الفهرس 
Taxonomy & Classification
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Abstract
The present work was carried out to study the histopathologicai lesions that occur in heterophyiasis on the different organs of the experimentally infected dogs subsequent to consumption of infective encysted metacercariae. Also to evaluate the therapeutic effect of mirazid .(myrrh or Commiphora molmol) and propolis (a natural bee product) compared to praziquantel in treatment of animals experimentally infected with heterophyids.
The present study was conducted on twenty eight dogs which were divided into two groups as follows:
©Groiif) |: Consisted of 8 dogs which were further subdivided into the following subgroups:
- Subgroup la: 4 dogs infected with approx. 500 encysted metacercariae of heterophyids. 2 dogs were sacrificed at 2 and 3 weeks after infection. This group was subjected to histopathologicai examination of different organs and to tissue digestion.
~ Subgroup lb (negative control group): 4 dogs were left without infection as negative control.
# i^rouj) jj : Consisted of 20 dogs infected with approx. 500 encysted metacercariae of heterophyids and were further subdivided into the following subgroups:
- Subgroup TIa (positive control group): It was formed of 5 dogs which
were left without treatment.
- Subgroup lib: It was formed of 5 dogs treated with PZQ at a dose of 25 mg/kg for 2 consecutive days.
- Subgroup lie: It was forrnedxrf 5 clogs treated with MZ at a dose oi 500 mg/kg/day ior 5 consecutive days.
- Subgroup lid: It was formed of 5 dogs treated with propolis at a dose of 350 mg/kg/day .for 5 consecutive days.
In order to evaluate the efficacy of the tested drugs the following was done:
1 -Farasitoiogieai parameters:
A- Egg count per grain of stool.
B~ Worm count recovered from the small intestine.
2~Histopathoiogica,l examination: Samples were taken from small
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intestines, mesenteric lymph nodes, lungs, hearts, kidneys, livers and brains and stained with Hematoxylin and Eosin stain. Samples from small intestines and livers were also stained with Masson’s Trichrome technique.
3- Histocheinical examination: samples were taken from small intestines and livers and stained with Periodic Acid Schiff
4- BIood examination:
A- Determination of total leucocytie count.
B~ Determination of leucocytic cell differentials.
C~ Determination of serum biochemical values (aspartate aminotransferase (AST), alanine aminotransferase (ALT), total protein (TP), albumin, blood urea nitrogen (BUN) and creatinine.
from the present study, the following results were obtained;
❖ Egg count per gram of stool (EPG):
The mean ±SD was found to be significantly lower in clogs treated with PZQ and MZ than control +ve one in the V1 day after treatment. Trie mean ±SD was significantly lower in PZQ than MZ group.
At the 3ra and 5tn days after treatment, there were significant lower levels of EPG in each of PZO. MZ and propolis groups than control ~f vc one, There was complete disappearance of eggs in stools of PZQ treated dogs.
Worm count recovered from the small intestine (WRR):
There were significantly lower WRR in each of PZQ, MZ and propolis groups than control +ve one. The mean ±SD was significantly lower in PZQ than MZ and propolis groups.
❖ H is to pa t h o I og ie a I examination:
* Intestine:
In i n fee ted non treated groups, after 2 weeks, the affected villi showed exfoliation of the epithelial layer. Increased goblet cells were detected in diffuse manner all over the lining mucosal epithelium, associated with edema and inflammatory cells infiltration, mainly lymphocytes, in the underlying lamina propria of the villi. Congestion and dilatation were noticed in the blood vessels of the sub mucosa. Also, there was lymphoid hyperplasia of Peyer’s patches. After 3 weeks, all previous findings became more severe. Some villi showed complete exfoliation of the epithelial layers & broad surface of mucosa was covered with necrotic debris. Severe destruction or loss of villi in some portions of small intestine was also noticed. In PZQ group, the epithelial lining of the villi appeared nearly normal. In MZ group, the epithelial lining showed minimal shedding in some \ dh together with e-A-reased goblet ceils, inflammatory cells infiltration in the lamina propria of the
villi and lymphoid hyperplasia in Peyefs patches. In propolis group, the
it
epithelial lining showed minimal shedding in the villi; some areas showed intact epithelium and increased goblet cells. There were inflammatory cell infiltrations in the lamina propria with lymphoid depletion in Peyefs patches. By Masson’s tri chrome technique, the connective tissue (C.T’) was demonstrated as very thin collagenous fibers in the core of the villi and sub mucosa in the sections of control normab intestine, The sections of intestine of infected non treated dogs showed submucosal fibrosis. None of the treated groups showed fibrosis,
* Lymph node:
In infected non treated groups, after 2 weeks, there was lymphoid hyperplasia in the cortex. After 3 weeks, there were exiravasated red blood cells and haemosiderosis in the medullary portiopf Some follicles in the cortex showed lymphoid depletion associated with sub capsular and medullary oedema. In PZQ group, no histopathologicai findings were observed. In MZ group, the sub cortical and medullary sinuses were dilated and filled with mononuclear cells. In propolis group, some lymphoid follicles showed depletion.
* Liver:
In infected non treated groups, after 2 weeks, there were degeneration of hepatocytes (in the form of vacuolation of hepatocytes), moderate congestion in the central vein, portal tract and blood sinusoids and activation of kupffer cells. After 3 weeks, the hepatocytes showed degenerative changes associated with severe dilatation and congestion of the central veins and sinusoids. Small focal necrotic areas were observed.
