谷氨酸脱羧酶lmo2363基因缺失对单增李斯特菌致病性的影响  

Effect of deletion of glutamate decarboxylase lmo2363 gene on pathogenicity of Listeria monocytogenes

作　　者：芝吉 曹青 赵学慧 张浩浩 范子秋 马永辉 邓静 何曾文 马金锐 张坤中 崇倩 薛惠文[1] 苟惠天[1] ZHI Ji;CAO Qing;ZHAO Xuehui;ZHANG Haohao;FAN Ziqiu;MA Yonghui;DENG Jing;HE Zengwen;MA Jinrui;ZHANG Kunzhong;CHONG Qian;XUE Huiwen;GOU Huitian(College of Veterinary Medicine,Gansu Agricultural University,Lanzhou 730070,China)

机构地区：[1]甘肃农业大学动物医学院,甘肃兰州730070

出　　处：《畜牧与兽医》2025年第4期93-100,共8页Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金项目(31960726,32060822,31560700);国家重点研发计划项目(2019YFC1605705);甘肃农业大学青年导师扶持基金项目(GAU-QDFC-2020-10);甘肃省重点研发计划项目(20YF8FA136);张家川揭榜挂帅项目(ZC-STK-2023A-030)。

摘　　要：旨在探究谷氨酸脱羧酶lmo2363基因缺失对单增李斯特菌(Listeria monocytogenes,LM)致病性的影响。以LM参考株ATCC 19111为对照,用荧光显微镜观察分离株LM83-1、缺失株LM83-1Δlmo2363、回补株CLM83-1Δlmo2363的生物被膜形成;采用人结肠腺癌细胞Caco-2和小鼠巨噬细胞RAW264.7进行黏附侵袭和胞内增殖试验;对LM83-1和LM83-1Δlmo2363转录组测序分析,筛选两者之间的差异表达基因,对差异表达基因进行GO功能富集分析和KEGG代谢通路富集分析。结果:与LM83-1相比,在培养24、48 h时,LM83-1Δlmo2363的生物被膜结构疏松,形成量下降,CLM83-1Δlmo2363在各时间段生物被膜形成与LM83-1相同;LM83-1Δlmo2363对Caco-2细胞的黏附率增强,侵袭率下降,CLM83-1Δlmo2363对Caco-2细胞的黏附率和侵袭率与LM83-1无显著差异;在4、8、12、24 h时,LM83-1Δlmo2363在巨噬细胞胞内增殖能力减弱,在4 h时CLM83-1Δlmo2363在巨噬细胞胞内增殖能力减弱;转录组测序显示,与LM83-1相比,缺失株LM83-1Δlmo2363中共有136个基因表达差异显著,其中上调基因36个,下调基因100个;GO功能富集分析显示,差异表达基因主要富集在乙醇胺解胺酶活性、细菌型鞭毛基体、乙醇胺分解代谢过程、铁离子稳态等GO条目中;KEGG代谢通路富集分析显示,差异表达基因主要富集在磷酸转移酶系统、淀粉和蔗糖代谢、鞭毛组装等通路中。本研究表明,谷氨酸脱羧酶lmo2363基因可能通过参与铁稳态、鞭毛组装、磷酸转移酶系统等生物学过程在LM生物被膜形成和体外细胞感染中发挥重要作用,研究结果为进一步探究谷氨酸脱羧酶的功能奠定基础。The aim of this study was to investigate the effect of the deletion of glutamate decarboxylase lmo2363 gene on the pathogenicity of Listeria monocytogenes.In here,the reference strain of Listeria monocytogenes ATCC 19111 was used as the control.The biofilm formation of LM83-1,LM83-1Δlmo2363 and CLM83-1Δlmo2363 was observed by fluorescence microscopy.Caco-2 cells and RAW264.7 cells were used for adhesion,invasion and intracellular proliferation tests of the above bacterium.Transcriptome sequencing of LM83-1 and LM83-1Δlmo2363 was performed to screen the differentially expressed genes between them,and GO functional enrichment analysis and KEGG met⁃abolic pathway enrichment analysis were performed for differentially expressed genes.The results showed that,compared with LM83-1,the biofilm structure of LM83-1Δlmo2363 decreased and the biofilm formation of CLM83-1Δlmo2363 was the same as that of LM83-1 at 24 h and 48 h.The adhesion rate of LM83-1Δlmo2363 to Caco-2 cells was enhanced,and the invasion rate was decreased.The adhesion rate and invasion rate of CLM83-1Δlmo2363 to Caco-2 cells were not significantly different from those of LM83-1.The proliferation of LM83-1Δlmo2363 in macrophages decreased at 4,8,12 and 24 h,and the proliferation of CLM83-1Δlmo2363 in macrophages decreased at 4 h.Transcriptome sequencing showed that 136 genes of the deletion strain LM83-1Δlmo2363 were significantly different from those of LM83-1,including 36 up-regulated genes and 100 down-regulated genes.The GO functional enrichment analysis showed that the differentially ex⁃pressed genes were mainly concentrated in GO items such as ethanolamine lysozyme activity,bacterial flagella substrate,ethanolamine catab⁃olism,iron ion homeostasis,etc.The enrichment analysis of the KEGG metabolic pathways showed that the differentially expressed genes were mainly concentrated in the phosphotransferase system,starch and sucrose metabolism,and flagella assembly.These results suggested that the glutamate decarboxylase lmo2363 gene may pla

关 键 词：单增李斯特菌 谷氨酸脱羧酶 生物被膜 体外细胞感染 转录组测序 

分 类 号：S852.612[农业科学—基础兽医学]