非编码小RNA FrnS基因缺失对鼠伤寒沙门菌致病性的影响

doi:10.16431/j.cnki.1671-7236.2025.10.036

摘要/Abstract

摘要： 【目的】通过构建非编码小RNA(sRNA)FnrS基因缺失株，探究FnrS基因对鼠伤寒沙门菌致病性的影响。【方法】利用λ-Red同源重组技术构建鼠伤寒沙门菌FnrS基因缺失株ΔFnrS，并测定其遗传稳定性、生长曲线及生化特性；通过检测菌株对小鼠的半数致死剂量(LD₅₀)、组织载菌量及病理组织切片观察分析FnrS基因对鼠伤寒沙门菌致病性的影响。【结果】成功构建FnrS基因缺失株ΔFnrS，其能稳定遗传。生长曲线和生化试验结果显示，FnrS基因缺失不影响鼠伤寒沙门菌的生长能力及生化特性。小鼠致病性试验结果显示，缺失株ΔFnrS对小鼠的LD₅₀为1.06×10⁵ CFU，显著高于亲本株和回补株(P＜0.05)；缺失株ΔFnrS与亲本株感染后小鼠存活率无显著差异(P＞0.05)；缺失株ΔFnrS株感染后7、9 d，小鼠肝脏载菌量显著低于亲本株和回补株(P＜0.05)，感染后5、7、9 d小鼠脾脏载菌量显著低于亲本株和回补株(P＜0.05)。【结论】FnrS基因缺失降低了鼠伤寒沙门菌的致病能力，研究结果为鼠伤寒沙门菌致病机制的深入研究奠定了基础。

关键词: 鼠伤寒沙门菌; sRNA; FnrS基因; 致病性

Abstract: 【Objective】 This study aimed to explore the effect of FnrS gene on the pathogenicity of Salmonella Typhimurium by constructing non-coding small RNA (sRNA) FnrS gene deletion strain.【Method】 The λ-Red homologous recombination technology was used to construct the FnrS gene deletion strain ΔFnrS of Salmonella Typhimurium,and its genetic stability,growth curve and biochemical characteristics were determined.The effect of FnrS gene on the pathogenicity of Salmonella Typhimurium was analyzed by detecting the median lethal dose (LD₅₀) of the strain on mice,the bacterial load of the tissue and the pathological tissue sections.【Result】 The FnrS gene deletion strain ΔFnrS was successfully constructed and it could be stably inherited.The growth curve and biochemical test results showed that the deletion of FnrS gene did not affect the growth ability and biochemical characteristics of Salmonella Typhimurium.The results of the pathogenicity test in mice showed that the LD₅₀ of deletion strain ΔFnrS for mice was 1.06×10⁵ CFU，which was significantly higher than that of parental and complement strains (P＜0.05).There was no significant difference in the survival rate of mice between deletion strain ΔFnrS and parental strain (P＞0.05).At 7 and 9 days after infection with deletion strain ΔFnrS,the bacterial load in the liver of mice was significantly lower than that of parental and complement strains (P＜0.05).At 5，7 and 9 days after infection，the bacterial load in the spleen of mice was significantly lower than that of parental and complement strains (P＜0.05).【Conclusion】 The deletion of FnrS gene reduced the pathogenicity of Salmonella Typhimurium.This results laid a foundation for the further research of the pathogenic mechanism of Salmonella Typhimurium.

Key words: Salmonella Typhimurium; sRNA; FnrS gene; pathogenicity

中图分类号:

S852.61

宁程程, 田路路, 冯凯, 陈英, 艾佳吴怡, 柳旭伟, 杨力伟, 张再超. 非编码小RNA FrnS基因缺失对鼠伤寒沙门菌致病性的影响[J]. 中国畜牧兽医, 2025, 52(10): 4912-4919.

NING Chengcheng, TIAN Lulu, FENG Kai, CHEN Ying, AI Jiawuyi, LIU Xuwei, YANG Liwei, ZHANG Zaichao. Effect of sRNA FrnS Gene Deletion on the Pathogenicity of Salmonella Typhimurium[J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(10): 4912-4919.

