Republic of the Philippines

NUEVA ECIJA UNIVERSITY OF SCIENCE AND

TECHNOLOGY
Gabaldon Campus
College of Agriculture

MAJOR COURSE 1

Beneficial Arthropods
and Microorganisms
Lecture Manual
 VICTOR PAULO D. DACUMOS
Assistant Professor II
TABLE OF CONTENTS

CHAPTER PAGE
Nature of Biological Control Agents …………………………………………………………. 26
A. Parasitoids, Pathogens, and Predators of Insects and Vertebrate Pests ……………….. 26
B. Antagonists of Pathogens ……………………………………………………………… 27
C. Biological Control Agents of Weeds ………………………………………………….. 28
II. Assessment, Mass Production and Field Releases …………………………………………… 30
A. Assessment of Parasitism ……………………………………………………………… 30
B. Nutrition and Mass Production of Biological Agents …………………………………. 31
III. Methods and Approaches to Biological Control …………………………………………..……
33
A. Quarantined and Exclusion ………………………………………………….……….. 33
B. Use of Resistant Host Plants ….……………………………………………………… 39
C. Destruction by Cultural Management or Direct Removal of Infected/
Affected Hosts …………………………………………………………………………. .. 44
D. Conservation and Augmentation ……………………………………………………. 46
E. Integration with other Control Tactics for Pest Management ……………….………… 49

REFERENCES……………………………………………………………………………... 51
 CHAPTER III. Nature of Biological Control Agents

A. Parasitoids, Pathogens, and Predators of Insects and Vertebrate Pests

Types of Natural Enemies

Parasites, pathogens, and predators are the primary groups used in biological control of
insects and mites. Most parasites and pathogens, and many predators, are highly specialized and
attack a limited number of closely related pest species. Learn how to recognize natural enemies by
consulting resources such as the Natural Enemies Handbook and the Natural Enemies Gallery.

Parasites

A parasite is an organism that lives and feeds in or on a host. Insect parasites can develop
on the inside or outside of the host’s body. Often only the immature stage of the parasite feeds on
the host. However, adult females of certain parasites (such as many wasps that attack scales and
whiteflies) feed on and kill their hosts, providing an easily overlooked but important source of
biological control in addition to the host mortality caused by parasitism.

Although the term “parasite” is used here, true parasites (e.g., fleas and ticks) do not
typically kill their hosts. Species useful in biological control, and discussed here, kill their hosts;
they are more precisely called “parasitoids.”

Most parasitic insects are either flies (Order Diptera) or wasps (Order Hymenoptera).
Parasitic wasps occur in over three dozen Hymenoptera families. For example, Aphidiinae (a
subfamily of Braconidae) attack aphids. Trichogrammatidae parasitize insect eggs. Aphelinidae,
Encyrtidae, Eulophidae, and Ichneumonidae are other groups that parasitize insect pests. It’s
important to note that these tiny to medium-sized wasps are incapable of stinging people. The most
common parasitic flies are the typically hairy Tachinidae. Adult tachinids often resemble house
flies. Their larvae are maggots that feed inside the host.

Pathogens

Natural enemy pathogens are microorganisms including certain bacteria, fungi, nematodes,
protozoa, and viruses that can infect and kill the host. Populations of some aphids, caterpillars,
mites, and other invertebrates are sometimes drastically reduced by naturally occurring pathogens,
usually under conditions such as prolonged high humidity or dense pest populations. In addition
to a naturally occurring disease outbreak (epizootic), some beneficial pathogens are commercially
available as biological or microbial pesticides. These include Bacillus thuringiensis or Bt,
entomopathogenic nematodes, and granulosis viruses. Additionally, some microorganism

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byproducts, such as avermectins and spinosyns are used in certain insecticides; but applying these
products is not considered to be biological control.

Predators

Predators kill and feed on several to many individual prey during their lifetimes. Many
species of amphibians, birds, mammals, and reptiles prey extensively on insects. Predatory beetles,
flies, lacewings, true bugs (Order Hemiptera), and wasps feed on various pest insects or mites.
Most spiders feed entirely on insects. Predatory mites that feed primarily on pest spider mites
include Amblyseius spp., Neoseiulus spp., and the western predatory mite, Galendromus
occidentalis.

B. Antagonists of Pathogens

Antagonists

Antagonists of plant disease and food spoilage microorganisms are not yet well understood.
However, the research that has been done has yielded exciting and promising results, and the study
of antagonists has become a rapidly expanding field in plant pathology.

Worldwide, diseases of crop plants cause losses estimated to be 12%, and post harvest
losses due to food spoilage have been estimated to be between 10% and 50%. In the United States,
these figures are estimated to be 12% and 9%, respectively. Finding ways to prevent
microorganisms from causing these losses would help ensure a stable food supply for the world's
ever expanding population. Outside of agriculture, diseases can cause the destruction of entire
stands of plants in marshes, forests, or other natural settings, and in other plant systems.

