MYCOTIC INFECTION IN SOME ECONOMICALLY IMPORTANT FRESHWATER FISHES

WORLD JOURNAL OF PHARMACY AND PHARMACEUTICAL SCIENCES Mastan et al. SJIF Impact Factor 5.210 Volume 4, Issue 11, 1449-1456 Research Article ISSN 2278 4357 MYCOTIC INFECTION IN SOME ECONOMICALLY IMPORTANT FRESHWATER FISHES S. A. Mastan 1*, S. A. Begum 2, Md. Osman Ahmed 3 and S. Shamshad 4 1 Post Graduate Department of Biotechnology, PG Courses and Research Centre, DNR College, Bhimavaram-534 202. 2 Department of Biological Sciences, D N R College of Education, Bhimavaram-534 202. 3 Department of Zoology, (UG & PG) Osmania College (Autonomous), Kurnool-518 004. 4 Department of Zoology, KVR Govt Degree College for Women, Kurnool-518 004. Article Received on 07 Sept 2015, Revised on 01 Oct 2015, Accepted on 24 Oct 2015 *Correspondence for Author S. A. Mastan Post Graduate Department of Biotechnology, PG Courses and Research Centre, DNR College, Bhimavaram-534 202. ABSTRACT A total five species of fungi were isolated from diseased fishes collected from local water bodies and fish markets of Gudiwada and Kaikalur mandals of Krishna district, Andhra Pradesh. The isolated fungal pathogens were belongs two genera and five species and were identified as Saprolegnia parasitica, S.diclina, Achlya americana, A.flagellata and A.prolofera. They were isolated from eight species of fresh water fishes namely Labeo rohita,catla catla,channa striatus, C.punctatus, Clarias batrachus, Mystus cavasius, M.seenghala and Tilapia mossambica. S.parasitica was associated with six species of fishes while A. prolifera was associated two fish species only. The maximum percentage (7.9%) of infection was observed in Channa striatus and minimum (2.5%) was observed in Tilapia mossambica. The artificial infection trails indicates that Saprolegnia parasitica is found to be more virulent to the freshwater fishes. KEYWORDS: Mycotic infection, Fungi, Fishes. INTRODUCTION Most fungal infections only attack the external tissue and few fungal infections that will infect the internal organs of fishes. [1] Aquatic fungi are responsible for significant infections, involving both cultured and wild fish and their eggs, particularly in aquaculture. The members of Oomycetes are saprophytic opportunists multiplying on fish that are physically www.wjpps.com Vol 4, Issue 11, 2015. 1449

injured, stressed or infected. [2] They are generally considered agents of secondary infections arising from conditions such as bacterial infection, poor husbandry practices, and infestations by parasites and social interactions. However, there are several reports of Oomycetes as infectious agents of fish and their eggs. [3,4,5,6,7,8,9,10,11,12] The present paper communicates the Mycotic infection in some freshwater fishes of Andhra Pradesh, India. MATERIALS AND METHODS Collection of diseased fish Fungal infected fishes were collected from cultured ponds and local fish markets of Gudiwada and Kaikalur mandals of Krishna Dist, A.P. A total of 942 fishes were screened. The infected fishes were brought to the laboratory in the living condition and kept in glass aquarium, of the size of 90x45x45 cm filled with clean fresh water. The dead as well as living fishes were examined grossly for lesions and ulcerations. Isolation of fungi Fungal isolation from diseased fishes was carried out by taking small pieces of muscles about 2 mm in diameter from infected portions of the body. They were then washed thoroughly with sterilized distilled water to remove the unwanted micro organisms adhered on the surface. These tissues were then inoculated over the Sabourauds Dextrose Agar (SAD) and Potato Dextrose Agar (PDA) plates. Alternatively small pieces of mycelia taken out from infected parts of fish body were washed thoroughly with distilled water. They were placed in a Petri dish containing 20-30 ml distilled water and baited on Hemp seeds. These Petri dishes were incubated at 15 0 C to 22 0 C for a week. Pure and bacteria free cultures were prepared by using the methods of Coker [13] Johnson [14] and Scott. [15] Identification of fungi was done on the basis of their vegetative and reproductive characters using the monographs of Coker [13] and Khulbe. [16] Artificial infection trials In order to demonistrate the pathogenicity of the isolates obtained from the naturally infected fishes, experimental infection trails were conducted in the laboratory. Isolated species of fungi viz. S. parasitica, S.diclina, Achlya americana, A.flagellata and A.prolifera were tested separately on the fingerlings of different species of fishes, having average size and weight 8.16 ± 0.13 cm and 12.5 ± 0.28 gm, respectively. Experimental infection trails were carried out by employing the methodology of Scott and O warren. [3] Covered glass troughs (12 x 9) wrapped in aluminum foils were sterilized in hot air oven at 120 0 C tem for 24 hours. Filtered www.wjpps.com Vol 4, Issue 11, 2015. 1450

