10. Malakulantay dilan xoolohaada kale sanadkii lasoo dhaafay ?(hadii jawaabtu
tahay maya ugudun suaasha 13)

Haa………………..

Maya………………..

11. Hadii jawaabtu suasha kore tahay haa waa xoolahee kuwa dilanka aad ku
aragtay waana meeqaa?

Lo’oda……………………………………………………………………………
………

Geela………………………………………………………………………………
………

Kuwakale…………………………………………………………………………
………

133
12. Maku soo dartay xoolo cusub arigaada iyo idahaaga sanadkii lasoo
dhaafay?(hadii jawaabtu tahay maya ugudub suaasha 17)

Haa……………..

Maya…………….

13. Meeqeey ahaayeen ?

Idaha……………………………………………………………………….

Ariga…………………………………………………………………

14. Xageed ka keentay xooloha?

Suuqa……………

Dariska/jiiranka…………

Xoolodhaqato kale………..

Meelo kale(fadlan sheeg)………….

15. Xooloha cusub mala baaray , gees malooxiray inta aan lagu darin xooloha?

Haa,………….

Maya………….

16. Maudhiibtay doorodaada(mida cagaha waaweyn) xoolo dhaqato kale sii eey
utarmaan sanadkii lasoo dhaafay ?

Haa……………..

134
Maya……………

17. Makatalaashay cudurada dilaaga ah xooloda sanadkii lasoo dhaafay?.(hadii


jawaabtu tahay maya ugudub su’aasha 20)

Haa

Maya

18. Hadii jawaabtu suaahsa kore tahay haa ,Diis kamid ma’aheed kuwa aad
katalaashay xooloha?

Haa

Maya

19. Dhaqtarka xooloha maku waaniyay sanadkii lasoo dhafay sidii aad udhaqi
laheed ariga iyo idaha?

Haa………………….

Maya………….

AQOONTA XOOLALEYDA ,SIDEY U ARKAAN ,QAABKEY ULA


DHAQMAAN CUDURKA DIIS .SU’AALO GUUD IYO KUWA DADKA KU
SAABSAN

20. Ma maqashay cudur la yiraahdo Diis ?( jawaabta hadey maya tahay ,su’aasha
meeshaa ayey ku egtahay )

Haa………..

Maya……………

135
21. hadeey jawaabta haa tahay maxaa xoolaha cudurkaan u keeno ?

Germs………………

Kala dhaxlid……………….

Sixir………………….

Ma aqaano………………..

Kuwa kale ( cadeey)………………………………………

22. Fikirkaada , cudurka Diis xayawaanadee ayuu ku dhacaa ?( dooro inta la


sheegay)

Ari……………

Ido…………………….

Lo’……………………

Geel ……………………

Kuwa kale (
sheeg)…………………………………………………………………..

23. Fikirkaada , calaamadahee lagu garanayaa Diis marku ariga iyo idaha yo (dooro
kulli)

Abortions/dilan………………….

Retained after birth/mandheer kuharto……………………..

Orchitis/sheelo…………………………………….
136
Hygroma……………………….

Metritis……………………….

Arthritis/cudurka jilbaha……………….

Mastitis/cudurka candhada xoolaha kagalo ………………..

Kuwakale (fadlan sheeg):……………….

24. Xoolaha ma u gudbin karaan dadka cudurka Diis?( maya/ma aqaano ubood
su’aasha 34)

Haa

Maya /ma aqaano

25. Hadii jawaabta kore eey haa tahay , sidee? (dooro inta la sheegay)

Ka dhalin xoolo gacmihiisoo banaan

Gacantaada oon galoofis lahayn in aad xayawaan yar oo dhicis ku qabato.

Cabid caano aan la karkarin

Isticmaalid wax yaabo laga hagaajiyay caano aan karkarsaneyn

Xoolo qalid

Xoolo maalid

Hilib aan la Karin cunidiisa

Xoolo la noolaasho

137
Kuwa kale (sheeg): …………………………………………………………

26. Waligaa cudurka Diis makuugu dhacay ?

Haa

Maya / ma aqaano

27. Qof familkaada ka mid ah waligiisa ma qaaday cudurka Diis ka ?

Haa

Maya / ma aqaano

28. Qof aan familkaada ahayn waligaa Diis maku aragtay ?

Haa

Maya / ma aqaano .

29. fikirkaaga , calaamadahee lagu arkaa dadka markuu cudurka Diis ku dhaco ?

Qandho

Madax xanuun

Daal

qarqar

xanuun aan la garaneynin/ caajis

cuntadoo ka istaagta

138
Kalagoosyadoo xanuuno

Dhabar xanuun

Habeen dhidid

Shuban

Lalabo

Matag

Indhaha oo aragtida gaabiyo

Ilmo dilan

Raaya xanuun

30. Marki kugu horeysay Diis inteed ka baratay? ( qor dhamaan inta la sheegay)

Newspapers and magazines

Radio

TV

Dhaqaatiirta xoolaha

Waraaqaha darbiyada lagu dhajiyo ama dadka loo qeybiyo

Shaqaalaha caafimaadka xoolaha ee bulshada dhaxdeeda

Familka , saaxiib , dariska ama qof wax aad wada barataan

139
Hogaamiyayaasha diimaha

Macaliminta

Markuu cudurka igu dhacay .

Kuwa kale ( fadlan sharax) ………………………………………….

31. Fikirkaada , sheeg qatarta uu cudurka Diis uu ku hayo xoolaha eeriyadaada? (


dooro mal mid)

Halis

Halis aan sidaa ubadneyn

Halis ma jirto

32. Fikirkaaga , halis intee la eg ayuu cudurka Diis uu ku hayaa dadka eeriyadada? (
dooro mid )

Halis

Dhaxdhaxaad /halis aan sidaa ubadneyn

Halis ma jirto

33. Sided uga hortagtaa si eeysan xoolaha u qaadin Diis ? ( qor inta la sheegay)

Talaal

Ka takoorid inta kale

War galin dhaqaatiirta xoolaha

140
Ma aqaao

Kuwa kale ( sheeg


)…………………………………………………………………

34. Diis dadka malaga daaweyn karaa? ( jawaabta hadey maya tahay ubood S40)

Haa

Maya /ma aqaano

141
35. Hadey jawaabto haa tahay side loo daaweyn karaa ? (qor inta la sheegay)

Daawo dhaqameed

Nasiino ayadoon daawo la qaadan

Salaad/duco

Daawo eey kuu soo qoreen goobo caafimaad

Ma aqaano

Kuwa kale
(Sheeg)……………………………………………………………………………
……

Sida loo arko iyo hab dhaqanka ku aadan Diis

36. Talaabo noocee ah ayaad qaadan laheyd hadi xoolahaaga laga heli lahaa ama
looga shakiyo iney Diis qabaan ?(Qor kuli waxa la sheegay).

Qal oo isticmaal hilibkooda

Xoolaha ka gad dariska

Kawaanka ka iibi xoolaha

Suuqa ku iibi

Uyeer dhaqtar xayawaan

Wax ha suubin

142
Kuwa kale( xus)
……………………………………………………………………………

37. Talaabo noocee ah ayaad qaadan lahayd oo ku aadan ilma dilanka xoolahaada?(
qor inta la magacaabay)

Dhaqtar xoolo uyeerid

Adiga daaweyso

Xoolaha intooda kale gooni uga bixi

Iibi oo qal

Wax ha suuban

Kuwa kale (
xus)…………………………………………………………………………….43:
Maxaad suubin lahayd hadaad moodo in aad qabto calaamadaha Diis? (qor inta
ku haboon)

Goob caafimaad aadid

Farmashye aadid

Daawo daqameed

Quraan aqris / duco saarid

Kuwa kale ( xus)


…………………………………………………………………………

143
38. Ma caawisay lax ama ri ama xoolahaada qeyb ka mid ah mar eey dhalayeen/
ilmo kasoo dilmeen/ mandheerta ka soo saartay sanadkii hore ?( hadey jawaabta
maya tahay / ma aqaano u bood S47)

Haa

Maya / Ma aqaano

39. Ma isticmaashaa gloves ama masks marka aad mandheerta kasoo jiideysid ama
aad ka dhalineysid ama ilmaha dilanka kasoo jiideysid ?

Haa

Maya / ma aqaano

40. Maxaad ku suubisaa dhiciska soo dilmo iyo mandheerta ?

Gubid

Qashinka ku darid

Duugid

41. Maka qeyb gashay qalid / kawaan geyn xoolo lixdii bil ee lasoo dhaafay ?

Haa

Maya / ma aqaano

42. Maka qeyb qaadatay soo saarid caano aanan karkarsaneyn wax laga suubiyay
lixdii bil ee laga soo gudbay ?

Haa

144
Maya /ma aqaano

43. Caano aan la karkarin ma isticmaashay lixdii bil ee lasoo dhaafay ?

Haa

Maya /ma aqaano

44. Ma isticmaashay waxyaalo laga suubiyay caano aan la karkarin lixdii bil ee ugu
danbeysay

Haa

Maya

Diis , aqoonisteeda iyo ilaha warbaxineed

45. Ma umaleynee in aad si fiican u taqaano Diis?

Haa

Maya

46. Ma rabtaa in warbixin dheeri aad ka hesho Diis ?

Haa

Maya

47. Maxey yihiin ilaha warbixin ee aad is leedahay warbixin fiican ayey dadka
kaheli karaan oo ku saabsan Diis?( sheeg 3 qeyb ee ugu tayada roon).

