"The study was conducted to investigate the histological changes of the gills infection 
by myxosporean parasites in Labeo rohita from Myit- Too-Myit-Sut In (lake). The study 
period lasted from March 2019 to January 2020. The occurrences of histological 
alterations in the gills were hyperplasia that caused shortening and fusion of lamellae, 
epithelium separation, curling of lamellae, blood cell congestion and dilation of gill 
filament. At the present study, the plasmodia of myxosporean parasites were found 
within the lamellae (Intralamellar type) and between the gill lamellae (Interlamellar 
type). Histopathological changes of gills tissues indicated that the myxosporean 
parasites were highly pathogenic to host fish. 

Nu Nu Wai

Thant Zin

English

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Yadanabon University Research Journal 2023, Vol.13 , No.2                                              143  Histopathological Changes in the Gills of Labeo rohita (Hamilton, 1822) Infection of Myxosporean Parasites at Myit-Too-Myit-Sut In (Lake), Amarapura Township, Mandalay Region Nu Nu Wai, Thant Zin  Abstract The study was conducted to investigate the histological changes of the gills infection by myxosporean parasites in Labeo rohita from Myit- Too-Myit-Sut In (lake). The study period lasted from March 2019 to January 2020. The occurrences of histological alterations in the gills were hyperplasia that caused shortening and fusion of lamellae, epithelium separation, curling of lamellae, blood cell congestion and dilation of gill filament. At the present study, the plasmodia of myxosporean parasites were found within the lamellae (Intralamellar type) and between the gill lamellae (Interlamellar type). Histopathological changes of gills tissues indicated that the myxosporean parasites were highly pathogenic to host fish. Keywords: Histological changes, Myxosporean parasites, fish, gill filament, Myit-Too-Myit-Sut In (lake)  Introduction Fish is a particularly rich source of high quality protein, a range of micronutrients (eg. calcium, zinc, iron, vitamins) and fatty acids essential for human health and development (Belton and Thilsted, 2014). Development of aquaculture during the last decade have resulted in much greater attention paid to problem posed by parasites and their impact on the fish health leading to constraints in the productivity of aquaculture (Kaur and Singh, 2018).  Parasites are capable of causing damage to the fish through injury to the tissues or organs. Fish parasites result in economic losses or mortality, treatment expenses, growth reduction during and after outbreak of disease (Roberts, 2001). Myxozoan infections result in large scale histopathological features according to the infection stage and host susceptibility. Histozoic parasite species elicit greater pathological response compared to coelozoic species (Feist and Longshaw, 2008). Gills are specialized tissues for gaseous exchange, circulation, ionic balance, hormone production and nitrogenous waste excretion (Pelster, 2010). Gills of fish are important sites for prevalence of disease.  In Indian major carps, gill diseases are very common (Sanaullah and Ahmed, 1980).   Lecturer, Dr, Department of Zoology, Yadanabon University  Professor, Dr, Curriculum Central Unit, Department of Education Research, Planning and Training Yangon 144                                                                   Yadanabon University Research Journal 2023, Vol.13 , No.2 There are several earlier reports of histopathological studies of myxosporidian gill infections from different localities (Shulman, 1957; Dykova and Lom, 1988; Molnar, 1979; Sanaullah and Ahmed, 1980). In Myanmar, histopathological studies of myxosporean parasites in the gill infections from different fish species were reported in Laboe rohita (Moe Kyi Han, 2006), in Cyprinus carpio (Sein Sein Myint, 2006) and in Cirrhinus mrigala (Pa Pa Win, 2007). Histological alterations of animal tissues provide a rapid method to show effects of irritants and pathogens in different tissues and organ (Bernet et al., 1999). Very often, histopathological examination allows to identify the parasite species involved, the area of pathological lesions, possible complications of bacterial or viral origin, and the outlook of treatment (Soltysiak et al., 2014). Myanmar’s farmed fish exports are dominated by rohu, along with two other “Indian major carp species; mrigal and Catla catla (Belton et al., 2015). Thus, Labeo rohita shows the important role for economic commercial value in Myanmar’s market. However, natural populations of the fish are rapidly decreasing due to parasitic infections that cause potentially disease. The objectives of the present study were to study the histological changes induced by myxosporean infection in the gills of the selected fish species and to confirm the location of the parasitic infection in the gills of fish hosts.    