sterile lake water was filled aseptically in to each trough. An aerator was used to aerate the water throughout the experiment. Six fungal inoculated blocks (1.0 cm 2 ) of SPS agar/ Potato Dextrose Agar (PDA) medium were placed at different sites in the trough. Six uninoculated blocks of the same agar medium were placed in another trough which was used as control. After 48 hours, when spores developed, experimentally injured fishes were placed in these troughs. Six fishes of each species were kept in each trough. All the experiments were conducted at 20.0 0 C to 25.0 0 C temperature in triplicate sets. RESULTS AND DISCUSSION In the present study, it has been observed that a total 157 fishes were infected with fungus. Among the fishes examined, maximum infection (7.9 %) was noticed in Channa striatus, while minimum infection (2.5%) was noticed in Tilapia mossambica (Fig-1). The fungal species isolated from infected fishes are Saprolegnia parasitica (38%), S.diclina (31%), Achlya americana (10%), A.prolifera (14%) and A.prolofera (7%). The details of isolation of fungal species from fishes are presented in Table-1&Fig-2. Saprolegnia parasitica was isolated from eight species of fishes such as Labeo rohita,catla catla,channa striatus, C.punctatus, Clarias batrachus, Mystus cavasius, M.seenghala, Tilapia mossambica. S.diclina was isolated from Channa striatus, C.punctatus, Clarius batrachus, Mystus seenghala and Tilapia mossamica. Achlya Americana was isolated from Channa striatus, C.punctatus and Mystus cavasius while A. prolifera was isolated from C.stratius, C.punctatus and Clarias batrachus. The infected fishes showed clinical symptoms such as lethargic behavior, de-colouration of body, grayish cotton like patches on the surface of body. In most of fishes formation of hemorrhagic lesions and ulcerations were observed on head and tail region. In some fishes especially C.punctatus tail was found completely lost. The results of artificial infection trails are presented in Tables 2-6. The artificial infection trails indicated that all fungal species were pathogenic to test fishes. But S. parasitica is highly pathogenic to test fishes and it causes infection within eighteen hours. In India the mycological studies were initiated by Chidambaram [17] who observed red patches on the body of Osphronemus gouramy due to Saprolegnia species. Tiffney [18] was the first to demonstrate the ability of Saprolegnia parasitica (Coker) to parasitize a wide range of fishes and amphibians and emphasized the fact that the injury greatly lowers the resistance of hosts to fungal infections. Vishniac and Nigrelli [19] conducted laboratory experiments and demonstrated the parasitic ability of sixteen species of aquatic fungi belonging to seven genera of Saprolegniaceae. Scott [20] demonstrated that Saprolegnia parasitica, Saprolegnia ferax, Saprolegnia diclina, www.wjpps.com Vol 4, Issue 11, 2015. 1451