Newspapers and magazines

145
Radio

TV

Dhaqaatiirta xoolaha

Waraaqaha darbiyada lagu dhajiyo ama dadka loo qeybiyo

Shaqaalaha caafimaadka

Familka , saaxiib , dariska ama qof wax aad wada barataan

Hogaamiyayaasha diimeed

Macaliminta

Kuwa kale (fadlan sharax) ………………………………………….

146
Appendix 4: Translated consent form in Somali Language

LIFAAQA 3: Foomka ogolaanshaha

CINWAANKA BAARISTA

Sero –prevalence iyo xaqiiqooyinka la xiriira ee Diis ka ariga iyo idaha , Degmada
Gaarisa

Hordhac:

Diis wuxuu ka midyahay cudurada loo yaqaano Zoonotic caafimaadka bulshadana


muhiim u ah . xoolaha ayaa u gudbiya bini aadamka marka eey dadka isticmaalaan
xoolo ama waxyaalo laga suubiyay xoolo qabay cudurka .

Ujeedada baaritaanka:

Waxaan kaa codsaneynaa kaqeybqaadahsada baaritaanka ee ujeedkiisu yahay in la


qiimeeyo faafida iyo xaqiiqooyinka la xiriira cudurka Diis ee ku dhacay idaha iyo ariga
ku nool Dagmada Gaarisa .

Waxa laga filanaayo baaristan :

Waxaan rajeyneynaa in xoolihiina qaar aan tijaabino in eey udhiban yihin Diis . hadaad
nagala qeyb qaadato baaritaankan , waxaan dhiig yar ka qaadeynaa xoolaha aan
dooranay kuwaas oon ka baareyno Diis , ka dib waxaan ku weydiineynaa suaalo dhowr
ah oo ku saabsan xanaanada xoolaha iyo aqoonta aad uleedahay, sifooyinka iyo
practiska ku saabsan Diis , baaritaanka natiijadiisa waxaa si dhaqsa leh loogu gudbinaa
dhaqaatiirta xoolaha ee degmada gaarisa kuwaasoo idinka idinsoo gudbin doono idinna
raacinayo waanooyin iyo waxyaalo kale oo loo baahan yahay .

Qatarta :

147
Ma jiraan Qatar la filanayo iney keento ka qeybqaadashada baaritaankam , si kastaba ha
ahaatee dilaac yar ama diig yar inuu ka soo daato ayaa laga yaabaa ariga ama idaha
marka dhiiga tijaabada laga qaadayo.

Faaiidooinka :

Natiijada kasoo baxeyso baaritaankaan waxaa loo gudbinayaa dadka uu quseeyo si eey u
qaataan talaabooyinka ku haboon oo kasoo bixi doono baaritaanka.

Kalsooni:

Warbixin kasta oo dhankaaga ka timaato waa la xifdinayaa waxaa loo isticmaalaaya


dano baaris oo kaliya , natiijada kasoo baxdo baaritaankaan waxaa lagu daabacayaa
scientific journals ama waxaa loo gudbin dhaqtar ama kulan dhaqaatiir xooleed laakin
qofka aad tahay waa la qarinayaa.

148
Fursada aad heysato

Shaqadada barista ah waqtigaad doonto ayaad ka bixi kartaa wax dhibaata ahna ma
jirayaan.

Ogolaanshaha baaritaanka :

Baaritaankaan waxaa ogolaaday gudiga:

The Kenyatta National Hospital/University of Nairobi-Ethics and Research Committee


(KNH/UON-ERC)

P O BOX 20723-00202, Nairobi, Kenya

Email: uonknh_erc@uonbi.ac.ke

Tel: (254-020) 2726300 ext 44355 for UoN or 726300-9 for KNH

149
Iyo

Board of Post graduate studies

Jomo Kenyatta University of Agriculture and Technology

P.O. Box 62,000, Juja, Kenya

Ogolaansho:

Si wacan baa leygu sheegay baaritaanka ,qatarta iyo faaiidooyinka , waxaan heystay
fursad aan suaalo ku weydiin karay kuwaa oo jawaabo aan ku qancay lahaa ,sidaa
darted waxaan ogolahay inaan si voluntarily ah uga qeybqaato baaritaankan

Magaca ka qeyb qaataha


…………………………………………………………………………

Saxiixa / suulka ka qeybqaataha


…………………………………………………………………

Taariikhda
………………………………………………………………………………………..

Magaca baaraha /caawiye bare


…………………………………………………………………..

Saxiixa
…………………………………………………………………………………………...

Taariikhda
……………………………………………………………………………………….

150
Signature………………………………………………..
Date…………………………………

151
Appendix 5: Manuscript on Sero-prevalence and herd level factors associated with
Brucellosis in sheep and goats in Garissa County, 2013.

Manuscript Published in BMC Veterinary

Title: Estimated Brucellosis sero-prevalence at individual animal and flock level in


Sheep and Goats and factors associated with flock level sero-positivity in Garissa
County, Kenya-2013

Running Title: Brucellosis Sero-prevalence and associated factors, Kenya

Author: Mark Obonyo1, 2, 3§ and Gideon Kikuvi2

Author Affiliations:

1
Field Epidemiology and Laboratory Training Program, Ministry of Health, Kenya

2
Jomo Kenyatta University of Agriculture and Technology, School of Public Health

3
Ministry of Agriculture, Livestock and Fisheries, Directorate of Veterinary Services,
Kenya

§
Corresponding author

Email addresses:

MO: mobonyo@feltp.or.ke

GK: kikuvi@yahoo.com

152
Abstract

Background: Brucellosis, a zoonosis of major public health importance, is endemic in


livestock in Kenya. A cross-sectional epidemiological study was carried out to estimate
the sero-prevalence of brucellosis and identify factors associated with flock level sero-
positivity in small ruminants in Garissa County of North Eastern Kenya.

Methods: A total of 2,400 sera from 120 flocks were collected from sheep and goats
which were randomly selected using a multi-stage sampling technique and data on
potential flock level factors were collected from livestock owner or their representative
≥15 years using a pre-tested structured questionnaire. The sera were analyzed using
Rose Bengal Plate Test (RBT) and sero-positive reactors confirmed by Complement
Fixation Test (CFT) using serial interpretation. We considered a sample to be positive
when both tests results were positive and a flock was considered positive when a single
animal within the herd tested positive on both tests. Multivariate logistic regression was
used to investigate for independent factors associated with flock brucellosis sero-
positivity in small ruminants.

Results: The overall sero-prevalence of brucellosis at individual animal-level was


20.0% [(394/2400); (95% CI: 18.2% to 22.0%)]; in goats 24.3% [(290/1471); (95% CI:
21.8% to 27.1%)] and sheep 12.5% [(104/979) (95% CI: 10.2% to 15.2)]. Overall flock-
level sero-prevalence was 65.8% [(62/120) (95% CI: 54.3% to 77.2%)]. Seeking
veterinary services [aOR=0.30 (95% CI: 0.12 to 0.76], introduction of new animals into
the flock [aOR=8.0 (95% CI: 3.09 to 20.70] and experiencing abortions in the flock
[aOR=3.43 (95% CI: 1.33 to 8.88] were independently associated with brucellosis flock
sero-positivity in small ruminants.

Conclusions: The study highlights considerable high sero-prevalence of brucellosis and


factors that contributes for its transmission in small ruminants in Garissa County. This

153
poses potential public health threat associated with zoonotic transmission. We
recommend a one health approach for effective control of brucellosis in this region.

Key words: Brucellosis, Risk factors, Sero-positivity, Kenya, Garissa, Small Ruminants

Background

Brucellosis, more so that caused by B. melitensis remains one of the most


common zoonotic diseases worldwide accounting for an estimated 500,000 human cases
reported annually [1]. The true incidence of human brucellosis worldwide is estimated to
vary from <0.03 to >160 per 100,000 population [1, 2]. The bacteria is classified by the
US Centers for Disease Control and Prevention as a category (B) pathogen which can be
developed into a bio-terrorist agent of mass human destruction with a potential of
aerosol transmission [3].

Brucellosis infection in small ruminants causes heavy economic losses due to


production and reproduction losses resulting from mass abortions, neonatal losses,
reduced fertility, and decreased milk production, high cost of veterinary care and costs
associated with replacement animals. In addition, the disease also hinders free animal
movement due to imposition of quarantines to affected farms and it is a great
impediment for international trade of animals and their products [4]. The disease in
humans is associated with high direct and indirect medical and non-medical expenses
arising from high cost of treatment of primary disease and management of sequelae as
well as productivity losses [5].

