Objective:To compare the applicability of the SYBK Grcen-Ⅰ assay with the standard schizont maturalion assay,for determination of sensitivity of Plasmodium vivax(P.vivax) to chloroquine and a new antifolale WR 99210....Objective:To compare the applicability of the SYBK Grcen-Ⅰ assay with the standard schizont maturalion assay,for determination of sensitivity of Plasmodium vivax(P.vivax) to chloroquine and a new antifolale WR 99210.Methods:The study was conducted at Mae Tao Clinic for migrant workers,Tak Province during April 2009 to July 2010.A total of 64 blood samples(1 mL blood collected into sodium heparinized plastic tube) were collected from patients with monoinfection with P.vivax malaria prior to treatment with standard regimen of a 3-day chloroquine. In vitro sensitivity of P.vivax isolates was evaluated by schizont maturation inhibition and SYBR Green-Ⅰ assays.Results:A total of 30 out of 64 blood samples collected from patients with P.vivax malaria were successfully analyzed using both the microscopic schizont maturation inhibition and SYBR Green-I assays.The failure rates of the schizont maturation inhibition assay(50%) and the SYBR Green-I assay(54%) were similar(P=0.51).The median IC_(10)s,IC_(50)s and IC_(90)s of both chloroquine and WR99210 were not significantly different from the clinical isolates of P.vivax tested.Based on the cut-off of 100 nM,the prevalences of chloroquine resistance determined by schizont maturation inhibition and SYBR Green-I assays were 19 and 11 isolates,respectively.The strength of agreement between the two methods was very poor for both chloroquine and WR992I0.Conclusions:On the basis of this condition and its superior sensitivity,the microscopic method appears better than the SYUK Green-I Green assay for assessing in vitro sensitivity of fresh P.vivax isolates to antimalarial drugs.展开更多
Background:Plasmodium vivax remains the predominant species at the China–Myanmar border,imposing a major challenge to the recent gains in regional malaria elimination.To closely supervise the emerging of drug resista...Background:Plasmodium vivax remains the predominant species at the China–Myanmar border,imposing a major challenge to the recent gains in regional malaria elimination.To closely supervise the emerging of drug resistance in this area,we surveyed the variations in genes potentially correlated with drug resistance in P.vivax parasite and the possible drug selection with time.Methods:A total of 235 P.vivax samples were collected from patients sufering uncomplicated malaria at Yingjiang,Tengchong,and Longling counties,and Nabang port in China,Yunnan province,and Laiza sub-township in Myanmar,from 2008 to 2017.Five potential drug resistance genes were amplifed utilizing nested-PCR and analyzed,including pvdhfr,pvdhps,pvmdr1,pvcrt-o,and pvk12.The Pearson’s Chi-squared test or Fisher’s exact test were applied to determine the statistical frequency diferences of mutations between categorical data.Results:The pvdhfr F57I/L,S58R,T61M and S117T/N presented in 40.6%,56.7%,40.1%,and 56.0% of the sequenced P.vivax isolates,and these mutations signifcantly decreased with years.The haplotype formed by these quadruple mutations predominated in Yingjiang,Tengchong,Longling and Nabang.While a mutation H99S/R(56.6%)dominated in Laiza and increased with time.In pvdhps,the A383G prevailed in 69.2% of the samples,which remained the most prevalent haplotype.However,a signifcant decrease of its occurrence was also noticed over the time.The S382A/C and A553G existed in 8.4% and 30.8% of the isolates,respectively.In pvmdr1,the mutation Y976F occurred at a low frequency in 5/232(2.2%),while T958M was fxed and F1076L was approaching fxed(72.4%).The K10 insertion was detected at an occurrence of 33.2% in pvcrt-o,whereas there was no signifcant diference among the sites or over the time.No mutation was identifed in pvk12.Conclusions:Mutations related with resistance to antifolate drugs are prevalent in this area,while their frequencies decrease signifcantly with time,suggestive of increased susceptibility of P.vivax parasite to antifolate drugs.Resistance to chloroquine(CQ)is possibly emerging.However,since the molecular mechanisms underneath CQ resistance is yet to be better understood,close supervision of clinical drug efciency and continuous function investigation is urgently needed to alarm drug resistance.展开更多
The dihydrofolate reductase(dhfr)and dihydropteroate synthetase(dhps)genes of Plasmodium vivax,as antifolate resistance-associated genes were used for drug resistance surveillance.A total of 375 P.vivax isolates colle...The dihydrofolate reductase(dhfr)and dihydropteroate synthetase(dhps)genes of Plasmodium vivax,as antifolate resistance-associated genes were used for drug resistance surveillance.A total of 375 P.vivax isolates collected from different geographical locations in China in 2009–2019 were used to sequence Pvdhfr and Pvdhps.The majority of the isolates harbored a mutant type allele for Pvdhfr(94.5%)and Pvdhps(68.2%).The most predominant point mutations were S117T/N(77.7%)in Pvdhfr and A383G(66.8%)in Pvdhps.Amino acid changes were identified at nine residues in Pvdhfr.A quadruple-mutant haplotype at 57,58,61,and 117 was the most frequent(57.4%)among 16 distinct Pvdhfr haplotypes.Mutations in Pvdhps were detected at six codons,and the double-mutant A383G/A553G was the most prevalent(39.3%).Pvdhfr exhibited a higher mutation prevalence and greater diversity than Pvdhps in China.Most isolates from Yunnan carried multiple mutant haplotypes,while the majority of samples from temperate regions and Hainan Island harbored the wild type or single mutant type.This study indicated that the antifolate resistance levels of P.vivax parasites were different across China and molecular markers could be used to rapidly monitor drug resistance.Results provided evidence for updating national drug policy and treatment guidelines.展开更多
