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目的单核细胞增生性李斯特菌(Listeria monocytogenes,Lm)对宿主的粘附、侵袭作用与其多种内化素蛋白密切相关。本研究以4b型菌株LmNTSN的內化素基因NTSN_0462为研究对象,初步探究其致病作用。方法利用同源重组技术构建该基因缺失的突变株,研究0462基因对NTSN在生长、细胞侵袭和体内定植中的作用。人结肠腺癌细胞Caco-2、人肝癌细胞HepG2的侵袭率,以及BALB/c小鼠体内肝、脾脏中定植能力的差异。结果缺失株的生长曲线结果显示,NTSN_0462基因对Lm在BHI培养基中的生长及代谢未造成明显影响。以人结肠腺癌细胞Caco-2、人肝癌细胞HepG2进行的体外试验表明,缺失株的侵袭能力低于野生株(P<0.001)。以BALB/c小鼠进行的体内试验显示,缺失株在肝脏中定殖的能力显著低于野生株(P<0.001),在脾脏中无统计学差异。结论 NTSN_0462基因在LmNTSN入侵肝脏和定植中发挥重要作用,是侵袭相关的重要毒力因子。
Aim The adhesion and invasion of Listeria monocytogenes (Lm) to its host are closely related to its multiple endothelin proteins. In this study, the endonuclease gene NTSN_0462 of LmNTSN strain 4b was used as the research object to investigate its pathogenicity. Methods The homologous recombination technique was used to construct the mutant with deletion of the gene and the role of 0462 gene in the growth, invasion and in vivo colonization of NTSN. Colon adenocarcinoma Caco-2 and human hepatoma HepG2 in BALB / c mice and the ability of colonization in the liver and spleen of BALB / c mice. Results The growth curve of the deletion strain showed that NTSN_0462 had no significant effect on the growth and metabolism of Lm in BHI medium. In vitro tests with human colon adenocarcinoma Caco-2 and human hepatocellular carcinoma HepG2 showed that the invasive strains were less invasive than the wild-type (P <0.001). In vivo tests in BALB / c mice showed that the abilities of colonies in the liver of the deletion strain were significantly lower than those of the wild strain (P <0.001), and there was no significant difference in the spleen. Conclusion The NTSN_0462 gene plays an important role in invasion of liver and colonization of LmNTSN and is an important virulence factor related to invasion.