Brucellosis is endemic is several parts of Kenya and there is increasing evidence


that the incidence and prevalence is increasing both in humans and animals more so in
pastoral regions [6-8]. Since domestic animals are the source of infection to humans,
increasing incidence and prevalence in humans is perhaps a reflection of a similar trend
in domestic animals. Livestock plays a crucial role in the livelihood of the majority of

154
residents of Garissa who are predominantly nomadic pastoralists keeping mainly sheep,
goats, cattle and camels. It is estimated that livestock sector provides employment to
almost 95% of the residents of Garissa County and it is the main source of milk and
meat and plays an important role in many socio-cultural traditions in this set up. Sheep
and goats are considered as cash banks which can easily be liquidated to meet urgent
family needs. Between 2005 and 2007, the sector generated an estimated 1.8 billion
Kenya shillings from direct sales in domestic and overseas markets [9]. However this
dependence on livestock makes these people vulnerable to zoonotic diseases. Some of
the socio cultural practices such as consumption of raw milk has been shown to make
residents of Garissa at greater risk of infection with brucellosis [6].

Although there are no published studies that incriminate Brucella species as


cause of abortion in goats and sheep in Garissa County, brucellosis has been suspected
on basis of clinical grounds. Unfortunately, reliable data on brucellosis in Kenya is
scarce and data on sero-prevalence and risk factors associated with small ruminant
brucellosis in Garissa County is unknown. Understanding of sero-prevalence of
brucellosis, geographical distribution and risk factors for its transmission in mall
ruminants is vital for designing effective control strategy [10]. This study aimed at
determining the sero-prevalence of brucellosis in sheep and goats at individual animal
and flock level and to identify factors associated with brucellosis at flock level with an
aim of providing evidence based information to inform prevention and control strategies
in both domestic animals and humans in this region.

Materials and Methods

Study site and study population

We conducted the study in Garissa County which is classified as arid and semi-
arid and has an altitude range of between 70m and 400m above sea level. Human
population is estimated at 623,060 and combined sheep and goat population at 1,322,457

155
according to the 2009 Kenya housing and population census. The region has two rain
seasons (April to May and October to December) with annual mean precipitation of
225mm. The area is characterized by hot and dry weather throughout the year and daily
temperatures are typically above 30 °C (86 °F), while at night, they can fall to 20 °C (68
°F). Indigenous people of Somali origin account for the majority of the population in
Garissa County [9, 11].

Study Design

We conducted a cross-sectional study between October and November 2013 in


flocks that we randomly selected from 36 sub-locations in Garissa Township and
Balambala sub-counties of Garissa County. We calculated sample size using Thrushfield
formula for simple random sampling [12] as shown below:

We assumed an expected flock prevalence of 16% in goats [7], a 10% level of precision
of the estimate, a confidence interval of 95% and a design effect of two because of the
multi-stage sampling approach used to give a minimum sample size of 103 flocks. In
addition, we applied a 10% contingency to give rise to 114 flocks.

We obtained a list of households from all the sub-locations in Garissa and


Balambala sub-counties with estimated numbers of sheep and goats owned by each
household. This was achieved with assistance from the local national government
administrators, community elders, community animal health workers (CAHWs) and the
Garissa County veterinary and livestock production office. We considered the sheep and
goats flocks as the primary sampling unit (PSU) and used the household lists as proxy
for the number of flocks in each sub-location. Due to logistic issues, we randomly
selected a third of the sub-locations (12 sub-locations) for inclusion into the study. We
randomly selected the number flocks to sample in each sub-location proportionate to the
156
number of flocks in each sub-location. Once we determined the number of flocks per
sub-location to be included in the study, we employed systematic random sampling
(SRS) technique to select flocks within each sub-location for sampling using the
livestock owners’ list as sampling frame. We attempted a simple random sampling to
select individual animals within the flock for blood sample collection. We sampled up to
a maximum of 20 animals per flock and we selected sheep and goats for sampling
proportionate to their numbers within the flock. Since sheep and goats are raised
together as one flock in this set up, the same sample size applied to both. We included
flocks whose owners (or representative) were willing to participate in the study; allowed
blood samples to be collected from the flock and were ≥15 years of age. We replaced
flocks whose owners/representative not meeting the inclusion criteria with another flock
among the remaining flocks in the list from the same sub-location to achieve the
required sample size for a particular sub-location.

Sample collection

The owner and field assistants individually restrained sheep and goats selected
for sample collection and we aseptically collected10–15 ml of blood into plastic
vacutainers®. We allowed the vacutainers® to stand for approximately 15 to 30 minutes
in a rack in a slanting manner at ambient temperature to separate serum from clot. We
harvested the sera using disposable plastic Pasteur pipettes and dispensed into Eppendorf
tubes and stored in a cool-box containing ice cubes in the field. We transported the
Eppendorf tubes to the laboratory where the specimens were stored in a freezer at -20°C
until used for serological testing.

Sample analysis

We screened for presence of antibodies in collected sera specimens using Rose


Bengal Plate test (RBT) and complement fixation test (CFT). The sensitivity and
specificity of RBT were 89% and 97%, respectively and that of CFT were 88% and

157
100%. The CFT and RBT test antigens (Brucella arbortus strain 99), control sera and
other reagents were obtained from Atlas medical, William James House, Cowley Rd.
Cambridge Cb4, 4WX and sensitized sheep red blood cells (SRBC) were obtained from
the central veterinary laboratory (national veterinary reference laboratory in Kenya). The
sera specimens were tested serially first using RBT then CFT for those that tested
positive on RBT. An animal was considered positive if the serum specimen tested
positive on both RBT and CFT whereas a flock was considered positive if at least a
single serum specimen from an animal within the flock tested positive on both RBT and
CFT. For RBT, Rose Bengal test antigen was prepared from killed standard strain of B.
abortus strain 99 and stained with Rose Bengal dye, in an acidic buffer pH 3.65. Serum
samples and the antigen were left at room temperature for an hour before the test
commenced. We shake the bottle containing the antigen so that the suspension was
homogeneous. We thereafter added 30µl of the sample after swirling for a minute onto a
white tile and same volume of antigen alongside the antigen spots using a micro-pippete.
We then thoroughly mixed the sera and antigen using a wooden splint, using one
wooden splint for each test, until a circular zone of approximately 2 cm was formed. We
thereafter rocked the white tile both clockwise and anti-clockwise for four minutes
(timing was done using a laboratory buzzer). We then observed for any agglutination
(due to antigen and antibody complex formation) in a well-lit place to avoid false
positive reading due to formation of fibrin. We used a magnifying glass to examine
those tests that were suspected to have micro-agglutination. We considered a positive
test results as any visible agglutination and negative results as absence of any visible
agglutination. We tested a control serum that provides minimal agglutination every day
before the actual testing began to verify the sensitivity of the test conditions. For CFT,
we diluted the test serum and appropriate working standards with equal volume of
veronal buffered saline in small tubes and incubated at 58°C for 50 minutes to inactivate
the native complement. We thereafter dispensed 25µl of diluted test serum on a round
bottom 96 well micro-titre on the first and second rows of the well. We then added 25µl
of the veronal buffered saline to all wells except those on the first row. We applied
158
serial doubling dilution by transferring 25µl of serum from third row onwards and
discarded 25µl of the mixture in last row. We carried out the serial dilution four times.
We then dispensed 25µl of antigen to each well except in the first row followed by 25µl
of complement to each well. We set up control wells having diluent only; control wells
having complement and diluent; and control wells having diluent, complement and
antigen each at 75µl volume in each control wells. We tested control serum that results
into a minimum positive reaction for each set of tests to ascertain the sensitivity of test
conditions. We then incubated the plates at 37°C for 30 minutes and thereafter added
25μl of sensitized sheep red blood cells (SRBC) to each well. We then re-incubated the
plates at 37°C for 30 minutes. Thereafter we centrifuged the plates for 100rpm for 10
minutes to allow the SRBC that did not undergo hemolysis to settle. We compared
degree of hemolysis with standard s corresponding to 0, 25, 50, 75 and 100% hemolysis.
We also checked for the absence of complementary activity for serum in the first row.
We considered sera samples having SRBCs sedimentation at a dilution ≥ 1:5 to be
positive for Brucellosis. We conducted both RBT and CFT tests as described and
recommended by OIE [13].

We determined the true prevalence (TP) at individual animal level and flock level using
a formula by Rogan and Gladen [14] and combined test sensitivity and specificity of the
two tests when interpreted serially which we calculated as 78% and 100% respectively
[15].

