HighlightsA novel conjugative plasmid pHJ90-cfr carrying the multiresistance gene cfr was characterized in Proteus vulgaris.A new IS5-family member,ISPmi4,was identified for the first time.Both plasmids and ICEs were ...HighlightsA novel conjugative plasmid pHJ90-cfr carrying the multiresistance gene cfr was characterized in Proteus vulgaris.A new IS5-family member,ISPmi4,was identified for the first time.Both plasmids and ICEs were vital mobile genetic elements for horizontal transmission of cfr gene in Proteus species.展开更多
The escalating global dissemination of plasmid-mediated antibiotic resistance poses a formidable threat to global health.Conjugation stands as a pivotal mechanism for horizontal gene transfer among bacterial populatio...The escalating global dissemination of plasmid-mediated antibiotic resistance poses a formidable threat to global health.Conjugation stands as a pivotal mechanism for horizontal gene transfer among bacterial populations,facilitating the spread of antibiotic resistance genes(ARGs).Microelectrolysis has garnered attention as an efficacious strategy for mitigating antibiotic concentrations in wastewater,yet its potential impact on ARG horizontal transfer remain largely unexplored.This comprehensive investigation unveils that microelectrolysis not only influences but significantly accelerates the conjugative transfer of ARG-harboring plasmids.Remarkably,this phenomenon is corroborated at the microbial community scale,underscoring its ecological relevance.Alarmingly,the study highlights the vulnerability of intestinalmicroorganisms to acquire antibiotic resistance under electrolytic stimulation,posing heightened risks to both animal and human health.Delving deeper,the study elucidates the underlyingmechanisms responsible for this enhanced conjugative transfer.It reveals that microelectrolysis augments the abundance of mating-competent cells,triggers the generation of reactive oxygen species,inflicts modest membrane damage,and upregulates the expression of genes critical for conjugation.These findings collectively contribute to a more profound comprehension of the environmental dissemination dynamics and associated public health implications of ARGs in the context of wastewater treatment employing microelectrolytic technologies.展开更多
The objective of this study is to quantitatively reveal the main genetic carrier of antibiotic resistance genes(ARGs)for blocking their environmental dissemination.The distribution of ARGs in chromosomes,plasmids,and ...The objective of this study is to quantitatively reveal the main genetic carrier of antibiotic resistance genes(ARGs)for blocking their environmental dissemination.The distribution of ARGs in chromosomes,plasmids,and phages for understanding their respective contributions to the development of antimicrobial resistance in aerobic biofilm consortium under increasing stresses of oxytetracycline,streptomycin,and tigecyclinewere revealed based on metagenomics analysis.Results showed that the plasmids harbored 49.2%-83.9%of resistomes,which was higher(p<0.001)than chromosomes(2.0%-35.6%),and no ARGs were detected in phage contigs under the strict alignment standard of over 80%identity used in this study.Plasmids and chromosomes tended to encode different types of ARGs,whose abundances all increased with the hike of antibiotic concentrations,and the variety of ARGs encoded by plasmids(14 types and 64 subtypes)was higher than that(11 types and 27 subtypes)of chromosomes.The dosing of the three antibiotics facilitated the transposition and recombination of ARGs on plasmids,mediated by transposable and integrable transfer elements,which increased the co-occurrence of associated and unassociated ARGs.The results quantitatively proved that plasmids dominate the proliferation of ARGs in aerobic biofilm driven by antibiotic selection,which should be a key target for blocking ARG dissemination.展开更多
