Background Culex pipiens pallens and Culex pipiens quinquefasciatus are the dominant species of Culex mosquitoes in China and important disease vectors.Long-term use of insecticides can cause mutations in the voltage-...Background Culex pipiens pallens and Culex pipiens quinquefasciatus are the dominant species of Culex mosquitoes in China and important disease vectors.Long-term use of insecticides can cause mutations in the voltage-gated sodium channel(vgsc)gene of mosquitoes,but little is known about the current status and evolutionary origins of vgsc gene in different geographic populations.Therefore,this study aimed to determine the current status of vgsc genes in Cx.p.pallens and Cx.p.quinquefasciatus in China and to investigate the evolutionary inheritance of neighboring downstream introns of the vgsc gene to determine the impact of insecticides on long-term evolution.Methods Sampling was conducted from July to September 2021 in representative habitats of 22 provincial-level administrative divisions in China.Genomic DNA was extracted from 1308 mosquitoes,the IIS6 fragment of the vgsc gene on the nerve cell membrane was amplified using polymerase chain reaction,and the sequence was used to evaluate allele frequency and knockdown resistance(kdr)frequency.MEGA 11 was used to construct neighborjoining(NJ)tree.PopART was used to build a TCS network.Results There were 6 alleles and 6 genotypes at the L1014 locus,which included the wild-type alleles TTA/L and CTA/L and the mutant alleles TTT/F,TTC/F,TCT/S and TCA/S.The geographic populations with a kdr frequency less than 20.00%were mainly concentrated in the regions north of 38°N,and the geographic populations with a kdr frequency greater than 80.00%were concentrated in the regions south of 30°N.kdr frequency increased with decreasing latitude.And within the same latitude,the frequency of kdr in large cities is relatively high.Mutations were correlated with the number of introns.The mutant allele TCA/S has only one intron,the mutant allele TTT/F has three introns,and the wild-type allele TTA/L has 17 introns.Conclusions Cx.p.pallens and Cx.p.quinquefasciatus have developed resistance to insecticides in most regions of China.The neighboring downstream introns of the vgsc gene gradually decreased to one intron with the mutation of the vgsc gene.Mutations may originate from multiple mutation events rather than from a single origin,and populations lacking mutations may be genetically isolated.展开更多
Background Overseas imported dengue fever is an important factor in local outbreaks of this disease in the mainland of China.To better prevent and control such local outbreaks,the epidemiological characteristics and t...Background Overseas imported dengue fever is an important factor in local outbreaks of this disease in the mainland of China.To better prevent and control such local outbreaks,the epidemiological characteristics and temporal-spatial distribution of overseas imported dengue fever cases in provincial-level administrative divisions(PLADs)where dengue fever is outbreak in the mainland of China were explored.Methods Using the Chinese National Notifiable Infectious Disease Reporting Information System(CNNDS),we identified overseas imported dengue fever cases in dengue fever outbreak areas in the mainland of China from 2005 to 2019 to draw the epidemic curve and population characteristic distribution of overseas imported cases in each PLAD.Based on spatial autocorrelation analysis of ArcGIS 10.5 and temporal-spatial scanning analysis of SaTScan 9.5,we analyzed the temporal-spatial distribution of overseas imported dengue fever in dengue fever outbreak areas in the mainland of China.Results A total of 11,407 imported cases,mainly from Southeast Asia,were recorded from 2005 to 2019 in these 13 PLADs.Of which 62.1%were imported into Yunnan and Guangdong Provinces.Among the imported cases,there were more males than females,mainly from the 21–50 age group.The hot spots were concentrated in parts of Yunnan,Guangdong and Fujian Provinces.We found the cluster of infected areas were expanding northward.Conclusions Based on the analysis of overseas imported dengue cases in 13 PLADs of the mainland of China from 2005 to 2019,we obtained the epidemiological characteristics and spatial distribution of imported dengue cases.Border controls need to pay attention to key population sectors,such as 21–50 years old men and education of key populations on dengue prevention.There is a need to improve the awareness of the prevention and control of imported cases in border areas.At the same time,northern regions cannot relax their vigilance.展开更多
