Biodiversity and evolution of Imjin virus and Thottapalayam virus in Crocidurinae shrews in Zhejiang Province, China.

The recent discovery of numerous hantaviruses in insectivores has provided a new view of hantavirus biodiversity and evolution. To determine the presence and genetic diversity of Imjin virus (MJNV) and Thottapalayam virus (TPMV) in insectivores in Zhejiang Province, China, we captured and performed virus screening of 32 Ussuri white-toothed shrews (Crocidura lasiura) and 105 Asian house shrews (Suncus murinus) in different coastal regions. Hantavirus genome (S, M, and L segments) sequences were successfully recovered from one Ussuri white-toothed shrew and seven Asian house shrews. Phylogenetic analysis revealed that the virus carried by the Ussuri white-toothed shrew was most closely related to MJNV, but with >15% nucleotide sequence difference, suggesting that it represents a new subtype. The hantaviruses carried by Asian house shrews were closely related to the TPMV variants found in the same geographic area, but more distantly related to those sampled in India and Nepal. Additionally, the TPMV sequences obtained in this study, as well as those found previously in this area, could be divided into three lineages reflecting their geographic origins, indicative of largely allopatric evolution. Overall, our data highlights the high genetic diversity of insectivore-borne hantaviruses in China, suggesting that more may be discovered in the future.

Molecular survey of hard ticks in endemic areas of tick-borne diseases in China.

Over the past several years, there was a substantial increase in the number of cases of known and novel tick-borne infections in humans in China. To better understand the ticks associated with these infections, we collected hard ticks from animals or around livestock shelters in 29 localities in 5 provinces (Beijing, Henan, Hubei, Inner Mongolia, and Zhejiang) where cases of tick-borne illness were reported. We collected 2950 hard ticks representing 7 species of 4 genera (Dermacentor sinicus, Haemaphysalis flava, Haemaphysalis longicornis, Ixodes granulatus, Ixodes persulcatus, Rhipicephalus microplus, and Rhipicephalus sanguineus). These ticks were identified to species using morphological characters initially. We then sequenced the mitochondrial small subunit rRNA (12S rRNA) gene, cytochrome oxidase subunit 1 (COI) gene, and the second internal transcribed spacer (ITS2) gene of these ticks, and conducted phylogenetic analyses. Our analyses showed that the molecular and morphological data are consistent in the identification of the 7 tick species. Furthermore, all these 7 tick species from China were genetically closely related to the same species or related species found outside China. Rapid and accurate identification and long-term monitoring of these ticks will be of significance to the prevention and control of tick-borne diseases in China.

[Study on the molecular characteristics of Japanese encephalitis virus living in vector mosquitoes, in Zhejiang province, 2009 - 2010].

OBJECTIVE: To investigate the molecular characteristics of Japanese encephalitis virus (JEV) living in vector mosquitoes, from Zhejiang province. METHODS: A total of 13 620 mosquitoes were collected from the monitoring stations located in Cixi city and Xianju county in Zhejiang province, in July and August, 2009 - 2010. Nucleic acid of JEV from the mosquitoes was monitored by using real-time RT-PCR. The virus strains were isolated with BHK-21 cell line, with E genes of the isolated viruses amplified, sequenced and their phylogeny and homology analyzed. RESULTS: The positive rates of JEV for those mosquitoes collected in the stations of Cixi and Xianju were 17.0% (27/159) and 3.4% (1/29), respectively. Twenty-two JEV strains were isolated, accounted for 15.4% among the 143 batches of mosquitoes collected in 2010. All E genes in the 6 sequenced virus isolates contained 1500 nucleotides encoding 500 amino acids, in which no inserts and deletions were identified. The identity rates of nucleotide and amino acid in E gene were 99.2% - 99.8% and 100.0% among the 6 JEV strains isolated from Zhejiang, 99.1% - 99.3% and 99.2% - 99.8% between the Zhejiang strains in 2009 - 2010 and the Zhejiang strains in 2007 - 2008, respectively, 87.6% - 88.0% and 97.8% between the 6 Zhejiang strains and the vaccine strain SA14-14-2 of JEV, respectively. The phylogeny tree of E gene indicated that the JEV isolates in Zhejiang during 2009 - 2010 was located in the branch of the genotype I. CONCLUSION: Mosquitoes collected from Cixi and Xianju areas carried JEV, with the rate of JEV in Cixi higher than in Xianju. All the Zhejiang isolates in 2009 - 2010 were proven to be the genotype I of JEV.

Hantaviruses in small mammals and humans in the coastal region of Zhejiang Province, China.

To gain more insights into the epidemiology of hantaviruses in the coastal region of Zhejiang Province, China, the morbidity and mortality of hemorrhagic fever with renal syndrome (HFRS) were analyzed in two coastal areas: Cixi (hilly terrain) and Wenzhou (mountainous terrain). More HFRS cases have been reported in Cixi than in Wenzhou. Annual incidence rate of HFRS in Cixi had been on the level of approximately 1.5/100,000 from 1968 (when the first HFRS case was reported) to 2007, with the highest incidence rate of 8.54/100,000 in 1999. The annual incidence rate in Wenzhou has been relatively low, less than 0.5/100,000 since the first HFRS case was reported in 1981. A total of 461 rodents and 199 shrews were captured in these two areas. Hantavirus antibodies were detected in 16 of 241 (6.64%) Rattus norvegicus and 13 of 122 (10.66%) R. flavipectus. Interestingly, hantavirus antigens were identified in 6 of 196 (3.06%) Suncus murinus. Genetic analysis showed that partial M and S segment sequences recovered from rats in the two regions belong to Seoul virus (SEOV) and can be assigned into two genetic lineages. SEOV variants that belong to these two lineages of viruses are distributed widely in China and have been found outside China. As most trapped rodents were rats and SEOV was the only hantavirus detected, these results suggested that SEOV plays an important role in human hantavirus infections. They also reinforce the need for vigilance in preventing HFRS caused by hantaviruses in the coastal region.

[Study on the molecular epidemiology of hantavirus carried by rodent hosts in Cixi, Zhejiang province].

OBJECTIVE: To collect more data on the epidemiology of hantavirus in rodents in Cixi, Zhejiang province. METHODS: Rodents were captured in Cixi, where hemorrhagic fever with renal syndrome (HFRS) appeared endemic. Hantavirus antigens in the rat lungs were detected by immunofluorescence assay. Partial S segment sequences were amplified by reverse transciption-polymerase chain reaction (RT-PCR) and then sequenced. The phyologenetic trees were constructed by maximum likelihood method to detect the genetic characteristics of hantavirus. RESULTS: A total of 243 rodents were trapped in the epidemic areas, and hantavirus antigens were identified in 7 out of these lung samples (2.88%). Partial S segment sequences (620-999 nt) were recovered from 6 samples and sequenced. Data from phylogenetic analysis of these S segment sequences indicated that all viruses belonged to Seoul virus (SEOV), despite the origins of sources were either from Rattus norvegicus or from R. flabipectus. These viruses could further be divided into two distinct lineages but the viruses carried by R. norvegicus were different from those carried by R. flabipectus. CONCLUSION: Two distinct lineages of SEOV had been cocirculating in rodents in Cixi.