Materials and Methods Study Area  Myit-Too-Myit- Suit In (lake) was located between Latitude 21°51′10′′ N and 21°51′14.16′′ N and Longitude 96°01′25′′ E and 95°59′41.85′′ E in Amarapura Township, Mandalay Region (Fig 1). Study period  The study period lasted from March 2019 to January 2020. Collection of fish Specimens  Five specimens of Labeo rohita was collected seasonally during the study period from Myit-Too-Myit-Suit In. Total length, standard length and body weight of collected fish specimens were measured. Then, the gills were taken from the head and they were fixed in 10% formaldehyde solution and carried to the laboratory of Zoology Department, University of Mandalay. Preparation and examination of myxosporean parasites  For histopathological study, myxosporean infected gills of fish species fixed in 10% formaldehyde solution and dehydration, clearing and infiltration were made by the tissue processing. Paraffin wax sections of tissues were cut at 5 µ thickness using a rotary microtome. Wax sections affixed to the sides were subjected to heat at 60 ̊ C for overnight and then stained with Hematoxylin and Eosin (H&E). The routine procedures for H&E staining include deparaffinization in xylene series, rehydration in descending alcohol series, staining in Hematoxylin, differentiation in 0.5% acid alcohol, bluing Scott's tap water, staining in Eosin, dehydration in ascending alcohol series and cleaning in xylene series. These slides were subsequently mounted into Balsam Canada. Yadanabon University Research Journal 2023, Vol.13 , No.2                                              145  Myxosporean uninfected gills of fishes were also processed for the histological slides to be used as control. All of these histological slides were examined under light microscope and photomicrographs which were taken with digital camera attached to Meiiji Biological Microscope (MT 4300H). The method used in this study was according to the histological method of Cambell (cited by Kolmer et al., 1969).                            Figure 1. The map of Myit-Too-Myit-Sut In (Lake), Amarapura Township, Mandalay Region        (Source: Department of Geography, Mandalay University)  Results  Histopathological changes in the gills of Labeo rohita  In a normal gill, the primary and secondary gill lamellae were arranged systematically with the secondary lamellae on each of the primary lamella. (Plate 2. A)  The histological changes of infected gills in Labeo rohita were observed. Hyperplasia and fusion of secondary lamellae were found in many gill filaments (Plate 2 B and C). In some gills, dilation of filament and epithelium separation of gill lamellae were observed (Plate 2 B). In many infected gills, blood congestion and defroming were observed (Plate 3 G and I).  Plasmodium of Myxobolus sp. and Thelohanellus sp. were found in several gill filaments (Plate 3 H and I). Infection of plasmodium caused destruction of adjacent gill lamellae (Plate 2 F). Plasmodium attached to the tip of gill lamellae caused the hyperplasia of gill lamellae (Plate 2 C).In infected gills, plasmodium attached between the gill lamellae were observed (Plate 3 J). In some cases, individual spores were found at the tip of gill lamellae (Plate 2 D) (Table 1).  Localization of plasmodia in the gills of Labeo rohita The infected gills, plasmodia with mature spore and individual spores were observed. Plasmodia of Thelohanellus sp. attached within the gill lamellae (Intralamellar type) (Plate 2 E and F). In some cases, plasmodia were found between the gill lamellae (Interlamellar type) (Plate 3  I and J) (Table 1). 146                                                                   Yadanabon University Research Journal 2023, Vol.13 , No.2 Table 1.  Histopathological changes in the gills of Labeo rohita infected by myxosporean parasites from Myit-Too-Myit-Sut In (lake) during the study period Sr no. Histopathological changes of gill tissues Rainy Season Cold Season Hot Season 1 2 3 4 5 1 2 3 4 5 1 2 3 4 5 1 Spore - - - - - - + + - - + + - + - 2 Plasmodia - + - ++ - + ++ + - - ++ + - ++ + 3 Hyperplasia ++ +++ ++ +++ + +++ ++ ++ + + + ++ + +++ ++ 4 Fusion of lamellae + ++ - +++ + ++ ++ + + ++ + +++ + ++ ++ 5 Shortening of lamellae + ++ + ++ - + + - - + + ++ + + + 6 Epithelium separation - - - + - + + - - + + + - ++ + 7 Dilation of filament + - ++ + - + - - - + - - + ++ - + = low (1-3), ++ = medium (3-5), +++ = high (5 above)             Plate 1. Myxosporean parasites infected in the gills of Labeo rohita from Myit-Too-Myit- Suit In (lake)                          A. Normal uninfected gill (x 100) B. Shortening and complete fusion of gill lamellae (black arrow), dilation of filament (blue arrow) and epithelium separation (red arrow) (x 100)  A. Labeo rohita B. Myxobolus sp. C. Thelohanellus sp.  Yadanabon University Research Journal 2023, Vol.13 , No.2                                              147                                   Plate 2. Histopathological changes in the gills of Labeo rohita from Myit-Too-Myit- Suit In (lake)            C. Plasmodium (oval shape) attached to the tip of gill lamellae (black arrow), hyperplasia and fusion of lamellae (blue arrow) (x 400)  D. Escaped spore of Thelohanellus sp. attached to the tip of gill lamellae (black arrow) and hyperplasia of gill lamellae (x 400)    E. Plasmodium (oval shape) of Thelohanellus sp. within the gill lamellae (black arrow )(x 400)  F. Plasmodium (rounded shape) of Thelohanellus sp. within the gill lamellae (black arrow) destroyed the adjacent gill lamellae (x 400)  G. Plasmodium (oval shape) of Thelohanellus sp. within the gill lamellae (black arrow) and blood congestion (blue arrow) (x 400) H. Plasmodium (rounded shape) of Thelohanellus sp. within the gill lamellae (black arrow) (x 400)  148                                                                   Yadanabon University Research Journal 2023, Vol.13 , No.2                         Plate 3. Histopathological changes in the gills of Labeo rohita from Myit-Too-Myit- Suit