Data collection

We administered a pre-tested structured questionnaire to the flock owner or their


representative immediately after bleeding the animals to collect information on potential
flock level factors associated with small ruminant brucellosis. We interviewed the

159
participants in their local language by assistance from a trained research assistant. We
translated the questionnaire from English to Somali language (local language) and back
translated to English using professional independent translators to ensure consistency,
clarity and socio-cultural acceptability by the community. We pre-tested the
questionnaire on five flocks and made adjustments, additions and modifications to the
questions. The flock level factors we collected included: flock size; contact with wildlife
and wildlife abortus; introduction of new animal into the flock in the past one year;
keeping other livestock apart from sheep and goats; history of abortion in livestock kept
in past one year; seeking veterinary services and lending male animals to other farmers
for breeding.

Data management

We entered cleaned and analyzed data from questionnaires on flock-level factors


and laboratory results using Epi Info 7 (CDC, Atlanta, GA, USA) and Ms. Excel 2007
(Microsoft, Seattle, WA, USA). We calculated descriptive statistics for relevant
variables. For proportions, we estimated the 95% confidence interval (CI) using the
exact binomial test. To evaluate the association between the flock level factors and
Brucella sero-positivity, we carried out univariable analysis where we calculated
prevalence odds ratios (PORs), 95% confidence intervals (95% CI) and p-values. Factors
with p-value of ≤ 0.05 were considered statistically significant. We used unconditional
multiple logistic regression employing a stepwise forward selection approach to identify
independent factors associated with flock level Brucella sero-positivity. We calculated
adjusted odds ratios (aOR), 95% CI and p-values. For selection of independent variables
for inclusion into the initial multiple logistic regression model, the entry criterion was p-
value ≤ 0.20. We developed the model by dropping the least significant independent
variable until all the remaining predictor variables were significant (p-value ≤ 0.05). All
biologically and statistically plausible two-way interactions between variables remaining
in the final model was tested and retained if significant.

160
Results

Individual animal level sero-prevalence

We sampled a total of 2,400 animals composed of 979 (41%) sheep and 1471
(59%) goats from 120 flocks having a total of 12,945 animals. The median flock size
was 112 animals (range: 58 to 140 animals). Of the 2,400 sera samples examined, 437
(18.2%); (95% CI: 16.7% to 19.8%) were positive for Brucella antibodies on RBT. Of
the 437 samples positive on RBT, 138/979 (14.1%); (95% CI: 12.1% to 16.4%) were
from sheep and 299/1471 (20.3%); (95% CI: 18.4% to 22.5%) were from goats. Of the
samples testing positive on RBT, 394/2400 (16.4%); (95% CI: 15.0% to 18.0%) tested
positive on CFT of which 104/979 (10.6%); (95% CI: 8.8% to 12.7%) were from sheep
and 290/1471 (19.7%); 95% CI: 17.8% to 21.8%) from goats.

Adjusted overall true sero-prevalence of Brucellosis at individual animal level in


sheep and goats was 20.0% (95% CI: 18.2% to 22.0%); and in goats it was 24.3% (95%
CI: 21.8% to 27.1%) and sheep 12.5% (95% CI: 10.2% to 15.2%). At individual animal
level, highest apparent sero-prevalence of 37.8% [(68/180); (95% CI: 30.92% to
45.03%)] was recorded in Abdisimit; 25.4% [(61/240); (95% CI: 20.21% to 31.21%)] in
Ohio and 22.5% [(63/280); (95% CI: 17.9% to 27.67%)] in Kuno. The lowest apparent
sero-prevalence of 8.5% [(17/200); (95% CI: 5.2% to 13%)] were recorded in Sikley and
2% [(6/300); (95% CI: 0.82% to 4.11%)] in Balambala (Table 1).

Flock level sero-prevalence

At the flock level, the overall apparent flock sero-prevalence was 51.7%
(62/120); 95% CI: 42.8% to 60.4%) and the overall true flock sero-prevalence of
brucellosis was 65.8% (95% CI: 54.3% to 77.2%). The apparent flock level sero-
prevalence varied by sub-location with the highest 85.7% [(6/7); (95% CI: 47.0% to

161
99.3%)] recorded in Galbet and the lowest 13.3% [(2/15); (95% CI: 2.3% to 37.5%)]
recorded in Balambala (Table 2).

Flock level factors associated with Brucellosis sero-positivity

In determining both the risk factors and protective factors associated with
brucellosis flock sero-positivity, we examined 10 factors at the univariate analysis of
which five variables (introduction of new animal in the flock, experiencing abortion in
sheep and goats flock, contact with other flocks, lending male animals to other flocks for
breeding purposes and seeking veterinary services) were significantly associated with
brucellosis flock sero-positivity with p values ≤ 0.05. The following factors were
independently associated with brucellosis sero-positivity in sheep and goats at flock
level: introduction of new animals into the flock [adjusted odd ratio (aOR) = 8.0; 95%
CI: 3.1 to 20.7], experiencing abortion in sheep and goats flock (aOR = 3.4; 95% CI: 1.3
to 8.9) and seeking of veterinary services (aOR = 0.3; 95% CI: 0.1 to 0.8) (Table 3).

Discussion

We report the serological analysis of exposure to Brucella spps in sheep and


goats and determination of factors associated with sero-positivity at the flock level from
a population based sample of sheep and goats flocks in Garissa County. The study
estimated the true sero-prevalence of Brucella spps in sheep and goats both at the
individual animal and flock level incorporating the sensitivity and specificity of the
diagnostic tests used and the uncertainties in these tests. The large uncertainty in the
estimates of sero-prevalence especially at the flock level could be attributed to the small
sample size of flocks included in this study from sub-locations sampled. However, the
result from this study does confirm that there is very high level of probability that the
sheep and goats flocks in Garissa have been exposed to Brucella spp. The study also
highlights the need for adjusting for the sensitivity and specificity of serological tests

162
used in serological surveys for reliable interpretation of results. This also provides
unbiased estimates of sero-prevalence estimates.

Brucellosis sero-prevalence in the sheep and goats population in Garissa County


appears to be high at 20.0% individual animal level and 65.8% at flock-level. Several
studies have found very variable small ruminant brucellosis sero-prevalence at
individual and flock-level across various African countries. Estimates include animal-
level sero-prevalence of between 0.4% and 9.7%, and 15% flock-level sero-prevalence
in Ethiopia [16-29]; 16% individual animal and 20% flock-level sero-prevalence in
Cameroon [30]; between 0% and 8.2% individual animal and 0% and 56.3% flock sero-
prevalence in Eritrea [31]; between 0.4% and 3.6% individual animal and 17.8% flock-
level sero-prevalence in Niger [32]; between 0.44% and 12.1% individual animal level
in Egypt [33, 34]; individual animal level of 3.3% in sheep and 4.5% in goats in Nigeria
[35]; between 0.9% and 22% individual animal level in Sudan [36-38] and in Uganda,
4% individual animal and 13% flock-level sero-prevalence in goats were estimated [39].
In Kenya, studies done in Kiambu, Kajiado and Marsabit Counties estimated animal-
level sero-prevalence ranging from 1.3% to 16.1% and flock level sero-prevalence
ranging from 5.6% to 68% [7]. Another study done in Kenya in Baringo County
estimated an individual animal sero-prevalence of 13.04% in goats and 8.23% in sheep
[8]. These variations in estimates of sero-prevalence of brucellosis in small ruminants
may be largely attributed to the varied animal husbandry systems, various agro-
ecological zones in which the studies were carried out, the sampling methodology
employed and diagnostic tests used. These factors have been shown to contribute to
variation in the obtained sero-prevalence of brucellosis infection in livestock among
different researchers [40-43]. However the high sero-prevalence of brucellosis in sheep
and goats in Garissa County portends greatest probability of zoonotic transmission of
brucellosis to humans.

The introduction of new animals into the sheep and goats flock from unscreened
flocks was significantly associated with flock seropositivity in this study. This was in
163
agreement with several other studies which noted that introduction of animals from non-
free brucellosis flocks or from flocks of which their brucellosis status was unknown was
significantly associated with brucellosis in sheep and goat flocks and in cattle [39, 44-
48]. Other studies suggest that the introduction of infected animals can lead to an
increase in the individual level prevalence due to the fact that the longer the animals stay
in the flock and they are in contact with rest of the flock, the higher the risk of spread of
brucellosis would be [49, 50]. Practices that involve movement of animals between
flocks have also been found likely to be risky and potentiate transmission of brucellosis
between flocks. Evidence suggests that failure of most brucellosis control strategies
could be attributed to the lack of control in the movement of animals and measures to
avoid introduction of infected animals by maintaining a completely closed flock or by
carefully screening purchased animals before introducing them into the flock, a practice
that is very rare in pastoral communities, is perhaps one of the effective control
strategies for small ruminants brucellosis [39, 44, 51].

