Association between IL-10 gene polymorphisms and the outcomes of hepatitis C virus infection among high-risk populations in Jiangsu province / 中华流行病学杂志

Objective To explore the relationship between interleukin (IL)-10 gene polymorphisms and the susceptibility or the outcomes of HCV infection among high-risk populations in Jiangsu province.Methods IL-10 gene SNPs were detected in 1555 subjects including 264 self-limited HCV infections.371 persistent HCV infections and 920 healthy controls were selected through Taqman-MGB.Results After adjusted for cofounders as sex,age and high-risk population,data from logistic regression analysis showed that the distribution of IL-10 genotypes among the controls,spontaneous clearances and those with persistent infections did not show much differences.Results from further stratified analysis showed that,at the position of-819T/C,when compared with TT genotype,TC genotype had a significantly increasing chance of self-limited HCV infection among middle-aged,females and paid blood doners (adjusted OR values and 95％ CI were:2.160,1.163 4.011 ；1.693,1.066-2.688 and 4.084,1.743-9.570).It also had a lower risk of progressing to persistent HCV infection among those paid blood doners (the adjusted OR values and 95％CI were:0.312,0.130-0.747 ).CC genotype had a higher chance of self-limited HCV infection among people underwent blood dialysis (the adjusted OR values and 95％CI were:2.120,1.071 -4.197).Results also showed a decreased risk of progressing to persistent infection among paid blood doners (the adjusted OR values and 95％CI were:0.156,0.043-0.566).At the position of -592A/C,when compared to AA genotypc,the AC genotype had a significantly increasing chance of self-limited HCV infection among middle-aged,femalcs and paid blood doners (the adjusted OR values and 95％ CI were:2.176,1.173-4.037；1.659,1.055-2.607;3.704,1.625-8.443) but had an increased risk of persistent HCV infection among females (the adjusted OR values and 95％CI were:1.525,1.017-2.286).AC genotype showed an increased opportunity to progress to HCV persistent infection among drug users (the adjusted OR values and 95％CI were:1.845,1.122-3.034) but had a reduced risk of progressing to HCV persistent infection among paid blood doners (the adjusted OR values and 95％CI were:0.361,0.155-0.841 ).CC genotype had an increased opportunity to self-linited HCV infection as well as having a dccreased risk of progressing to persistent infection among paid blood donets (the adjusted OR values and 95％CI were:3.125,1.016-9.605；0.218,0.063-0.748).At the position of-1082A/G,AG/GG genotypcs had an increased chance of self-limited infection among blood doners (the adjusted OR values and 95％CI were:3.780,1.620-8.820).Conclusion IL-10-819T/C,-592A/C,-1082A/G SNPs might be related with the susceptibility and the outcomes of HCV infection among populations at high risk.

A study of immunoassay by using quantum dots to detect Hantavirus infection / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To develop a new method to detect anti-Hantavirus IgG antibodies (HV IgG) based on quantum dots (QDs) and indirect immune technique.</p><p><b>METHODS</b>The carbodiimide crosslinking method was used to couple protein G and goat antihuman IgG on the surface of water-solubility QDs. The coverglass covered HV antigen was used as carrier, and QDs-PG-IgG conjugates was used as labeled second antibody to detect the HV-IgG in the serum samples. The detecting conditions were optimized.</p><p><b>RESULTS</b>The optimum reaction time, pH and goat antihuman IgG concentration for conjugating the QDs with goat antihuman IgG were 6.0, 2h, and 20 microg/ml, respectively. The optimum working dilution of QDs-PG-IgG conjugates was 1: 200. The detection limit of the serum samples was about 1: 1280 dilution.</p><p><b>CONCLUSION</b>The method established in this study has been demonstrated to be a specific, sensitive, rapid test for detecting HV antibodies, laying the foundation of single molecule detection. The anti-fluorescence quenching ability of this method was significant improved.</p>

Study on the transmission of Hantaan virus and Orientia tsutsugamushi by naturally dual infected Leptotrombidium scutellare through stinging / 中华预防医学杂志

