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Cryptosporidium molnari n. sp. is described from two teleost fish, the gilthead sea bream (Sparus aurata L.) and the European sea bass (Dicentrarchus labrax L.). The parasite was found mainly in the stomach epithelium and seldom in the intestine. Oocysts were almost spherical, with four naked sporozoites and a prominent residuum, and measured 3.23-5.45 × 3.02-5.04 (mean 4.72 × 4.47) μm in the type host, gilthead sea bream (shape index 1-1.17, mean 1.05). Sporulation was endogenous, as fully sporulated oocysts were found within the fish, both in the stomach epithelium and lumen, and in faeces. Oocysts and other stages of C. molnari fit most of the diagnostic features of the genus Cryptosporidium, but differ from hitherto described species, including piscine ones. All stages were located within a host contributed parasitophorous vacuole lined by a double host microvillar membrane. Merogonial and gamogonial stages appeared in the typical extracytoplasmic position, whereas oogonial and sporogonial stages were located deeply within the epithelium. Ultrastructural features, including the characteristic contact zone of the parasite with the host epithelial surface, were mostly coincident with those of other Cryptosporidium spp. Mitochondria were found in dividing meronts, merozoites, microgamonts and sporozoites. Pathological effects were more evident in gilthead sea bream, which also exhibited a clearly higher prevalence (24.4 versus 4.64% in sea bass). External clinical signs, consisting of whitish faeces, abdominal swelling and ascites, were rarely observed, in contrast with important histopathological damage. The wide zones of epithelium invaded by oogonial and sporogonial stages appeared necrotic, with abundant cell debris, and sloughing of epithelial cells, which detached to the lumen. No inflammation reaction was observed and the cellular reaction was limited to the cells involved in the engulfing of intraepithelial stages and debris, probably macrophages., This work was supported by research grants from the Spanish Ministerio de Educación, Cultura y Deporte MAR-98/1000 AGF95-0058, and FEDER Program 1FD97-0979. We are thankful to technicians from the Electron Microscopy Services of the University of Barcelona and Valencia and J. Monfort from IATS for histological processing. The authors want to thank Dr L. Ortega from the Departamento Sanidad Animal, Facultad de Veterinaria, Universidad Complutense de Madrid for his contribution to this work.

The possible interaction of Cryptosporidium molnari and bacteria in gilthead sea bream (Sparus aurata) was studied. Epidemiological data from a pathological survey under farm conditions were analyzed. In addition, parasite and bacteria burdens were studied in experimental models in which naturally and experimentally parasitized fish were challenged with a particular strain of Vibrio harveyi (H57). All the bacteria species present were studied. Under farm conditions, the parasite was more prevalent when mortality or morbidity cases (study C) occurred than in randomly sampled fish (study B). In study C, parasite abundance was significantly higher in bacteria-negative fish, and total bacteria abundance was significantly higher within non-parasitized fish. V. harveyi and V. splendidus were the most prevalent among bacteria carriers in studies B and C, respectively. In study C, among bacteria carriers, most isolates were slightly more prevalent in parasitized than in non-parasitized fish. Two groups (G1, G2) of naturally parasitized fish were inoculated with H57 by intracoelomic injection (ICI) and by oral intubation (OI). H57 was recovered only in G1 inoculated fish, which had a significantly higher basal abundance of total bacteria, and where the only ones with mortalities. In G1, the mortality rate and the prevalence of other V. harveyi strains different from the H57 molecular type were higher in ICI than in OI fish, and the total bacteria abundance was also significantly higher in ICI fish. C. molnari abundance was significantly higher in G1 than in G2, and also in OI than in ICI fish within G1. When H57 was IC inoculated to fish (G3, from the same farm as G2) experimentally infected with C. molnari, H57 was not recovered from any fish. A low mortality was recorded, and only in those fish inoculated with both pathogens. Also in these fish, the prevalence of infection of C. molnari was higher and histopathological damage to the stomach was greater than in fish inoculated only with the parasite. Therefore, the impact of the parasite would be reduced notably when the bacterial burden or the intensity of parasite infection are low (G2, G3)., Funding for this study was provided by the Spanish Ministerio de Ciencia y Tecnología (project no. AGL-2002-0475-C02). We thank two anonymous reviewers for their valuable comments on the manuscript. The authors are also grateful to M. Alonso and P. Cabrera for excellent assistance in experimental infections, to J. Monfort for histological processing, and to Tinamenor S.A. for providing parasite-free fish.