Epizootic Ulcerative Syndrome Research Articles

Molecular characterization of Aeromonas hydrophila isolates from diseased fishes in district Kasur, Punjab, Pakistan

Abstract Pakistan is an agricultural country and fisheries play a very important role in the economic development of the country. Different diseases are prevalent in Pakistani fish but information related to the causative agents is not well-known. Keeping in view the significance of bacterial pathogens as the causative agents of multiple fish diseases, the present study was conducted for identification, characterization and analysis of virulence genes of Aeromonas spp. isolated from diseased fishes. A total of fifty fish samples having multiple clinical indications were collected from different fish farms of district Kasur, Punjab Pakistan. For isolation of Aeromonas spp. samples were enriched and inoculated on Aeromonas isolation medium. Isolates were identified and characterized by different biochemical tests, Analytical Profile Index (API) 20E kit and Polymerase Chain Reaction (PCR) assays. All isolates were screened for three putative virulence genes including aerolysin (aer), haemolysin (hyl) and heat labile cytotonic enterotoxin (alt). Seven isolates of Aeromonas (A.) hydrophila were retrieved and identified based on API 20E. These isolates were further confirmed as A. hydrophila on the basis of PCR assays. Three isolates were detected positive for the presence of virulence genes (alt and hyl). Whereas aerolysin (aer) gene was not present in any of A. hydrophila isolates. The present study confirmed A. hydrophila as the causative agent of epizootic ulcerative syndrome and motile Aeromonas septicemia in fish farms of district Kasur, Punjab Pakistan. Moreover, detection of two virulence genes (alt and hyl) in A. hydrophila isolates is a threat for fish consumers of study area.

Assessment of Epizootic Ulcerative Syndrome (EUS) in Estuarine Fishes Collected from Arasalar Estuary, Puducherry State, India

Fishes form an important source of cheap protein for people of many countries. The majority of the global production comes from freshwater aquaculture (58%) followed by marine (36%) and brackish water (6%). With an average increase in aquaculture production of 11% during the last decade, aquaculture has become the fastest growing sector of the world food economy However, fish diseases is now posing a great threat to achieve maximum production and has become a limiting factor to the success of aquaculture. Among the various diseases, Epizootic Ulcerative Syndrome (EUS) is a disease affecting a wide range of wild and farmed freshwater and estuarine fish which has spread across south-east Asia and India. From the fish survey, Food Fishes were collected from the Arasalar estuary, Puducherry State, India. The fish caught were identified and average 200 species of each fish were examined for EUS (Epizootic ulcerative syndrome) infection for the four seasons of the year (Pre-summer, Summer, Post-summer and Rainy season). The results of the number of fishes with EUS that could be identified in the different seasons are reported. Of the 75 fishes, that were examined, 12 were affected with EUS. This shows that 16% of the fishes were subjected to infection.
 Among Lizaparsia, 11 species were infected representing 14.6% of infection and among Mystus, seven fishes were infected forming 9.3% of infection. With regard to Oreochromis mossambicus, only four species were infected forming 5.3% of infection. With regard to Glossogobius giuris, nine species were infected thus 12% of infection. Nevertheless, during this period, the range of infection ranged from nil to 16%.
 During the pre-summer season, the percentage of infection again ranged from nil but to 12%. The maximum infection was noticed in Liza parsia (9 fishes) representing 12% infection, Puntius filamentosus (8 fishes) recording 10.6% followed by Glossogobius and Mystus (8%) and Oreochromis mossambicus (1.33%) infection respectively. During the summer and the post-summer season, none of the fishes that were investigated recorded any signs of EUS. Thus, an overall comparison reveals that the highest percentage of EUS was recorded in the rainy season followed by the pre-summer season and least in summer and post-summer season.

A case study of epizootic ulcerative syndrome (EUS) caused by Aphanomyces invadans in eastern mosquitofish (Gambusiaholbrooki) from the headwaters of Charleston Harbor, South Carolina.

A case study of epizootic ulcerative syndrome (EUS) caused by Aphanomyces invadans in eastern mosquitofish (Gambusiaholbrooki) from the headwaters of Charleston Harbor, South Carolina.

