Effects of siRNA silencing on the susceptibility of the fish cell line CHSE-214 to Yersinia ruckeri.

Yersinia ruckeri is a facultative intracellular enterobacterium mostly known as the causative agent of enteric redmouth disease in salmonid fish. In the present study, we applied RNA inhibition to silence twenty pre-selected genes on the genome of a fish cell line (CHSE-214) followed by a gentamicin assay to quantify the effect of silencing on the cells' susceptibility to infection and found that silencing of 18 out of 20 genes significantly reduced the number of Y. ruckeri recovered. These findings improve our understanding of the infection process by Y. ruckeri and of the interactions between this bacterial pathogen and host cells.

Isolation and characterization of a novel reovirus from white bream Blicca bjoerkna.

During a fish health inspection in the Viennese waterway 'Old Danube', a virus was isolated exclusively from white bream Blicca bjoerkna (L.) (formerly Abramis bjoerkna L.), one of the most abundant cyprinids present and not known as a host species for this virus. The virus preferentially replicated in cultures of the epithelioma papulosum cyprini cell line where focal plaques of infection developed slowly. Examination of infected cell cultures by electron microscopy revealed non-enveloped 60 to 70 nm icosahedral virions that had characteristic multiple segregated protrusions of their outer capsid. A partial RNA-dependent RNA polymerase gene sequence was obtained and a BLAST search indicated 76% identity to golden shiner reovirus and grass carp reovirus. These results suggested that the virus belonged to the genus Aquareovirus (Family Reoviridae). Phylogenetic analysis placed the isolated virus within a clade of the species Aquareovirus C species. Accordingly, the virus was tentatively designated as white bream reovirus (WBRV) strain A-127/06 within the species Aquareovirus C.

Direct detection of unamplified spring viraemia of carp virus RNA using unmodified gold nanoparticles.

Spring viraemia of carp (SVC) is a viral disease that mainly affects carp Cyprinus carpio and other cyprinid fish, causing severe economic losses. Rapid detection and identification of spring viraemia of carp virus (SVCV) is crucial for effective disease management. Recent advances in nanoscience are having a significant impact on many scientific fields, especially biodiagnostics, where a number of nanoparticle-based assays have been introduced for biomolecular detection. Single- and double-stranded oligonucleotides can be adsorbed on gold nanoparticles (AuNPs) in colloidal solution under certain conditions. We exploited this phenomenon to develop a specific hybridization assay for direct detection of SVCV-RNA without prior amplification. The result of the hybridization process could be detected visually within 1 min when the colour of the reaction mixture changed from red to blue (positive reaction) or remains red (negative). The lower detection limit of the assay was estimated to be 10-3 TCID50 ml-1 SVCV-RNA, and it has the feasibility to detect the target virus-RNA in clinical specimens without previous amplification. In order to obtain an indication of the assay's performance on clinical samples we compared the optimized assay with nested RT-PCR in detection of SVCV-RNA in infected fish samples. The concordance of the 2 methods was defined as 100% when compared to nested RT-PCR positive and negative samples. The SVC-AuNPs assay requires only 15 min, eliminates the need for thermal cycling or detection instruments and is a specific and rapid tool for detection of SVCV-RNA directly from clinical samples.

"Mid-G" region sequences of the glycoprotein gene of Austrian infectious hematopoietic necrosis virus isolates form two lineages within European isolates and are distinct from American and Asian lineages.

Infectious hematopoietic necrosis virus (IHNV) is one of the most important pathogens of salmonid fish. In this study a comprehensive phylogenetic analysis of the genetic evolution and variety of Austrian IHNV strains, as well as selected strains ensuring worldwide coverage, is presented. The phylogenetic investigation is based on sequences comprising the "mid-G" region of the G gene, and it includes all currently available IHNV sequences of the G gene with a length of at least 615 bp. Austrian IHNVs are located--together with other European IHNV isolates--in two clusters of genogroup M (M-Eur1 and M-Eur2) and are clearly separated from American and Asian lineages. The genetic clustering, however, could not be linked to certain clinical symptoms or significant differences in the mortality rates.

Myxobolus cycloides on the swimbladder of chub Leuciscus cephalus: a controlled, host-specific localisation.

Of 150 wild stock chub, Leuciscus cephalus L. captured in Lower Austrian watercourses, 112 revealed disc like plasmodia of Myxobolus cycloides Gurley, 1893 on the caudal chamber of the swim bladder. Other cyprinid species from the same waters lacked M. cycloides or other myxosporeans in this specific localisation. In chub, the intensity of infection (number of discs on the swim bladder) showed a logarithmic, age-dependent increase. The plasmodia of M. cycloides were situated in the connective tissue--mainly along blood vessels--and exhibited a delicate envelope of host tissue, thus forming a characteristic myxosporean cyst. Occasionally single trophozoites seemed to merge. A general process of fibroblast proliferation leading to encapsulation and degradation of the parasite was observed. This process was initiated by the formation of small multiple encapsulations within the spore containing trophozoid, before thickening of the outer cyst wall occurred. The general non-inflammatory course of the M. cycloides infection, and the obvious good health of the investigated chub suggest that this myxosporean in its host specific localisation cannot be regarded as a serious pathogen--on the contrary: parasite multiplication and degradation seemed to occur in a well-defined equilibrium controlled by the host fish.