Isolation and partial characterization of a novel porcine astrovirus.

Astroviral infection has been described as one of the causes of porcine diarrhoeal disease. Here we describe the detection of astrovirus-like particles by electron microscopy in a diarrhoeal specimen. Furthermore, a cytopathic virus was isolated and propagated in an established porcine kidney cell line, PK-15. Reverse transcription and PCR performed with astrovirus-specific primers amplified a product with the expected size. Sequencing of the PCR product revealed that the virus observed by electron microscopy and propagated in the porcine cell line is an astrovirus, showing 86% identity at the nucleotide level with the only known porcine astrovirus, PAstV. Phylogenetic analysis clustered the novel isolate, Sb4685, together with PAstV in a broad clade comprising mammalian astroviruses.

Comparing electron microscopy and a competitive blocking ELISA in the detection of rotaviruses in porcine faeces.

Analysis of 476 faecal samples from diarrhoeic piglets was performed using electron microscopy (EM) and a competitive blocking (CB)-ELISA based on monoclonal antibodies to the VP6 protein of group A rotavirus. Rotavirus was detected by EM and/or CB-ELISA in 111 (23.3%) samples. Of these, all groups of rotavirus were identified in 83 (74.8%) samples by EM (EM+), while group A rotavirus was identified in 90 (81.1%) samples by CB-ELISA (ELISA+). However, only 62 (55.9%) of samples were positive using both detection methods. The finding of 28 (25.2%) EM-/CB-ELISA+ samples illustrated the high sensitivity of the CB-ELISA method. On PCR analysis, groups B and C rotavirus was found in 3 and 16 of 19 EM+/CB-ELISA- samples, respectively. Although the study illustrates the high sensitivity of a CB-ELISA in rotavirus detection, the findings highlight the need to use a range of diagnostic methods in detecting these viruses in clinical samples.

Avipoxvirus in blackcaps (Sylvia atricapilla).

From July to September 2005, 1075 wild birds of 37 species were mist-netted at a location in the north-eastern part of the Czech Republic. The birds were examined for the presence of avipoxvirus lesions. This was demonstrated by electron microscopy in skin lesions in nine of 244 blackcaps (Sylvia atricapilla) examined (4% prevalence). Blackcaps skin bioptates were processed using the ultrathin section method. In skin bioptates, avipoxviruses were demonstrated in intracytoplasmic inclusions where, in addition to mature viruses, lipids and filamentous structures concentrated into large circular formations were found. The so-called additional inclusions were also found. These did not contain any virus components, and they served as the precursor of A-type intracytoplasmic inclusions. Blackcap avipoxvirus was isolated by passage on the chorioallantoic membrane of 9-day-old chicken embryos. The virus was successfully adapted after 11 passages (each passage lasted 5 to 7 days), at which time a marked changes in the form of tiny nodules 2 to 3 mm in diameter were observed on the chorioallantoic membrane. Further identification of field isolates and of the cultured virus was carried out using polymerase chain reaction and sequencing. Sequences were compared with consensus sequences of both canarypoxviruses and fowlpoxviruses. Our sequence was found to be 98.8% identical to the canarypox consensus sequence, but only 63% identical to the fowlpox consensus sequence. Our avipoxvirus sequence was proven to be significantly more closely related to canarypoxviruses than to fowlpoxviruses also by phylogenetic analysis.