Background The current prevention and control strategy forAedes albopictus heavily relies on comprehensive management, such as environmental management and chemical control. However, the wide application of pyrethroid...Background The current prevention and control strategy forAedes albopictus heavily relies on comprehensive management, such as environmental management and chemical control. However, the wide application of pyrethroids has facilitated the development of insecticide resistance, primarily via mutations in the voltage-gated sodium channel (VGSC) gene. This study aims to develop a novel strategy for detecting mutations in the VGSC gene inAe. albopictus using multiplex PCR-mass spectrometry (MPCR-MS) minisequencing technology.Methods We established a new strategy for detecting mutations in the VGSC gene inAe. albopictus using MPCR-MS minisequencing technology. MPCR amplification and mass probe extension (MPE) were first used, followed by single nucleotide polymorphism (SNP) typing mass spectrometry, which allows the simultaneous detection of multiple mutation sites of the VGSC gene in 96 samples ofAe. albopictus. A total of 70 wild-collectedAe. albopictus were used to evaluate the performance of the method by comparing it with other methods.Results Three target sites (1016, 1532, 1534) in the VGSC gene can be detected simultaneously by double PCR amplification combined with matrix-assisted laser desorption ionization-time-of-flight mass spectrometry, achieving a detection limit of 20 fg/μl. We applied this method to 70 wild-collectedAe. albopictus, and the obtained genotypes were consistent with the routine sequencing results, suggesting the accuracy of our method.Conclusions MPCR-MS minisequencing technology provides a sensitive and high-throughput approach toAe. albopictus VGSC gene mutation screening. Compared with conventional sequencing, this method is economical and time-saving. It is of great value for insecticide resistance surveillance in areas with a high risk of vector-borne disease.展开更多
Introduction:Mosquito-borne flavivirus can lead to serious infectious diseases worldwide and cause high mortality and disability.In order to strengthen epidemiological investigation of flavivirus and meet the needs of...Introduction:Mosquito-borne flavivirus can lead to serious infectious diseases worldwide and cause high mortality and disability.In order to strengthen epidemiological investigation of flavivirus and meet the needs of early warning of diseases,a simple,rapid,and sensitive detection method needs to be established to prevent and control mosquito-borne diseases.Methods:Using the NS5 gene of flavivirus in GenBank,3 universal primers targeting the conserved regions were designed.The complementary DNAs(cDNAs)of Japanese encephalitis virus(JEV),dengue virus(DENV),West Nile virus(WNV),and yellow fever virus(YFV)were used as the template to optimize the reaction conditions.A heminested reversetranscriptase polymerase chain reaction(hnRT-PCR)was established to verify the sensitivity and specificity of this method and to detect field-caught mosquitoes.Results:Our results showed that this method exhibited better specificity,higher sensitivity,and the ability to detect multiple viruses simultaneously.The lowest detection limit of JEV,DENV-2,YFV,and WNV was 3×10^(4),3×10^(6),3×10^(5),and 3×10^(4)copies/μL,respectively.Mosquito-borne flavivirus was successfully detected in the field-caught mosquito samples using the method established in this study.Discussion:The hnRT-PCR method established in this study can be employed for the rapid detection of flavivirus and provide technical support for early and rapid diagnosis of mosquito-borne flavivirus.展开更多
基金funded by the National Science and Technology Major Project of China(No.2018ZX10101002-002)
文摘Background Culex pipiens pallens and Culex pipiens quinquefasciatus are the dominant species of Culex mosquitoes in China and important disease vectors.Long-term use of insecticides can cause mutations in the voltage-gated sodium channel(vgsc)gene of mosquitoes,but little is known about the current status and evolutionary origins of vgsc gene in different geographic populations.Therefore,this study aimed to determine the current status of vgsc genes in Cx.p.pallens and Cx.p.quinquefasciatus in China and to investigate the evolutionary inheritance of neighboring downstream introns of the vgsc gene to determine the impact of insecticides on long-term evolution.Methods Sampling was conducted from July to September 2021 in representative habitats of 22 provincial-level administrative divisions in China.Genomic DNA was extracted from 1308 mosquitoes,the IIS6 fragment of the vgsc gene on the nerve cell membrane was amplified using polymerase chain reaction,and the sequence was used to evaluate allele frequency and knockdown resistance(kdr)frequency.MEGA 11 was used to construct neighborjoining(NJ)tree.PopART was used to build a TCS network.Results There were 6 alleles and 6 genotypes at the L1014 locus,which included the wild-type alleles TTA/L and CTA/L and the mutant alleles TTT/F,TTC/F,TCT/S and TCA/S.The geographic populations with a kdr frequency less than 20.00%were mainly concentrated in the regions north of 38°N,and the geographic populations with a kdr frequency greater than 80.00%were concentrated in the regions south of 30°N.kdr frequency increased with decreasing latitude.And within the same latitude,the frequency of kdr in large cities is relatively high.Mutations were correlated with the number of introns.The mutant allele TCA/S has only one intron,the mutant allele TTT/F has three introns,and the wild-type allele TTA/L has 17 introns.Conclusions Cx.p.pallens and Cx.p.quinquefasciatus have developed resistance to insecticides in most regions of China.The neighboring downstream introns of the vgsc gene gradually decreased to one intron with the mutation of the vgsc gene.Mutations may originate from multiple mutation events rather than from a single origin,and populations lacking mutations may be genetically isolated.
