Objective:To identify members of genera of rickettsia and O.tsutsugammhi simultaneously.Methods:Rapid and duplex and nested PCR methods have been established by designing primers based on the conserved regions of heat...Objective:To identify members of genera of rickettsia and O.tsutsugammhi simultaneously.Methods:Rapid and duplex and nested PCR methods have been established by designing primers based on the conserved regions of heat shock protein GroEL gene.345 mouse viscera samples including liver,spleen and kidney,96 Xenopsylla cheopis and 32 chiggers collected from Hongta areas of Yuxi city,Yunnan province were tested by the new PCR methods.Results:The result of the study showed that the new PCR methods could identify most members of genera -Rickettsia and Orientia simultaneously with 100%specificity and its sensitivity could test one copy per microliter.The results of detection prevalence of rickettsioses in mouse,flea and mites DNA samples showed that the total rickettsia infection rate in mouse was 34.78%(120/345).The total infection rates in R.typhi,O.t Karp and R.sibirica of mouse samples were 28.12%(97/345),19.71%(68/345) and O. 29%(1/345) respectively.Co-infection rates in R.typhi and 0.t Karp of mouse samples were 13.33%(46/ 345).O.t Karp type has been the main epidemic strain in these areas.Conclusion:We concluded that this PCR method could be used to detect multi-genera rickettsia simultaneously.Molecular evidences provided in this and previous studies strongly support that Hongta areas of Yuxi city are a natural focus for typhus and scrub typhus with the common occurrence of their confection.展开更多
目的利用环介导等温扩增(LAMP)可视化检测技术,建立一种灵敏、特异和简便的恙虫病检测方法。方法利用LAMP在线引物设计软件Primer Explorer V4,以恙虫病东方体热休克蛋白groEL基因作为靶序列,设计扩增引物,优化反应条件。利用恙虫病东...目的利用环介导等温扩增(LAMP)可视化检测技术,建立一种灵敏、特异和简便的恙虫病检测方法。方法利用LAMP在线引物设计软件Primer Explorer V4,以恙虫病东方体热休克蛋白groEL基因作为靶序列,设计扩增引物,优化反应条件。利用恙虫病东方体标准株评估灵敏度和特异度;收集42份恙虫病血样,分别采用LAMP、普通PCR和实时荧光定量PCR方法进行验证。结果建立的LAMP方法可在61~65℃80 min内完成恙虫病东方体的快速检测,最低检测限为10拷贝/μL,高于普通PCR方法2个数量级,高于荧光定量PCR方法1个数量级;对8种常见的自然疫源性疾病病原体检测均为阴性,特异度为100%;对42份恙虫病血样采用3种方法检测,阳性率一致,均为21.4%(9/42)。结论本研究建立的恙虫病东方体LAMP可视化检测反应快速,操作简便,灵敏特异,可在基层医疗卫生机构运用推广。展开更多
Salmonella infection, ranging from mild, self-limiting diarrhea to severe gastrointestinal, septicemic disease and enteric fever, is a global health problem both in humans and animals. Rapid development of microbial d...Salmonella infection, ranging from mild, self-limiting diarrhea to severe gastrointestinal, septicemic disease and enteric fever, is a global health problem both in humans and animals. Rapid development of microbial drug resistance has led to a need for efficacious and affordable vaccines against Salmonella. Microbial heat shock proteins （HSPs）, including HSP60 and HSP70, are the dominant antigens that promote the host immune response. Co-administration of these antigens with cytokines, such as IL-22, which plays an important role in antimicrobial defense, can enhance the immune response and protection against pathogens. Therefore, the aim of the present study was to determine the immunogenicity of rGroEL （Hsp60） of S. Typhi, alone or administered in combination with murine rlL-22, and its protective efficacy against lethal infection with Salmonella, in mice. There was appreciable stimulation of the humoral and cell-mediated immune responses in mice immunized with rGroEL alone. However, co-administration of rGroEL with rlL-22 further boosted the antibody titers （IgG, IgG1 and IgG2a）, T-cell proliferative responses and the secretion of both Thl and Th2 cytokines. Additionally, rGroEL alone accorded 65%-70% protection against lethal challenge with S. Typhi and S. Typhimurium, which increased to 90% when co-administered with rlL-22.展开更多