Presence of female animals who had aborted in the flock was found to be
significantly associated with flock sero-positivity. Abortion in livestock represents the
major complaint attributed to Brucella infections [39, 52-55]. Females infected with
Brucella spps are known to shed highly concentrated volumes of Brucella organisms in
their milk, placental membranes and aborted fetuses and continue shedding the
organisms for several months resulting into environmental contamination [4]. As a
result, there is enhanced high risk of transmission of Brucella organisms between
animals of the same flock and other flocks during free mixing in grazing and watering
places. There is also enhanced zoonotic transmission to humans. During this study, we
also assessed the livestock owners knowledge attitude and practices towards brucellosis
as reported elsewhere [56] where most of those interviewed reported assisting animals
during abortions and birthing processes and handling fetal materials without any
protective clothing putting them at risk of coming down with the disease should such
materials be contaminated with Brucella organisms. Similarly the owners reported

164
consuming unpasteurized milk and milk products and just dumping aborted fetuses and
fetal membranes, practices which clearly shows their ignorance of the epidemiology of
the disease.

Lending male animals to other flocks for breeding purposes and seeking
veterinary services in the past year were also factors that were significantly associated
with brucellosis flock sero-positivity with the former being a risk factor and the latter a
protective factor. Lending of male animals for breeding has been identified in other
studies to be a risk factor associated with Brucella sero-positivity in animals [57, 58].
Although venereal route is not considered an important route for Brucella transmission
in small ruminants under natural conditions, practices that involve movement of animals
between flocks are considered risky due to potential of mechanical transmission [4, 59].

We utilized a cross-sectional study design which was not ideal for investigation
of flock level factors associated with brucellosis in sheep and goats in this region. The
study design could not permit the investigators to determine when the animals within the
flocks classified as positive were first exposed or when the Brucella organisms were first
introduced into the flock. This may have led to misclassification of the flock level
factors associated with seropositivity. However, since the study aimed to identify
associations and not cause-effect relationships; we deemed that the design was sufficient
in identifying the flock level factors associated with Brucellosis in this region despite the
small numbers of flocks included in the study. We only focused on small ruminants’
brucellosis despite the fact that other livestock species such as such as cattle and camels
which are also reared in this community were never studied and they are deemed to be
susceptible to brucellosis. We were also not able to identify the various species and
biovars of Brucella spps circulating in in sheep and goats as evaluate the socio-
economic impact of the disease in limiting livestock production.

Conclusion and Recommendation

165
Despite the limitations highlighted above, the present study confirms a
considerably high seroprevalence of Brucellosis both at individual animal and flock
level in sheep and goats. The introduction of new non-quarantined animals with
unknown brucellosis status into the flock, occurrence of abortions within the flock
(which is could be a pointer to brucellosis infection) and lending of male animals for
breeding purposes were significant risk factors associated with the spread of the disease
within and between flocks whereas seeking veterinary care as found to be a protective
factor. Necessary measures should therefore be taken to prevent the transmission of
brucellosis to the human population. Future studies in this region should holistically
focus on all animal species including humans to provide a more comprehensive picture
of the disease epidemiology and socio-economic implications.

The pastoral lifestyle of the community in the study area makes brucellosis
control very challenging not only because of the number and complexity of risk factors
involved, but also because the risk factors are tightly linked and often inherent to the
livestock production practices. The above factors when combined with the close
interaction between the people and their livestock coupled with the socio-cultural
lifestyle and poor knowledge attitude and practices [56] evidenced in this study, poses a
serious public health concern and has been known to enhance transmission of brucellosis
to the human population. These reasons and make control of brucellosis both in
livestock and humans challenging.

There are no officially coordinated program for control of brucellosis in Kenya


and at large, greater horn of Africa where Kenya belongs. However, the African Union
Inter-African Bureau on Animal Resources (AU-IBAR) developed a strategy dubbed
“Standard Methods and Procedures (SMPs) for control of Brucellosis in the Greater
Horn of Africa” which outlines minimum standards, methods and procedures on various
subject areas such as surveillance, laboratory procedures and disease control that must
be met for harmonized regional control of the disease [60]. If fully adopted, these SMPs
will go a long way towards control of livestock brucellosis in the greater horn of Africa.
166
However for the full potential of the SMPs to be realized, there is need to evaluate
attitudes of communities involved and their roles need to be clearly defined in the
various control strategies. For effective control of this disease in this region, a “One
Health” approach is recommended.

Abbreviations

RBT: Rose Bengal Plate Test; CFT: Complement Fixation Test; POR: Prevalence odds
ratio; aOR: Adjusted odds ratio; AU-IBAR: African Union Inter-African Bureau on
Animal Resources; SMP: Standard Methods and Procedures

Declarations

Acknowledgements

We would like to extend our thanks to Kenya Field Epidemiology and Laboratory
Program (K-FELTP) for financial support to carry out the study. The authors would also
wish to acknowledge all the staff of the Department of Veterinary Services and
Livestock production of Garissa County and staff at Regional Veterinary Investigation
Laboratory in Garissa for their cooperation and untiring efforts towards the success of
this study. We would also acknowledge the study participants and the community
leaders for support provided during the study.

Funding

This study was fully funded by Kenya Field Epidemiology and Laboratory Training
Program who provided full scholarship for the first author to pursue a master’s degree in
applied epidemiology from Jomo Kenyatta University of Agriculture and Technology.
The funding body of this study did not participate in the design or conclusion of the
study.

167
Availability of data and materials

The datasets used and/or analyzed during the current study are available as Additional
file 1.

Authors’ Contribution

Both the authors’ conceived and designed the study. MO collected, cleaned the data,
performed the experiments and analyzed the data. MO drafted the manuscript. GK
helped with the interpretation of the results. All authors’ read, critically reviewed and
approved the final manuscript.

Ethics approval and consent to participate

We sought and obtained approvals for the study protocol from Board of post graduate
studies of Jomo Kenyatta University of Agriculture and Technology (JKUAT) and we
obtained ethical clearance from Kenyatta National Hospital/University of Nairobi-Ethics
and Research Committee (KNH-UoN ERC) under approval number P248/05/2013. We
explained the study objectives to all potential study participants in their local language
(Somali) and obtained informed oral consent from all those who agreed to participate in
the study. We obtained ethical approval for verbal consent from KNH-UoN ERC and
documented consent from each consenting study participant on the questionnaire for
data collection. We independently interviewed each study participant and measures were
taken to ensure that collected data was properly stored, secured and confidentiality
maintained.

Consent for publication

Not applicable.

Competing interests

168
The authors declare that they have no personal or financial competing interests that may
bias publication of this manuscript.

Author Information:

Dr. Mark Obonyo-Field Epidemiology and Laboratory Training Program, Ministry of


Health, Kenya and Ministry of Agriculture, Livestock and Fisheries, Directorate of
Veterinary Services, Kenya

Prof. Gideon Kikuvi-Jomo Kenyatta University of Agriculture and Technology, School


of Public Health

169
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Appendix 6: Manuscript on Brucellosis KAP survey

Published in International Journal of Innovative Research and Development, September,


2015 Vol 4 Issue 10

Title: Knowledge, Attitude and Practices towards Brucellosis among pastoral


community in Kenya, 2013

Running Title: Pastoralism, Brucellosis and Knowledge, Attitude and Practices

Author: Mark Obonyo1, 2ꭞ and Waqo Boru Gufu1, 3, Gideon Kikuvi2

Affiliations:

1
Field Epidemiology and Laboratory Training Program

2
Jomo Kenyatta University of Agriculture and Technology

3
Ministry of Health, Kenya

ꭞCorresponding author: markobonyo@yahoo.com

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Abstract

Background: Brucellosis is a global zoonotic disease and a major public and animal
health problem in many parts of the world, particularly in pastoral set up where livestock
is a major source of livelihood and food. Effective prevention and control of Brucellosis
depends on knowledge, attitude and practices of the community. This study aimed to
assess the knowledge, attitudes and practices related to Brucellosis among pastoralists in
Garissa.

Methods: The study was based on a cross-sectional study design, using a multistage
sampling technique and a structured questionnaire was administered using a face-to-face
interview to farmers aged 15 years and above.

Results: A total of 120 pastoralists were interviewed of which 90 (75%) were male;
median age was 16 years (Range: 15 – 70 years); 102 (85%) were aged below 35 years
and 95 (79%) had heard of Brucellosis. Among those aware of Brucellosis, 17 (18%)
mentioned bacteria/germ as cause and 44 (46%) were informed through community
health workers. Abortion was mentioned by 56 (59%) of respondents as main clinical
sign of Brucellosis in animals. Sixty-seven (71%) knew Brucellosis as zoonotic disease
of which 55 (82%) mentioned drinking of raw milk as main route of transmission. Fever
was mentioned by 71 (75%) as main clinical symptom. Regarding attitudes and
perceptions, 13 (14%) knew that Brucellosis could be prevented in animals; 33 (35%)
knew that it could be treated in humans; only eight (8%) would visit a health facility if
they suspected Brucellosis and 44 (46%) would do nothing if they had aborting animal
in their herd. Regarding practices, 91 (96%) consumed raw milk in past year; 72 (76%)
assisted an animal during birthing process of which 61 (75%) disposed fetal materials by
dumping; and 34 (36%) participated in slaughtering an animal.

