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Six myxosporidian species were found in chub (Leuciscus cephalus) originating from Lower Austrian rivers. The frequency of the parasites and their localization was recorded. In all chub, independent of size and origin, Myxobolus cyprini occurred predominantly in the macrophage centres (MCs) of the haematopoietic organs, spleen and kidney. Exclusively in the head kidney of young fish not yet described vermicular plasmodia containing spores of M. cyprini were found. In muscle tissue the prevalence of M. cyprini was comparatively low. Other species of Myxobolus characterized by plasmodial cysts frequently occurred in gills and swimbladder but were rarely detected, and only in small numbers, in the haematopoietic organs. The number of M. cyprini spores and the relative volume of MCs in the haematopoietic organs were estimated in order to examine possible correlations. Significant interrelated changes were found only in juvenile fish up to a size of 15 cm. In bigger fish, the number and size of macrophage aggregates were highly variable and independent of infection intensity and fish size, but the number of spores never exceeded that of the aggregated macrophages. The data suggest that due to an early date of infection M. cyprini is the only species which is closely associated with macrophage aggregation.

Summary Between April and November 2003, parasitological examinations of the nase Chondrostoma nasus L. and the chub Leuciscus cephalus L. from the neighbouring Melk and Pielach rivers in Lower Austria were conducted. Amongst various gill parasites, Lamproglena pulchella Nordmann 1832 was detected on both fish species, which was the first record of this parasitic crustacean in Austria. Physico‐chemical examinations of water samples of the two rivers were carried out during the same period. The results indicate that general water parameters in the Melk were subjected to more vigorous changes than in the Pielach. Critical temperature levels and ammonia concentration as well as drastic changes in the ionic composition occurred more frequently in the Melk River. The observed distribution of L. pulchella indicates its sensitivity to such stress factors: there was no evidence of the parasite in the Melk River until late November and it only then occurred on the gills of L. cephalus with a prevalence of 20% and a mean intensity of 2. In the Pielach River, infestation on chub had already occurred in June with a prevalence of 40% and a mean intensity of 3, rising to 60% and 7 in November; 45% of the nase was also infested in November at a mean intensity of 3.

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 TCID 50 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. KEY WORDS: SVCV · AuNPs · Colorimetric detection · Diagnosis Resale or republication not permitted without written consent of the publisherDis Aquat Org 100: [3][4][5][6][7][8][9][10] 2012 rescence, immunoperoxidase, and ELISA (Amos 1985, OIE 2009). Rapid ad vances in molecular biology techniques have led to the development of several molecular methods for SVCV detection, including hybridization assays, RT-PCR, nested RT-PCR (nRT-PCR) and real-time PCR (Oreshkova et al. 1999, Koutná et al. 2003, Yue et al. 2008. Although these assays are specific and sen sitive, they are time consuming and require sophis ticated apparatus and complex post-run manipu lations.Recently, noble metal nanoparticles, particularly gold nanoparticles (AuNPs), have been introduced as a promising approach to the development of the next generation of diagnostic assays (Mirkin et al. 1996, Storhoff et al. 2000. AuNPs have become an important alternative as imaging agents due to their noncytotoxicity, facile immunotargeting and nonsusceptibility to photobleaching or chemical/thermal de naturation, a problem commonly associated with dyes (Jain et al. 2006). Advances in functionalizing particles with oligonucleotides and tailoring their surface properties have paved the way to design a series of new and practical systems for nucleic acid detection (Mirkin et al. 1996, Elghanian et al. 1997, Storhoff et al. 2000, Daniel & Astru...

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 noninflammatory 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 welldefined equilibrium controlled by the host fish.

Using a PCR that amplifies a region of the thymidine kinase (TK) gene, an epidemic spread of koi herpesvirus (KHV) was determined in koi carps in Austria in 2007. A total of 15 virus samples from different locations in Austria were analyzed to determine their genetic relatedness following PCR and nucleic acid sequencing of the open reading frame 40 (ORF40) region of the KHV genome. ORF40-specific PCR amplification products that were obtained from tissue samples shared 100% nucleotide sequence identity with the published sequence of the Japanese strain of KHV. The ORF40 sequence of one isolate from the UK that was included in the present study was 100% identical with the published sequence of an Israeli strain of KHV. This is the first study that used a larger number of samples and a PCR method, which allowed distinguishing all 3 strains of KHV. The present investigation provides information on the epidemiology of KHV infections in Europe and describes a useful molecular tool for epidemiological studies. KEY WORDS: Cyprinus carpio koi · Koi herpesvirus · Sequence analysis · ORF40Resale or republication not permitted without written consent of the publisher Dis Aquat Org 88: [267][268][269][270] 2010 of 156 presumably protein-encoding open reading frames (ORFs). In each strain, several genes were fragmented by frameshifts which are likely to render the encoded proteins nonfunctional. Most of the affected genes encode predicted membrane glycoproteins and at least some of them occurred in vivo, suggesting a role of gene loss in the emergence of KHV disease (Aoki et al. 2007). The 3 available full-length genomes were aligned and inspected visually for differences among the strains. Since the published sequences of the J, U and I isolates differ in ORF40, we decided to use this region to compare samples collected in 2007. Moreover, ORF40 encodes a predicted membrane glycoprotein and is broken at least twice in the same place in J, U and I, once more in J, and frameshifted once more in U (Aoki et al. 2007). In general, membrane proteins are relevant both for virus-host cell interactions and for the host immune response, and complete sequence analysis revealed 27 predicted membrane proteins. However, none of them was identified and functionally characterized, and it is still not known which of them are major components of the virion envelope. Until now, only ORF81 was shown to encode one of the membrane proteins of KHV (Rosenkranz et al. 2008). MATERIALS AND METHODSIn the present study, we investigated the nucleotide sequences of the ORF40 region of 15 KHV samples from Austria and 1 isolate from the UK. In order to determine their genetic relatedness, we compared their ORF40 nucleotide sequences with those of completely sequenced KHV isolates from Japan, Israel and the USA.Some characteristics of the samples and the infected fish used in the study are listed in Table 1. Most samples have been prepared from single fish mortalities. In cases where organs from more than one fish were pooled, all fish displaye...

Infectious hematopoietic necrosis virus (IHNV) is

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.