Pathology of Heterakis gallinarum in the ceca of naturally infected chicken (Gallus domesticus)

Present study was aimed to examine the pathology of nematode (Heterakis gallinarum) in the ceca of local chicken (Gallus domesticus). A total of 90 chickens (Gallus domesticus) were randomly purchased from different villages and local markets of District Hyderabad, Pakistan. Birds were dissected and different parts of intestine were recovered. Out of 90 birds 83 were found infected with a variety of helminth parasites. The sex wise nematode Heterakis gallinarum was collected from cecum of infested chickens. Tissues from infected ceca were fixed in Bouin’s fluid for 24 hours for histopathological examinations using routine histological techniques, 6 μm thick sections were stained by using principal staining procedure of hematoxylin and eosin (H&E). The parasitic Heterakis gallinarum prevalence was 20% of infected chickens. Histopathological findings revealed sever damage of cecal architecture, necrosis of lamina propria, destruction of intestinal glands. Infected cecum was showing muscular alterations like vacuolation and tearing of muscles. Further investigations are suggested to reduce the risk factors of nematode infestation in birds which will ultimately minimize further losses.


Introduction
Poultry farming is one of the most important sectors of agricultural industries throughout world providing high energy food meat & eggs and they are considered rich energy nutritional source of proteins [1]. Poultry farming is an essential business sector play a significant role in enhancement of the social-economic status of rural communities. Annually more than 50,000 million chickens are reared for their meat [2, 3]. In developing countries like Pakistan majority of families from rural communities keep chickens flocks in free range scavenging system. Due to the free range scavenging system, the chickens are susceptible to parasitic infestations from their habitats, particularly gastrointestinal parasitic infections which results heavy financial losses. These parasitic infestations are causing severe health problems of growth, egg laying performance and mortality. Beside the viral and bacterial infection poultry birds have also been affected by parasitic infections Keeping in view of significant medical and socio economic importance of poultry sector, present study was carried out to evaluate the intestinal damage caused by nematode parasite Heterakis gallinarum in naturally infected chicken. So that to expand knowledge of pathogenicity and possible attention may be given towards the management of free ranges chicken.

Materials and methods
Adult free range local chickens (Gallus domesticus) were purchased from local markets and different villages of Hyderabad, Sindh, Pakistan. The obtained birds were slaughtered at parsititology laboratory, Departement of Zoology, University of Sindh, Jamshoro, Pakistan. There after birds were dissected for the collection of gastrointestinal parasites and searching of helminthes was carried out by stereoscopic microscope and magnifying glass by applying method of Dharejo [13]. The recovered parasites from the caecum of each dissected bird were kept in normal saline solution. A heated mixture of 70% ethanol and 5% glycerin was used for stretching and killing of parasites then same parasites were shifted in 70% ethanol and glycerin by 1:1 ratio for further process. Parasites were mounted in glycerin for identification. For histopathological studies the affected pieces of parasitized ceca were fixed in Boun's fluid for 24 hours and further processed by using routine histological techniques [14]. These pieces were processed in paraffin wax for embedding and 6 µm thick sections were cut on manual Rotary microtome then stained in (H&E) heamatoxylene and eosin. Finally stained sections were mounted in Canada balsam. Photomicrographs of the selected sections were taken with help of camera Olympus Dp12.

Results and discussion
The nematode parasite Heterakis gallinarum has widely host distribution in birds and a little work related to pathology is often reported. In present study one species of nematode parasite was recovered from the cecum of infested birds. The Parasite was identified as Heterakis gallinarum on the basis of structural characters by the help of keys and literature Souls by (1982) and their prevalence was 20 % among recovered helminthes. Free range scavenging system and contaminated food are considered as major contributing factors for infestation of Heterakis gallinarum in poultry [15]. Only one nematode species was recovered from the exotic chickens during wet and dry seasons in scavenging birds and no occurrences of trematode was found [16]. Histopathology of Heterakis gallinarum in common pheasants show high prevalence of necropsies infection with gross lesion, obstruction, thickening and petechial mucosa, bowel obstructions and nodular granulomas in the ceca. Sub mucosal examination confirms chronic inflammation of cecum, hemosiderosis, granulomas, and leiomyomata. The immature H. gallinarum was recovered from the muscular and serosal region of intestine. These pathological lesions in birds reveal severe sub mucosal damage caused by H. gallinarum that results numerous necroses interrelated with cholesterol cleft and neoplastic nodules [17]. In the present histological investigations ceca of noninfected chicken showed normal sub mucosal and serosal region (Figure 1). Ceca also exhibited normal architecture of columnar epithelia contain enterocytes with microvilli and goblet cells, lamina propria mucosae and crypts of lieberkuhn ( Figure  2).

Figure 3. Section of infected ceca of chicken showing intestinal glands which are totally fused and filled with inflammatory cells. Distortion of muscular layer is obvious. Note two arteries with obscure morphology X100
Whereas ceca of infected chicken with Heterakis gallinarum represents fused architecture (Figure 3). Consequently these intestinal glands fused and come out as a cellular mass (Figure 4) and degeneration of intestinal glands, necrosis and infiltrated caecal lamina propria were also observed ( Figure 5). The infiltrated colon lamina proparia present in the intestine of duck infected by Heterakis gallinarum [18]. Nematode parasite cause hemorrhagic enteritis and sinusitis in the intestine therefore sever destruction and deterioration of microvilli observed in duck flocks [19].
The Heterakis gallinarum parasites cause inflammation, damage of intestinal glands and muscularis mucosae in pigeons [20]. The findings of present investigations reflect that Heterakis gallinarum exerted alterations in muscular layer along with vacuolation in the ceca of infected birds (Figure 6-7). The occurrence of intestinal infection associated with H. gallinarum shows severe inflammatory reaction in cecum [21,22]. Pathological alterations detect tiny white circumscribed nodules measuring 2-3 mm in the intestinal mucosa in case of H. gallinarum infestations [23,24]. In accordance to previously reported results in birds, findings of present study revealed, narrow, blocked and distorted blood vessels in muscular layer of ceca (Figure 8). Histopathological results of present experimental study on Heterakis gallinarum in the ceca of infected birds reveal widerange muscular alterations and damage affecting entire structure of gastro intestine. Hence, proper knowledge and advanced poultry farming practices are recommended to reduce the nematode parasitism in local birds to avoid further economical losses.