Herpes simplex virus thymidine kinase(HSVtk)gene therapy is a promising strategy for glioblastoma therapy.However,delivery of plasmid DNA(pDNA)encoding HSVtk into the brain by systemic administration is a challenge si...Herpes simplex virus thymidine kinase(HSVtk)gene therapy is a promising strategy for glioblastoma therapy.However,delivery of plasmid DNA(pDNA)encoding HSVtk into the brain by systemic administration is a challenge since pDNA can hardly penetrate the bloodbrain barrier.In this study,an exosome-membrane(EM)and polymer-based hybrid complex was developed for systemic delivery of pDNA into the brain.Histidine/arginine-linked polyamidoamine(PHR)was used as a carrier.PHR binds to pDNA by electrostatic interaction.The pDNA/PHR complex was mixed with EM and subjected to extrusion to produce pDNA/PHR-EM hybrid complex.For glioblastoma targeting,T7 peptide was attached to the pDNA/PHR-EM complex.Both pDNA/PHR-EM and T7-decorated pDNA/PHR-EM(pDNA/PHREM-T7)had a surface charge of–5 mV and a size of 280 nm.Transfection assays indicated that pDNA/PHR-EM-T7 enhanced the transfection to C6 cells compared with pDNA/PHREM.Intravenous administration of pHSVtk/PHR-EM-T7 showed that pHSVtk/PHR-EM and pHSVtk/PHR-EM-T7 delivered pHSVtk more efficiently than pHSVtk/lipofectamine and pHSVtk/PHR into glioblastoma in vivo.pHSVtk/PHR-EM-T7 had higher delivery efficiency than pHSVtk/PHR-EM.As a result,the HSVtk expression and apoptosis levels in the tumors of the pHSVtk/PHR-EM-T7 group were higher than those of the other control groups.Therefore,the pDNA/PHR-EM-T7 hybrid complex is a useful carrier for systemic delivery of pHSVtk to glioblastoma.展开更多
The inappropriate use of cephalosporins lead to the occurrence and global spread of bacteria resistant to these antimicrobials.In this study,we isolated four Escherichia albertii strains from broilers in eastern China...The inappropriate use of cephalosporins lead to the occurrence and global spread of bacteria resistant to these antimicrobials.In this study,we isolated four Escherichia albertii strains from broilers in eastern China.The antimicrobial susceptibility and genomic characterization of these E.albertii isolates were determined.Our results revealed that these four E.albertii isolates exhibited resistance to tetracyclines,chloramphenicol,β-lactams,aminoglycosides,polymyxin B,sulfonamides,quinolones,and other antimicrobials.Among them,EA04 isolate was multidrug resistant and harbored extended-spectrumβ-lactamases(ESBL)genes blaCTX-Mand blaTEM.Whole genome sequencing and core-genome multilocus sequence typing(cgMLST)based on all ST4638 E.albertii for EA04 inferred highly probable epidemiological links between selected human isolates.Additionally,the ESBL genes blaTEM-141and blaCTX-M-55were coexistent in an approximately 75 kb Inc FII plasmid pEA04.2 in EA04.Comparative analysis indicated that genes blaTEM-141and blaCTX-M-55were located in IS15-blaCTX-M-55-wbu C-blaTEM-141-IS26 region,which similar structures were identified in various bacteria.Furthermore,the plasmid pEA04.2 could be transferable to E.coli EC600 and lead to the resistance to third-generation cephalosporins.These results suggested that chicken potentially serve as a reservoir for multidrug resistant E.albertii,which increases the risk of horizontal transfer of antimicrobial resistance between humans,animals and environment.展开更多
Objective Our study aimed to conduct genomic characterization of Salmonella strains carrying the blaNDM-1 gene in the intestinal tract of healthy individuals.The objectives were to underscore the importance of genomic...Objective Our study aimed to conduct genomic characterization of Salmonella strains carrying the blaNDM-1 gene in the intestinal tract of healthy individuals.The objectives were to underscore the importance of genomic surveillance for drug resistance in both commensal and pathogenic bacteria among healthy populations,and to establish protocols for regulating drug resistance plasmids based on the completion of a comprehensive map of drug resistance plasmid genomes.Methods We performed antimicrobial susceptibility testing and employed second-and third-generation sequencing techniques to analyze Salmonella strains harboring the blaNDM-1 gene,to surveil drug-resistant bacteria in the intestines of healthy subjects.Sequence comparison was conducted using both core-and pan-genome approaches.Concurrently,conjugation experiments were carried out to assess the efficiency of plasmid transfer.Results We isolated a carbapenem-resistant Salmonella enterica serovar Typhimurium strain from a healthy food worker in China.This strain harbored an IncHI2/IncHI2A plasmid carrying blaNDM-1 along with multiple antibiotic