Conclusions: The study indicates that Brucellosis remains a major public health
problem among the pastoralists in this area. Though the community has fair knowledge
on Brucellosis, attitudes, perceptions and practices are poor. The study highlights the

179
importance of increased provision of information about knowledge attitude and practices
regarding Brucellosis in this area as one of the major strategies in prevention and control
of Brucellosis.

Key words: Knowledge; Attitude; Practice; Pastoralism; Brucellosis

180
Introduction

Brucellosis remains amongst the most normally disregarded zoonotic diseases


worldwide[1, 2]. The true incidence of Brucellosis in human and animals worldwide is
obscure and the occurrence is expanding in low and middle income nations like Kenya
[3, 4]. The bacterial pathogen is considered by US Centers for Disease Control and
Prevention (CDC) as a category (B) pathogen that has potential for improvement as a
bio-terrorism weapon with a capability of airborne transmission [5].

In animals, Brucellosis is thought to be a group or herd issue spread inside of the herd
fundamentally by ingestion of contaminated materials. Venereal infection can likewise
happen, primarily with B. Suis. Congenital (in utero) or perinatal infection might
likewise happen often resulting into latent infection. Spread between herds normally
happens by introduction of asymptomatic chronically sick animals. Initial infection in
female animals results in abortion and in long term, delayed or permanent infertility. The
disease is considered chronic and infected animals continue to shed Brucella organisms
following abortions, after subsequent parturitions and also in milk and colostrum [6].

Human transmission occurs through breaks in the skin following direct contact with
contaminated animal tissues like blood, urine, vaginal discharges, aborted fetuses or
placentas. Foodborne transmission occurs more often from consumption of raw milk and
raw milk products like cheese and yoghurt. However once in a while eating raw meat
from infected animals may result into infection. Brucellosis is considered an
occupational hazard and airborne transmission has been documented among personnel
working in laboratories and among abattoir workers. Accidental inoculation with live
vaccine (such as B. abortus Strain 19 and B. melitensis Rev.1) can likewise happen.
Cases of venereal and congenital infections are also known to occur in humans [7, 8].
Incubation period following infection with Brucellosis in human varies from few days to
several years. This is followed by clinical signs and symptoms mostly characterized by
intermittent or undulant fever, headaches, weakness, profuse sweating, chills, depression
and weight loss [9].

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Brucellosis represents various difficulties in designing effective prevention and control
programs. This is primarily due to the chronic or asymptomatic nature of the disease in
both animals and humans, varied incubation period and often lack of laboratory
confirmation[10, 11]. In pastoral communities where Brucellosis is of most prominent
significance due to close interaction between the pastoralists and their animals,
population of animals in such set up are usually ill defined or unknown [10]. In Kenya,
Brucellosis is endemic in several parts of the country and evidence exists of increasing
incidence and prevalence in both human and animals especially in pastoral areas in
Kenya [12, 13]. Given that infected animals are the source of human infection, the
increasing prevalence of human Brucellosis probably reflects a similar trend in domestic
animals. Due to nomadic pastoral lifestyle of the study community there is frequent
mixing of animals on common grazing grounds or at water sources ensures maintenance
of infection within and between herds. The eating habits and lifestyle of pastoralist also
enhance transmission of Brucellosis in humans thus making control of Brucellosis both
in livestock and humans challenging [14, 15]. Effective Brucellosis control programs
depends on understanding of prevalence, geographical distribution, risk factors for
transmission and knowledge, attitude and practices of livestock owners [14]. Limited
information is available on knowledge, attitude and practices of livestock farmers in this
set up. This study aimed at assessing the knowledge, attitude and practices of sheep and
goat farmers and owners in Garissa County in order to provide evidence based
information geared towards prevention and control of Brucellosis both in animals and
humans.

Methods

Study area and population

The study was conducted in the pastoral County of Garissa which is low lying with
altitudes ranging between 70m and 400m above sea level. The county is generally semi-
arid and receives annual rainfall of between 150mm and 300mm. The communities in
this region derive their livelihoods by selling livestock and livestock products and by

182
product, and recently they have started growing food crops especially along river Tana
which traverses the County. The rainfall in this area is unreliable and main sources of
water for both livestock and humans are mainly permanent water points like bore holes,
dams and seasonal shallow wells. The temperatures in the county are high ranging from
20oC to 38oC [16]. Human population is estimated as 623,060 and combined sheep and
goat population is 1,322,457 animals [17] .

Sampling Procedure and sample size determination

Between October and November 2013, a cross sectional survey was conducted in
randomly selected households located in 36 sub-locations of Garissa County to assess
the knowledge and perception of the communities about Brucellosis. With the help of
the Local national government administrators, local elders, community animal health
workers and veterinary office, the research obtained a list of households from all the
sub-locations with estimated numbers of sheep and goats owned by each household. The
primary sampling unit was the sheep and goats herds and the household lists acted as
proxy for the number of herds in each sub-location. Due to logistic issues, a third of the
sub-locations (12 sub-locations) from the 36 listed were selected using simple random
sampling technique. Number of herds to sample in each sub-location was randomly
selected proportionate to the number of herds in each sub-location. Once the number of
herds per sub-location was determined, a systematic random sampling technique was
used to select herds within each sub-location for sampling using the generated livestock
owners list as a sampling frame. Study eligibility was based on willingness to be
interviewed and being more than or equal to 15 years. Any livestock owner or
representative not meeting any of the criteria was replaced with another farmer or
representative from the sub-location list until the desired sample size in each sub-
location was achieved. Sample size was estimated at 114 herds using a 16% herd
prevalence of Brucellosis in goats in Kajiado County [13], a 10% level of precision of
the estimate, a design effect of 2 due to multi-stage sampling technique employed.
Thrushfield formula for simple random sampling was employed [18].

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Data management

Information on knowledge about Brucellosis which included awareness of Brucellosis,


sources of information on awareness, causes , awareness of Brucellosis as a zoonotic
disease, mode of transmission in both animals and humans, signs and symptoms in
animals and human and prevention and control measures in animals and treatment in
humans. Information collected on attitude and perceptions included attitude and
perception on seriousness of Brucellosis in human and animals, attitude towards
prevention of Brucellosis in animals and treatment in humans, attitude towards aborting
animals and attitude and perceptions when someone suspects to have Brucellosis.
Information on practices included consumption of raw milk or milk products made from
raw milk, participation in slaughtering or butchering an animal, assisting an animal
during birth or removal of retained placenta or abortion and method of dumping of foetal
materials after birth or abortion. Information on the socio-demographic characteristics of
the participants was also included in the questionnaire. The questionnaire was translated
from the original English version into the local language (Somali) and back translated to
English by independent persons to ensure consistency, clarity and socio-cultural
acceptability in the community. During pre-testing, additional information was gathered
and some of the questions were modified. The participants were interviewed in their
local language by the principal investigator and trained research assistant. We entered
and cleaned the data using Microsoft Excel 2010 (Microsoft, Seattle, WA, USA), and
analyzed using Epi Info version 7 (CDC, Atlanta, GA, USA) and Microsoft Excel 2010.
We calculated proportions for categorical variables and means and medians for
continuous variables.

Ethical issues

The study protocol approval was sought and obtained from Board of post graduate
studies of Jomo Kenyatta University of Agriculture and Technology (JKUAT) and
ethical clearance was sought and obtained from Kenyatta National Hospital/University
of Nairobi-Ethics and Research Committee (KNH-UoN ERC). The study objective was

184
explained to participants in their local language (Somali) and informed oral or written
consent was obtained from each study participant who agreed to participate. Each
participant was interviewed independently and measures were taken to assure collected
data were properly stored, secured and confidentiality maintained.

Results

Socio-demographic characteristics of participants

A total of 120 participants were interviewed to assess their knowledge, attitude and
practices towards Brucellosis. The median age of the study participants was 16 years
(Range: 15 – 70 years), with 102 (85%) aged below 35 years. There were 90 (75%)
males; and 92 (77%) had no formal education. In regard to primary role of the study
participants in management of the herd, 58 (48%) were herd owner, 38 (32%) were
herders and 24 (20%) were involved mostly in milking animals in the herd. Eighty-three
(69%) of the participants were married and 37 (31%) were single (Table 1).

Awareness and cause of Brucellosis in animals and humans

Among the study participants, 95 (79%) had heard of Brucellosis, 17 (18%) mentioned
germs/bacteria as the cause of Brucellosis, 38 (40%) did not know the cause, 14 (15%)
mentioned food, and 13 (14%) mentioned water and wild animals. Among those who
had heard of Brucellosis, 44 (46%) was through community health workers, 19 (20%)
from a family member, 19 (20%) religious leaders, eight (8%) from veterinary staff and
five (5%) from local FM stations (Table 2).