Knowledge of the interactions among microorganisms and ways to manipulate microbiota is

growing as research in this field rapidly expands. Antagonists have been successfully used to
suppress tomato mosaic, foot and butt rot of conifers, citrus tristeza disease, and crown gall of
several crops. Seeds have been coated with antagonists that reduce infection by pathogens and also
enhance plant growth. Brown rot of peaches in storage was controlled under simulated commercial
conditions by incorporating the antagonist Bacillus subtilis into wax used in the packing process.
Inoculation of hosts with antagonists has been used with good results against a common fungal
pathogen of conifers and chestnut blight. The future also holds much promise for the suppression
of plant-parasitic nematodes by microbiota.

Growers have applied antagonists to the above-ground parts of plants, to the soil (and roots), and
to plant seeds. The above-ground environment is the least stable for antagonists because of the

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extreme variability in moisture and nutrients. Soil is a more stable environment for microbiota, but
soil in most fields is generally nutrient poor, pH may range from 4-8, and temperatures and
moisture may vary widely. In contrast, greenhouse planting mixes can be managed more
effectively to promote antagonist colonization. Finally, it is practical to treat seeds to favor
microbial antagonists.

To be most effective, antagonists of plant disease and food spoilage should be:

• genetically stable
• effective at low concentrations
• easy to culture and amenable to growth on an inexpensive medium
• effective against a wide range of pathogens in a variety of systems
• prepared in an easily distributable form
• non-toxic to humans
• resistant to pesticides
• compatible with other treatments (physical and chemical)
• non-pathogenic against the host plant

C. Biological Control Agents of Weeds

Biological weed control involves using living organisms, such as insects,

nematodes, bacteria, or fungi, to reduce weed populations. In nature, plants are controlled
biologically by naturally occurring organisms. Plants become pests- and are labelled "weeds"
- when they run rampant because their natural enemies become ineffective or are non-existent.
The natural cycle may be interrupted when a plant is introduced into a new environment,
or when humans disrupt the ecological system.
Biological control of weeds is the deliberate use of natural enemies to reduce the density
of a particular weed to a tolerable level The objective of biological weed control is not eradication
but simply the reduction of the weed population to an economically low level In fact for
biological control to be continuously successful, small numbers of the weed host must always be
present to assured the survival of the natural enemy.
Weeds are plants growing where they are not wanted" and most of our serious weeds have
been introduced from other parts of the world. Of the 107 noxious Canadian weeds, 78 have been
introduced from Europe or Asia. One of the reasons why these plants are so noxious in their new
habitat is that their natural enemies are often absent. Biological control has most frequently been
applied against these alien weeds and attempts are made to restore the natural control of these
weed pests by introducing one or more host-specific, damaging natural enemies from the native
region of the weed.

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Methods Used in Biocontrol of Weeds
The classical approach to biological control of weeds involves the introduction of
hostspecific natural enemies of allien weeds. Recently the approaches utilized in biocontrol
programs have been expanded to include two other methods:
1. augmentation of natural enemy populations;
2. application of "biological herbicides".

Augmentation includes the periodic release and/or distribution of natural enemies. Work is
presently being conducted in the United States to determine the effect of mass rearing of a native
moth Bactra verutana) in the laboratory and releasing it in fields of yellow nutsedge Cyperus
esculentus) (souchet comestible). The application of spore suspensions of plant pathogens as
'biological herbicides' is discussed in more detail later.

Biological Control Agents

Insects have been most frequently used as biological control agents of weeds and this will
likely continue. The reasons are that there have been major successes using phytophagous insects
and almost all of the scientists working in biocontrol of weeds are entomologists. However, recent
research has demonstrated the potential of other organisms, including plant pathogens, nematodes,
and fish.

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 CHAPTER IV. Assessment, Mass Production and Field Releases

A. Assessment of Parasitism
Parasitism is a pervasive phenomenon in nature involving, either as hosts or as parasites,
virtually all species on earth. By definition, parasites are costly for their hosts as they divert
resources for their growth, reproduction and survival with no rewards for the hosts. Parasitism is
not restricted to a few taxonomic groups since a parasitic life style has evolved in viruses, bacteria,
protozoa, invertebrate and vertebrate metazoan. This taxonomic diversity is, of course, also
associated with a tremendous diversity of life cycles, host exploitation strategies, transmission
modes and virulence levels. Given the cost of parasitism, hosts are expected to evolve defense
mechanisms aiming at limiting the negative effect of parasitism on their fitness. In agreement with
this view, hosts have evolved a series of morphological, physiological, behavioral adaptations to
fight off parasitic attacks. On their side, parasites have responded to the selection pressures exerted
by their hosts by evolving counteradaptations to overcome host defenses. These cycles of host–
parasite adaptations–counteradaptations define the so-called coevolutionary process, one the most
prominent characteristics of host–parasite interactions.