<p><b>OBJECTIVE</b>To investigate whether Leptotrombidium scutellare could be naturally infected by both Hantaan virus (HV) and Orientia tsutsugamushi (OT) and transmission status by stinging.</p><p><b>METHODS</b>3459 Leptotrombidium scutellares from mice bodies and 3265 which were free were collected in the epidemic area of hemorrhagic fever with renal syndrome (HFRS) and tsutsugamushi disease.15 days later, the suspensions of lung and spleen of mice with 6 in a group stung by 1, 5 or 10 infected mites were injected intra-cerebrally into other mice for the detection of HV and OT in the next 6 generations of the mice, with immunofluorescent antibody technique (IFAT) and Giemsa staining technique. The passages of Vero-E6 cells inoculated on the aseptic filtrations from different number of infected mites were used to detect HV and OT pathogens. HV-RNA and OT-DNA were detected by PCR.</p><p><b>RESULTS</b>After passage, HV positive mouse body mite group out of both 5 and 10 mites in the sixth generation, OT positive mouse body mite group out of the 10 mites in the sixth generation, both HV and OT positive mouse body mite group out of 1 mite in the fifth and sixth generation, both HV and OT positive mouse body mite group out of 5 and 10 mites in the sixth generation, and free mites group out of 1, 5 and 10 mites in the sixth generation, were found one mouse infected by both HV and OT, respectively. Out of the fourth generation of Vero-E6 cells, one sample was found both HV and OT positive out of 5 and 10 HV and OT mouse body mite group, respectively. In the sixth generation, both HV and OT positive cells were detected in one mouse mite group and the 1, 5, 10 free mite groups, respectively. HV-RNA and OT-DNA were all detected by PCR.</p><p><b>CONCLUSION</b>Both HV and OT could be coexisted in wild Leptotrombidium scutellare and transmitted by stinging.</p>

Molecular evolution analysis of hantaviruses in Zhejiang Province / 病毒学报

In order to analyze the molecular epidemiology of Hantavirus (HV) in Zhejiang Province, the complete M and S genome sequences of 12 HV strains from different hosts and locations in Zhejiang Province of China during the period of 1981-2007 were analyzed on genetic evolution by DNAstar and MEGA 4.0 software in this research. Phylogenetic analyses revealed that HTN and SEO strains were co-circulating in Zhejiang Province, and the difference in sequence similarity and the phylogeny was closely related to the isolated regions, but had no distinct relationship with the isolate year and the host, indicating a relationship between epidemiology of HFRS and the distribution region, especially in HTNV. The isolates in the same region could be assigned in same or near phylogenetic clade sharing high sequence similarity. Interestingly, the Gou3 strain and ZJ5 strain isolated from Jiande region in Zhejiang Province formed a distinct phylogenetic lineage in SEOV clade, and different from the other SEOV variants outside China. We believed that the special SEOV variants were distributed in Jiande region.

Expression of partial S gene of Hantavirus Z10 in Pichia pastoris / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>The purpose of this study is to express partial S gene of Hantavirus Z10.</p><p><b>METHODS</b>The 300 bp S gene of Z10 strain was synthesized by using a successive PCR method for the optimal expression in Pichia pastoris and subcloned into pMD19-T. The SP300 gene was constructed into pPICZaA and sequenced. The recombinant pPICZaA-SP300 and pPICZaA-S300 was transformed into Pichia with LiCI.</p><p><b>RESULTS</b>The recombination Pichia were cultivate, and expressed the SP300 or S300 gene induced in Pichia by methanol.</p><p><b>CONCLUSION</b>The nucleocapsid secreted from the Pichia can be detected by ELISA and WesternBlot.</p>

Study on the difference of genes and the type identification of hantavirus from Lishui,Zhejiang province / 中华流行病学杂志

Objective To isolate hantavirus from Lishui county-one of the epidemic regions for hemorrhagic fever with renal syndrome(HFRS),in Zhejiang province,and to identify the serotype and molecular/biological characteristics of a new HTN subtype hantavirus(HV)strains,hopefully to provide evidence for HFRS prevention and therapy.Methods Data on the host animals was collected from Lishui,Zhejiang province in 2007.Direct immunofluorscece assay was adopted to determine HFRS antigens and the lung tissues from HV infeeted Vero-E6 cells for HV isolation,then total RNA was extracted from Hantavirus Lishui strains and amplified by RT-PCR M,S segments of strains genome were also clened and sequenced and compared with those of other strains of HV Results 2 strains virus(ZLS6-11 and ZLS-12)Were successfully isolated from 7 positive lung samples of mice and were identified as HTNV by anti-McAb and phylogenetic analysis.With sequence compation.We found that 2 strains with complete M and S segment had higher homology with HTN-type strains than with other types of HV,but 13.4%-20.7%and 10.3%-16.1%of the genes were found which were difierent from HTNV.The phylogenetic trees constructed by complete S and M segment showed that ZLS6-11 and ZLS-12 strains were located in HTNV group,and structured independent embranchment.Conclusion ZLS6-11 and ZLS-12 Strains were believed to belong to HTN-type and phylogenetically different from the HTNV.