基金Supported by the Commission on Higher EducationMinistry of Education of Thailand and Thailand National Research University(NRU)
文摘Objective:To compare the applicability of the SYBK Grcen-Ⅰ assay with the standard schizont maturalion assay,for determination of sensitivity of Plasmodium vivax(P.vivax) to chloroquine and a new antifolale WR 99210.Methods:The study was conducted at Mae Tao Clinic for migrant workers,Tak Province during April 2009 to July 2010.A total of 64 blood samples(1 mL blood collected into sodium heparinized plastic tube) were collected from patients with monoinfection with P.vivax malaria prior to treatment with standard regimen of a 3-day chloroquine. In vitro sensitivity of P.vivax isolates was evaluated by schizont maturation inhibition and SYBR Green-Ⅰ assays.Results:A total of 30 out of 64 blood samples collected from patients with P.vivax malaria were successfully analyzed using both the microscopic schizont maturation inhibition and SYBR Green-I assays.The failure rates of the schizont maturation inhibition assay(50%) and the SYBR Green-I assay(54%) were similar(P=0.51).The median IC_(10)s,IC_(50)s and IC_(90)s of both chloroquine and WR99210 were not significantly different from the clinical isolates of P.vivax tested.Based on the cut-off of 100 nM,the prevalences of chloroquine resistance determined by schizont maturation inhibition and SYBR Green-I assays were 19 and 11 isolates,respectively.The strength of agreement between the two methods was very poor for both chloroquine and WR992I0.Conclusions:On the basis of this condition and its superior sensitivity,the microscopic method appears better than the SYUK Green-I Green assay for assessing in vitro sensitivity of fresh P.vivax isolates to antimalarial drugs.
文摘Background:Plasmodium vivax remains the predominant species at the China–Myanmar border,imposing a major challenge to the recent gains in regional malaria elimination.To closely supervise the emerging of drug resistance in this area,we surveyed the variations in genes potentially correlated with drug resistance in P.vivax parasite and the possible drug selection with time.Methods:A total of 235 P.vivax samples were collected from patients sufering uncomplicated malaria at Yingjiang,Tengchong,and Longling counties,and Nabang port in China,Yunnan province,and Laiza sub-township in Myanmar,from 2008 to 2017.Five potential drug resistance genes were amplifed utilizing nested-PCR and analyzed,including pvdhfr,pvdhps,pvmdr1,pvcrt-o,and pvk12.The Pearson’s Chi-squared test or Fisher’s exact test were applied to determine the statistical frequency diferences of mutations between categorical data.Results:The pvdhfr F57I/L,S58R,T61M and S117T/N presented in 40.6%,56.7%,40.1%,and 56.0% of the sequenced P.vivax isolates,and these mutations signifcantly decreased with years.The haplotype formed by these quadruple mutations predominated in Yingjiang,Tengchong,Longling and Nabang.While a mutation H99S/R(56.6%)dominated in Laiza and increased with time.In pvdhps,the A383G prevailed in 69.2% of the samples,which remained the most prevalent haplotype.However,a signifcant decrease of its occurrence was also noticed over the time.The S382A/C and A553G existed in 8.4% and 30.8% of the isolates,respectively.In pvmdr1,the mutation Y976F occurred at a low frequency in 5/232(2.2%),while T958M was fxed and F1076L was approaching fxed(72.4%).The K10 insertion was detected at an occurrence of 33.2% in pvcrt-o,whereas there was no signifcant diference among the sites or over the time.No mutation was identifed in pvk12.Conclusions:Mutations related with resistance to antifolate drugs are prevalent in this area,while their frequencies decrease signifcantly with time,suggestive of increased susceptibility of P.vivax parasite to antifolate drugs.Resistance to chloroquine(CQ)is possibly emerging.However,since the molecular mechanisms underneath CQ resistance is yet to be better understood,close supervision of clinical drug efciency and continuous function investigation is urgently needed to alarm drug resistance.
基金supported by the Natural Science Foundation of Shanghai(No.18ZR1443400)the Fifth Round of Three-year Public Health Action Plan of Shanghai(No.GWV-10.1-XK13)the National Important Scientific&Technological Project(No.2018ZX10101002-002).
文摘The dihydrofolate reductase(dhfr)and dihydropteroate synthetase(dhps)genes of Plasmodium vivax,as antifolate resistance-associated genes were used for drug resistance surveillance.A total of 375 P.vivax isolates collected from different geographical locations in China in 2009–2019 were used to sequence Pvdhfr and Pvdhps.The majority of the isolates harbored a mutant type allele for Pvdhfr(94.5%)and Pvdhps(68.2%).The most predominant point mutations were S117T/N(77.7%)in Pvdhfr and A383G(66.8%)in Pvdhps.Amino acid changes were identified at nine residues in Pvdhfr.A quadruple-mutant haplotype at 57,58,61,and 117 was the most frequent(57.4%)among 16 distinct Pvdhfr haplotypes.Mutations in Pvdhps were detected at six codons,and the double-mutant A383G/A553G was the most prevalent(39.3%).Pvdhfr exhibited a higher mutation prevalence and greater diversity than Pvdhps in China.Most isolates from Yunnan carried multiple mutant haplotypes,while the majority of samples from temperate regions and Hainan Island harbored the wild type or single mutant type.This study indicated that the antifolate resistance levels of P.vivax parasites were different across China and molecular markers could be used to rapidly monitor drug resistance.Results provided evidence for updating national drug policy and treatment guidelines.