resistance genes(ARGs).Our findings highlight the potential for asymptomatic carriers to facilitate the transmission of ARGs.Pan-genomic analysis revealed that blaNDM-1-positive plasmids could traverse bacterial species barriers,facilitating cross-host transmission.Conclusion This study marks the first detection of blaNDM-1 in Salmonella strains isolated from healthy individuals.We underscore the risk associated with the transmission of conjugative hybrid plasmids carrying blaNDM-1,which have the potential to be harbored and transmitted among healthy individuals.Enhanced surveillance of drug-resistant pathogens and plasmids in the intestinal microbiota of healthy individuals could provide insights into the risk of ARG transmission and pathways for population-wide dissemination via ARG transfer factors.展开更多
目的建立可检测早期区域1A(early region 1A,E1A)蛋白和猿猴病毒40大T抗原(simian virus 40 large T anti-gen,SV40LTA)残留DNA的质粒参考品,并探索基于数字PCR法对质粒参考品拷贝数标定的分析技术,从而准确定量质粒参考品,为生物制品...目的建立可检测早期区域1A(early region 1A,E1A)蛋白和猿猴病毒40大T抗原(simian virus 40 large T anti-gen,SV40LTA)残留DNA的质粒参考品,并探索基于数字PCR法对质粒参考品拷贝数标定的分析技术,从而准确定量质粒参考品,为生物制品中宿主DNA残留检测提供新的思路。方法构建质粒pUC19-E1A-SV40LTA,全基因合成后进行扩增,以得到足够的质粒参考品;采用数字PCR仪,分别用E1A和SV40LTA引物探针体系的双荧光通道对其进行拷贝数检测;将数字PCR标定拷贝数的参考品应用于qPCR中,建立检测E1A和SV40LTA特定序列的方法。对建立的qPCR体系进行质粒参考品线性、准确性、定量限、精密性、特异性、适用性、稳定性验证,并应用建立的qPCR体系对重组腺病毒(recombinant adenovirus,rAdV)、重组腺相关病毒(recombinant adeno-associated virus,rAAV)样本进行检测。结果用数字PCR双荧光通道检测质料参考品的E1A和SV40LTA靶点的拷贝数基本一致,其中用E1A引物探针体系检测值为3.55×10^(9)copies/μL,SV40LTA引物探针体系检测值为3.48×10^(9)copies/μL,这两种检测值的变异系数(CV)为1.42%,取平均值3.51×10^(9)copies/μL作为数字PCR的标定值。用数字PCR标定拷贝数建立的qPCR体系线性、准确性、定量限、精密性、特异性、适用性、稳定性良好,检测rAdV、rAAV样本的回收率均在70%130%之间。结论建立了可检测生物制品中E1A和SV40LTA残留的质粒参考品,并引入了更为灵敏和准确的数字PCR方法对其进行定量。该参考品可应用于以HEK293、HEK293T为宿主细胞生产的基因治疗产品中E1A和SV40LTA残留DNA检测。展开更多
[Objective] The pathogenic Escherichia coli in musk deer was classified at molecular level to provide basic materials for molecular epidemiology of pathogenic Escherichia coli in musk deer. [Method] Plasmids from 24 p...[Objective] The pathogenic Escherichia coli in musk deer was classified at molecular level to provide basic materials for molecular epidemiology of pathogenic Escherichia coli in musk deer. [Method] Plasmids from 24 pathogenic Escherichia coli in musk deer were extracted by the Lysis Triton method, and then identified by single enzyme digestion with three endonucleases of Hind Ⅲ, EcoR Ⅰ and BamH Ⅰ. [Result] The yield rate of plasmids was 91.6%, and 24 pathogenic Escherichia coli in musk deer had the identical or similar plasmid profiles. [Conclusion] Plasmid DNA analysis offers scientific basis for molecular epidemiology of pathogenic Escherichia coli in musk deer in Sichuan Institute of Musk Deer Breeding.展开更多
Expression of rol genes from Ri plasmid of Agrobacterium rhizogenes not only leads to the excessive formation of adventitious roots, but also exhibits various genetically modified characteristics that have bro...Expression of rol genes from Ri plasmid of Agrobacterium rhizogenes not only leads to the excessive formation of adventitious roots, but also exhibits various genetically modified characteristics that have broad prospects for the application of plant genetic improvement. Since the 1980s of the last century, much progress has been made in the studies of A. rhizogenes, in particular the agropine type Ri plasmid rol genes and their applications for plant genetic improvement, which involves the structure and function of Ri plasmid, the characters of rol genes, the influence of rol genes expression on plants growth and development, and the applications of rol genes for genetic improvement of forest tree. In this paper, the advances in this field are reviewed and the existing problems about the application of rol genes for genetic improvement of forest tree are also discussed.展开更多
基金supported by the National Key Research and Development Program of China(2022YFF0710505)the Central Public-interest Scientific Institution Basal Research Fund,China(1610302022001)。
文摘HighlightsA novel conjugative plasmid pHJ90-cfr carrying the multiresistance gene cfr was characterized in Proteus vulgaris.A new IS5-family member,ISPmi4,was identified for the first time.Both plasmids and ICEs were vital mobile genetic elements for horizontal transmission of cfr gene in Proteus species.
