Arboviruses Investigation In Xinjiang

Xinjaing is the largest provincial-level administrative region in China.It is adjacent to five provinces in China and borders on eight countries.Xinjiang is divided into North and South Regions by Tianshan Mountain which lies across the middle of Xinjiang.There are remarkable differences on aspects of climate,geography, environment and species distribution between North and South Regions of Xinjiang. Many species of arthropods are distributed in Xinjiang.Mosquitoe is major vector, which might transmit arboviruses and cause zoonotic diseases.Recent years several arboviruses such as Sindbis,Eastern equine encephalitis,Western equine encephalitis have been isolated from Xinjiang.But until now there were still no reports of population-related diseases.However,the viral encephalitis epidemiological surveillance in Xinjiang showed that there were unknown fever and viral encephalitis cases during summer.In addition,most provinces report Japanese encephalitis cases during summer except in Xinjiang.The information showed that there is significant difference between Xinjiang and other parts of China.So we carried out a systematic investigation of arbovirus etiology and correlated human infection with seroepidemiological studies.In this stydy,system investigation of arbovirus was carried out to clearly describe the existence of the species of arbovirus and distribution.Further study also identify the taxonomy and characteristics of molecular biology of the newly isolates. Finally,correlation between the arbovirus and human illness was stydied.1.Arbovirus investigation in XinjiangMosquito samples were collected in the villages of 11 counties belonged to Kashi, Hetian,Bayinguoleng,Aletai,Yili,Tacheng,Hami,from July to Septrmber in 2006, 2007 and 2008.21405 individuals had been collected.The mosquitoes were sorted into 222 pools for virus isolation.Total 50 virus isolates were obtained.47 isolates were observed to cause CPE in C6/36 cells only,while the other 3 isolates could cause CPE in both C6/36 cells and BHK-21 cells.All the isolates were identified by using morphological,serological and molecular biological methods.They are 3 Tahyna viruses(TAHV),41 Liaoning viruses,and 6 strains of unidentified virus.2.Molecular analysis of the new isolatesIn this study 3 strains of TAHV isolated from Kashi and Bayinguoleng area were sequenced.Phylogenetic analysis and differences of nucleotides and amino acids were carried out.The results showed that S segment had the highest homology among all the 3 segments,which were 98.3%-100%for nucleotide and 98.3%-100%for amino acids respectively,all the Chinese strains are between 99.2-99.6%for nucleotide and100%for amino acid.There are 81-97.4%and 97.8-100%similarity for L segment. M segment is the lowest similiarity one among all 3 segments.All these 3 Chinese strains showed higher identity than Europe strains.So it was concluded that there are correlation between the geographical distribution and the genomic character.41 strains of LNV were isolated from Kashi,Bayinguoleng and Aletai region. Segment 10 and segment 12 of LNV were sequenced.The phylogenetic analysis showed that LNV isolates from Xinjiang showed high similiarity than Jilin isolates. The identities of the 10^th segment between all Xinjiang strains were more than 94%in nucleotide and only 75%with Jilin strains.4 subtypes of LNV were classified based on nucleotide sequence of segment 10.All strains from southern region of Xinjiang belong to genotypeâ… .Genotypeâ… andâ…¡have been isolated from Aletai.Strains from Jilin were classified as genotypeâ…¢andâ…£.Genotypeâ…¡,â…¢andâ…£have some consistency mutation.So correlation may exist between all these subtypes.3.Arbovirus and correlation with illness among the local population in Xinjiang.Among them,742 cases were collected from Kashi,IgM positive rate was 5.26% (39/742),and IgG positive rate was 18.27%(59/323),no IgM positive were found in northern part of Xinjiang among 222 cases in Yili.The results indicated that in southern part of Xinjiang there were TAHV cycle and local people may be infected during Sumer and for northern part of Xinjiang,due to the Tianshan Mountain,TAHV may be not so active and few people were infected with it.To determine whether the initial illness of the patients was correlated with TAHV infection,in December 2007 second serum samples from 10 of the IgM positive patients.These paired serum specimens were tested by serum dilution neutralization test with XJ0625 virus.Most of the samples contained neutralizing antibody with a titer of 40 or 80.The results indicated that human infection with TAHV is common in the area.Furthermore,the result of IFA test coordinate with the NT test,so it may be used as a preliminary screening test.Sera samples of acute and convalescent phase of unknown encephalitis patients were screened by IFA test for several kinds of arbovirus.The results showed all the sera samples have IgG and IgM against flavivirus with a high titer and no positive results for non-flavirus.Combination with the features of clinical manifestations,prognostic and immunological responsds,it was concluded that pathogen of the epidemic may be a kind of unknown flavivirus.In the study,an pathogen investigation on an encephalitis outbreak in south Xinjiang in the summer of 2004 was conducted.The results showed only 2cases had 4 fold rises among all six pairs of acute and convalescent serum specimens collected during the disease.And the neutralizing antibody titer against JEV were very low.The remaining four pairs of specimens of JEV neutralizing antibodies were negative, according to the results,it was not sufficient to take JEV as the pathogen for the endemic.So,a wide range of arbovirus antibody screening was carried out.The results showed that IgM and IgG antibody against JEV,DENV,WNV,SLEV and other flavirus were positive and negative for TAHV and other alphavirus.Taking into account serological cross-reaction between flavivirus,these results prompted only there may be some flavivirus infection.But the real pathogen is still not certain.It is recommended that NT test against a variety of flavivirus should be carried out.In addition,641 cases of patients with unknown fever in northern and southern Xinjiang were collected in 2008 to carried out the IgM antibody screening against WNV,DENV and JEV.The results showed that 11 cases has IgM antibody against WNV,19 cases against DENV and 4 cases against JEV.These results indicated the existence of a wide range of the flavivirus in Xinjiang and may cause human diseases in summer.The importance of the studyIn summary,arbovirus investigation in Xinjang was carried out in this study during 2006,2007 and 2008.3 strains of TAHV and 41 strains of LNV were isolated from mosquitoe pools.The isolation of TAHV from southern region of Xinjiang indicated this virus prevelant in local area and no isolation from northern region of Xinjiang indicated that this virus may not active there.Molecular character of all the viruses indicated that there may be several subtypes existed in Xinjiang. Seroepidemiological results showed TAHV may be prevelant in local area and unknown fever patients may be infected by such virus.LNV existed in most region of Xinjiang,the sequence of all the strains were relatively conserved.Phylogenetic results showed four subtypes can be classified based on segment 10.Isolates from the Xinjiang region,and Jilin exist significant regional comparison.Isolates from Aletai showed relatively high homology and the characteristic consensus sequences with Jilin isolates.It suggested that there may be similar virus between Jilin and Xinjiang.These findings give us a clue to strengthen arbovirus surveillance in order to prevent arbovirus diseases.Identification and molecular analysis of TAHV and LNV isolates isolated from Xinjiang can help to know more about the virus characterization, especially the molecular difference from foreign isolates,and can provide basic information for further study of the arboviruses distributed in Chinese territory.