The bile ducts showed hyperplasia with fibrosis in the porta.! area. In PZQ group, the liver appeared nearly normal In MZ group, the picture improved but still there were few areas that showed degeneration in hepatocvies, activation of kupfier cells and dilatation in sinusoids. Neither fibrosis nor foci of hepatic necrosis were seen in this group, hi propolis group, there were proliferation of kupffer cells and minimal changes in hepatocytes. By MassoiPs trichrome technique., there were periportal and perivascular fibrosis. No fibrosis could be detected in all treated groups.
* Kidney;
In infected won treated groups, after 2 weeks, there were degenerative changes in the epithelial cells lining the renal tubules in the form of pyknosis of nucleus and/or vacillation. The glomeruli showed variation in size and some of them were degenerated. After 3 weeks, the kidney exhibited the same changes besides there were congestion and haemorrhage in renal tubules. Peri glomerular inflammatory cells infiltration and fibrosis were also noticed. In PZQ group, sections in the kidney showed great restoration of the normal structure with glomeruli and tubules, more or less like the normal. In MZ arid propolis groups, the kidney sections showed restoration of the normal glomeruli and tubules in some areas, while other areas showed minimal degenerative changes in the tubules and atrophy in some glomeruli,
* Lung;
In infected non treated groups, after 2 weeks, no changes could be detected. After 3 weeks, there was thickening of the wall of the alveoli by inflammatory cells mainly mononuclear cells (macrophages and lymphocytes). In few areas there was rupture of the wail of the alveoli.
Minimal congestion in perialveolar blood vessels vv o noticed. In PZQ, MX and propolis groups, no histopathologicai changes could be delected.
* Heart:
In infected mm treated groups, after 2 weeks, no changes could be detected. After 3 weeks, myocardial muscles suffered from degeneration and necrosis in which the muscle bundles appeared swollen and separated, loss of striation and nuclear degeneration accompanied by mild lymphocytic aggregations. There were congestion and haemorrhage in some areas. In PZQ„ MZ and propolis groups, no histopathologicai changes could be detected.
* Brain:
In infected non treated groups, no histopathologicai changes could be detected neither after 2 weeks nor 3 weeks.
♦♦♦ IlistoehemicaJ examination: Intestine:
In infected non treated groups, intestines showed faintly stained brush borders of the absorptive columnar cells, while goblet cells appeared as deeply purple coloured dots. In PZQ group, there was marked restoration of glycogen in the brush borders of the absorptive columnar cells simulating normal. In MZ group, there was tendency towards restoration of glycogen inclusions reflected by moderate PAS reactivity in the brush borders of the absorptive columnar cells. In propolis group, there was tendency towards restoration of glycogen inclusions reflected by minimal to moderate PAS reactivity in the brush borders of the absorptive columnar cells.
* Liver:
In livers of infected non treated groups, there was marked depletion of glycogen contents reflected by weak PAS reactivity as compared with control group, In PZQ group, there was glycogen staining of marked degree in most hepatocytes. The picture was more or less simulating the control. In MZ group, there was a tendency towards restoration of glycogen contents reflected by moderate PAS reacti vity. In propolis group, there was a tendency towards restoration of glycogen contents reflected by minimal to moderate PAS reactivity.
Blood examination:
Concerning total leucocytic count M0Jcell/uL, the mean ±SD was significantly higher in control +ve group than control ve group. The mean ±SD was significantly lower in PZQ. MZ and propolis groups than control +ve group.
Regarding neutrophilic and lymphocytic count MO’cell/pL the mean /
±SD was significantly higher in control +ve group than control -ve group. In PZQ, MZ and propolis groups the mean ±SD was significantly lower than control +ve group.
Regarding monocytes and eosinophilic count, there was no significant change between control +ve as compared with control -ve group. Also, between control +ve group and other treated groups.
Regarding ALT, AST, BUN and creatinine, the mean ±SD was significantly higher in control +ve group than control -ve group. While, there was no significant difference in the mean values of TP and albumin between control +ve and control -ve groups. There were significantly lower mean values of ALT, AST, BUN and creatinine in PZQ group than control we group. While, there was no significant difference in the mean values of TP and albumin.
There were significantly rower mean values os ALT, AS i ami creatinine in MZ group than control +ve group. While, there was no significant difference in the mean values of BUN. TP and albumin. There were significantly lower mean values of ALT and AST’ in propolis group than eon f ro! +ve group. Whi le, there was no significant di fference in the mean values of BUN, creatinine, TP and albumin.
❖ Scanning electron microscopic study:
No worms were detected in the small intestines of PZQ treated
dogs and so no SEM could be done, On the other hand, the adult
heterophy.ids obtained from the intestines of dogs treated with MZ,
appeared contracted with the tegument showing irregularity and
discontinuity. Also the tegument showed more or less loss of the spines
with few vesicle formations. In propolis group, the worms showed
complete loss of tegumental spines. Erosion of the surface had occurred
to such extent that in some areas no tegument remained and even some
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areas showed only a mass of fibrous structures. Some areas were observed to be covered with debris which may be consisted of numerous blebs.
the present study demonstrated that;
L Treatment of heterophvid-infected dogs with PZQ resulted in complete disappearance of eggs from stool on the 3IU day after treatment. While in case of using MZ and propolis, there were decrease in EPG alone the davs of treatment but not eradicated completely.
2. PZQ showed higher efficacy than MZ and propolis as it caused complete eradication of worms from intestines.
3, Heterophyid Infection caused significant changes in all examined