In (lake)  Discussion In the present study, the gills of Labeo rohita obtained from the study area were found to be affected by the myxosporean parasites under histopathological observation. In the infected fish host, the gills were found to have dilation of blood vessel, separation of epithelial cells in the filament, hyperplasia, fusion and shortening of gill lamellae. Rukyani (1990) also described that Myxobolus infection in common carp was reported to result in lesion, inflammation, congestion and hyperplasia in the gills. In the present study, the plasmodium of myxosporean parasites infected the gills of L. rohita from the study area. Hyperplasia, separation and dilation of gill filament, shortening and fusion of gill lamellae were observed. The present observation correlated with the results of I.Plasmodium (oval shape) of Thelohanellus sp. found between the gill lamellae (black arrow) and deforming the gill lamellae (x 400)  J. Plasmodium (rounded shape) of Thelohanellus sp. between the gill lamellae (black arrow) and distortion of adjacent lamellae (x 400)  K. Plasmodium (oval shape) of Myxobolus sp. between the gill lamellae (blue arrow) and hyperplasia (black arrow) (x 100)  L. Hyperplasia and fusion of lamellae (blue arrow) and dilation of filament (black arrow) (x 400)   Yadanabon University Research Journal 2023, Vol.13 , No.2                                              149  histopathological studies done by Kaur and Katoch (2014) who stated that the gills of L. rohita were infected by Thelohanellus bifurcate where several gill lamellae were fused and destruction of gill lamellae were reported. Kaur et al. (2015) also described that the presence of spore within the entire length of the gill filament in Labeo rohita, the blood vessels were dilated and bounding endothelial cells were highly compressed. Molnar and Eszterbauer (2015) stated that myxosporean plasmodia might develop in or among gill lamellae, in gill filaments and inside the cartilaginous gill arch. This statement agreed with the present study. Almost all of infected fish, the myxosporean parasites were found in the gill filaments and gill lamellae. The location of myxosporean parasites had been reported by many researchers. Ahmad and Kaur (2017) reported that the intralamellar vascular type in Cirrhinus mrigala and histological changes also revealed hypertrophy, lifting and hyperplasia of epithelial cell within the gill lamellae. Gupta and Kaur (2017) observed that the gills were infected with Thelohanellus theinensis as interlamellar type and resulting the hyperplasia and the dilation of the lamellae.  Sandar Win (2008) described that the location of plasmodia in the gills of Tilapia were observed to be three type; epithelial filamental type, interlamellar type and intralamellar type. These plasmodia type are very similar to the present work. However, the host fish species are different from the present study. Sanaullah and Ahmed (1980) reported that parasitic cysts were observed in the distal tip of the primary gill lamellae along their length. The cysts in the proximal region of the filament were generally much smaller than those in the distal region. This finding agreed with the present study. In some infected fish, the plasmodium of myxosporean parasites attached at the tip of gill lamellae. Kaur and Kotach (2014) observed that the rupturing of cysts can also lead to hemorrhages and may result in considerable loss of respiratory surface. This observation agreed with the present study. In some cases, plasmodium of Myxobolus sp. ruptured and released many spores causing the deformation of gill lamellae. The gills of fish are an important site for disease production because they are a rich source of blood, an important media for the infectious agents. Thus, the histological study of gills indicated the myxosporean parasites actually pathogenic to gill of host fish. The present study revealed that myxosporean parasites were potentially pathogenic to host fish. The pathological changes of gills indicated the health status of the fish species from the study area.     150                                                                   Yadanabon University Research Journal 2023, Vol.13 , No.2 Acknowledgements We are greatly indebted to Dr Tint Moe Thu Zar, Rectors and Dr Thwe Linn Ko, U Khin Myot, Dr Myint Myint Oo and Dr Khin Maw Maw Soe, Pro-Rectors, Yadanabon University for their acceptance and permission to carry out this research. I strongly wish to thanks Dr Aye Aye Cho, Professor and Head, Department of Zoology, Yadanabon University, for her permission to conduct this research and guidance, advice and encouragement. I would like to express my thanks to Professor Dr Thi Thi Han, Department of Zoology, Yadanabon University, for her advice and encouragement.  References Ahmad, I. and Kaur, H., 2017. Redescription and histopathology of two species of Myxozoans infecting gills of fingerlings of indian major carps. 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Histopathological Changes in the Gills of Labeo rohita (Hamilton, 1822) Infection ofMyxosporean Parasites at Myit-Too-Myit-Sut In (Lake), Amarapura Township,Mandalay Region

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Histopathological Changes in the Gills of Labeo rohita (Hamilton, 1822) Infection ofMyxosporean Parasites at Myit-Too-Myit-Sut In (Lake), Amarapura Township,Mandalay Region

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