PENGUJIAN PENYAKIT KOMODITAS IKAN AIR TAWAR PADA LALU LINTAS DOMESTIK DI STASIUN KARANTINA IKAN JAMBI

Increased trade flows of international and domestic fishery coomodities in the opportunity to enter and spread Hama penyakit Ikan Karantina (HPIK) caused by pathogenic organism, namely bacteria, parasites, fungi, and viruses. This study aims to identify whether freshwater fish commodities in domestic traffic are infected with parasites, the bacteria Edwardsiella ictaluri, TiLV (Tilapia lake Virus) and the fungus Aphanomyces invadans at the Fish Quarantine Stasion, Quality Control and Safety of Fishery Products Jambi. Fish samples consist of 15 fish samples to be tranported. Based on results of the examinatoin of fish diseases, Parasite Examination there were 4 parasites with 3 types of parasites, namely 2 parasites of Dactylogyrus sp. On the gills of Tilapia and catfish. One (1) parasite Gyrodactylus sp. On fish slime Stripe 3, and 1 parasite Camallanus sp.in the intestines of betta fish; on bacterial examination of the 4 samples of fish examined, Edwardsiella ictaluri bacteria were nof found; on virus examination the result were negative for TiLV (Tilapia lake Virus); on examination of the fungus with the PCR method, negatif result were obtained for the fungus Aphanomyces invadans which is the cause of EUS (Epizootic Ulcerative Syndrome).

Global Pattern of Epizootic Ulcerative Syndrome Outbreaks in Freshwater Farmed and Wild Fish from 2005 to 2012

Epizootic ulcerative syndrome (EUS) is an infectious disease of freshwater farmed and wild fish caused by a fungus, Aphanomyces invadans. This disease is an important economic disease causing huge socio-economic losses to fish farmers across the world. While this is an important fish disease, its global epidemiology is poorly understood. The objective of this study is understand the global spatiotemporal epidemiology of EUS outbreaks from 2005 to 2022. The data were retrieved from open access database of the World Organization for Animal Health (WOAH). Results showed that there were a total of 277 outbreaks of EUS reported to WOAH from 2005 to 2022 from 15 countries and territories with 76% outbreaks reported from African countries. Democratic Republic of Congo reported the highest number of EUS outbreaks. Higher number of outbreaks were reported during 2014 to 2017. The observed epidemiological pattern might be biased due to the underreporting of the disease. Nevertheless, it offers important insights to understand the latest global pattern of EUS. We suggest to increase country level awareness on EUS so that outbreaks are reported in a timely manner.

Development of a rapid and sensitive real-time diagnostic assay to detect and quantify Aphanomyces invadans, the causative agent of epizootic ulcerative syndrome.

The oomycete Aphanomyces invadans causes epizootic ulcerative syndrome (EUS), a World Organization for Animal Health (WOAH)-listed disease that has seriously impacted a wide range of fish worldwide. Currently, only three conventional polymerase chain reaction (PCR) assays are recommended for the detection of A. invadans. The robust quantitative PCR (qPCR) assay has recently become more important due to its highly accurate nature and the applicability of qPCR-based environmental DNA (eDNA) detection in the monitoring of pathogens in aquatic environments. Therefore, in this study, we developed a novel TaqMan probe-based qPCR method to sensitively and quantitatively detect A. invadans. The assay limit of detection was determined using 10-fold serial dilutions of linearized A. invadans plasmid. Assay sensitivity was assessed in the presence of interfering substances and compared to three WOAH-listed primers using the mycelia and zoospores of A. invadans with and without fish muscle tissue. The assay specificity was also theoretically and experimentally assessed against other oomycetes, fish muscle tissue, and water samples. The assay's repeatability and reproducibility were determined. In this study, the limit of detection of the developed assay was 7.24 copies of A. invadans genomic DNA per reaction (95% confidence interval (CI): 2.75 to 19.05 copies/reaction). The assay showed the same sensitivity in the presence of other substances. Compared to the WOAH-recommended PCR assays, this assay had 10-times higher sensitivity for all tested samples. There were no cross-reactions with other closely related oomycetes, fish muscle, or water samples, indicating that the assay was highly specific for A. invadans. The repeatability and reproducibility tests showed little variation, ranging from 0.1-0.9% and 0.04-1.1%, respectively, indicating the high consistency, repeatability, and reliability of the developed assay. This highly rapid, sensitive, specific, and consistent EUS qPCR assay would be of importance in transboundary disease management and the monitoring of pathogens in aquatic environments.

