Cryptosporidium sp. (Apicomplexa) from cultured turbot Psetta maxima

During a survey conducted in a fish farm located in NW Portugal Cryptosporidium sp. sporogonial stages were detected in bile ducts epithelial cells of turbot (Psetta maxima). Fibrosis surrounding the infected bile ducts and necrosis of adjacent hepatic cells were observed. *Corresponding author’s E-mail: [email protected] The turbot, Psetta maxima (Linnaeus, 1758) is a very important marine fish farming species in Europe. In the last years it is estimated that the European production of turbot is about 6.000 tonnes per year (FAO Fishery Statistic, 2002) being the main producer countries located in Southwestern Europe. In Portugal production is about 350 to 400 tonnes per year. One of the major problems to the intensive culture of this fish species is the susceptibility to several pathogens. Fish coccidia have a considerable pathogenic potential, especially in intensive fish cultures where it can quickly spread and cause high losses (Lom & Diková, 1992). Cryptosporidium molnari AlvarezPelliteiro & Sitjà-Bobadilla, 2002, a parasite frequently found in young gilthead sea bream (Sparus aurata L.) and European sea bass (Dicentrarchus labrax L.), and Cryptosporidium scophthalmi Alvarez-Pellitero, Quiroga, SitjàBobadilla, Redondo, Palenzuela, Pádrós, Vásquez & Neto, 2004 from cultured turbot are coccidian species that cause important histopathological damage in the stomach and intestinal epithelial cells and are associated with trickling mortalities and retarded growth (Alvarez-Pelliteiro & Sitjà-Bobadilla, 2002; Alvarez-Pellitero et al., 2004; Sitjà-Bobadilla et al., 2006). This paper reports the first occurrence of a bile duct Cryptosporidium species in a turbot population also infected with a bacterium and a parasite ciliate. From March 2004 to February 2005 surveys were conducted in a turbot farm located in NW Portugal in order to determine the cause/ s of mortality. Around 110 fishes were examined (total weight 20 to 1020g). Moribund fishes were killed by spinal cord severance and necropsy. For bacteriological studies swabs from several organs were taken Bull. Eur. Ass. Fish Pathol., 29(1) 2009, 35 Figures 1 to 3. Developmental stages of Cryptosporidium sp in the bile ducts epithelial cells of turbot, Psetta maxima. Figure 1. Fibrosis (F) and necrotic hepatic tissue (NT) in turbot liver infected with Cryptosporidium sp. Figure 2. Mature oocysts with 4 sporozoites (arrows). Figure 3. Oocysts in different stages of sporogony and a free sporozoite (arrow). (H&E;). to different medium culture. Simultaneously squash smears and samples for histology were taken from different organs. Histological material were fixed in 10% buffered formalin, embedded in paraffin, routinely processed, sectioned at 2 μm and stained with haematoxylin and eosin (H&E;). Macroscopically fish were emaciated, with darkened and ulcerated skin, haemorrhages Bull. Eur. Ass. Fish Pathol., 29(1) 2009, 36 authors; however due to the scarcity ofavailable material a specific identification ofthis parasite was not possible. As fishes wereinfected with several pathogens it is notknown the effects and possible mortalitiescaused by this cryptosporidiosis. ReferencesAlvarez-Pellitero P & Sitja-Bobadilla A (2002).Cryptosporidium molnari n. sp (Apicomplexa :Cryptosporidiidae) infecting two marine fishspecies, Sparus aurata L. and Dicentrarchuslabrax L. International Journal for Parasitology32, 1007-1021. Alvarez-Pellitero P, Quiroga MI, Sitja-Bobadilla A, Redondo MJ, Palenzuela O,Padros F, Vazquez S & Nieto JM (2004).Cryptosporidium scophthalmi n. sp(Apicomplexa : Cryptosporidiidae) fromcultured turbot Scophthalmus maximus. Lightand electron microscope description andhistopathological study. Diseases of AquaticOrganisms 62, 133-145. Lom J & Dykova I (1992). “Protozoanparasites of fishes” Elsevier, Amsterdam. Ramos MF, Costa AR, Barandela T, Saraiva A& Rodrigues PN (2007). Scuticociliateinfection and pathology in cultured turbotScophthalmus maximus from the north ofPortugal. Diseases of Aquatic Organisms 74, 249-253. Sitja-Bobadilla A, Pujalte MJ, Macian MC,Pascual J, Alvarez-Pellitero P & Garay E(2006). Interactions between bacteria andCryptosporidium molnari in gilthead seabrearn (Sparus aurata) of under farm andlaboratory conditions. Veterinary Parasitology142, 248-259.at the base of the fins, exophthalmia, withdistension of the abdominal cavity andascites. Examined fishes were infected byseveral pathogens. Gram-positive cocciidentify as Streptococus parauberis weredetected in several organs. The detailedidentification and the pathology caused bythis bacterium will be reported elsewhere.Scuticociliate belonging to the Philasteridawere detected in skin, gills and ascitic fluid(see Ramos et al., 2007). Histological sectionsof the liver showed the occurrence of ovoidoocysts in different stages of sporulationinside a parasitophorous vacuole in bile ductsepithelial cells (Figure 1). Mature oocysts, 4to 8 μm long, contained four naked elongatesporozoites (Figure 2). In some casessporozoites were released from the oocysts(Figure 3). Other developing stages were notobserved. Levels of infection were notdetermined as this parasite was observedaccidentally. Fibrosis surrounding the infectedbile ducts and necrosis of adjacent hepaticcells were observed (Figure 1). Epithelial bileducts host cells seem not to be seriouslydamaged. The absence of sporocysts and the presenceof 4 sporozoites in the oocysts are features ofthe genus Cryptosporidium Tyzzer, 1907.According to Lom & Dyková (1992) the siteof infection of piscine coccidia is very oftenextra-intestinal. As far as we know this is thefirst report of a piscine Cryptosporidiumdetected in bile ducts. Recently Alvarez-Pellitero et al. (2004) described C. scophthalmifrom intestine and stomach of turbot fromdifferent farms on NW Spain coast. Theoocysts of the Coccidia detected in this studywere bigger than that ones described by these