Gross and Histopathological Lesion Induced on the Gills Ofbarbus Fish by Monogenea Trematods Habiteting in Western Part of Lake Tana, Amhara Region, Ethiopia

Back ground: Gill parasitic is common on cultured and wild �sh. Many of these species have long been recognized to have the potential to affect the growth, fecundity and survival of hosts.the objective of the study were to determine the prevalence and identify, gross and histopathological lesion of gills caused by Dactylogyrus and Gryodactylus spp. parasitic infection on naturally infected barbus �sh. Result: A cross sectional study was conducted from in Lake Tana, amhara region. Three hundred eight four gill specimens were collected from barbus �sh and of these, 22 (5.7%) of �sh were infected with monogenean parasites. Higher prevalence of Dactylogyrus spp. (4.86%) was recorded than Gryodactylus sp (1.04). Large size �shes ( ≥ 20cm) were more susceptible to Dactylogyrus spp and Gryodactylus spp. However, highest prevalence of Dactylogyrus sp. was found in large �sh size (6.85%). Gryodactylus sp. was not recorded in all small size �sh. Descriptive statistics such as percentages was used to describe the nature and the characteristics of the disease. At gross examination of gills, hyperaemia and swollen, excess mucus secretion, paleness, congestion of branchial blood vessel were observed. Gills specimens of infected �sh were �xed with10% formalin, for further identi�cation and stained with the haematoxylin-eosin. Histopathological changes included hyperplasia, congestion and mucous cell proliferation of the gill epithelium and damaged primary and secondary lamellae, the uplifting of respiratory epithelial wall and damaged pillar cells. Conclusion


Background
Gill parasites are common on cultured and wild sh.Many of these species have long been recognized to have the potential to affect the growth, fecundity and survival of hosts [1] Moreover, extensive tissue damage resulting from the feeding and attachment of these parasites have been reported in several species of sh [2].Among the comment sh parasite, Dactylogyrus sp. was mostly observed in gills.Dactylogyrus sp have a series of hooks that enable them to attach while feeding.Most species are hostand site-speci c, requiring only one host to complete an entire life cycle.Fish parasites result in huge economic losses as they increase mortality and also increase farm inputs via increased treatment expenses and cause reduction in growth rate due to the parasitic disease outbreak [3].Also there was no research work conducted on gross and histopathological analysis on lesions caused by monogenea parasite of barbus sh at Lake Tana.Therefore, the objectives of this study were: General objective: The general aim of the study was to determine the pathological effect of gill trematode infection and the prevalence of gill Dactylogyrus spp and Gryodactylus spp of barbus sh at western part of the Lake Tana, Amahara region.Speci c objectives: To determine the prevalence of gill Dactylogyrus spp and Gryodactylus spp parasites in barbus sh in western part of LakeTana, to identify the type of Dactylogyrus spp and Gryodactylus spp parasites affecting gills on barbus sh in western part of Lake Tana and to determine the gross and histopathological lesions caused by the identi ed parasite of gills in barbus sh at western part of Lake Tana.

Results
A total number of 384 laebo barbus sh were observed in Lake Tana.Out of the 384 sh examined, 22 (5.72) were infested with monogenean parasites where the prevalence of Dactylogyrus sp. was 4.86 % while that of Gyrodactylus sp. was 1.04% (Table 1).

Histopathological lesions
Microscopic pathological changes were observed on the gills of parasitized sh, and they were comprised of fusion, curling and necrotic change the gill laments.The proliferation of bronchial tips, atrophy, complete loss of gill lamellar epithelium, hyperplasia and fusion of secondary gill lamellae were recorded.
Uplifting of primary and secondary gill lamellae epithelium was also observed.Leucocytes in ltration, telangiectasia in secondary lamellae, congestion on the gill laments and mucous cells proliferation were also observed during hitopathological examination.

Discussions
In present study, the histopathological effect caused by monogenea is mainly due to their morphological structures and specialized mode of attachment with host tissues.Infestation of the sh with Gryodactylus and Dactylogyrus, resultes severe damage to gills lamellae may be similar to lesions that were reported by [4].
In this study the Parasites attach gill epithelium and destruction of the cells or by congesting blood from the ruptured blood vessels.Gill epithelial cells are mechanically injured by the hooks of identi ed parasites.The blood expelled from this would be fed by the parasite and activity of some parasites can lead to bacterial or fungal secondary infection and causes mass mortality in cultured and wild situation and consequently sh respiration will be impaired, which results in reduced feeding, weight loss and general deterioration of health.Decreasing the body weight and condition factor, sever changes in osmoregulation or respiratory dysfunction and nally death may be observed in infection with Gryodactylus and Dactylogyrus parasite these study agree with [5].
The histopathological lesion which is observed in this study include lamellar fusion, hyperplasia, and aneurysm.Some alternations in blood vessel may occur when sh suffer from severe types of stress.In this case, damaged pillar cells can result in an increased blood ow inside the lamellae and cause blood congestion or even an aneurysm [6].In the present study of monogenea infection, gill in ammation, swollen, hyperemic, hemorrhage, and excessive mucus production were recorded in infected barbus sh gill tissue.Leads to, blockage of blood vessels leading to respiratory and osmoregulatory failure according to [7].
In the present study, necrosis lamellae were observed in barbus sh infected with Gryodactylus and Dactylogyrus spp of monogenean parasites.The mechanism by which the destruction of the branchial vessels may happen by the parasite, where the blood pressure is low and no extensive hemorrhages are caused and the very short clotting time of blood brings about rapid occlusions of the vessel then thrombus is formed resulting in ischemia, which in turn leads to necrosis, these result similar with [8] and [9].In the present study, reduced chloride cells were seen.The infection of identi ed parasite in sh gills has an impact on the host's ability to regulate its ion balance.These ndings agree with reports of [10] and [11].These parasites reduced the number of chloride cells which are the main site of ion absorption and secretion [12].
In the present study heamorrhage was observed.Gryodactylus and Dactylogyrus parasitic damage the gills by feeding on the delicate tissue of the gill lamellae or on the blood circulating within the lamellae, leading to a loss of respiratory surface area, extensive gill damage and severe hemorrhage, with in ammation.Blood vessels in the gill laments are blocked and this leads to atrophy of gill tips .thesendings is agree with [13]and [14].