Study of epizootic ulcerative syndrome on hematological responses on Fish Channa punctatus

The present paper deals with the study of epizootic ulcerative syndrome on Hematological responses on Fish Channa punctatus. A number of fishes have been found to suffer from this epidemic disease. We are observed that the decreases was due to microcytic anaemia which in torn was due to epizootic ulcerative syndrome. Decrease in count of basophils, neutrophils and eosinophils in fish Channa punctatus to malathion toxicity was also observed.

Ciliated parasites as the primary agents for early epizootic ulcerative syndrome (EUS) in yellow catfish, Tachysurus fulvifraco

Epizootic ulcerative syndrome (EUS) is an emerging disease that accounts for the high morbility and mortality in yellow catfish, Tachysurus fulvifraco. Though the EUS of T. fulvifraco was most frequently supposed to be associated with bacterial infection, the causative agent for the disease is still not completely determined. To address this, an epidemiological investigation on EUS was carried out from 2019 to 2022 in Hubei province, one of the main culture areas of T. fulvifraco in China. It showed the outbreaks of EUS were mainly occurred in March and April, during which the water temperature ranged from 13 °C to 20 °C. At the early stage of EUS, the diseased T. fulvifraco typically aggregrated, with circular-hole shaped ulcers on skin. Gross examinations on the T. fulvifraco of early EUS revealed that the disease was commonly accompanied by ciliates infections. To further illustrate, recent diagnoses on T. fulvifraco of early EUS from Wuhan city were recorded in details. Critically, 78.1% of the specimens were heavily infected with Ichthyophthirius multifiliis and/or Epistylis horizontalis. Histopathological analyses revealed that these ciliated parasites infection caused severe damages on T. fulvifraco skin and gills. In comparison, only 12.5% of the T. fulvifraco specimens were infected with the bacterium, Aeromonas versonii. Taken together, our observations indicated that the ciliated parasites were the primary agents for early EUS, that contribute to the formation of ulcers on T. fulvifraco. These data facilitate us to understand the epidemic of ESU in farmed T. fulvifraco, and further explore the effective treatments to the disease.

Economically important freshwater fishes infected with fungi causes EUS

The aim of this study was to investigate fungal infections in Channa punctatus and Channa striatus twenty specimens of each species were studied for the presence of fungal infections. Infected fish’s shows clinical signs such as fungal growth on skin, fins, eyes, eroded fins and scales, hemorrhages on body surface and abdominal distension. The ability of aquatic fungi to cause diseases in fish is well known potentially all freshwater fishes their incubating eggs are susceptible to fungal infection. The most common fungal diseases of fish are saprolegniasis, disease caused by Achlya, branchiomycosis, epizootic ulcerative syndrome (EUS). Saprolegniasis is one of the most common infections of freshwater fishes in warm temperature and tropical waters; distributed throughout worldwide. Although there are many reports of different Achlya species infecting fish, there is no consistent and regularly observed clinical condition such as occurs with Saprolegnia. EUS is a seasonal epizootic condition of great importance in wild and farmed freshwater.

Immunomodulatory Potency of Eclipta alba (Bhringaraj) Leaf Extract in Heteropneustes fossilis against Oomycete Pathogen, Aphanomyces invadans

Aphanomyces invadans is an aquatic oomycete pathogen and the causative agent of epizootic ulcerative syndrome (EUS) in fresh and brackish water fish, which is responsible for severe mortalities and economic losses in aquaculture. Therefore, there is an urgent need to develop anti-infective strategies to control EUS. An Oomycetes, a fungus-like eukaryotic microorganism, and a susceptible species, i.e., Heteropneustes fossilis, are used to establish whether an Eclipta alba leaf extract is effective against the EUS-causing A. invadans. We found that treatment with methanolic leaf extract, at concentrations between 50-100 ppm (T4-T6), protects the H. fossilis fingerlings against A. invadans infection. These optimum concentrations induced anti-stress and antioxidative response in fish, marked by a significant decrease in cortisol and elevated levels of superoxide dismutase (SOD) and catalase (CAT) levels in treated animals, as compared with the controls. We further demonstrated that the A. invadans-protective effect of methanolic leaf extract was caused by its immunomodulatory effect and is linked to the enhanced survival of fingerlings. The analysis of non-specific and specific immune factors confirms that methanolic leaf extract-induced HSP70, HSP90 and IgM levels mediate the survival of H. fossilis fingerlings against A. invadans infection. Taken together, our study provides evidence that the generation of anti-stress and antioxidative responses, as well as humoral immunity, may play a role in protecting H. fossilis fingerlings against A. invadans infection. It is probable that E. alba methanolic leaf extract treatment might become part of a holistic strategy to control EUS in fish species.

