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. 2012 Mar 27:9:73.
doi: 10.1186/1743-422X-9-73.

Isolation and identification of a distinct strain of Culex Flavivirus from mosquitoes collected in Mainland China

Wang Huanyu  1 , Wang Haiyan, Fu Shihong, Liu Guifang, Liu Hong, Gao Xiaoyan, Song Lizhi, Simon Rayner, Xu Aiqiang, Liang Guodong
Affiliations

Isolation and identification of a distinct strain of Culex Flavivirus from mosquitoes collected in Mainland China

Wang Huanyu et al. Virol J. .

Abstract

Background: Culexflavivirus (CxFV) is an insect specific virus that has been isolated from Culexpipiens, Culexquinquefasciatus, Culextritaeniorhynchus and other Culex mosquitoes. It is a novel flavivirus isolated in Asia, North America, Central America and Africa. Phylogenetic analysis indicates that, based on the envelope gene (E gene) sequence, the worldwide CxFV strains can be divided into two genotypes.

Result: A virus (SDDM06-11) was isolated from Culexpipiens collected in Shandong Province, China in 2006. The strain caused cytopathic effect (CPE) in Aedesalbopictus (C6/36) cells by 3 days post-infection and immunofluorescence assay (IFA) showed a reaction with Japanese encephalitis virus (JEV) polyclonal antibodies. Phylogenetic analysis of the E gene sequence showed CxFV formed two genotypes with the SDDM06-11 strain assigned to genotype 1. Analysis of the E gene nucleotide homology showed the virus possessed characteristic amino acids at specific sites. The nucleotide homology of the open reading frame (ORF) was 94.8%-95.1% between SDDM06-11 and isolates from Japan, Iowa and Texas, and 90.2%-90.5% between SDDM06-11 and isolates from Uganda and Mexico.

Conclusion: In this paper we report the first isolation and identification of an isolate of CxFV in mainland China. Phylogenetic analysis indicates the isolate belongs to genotype 1. Our findings provide insight into the occurrence of CxFV in Culex mosquito populations and its distribution on a global scale.

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Figures

Figure 1
Figure 1
Phase contrast photomicrographs of control and infected C6/36 cells after infection. (A) Control cells. (B) Cells 3 days after infection with SDDM06-11. Note the extensive cell fusion and syncytia formation in the infected cells. Microscope settings Ocular: 10; Lens: 10X. Scale bar, 50 μm.
Figure 2
Figure 2
Immunofluorescence assay (IFA) of SDDM06-11 using JEV polyclonal antibody (JEV-GSS). (A) C6/36 cell control (uninfected) stained by IFA, using JEV polyclonal antibody (JEV-GSS). (B) SDDM06-11 antigen in infected C6/36 cells, as detected by IFA, using JEV polyclonal antibody (JEV-GSS). Scale bar, 50 μm.
Figure 3
Figure 3
Phylogenetic analysis based on the E gene of CxFV strain isolated in China. Phylogenetic tree generated using the ML method. The tree was rooted by using CRFV (GenBank accession no. NC001564) sequence as the outgroup. Horizontal branch lengths are proportional to genetic distance; scale bars indicate a genetic distance of 10-nt substitutions per site. Isolate obtained in Shandong Province is shown in boldface and blue. Isolates from United States are shown in red and isolates from Japan and Indonesia are shown in green. See Table 2 for sequence information. CN: China; US: United States; JP: Japan; ID: Indonesia.
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