Parasitic Relationships
A parasitic relationship is one in which one organism, the parasite, lives off of another
organism, the host, harming it and possibly causing death. The parasite lives on or in the body of
the host.
A few examples of parasites are tapeworms, fleas, and barnacles. Tapeworms are
segmented flatworms that attach themselves to the insides of the intestines of animals such as cows,
pigs, and humans. They get food by eating the host's partly digested food, depriving the host of
nutrients. Fleas harm their hosts, such as dogs, by biting their skin, sucking their blood, and causing
them to itch. The fleas, in turn, get food and a warm home. Barnacles, which live on the bodies of
whales, do not seriously harm their hosts, but they do itch and are annoying.
Usually, although parasites harm their hosts, it is in the parasite's best interest not to kill
the host, because it relies on the host's body and body functions, such as digestion or blood
circulation, to live.
Some parasitic animals attack plants. Aphids are insects that eat the sap from the plants
on which they live. Parasitic plants and fungi can attack animals. A fungus causes lumpy jaw, a
disease that injures the jaws of cattle and hogs. There are also parasitic plants and fungi that attack
other plants and fungi. A parasitic fungus causes wheat rust and the downy mildew fungus attacks
fruit and vegetables. Some scientists say that one-celled bacteria and viruses that live in animals
and harm them, such as those that cause the common cold, are parasites as well. However, they are
still considered different from other parasites. Many types of parasites carry and transmit disease.
Lyme disease is transmitted by deer ticks.
A parasite and its host evolve together. The parasite adapts to its environment by living
in and using the host in ways that harm it. Hosts also develop ways of getting rid of or protecting

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themselves from parasites. For example, they can scratch away ticks. Some hosts also build a
symbiotic relationship with another organism that helps to get rid of the parasite. Ladybugs live on
plants, eating the aphids and benefiting by getting food, while the plant benefits by being rid of the
aphids.
Formula for Assessment of % Parasitism
No. of Infected Samples
% Parasitism= __________________________ x 100
Total No. of Samples Observed

B. Nutrition and Mass Production of Biological Agents

Mass Production of Beneficial Organisms: Invertebrates and Entomopathogens is an
essential reference and teaching tool for researchers in developed and developing countries
working to produce "natural enemies" in biological control and integrated pest management
programs. As we become aware of the negative impact of pesticides in human health and on the
environment, interest is rapidly increasing in developing biological pest control alternatives.
Tremendous advances have been made in beneficial organism technology, such as insect predators
and parasitoids, mite predators, entomopathogenic nematodes, fungi, bacteria, and viruses.
However, developing techniques to mass produce these biological control agents is not enough if
the cost of commercialization is prohibitive. Advancing mass production to the level of economic
feasibility is critical, so these new technologies can compete in the open market. This book
educates academic and industry researchers, and enables further development of mass production
so new technologies can compete in the open market. It is also an excellent resource for those
researching beneficial arthropod mass production and technologies for other uses, including for
study and application in biotechnology and biomedical research.
Because parasitoids are living animals, nutrition inevitably plays an important role in
augmenting these natural enemies of pests. For nutrition is about nourishment; that is, it is the
action or processes of transforming substances found in foodstuff into body materials and energy
to do all the things attributed to life. Nutritional requirements depend on the synthetic abilities of
the organism and the basis is genetical. Therefore, through nutrition we have a direct and essential
connexion between an environmental factor, foodstuff, and the vital processes of the insect
organism. A comprehensive knowledge of the nutritional ecology of insects is a requirement to
understand their biology, behavior, and impact in natural ecosystems and agro-ecosystems.
Although, major insect nutritional requirements for growth and reproduction are known for years,
only a few dozen insects have been studied to learn their nutritional requirements. Extraordinary
nutritional requirements were found in a few insects from even these few studies. It is important
to understand comparative nutritional requirements in order to study insects. In this review,
emphasis will be placed on insect nutritional requirements for natural enemies.

Key biological characteristics to be understood when rearing an insect pest and its
biological control agent

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 • Reproductive strategy: for example, does the insect reproduce sexually?

• Natural sex ratio

• Nutrient requirements at different life stages

• Fecundity

• Longevity of different life stages

• Temperature requirements

• Daylength requirements CHAPTER V. Methods and Approaches to Biological

Control

A. Quarantine and Exclusion

The concept of quarantining materials and organisms to keep unwanted elements from
entering new areas originated almost simultaneously with the distinction between "valuable and
possibly injurious" (Fisher & Andres 1999). Much of the information on excluding unwanted
plants and plant pests from world commerce through the use of quarantine has been assembled by
Kahn (1988) Regulations governing the arrival of questionable materials to a country, state,
province or county arose along with the establishment of quarantine facilities where transported
items could be examined before passing on to their destination and assimilation into the general
economy and environment. Ooi (1986) gave an overview of biological control quarantine from the
perspective of developing countries. The process of seeking effective natural enemies and
importation is highly involved and demands many considerations (Legner, 1986 ).
The primary function of a biological control quarantine facility is to provide a secure area
where the identity of all incoming biological control candidates can be confirmed and undesirable
organisms, especially hyperparasitoids, parasitoids of predators, and extraneous host or host plant
material, can be eliminated. In fact the quarantine laboratory often represents the last chance to
study and evaluate potential biological control agents in the sequence of collection, importation
and liberation.
The number of quarantine facilities in the United States, which are certified to handle
incoming shipments of beneficial organisms, has increased from four to 26 over the past 40 years.
In addition to 24 listed by Coulson & Hagen (1985), new quarantine facilities have been
constructed at Montana State University, Bozeman for phytophagous insects and the University of
California, Riverside for nematodes. New or expanded quarantine facilities have been constructed
in a number of other countries (e.g., Australia, Great Britain, Mexico, Germany and Thailand).
The steady increase in quarantine need and capacity is due on part to an increased interest
in biological and non-polluting methods of pest control, and the desire to expand on the many
successes already achieved through the importation of exotic natural enemies. Also, there is an
increase in new pests that are transported throughout the world and which are amenable to
biological as well as the stricter prerelease information requirements on behavior and safety of
biological control candidates. A lengthy study on candidates often ties up quarantine areas thereby