Epidemiology of epizootic ulcerative syndrome (EUS) in fish in the main water bodies of the Kavango–Zambezi and Great Limpopo transfrontier conservation areas of Zimbabwe

A three-year study investigated the epidemiology of epizootic ulcerative syndrome (EUS) in fish from Kavango-Zambezi (KAZA) and Great Limpopo (GL) transfrontier conservation areas of Zimbabwe. A total of 38 sites comprising 27 wild fisheries and 11 aquacultures, from Mashonaland West, Matabeleland North and South, and Midlands were selected. Of the 27 wild fisheries, EUS-positive fish were detected from 9 (33.3%) and none from the 11 aquacultures. No positive cases were detected from Nile tilapia (Oreochromis niloticus) from both aquacultures and wild fisheries. A total of 9.9% (239/2423) fish from the nine positive fisheries had typical EUS lesions, and infection was confirmed in 15 species. Prevalence was significantly higher (p < 0.05) in KAZA (11.5%; 95% CI: 9.6-13.4) compared with GL (8.6%; 95% CI: 7.1-10.1). The most affected were Clarias, followed by Barbus and Oreochromis species. Most cases (>80%) were reported in winter when ambient temperature was low. Further studies are required to determine water parameters associated with EUS outbreaks. These results suggested that the African sharptooth catfish (Clarias gariepinus) could be used potentially as an indicator species for EUS surveillance programmes. Thus, implementation of surveillance and biosecurity programmes that take into consideration the epidemiology of EUS will be beneficial.

Chitosan-polymer based nanovaccine as promising immersion vaccine against Aeromonas veronii challenge in red tilapia (Oreochromis sp.)

Red tilapia (Oreochromis sp.), one of the important freshwater fish species in fish farming in Thailand, has for long been suffering from a serious bacterial disease named epizootic ulcerative syndrome and hemorrhagic septicemia. The disease is mainly caused by Aeromonas veronii. Vaccine is proposed to be a major impact tool for sustainable control and prevention strategies. Vaccination by immersion has many benefits over injection. However, the conventional immersion method suffers from a low potency due to the inefficient uptake of antigens across mucosal tissue. Here, we developed a chitosan-polymer based nanovaccine together with an efficient delivery vehicle to enhance the immunogenicity of immersion vaccination, increasing bioavailability and inducing local immune responses during transit to mucosal inductive immune sites. The physiochemical properties of nanovaccine, which was modified on surface particle by using a mucoadhesive polymer, were assessed for size, zeta potential, and particle distribution. Our study demonstrated by SEM image and microscopic fluorescence image that nanovaccine greatly increased the binding and penetrating ability into gills when compared with formalin killed vaccine. The nano-sized particles were well dispersed in water and trapped in core nanoparticle as confirmed by TEM image. The efficacy of vaccine was performed by immersion challenge with virulent A.veronii after 30 days post vaccination in tilapia. The result revealed a high level of mortality in the control, empty-polymeric nanovaccine and formalin killed bacterin vaccine groups. A high relative percentage survival (RPS) of vaccinated fish was noted with chitosan-polymer based nanovaccine. Our studies indicated that this chitosan-polymer based nanovaccine derived from cell fragments and supernatant was the improved version of the conventional formalin killed vaccine. The chitosan polymer based particle could increase the efficacy of nanovaccine toward the target mucosal membrane and enhance protection against A. veronii infection in red tilapia.

Histopathology of the fish infected with the epizootic ulcerative syndrome in Eastern Nepal

Epizootic ulcerative syndrome (EUS) is the major disease of fish which causes high mortality of fish in eastern Nepal since its initial outbreak. Of the 445 EUS infected fish samples, the suspected fungus was identified in eight spp. during the five years study period following standard methods to isolate, culture and identification. The sections of muscles, livers, and kidneys of infected fish samples showed the outbreak of EUS due to the presence of Aphanomyces spp. A complete loss of epidermis and the underlying musculature was replaced by granulomatous and inflammatory tissues. In some areas, myonecrosis and fungal hyphae, black stained with GMS, reflected a positivity for the presence of fungus. Pathogenicity test in Heteropneustes fossilis showed some lesions and 43.33% mortality when isolated zoospores were injected into experimental fish samples.

