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A small-scale piglet-producing farm in Styria, in southeastern Austria, reported major piglet losses from CT in January 2015. Animals from this farm were examined and samples were taken. The CT-affected piglets showed a severe lateral shaking and were often incapable of sucking milk, which led to elevated preweaning death rate. The prevalence of CT varied from 20% to 100% within the affected litters. Pathological examinations were performed confirming the presence of typical CT A-II lesions.
We tested various samples from the farm using an atypical porcine pestiviruses (APPV)-specific TaqMan probe-based reverse transcription PCR (RT-PCR) but obtained negative results. However, we obtained an amplicon of appropriate length from CT-piglet serum samples by using a novel panpestivirus RT-PCR (PPF 5′-GTKATHCAATACCCTGARGC-3′ and PPR 5′-GGRTTCCAGGARTACATCA-3′), which enables detection of CSFV, BVDV-1, BVDV-2, BDV, BV, and APPV. Surprisingly, sequencing of this RT-PCR product yielded an unknown sequence with a noninterrupted open reading frame. An initial BLAST search resulted in “no significant similarity found,” but the translated sequence of 270 aa aligned well with different pestiviruses. The viral RNA was detected in all samples (serum, tonsils, lung, liver, spleen, and central nervous system material) of CT-affected piglets and their littermates from the farm.
After inoculating SK-6 cells with serum samples of affected piglets, we detected the amplification of this unknown pestivirus using RT-PCR. We provisionally termed the agent “Linda” (lateral-shaking inducing neurodegenerative agent) to avoid confusion with other pestiviruses. We determined the full genome of Linda virus (LV) and determined the length of the LV genome. We performed a phylogenetic analysis of LV demonstrating LV’s divergence from other pestiviruses. We found the identity between LV, approved pestiviruses, and APPV to be only 60%, but we found an identity of 68% between LV and BV. Comparison of the pestiviral polyprotein sequences yielded an amino acid identity of 69% between LV and BV and of <54% with all other pestiviruses.
Conclusions
A previously unknown pestiviral agent was found in CT-affected piglets on a farm in Austria. We observed a severe hypomyelination in the entire white matter of the spinal cord and detected pestivirus antigen in the brain of the CT-affected piglets, suggesting a causal relationship between infection and lesions. Analyses of the assembled genome allowed an unambiguous assignment of LV within the genus Pestivirus with regard to the presence of pestivirus-specific genes (Npro and Erns) and sequence homology to other pestiviruses. In contrast to atypical porcine pestiviruses (APPV), which hardly infects cultured cells at all, LV could be easily propagated on porcine cell lines without the need for adaptation, similar to what has been reported for BV. We suggest that LV likely shares a common ancestor with BV.
After its description ≈10 years ago, BV was intensively sought worldwide, but the virus has never been detected outside Australia. The broad cell culture tropism of BV led to the hypothesis that the virus recently jumped from another species to porcine hosts. The discovery of a related pestivirus with substantial sequence divergence in swine on a different continent suggests that both viruses probably have a porcine origin. The identification of the cross-reacting E2-specific monoclonal antibody 6A5 indicates that a cross-reactivity to related pestiviral proteins exists, which might interfere with the serologic testing for CSFV. Further work is needed to investigate the prevalence and the epidemiology of LV in Europe and to assess its virulence in controlled animal experiments.
Lamp B, Schwarz L, Högler S, Riedel C, Sinn L, Rebel-Bauder B, et al. Novel Pestivirus Species in Pigs, Austria, 2015. Emerg Infect Dis. 2017;23(7):1176-1179. https://dx.doi.org/10.3201/eid2307.170163