Bull. Eur. Ass. Fish Pathol., 30(4) 2010, 146 Centrocestus formosanus metacercarial infection of four ornamental fish species imported into Iran

S. M. Mood1, H. A. Ebrahimzadeh Mousavi1*, B. Mokhayer1, M. Ahmadi1, M. Soltani1 and I. Sharifpour2

1 Department of Aquatic Health and Diseases , Faculty of Veterinary Medicine , University of Tehran; 2 Iranian Fisheries Research and Training Organization , Department of Fish Disease,Tehran, Iran

Abstract During an investigation into parasites of 11 most common imported ornamental fish species to Iran, metacercariae of the heterophyid trematode Centrocestus formosanus were found encysted in the gills of four fish species including Dwarf gourami (Colisa lalia), Goldfish (Carassius auratus), Red fin shark (Labeo erythrurus) and Arowana (Osteoglossum bicirrhosum). This is the first report of the parasite in Iran. The prevalence of metacercarial cysts ranged from 5% to 60% depending on fish species. The most infected species was Red fin shark.

Introduction In recent years, significant development in 1983; Madhavi, 1986), United States (Mitchell the ornamental fish industry in Iran has been et al., 2000) and other parts of the world (Vélez- observed. The majority of these ornamental Hernández et al., 1998; Hernández et al., 2003; fish are imported from countries of Southeast Salgado-Maldonado et al., 2005). In Turkey, Asia. New fish parasites have been introduced Yildiz (2005) first recovered the metacercariae to Iran through this trade (Mousavi, 2003). One of C. formosanus from the gills of ornamental of these high risk pathogens is Centrocestus fish imported from Singapore. The occurrence formosanus (Nishigori, 1924). This species was of C. formosanus has been not documented in originally described from Taiwan and is widely Iran. In this project we aimed to determine the distributed in Asia (Scholz, 2000). C. formosanus prevalence of this parasite in the most common is a trematode that produces severe alterations freshwater species of ornamental fish imported in the gills of many freshwater fishes, most in to Iran. notably cartilage proliferation around the encysted trematode in the branchial tissues. It Materials and methods has caused significant losses among cichlids, Between May 2008 and April 2009 a total characids, and cyprinids. The parasite has been of 550 specimens of the most frequently found in the gills of many fish species from imported species (50 samples of each species) numerous Asian countries (Yanohara & Kagei, including guppy (Poecilia reticulate), dwarf

* Corresponding author’s email: [email protected] Bull. Eur. Ass. Fish Pathol., 30(4) 2010, 147 gourami (Colisa lalia), goldfish (Carassius Results auratus), red fin shark (Labeo erythrurus), barb Four fish species out of 11 were infected with (Capoeta tetrazona), red-eye tetra (Moenkhausia metacercarial cysts of C. formosanus. The sanctaefilomenae), sucker catfish (Hypostomus encysted parasite was observed in the gill of 42 plecostomus), oscar (Astronatus ocellatus), sample (7.63%). Among the infected species, a pangasius sutchi (Pangasius hypophthalmus), variable prevalence rate was observed (Table angelfish (Pterophyllum scalare) and arowana 1). Excysted metacercariae were elongate and (Osteoglossum bicirrhosum) were obtained pyriform in shape, and completely covered from 5 commercial suppliers and taken alive with tegumental spines. The most parasitized to the lab. Fish were killed by incision using species was Red-fin shark. The important a scalpel into brain and subsequently the morphological features included the presence gills were dissected, placed in Petri dishes of 32 circumoral spines arranged in two and examined using a stereoscope. Cysts of rows and an X-shaped excretory bladder. infected gills (Figure 1A) were popped open Histopathological section of infected gills (by using fine needle forceps) in saline solution shows that parasite metacercariae lodged next releasing the live metacercaria (Figure 1B) to the filaments cartilage and caused intensive and observed under high magnification light inflammatory response including: Hyperemia, microscope. Photo and films were taken (Sony, hyperplasia of the cartilage of the primary SSC-DC80P-microscope digital camera). For lamellae that envelops the metacercarial cysts, histopathological examination, the gills of each secondary lamellae fusion, epithelial liVing, fish were fixed in 10% buffered formalin. The proliferation of fibroblasts and chondroblasts. fixed material was embedded in paraffin wax and serially sectioned. The 5 μm thick sections Discussion were stained with haematoxylin and eosin Centrocestus formosanus is so far known to (H&E). be distributed in Taiwan, China, Japan, the

Figure 1. C. formosanus. A: Metacercarial cysts (arrow) in infected gills (x4). B: Two row circumoral spines in metacercaria (x40) Bull. Eur. Ass. Fish Pathol., 30(4) 2010, 148

Table 1. PF: parasitized Fish; EF: Examined Fish , Prevalence (%) of the metacercarial cysts of C. formosanus in infected species

Species Common name PF/EF Prevalence(%) Labeo erythrurus Red- fin shark 28/550 60 Carassius auratus Goldfish 12/550 25 Colisa lalia Dwarf gourami 9/550 10 Osteoglossum bicirrhosum Arrowana 3/550 5