基金supported by Jiangsu Agriculture Science and Technology Innovation Fund(No.CX(22)3001)。
文摘The escalating global dissemination of plasmid-mediated antibiotic resistance poses a formidable threat to global health.Conjugation stands as a pivotal mechanism for horizontal gene transfer among bacterial populations,facilitating the spread of antibiotic resistance genes(ARGs).Microelectrolysis has garnered attention as an efficacious strategy for mitigating antibiotic concentrations in wastewater,yet its potential impact on ARG horizontal transfer remain largely unexplored.This comprehensive investigation unveils that microelectrolysis not only influences but significantly accelerates the conjugative transfer of ARG-harboring plasmids.Remarkably,this phenomenon is corroborated at the microbial community scale,underscoring its ecological relevance.Alarmingly,the study highlights the vulnerability of intestinalmicroorganisms to acquire antibiotic resistance under electrolytic stimulation,posing heightened risks to both animal and human health.Delving deeper,the study elucidates the underlyingmechanisms responsible for this enhanced conjugative transfer.It reveals that microelectrolysis augments the abundance of mating-competent cells,triggers the generation of reactive oxygen species,inflicts modest membrane damage,and upregulates the expression of genes critical for conjugation.These findings collectively contribute to a more profound comprehension of the environmental dissemination dynamics and associated public health implications of ARGs in the context of wastewater treatment employing microelectrolytic technologies.
基金supported by the National Natural Science Foundation of China(Nos.52091545 and 51978645).
文摘The objective of this study is to quantitatively reveal the main genetic carrier of antibiotic resistance genes(ARGs)for blocking their environmental dissemination.The distribution of ARGs in chromosomes,plasmids,and phages for understanding their respective contributions to the development of antimicrobial resistance in aerobic biofilm consortium under increasing stresses of oxytetracycline,streptomycin,and tigecyclinewere revealed based on metagenomics analysis.Results showed that the plasmids harbored 49.2%-83.9%of resistomes,which was higher(p<0.001)than chromosomes(2.0%-35.6%),and no ARGs were detected in phage contigs under the strict alignment standard of over 80%identity used in this study.Plasmids and chromosomes tended to encode different types of ARGs,whose abundances all increased with the hike of antibiotic concentrations,and the variety of ARGs encoded by plasmids(14 types and 64 subtypes)was higher than that(11 types and 27 subtypes)of chromosomes.The dosing of the three antibiotics facilitated the transposition and recombination of ARGs on plasmids,mediated by transposable and integrable transfer elements,which increased the co-occurrence of associated and unassociated ARGs.The results quantitatively proved that plasmids dominate the proliferation of ARGs in aerobic biofilm driven by antibiotic selection,which should be a key target for blocking ARG dissemination.
基金supported by the Individual Basic Science&Engineering Research Program(NRF-2022R1A2B5B01001920)through the National Research Foundation,funded by the Ministry of Science and ICT in Korea.