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increasing the need for greater quarantine capacity to avoid limiting the amount of materials that
can be handled. For example, in the United States to prove the environmental safety of plantfeeding
arthropods for the biological control of weeds can include studies with as many as 10-20 North
American native plant species related to the target weed. When such studies are not permitted or
feasible in the country of origin of the biological control phytophage, these tests must be conducted
in a domestic quarantine facility. Similarly, testing parasitoids against indigenous insect species
which have been declared legally threatened or endangered and which may be present in areas near
or contiguous to insect pest infested agricultural crops that are targeted for parasitoid release, may
not only require more quarantine space but also delay or prevent the colonization of newly
imported organisms. The longer the imported organisms remain in quarantine before these tests
can be conducted, the greater the risk that subtle genetic changes occur, altering the potential fitness
of the organism.
Increasing concern over the quality of the environment is also causing a proliferation of
regulations governing the importation and liberation of beneficial organisms. Explorer collectors,
quarantine officers and project scientists must spend increasing time to study and comply with
domestic and foreign regulations that cover importation, exportation and liberation of biological
control agents. Air travel has reduced the amount of time required to move biological control
agents from one continent to another, but the proliferation of international airports has spawned a
logistical confusion of unpredictable package routing, delayed agricultural and customs inspection
and unscheduled reloading and shipment to the final destination. Frequently material arrives dead
or in a weakened state and on occasion may never arrive.
The safety record for international transport of beneficial organisms has been very good.
Worldwide there have been remarkably few escapes when considering the hundreds of species and
millions of specimens which have been processed. This safety record is a result of the surprisingly
uniform set of international protocols and procedures that are shared by quarantine personnel and
regulatory officials in each country. For an overview of the history and continuing role of
quarantine within the context of classical biological control see Coulson & Soper (1989).
There are several statutory and technological elements of which the explorer, collector and
shipper should be aware and which shape the operation of the quarantine laboratory, beginning
with the collection, selection and packaging of exotic biological control candidates in their native
habitat or country of origin until their release from quarantine or termination of the study. These
include national and state regulations (including required permits) as they pertain to the
certification of quarantine facilities and the importation, handling and release of natural enemies.
Also included are quarantine laboratory design and equipment, personnel and operating procedures
(Fisher 1964, Creager 1987).

Establishing Quarantine Facilities

A quarantine basically provides a tight security room for opening and examining incoming
shipments of beneficial organisms prior to release to other laboratories for further study or to
cooperators for field liberation. United States Department of Agriculture certified quarantine
facilities may range from a one- or two-room unit in an existing building to multi-room complexes
designed to meed specific quarantine needs, such as screening. A primary quarantine facility is one

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certified by the U. S. Dept. of Agriculture (APHIS) to receive direct shipments from foreign
sources which may contain live pest host material as well as the candidate natural enemies.
Secondary quarantine facilities may handle only those biological control shipments previously
processed through a primary quarantine laboratory or which are free of live, exotic pest species,
but which may contain hyperparasitoids of entomopathogens still to be screened out prior to
liberation.
Location and Utilities.--Primary quarantine laboratories are preferably located near a major
port of entry, such as an international airport. This proximity becomes increasingly important in
proportion to the number of shipments received throughout the year. The quarantine facility should
be physically located where water, electricity, natural gas or propane, road access, etc. are
available. A standby electrical generator powered by natural gas or propane is considered essential
for supplying power to selected circuits during power outages. Less obvious concerns are freedom
from windborne pollutants such as industrial smoke, dust and pesticide drift. A telephone for
communicating with collectors worldwide and between quarantine personnel and federal or state
regulatory personnel is essential. Ready access to a telex or FAX transmitting unit is highly
desirable.
Structural Design.--Details of the quarantine building itself are dictated largely by local
construction codes. Key features of all quarantine laboratories are the sealed nature of the rooms
or buildings, a vestibule system for entry and exit with positive closure doors, and a network of
filters through which air enters and leaves the facility. Leppla & Ashley (1978) show diagrams of
floor plants of five biological control quarantine facilities in the United States.
In order to reduce heat and cooling costs, the walls and ceilings should be well insulated.
To minimize transfer of heat in or out of the building, as well as to deter window breakage,
doubleglazed or thermopane windows are advised. The outer panes should be of tempered and/or
wirereinforced glass. If vandalism is considered a problem, the quarantine facility should be
encircled with a sturdy fence at least two meters in height. Added precautions include alarms that
signal unlawful entry and fire.
In the United States, the USDA APHIS Biological Assessment Support Staff (BASS)
approves the design of new and or modifications of established quarantines. Final certification
includes an onsite verification by an APHIS official to confirm compliance with structural and
operational criteria. Additional information on quarantine structural criteria may be obtained from
the USDA, Animal Plant Health Inspection Service (APHIS), Hyattsville, MD. 20782.
The structural criteria vary according to the kinds of beneficial organisms to be handled
and the risks posed to the environment:
Arthropods: The handling of beneficial parasitoids, predators and phytophagous arthropods
requires rooms with temperature, relative humidity, light and air exchange control systems to meet
the environmental needs of the different species. When such requirements cover a relatively narrow
range of environmental parameters, the heating, ventilation, air conditioning, air delivery systems
can be relatively simple. On the other hand, if several species of beneficial arthropods having
widely divergent environmental requirements must be handled simultaneously, each room will
require special controls to provide the variety of rearing conditions. The diversity of conditions
can be greatly increased by the use of individual temperature and environmental chambers.