Influence of F protein of hepatitis C virus subtype 1b inhibits on human hepatocellular carcinom HepG2 cell apoptosis / 中华流行病学杂志

Objective To investigate the effects of F protein of hepatitis C virus subtype lb on the apoptosis of human hepatocellular carcinom HepG2 cells. Methods HepG2 cells were transfected with recombinant plasmid pcDNA3.0-F-EGFP and pcDNA3.0-F-EGFP-HepG2 strain was exposed to Act-D and tumor necrosis factor a (TNFα) treatment in order to induce cell apoptosis with positive control pcDNA3.0-C-EGFP-HepG2, negative control pcDNA3.0-C-EGFP-HepG2 and blank control HepG2.Annexin V-FITC/PI of flow cytometry was performed to determine the number of apoptotic cells. DNA Ladder was used to observe the isolation of apoptotic DNA fragments in the apoptotie cells. Results pcDNA3.0-F-EGFP- HepG2 cell strain showed a much delayed apoptosis as well as obviously lowering the apoptotic rate when compared with the pcDNA3.0-HepG2 strain and HepG2 strain (P<0.001).Conclusion The introduction and expression of extraneous gene (the F gene of hepatitis C virus subtype 1b) could significantly inhibit the apoptosis of HepG2 cells.

Molecular characterization and sequence comparison of the M and S segments of Hantavirus ZJ5 strain / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>In order to understand the molecular characters of Hantavirus ZJ5 strain, its complete M and S genome were sequenced and compared with that of other hantavirus strains.</p><p><b>METHODS</b>We prepared the total RNA from ZJ5. Infected cells and the raw or purified RT-PCR product was cloned and sequenced.</p><p><b>RESULTS</b>With sequence compation, we found ZJ5 strain complete M and S segment had higher homology with SEO-type strains than other type of HV, but differential genes were 11.7%-19.2% and 6.7%-14.5% from SEOV. The phylogenetic trees constructed by complete M ind S segment showed that ZJ5 strain was located in SEOV group, and structured alone embranchment.</p><p><b>CONCLUSION</b>The ZJ5 strain was believed to belong to SEO-type virus,and suggest that ZJ5 strain is a new subtype S SEOV group,and structured alone embranchment.</p><p><b>CONCLUSION</b>The ZJ5 strain was believed to belong to SEO-type virus, and suggest that ZJ5 strain is a new subtype from other SEO viruses.</p>

Study on the molecular epidemiology of Hantavirus carried by rodent hosts in Cixi, Zhejiang province / 中华流行病学杂志

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(RTPCR)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.fzabipectus.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.

Molecular epidemiological study on the host and role of the Hantavirus and Orientia tsutsugamushi in the same epidemic area / 中华预防医学杂志

<p><b>OBJECTIVE</b>To investigate whether Hantavirus (HV) and Orientia tsutsugamushi ( OT) can naturally infect and coexist in their host and role.</p><p><b>METHODS</b>By field epidemiological study, Leptotrombidium scutellare (3829) was collected and separated from mice(166) in epidemic areas. The cells of mites separated from their host and role were cultured. PCR was used to detect HV-RNA and OT-DNA in the cell culture.</p><p><b>RESULTS</b>In 105 Apodemus agrarius, 3 HV-RNA positive, 2 OT-DNA positive and 2 coinfection with HV and OT were detected;in 41 Brown rattus, 2 HV-RNA positive, 1 OT-DNA positive and 1 co-infection with HV and OT were detected. From 15 mites co-infected with HV and OT, 2 strains of HV pathogen, 2 strains of OT pathogen were separated and 1 HV and OT pathogen in the same mite were separate.</p><p><b>CONCLUSION</b>The study demonstrates that co-infection of HV and OT did simultaneously exist in wild Leptotrombidium scutellare. This theory has some significance to the epidemic and precaution of HV and OT.</p>

Detection of serum anti-F antibody in hepatitis C virus infected patients / 中华流行病学杂志