Saprolegnia monoica, Achyla bisexualis and some non fruiting isolates of Saprolegnia could parasitize wounded platy fish under controlled conditions. Sati and Khulbe [21] carried out host range studies with Saprolegnia diclina on nine species of cold water fishes namely Barilius bendelisis, Carassius auratus, Cyprinus carpio, Nemachelius rupicola, Puntius conchonius, Puntius ticto, Schizothorax palgiostomus, Saprolegnia richardsoni and Tor tor. The experimental infection of Saprolegnia on different species of fishes has also been reported by Qureshi et al., [22] Hatai et al., [23] reported the pathogenicity of Aphanomyces species on Dwarf gourami. Kitanchroen and Hatai [9] have conducted experimental infection trials with Saprolegnia sp. on Rainbow trout eggs. Ramaiah [24] has reviewed the fungal diseases of algae, marine fishes, and shrimps corals. Zafar Iqbal and Saira Saleemi [26] have isolated pathogenic fungi from freshwater fish, Catla catla. They have isolated Aspergillus spp., Blastomyces sp., Penicilium sp. and unidentified fungal hyphe from the Catla catla of Pakistan. In the present study mycological examination of infected fishes revealed the presence of sixteen isolates of five species viz. S. diclina, S. ferax, S. hypogyana, S. parasitica and A. americana. All the species of Saprolegnia are found to be virulent for fishes. This observation is in agreement with the finding of Scott and O Bier [3] who have reported that the species of fungi, S. parasitica is found to be the most destructive. This finding confirms with the reports of Hatai and Hoshiai [26] who have reported that the infection caused by Saprolegnia parasitica in salmon resulted mass mortality. Both the scaly and non scaly fishes were found to be equally susceptible to the species of fungi tested. S. hypogyana was isolated from and tried on Channa straitus, also showed its wide range on fishes. The same is also reported by Chauhan and Qureshi. [27] Qureshi et al., [28] have carried out pathogenicity studies with various species of Saprolegnia on different species of fishes of central India. S.NO Table: 1 Details of fungal species isolated from diseased fishes Isolated fungal species infected fishes 1 Saprolegnia parasitica 2 S.diclina Infected fishes from which fungal pathogens isolated Labeo rohita, Catla catla, Channa striatus, C.punctatus,Clarias batrachus, Mystus cavasius, M.seenghala, Tilapia mossambica Channa striatus, C.punctatus, Clarias batrachus, M.seenghala, Tilapia mossambica 3 Achlya americana Channa striatus, C.punctatus,Clarias batrachus, Mystus cavasius, 4 A.flagellata Channa striatus, C.punctatus, Mystus cavasius, 5 A.prolifera Channa striatus, C.punctatus,Clarius batrachus www.wjpps.com Vol 4, Issue 11, 2015. 1452

Table 2: Artificial infection trails conducted with S. parasitica on different species of fishes S.NO Fish species No. of fish used Infection in hrs Death in hrs 1 Labeo rohita 6 24 40 2 Catla catla 6 18 32 3 Channa striatus 6 18 30 4 C.punctatus 6 18 36 5 Clarias batrachus 6 32 38 6 Mystus cavasius 6 24 40 7 M.seenghala 6 24 38 8 Tilapia mossabica 6 24 40 Table 3: Artificial infection trails conducted with S.diclina on different species of fishes S.NO Fish species No. of fish used Infection in hrs Death in hrs 1 Labeo rohita 6 30 40 2 Catla catla 6 32 42 3 Channa striatus 6 24 36 4 C.punctatus 6 24 36 5 Clarias batrachus 6 30 38 6 Mystus cavasius 6 36 42 7 M.seenghala 6 38 42 8 Tilapia mossabica 6 36 42 Table 4: Artificial infection trails conducted with A. americana on different species of fishes Table 5: Artificial infection trails conducted with A. flagelleta on different species of fishes. S.NO Fish species No. of fish used Infection in hrs Death in hrs 1 Labeo rohita 6 32 45 2 Catla catla 6 32 45 3 Channa striatus 6 26 38 4 C.punctatus 6 28 40 5 Clarias batrachus 6 30 42 6 Mystus cavasius 6 38 45 7 M.seenghala 6 38 45 8 Tilapia mossabica 6 40 48 S.NO Fish species No. of fish used Infection in hrs Death in hrs 1 Labeo rohita 6 36 48 2 Catla catla 6 34 48 3 Channa striatus 6 30 52 4 C.punctatus 6 32 48 5 Clarias batrachus 6 38 48 6 Mystus cavasius 6 32 45 7 M.seenghala 6 32 45 8 Tilapia mossabica 6 36 48 www.wjpps.com Vol 4, Issue 11, 2015. 1453

