Fungal infection in an imported Ornamental fish Fantail, (Carassius auratus L.)

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1 BIOLOGIA (PAKISTAN) June, 2015, 61 (1), PKISSN (Print) ISSN X (On-Line) Fungal infection in an imported Ornamental fish Fantail, (Carassius auratus L.) ZAFAR IQBAL & HUSNA NOREEN Department of Zoology, University of Punjab, Quaid-E-Azam campus, Lahore, Pakistan ABSTRACT In the present study fungal diseases in pet fish fantail are investigated by isolating the fungi from the fish. The Infected fish showed eroded fins, and wounds on the body. Twenty out of thirty (66%) fish specimen showed fungal infection. Material from diseased area of fish body was inoculated (in Laminar flow cabinet) on Malt extract agar and Sabouraud dextorse agar and incubated at 27±2ºC for 5-6 days. Fungal genera isolated from fish were, Penicillium sp. and Fusarium sp. (3.3% each), Rhizopus sp. (6.6%), Mucor sp. and Alternaria sp. (10% each) and Aspergillus sp. (100%). Abdomen (100%) and gills (65%) were the most affected areas followed by fins ( %) and head (6.0%). Infection on anterior area and posterior parts of the fish had no difference (p=0.05). Pure culture showed uniform colonies of fungi, indicating the growth of one species of fungus. The results of present study shows that imported infected fish may have adverse effects on the emerging pet fish trade in Pakistan. Key Words: Ornamental fish, fungal infection, Aspergillus sp, Fusatium sp. Alternaria sp INTRODUCTION Worldwide trade of ornamental fishes is one of the major causes of introduction of serious pathogens in importing countries. Such fishes in tanks and during shipment are infected by various types of pathogens (Conroy et al. 1982). Imported fish may carry harmful parasites and pathogens. Such pathogens are reported from imported fishes in many countries such as: Germany, Norway, Australia, Brazil, Korea, Sri Lanka as well as in Pakistan (Iqbal & Haroon, 2014, Iqbal & Mumtaz, 2013, Iqbal & Sajjid, 2013). Fungi cause superficial and systemic type of infection in the host fish. According to report of Czeczuga et al. (2004) Saprolegnia salmonis and Aphanomyces spp. were isolated from eggs of a fish Coregonus sp. Fungal infection in rainbow trout, Oncorhynchus mykiss eggs was studied by Ebrahimzadeh et al. (2007) in Iran. They indicated that Saprolegnia parasitica was the most important species causing infestation. Water quality unsuitable for fish; facilitate growth of fungi in earthen ponds. Saprolegnia sp., and Achyla sp. has been recorded from two commercial fishes C. idella and C. catla (Iqbal et al., 2012a). Five fungal species have also been isolated from Labeo rohita, H. molitrix and goldfish (Iqbal et al., 2012b). In another study Haroon et al, (2014) has described fungal infection in ornamental fishes which result in economic loss. Hence, the goal of this study was isolation of fungi from fantail, a freshwater pet fish. MATERIALS AND METHODS The experimental fish specimens were obtained from a local market in Lahore. The fish were kept in Fish Disease and Health Management laboratory, Zoology Department, University of the Punjab, Lahore in glass aquarium (capacity 40 liters) containing freshwater at 25-28ºC with continuous oxygen supply. Morphometric readings and health status of fish was observed. Fantail ranged in total length from cm and body weight was g. Two types of agar; Malt Extract Agar, Sabouraud Agar MEA and SDA (Oxoid, UK) was used to make culture plates. Isolation and identification of fungi from fish and its pure culture was carried out following Iqbal & Saleemi (2013). Difference in infection level on posterior and anterior part of the body of fish was calculated by Chi-square test. Posterior part included area beyond middle of the body and anterior part comprise area prior to pelvic fin. RESULTS AND DISCUSSION Infected fish (n=20, 66%) showed pathological signs like, ruptured dorsal as well as anal fins. There were bloody wounds on body and in caudal region. One hundred sixty six SDA and one hundred thirty four MEA culture plates were inoculated from infected fishes (Table 1). Six fungal genera: Aspergillus sp; Fusarium sp. Rhizopus sp. Alternaria sp. Mucor sp. and Penicillium sp. isolated from infected fish (Fig.1ABC (Aspergillus sp.); 2AB (Mucor sp.); 3 (Aspergillus sp.); 4 (Fusarium sp.); 5 (Rhizopus sp.); 6 (Alternaria sp.). Infection of various fungal genera was variable; Aspergillus 100%, Alternaria sp. and Mucor 10%. Rhizopus sp. 6.6%, Fusarium sp. and Penicilium sp. 4% infection. Both posterior and anterior part of the fish had no significant difference of infection (p = 0.05, χ² = 5.99). Gills and eyes were infected by two fungal species. Caudal fin harboured four fungal species *Corresponding author: dr.zafariqbal.pu@gmail.com.

