Acta Parasitologica Globalis 2 (2): 34-39, 2011

ISSN 2079-2018
IDOSI Publications, 2011

Advanced Approach in Differentiation Study in

Hymenolepis nana and H. Diminuta by Scanning Electron Microscopy
1

M.S. Mahmoud, 1A.H. El Namaky, 1O.M. Kandil, 1N.A.T. Allam, 2A.A. Hasan and 1H.M. Ashry
1

Department of Parasitology and Animal Diseases, National Research Center, Giza, Egypt
Department of Parasitology, National Hepatology and Tropical Medicine Research Institute, Cairo, Egypt

Abstract: Scanning electron microscopy (SEM) succeeds in revealing novel microtopographical features of the
host-parasite relationship, as well as proving invaluable in helminthes taxonomy. Hymenolepis nana and
H. diminuta are commonly maintained in laboratory rodents and used in many experimental model systems of
tapeworms infections. In the present work, the length of worm of Hymenolepis nana and H. diminuta were
approximately up to 1-3 cm 1mm and 6 cm 3.5 mm, respectively. The SEM revealed that the scolex of H. nana
and diminuta appears roughly rectangular and is provided with an armed suker and rostellum. The rostellum
is provided with hymenolepid hook in H. nana and an unhooked rostellum in H. diminuta. These suckers
occupy the corners of the rectangular scolex. Fine structural of the tegumental surface of both H. nana and
H. diminuta is densely covered with microtriches which are of the same shape and were never seen to
be branched. SEM of intestinal mucosa of infected rat with H. nana revealed alteration in villous architecture.
The purpose of this study is to use SEM technique as advanced approach for differentiating between H. nana
and H. diminuta.
Key words: Scanning electron microscopy % SEM % Rat % H. nana % H. diminuta
INTRODUCTION

Few literature are found concerning the surface of

Hymenolepis by SEM. Fine structural studies of the
tegumental surface of cestodes have demonstrated that
the surface cytoplasm is extended as microtriches,
consisting of cylindrical cytoplasmic bases capped by
dense structures termed "shafts" [9].
SEM studies of cestode surface have shown that it
provides as a suitable technique for obtaining information
regarding the three dimentional relationship of surface
structures. Berger and Mettrick [10] reported
polymorphism in the microtriches from H. diminuta,
H. microstoma and H. nana by SEM. They suggested that
the microtriches played a role in locomotion of these
organisms within the host's gut.
Several workers have examined the pathological
effects of larval cestodes in man and mammals [11, 12].
Martin and Holland [13] examined by SEM the intestinal
morphology of rats given 1, 10, 100 cystecercoids of H.
diminuta, they found that the presence of this tapeworm
cause extensive villous atrophy and fusion. Also, they
observed colonization of the upper region of ileum
by long filamentous bacteria in rats infected with
H. diminuta.

Hymenolepis nana and H. diminuta are the most

common cestodes in humans, domestic and wild rats,
mice and human [1]. The body of tapeworms is
differentiated into an expanded anterior end, scolex,
bearing hold fast organs, un-segmented neck and a
strobila which is usually very elongated and
ribbon-like, ranging from few millimeters to 10-20 m
or more, divided into 4-4000 segments or proglottides
[2-4].
The cestode tegument is, however, unique in bearing
microvilli (microtriches) apparently serving to increase the
area of absorptive surface. Each mature segment has a
complete set of male and female genitalia. Immature newly
formed segments are nearer to the neck, mature segments
are more posterior and those at the end are gravid
segments [1, 5].
Scanning electron microscopy succeeded in revealing
novel microtopographical features of the host-parasite
relationship, as well as proving invaluable in helminthes
taxonomy and in assessing the efficacy of test substance
in drug screens [6-8].

Corresponding Author: Mona Said Mahmoud, Department of Parasitology and Animal Diseases, Veterinary Research Division,
National Research Centre, Postal code: 12622, Giza, Egypt.