Table 6: Artificial infection trails conducted with A.prolifera on different species of fishes S.NO Fish species No. of fish Infection in Death in used hrs hrs 1 Labeo rohita 6 36 45 2 Catla catla 6 38 48 3 Channa striatus 6 30 48 4 C.punctatus 6 30 48 5 Clarias batrachus 6 32 45 6 Mystus cavasius 6 40 42 7 M.seenghala 6 42 48 8 Tilapia mossabica 6 40 45 Fig.1: Percentage of fungal infection of different fish species Fig 2: Percentage of fungal species isolated from diseased fishes. www.wjpps.com Vol 4, Issue 11, 2015. 1454

CONCLUSION The incidences of fungal infections varied with fungal species and the season of the year. The highest prevalence of infection was observed in colder months from November to January and the lowest during February to July. Low temperature is conducive for pathogenic potentiality of the water molds, retardation of pathogenic potentiality of the water molds at higher temperature above 28ºC, which cannot infect and require a certain period for multiplication. In the present study, it was observed that S. parasitica and A. americana were found to be the most common water molds responsible for the fungal infections to freshwater fishes. Saprolegnia parasitica is found to be more virulent for freshwater fishes. REFERENCES 1. Vinay Varma. Fungus disease, diagnosis and treatment, Veterinary World, 2008; 1(2): 62 2. Pickering AD, Willoughby LG. Saprolegnia infection of salmonid fish. In: 50 th Annual Report, Institutes of freshwater Ecology, Windermere Laboratory, England, 1982; 38-48. 3. Scott WW, O Bier, AH. Aquatic fungi associated with diseased fish and fish eggs. Prog. Fish Cult, 1961; 24: 3-15 4. Bhargava KS, Swarup K, Singh CS. Fungi parasitic on certain fresh water fishes of Gorakhpur, Indian, Biologist, 1971; 3: 65-69. 5. Willoughby LG. Saprolegniasis of Salmonid fish in Windermere, a critical analysis, J Fish Dis, 1978; 1:51-67. 6. Sati SC. Two fungal parasites of the eggs of Tor putitora Himalaya, 1991; 57: 95-96. 7. Walser CA, Phelps RP.The use of Formalin and iodine to control Saprolegnia infections on Channel catfish, Talurus punctatus eggs, J Appl Aqua,1993; 3: 269-278 8. Khulbe RD, Joshi C, Bisht GS. Fungal diseases of fish in Nanak Sagar, Nainital, India Mycopathol, 1995; 130: 71-74. 9. Kitancharoen N, Hatai K. Experimental infection of Saprolegniasis in rainbow trout eggs, Fish patho, 1996; 31(1): 49-50. 10. Sindhu, Rajender K, Khulbe RD. A survey of impact and evaluation fisheries in Kumann Himalayas, India with special reference to micro organism, Cur Sci, 1998; 75(12): 1303-1308. 11. Vikas Salgotra, Mastan SA, Qureshi TA (2005). Incidences of Saprolegniasis in fishes of Hataikhada reserviour, Bhopal, Indian J fish, 2005; 52(3): 367-37 12. Rekha Chauhan.Fungal attack on Tilapia mossabicus in culture ponds, leading to mortality of fishes., 2014; 5(7): 425-428. www.wjpps.com Vol 4, Issue 11, 2015. 1455

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