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However, such units should be viewed as temporary at best because of their limited size and the
restricted numbers of organisms which can be produced in them. Quarantines for handling
phytophagous arthropods need one or more glasshouse containment areas directly accessible from
the quarantine laboratory.
Pathogens of Pest Arthropods: Information on pathogen quarantine construction and
operation may be obtained from the Biological Assessment and Taxonomic Support Group, Plant
Pest Quarantine, APHIS, USDA, Federal Center Bldg., Room 625, 6505 Belcrest Rd., Hyattsville,
MD 20782, Tel. (301) 436-5215.
Pathogens of Weeds: Melching et al (1983) discuss criteria for handling plant pathogens.
The buildings at the USDA/ARS Plant Disease Research Laboratory, Frederick, MD are sealed
and air conditioned via a tandem set of filters designed to remove particles larger than 0.5 lm.
Exhaust air is also passed through a third, deep-bed filter before discharge to the outside. Each
filter is capable of removing airborne bacteria or fungal spores. The air pressure within the unit is
negative to the outside atmosphere, so in the event of any leakage, the air would flow inward.
Waste water is sterilized before discharge from the area. Workers must shower before leaving the
laboratory, leaving their laboratory garments inside the quarantine. To minimize cross
contamination between study areas, the laboratory and greenhouse are divided into a series of
cubicles of varying size. Some of the work in progress at this facility is described by Bruckart &
Dowler (1986). Another description of a facility designed to contain weed plant pathogens is given
by Watson & Sackston (1985). A much simplified pathology quarantine, which incorporates all
the essential features of the above units, is that described by Inman (1970), who converted a room
in an older building into a functioning quarantine.
Nematodes: Certain species of nematodes attack a narrow range of introduced weeds.
Others are narrowly host specific, or pathogenic, on pest arthropods. Since beneficial as well as
phytophagous pest species of nematodes are closely tied to the soil environment, the safe handling
of imported species requires a quarantine facility capable of handling and sterilizing plants and
soil. In the unique Isolation and Nematode Quarantine Facility recently constructed at the
University of California, Riverside, soil containment is the primary concern. Security measures
include restricted entry, use of disposable shoe covers, arthropod control and stringent disposal
methods.
Equipment and Amenities.--The kinds of these items needed in quarantine will vary
depending on the class of organism and the studies to be made. Most are standard items in
entomological laboratories and include various dissecting tools, holding cages, micro habitat
monitoring equipment and illuminators. An olfactometer and video recording equipment for
studying the behavioral biologies and host relationships of organisms are also useful. Three main
categories of equipment are (1) hardware (cages, microscopes, temperature cabinets), (2)
reference items (literature files, identified voucher specimens, records, and (3) cleaning and
disposal equipment.
A quarantine laboratory to process incoming shipments solely for identification
requires only a handling cage, a microscope, identified reference specimens and other
identification aids, and containers for reshipment. When maintaining entomophagous or
phytophagous arthropods throughout their life cycles, hosts and host plants in various stages of

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development, several sizes of cages and special lighting, temperature and humidity controls are
necessary.
There is a need to avoid overstocking with equipment as quarantine space is often
limited. Ample enclosed storage within the facility should be provided to keep work surfaces
clean and free of clutter. Normally, equipment used in quarantine should remain inside the
facility.
Handling Cages: A cage design that has proven highly satisfactory in handling
arthropods for over 40 years measures ca. 55 cm high, 44 cm deep and 46-60 cm wide, and is
constructed of wood with a glass top and fine meshed cloth or screen on the backside. It has a
door in front equipped with paired, cloth sleeves which allow easy, yet effective escape proof
access to the cage's contents. One variation of this cage is of lucite plastic, the architecture speeds
the handling and recollection of large numbers of organisms by making them more accessible to
the paved openings at the front. The lucite cage's limited ventilation may allow moisture to
condense on interior surfaces if large volumes of fresh plant material are held, but presents no
problem when samples are processed quickly.
When processing an incoming shipment, the handling cage should be equipped inside
with a knife or scissors to open packages, tweezers, an aspirator and camel's hair brush to use in
the transfer or capture of the organisms, vials and cartons to hold the organisms, and paper and
pencil for recording the number of organisms and other observations. A CO2 unit for
anesthetizing organisms while in the handling cage or even refrigeration of the package prior to
placement in the cage will reduce their activity and facilitate identification, sorting and
processing.
Microscope: A binocular dissecting microscope (10-60X) and high quality illuminator
(fiber optic) is usually adequate for assessing the general conditions of quarantined arthropod
material, including identification and the sexing of specimens. A microscope mounted on a
pedestal with an adjustable arm is versatile and can also be used to view organisms on plants and
in cages. In addition a second microscope, such as one with phase contrast adaptation capable of
detecting entomopathogens may be needed to maintain healthy cultures (Poinar & Thomas 1978).
Identified Voucher Specimens: The availability of identified reference specimens to
compare with incoming material can greatly increase the speed and accuracy of workers as they
select specimens for release or further study. Often a single box or, at most, a small cabinet with
several trays of specimens, plus a file containing taxonomic keys and other aids will suffice.
Reference Files: The following references and information have proved useful in the
operation of the quarantine facility: Borer & Delong (1970), DeBach (1964), Clausen (1940),
Clausen (1978), King & Leppla (1984), Peterson (1959, 1960), Poinar (1977), Poinar & Thomas
(1978), Waage & Greathead (1987).
Lights: Daylight-fluorescent and the halide type plant growth lamps have proven
satisfactory for indoor plant culture and greenhouse containment areas that require supplemental
light. Heat from halide lamps may be of value in speeding plant growth, but harmful if excessive
heat buildup is undesirable. These lamps also require special wiring and circuitry. Time switches
are needed to simulate day length. The entry vestibules of the quarantine laboratory should be