Conclusions
The major features of Gryodactylus and Dactylogyrus parasite spp.infections on the gill laments of the sh in this study mostly include destruction of the gill laments and lamellae, exerted by feeding of the parasite and the result in hyperplasia and thickening of the epithelial cells reducing the surface area for effective respiration.Gill damage could result in loss of gill surface area for respiration, which would lead to suffocation particularly at high water temperatures.The histoarchitectural change in gills hinders the oxygen intakes and in the long term causes huge mortality in aquaculture practices.According to the result of the gross and histopathological changes induced by the infections of parasites would adversely affect the proper functioning of the gills of infected sh.Further study should be warranted on the prevalence and effect of other parasites.

Study population
The study animals were sh of barbus (Labeo barbus) that was collected from Lake Tana, Amahara region, Ethiopia.Sampled shes were selected by simple random sampling technique.Large, medium and small sized shes were included in my study.Large, medium and small sized shes were selected by simple random sampling techniques associations that are found in western part of Lake Tana.The study population, small (1 to 10 cm), medium (10.5 to 20cm) and large size shes (≥20 cm) were categorized according to [15].

Sample collection techniques, gross and histopathological examination
A total of 384 randomly selected sh, Labeo barbus, were sampled and examined.The length, date and site of collection of host specimens were recorded.Gross lesion and health conditions of each specimen were recorded and monogenean parasite (Gryodactylus and Dactylogyrus ssp.) of gills were collected with forceps.Postmortem examination of gills of the investigated shes was examined according to [16].Damaged sh tissues (gills) were taken from the parasite attachment area of infested shes and were cut out in fresh condition xed in 10% buffered neutral formalin.In addition, monogeneans parasites of gills were collected and preserved10% buffered neutral formalin.Then, the necessary information were leveled on the sampling bottles.Finally, the samples were transported to Faculty of Veterinary Medicine Pathology and parasitology laboratory, University of Gondar, for identi cation of parasites and histopathological examination of tissue specimens.After used the xative, the tissue were washed in running water for 24hr to remove the xative entirely .The tissues were then processed using a 18 hr automatic tissue processor .The tissue processor contains 12 beakers (8 glass beakers contains alcohol, 2 glass beakers contains xylene and 2 glass beakers containing para n wax).
The tissues after being processed are embedded using an automatic embedding centre.Embedding is a process of submerging a tissue in a metal plastic disposable embedding mould containing molten para n wax, which became solidi ed when it was cold.This formed a support medium for the tissue during sectioning.The blocked tissues were sectioned at 4-5 microns a rotary microtome (LEITZ 2535, Germany), and oated into pre coated slides and were placed in a clean grease free slide which was then placed on a hot plate for 30 min in order for section to adhere to the slides.The staining method used was the H&E staining method.Standard histological procedure was followed as described by [17].This method was used in order to demonstrate the general structure of the tissues.These were then dewaxed in xylene.The processed section were later taken to water by using descending grades of alcohol, that is, from absolute alcohol, 95% alcohol, 70% alcohol,50 %,for 5 min each and rinse tap water for 10 min.It was stained in haematoxylin for 6 min, then put in running water for 20 min.This was counter stained in 1% Eosin for 15 min and, then dehydrated using ascending grades of alcohol (95% alcohol and absolute alcohol).These were cleared in xylen, mounted using D.P.X (a mountant) and viewed under light microscope [18].Lesions were described and scored as none for no lesion, mild, or severe depends on the type of lesion [19].Preserved in 10% formaline and labeled with all necessary information for further identi cation.The collected parasites placed on glass micro-scope slides with a drop of 10% formalin and slightly compressed between a slide and a cover slip prior to being examined under stereomicroscope (4x,10x.40x)and observe morphological feature of the parasite .The monogeneas parasites were identi ed microscopically using the identi cation guideline of [20].

Data analysis
The data were entered and managed in Microsoft Excel.All the data analysis was done by SPSS software version 12. Descriptive statistics was applied for the analysis of the data obtained.Descriptive statistics such as percentages were used to describe the nature and the characteristics of the data.The prevalence of monogenea parasite (Gryodactylus and Dactylogyrus spp.) was analyzed using Chi-square test and <0.05 was considered as signi cant.

Figures
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Figure 4 Normal
Figure 4

Table 1
Prevalence of Gyrodactylus sp and Dactylogyrus sp in barbus sh in the study site.