[ Objective ] The paper aimed to establish a real-time fluorescent quantitative PCR （qPCR） detection method for Pineapple mealybug wilt associated vi- rus-3 （ PMWaV3 ）. [ Method] Specific TaqMan probe and primers ...[ Objective ] The paper aimed to establish a real-time fluorescent quantitative PCR （qPCR） detection method for Pineapple mealybug wilt associated vi- rus-3 （ PMWaV3 ）. [ Method] Specific TaqMan probe and primers were designed and synthesized according to the conserved sequence of coat protein（CP） gene of PMWaV-3, and the standard curve was established after optimizing the amplification condition of qPCR. [ Result] The results showed that the method was specific for the detection of PMWaV-3, and the sensitivity of the present method was about 10 times higher compared to general RT-PCR ; the variation coefficients of intra- assay and inter-assay were less than 1.73, respectively. [ Conclusion] The qPCR is an easy, fast and reliable method for quantitative detection of PMWaV-3.展开更多
Objective:To study evolutionary relationship of the 5'untranslated regions(5'UTRs) in low passage dengue3 viruses(DEN3) isolated from hospitalized children with different clinical manifestations in Bangkok dur...Objective:To study evolutionary relationship of the 5'untranslated regions(5'UTRs) in low passage dengue3 viruses(DEN3) isolated from hospitalized children with different clinical manifestations in Bangkok during 24 year-evolution(1977-2000) comparing to the DEN3prototype(H87).Methods:The 5'UTRs of these Thai DEN3 and the H87 prototype were amplified by RT-PCR and sequenced.Their multiple sequence alignments were done by Codon Code Aligner v 4.0.4 software and their RNA secondary structures were predicted by MFOLD software.Replication of five Thai DEN3 candidates comparing to the 1187 prototype were done in human(HepG2) and the mosquito(C6/36) cell lines.Results:Among these Thai DEN3,the completely identical sequences of their first 89 nucleotides,their high-order secondary structure of 5'UTRs and three SNPs including the predominant C90 T,and two minor SNPs including A109 G and A112 G were found.The C90 T of Thai DEN3.Bangkok isolates was shown predominantly before 1977.Five Thai DEN3 candidates with the predominant C90 T were shown to replicate in human(HepG2) and the mosquito(C6/36) cell lines better than the H87 prototype.However,their highly conserved sequences as well as SNPs of the 5'UTR did not appear to correlate with their disease severity in human.Conclusions:Our findings highlighted evolutionary relationship of the completely identical 89 nucleotide sequence,the high-order secondary structure and the predominant C90 T of the 5'UTR of these Thai DEN3 during 24 year-evolution further suggesting to be their genetic markers and magic targets for future research on antiviral therapy as well as vaccine approaches of Thai DEN3.展开更多
AIM:To evaluate steatosis,insulin resistance(IR)and patatin-like phospholipase domain-containing 3(PNPLA3) and their relation to disease progression in hepatitis B and C viruses(HCV-HBV) coinfected patients.METHODS:Th...AIM:To evaluate steatosis,insulin resistance(IR)and patatin-like phospholipase domain-containing 3(PNPLA3) and their relation to disease progression in hepatitis B and C viruses(HCV-HBV) coinfected patients.METHODS:Three hundred and thirty patients with biopsy proven chronic hepatitis were enrolled:66 had HBV-HCV,66 HBV and 198 HCV infection.Prevalence of steatosis,IR and PNPLA3 polymorphisms and their relation to anthropometric,biochemical,virological and histological parameters were evaluated.RESULTS:Prevalence of steatosis in group HBV-HCV was similar to that in HCV(47.0% vs 49.5%,respec-tively);group HBV showed the lowest steatosis(33.3%).Group HBV-HCV had a lesser degree of steatosis than HCV(P = 0.016),lower HCV RNA levels(P = 0.025) and lower prevalence and degree of IR(P = 0.01).PNPLA3 polymorphisms were associated with steatosis.Group HBV-HCV showed higher levels of liver fibrosis than group HCV(P = 0.001),but similar to that ob-served in HBV group.In HBV-HCV group,liver fibrosis was not associated with steatosis,IR or PNPLA3.HBV infection was the independent predictor of advanced liver fibrosis.CONCLUSION:HBV-HCV co-infected patients have lower degree of hepatic steatosis,IR and HCV RNA than HCV mono-infected;co-infected patients showed a more rapid liver fibrosis progression that seems to be due to the double infection and/or HBV dominance.展开更多
基金Supported by Applied Research and Industrialization Projects of Key Science and Technology Plan of Hainan Province(ZDXM20130031)Special Fund for Agro-scientific Research in the Public Interest(201203021)+1 种基金Scientific Operation Fund of Hainan Province(QCY[2013]131)Natural Science Foundation of Hainan Province(QK[2013]32)
文摘[ Objective ] The paper aimed to establish a real-time fluorescent quantitative PCR （qPCR） detection method for Pineapple mealybug wilt associated vi- rus-3 （ PMWaV3 ）. [ Method] Specific TaqMan probe and primers were designed and synthesized according to the conserved sequence of coat protein（CP） gene of PMWaV-3, and the standard curve was established after optimizing the amplification condition of qPCR. [ Result] The results showed that the method was specific for the detection of PMWaV-3, and the sensitivity of the present method was about 10 times higher compared to general RT-PCR ; the variation coefficients of intra- assay and inter-assay were less than 1.73, respectively. [ Conclusion] The qPCR is an easy, fast and reliable method for quantitative detection of PMWaV-3.
