目的:建立Taq M an-MGB探针Real-tim e荧光PCR快速检测单增李斯特菌技术。方法:在对单增李斯特菌invA序列进行分析、比较基础上,设计一对特异性Taq M an-MGB探针法引物及探针,通过Real-tim ePCR反应条件和反应体系的优化,实现对单增李...目的:建立Taq M an-MGB探针Real-tim e荧光PCR快速检测单增李斯特菌技术。方法:在对单增李斯特菌invA序列进行分析、比较基础上,设计一对特异性Taq M an-MGB探针法引物及探针,通过Real-tim ePCR反应条件和反应体系的优化,实现对单增李斯特菌的快速检测;用克隆到pMD18-T载体上的李斯特菌invA基因阳参片段及不同菌株、食品标本验证方法的特异性和敏感性。结果:方法的灵敏性高,其循环阈值与模板浓度的对数值具有很好的对应关系,最低可检测57个拷贝数,经18 h增菌,可检测低至4.5 cfu/m l的细菌;特异性强,检测6株单增李斯特菌标准株和100株单增李斯特菌样品分离株的PCR循环域值(CT值)均小于25,而90株威氏李斯特菌、英诺克李斯特菌、绵羊李斯特菌以及非李斯特菌PCR循环域值(CT值)大于35;快速,最快1 h可出结果,实际样品20 h可出结果,而常规细菌培养法需要一周以上;对103份速冻米面食品进行检测,13份阳性,与常规方法无显著性差异(P>0.05)。结论:Taq M an-MGB探针的单增李斯特菌Real-tim e荧光PCR检测方法具有特异性强,敏感性高,易操作等优点,可推广应用。展开更多
The food additive titanium dioxide is commonly utilized to enhance the appearance and flavor of food products.However,this substance has been linked to gastrointestinal disorders.The aim of this study was to investiga...The food additive titanium dioxide is commonly utilized to enhance the appearance and flavor of food products.However,this substance has been linked to gastrointestinal disorders.The aim of this study was to investigate the impact of dietary exposure of titanium dioxide E171 on Listeria monocytogenes infection in mice.Mice were mainly divided into the control,LM group(L.monocytogenes infection),and E171+LM groups(E171 supplementation before L.monocytogenes infection).Pre-exposure to E171 resulted in increased bacterial counts in the liver,spleen,ileum,colon,mesenteric lymph nodes,and feces of mice after L.monocytogenes infection.Moreover,E171 exposure increased the levels of pro-inflammatory cytokines while attenuating the levels of anti-inflammatory cytokines in mice infected with L.monocytogenes.Meanwhile,mice in the E171+LM group exhibited considerably more severe colonic inflammation and worse intestinal barrier function than mice in the LM group.The 16S rRNA gene sequencing revealed a shift in the composition of the gut microbiota of mice in the E171+LM group,characterized by a decrease in the relative abundance of Firmicutes and a decrease in the Firmicutes-to-Bacteroidetes ratio.The levels of acetate,butyrate,and isobutyrate were markedly decreased within the cecum of mice in the E171+LM group in comparison to mice in the LM group.In conclusion,these results suggest that E171 exposure could exacerbate L.monocytogenes infection in mice,which may provide useful information for future risk assessment of this commonly used food additive.展开更多
文摘目的:建立Taq M an-MGB探针Real-tim e荧光PCR快速检测单增李斯特菌技术。方法:在对单增李斯特菌invA序列进行分析、比较基础上,设计一对特异性Taq M an-MGB探针法引物及探针,通过Real-tim ePCR反应条件和反应体系的优化,实现对单增李斯特菌的快速检测;用克隆到pMD18-T载体上的李斯特菌invA基因阳参片段及不同菌株、食品标本验证方法的特异性和敏感性。结果:方法的灵敏性高,其循环阈值与模板浓度的对数值具有很好的对应关系,最低可检测57个拷贝数,经18 h增菌,可检测低至4.5 cfu/m l的细菌;特异性强,检测6株单增李斯特菌标准株和100株单增李斯特菌样品分离株的PCR循环域值(CT值)均小于25,而90株威氏李斯特菌、英诺克李斯特菌、绵羊李斯特菌以及非李斯特菌PCR循环域值(CT值)大于35;快速,最快1 h可出结果,实际样品20 h可出结果,而常规细菌培养法需要一周以上;对103份速冻米面食品进行检测,13份阳性,与常规方法无显著性差异(P>0.05)。结论:Taq M an-MGB探针的单增李斯特菌Real-tim e荧光PCR检测方法具有特异性强,敏感性高,易操作等优点,可推广应用。
基金the National Key Research and Development Program of China(No.2022YFD2100104)the Science and Technology Program of Shaanxi Province(No.2022JM-111)the Science and Technology Research Program of Liaoning Department of Education(No.J2020044),China.
文摘The food additive titanium dioxide is commonly utilized to enhance the appearance and flavor of food products.However,this substance has been linked to gastrointestinal disorders.The aim of this study was to investigate the impact of dietary exposure of titanium dioxide E171 on Listeria monocytogenes infection in mice.Mice were mainly divided into the control,LM group(L.monocytogenes infection),and E171+LM groups(E171 supplementation before L.monocytogenes infection).Pre-exposure to E171 resulted in increased bacterial counts in the liver,spleen,ileum,colon,mesenteric lymph nodes,and feces of mice after L.monocytogenes infection.Moreover,E171 exposure increased the levels of pro-inflammatory cytokines while attenuating the levels of anti-inflammatory cytokines in mice infected with L.monocytogenes.Meanwhile,mice in the E171+LM group exhibited considerably more severe colonic inflammation and worse intestinal barrier function than mice in the LM group.The 16S rRNA gene sequencing revealed a shift in the composition of the gut microbiota of mice in the E171+LM group,characterized by a decrease in the relative abundance of Firmicutes and a decrease in the Firmicutes-to-Bacteroidetes ratio.The levels of acetate,butyrate,and isobutyrate were markedly decreased within the cecum of mice in the E171+LM group in comparison to mice in the LM group.In conclusion,these results suggest that E171 exposure could exacerbate L.monocytogenes infection in mice,which may provide useful information for future risk assessment of this commonly used food additive.
