Molecular detection and genome analysis of circoviruses of European eel (Anguilla anguilla) and Sichel (Pelecus cultratus) from lake Balaton, Hungary

Balatoni angolnákból és gardából kimutatott cirkovírusok genomiális szintű összehasonlító vizsgálat

Molecular detection and genome analysis of circoviruses of European eel (Anguilla anguilla) from Lake Balaton

Circoviruses are small, non-enveloped viruses with a circular single-stranded (ss) DNA genome, which ranges from about 1.3-2.3 kb in size. The genome contains at least two open reading frames (ORF) arranged on different strands, these encode the replication (Rep) and capsid (Cap) proteins. Before the last decade, circoviruses were known only from birds and swine, but recently, due to the different metagenomic methods and PCR based diagnostic tests, numerous circovirus-related sequences were described from environmental samples, invertebrates and also from lower vertebrates (amphibian and fish). According to the increasing number of the detected piscine circoviruses, a monitoring program (Bioclimate) is carried out for screening circoviruses in Hungarian freshwater fishes. Samples were taken regularly also from Lake Balaton. In March 2014, fifteen living European eels (Anguilla anguilla) and two razorfish (Pelecus cultratus) were caught among other species at the Sió-floodgate sampling site. After euthanasia, tissue samples were collected from the gills, liver, spleen, kidney and intestine. The presence of circoviral DNA was detected by a broad-spectrum nested PCR, targeting the Rep gene. In case of positive results, rolling circle amplification and inverse nested PCR reactions were carried out to amplify the remaining part of the circular genomes. Six out of the fifteen eels, and one razorfish sample were found to be positive for circoviral DNA. Analysis of these fragments revealed the presence of three different circovirus-related rep-like sequences. One of them (representing with 3 samples) was identical with the previously described eel circovirus (EeCV) by Doszpoly et al. (2014). The whole genome amplification of these 3 samples was also successful. Comparing the partial rep-like sequences, two other samples were similar with 96% nucleotide identity to the above mentioned ones, and were identical to the rep-like sequence originating from razorfish. The 6th circovirus positive sample showed only 50% similarity to the EeCV, but it was found to be identical to the rep-like sequences of roaches (Rutilus rutilus) caught from Lake Balaton, recently. Circoviruses were described as host-specific or narrow host range microorganisms. To the best of our knowledge, this is the first report about the detection of the same circovirus related sequences in various, distantly related fishes and the occurrence of different rep-like sequences in the same fish species

Full genome sequence of a novel circo-like virus detected in an adult European eel Anguilla anguilla showing signs of cauliflower disease.

An adult European eel Anguilla anguilla, showing typical signs of the so-called cauliflower disease, was subjected to pathological and molecular virological examinations. Samples taken from internal organs and the polypoid proliferative tissue from the mouth were examined by PCR for the detection of several viruses. Positive results were obtained with a nested PCR targeting the rep gene of circoviruses. Analysis of the partial rep sequence indicated the presence of a putative novel circovirus, but attempts to isolate it remained unsuccessful. The missing part of the genome was acquired by an inverse nested PCR with 2 specific primer pairs, designed from the newly determined rep sequence, followed by genome walking. The circular full genome was found to consist of 1378 nt (GenBank accession no. KC469701). Two oppositely oriented open reading frames (ORFs) were present, of which one was unambiguously identified as a circoviral rep gene. However, the predicted product of the other ORF, though it is a clear positional counterpart of the cap genes, showed no obvious homology to any known circoviral capsid proteins. A stem-loop-like element in the intergenic region between the 5' ends of the ORFs was also found. Phylogenetic calculations indicated that the novel virus belongs to the genus Circovirus in the family Circoviridae. The relative amount of the viral DNA in the organ samples was estimated by quantitative real-time PCR. The results suggested that the examined fish was caught in an active viremic state, although the role of this circovirus in the etiology of the cauliflower diseases could not be ascertained

Balatoni angolnákból (Anguilla anguilla) és gardából (Pelecus cultratus) kimutatott circovírusok molekuláris elemzése

Balatoni angolnákban és gardában kimutatott cirkovírusok részleges és teljes genom alapú összehasonlító vizsgálat

Molecular detection and genome analysis of circoviruses of European eel (Anguilla anguilla) and sichel (Pelecus cultratus)

The prevalence and distribution of piscine circoviruses (CVs) were tested in a routine virus monitoring programme in Lake Balaton, Hungary. A high prevalence of European eel CV (EeCV) was found in the apparently healthy eel population (35.5%). The copy number of the viral DNA in different organs was determined by quantitative real-time PCR. The results suggested that some eel specimens were in active viraemic status despite their asymptomatic condition. Furthermore, a novel, previously undescribed CV was also detected in eel and sichel samples. Full genome characterisation confirmed that the virus represents a novel EeCV species (EeCV-2). The genome contains an integrated eel chromosome-derived fragment, suggesting that the original host of the virus was the eel and it probably emerged subsequently in the sichel by host switching. In some samples, an additional, 1,111-nt-long circular ssDNA was also observed involving a CV-like stem-loop structure and an ORF showing homology to CV capsid protein genes, without any sign of a replication initiator protein sequence

Atlantic salmon papillomatosis in Russia and molecular characterization of the associated herpesvirus.

Papillomatosis of Atlantic salmon Salmo salar has been reported for decades in Russia, Scandinavia and Scotland. The disease is typically benign although heavy losses have occasionally been reported. A herpesviral etiology has been suggested based on ultrastructural evidence; however, the virus has not been isolated or genetically characterized. In this study, we provide the first viral sequences detected in the papillomas from diseased Russian Atlantic salmon. Phylogenetic analyses, based on the partial sequences of the herpesviral polymerase and terminase genes, supported the virus as a novel member of the genus Salmonivirus within the family Alloherpesviridae. The sequences of the Atlantic salmon papillomatosis virus differ markedly from those of the 3 known salmoniviruses; therefore, the authors propose the species designation Salmonid herpesvirus 4 to be considered for approval by the International Committee on Taxonomy of Viruses