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equipped with blacklight traps to attract insects which may have inadvertently gained entry to the
vestibule either from the outside or from within the quarantine handling area.
Temperature Cabinets: These are essential for experiments requiring closely regulated
temperature cycles. Units with good records of reliability are preferred to minimize repairs.
Temperature and humidity recording devices also will be needed.
Refrigerator: Cooling incoming shipments of arthropods to ca. 5°C not only extends
the longevity of the organisms but facilitates handling during transfer and identification.
Refrigerators may range from small, under-the-counter units, to the large double door, restaurant
models when space permits. The latter can accommodate large packages, and are especially
useful when large amounts of material is being handled. Household refrigerators with
thermostatic controls adapted to operate at predetermined minimum temperatures also may be
used. Care should be taken when selecting a temperature for long term holding periods to avoid
excessive mortality.
Refrigerated Room: A built-in cold room is considered desirable if large amounts of
plant material or hibernating immature arthropods are to be held under simulated winter
conditions. For temporary storage of packaged dormant material, portable refrigerated walk-in
units can be rented.
Carbon Dioxide: Judiciously used, CO2 anesthetization (one minute maximum) can
facilitate the handling of both entomophagous and phytophagous arthropods. However, Nicolas
& Sillens (1989) pointed out that CO2 narcosis may have adverse short- and long-term effects.
Passing the gas over ether can extend anesthetization time. Because CO-2 is heavier than air,
caution must be used to avoid build-up in the bottoms of open containers. Carbon dioxide is best
supplied with portable cylinders provided with pressure regulators that are under the direct
control of the personnel using them. Central CO2 installations which serve several stations on the
other hand often develop leaks, or gas is wasted by forgotten taps.
Tools: A selection of hand tools such as hammers, screwdrivers, pliers and small
ladders are all useful for cage and equipment maintenance. Flashlights and fire extinguishers are
essential for emergency purposes.
Vacuum and Pressure Pumps: Such devices are useful when collecting large numbers
of living specimens and with olfactometer experiments. Air pressure is handy for cleaning cages,
aerating hydroponic tanks of aquatic plants, etc. Positive and negative air supplies can be from a
central source in the building, or provided by portable units.
Pest Control: Ants, whiteflies, aphids, spider mites, etc. frequently pose problems to
plant and insect cultures in quarantine. Control by nonpesticide methods is preferred (e.g., light
traps, sticky boards, soaps, biological control agents, handpicking of infested leaves).
Commercial insectaries and some farm and garden supply stores are sources of biological control
agents (Anon. 1989, Bezark 1989). If chemical sprays are to be used, a unit for confining the
treatments to the plants and to exhaust odors and drift outside the quarantine will be needed.
Insecticidal dusts should not be used. Boric acid powder can be used for cockroach control
(Ebeling 1978). [See section on contaminants: ENT229.17].
Cleaning and Disposal Equipment: Vacuum cleaners, brooms, sponges, mops and other
janitorial equipment are necessary. Most containment facilities are equipped with pass through

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steam autoclaves which allow direct removal of treated materials from quarantine without the
possibility of recontamination. For treatment of small amounts of material, regular household
ovens or specially constructed electrically heated chests may supplement the autoclave in purely
arthropod handling facilities. The steam autoclave is preferred if soil and other compacted
materials are used in containment. Pathogen infected waste material should be sterilized at 100°C
for sufficient time to permit adequate penetration of heat.
The usefulness of microwave ovens is questionable. Hertelandy & Pinter (1986a,b)
discussed the use of microwaves to control stored product pests, but the effectiveness in killing
spores of certain pathogens is doubtful.
Under no circumstances ought carpets be used to floor a quarantine laboratory, as the
fibers retain material that can infest cultures, and sterilization is practically impossible.
Records: A file cabinet is useful for keeping equipment operating instructions,
quarantine handling records, taxonomic keys, pertinent literature, correspondence and
appropriate phone numbers. The cabinet may be kept in a quarantine anteroom, but should be
readily accessible to workers. Computer equipment greatly facilitates record management and
the exchange of information among laboratories and regulatory agencies.
Communication Units: A telephone communication capability between personnel
working in quarantine and elsewhere is essential. An intercom system permitting nonmanual
operated response should minimize worker interruption.