文摘Background Overseas imported dengue fever is an important factor in local outbreaks of this disease in the mainland of China.To better prevent and control such local outbreaks,the epidemiological characteristics and temporal-spatial distribution of overseas imported dengue fever cases in provincial-level administrative divisions(PLADs)where dengue fever is outbreak in the mainland of China were explored.Methods Using the Chinese National Notifiable Infectious Disease Reporting Information System(CNNDS),we identified overseas imported dengue fever cases in dengue fever outbreak areas in the mainland of China from 2005 to 2019 to draw the epidemic curve and population characteristic distribution of overseas imported cases in each PLAD.Based on spatial autocorrelation analysis of ArcGIS 10.5 and temporal-spatial scanning analysis of SaTScan 9.5,we analyzed the temporal-spatial distribution of overseas imported dengue fever in dengue fever outbreak areas in the mainland of China.Results A total of 11,407 imported cases,mainly from Southeast Asia,were recorded from 2005 to 2019 in these 13 PLADs.Of which 62.1%were imported into Yunnan and Guangdong Provinces.Among the imported cases,there were more males than females,mainly from the 21–50 age group.The hot spots were concentrated in parts of Yunnan,Guangdong and Fujian Provinces.We found the cluster of infected areas were expanding northward.Conclusions Based on the analysis of overseas imported dengue cases in 13 PLADs of the mainland of China from 2005 to 2019,we obtained the epidemiological characteristics and spatial distribution of imported dengue cases.Border controls need to pay attention to key population sectors,such as 21–50 years old men and education of key populations on dengue prevention.There is a need to improve the awareness of the prevention and control of imported cases in border areas.At the same time,northern regions cannot relax their vigilance.
基金National Science and Technology Major Project of China(No. 2018ZX10101002-002)。
文摘Background The current prevention and control strategy forAedes albopictus heavily relies on comprehensive management, such as environmental management and chemical control. However, the wide application of pyrethroids has facilitated the development of insecticide resistance, primarily via mutations in the voltage-gated sodium channel (VGSC) gene. This study aims to develop a novel strategy for detecting mutations in the VGSC gene inAe. albopictus using multiplex PCR-mass spectrometry (MPCR-MS) minisequencing technology.Methods We established a new strategy for detecting mutations in the VGSC gene inAe. albopictus using MPCR-MS minisequencing technology. MPCR amplification and mass probe extension (MPE) were first used, followed by single nucleotide polymorphism (SNP) typing mass spectrometry, which allows the simultaneous detection of multiple mutation sites of the VGSC gene in 96 samples ofAe. albopictus. A total of 70 wild-collectedAe. albopictus were used to evaluate the performance of the method by comparing it with other methods.Results Three target sites (1016, 1532, 1534) in the VGSC gene can be detected simultaneously by double PCR amplification combined with matrix-assisted laser desorption ionization-time-of-flight mass spectrometry, achieving a detection limit of 20 fg/μl. We applied this method to 70 wild-collectedAe. albopictus, and the obtained genotypes were consistent with the routine sequencing results, suggesting the accuracy of our method.Conclusions MPCR-MS minisequencing technology provides a sensitive and high-throughput approach toAe. albopictus VGSC gene mutation screening. Compared with conventional sequencing, this method is economical and time-saving. It is of great value for insecticide resistance surveillance in areas with a high risk of vector-borne disease.
基金the National Important Scientific&Technology Project(2018ZX10101002-002)China Prosperity Strategic Programme Fund(SPF)2015-16(15LCI1).
文摘Introduction:Mosquito-borne flavivirus can lead to serious infectious diseases worldwide and cause high mortality and disability.In order to strengthen epidemiological investigation of flavivirus and meet the needs of early warning of diseases,a simple,rapid,and sensitive detection method needs to be established to prevent and control mosquito-borne diseases.Methods:Using the NS5 gene of flavivirus in GenBank,3 universal primers targeting the conserved regions were designed.The complementary DNAs(cDNAs)of Japanese encephalitis virus(JEV),dengue virus(DENV),West Nile virus(WNV),and yellow fever virus(YFV)were used as the template to optimize the reaction conditions.A heminested reversetranscriptase polymerase chain reaction(hnRT-PCR)was established to verify the sensitivity and specificity of this method and to detect field-caught mosquitoes.Results:Our results showed that this method exhibited better specificity,higher sensitivity,and the ability to detect multiple viruses simultaneously.The lowest detection limit of JEV,DENV-2,YFV,and WNV was 3×10^(4),3×10^(6),3×10^(5),and 3×10^(4)copies/μL,respectively.Mosquito-borne flavivirus was successfully detected in the field-caught mosquito samples using the method established in this study.Discussion:The hnRT-PCR method established in this study can be employed for the rapid detection of flavivirus and provide technical support for early and rapid diagnosis of mosquito-borne flavivirus.