70 Z. IQBAL & H. NOREEN BIOLOGIA (PAKISTAN) (Aspergillus sp. Alternaria sp. Fusarium sp. Mucor sp.). The fins were highly affected sites. Concurrent infection such as: Aspergillus and Mucor sp.

Four fungal genera were isolated from head, eyes, and gills (Aspergillus sp. Rihzopus sp. Mucor sp. Penicillium sp.) Table 1.

2 70 Z. IQBAL & H. NOREEN BIOLOGIA (PAKISTAN) (Aspergillus sp. Alternaria sp. Fusarium sp. Mucor sp.). The fins were highly affected sites. Concurrent infection such as: Aspergillus and Mucor sp. (in eyes) and Aspergillus and Rhizopus on gills and pectoral fins were common. Aspergillus was the most common fungus. which was isolated from all parts of fish. Four fungal genera were isolated from head, eyes, and gills (Aspergillus sp. Rihzopus sp. Mucor sp. Penicillium sp.) Table 1. Infection around head area may prove dangerous, because treatment of such fish is not possible as the hyphae penetrate from eyes into head and brain. This type of infection causes complete blindness of eyes as reported by Srivastara (2009). The results of this study are comparable to the finding of Haroon et al., (2014), who reported isolation of five fungal genera; Aspergillus sp. Rhizopus sp. Mucor sp. Penecillium sp. and Alternaria sp. from goldfish C.auratus. The most affected part was caudal fin of the fish. Caudal fin has been highly affected in Mystus seenghala and Puntius sarana as reported by Chauhan et al. (2014). They isolated three fungal genera from this site; Aspergillus niger, A. fumigatus, A. sydowii. Fin infection is generally less serious. But, high infection may cause complete damage of the fins (Iqbal et al, (2012a). Dorsal fin infection of Catla catla has also been reported by Iqbal & Saleemi (2013). Damage of dorsal and caudal fins of Poecilla reticulata is already described by Haroon et al. (2014). Aspergillus niger is pathogenic to human and is environmental contaminant (Alinezhad et al., 2011, Saleemi et al., 2012). The fungal species isolated in this study are considered as normal mycoflora but become pathogenic under environmental stress conditions. Our findings are comparable to Rafai et al. (2010). They isolated Saprolegnia, Aspergillus, Fusarium, Mucor, Pencillium, Rhizopus from Oreochromis sp. and Clarias gariepinus. Adb-El-Ghany et al. (2014) reported three species of Fusarium from a fish, Symphysodon sp. Chauhan et al. (2014) described three fungal species, Aphanomyces laevis, A. niger and Saprolegnia from goldfish. Environmental factors influence level of infection (Fadaeifard et al., 2011). Isolated fungi observed from fantail is considered non pathogenic, but are responsible to produce diseases under particular predisposing environmental conditions. Such as, water temperature, moisture in feed which contaminate it and increase risk of fungal infection in fish. According to Chauhan et al., (2014) change in water temperature and poorly maintained aquarium causes zoospores to grow which facilitate fungal attack on fish. This study confirms that imported fantail was having fungal infection. The control on import of diseased fish into the country is urgently needed. This step will reduce the chance of spreading fungal infection in the native fish species. Acknowledgements Our thanks to University of the Punjab, for financial support for this study and Chairman Department of Zoology for providing facilities to complete this work. Fig., 1A: Colony of Aspergillus sp. on SDA isolated from caudal fin of fish No. 3. Fig., 1B: Pure culture of fungus made from plate 1A.

VOL. 61 (1) FUNGAL INFECTION IN FISH FANTAIL

, 1C: Trypanblue stained Aspergillus sp.

3 VOL. 61 (1) FUNGAL INFECTION IN FISH FANTAIL 71 Fig., 1C: Trypanblue stained Aspergillus sp. isolated from plate 1B. Fig., 2B: Trypanblue stained Mucor sp. isolated from plate 2A. Fig., 2A: Pure colony of Mucor sp. grown on SDA from eye of fish No.2. Fig., 3: Aspergillus sp. isolated from fungal colony grown on MEA inoculated from pectoral fin Fish No.30

72 Z. IQBAL & H. NOREEN BIOLOGIA (PAKISTAN) Fig., 4: Fusarium sp. isolated from caudal fin of fish No. 15 Fig.

isolated from caudal fin of fish No. 1. Table I: Culture plates prepared from each organ and fungal genera isolated Sr. No. Organ Fish No.