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B. Use of Resistant Host Plant

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C. Destruction by Cultural Management or Direct Removal of Infected/ Affected Hosts
Cultural method of pest control
These methods are aimed either at reducing the sources of inoculum or at reducing the
exposure of plants to infection. Its primary objective is the prevention of pest damage and not the
destruction of an existing and damaging pest population.
1. Good soil preparation
This is the first important element in pest control strategy. A healthy soil means healthy plants
which are relatively more resistant to pests. A soil rich in humus hosts a wide variety of beneficial
microflora that trap nematodes and destroy or keep in dormancy disease organisms, thereby
encouraging beneficial insects.

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2. Use of indigenous varieties
Traditional varieties are hardier and relatively more resistant to pests. They can withstand harsh
environmental conditions better than modem hybrids.
3. Pest control through the use of mesh screen (nylon nets)
Younger plants are usually preferred by insects and they suffer significantly from such attacks when
compared to older plants. Therefore, a single netting over the plants during the first 3045 days of
their growth can reduce pest damage. Also, the net helps diffuse sunlight thereby improving the
quality of some vegetables. Finally, the net breaks the impact of raindrops thus (i) reducing physical
damage to the plant and (ii) reducing soil erosion from the beds.
4. Roguing or Pruning
Removal of diseased plants or plant parts prevents the spread of microorganisms to uninfected
areas.
5. Intercropping with aromatic herbs
Several types of odorous plants can be grown together with the main crop to repel insects. The
following are some examples:
Allium cepa (onion)
Hyptis suaveolens (bush-tea bush)
Allium odorum (leek)
Mentha cordifolia (mint)
Allium sativum (garlic)
Ocimum basilicum (sweet basil)
Artemisia vulgaris (mugwort, worm wood)
Ocimum sanctum (sacred basil)
Coleus amboinicus (oregano)
Tagetes spp. (marigold)
6. Encouraging insect predators
Pests can be controlled by their natural enemies. By growing a variety of flowering plants,
specifically those belonging to Umbelliferae family, such as, fennel (Foeniculurn vulgare) and
celery (Apium graveolens), insect predators will be attracted to stay in the garden. These beneficial
insects feed on pests, keeping the pest population below economic injury level.

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7. Multiple cropping
This provides genetic diversity to minimize pest increase. Variation in susceptibility among species
or varieties to a particular disease is great. Given abundant hosts of a single species or variety, a
pest could easily be spread from host to host. When the number of hosts declines, the pest incidence
will also decrease for lack of necessary food for the organism.
8. Crop rotation
This is a practice of following a crop susceptible to a pest by a resistant crop. There is no build-up
of the organism to a high level since the growth cycle of the organism has been broken.
D. Conservation and Augmentation
Conservation
Pesticides kill beneficial predators, parasites and pathogens as well as pests, and can cause
outbreaks of secondary pests or rapid resurgence of pests that were initially suppressed. Using
non-chemical control methods, or pesticides which kill only the target pest, protects natural
enemies. Some easily seen predators are spiders, lacewings, lady beetles, ground beetles, rove
beetles, syrphid flies, flower flies, hover flies, true bugs (including minute pirate bugs, big-eyed
bugs and damsel bugs), predatory mites and even fire ants. However, many important natural
enemies are rarely seen, such as parasitic wasps and flies (more than 8,500 species), nematodes
and pathogenic bacteria and fungi.

Conservation-avoid harming natural enemies complex. Provide nesting sites, alternate hosts,
foods, refugia, over wintering sites, and other needs for natural enemies.
Conservation biological control is the implementation of practices that maintain and enhance the
reproduction, survival, and efficacy of natural enemies (predators, parasitoids, and pathogens) of
pests.

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Figure 1.Nesting sites and flowering plants
Augmentation
The release of natural enemies (predators, parasites and pathogens) to control pests is a
type of biological control called augmentation. This approach uses commercially available species
that are applied in a timely manner to prevent population increases, or to suppress a pest population.
Natural enemies can be released all at once or over time to suppress pests or keep their
numbers low. Also, the environment can be enhanced to favor natural enemies. Although research
has shown that releases of natural enemies can be very effective in greenhouses and interiorscapes,
outdoor releases are affected by unpredictable environmental conditions. Furthermore, if a second
pest is unaffected by the released organism, pesticides used to control the second pest often
eliminate the natural enemy of the first pest. Specific recommendations for Texas are still being
developed.
The application of microorganisms in a manner similar to conventional pesticides is a type
of augmentation. These products are referred to as “microbial insecticides.” Several products
available contain varieties of the bacterium, Bacillus thuringiensis, which controls certain
caterpillars, beetles and flies but does not affect other arthropods. Microbial insecticides are
relatively slow acting and are most effective if applied when pest numbers are low and pests are in
early stages of development.