Knowledge of respondents’about the animals affected and signs/symptoms of


Brucellosis in animals

In regard to animal species affected by Brucellosis, 62 (65%) mentioned goats, 47 (49%)


sheep, 45 (47%) cattle and 32 (34%) camels. Fifty-six (59%) of respondents mentioned
abortion as most common sign, 21 (22%) mentioned retaned placenta, 20 (21%) swollen

185
joints or hygroma and 11 (12%) mentioned mastitis or swollen udder and teats (Table
3).

Knowledge of respondents on modes of transmission of Brucellosis to humans

Concerning Brucellosis being zoonotic disease, 67 (71%) of the respondents knew of


this. Among these, 55 (82%) mentioned drinking raw milk as most common mode of
transmission of Brucellosis from animals to humans, followed by eating milk products
from raw milk mentioned by 27 (41%) respondents. The least mode of transmission of
Brucellosis from animals to humans was slaughtering animals mentioned by 17 (26%)
respondents (Figure 2).

Knowledge of respondents on signs/symptoms of Brucellosis in humans

Forty-six (48%) of the respondents knew a family member who had been diagnosed with
Brucellosis in the past, 43 (45%) knew somebody who is not a family member who had
been diagnosed with Brucellosis and 38 (40%) respondents had themselves been
diagnosed with Brucellosis in the past. Concerning signs and symptoms in humans, 71
(75%) of respondents mentioned fever, 56 (59%) joint pains, 48 (51%) muscle pains, 45
(47%) loss of appetite and 38 (40%) chills (Table 4).

Respondents’ attitude/perception towards Brucellosis

A total of 63 (67%) of the respondents thought that Brucellosis is a serious disease in


animals whereas 61 (64%) thought that it is a serious disease in human. Only 13 (14%)
respondents thought that Brucellosis can be prevented in animals. Among these, six
(46%) mentioned vaccination, four (39%) contacting veterinary office and three (31%)
isolation of sick/aborting animals. In regard to treatment of Brucellosis in humans, 33
(35%) thought that Brucellosis in human can be treated/cured. Out of these, 14 (42%)
mentioned visiting a health facility, eight (24%) seeking divine intervention (prayers),
six (18%) consuming herbal medicine and five (15%) would purchase medicine from a
local chemist. When confronted with an aborting animal in the herd, 44 (46%) will do

186
nothing, 16 (17%) would treat the animal with antibiotics, 11 (12%) will sell the
aborting animal, 10 (11%) will consult veterinary office for advise, eight (8%) will
isolate the animal from the herd and five (5%) will slaughter the animal (Table 5).

Respondents’ self-reported practices’ towards Brucellosis

Regarding practices towards Brucellosis, a total of 91 (96%) of the respondents


consumed raw milk in the past year, 72 (76%) assisted an animal during birthing process
or abortion or removal of retained placenta, 46 (48%) introduced new animals into sheep
and goat herd, 34 (36%) participated in slaughtering/butchering an animal, 30 (34%)
lend their male animals to other farmers for breeding and 13 (14%) consumed milk
products processed from raw milk and. Among those who assisted an animal during the
birthing process, 61 (75%) disposed of fetal material by dumping and none used any
protective clothing (Table 6).

Sources on more information on Brucellosis

A total of 92 (97%) of the respondents believed that they were not sufficiently informed
about Brucellosis and required more information on Brucellosis. The most favored mode
of receiving information on Brucellosis was through the local FM radio stations
mentioned by 36 (39%) of respondents, 23 (25%) favored religious leaders, 18 (20%)
local community meetings (barazas) and 15 (16%) community health
workers/community animal health workers (Figure 3).

Discussion

The results of this community-based cross-sectional study showed that Brucellosis is


known by the general community in the present study area, since more than three
quarters of the study respondents had heard of Brucellosis. This is similar to findings of
previous studies done in Uganda among pastoral communities living along lake Mburo;
in Egypt among cattle and Buffalo farmers in a village in Nile Delta region and among
small ruminant farmers in the peri-urban areas of Dushanbe Tajikistan in which 99.3%,

187
83.2% and 57% of the respondents’ had heard of Brucellosis. However, the awareness of
Brucellosis among study participants in Uganda and Egypt were higher compared to our
study but that in Tajikistan was lower [19-21]. In contrast to this finding, a study done
among small-scale dairy farmers in an urban and peri-urban area of Tajikistan and
another one done among urban and peri-urban dairy and non-dairy farming households
in Kenya found that most respondents had not heard of Brucellosis. In the Kenyan study,
30% of dairy respondents and 22% of non-dairy respondents knew of the existence of
Brucellosis whereas in Tajikistan 85% of the respondents had never heard of Brucellosis
[22, 23]. Perhaps an explanation as to why the pastoral communities are more aware of
Brucellosis compared to farmers in urban or peri-urban areas could be due to their close
proximity and interaction with animals resulting into in-built indigenous knowledge over
years which is subsequently passed down from one generation to the next. Despite a
higher proportion of the study participants had heard about Brucellosis, majority had
little or no knowledge about the cause of the disease. Less than a fifth of the participants
correctly mentioned germ/bacteria as cause of Brucellosis. Poor knowledge regarding
etiology of Brucellosis could negatively impact on respondents’ preventive and control
methods of Brucellosis in both humans and animals due to misconception on the cause.

The main sources of information on Brucellosis in this study area was community health
workers (CHWs) followed by family members. Contrary to this finding, the study in
Uganda and the two studies in Tajikistan found main source of information to be from
friends/relatives [19, 21, 22]. Few participants in the current study mentioned mass
media (radio/TV) as a source of information about Brucellosis, which was similar to the
studies in Uganda and Tajikistan. This findings implies the powerful role the community
health volunteers play in terms of relaying important health messages to nomadic
pastoralists in this area who in most circumstances have challenges in accessing basic
health care services. Deliberate moves should therefore be undertaken to incorporate the
two in all aspects of health care education for the pastoralists.

Based on results of this study, the respondents’ had basic knowledge about the animal
species affected and signs/symptoms of Brucellosis in animals. In this regard, about two

188
thirds mentioned goats, close to a half sheep and cattle, and majority were not aware that
camels could be affected. This findings contrast with the findings of studies in Tajikistan
[22] where 82% of respondents knew that cattle, sheep and goats could be affected and
the study in Egypt [20] in which 98.1% mentioned cattle, 86% sheep and 85% goats.
However, our study was fairly in agreement with another study in Tajikistan [21] in
which two thirds mentioned that all animals could be affected. With regards to clinical
signs of Brucellosis in animals, more than half of the respondents mentioned abortion as
the major clinical sign. This finding was in agreement with findings of a study done
among pastoralists in Kaduna state in Nigeria and the study in Egypt in which 94.4%
and 59.5% of respondents mentioned abortion as the major clinical sign [20, 24].
However our finding was different from that done in Tajikistan where only 11% of
respondents’ mentioned abortion as a clinical signs of Brucellosis in animals [21].
Knowledge of the animal species affected and signs/symptoms of Brucellosis in animals
are crucial because it positively impacts on farmers’ practices towards prevention and
control measures of Brucellosis in both animals and humans.

More than two third of our study participants knew that Brucellosis is a zoonotic disease,
findings which were similar to those in previous studies conducted in Tajikistan, Egypt,
Nigeria and Uganda [20-22, 25]. However, our findings contrasted those of studies done
in Ghana and Nigeria which found very low awareness of zoonotic nature of Brucellosis
[24, 26]. In our study, among those who were aware of the zoonotic nature of
Brucellosis, consumption of raw milk and raw milk products and handling of aborted
fetuses were the top most modes of transmission of Brucellosis from animals to humans.
The respondents’ response regarding consumption of milk as a mode of transmission
was comparable to findings in Egypt and Uganda [19, 20]. However in the current study,
the respondents’ had low awareness on other modes of transmission such as handling of
aborted fetuses and fetal membranes, consumption of raw or undercooked meat,
assisting animals during parturition and slaughtering animals; most of which have been
identified in many studies as major risk factors for transmission of Brucellosis from
animals to humans [27, 28]. Such low knowledge on mode of transmission of

189
Brucellosis from animals to humans has been documented elsewhere [21, 24, 26]. Good
knowledge of mode of transmission of Brucellosis from animals to humans has been
shown to have a protective effect towards human infection as shown in a hospital based
matched case control study done in Kyrgyzstan [29].

In the current study, the majority of the study participants identified fever, joint pains
and muscle pains in that order as the major signs and symptoms of Brucellosis. This was
consistent with the findings of a previous study in Kyrgyzstan where fever and joint pain
(locally known as “Tajik”) were mentioned as main signs and symptoms of Brucellosis
in humans [21] as well as a study in Nigeria where all respondents knew signs and
symptoms of Brucellosis in humans [25]. However, the finding of the current study is
different from the results of previous studies conducted in other parts of Nigeria and in
Ghana [24, 26] where almost all participants were not aware of signs and symptoms of
Brucellosis in humans. The respondents’ basic knowledge about the signs and symptoms
of Brucellosis in humans could have significant impact if the community knowledge is
enhanced thus reducing diagnosis and treatment delay which in the long run will prevent
sequelae and prolonged human suffering.