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Acta Parasitologica Globalis 2 (2): 34-39, 2011

The purpose of this study was to use the SEM

technique as a comparative diagnostic tool for
morphological differentiation between some surface
topography features of H. nana and H. diminuta. As well
as the pathological effects in intestinal morphology of
rats infected H. nana.

aminomethane meleat
solution
pH 7.4 [15].
The samples were rinsed for an hour in a fresh
phosphate buffer salin (pH.7.4) and dehydrated in an
ascending series of ethanol. The specimens were
placed in liquid CO2 then mounted on metal stubs
coated with gold and examined with a Joel SEM
(Joel Corp., Mikaka, Japan) operated at 15 Kv. This work
was carried out in central laboratory of national
research center, Cairo, Egypt.

MATERIALS AND METHODS

Parasites Samples Collection: H. nana eggs were
obtained from infected humans in National Hepatology
and Tropical Medicine Research Institute. Approximately
2000 H. nana eggs were inoculated into 5-week-old male
white mice [14]. Adult worms were dissected from the
small intestine approximately 14 days post-inoculation.
While hymenolepis species worms were obtained from
naturally infected Rattus norvegicus from Abu Rawash
area, Giza, Egypt. Rats were killed by cervical dislocation
and entire small intestine was removed from the gut and
rinsed in physiological saline.

RESULTS
SEM of H. Nana: The length worm is approximately 1-3
cm 1mm. The scolex appears roughly rectangular and is
provided with a comparatively large hooked rostellum.
The rostellum lies at the center of the scolex and is
provided with pointed and posteriorly directed hooks.
The neck and then is followed by gradually immature and
mature proglottides. Genital atria are unilateral; each has
a rounded opening and is surrounded with ordinary
microtriches. The entire tegumental surface is densely
covered with microtriches of uniform size and density.
The microtriches were overlapped that it was difficult
to observe their bases. Gravid segments are more
posterior (Fig1 a-e).

Scanning Electron Microscopy (SEM): The worms and

pieces of the intestinal tissue (which opened
longitudinally) were fixed at 4C for 24 hr in 5%
glutaraldehyde in
0.1
M tris-hydroxy methyl

Fig. 1:
a)
b)
c)
d)
e)

Scanning electron micrographs of H. nana:

Adult H. nana with scolex, neck and entire strobila (x 75).
The rostellum with a ring of hymenolipid hooks and muscular border covered with distinct microtriches (x1000).
Anterior part of H. nana (x180).
Segments with unilateral genital pore (x 300, x 2000).
Segment covered with distinct microtriches (x120, x 15000).
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Acta Parasitologica Globalis 2 (2): 34-39, 2011

Fig. (2): Scanning electron micrographs of H. diminuta:

a) Adult H. diminuta (x20).
b) Scolex provided with an armed sucker and rostellum, the muscular borderof sucker were covered with distinct
microtriches. (x450).
c, d) The genital pore was covered with microtiches (x4500, x 15000).
e) Integument of gravid segments (x40).
f) Excretory pore (x110).

Fig. 3: Scanning electron micrographs of the intestine of mice infected with H. nana:
a) Anterior region of the illum of mice show that the villi are reduced in height and fused to give irregularly
shaped ridges of tissue (x 2300).
b) Region of the ileum of mice with numerous long filamentous bacteria on the surface of mucosa associated with
the presence of worms (x65).
c, d) Damaged and fused epithelial cells which may the scolex of H. nana becomes attached to the wall of intestine
(x 800, x240).
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Acta Parasitologica Globalis 2 (2): 34-39, 2011

SEM of H. diminuta: The length of worm is approximately

up to 6 cm 3.5 mm. The scolex of H. diminuta is
provided with an armed sucker and rostellum as observed
by SEM is presented in. The suckers located bilaterally on
the dorsal and ventral surface. The suckers occupy large
portion of the available surface area, thus suggesting their
important functional role. The rostellum lies at the center
of the scolex and has no hooks. Genital atria are unilateral
and, similar to H. nana and surrounded with ordinary
microtriches. The scolex and proglottides are all covered
with posteriorly directed microtriches of uniform shape
(Figs. 2a-d).