Examples of augmentive biological control products:

Commercial products available for use in augmentive biological control include microbial
insecticides containing living pathogens (bacteria, fungi and viruses) and multicellular animals
(predators, parasites and nematodes). Other products occasionally used with biological control
agents include synthetic honeydew, flowers to attract and conserve beneficial insects in and around
pest-prone or pest-infested sites, and traps using colors or scents as attractants.

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Best use for augmentive products

• Purchasing and releasing natural enemies for pest suppression is a rapidly developing
technology but there is still much to be learned to assure effective use of these products. Results
are often difficult to evaluate and can be inconsistent because of differing conditions (e.g.,
environmental, meteorological, etc.). Natural enemies are living and their behavior under different
environmental conditions can influence the degree of pest control. Cost-effective use of augmentive
releases requires an understanding of the pest(s), natural enemies, economic goals and the
environment. Commercial uses often demand intensive monitoring or scouting of the cropping
system.

• Augmentive releases are meant to reduce populations at points in time. Releases at low pest
densities are more effective than attempts to reduce high pest densities. Action levels or economic
thresholds for release of natural enemies and effective release rate(s) have often not been
established through scientific research.
• Timing of the release of natural enemies is critical since most require some time to affect
the pest population. In addition, many natural enemies attack only certain life stages (e.g., egg or
larval stage) of the pest. Multiple releases may also be necessary to maintain pest suppression.

• Biological control using parasites is generally pest-specific. When multiple pests occur (e.g.
aphids, thrips, plus beetles), natural enemies are needed for each pest. In cases where natural
enemies are unavailable for augmentation, use of a selected pesticide that spares other natural
enemies may be necessary.

• Environmental conditions change dramatically and outdoor releases of natural enemies can
be negatively affected by high winds, rain, hot or cold weather and other insects in the ecosystem
(e.g., red imported fire ants). These factors are often unpredictable and result in erratic results from
releases. Release of appropriate natural enemies in greenhouses and interiorscapes often provide
more consistent results.

• Insecticide residues on the crop or site, or insecticide drift from adjacent areas, can remain
toxic to natural enemies long after the pesticide was applied. Residues should be mitigated prior to
releases.

What support can I expect from the companies selling these products?
Companies selling products and promoting their use should provide the consumer with
directions on how to use their products, and support their claims of product performance. Insectaries
and brokers, the companies producing and marketing parasites and predators, assure the delivery
of viable natural enemies of the stated species or strain. They usually do not guarantee results from
releases of these biological control agents even when used as directed. Although researchers and

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Extension faculty at The Texas A&M System are involved in evaluating some of these products,
suggestions for their most effective use are still being developed.

Are these products regulated by any laws?

Microbial insecticides (bacteria, fungi, viruses) are regulated like pesticides by the
Environmental Protection Agency (EPA) under the Federal Insecticide, Fungicide and Rodenticide
Act (FIFRA). Multicellular animals (arthropod predators, parasites, nematodes, etc.) are NOT
registered or regulated by the EPA under FIFRA. Complaints regarding product performance can
be reported to the Federal Trade Commission (FTC).
The user of purchase-and-release natural enemies must be aware of legal and biological
limitations of augmentive biological control methods. Just restricting frequent use of broad
spectrum insecticides often will allow a diverse group of naturally occurring beneficial organisms
to survive, sometimes profoundly impacting pest population densities. As the cost of natural enemy
products continues to decrease and delivery systems and methods are improved, the economic
feasibility of using these methods in commercial pest control will undoubtedly improve.

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E. Integration with other Control Tactics for Pest Management

Practical Uses of Biocontrol:

In private, commercial settings, the most common use of biocontrol is the augmentation method,
where natural enemies are purchased and released. Greenhouses are the most common place
where this method is used. However, because the natural enemies and pests are ALIVE and form
dynamic populations, success of biological control in suppressing pests requires attention to
certain important factors.
• Get the right enemy for the pest species.
• Release them at the right time in the pest population cycle.(Start when pest populations are
LOW so enemies have a chance!)
• Environmental conditions must be favorable (weather / temperature / moisture).
• Food sources must be present for the natural enemy (adults or larvae in some cases).

Advantages and Limitations of Augmentative Biological Control Advantages

of augmentative biocontrol include:

• Reduced reliance on chemical pesticides.

• On-going suppression of pests over time (vs. repeated spraying) They reproduce
themselves and keep working.
• Can you think of others?
Limitations of augmentative biocontrol include:
• More knowledge is required to use successfully.

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 • More attention to detail is required.
• Results are not immediate - it takes awhile before pest populations decline.
• Cost is usually higher than use of chemical pesticides

Figure 1. The relationship between biological control and other strategies for pest management.
The principal aim of the diagram is to place biological control in its larger IPM context. Placing
the order of the boxes as shown helps draw parallels between cultural control (agronomic practices
which target the pest itself) and conservation biological control (agronomic practices targetting the
pest’s natural enemies). Similarly, the close relationship between the use of conventional pesticides
and botanical pesticides (in biorational chemical agents) can be seen. We consider the use of
transgenic plants to be included in the box ‘host plant resistance’. (Modified from Nordlund,
1996)

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