文摘Herpes simplex virus thymidine kinase(HSVtk)gene therapy is a promising strategy for glioblastoma therapy.However,delivery of plasmid DNA(pDNA)encoding HSVtk into the brain by systemic administration is a challenge since pDNA can hardly penetrate the bloodbrain barrier.In this study,an exosome-membrane(EM)and polymer-based hybrid complex was developed for systemic delivery of pDNA into the brain.Histidine/arginine-linked polyamidoamine(PHR)was used as a carrier.PHR binds to pDNA by electrostatic interaction.The pDNA/PHR complex was mixed with EM and subjected to extrusion to produce pDNA/PHR-EM hybrid complex.For glioblastoma targeting,T7 peptide was attached to the pDNA/PHR-EM complex.Both pDNA/PHR-EM and T7-decorated pDNA/PHR-EM(pDNA/PHREM-T7)had a surface charge of–5 mV and a size of 280 nm.Transfection assays indicated that pDNA/PHR-EM-T7 enhanced the transfection to C6 cells compared with pDNA/PHREM.Intravenous administration of pHSVtk/PHR-EM-T7 showed that pHSVtk/PHR-EM and pHSVtk/PHR-EM-T7 delivered pHSVtk more efficiently than pHSVtk/lipofectamine and pHSVtk/PHR into glioblastoma in vivo.pHSVtk/PHR-EM-T7 had higher delivery efficiency than pHSVtk/PHR-EM.As a result,the HSVtk expression and apoptosis levels in the tumors of the pHSVtk/PHR-EM-T7 group were higher than those of the other control groups.Therefore,the pDNA/PHR-EM-T7 hybrid complex is a useful carrier for systemic delivery of pHSVtk to glioblastoma.
基金supported by the National Key Research and Development Program of China(2021YFD1800402)the National Natural Science Foundation of China(32172856,31972654 and 32302881)+1 种基金the Natural Science Foundation of Shanghai,China(22ZR1476100 and 23ZR1476600)the Agricultural Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciences(SHVRI-ASTIP-2014-8)。
文摘The inappropriate use of cephalosporins lead to the occurrence and global spread of bacteria resistant to these antimicrobials.In this study,we isolated four Escherichia albertii strains from broilers in eastern China.The antimicrobial susceptibility and genomic characterization of these E.albertii isolates were determined.Our results revealed that these four E.albertii isolates exhibited resistance to tetracyclines,chloramphenicol,β-lactams,aminoglycosides,polymyxin B,sulfonamides,quinolones,and other antimicrobials.Among them,EA04 isolate was multidrug resistant and harbored extended-spectrumβ-lactamases(ESBL)genes blaCTX-Mand blaTEM.Whole genome sequencing and core-genome multilocus sequence typing(cgMLST)based on all ST4638 E.albertii for EA04 inferred highly probable epidemiological links between selected human isolates.Additionally,the ESBL genes blaTEM-141and blaCTX-M-55were coexistent in an approximately 75 kb Inc FII plasmid pEA04.2 in EA04.Comparative analysis indicated that genes blaTEM-141and blaCTX-M-55were located in IS15-blaCTX-M-55-wbu C-blaTEM-141-IS26 region,which similar structures were identified in various bacteria.Furthermore,the plasmid pEA04.2 could be transferable to E.coli EC600 and lead to the resistance to third-generation cephalosporins.These results suggested that chicken potentially serve as a reservoir for multidrug resistant E.albertii,which increases the risk of horizontal transfer of antimicrobial resistance between humans,animals and environment.
基金supported by the National Key Research and Development Program of China(2022YFC2303900)the major projects of the National Natural Science Foundation of China(22193064)the Science Foundation(2022SKLID303)of the State Key Laboratory of Infectious Disease Prevention and Control,China。
文摘Objective Our study aimed to conduct genomic characterization of Salmonella strains carrying the blaNDM-1 gene in the intestinal tract of healthy individuals.The objectives were to underscore the importance of genomic surveillance for drug resistance in both commensal and pathogenic bacteria among healthy populations,and to establish protocols for regulating drug resistance plasmids based on the completion of a comprehensive map of drug resistance plasmid genomes.Methods We performed antimicrobial susceptibility testing and employed second-and third-generation sequencing techniques to analyze Salmonella strains harboring the blaNDM-1 gene,to surveil drug-resistant bacteria in the intestines of healthy subjects.Sequence comparison was conducted using both core-and pan-genome approaches.Concurrently,conjugation experiments were carried out to assess the efficiency of plasmid transfer.Results We isolated a carbapenem-resistant Salmonella enterica serovar Typhimurium strain from a healthy food worker in China.This strain harbored an IncHI2/IncHI2A plasmid carrying blaNDM-1 along with multiple antibiotic resistance genes(ARGs).Our findings highlight the potential for asymptomatic carriers to facilitate the transmission of ARGs.Pan-genomic analysis revealed that blaNDM-1-positive plasmids could traverse bacterial species barriers,facilitating cross-host transmission.Conclusion This study marks the first detection of blaNDM-1 in Salmonella strains isolated from healthy individuals.We underscore the risk associated with the transmission of conjugative hybrid plasmids carrying blaNDM-1,which have the potential to be harbored and transmitted among healthy individuals.Enhanced surveillance of drug-resistant pathogens and plasmids in the intestinal microbiota of healthy individuals could provide insights into the risk of ARG transmission and pathways for population-wide dissemination via ARG transfer factors.