Status of disease prevalence, drugs and antibiotics usage in pond-based aquaculture at Narsingdi district, Bangladesh: A major public health concern and strategic appraisal for mitigation

This research aimed to investigate the present status of disease prevalence and usage of aqua drugs for various aquaculture operations in the Narsingdi region of Bangladesh. Data were collected through the market survey, preset questionnaire interview, personal contact, and participatory rural appraisal tools. Amongst the respondents, the maximum percentages were found practicing mixed cultures of carp, tilapia, and pangas. The respondents suggested that epizootic ulcerative syndrome, saprolegniasis, streptococcosis, tail and fin rot and bacillary necrosis are common fish diseases in the area. About 140 drugs of different companies used in aquaculture for different purposes such as disease treatment, growth enhancement, water quality improvement, toxic gas removal, improvement of feed conversion ratio. Zeolite, rotenone, disinfectant, oxygen precursors, ammonia reducers, and probiotics were applied for pond preparation, water, and soil quality maintenance, while 30 different antibiotics were used for the purpose of treatment. Among the available antibiotics, oxytetracycline, ciprofloxacin, enrofloxacin, erythromycin, sulphadiazine, and trimethoprim were found extensively used by the fish farmers. Four enzymes and eighteen growth promoters were identified as being utilized to enhance digestion and boost up the production. This study elicited various issues connected with application and administration of such aqua chemicals, including farmers’ ignorance about their usage, proper doses, application methods, withdrawal period, and the human health concerns associated with their irresponsible use. However, the consequences of these chemical products to the environment, animal health, and human health required further study for the betterment of mankind.

Aphanomyces invadans: The Causative Agent of the Epizootic Ulcerative Syndrome in Albino Northern Snakeheads

AbstractA suspected epizootic ulcerative syndrome with a 50–70% morbidity rate broke out in farmed albino‐type (AT) Northern Snakeheads Channa argus at Jinming farm, Chao‐Yang Town, southwest China. To investigate the pathogen, diseased fish were sampled and laboratory examinations were conducted. Three specific staining methods (Grocott’s hexamine silver, glycogen–periodic acid–Schiff, and modified Gram’s stain) were also applied for a clear observation of fungi in tissues. Histopathological changes were studied to reveal the pathogenic mechanism. Results showed that Aphanomyces invadans was the causative agent of the disease in AT fish, judging from the fungus‐like hyphae, fungal granulomas, and 99% similarity of PCR products to A. invadans (MN814020.1). Among the three fungal staining methods, Grocott’s hexamine silver was the best; this method dyed the fungal hyphae black, while the surrounding muscle was dyed brown. Histopathologically, the affected skin and muscle exhibited remarkable dermatitis, myositis, and fungal granulomas, whereas the other tissues (e.g., gills, liver, spleen, kidneys, and intestines) showed histopathological changes related to osmotic imbalance. In conclusion, the causative agent of disease in AT Northern Snakeheads was A. invadans, which invaded the skin and muscle, causing ulcers in the skin.

Epizootic ulcerative syndrome

This datasheet on epizootic ulcerative syndrome covers Identity, Overview, Associated Diseases, Pests or Pathogens, Distribution, Hosts/Species Affected, Diagnosis, Pathology, Epidemiology, Prevention/Control, Further Information.

Aeromonas veronii isolated from climbing perch (Anabas testudineus) suffering from epizootic ulcerative syndrome (EUS)