基金supported by National Natural Science Foundation of China(No. 30771854)China-U.S.Collaborative Program on Emerging and Re-emerging Infectious Diseases(No. 1U2GGH000018-01)
文摘Objective:To identify members of genera of rickettsia and O.tsutsugammhi simultaneously.Methods:Rapid and duplex and nested PCR methods have been established by designing primers based on the conserved regions of heat shock protein GroEL gene.345 mouse viscera samples including liver,spleen and kidney,96 Xenopsylla cheopis and 32 chiggers collected from Hongta areas of Yuxi city,Yunnan province were tested by the new PCR methods.Results:The result of the study showed that the new PCR methods could identify most members of genera -Rickettsia and Orientia simultaneously with 100%specificity and its sensitivity could test one copy per microliter.The results of detection prevalence of rickettsioses in mouse,flea and mites DNA samples showed that the total rickettsia infection rate in mouse was 34.78%(120/345).The total infection rates in R.typhi,O.t Karp and R.sibirica of mouse samples were 28.12%(97/345),19.71%(68/345) and O. 29%(1/345) respectively.Co-infection rates in R.typhi and 0.t Karp of mouse samples were 13.33%(46/ 345).O.t Karp type has been the main epidemic strain in these areas.Conclusion:We concluded that this PCR method could be used to detect multi-genera rickettsia simultaneously.Molecular evidences provided in this and previous studies strongly support that Hongta areas of Yuxi city are a natural focus for typhus and scrub typhus with the common occurrence of their confection.
文摘目的利用环介导等温扩增(LAMP)可视化检测技术,建立一种灵敏、特异和简便的恙虫病检测方法。方法利用LAMP在线引物设计软件Primer Explorer V4,以恙虫病东方体热休克蛋白groEL基因作为靶序列,设计扩增引物,优化反应条件。利用恙虫病东方体标准株评估灵敏度和特异度;收集42份恙虫病血样,分别采用LAMP、普通PCR和实时荧光定量PCR方法进行验证。结果建立的LAMP方法可在61~65℃80 min内完成恙虫病东方体的快速检测,最低检测限为10拷贝/μL,高于普通PCR方法2个数量级,高于荧光定量PCR方法1个数量级;对8种常见的自然疫源性疾病病原体检测均为阴性,特异度为100%;对42份恙虫病血样采用3种方法检测,阳性率一致,均为21.4%(9/42)。结论本研究建立的恙虫病东方体LAMP可视化检测反应快速,操作简便,灵敏特异,可在基层医疗卫生机构运用推广。
文摘Salmonella infection, ranging from mild, self-limiting diarrhea to severe gastrointestinal, septicemic disease and enteric fever, is a global health problem both in humans and animals. Rapid development of microbial drug resistance has led to a need for efficacious and affordable vaccines against Salmonella. Microbial heat shock proteins （HSPs）, including HSP60 and HSP70, are the dominant antigens that promote the host immune response. Co-administration of these antigens with cytokines, such as IL-22, which plays an important role in antimicrobial defense, can enhance the immune response and protection against pathogens. Therefore, the aim of the present study was to determine the immunogenicity of rGroEL （Hsp60） of S. Typhi, alone or administered in combination with murine rlL-22, and its protective efficacy against lethal infection with Salmonella, in mice. There was appreciable stimulation of the humoral and cell-mediated immune responses in mice immunized with rGroEL alone. However, co-administration of rGroEL with rlL-22 further boosted the antibody titers （IgG, IgG1 and IgG2a）, T-cell proliferative responses and the secretion of both Thl and Th2 cytokines. Additionally, rGroEL alone accorded 65%-70% protection against lethal challenge with S. Typhi and S. Typhimurium, which increased to 90% when co-administered with rlL-22.