文摘目的建立可检测早期区域1A(early region 1A,E1A)蛋白和猿猴病毒40大T抗原(simian virus 40 large T anti-gen,SV40LTA)残留DNA的质粒参考品,并探索基于数字PCR法对质粒参考品拷贝数标定的分析技术,从而准确定量质粒参考品,为生物制品中宿主DNA残留检测提供新的思路。方法构建质粒pUC19-E1A-SV40LTA,全基因合成后进行扩增,以得到足够的质粒参考品;采用数字PCR仪,分别用E1A和SV40LTA引物探针体系的双荧光通道对其进行拷贝数检测;将数字PCR标定拷贝数的参考品应用于qPCR中,建立检测E1A和SV40LTA特定序列的方法。对建立的qPCR体系进行质粒参考品线性、准确性、定量限、精密性、特异性、适用性、稳定性验证,并应用建立的qPCR体系对重组腺病毒(recombinant adenovirus,rAdV)、重组腺相关病毒(recombinant adeno-associated virus,rAAV)样本进行检测。结果用数字PCR双荧光通道检测质料参考品的E1A和SV40LTA靶点的拷贝数基本一致,其中用E1A引物探针体系检测值为3.55×10^(9)copies/μL,SV40LTA引物探针体系检测值为3.48×10^(9)copies/μL,这两种检测值的变异系数(CV)为1.42%,取平均值3.51×10^(9)copies/μL作为数字PCR的标定值。用数字PCR标定拷贝数建立的qPCR体系线性、准确性、定量限、精密性、特异性、适用性、稳定性良好,检测rAdV、rAAV样本的回收率均在70%130%之间。结论建立了可检测生物制品中E1A和SV40LTA残留的质粒参考品,并引入了更为灵敏和准确的数字PCR方法对其进行定量。该参考品可应用于以HEK293、HEK293T为宿主细胞生产的基因治疗产品中E1A和SV40LTA残留DNA检测。
基金Supported by Youth Foundation of Education Department in Sichuan Province (07ZB060)Youth Science and Technology Innovation Fund in Sichuan Agricultural University~~
文摘[Objective] The pathogenic Escherichia coli in musk deer was classified at molecular level to provide basic materials for molecular epidemiology of pathogenic Escherichia coli in musk deer. [Method] Plasmids from 24 pathogenic Escherichia coli in musk deer were extracted by the Lysis Triton method, and then identified by single enzyme digestion with three endonucleases of Hind Ⅲ, EcoR Ⅰ and BamH Ⅰ. [Result] The yield rate of plasmids was 91.6%, and 24 pathogenic Escherichia coli in musk deer had the identical or similar plasmid profiles. [Conclusion] Plasmid DNA analysis offers scientific basis for molecular epidemiology of pathogenic Escherichia coli in musk deer in Sichuan Institute of Musk Deer Breeding.
文摘Expression of rol genes from Ri plasmid of Agrobacterium rhizogenes not only leads to the excessive formation of adventitious roots, but also exhibits various genetically modified characteristics that have broad prospects for the application of plant genetic improvement. Since the 1980s of the last century, much progress has been made in the studies of A. rhizogenes, in particular the agropine type Ri plasmid rol genes and their applications for plant genetic improvement, which involves the structure and function of Ri plasmid, the characters of rol genes, the influence of rol genes expression on plants growth and development, and the applications of rol genes for genetic improvement of forest tree. In this paper, the advances in this field are reviewed and the existing problems about the application of rol genes for genetic improvement of forest tree are also discussed.