3 Caudal fin 20 40 Aspergillus sp. Alternaria sp. Fusarium sp. Mucor sp. 4 Abdomen 22 44 Aspergillus sp.

4 72 Z. IQBAL & H. NOREEN BIOLOGIA (PAKISTAN) Fig., 4: Fusarium sp. isolated from caudal fin of fish No. 15 Fig. 5: Trypanblue stained Rhizopus sp. isolated from pectoral fin of fish No. 12. Fig. 6: Trypanblue stained Alternaria sp. isolated from caudal fin of fish No. 1. Table I: Culture plates prepared from each organ and fungal genera isolated Sr. No. Organ Fish No. of Fungal genera samples plates 1 Gills Aspergillus sp. Rhizopus sp. 2 Dorsal fin Aspergillus sp. 3 Caudal fin Aspergillus sp. Alternaria sp. Fusarium sp. Mucor sp. 4 Abdomen Aspergillus sp. 5 Eye Aspergillus sp. Mucor sp. 6 Pectoral fin Aspergillus sp. Rhizopus sp. 7 Pelvic fin Aspergillus sp. 8 Head 9 18 Penicillium sp. 9 Anal fin Aspergillus sp. Total 300

5 VOL. 61 (1) FUNGAL INFECTION IN FISH FANTAIL 73 REFERENCES Adb-El-Ghany, N.A., El-khatib N. R & Salman- Soad, S.A., Causes of mortality in Discus fish (Symphysodon) and trials for treatment. Egy. J. Aquac, 4(2): Alinezhad, S., Toluee, M., Kamalzadeh, A., Motalebi, A. A., Nazeri, M., Yasemi, M., Shams G. M., Tolouei, R & Razzaghi, A.M., Myobiota and aflatoxin B1 contamination of rainbow trout (Oncorhynchus mykiss) feed with emphasis to Aspergillus section Flavi. Iran. J. Fish., 10: Chauhan, R., Lone, S. & Beigh. A. H., Pathogencity of three species of Aspergullis (A.fumigatus, A. niger & A. sydowii) on some freshwater Fishes. Life Science Leaflets, 14: (Online print) Conroy, D.A., Morales, J., Pardomo, C., Ruit, R. A. & Santacana. J. A., The prevention and control of diseased conditions in South American ornamental fish. Riv. Ital. Piscicult. Ittiopatol, 17: Czeczuga, B., Kiziewiez, B & Godlewska, A., Zoosporic fungigrowing on egg of Coregonus larvaretus holstus. Thieneman, 1916.From lake wdzyde in Kazuby. Poilsh J. environ. Stud., 13: Ebrahimzadeh, M. H., Hoseinifard, S., Khosravi, A., Soltani, M & Yoosefian, M., Isolation and identification of saprophytic fungi from Rainbow trout infected eggs in farms of Mazandaran Province, Iran. Iran. J. Vet. Res., 62: Fadaeifard, F., Raissy, M., Bahrami, H., Rahimi, E & Najafipoor, A., Freshwater fungi isolated from eggs and broodstocks with anemphasis on saprolegnia in rainbow trout farms in West Iran. Afr. J. microbiol. Res., 4: Haroon, F., Iqbal, Z., Perviaz, K., & Khalid, A.N Incidence of Fungal Infection of Freshwater Ornamental Fish in Pakistan. Int. J. Agri. Biol., 16(3): Iqbal, Z., Asghar, M & Rubaba., 2012a. Saprolegniasis in two commercially important carps. Pakistan J. Zool., 44: Iqbal, Z., Sheikh, U. & Mughal, R., 2012b. Fungal infection in some economically important fresh water fishes. Pakistan Vet. J., 32: Iqbal, Z. & Mumtaz, R., Some fungal pathogens of an ornamental fish, Black Moor (Carassius auratus L.). Eur. J. Vet. Med., 2: 1-10 Iqbal, Z. & Sajjid. R Some pathogenic fungi parasitizing two exotic tropical ornamental fishes. Int. J. Agri. Biol., 15: Iqbal, Z & Saleemi, S., Isolation of pathogenic fungi from a freshwater commercial fish Catla catla. Sci. Intl, 25(4): Iqbal, Z & Haroon, F., Parasitic infections of Some Freshwater Ornamental fishes Imported to Pakistan. Pak. J. Zool., 46(3): Rafai, M.K., Laila, A.M., Amany, M.K. & Shimaa, M. K. A., The Assessment of Myotic Settlement of Freshwater Fishes in Egypt. J. Amer. Sci., 6: Saleemi, M. K., Khan, M. Z., Khan, A., Mehmood, M.A., Farooq, M., Hameed., S., Hassan, Z.U., Javed, M. R & Javed. I., Molecular identification of black Aspergilli isolated from poultry feeds by sequencing of its region. Pak. Vet. J., 32: Srivastara, R.C., Fish Mycopathology, Today and Tomorrow s printers and publishers New Dehli, India. 103 PP. Received: Revised: Accepted:

Pakistan Veterinary Journal

RESEARCH ARTICLE Pakistan Veterinary Journal ISSN: 0253-8318 (PRINT), 2074-7764 (ONLINE) Accessible at: www.pvj.com.pk Fungal Infections in Some Economically Important Freshwater Fishes Zafar Iqbal*, Uzma