<p><b>OBJECTIVE</b>To assess the prevalence of serum anti-F in patients with hepatitis C virus (HCV) infection and the distribution of anti-F.</p><p><b>METHODS</b>The recombinant protein (HCV-F/GST) was coated onto micro titer plates as antigen. Sera of 120 patients with hepatitis C virus infection, 15 patients with hepatitis B, 3 patients with hepatitis E and 10 normal sera were tested by indirect ELISA for detecting anti-F.</p><p><b>RESULTS</b>82 samples out of the 120 (68%) HCV infected patients exhibited a positive anti-F reaction, showing significant difference from the controls with no HCV infection (P < 0.01). Data from logistic analysis showed that the positive rate of anti-F was higher in patients over 50 year olds (OR = 6.675, 95% CI: 2.407-19.071). Patients of midrange, severe phase and hepatic cirrhosis had higher rate than the others (OR = 2.749, 95% CI: 1.470-5.141).</p><p><b>CONCLUSION</b>Prevalence and distribution of anti-F in Yixing hepatitis C patients was reported and which might be related to the progression of HCV infection.</p>

Detection the coinfection of Hantavirus and Orientia tsutsugamushi in primary cultured mite cells / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To investigate whether HV and Ot can coexist in their host (Leptotrombidium scutellare).</p><p><b>METHODS</b>Collecting the separate Leptotrombidium scutellare and the ones from mice in epidemic area. The cells of mites at larva, nymph, and adult stages were cultured and made into smear. In situ RT-PCR and PCR were used to detect and locate HV RNA and Ot DNA in the primary cultured cells.</p><p><b>RESULTS</b>Positive signals of HV RNA and Ot DNA distributed mostly in epithelial cells of digestive system and ovary cells of larva and nymph. The positive rate increased by the generation of passages.</p><p><b>CONCLUSION</b>Coinfection of HV and Ot did exist in wild Leptotrombidium scutellare.</p>

Study on the coinfection of Hantavirus and Orientia tsutsugamushi in tissue cell culture / 中华流行病学杂志

<p><b>OBJECTIVE</b>To investigate the possibility of Hantavirus (HV) and Orientia tsutsugamushi (Ot) coinfection in their hosts.</p><p><b>METHODS</b>HV and Ot were used to infect Vero E6 cells cultured in vitro singly, simultaneously or successively. Genes of HV and Ot were identified in different generation cells with RT-PCR.</p><p><b>RESULTS</b>Five experiment groups of infected Vero E6 cells were tested, the results were as follows: HV and Ot were both positive in infected Vero E6 cells passaged 2 times and the positive rate increased following the passaged times in HV and Ot infection groups, simultaneously or successively. However, in the groups which were infected with HV and Ot separately, the gene of HV or Ot could be detected in infected Vero E6 cells passaged only once and the positive rate increased following the times of the passaged. The positive rate was higher in the singly infected groups than in those infected simultaneously or successively.</p><p><b>CONCLUSION</b>Coinfection of HV and Ot did exist in the hosts while HV and Ot could inhibit each other in the initial infection stage.</p>

Study on hepatitis C virus genotyping in Yixing area, Jiangsu province / 中华流行病学杂志

<p><b>OBJECTIVE</b>To investigate the distribution of hepatitis C virus (HCV) genotypes in Yixing, Jiangsu province.</p><p><b>METHODS</b>Genotypes identification on sera samples were obtained from 158 donors who had already been anti-HCV positive through PCR method with type specific primer designed according to the sequence of 5'non-coding region (5'NCR). 5'NCR was also sequenced and compared with published date. Genotypes distribution was investigated in patients with different sex and clinical types of hepatitis C.</p><p><b>RESULTS</b>Of the total 158 patients, 95 were HCV RNA positive in which 80 patients having genotype 1b (80/95; 84.4%), 5 patients having genotype 2(5/95; 5.3%), 5 patients with 1b/2 mixed genotypes (5/ 95; 5.3%) and another 5 patients whose genotype undetermined. The difference on the distribution of HCV genotypes was significant between female and male patients (P < 0.05) but not in different kinds of hepatitis C patients.</p><p><b>CONCLUSION</b>Type 1b was the predominant HCV genotype in Yixing area.</p>

Detection of hantaan virus from gamasid mite and chigger mite by molecular biological methods / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the proliferation and location of hantaan virus (HV) in gamasid mites and chigger mites.</p><p><b>METHODS</b>HV RNA in gamasid mites and chigger mites were detected by reverse transcription, polymerase chain reaction (RT- PCR) and in situ hybridization.</p><p><b>RESULTS</b>The smallest quantity of mite from which HV RNA could be detected was 5 mites group. The titers of -and proliferated in mites HV RNA could be found in ovary cells and dug cells of gamasid mites and chigger mites by in situ hybridization.</p><p><b>CONCLUSIONS</b>The results showed that HV could be trans-stadially transmitted and proliferated in mites, and HV always located in ovary and dug organs of mites. These results provide direct evidence at molecular level for the role of gamasid mites and chigger mites as vectors in transmission of HV.</p>