基金supported by two research grants of Associate Professor Dr.W.Attatippaholkun:Grant No.493-5600-G-00-3461,Program in Science and Technology Cooperation,Human Capacity Development,Bureau for Global Programs,Field Support and Research,US Agency for International Development,Washington,DCThe Royal Golden Jubilee-Ph.D Program,Thailand Research Fund,Thailand
文摘Objective:To study evolutionary relationship of the 5'untranslated regions(5'UTRs) in low passage dengue3 viruses(DEN3) isolated from hospitalized children with different clinical manifestations in Bangkok during 24 year-evolution(1977-2000) comparing to the DEN3prototype(H87).Methods:The 5'UTRs of these Thai DEN3 and the H87 prototype were amplified by RT-PCR and sequenced.Their multiple sequence alignments were done by Codon Code Aligner v 4.0.4 software and their RNA secondary structures were predicted by MFOLD software.Replication of five Thai DEN3 candidates comparing to the 1187 prototype were done in human(HepG2) and the mosquito(C6/36) cell lines.Results:Among these Thai DEN3,the completely identical sequences of their first 89 nucleotides,their high-order secondary structure of 5'UTRs and three SNPs including the predominant C90 T,and two minor SNPs including A109 G and A112 G were found.The C90 T of Thai DEN3.Bangkok isolates was shown predominantly before 1977.Five Thai DEN3 candidates with the predominant C90 T were shown to replicate in human(HepG2) and the mosquito(C6/36) cell lines better than the H87 prototype.However,their highly conserved sequences as well as SNPs of the 5'UTR did not appear to correlate with their disease severity in human.Conclusions:Our findings highlighted evolutionary relationship of the completely identical 89 nucleotide sequence,the high-order secondary structure and the predominant C90 T of the 5'UTR of these Thai DEN3 during 24 year-evolution further suggesting to be their genetic markers and magic targets for future research on antiviral therapy as well as vaccine approaches of Thai DEN3.
文摘AIM:To evaluate steatosis,insulin resistance(IR)and patatin-like phospholipase domain-containing 3(PNPLA3) and their relation to disease progression in hepatitis B and C viruses(HCV-HBV) coinfected patients.METHODS:Three hundred and thirty patients with biopsy proven chronic hepatitis were enrolled:66 had HBV-HCV,66 HBV and 198 HCV infection.Prevalence of steatosis,IR and PNPLA3 polymorphisms and their relation to anthropometric,biochemical,virological and histological parameters were evaluated.RESULTS:Prevalence of steatosis in group HBV-HCV was similar to that in HCV(47.0% vs 49.5%,respec-tively);group HBV showed the lowest steatosis(33.3%).Group HBV-HCV had a lesser degree of steatosis than HCV(P = 0.016),lower HCV RNA levels(P = 0.025) and lower prevalence and degree of IR(P = 0.01).PNPLA3 polymorphisms were associated with steatosis.Group HBV-HCV showed higher levels of liver fibrosis than group HCV(P = 0.001),but similar to that ob-served in HBV group.In HBV-HCV group,liver fibrosis was not associated with steatosis,IR or PNPLA3.HBV infection was the independent predictor of advanced liver fibrosis.CONCLUSION:HBV-HCV co-infected patients have lower degree of hepatic steatosis,IR and HCV RNA than HCV mono-infected;co-infected patients showed a more rapid liver fibrosis progression that seems to be due to the double infection and/or HBV dominance.