The present study showed that a considerable proportion of the study respondents
perceived that Brucellosis was a serious disease in both animals and humans. However,
despite this high perception of risk, most respondents’ had unfavorable attitude towards
prevention of Brucellosis in animals and treatment of Brucellosis in suspected humans.
Regarding respondents’ opinion on actions most would take when confronted with an
aborting animal in their herd, majority would do nothing about it whereas others would
attempt treating the animal with antibiotics or sell the animal. Very few mentioned
isolation of the animal or seeking veterinary services. Failure to isolate suspected
animals has been cited as one of the major risk factors for transmission of Brucellosis
within and between herds as susceptible animals can become infected through contact
with infected animals aborted tissues or consumption of pasture or water contaminated
with aborted materials [30]. Frequent migration of pastoralists with their animals
increases the chances of different herds coming into contact with other potentially

190
infected herds thus spreading diseases [31, 32]. This is more important when considering
the high levels of infectiousness of Brucella species making practices such as sharing
grazing land and drinking water points by pastoral communities a major transmission
pathway of Brucellosis between different herds [33-35]

The study participants indicated that the communities in the present study area are
engaged in risky practices that could expose them to infection with Brucellosis. Nearly
all respondents consumed raw milk, about three quarter assisted animals during
abortions or parturition and handled aborted materials/fetal membranes and a third
participated in slaughtering or butchering an animal. Of those who assisted aborting
animals, three quarter dumped the aborted materials and none used any protective
clothing. Such risky practices have been shown to be important risk factors for
Brucellosis transmission to human [12, 27, 28, 36, 37]. Female animals infected with
Brucella spp. excrete high concentrations of the organism in their milk, placental
membranes and aborted fetuses [6, 30]. Goats have also been shown to have prolonged
secretion of Brucella organisms in milk compared to sheep [38]. Furthermore, Brucella
species have been shown to survive in aborted fetuses, manure and water for periods of
up to 150 to 240 days [39]. Therefore, there is a high risk of transmission of the
pathogen between animals and from animals to humans through direct contact with
contaminated materials such as fetal membranes, aborted fetuses, manure and other
animal products. Introduction of new animals into the herd without quarantine and
borrowing or lending breeding males to other farmers or even taking a female to be
served at a neighbor’s farm have been identified as major risk factors for transmission of
Brucellosis within and between herds as shown in studies in several places [40-46].

Limitations of the study

Although the present study provides important information on the knowledge, attitude
and perception of the pastoralists in Garissa County, it has limitations. The major
limitation of the study was the small sample size which could affect the power of the
study and external validity of the findings making it impossible to generalize findings

191
even to the whole of Garissa County except the villages which were included in the
study. Self-reporting on practices by the respondents was also subject to recall bias and
the face-to-face-interview situation, while enabling full response rates on all variables as
well as participation of livestock keepers most of whom were illiterate, might have
additionally enhanced this type of bias in assessing attitudes and behaviors.

Conclusion

The results of this study revealed that pastoralists in the study area had low level of
knowledge about the causative agent but some moderate knowledge on the main
symptoms of Brucellosis in animals and human. In addition, the study showed moderate
level of overall knowledge, unfavorable attitude and poor practices towards Brucellosis.
At present, there is no officially coordinated program for control of Brucellosis in
Kenya. Understanding of the knowledge, perceptions and practices have been defined as
important pillars regarding the feasibility and the acceptability of potential measures that
might be instituted. Enhanced public health education on the cause, symptoms and mode
of transmission of Brucellosis would be important towards the prevention and control of
Brucellosis in the present study area. This can be achieved by targeted messages in local
FM radios and integrating the community health volunteers in control and prevention
efforts. However for effective control of Brucellosis in the present study area, an
integrated approach should be promoted that takes into account the relationship between
humans, animals and environment in the context of ‘‘One Health approach’’.

Competing interests

The authors’ declare that they have no competing interests.

Acknowledgements

We would like to extend our thanks to Kenya Field Epidemiology and Laboratory
Program (K-FELTP) for financial support. We would also like to acknowledge the study

192
participants, the community leaders, County veterinary office in Garissa for support
provided during the investigations.

193
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Table 1: Distribution of demographic characteristics of respondents in Brucellosis


knowledge, attitude and practices assessment, Garissa 2013

(n=120)
Variable Frequency

n (%)
Age group
15-24 53 (44)
25-34 49 (41)
35-44 14 (12)
>45 4 (3)
Gender
Male 90 (75)
Female 30 (25)
Education level
None 92 (77)
Lower primary 25 (21)
Upper primary 3 (2)
Primary role in Herd
Herd owner 58 (48)
Herding 38 (32)
Milking 24 (20)
Marital status

199
Married 83 (69)
Single 37 (31)

Table 2: Distribution of responses of participants on awareness, cause and source of


information on Brucellosis, Garissa 2013

(n= 95)
Variable Frequency

n (%)
Heard of Brucellosisꭞ

Yes 95 (79)

No 25 (21)

Cause of Brucellosis

Don’t know 38 (40)

Bacteria/germs 17 (18)

Food 14 (15)

Wild animals 13 (14)

Water 13 (14)

Source of information on Brucellosis


Community Health Workers 44 (46)
Relatives/family member 19 (20)
Religious leaders 19 (20)
Veterinary staff 8 (9)

Local FM stations/media 5 (5)

200
ꭞSubsequent analysis based on those who had heard of Brucellosis

201
Table 3: Responses of participants on animal species affected by Brucellosis and
signs and symptoms of Brucellosis in animals, Garissa 2013

(n= 95)
Variable Frequency

n (%)
Animal species affected*
Goats 62 (65)
Sheep 47 (49)
Cattle 45 (47)
Camels 32 (34)
Signs and symptoms of
Brucellosis*
Abortions 56 (59)
Retained Placenta 21 (22)
Hygroma/Swollen joints 20 (21)
Swollen udder/Mastitis 11 (12)

*Multiple responses were permitted

Table 4: Responses of participants on Brucellosis diagnosis and signs and


symptoms in humans, Garissa 2013

(n= 95)
Variable Frequency

n (%)

202
Brucellosis diagnosis
Family member diagnosed with Brucellosis in the past 46 (48)
Person not family member/relative diagnosed with 43 (45)
Brucellosis
Respondent diagnosed with Brucellosis in the past 38 (40)
Signs and symptoms*
Fever 71 (75)
Joint pains 56 (59)
Muscle pains 48 (51)
Loss of appetite 45 (47)
Chills 38 (40)
Headache 37 (39)
Night sweat 33 (35)
Fatigue 29 (31)
Malaise 29 (31)
Vomiting 14 (15)
Painful scrotum in men 14 (15)
Diarrhoea 12 (13)
Blurred vision 9 (10)
Miscarriage in women 7 (8)
Nausea 4 (5)

*Multiple responses were permitted

203
Table 5: Responses of participants on attitude and perceptions towards Brucellosis,
Garissa 2013 (n= 95)
Attitudes and perceptions Frequency

n (%)
Attitude and Perception on Brucellosis seriousness
Serious Disease in Animals 64 (67)
Serious Disease in Humans 61 (64)
Attitude and perception towards Brucellosis prevention
in animals
Brucellosis can be prevented in animals 13 (14)
Prevention by vaccination 6 (46)
Prevention by contacting veterinary office 4 (31)
Prevention by isolation of sick and aborting 3 (23)
animals
Attitude and perceptions towards suspected human
Brucellosis
Brucellosis can be cured in humans 33 (35)
Seek Prayers 14 (42)
Visit health facility 8 (24)
Consuming herbal medicine 6 (18)
Visit local chemist and purchase medicine 5 (15)
Attitude and perceptions towards aborting animals
Do nothing 44 (46)
Treat aborting animals with antibiotics 16 (17)
Sell the animal 11 (12)
Inform veterinary office 10 (11)
Isolate the animal 8 (8)
Slaughter the animal 5 (5)

204
Table 6: Responses of participants on practices towards Brucellosis,
Garissa 2013 (n= 95)

Practices of respondents Frequency

n (%)
Consumption of raw milk 91 (96)
Assisted an animal during birthing/abortion/removal of 72 (76)
retained placenta
Disposal of fetal materials

Dumping 61 (75)

Burning 11 (25)

Burying 0 (0)

Use protective clothing 0 (0)

Participation in slaughtering/butchering an animal 34 (36)

Introduction of new animals into sheep and goat herd 46 (48)

Quarantine new animals 0 (0)

Lend male animals to other sheep and goat herds for 30 (32)
breeding
Processing of raw milk products 13 (14)

205
Figure 2: Responses of participants on mode of transmission of Brucellosis in
humans, Garissa 2013 (n= 67)

206
Figure 3: Responses of participants on preffered sources of more information on
Brucellosis, Garissa County, 2013 (n= 95)

207

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