Nairobi sheep disease virus RNA in ixodid ticks, China, 2013.

Shangshu Gong,Feng Wei,Limin Shang,Zedong Wang,Quan Liu,Biao He,Changchun Tu

doi:10.3201/eid2104.141602

Abstract

Nairobi sheep disease virus RNA in ixodid ticks, China, 2013.

Highlights

We describe a newly discovered Nairobi sheep disease virus (NSDV), named NSDV (China), identified by viral metagenomic analysis of ticks collected from the northeast region of the People’s Republic of China (Liaoning, Jilin, and Heilongjiang provinces) during May–July, 2013, and divided into 9 groups according to tick species and sampling sites
The isolate we identified, NSDV (China), is genetically divergent from the NSDVs of South Asia and Africa and is a novel strain, with H. longicornis likely the main vector
NSDV-positive samples were ground with cold serum-free minimum essential medium (MEM; Sigma-Aldrich) and centrifuged at 12,000 × g for 10 min at 4°C

Summary

Technical Appendix

Tick Collection, RNA extraction and Processing, Solexa Sequencing, and Analysis of Data. The viral RNAs and DNAs were extracted immediately using TRIzol (TaKaRa, Dalian, China) according to the manufacturer’s protocol. Total viral nucleic acids were dissolved in RNase-free H2O (TaKaRa) and used immediately for the following reverse transcription with SuperScript III reverse transcription (Invitrogen, Carlsbad, CA) using random primers according to the manufacturer’s protocol. To obtain sufficient viral nucleic acid, SISPA was employed to amplify the dscDNA with the Accuprime Taq DNA Polymerase System (Invitrogen) according to the manufacturer’s protocol. The purified PCR products of the 9 groups were pooled together and subjected to Solexa sequencing in one lane by the Beijing Genome Institute (BGI, Shenzhen, China)

Virus Isolation Procedure

Ixodes persulcatus

Findings

Contig Location