Three alpacas (Vicugna pacos) aged two to 22 months with a history of illthrift and diarrhoea were examined postmortem, and tissues were collected for histology, including immunohistochemical labelling for pestivirus antigen, virus isolation and TaqMan reverse transcriptase-pcr assay. Blood samples from two clinical cases and the remaining herd members were tested for bovine viral diarrhoea virus (bvdv) antibody by serum neutralisation, antigen detection and pcr assay. The three affected alpacas were positive for bvdv by pcr of splenic tissue and/or heparinised blood. Non-cytopathic bvdv was isolated from several tissues and plasma of two of the alpacas. dna sequencing and phylogenetic analysis of the viral genome from the pcr product showed that the bvdv was of subgenotype 1b. Immunohistochemical examination of brain tissue was positive in two cases, consistent with a persistent infection. bvdv antibodies were detected in 16 of 25 clinically unaffected alpacas. There was no evidence of persistent infection in the in-contact animals. The source of the infection was not determined.
To compare the real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR) assays used for the diagnosis of Porcine reproductive and respiratory syndrome virus (PRRSV), a Europe-wide interlaboratory ring trial was conducted. A variety of PRRSV strains including North American (NA) and European (EU) genotype isolates were analyzed by the participants. Great differences regarding qualitative diagnostics as well as analytical sensitivity were observed between the individual RT-qPCR systems, especially when investigating strains from the EU genotype. None of the assays or commercial kits used in the ring trial could identify all different PRRSV strains with an optimal analytical and diagnostic sensitivity. The genetic variability of the PRRSV strains, which is supposed to hinder the diagnostic of the RT-PCR because of mutations at the primer binding sites, was also confirmed by sequencing and subsequent phylogenetic analysis. In summary, a major problem in PRRSV diagnostics by RT-qPCR is false-negative results. To achieve maximum safety in the molecular diagnosis of PRRSV, the combined usage of different assays or kits is highly recommended.
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