Motile Aeromonads are responsible for various diseases in fishes that cause significant economic losses in aquaculture. Aeromonas spp. are also frequently isolated from fish suffering from Epizootic Ulcerative Syndrome (EUS). The present study was conducted to isolate and identify A. veronii from Vietnam climbing perch (Anabas testudineus) suffering from EUS, evaluate the pathogenicity of the isolates, and find out antibiogram profile and susceptibility of the isolates to medicinal plant extracts. Vietnam climbing perch (A. testudineus) suffering from EUS was collected from fish farms located at Savar, Dhaka and bacteria were isolated from the external ulcerative lesion, and kidney and liver of the infected fish on nutrient agar plates. Preliminary phenotypic identification of the bacterial isolates was performed following morphological, physiological and biochemical characterization tests. The 16S rRNA gene sequencing was accomplished for further molecular identification of five selected isolates. To evaluate the pathogenicity of the isolates in vivo artificial infection challenge test was conducted through immersion method. Another in vivo pathogenicity test was done at three different temperatures (18 °C, 28 °C, and 37 °C) to examine the effects of temperature on the pathogenicity of the isolates. Ten commercial antibiotics were used to find out the antibiogram profile of the isolates. Finally, susceptibility of the isolates was screened against 19 medicinal plants extracts. A total of 38 out of 46 isolates isolated from Vietnam climbing perch (A. testudineus) suffering from EUS were phenotypically identified as Aeromonas sp. Five randomly selected isolates were further confirmed as A. veronii by analyzing the 16S rRNA gene sequence homology. The pathogenicity of the isolates was found moderate to high (41%–80% mortality) in the artificially challenged fish. The highest and lowest mortality was obtained in the fish challenged with the isolates BFKA33 and BFKA12, respectively. The isolates were found resistant to several antibiotics, including amoxicillin, ampicillin, penicillin-G, and vancomycin. Nevertheless, extracts of three medicinal plant (Allium sativum, Syzygium aromaticum, and Tamarindus indica) demonstrated antibacterial activity to the isolates (BFKA12, BFKA15, BFKA18, BFKA29, and BFKA33).

Wetlands, Fishes and Pandemics with Special Reference to India

Water is life, life is water. Water is indispensably important for sustenance of life. Wetlands serve as potential water bodies, harbouring coveted bioresources, which sustain animal life. Fish is a significant bioresource for nutrition and avocation of the people. There are various types of wetlands in the Indian sub-continent. India has c 67,429 wetlands covering c 4.1 million ha. Concomitantly, c 21,723 living species of fish have been recorded out of 39,900 species of vertebrates. Of these, c 8411 are freshwater (FW) species and c 11,650 are marine. India recorded c 2500 species of fishes; of which, c 930 live in FW and c 1570 are marine. The hitherto unknown dreadful, virulent, enigmatic Epizootic Ulcerative Syndrome (EUS), has been sweeping the FW fishes in an epidemic dimension, unhindered, unimpeded and unabated, almost semi-globally; and, has been causing large-scale mortality among them, since 1988, rendering many of them endangered. Concomitantly, the outbreak of SARS-CoV-2, among the human, was first reported at Wuhan, China, in late December 2019. The first 54 reported cases of COVID-19 were observed in December 2019 at Wuhan, China, and this, subsequently, had spread across the globe. India has been facing much impacts of COVID19 pandemic since its inception in China.

Epizootic Ulcerative Syndrome (EUS) Fish Disease Chronology, Status and Major Outbreaks in the World

Abstract Epizootic Ulcerative Syndrome (EUS) has been causing large-scale mortality among the freshwater fishes of the globe since the 1070s. The symptoms include large haemorrhagic cutaneous ulcers, epidermal degeneration and necrosis followed by sloughing of scales. There have been many studies on EUS throughout the world. In India, since the initiation of EUS, in 1988, our study tried to reveal the aetiology of the disease through extensive and intensive studies on different aspects, like limnological, physical, chemical, bacteriological, fungal, viral including electron microscopic studies. Details of EUS investigation has been discussed in the present paper.

Outbreaks of epizootic ulcerative syndrome in Kerala, India following episodes of flooding.

Large-scale fish mortality was observed in flood-affected fish farms across several parts of Kerala following heavy rainfall in August 2018 and 2019-nearly 53% above the normal monsoon rain that the region receives. The affected fish had severe haemorrhages and ulcers, typical of the highly infectious disease epizootic ulcerative syndrome (EUS) caused by the water mould Aphanomyces invadans. In freshwater, snakeheads Channa spp. and in brackish water mullet (Mugilidae) and pearl spot (Etroplus suratensis) were severely affected. EUS was observed in 4 freshwater fishes for the first time: dotted sawfin barb Pethia punctata (Cyprinidae), Malabar leaffish Pristolepis malabarica (Pristolepididae), mahecola barb Puntius mahecola (Cyprinidae) and giant snakehead Channa pseudomarulius (Channidae). Histology and molecular diagnosis confirmed the cause of mortality to be EUS. Fungal hyphae were also observed in deeply ulcerated fish, revealed by lactophenol cotton blue staining. The severity of the EUS outbreak was linked to the sudden change in water quality associated with the flood, such as lower water temperature, and decreases in pH, total alkalinity and total hardness.