H. nana and H. diminuta, by SEM. He found that the

scolex, suckers, rostellum and strobilia are all covered with
dense populations of microtriches which are all of the
same size and shape except on the scolex and suckers
which are slightly more slender than those on the
strobilia. Presently study the microtriches appeared of
uniform shape and length along the strobilar surface and
were never seen to be branched.Moreover, Abouzakham
et al. [21] studied the cytological structure of tegument of
H. nana by SEM. They found that the surface of H. nana
is covered with dense populations of microtriches occur
on scolex proper, suckers and strobilia; with an average
density of 20/Fm2. On the other hand, Jha and Smyth [22]
reported polymorphic microtriches on the rostellum of
Echinococcus granulosus. In the present study neither H.
nana nor in H. diminuta this polymorphism was
observed. Andersen [23], using SEM compared the
surface structure of adults and larvae of three species of
Diphyllobothrium and reported the presence of regional
difference in microtiches appearance in the larva, but not
in the adult cestodes. Also, polymorphism was observed
in Diphyllobothrium.
In the present study the intestinal mucosa showed
alterations in villous architecture, formation of shallow
grooves and along filamentous bacteria also were
observed. This runs in full agreement with Martin and
Holland [13] who examined the intestinal morphology
of rats given cysticercoids of H. diminuta by SEM.
They found that the presence of this tapeworm cause
extensive villous atrophy and fusion. The most extreme
changes in mucosal architecture were observed adjacent
to the mature proglottides of the worm. Bacterial over
growth of the small intestine has occasionally been
reported [24] but the factors influencing this change in
habitat are not fully understood. Tannouk and Savage
[25] suggested that dietary conditions and stress may
affect bacterial colonization of the intestinal tract. Mettrick
[26] and Smyth [27] showed that H. diminuta has a high
rate of carbohydrate fermentation, which leads to the
release of several acids, including lactic acid and it is
possible that the presence of these substances may alter
the physiochemical conditions along the intestine and
thus facilitate colonization of the more anterior regions of
the gut by these microorganisms. Gross alterations of
intestinal mucosa ha observed in association with many
nematode parasites. Nippostrongylus brasiliensis in rats
[28,29], Trichinella spiralis in guinea pigs [30] and
Ascaris lumbricoides in man [31] all cause varying
degrees of villous atrophy and fusion. Flattening of
the villous surface has also been observed in mice
harboring H. microstoma [32].

SEM of Intestine Removed from Infected Mice with H.

nana: The intestine had showed some degree of villous
atrophy and fusion. The villi were reduced in height and
fused to form long ridges of tissue which gave a rippled,
wave-like appearance. Long filamentous structures
identified as bacteria were observed in large numbers, the
bacteria appeared to be attached to the entrocytes by
hold fast or attachment segments. However, small
depressions were occasionally observed in the mucosa of
infected animals and it is tentatively suggested that these
represent the attachment site of Hymenolepis. A shallow
groove was also seen in association with triangularshaped protuberances. Villi in the center of the groove
were further atrophied (Fig.3 a- d).
DISCUSSION
Scanning electron microscopy (SEM) succeeds in
revealing novel micro topographical features of the
host-parasite relationship, as well as proving invaluable
in helminthes taxonomy. H. nana and diminuta are
cosmopolitan in their distribution, but the former is more
common in warmer climates, while the latter is a common
parasite of rats and mice and only occasionally infects
man [16-18].In the present study, the length of full
strobilia of H. nana and diminuta were approximately
up to 1-3 cm 1mm and 6 cm 3.5 mm, respectively.
The SEM revealed that the scolex of H. nana and
diminuta appears rectangular and is provided with a
comparatively large rostellum and suckers. The rostellum
lies at the center of the scolex and is provided with
pointed and posteriorly directed hooks in H. nana and
has no hooks in diminuta. This is in agreement with the
view held by Rothman [19] and Ubelaker et al., [20]. In the
present work, the fine structural of the tegumental surface
of both H. nana and H. diminuta is densely covered with
microtriches. This is in agreement with Ashour [9]
who studied the surface ultra-structure of two cestodes,
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Acta Parasitologica Globalis 2 (2): 34-39, 2011

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