Philippines, Thailand, India, Turkey, Hawaii, freshwater ornamental fish imported into Iran. Vietnam, Croatia, USA, Mexico and Colombia Mousavi (2003) did not report C. formosanus in (Scholz, 2000). In the study of parasites of the previous study on ornamental fish parasite ornamental fish imported into Australia, Evans in Iran. To date, C. formosanus metacercariae & Lester (2001) had identified the parasites in have not been reported in native cultured Poecilia reticulata and Xiphophorus maculatus freshwater fishes in Iran. according to the two distinct features. Thilakaratne et al. (2003) had isolated C. Migration of cercariae through host tissues formosanus in gills of goldfish in Sri Lanka. Yildiz may produce minimal mechanical damage (2005) reported infection with metacercariae of and hemorrhaging (Hoffman 1999). C. C. formosanus in 4 species of ornamental fish formosanus, unlike most trematodes, produces imported into Turkey. Also the same infection serious damage in the gills of many freshwater in goldfish (Carassius auratus) imported into fishes (Mitchell et al., 2000). Histopathological Croatia have been reported by Gjurcevic (2007). section of infected gills shows that parasite The examined goldfish in our study were also metacercariae lodged next to the filaments parasitized by the C. formosanus. But it is the first cartilage and caused intensive inflammatory record of the metacercariae of C. formosanus in response including : hyperplasia of the

Figure 2. Histological section of infected gills. A: encysted metacercariae (white arrow) fusion of gill filaments (Black arrows) (x10) ; B: Epithelial liVing (White arrow) and hyperemia (Black arrows) (x40) Bull. Eur. Ass. Fish Pathol., 30(4) 2010, 149

Croatia. Helminthologia 44(4), 214-216. cartilage of the primary lamellae that envelops the metacercarial cysts, lamellar fusion, Mitchell AJ, Salmon MJ, Huffman DG, Goodwin epithelial liVing, proliferation of fibroblasts AE, and Brandt TM (2000). Prevalence and and chondroblasts that form a cartilaginous pathogenicity of a heterophyid trematode infecting the gills of an endangered fish, the encapsulation around the parasite resulting fountain darter, in two central Texas spring- destroying the normal gill structure and fed rivers. Journal of Aquatic Animal Health 12, reducing the surface area of the respiratory 283-289. epithelium. The same changes previously Mousavi HAE (2003). Parasites of ornamental described by other authors that reported C. fish in Iran. Bulletin of the European Association formosanus infection (described by Blazer and of Fish Pathologists 23(6), 297-300. Gratzek (1985), Vélez-Hernández et al. (1998), Mitchell et al. (2000), Yildiz (2005) and Gjurcevic Salgado-Maldonado G, Aguilar-Aguilar R, Cabanas-Carranza G, Soto-Galera E and (2007). Mendoza-Palmero C (2005). Helminth parasites in freshwater fish from the Papaloapan river In the present study, it has been confirmed basin, Mexico. Parasitology Research 96, 69–89. that C. formosanus is prevalent in freshwater Scholz T and Salgado-Maldonado G (2000). ornamental fish imported from Southeast Asia The introduction and dispersal of Centrocestus into Iran. According to the considerable role of formosanus (Nishigori, 1924) (Digenea: geographical transfer in alteration of infestation Heterophyidae) in Mexico: A review. American to infection and the risk of introduction of this Midland Naturalist 143, 185-200. new parasitic fauna to the native fish species, it Thilakaratne IDSIP, Rajapaksha G, Hewakopara seems important to quarantine and monitoring A, Rajapakse RPVJ and Faizal ACM (2003). the ornamental fish importation. Parasitic infections in freshwater ornamental fish in Sri Lanka. Diseases of Aquatic Organisms References 54, 157-162. Blazer VS and Gratzek JB (1985). Cartilage Vélez-Hernández EMF, Constantino-Casas proliferation in response to metacercarial LJ, García-Márquez and Sarabia O (1998). Gill infections of fish gills. Journal of Comparative lesions in common carp, Cyprinus carpio L., in Pathology 95, 273-280. Mexico due to the metacercariae of Centrocestus formosanus. Journal of Fish Diseases 21, 229-232. Erkin Koyuncu C (2009). Parasites of ornamental fish in Turkey .Bulletin of the European Association Yildiz HA (2005). Infection with of Fish Pathologists 29(1), 25-27. metacercariae of Centrocestus formosanus (Trematoda:Heterophyidae) in ornamental fish Evans BB and Lester RJG (2001). Parasites imported into Turkey. Bulletin of the European of ornamental fish imported into Australia. Association of Fish Pathologists 25(5), 244-246. Bulletin of the European Association of Fish Pathologists 21(2), 51-55.

Gjurcevic E, Petrinec Z, Kozaric Z, Kuzir S, Gjurcevic Kantura V, Vucemilo M and Dzaja P (2007). Metacercariae of Centrocestus formosanus in goldfish (Carassius auratus L.) imported into