Dabieshan tick virus (DTV) is a novel tick-borne virus with the potential to infect both animals and humans. It has been confirmed that DTV is widely distributed in Shandong and Zhejiang Provinces. In this study, a total of 389 ticks were sampled from Honghe city of Yunnan Province and Bijie city of Guizhou Province, and then divided into 148 pools according to the location and species. QRT-PCR and nested PCR were performed to confirm the presence of DTV. The results showed a minimum infection rate of 2.43% (5/206) in Yunnan Province and 3.28% (6/183) in Guizhou Province, respectively. Interestingly, DTV was identified in Rhipicephalusmicroplus for the first time besides Haemaphysalis longicornis. Phylogenetic analysis showed that DTV from Yunnan and Guizhou Provinces shared over 94% identity with isolates derived from Hubei and Shandong Provinces, and DTV was relatively conservative in evolutionary dynamics. These findings provide molecular evidence of Dabieshan tick virus in different species of ticks from unrecognized endemic regions and suggest that DTV may be widely prevalent in southwestern China.

BackgroundIn 2017–2018, a new highly pathogenic H5N6 avian influenza virus (AIV) variant appeared in poultry and wild birds in Asian and European countries and caused multiple outbreaks. These variant strains are different from the H5N6 virus associated with human infection in previous years, and their genetic taxonomic status and antigenicity have changed. Therefore, revision of the primers and probes of fluorescent RT-PCR is important to detect the new H5N6 subtype AIV in poultry and reduce the risk of an epidemic in birds or humans.MethodsIn this study, the primers and probes including three groups of HA and four groups of NA for H5N6 influenza virus were evaluated. Then a set of ideal primer and probes were selected to further optimize the reaction system and established a method of double rRT-PCR assay. The specificity of this method was determined by using H1~H16 subtype AIV.ResultsThe results showed that fluorescence signals were obtained for H5 virus in FAM channel and N6 virus in VIC channel, and no fluorescent signal was observed in other subtypes of avian influenza viruses. The detection limit of this assay was 69 copies for H5 and 83 copies for N6 gene. And, the variability tests of intra- and inter-assay showed excellent reproducibility. Moreover, this assay showed 100% agreement with virus isolation method in detecting samples from poultry.ConclusionThe duplex rRT-PCR assay presented here has high specificity, sensitivity and reproducibility, and can be used for laboratory surveillance and rapid diagnosis of newly emerged H5N6 subtype avian influenza viruses.