参考文献

[1] MOHAN A,MUNUSAMY C,TAN Y C,et al.Invasive Salmonella infections among children in Bintulu,Sarawak,Malaysian Borneo:A 6-year retrospective review[J].BMC Infectious Diseases,2019,19(1):330.
[2] 周南龙，丁永辉，李天森.鼠伤寒沙门菌毒力基因SptP的原核表达及生物信息学分析[J].中国畜牧兽医,2022,49(12):4734-4744. ZHOU N L,DING Y H,LI T S.Prokaryotic expression and bioinformatics analysis of Salmonella Typhimurium virulence gene SptP[J].China Animal Husbandry & Veterinary Medicine,2022,49(12):4734-4744.(in Chinese)
[3] LU J,WU H,WU S,et al.Salmonella:Infection mechanism and control strategies[J].Microbiological Research，2025,292:128013.
[4] DENG L,WANG S.Colonization resistance:The role of gut microbiota in preventing Salmonella invasion and infection[J].Gut Microbes，2024,16(1):2424914.
[5] 杨延，毛曌，李干武，等.致病性大肠杆菌非编码小RNA(sRNAs)的研究进展[J].中国预防兽医学报,2022,44(4):454-460. YANG Y,MAO Z,LI G W,et al.Research progress of non-coding small RNAs (sRNAs) in pathogenic Escherichia coli[J]. Chinese Journal of Preventive Veterinary Medicine,2022,44(4):454-460.(in Chinese)
[6] 陈斌杰，王亨，陈艳飞，等.沙门菌非编码小RNA RyhB研究进展[J].畜牧兽医学报，2019，50(1)：21-27. CHEN B J,WANG H,CHEN Y F,et al.Research progress on small non-coding RNA RyhB of Salmonella[J].Acta Veterinaria et Zootechnica Sinica,2019,50(1):21-27.(in Chinese)
[7] 张香美，许冬倩，闫洪波.细菌sRNA与群体感应系统相互作用研究进展[J].微生物学通报,2016,43(8):1822-1828. ZHANG X M,XU D Q,YAN H B.Research progress on interaction between bacterial sRNA and quorum sensing system[J].Microbiology China,2016,43(8):1822-1828.(in Chinese)
[8] 李瑞莲，陈杨.细菌非编码小RNA功能及作用机制研究进展[J].中国微生态学杂志,2014,26(11):1359-1361. LI R L,CHEN Y.The functions and mechanisms of small non-coding RNAs in bacteria:Research progress[J].Chinese Journal of Microecology,2014,26(11):1359-1361.(in Chinese)
[9] NEGRETE A,SHILOACH J.Improving E.coli growth performance by manipulating small RNA expression[J].Microbial Cell Factories，2017,16(1):198.
[10] KIM K,GOLUBEVA Y A,VANDERPOOL C K,et al.Oxygen-dependent regulation of SPI1 type three secretion system by small RNAs in Salmonella enterica serovar Typhimurium[J].Molecular Microbiology,2019,111(3)：570-587.
[11] FABREGA A,VILA J. Salmonella enterica serovar Typhimurium skills to succeed in the host:Virulence and regulation[J].Clinical Microbiology Reviews,2013,26(2):308-341.
[12] 孟霞，孟宪臣，王金秋，等.肠炎沙门菌非编码小RNA isrC基因缺失株的致病性[J].中国兽医学报，2015，35(7)：1095-1098. MENG X,MENG X C,WANG J Q,et al.Pathogenicity of the non-coding small RNA isrC gene deletion mutant in Salmonella Enteritidis[J].Chinese Journal of Veterinary Science,2015,35(7)：1095-1098.(in Chinese)
[13] DOS-SANTOS A P,FERRARI R G,CONTE-JUNIOR C A.Type three secretion system in Salmonella Typhimurium:The key to infection[J].Genes Genomics,2020,42(5):495-506.
[14] HUME P J,SINGH V,DAVIDSON A C,et al.Swiss army pathogen:The Salmonella entry toolkit[J].Frontiers in Cellular and Infection Microbiology,2017,7:348.
[15] NARM K E,KALAFATIS M,SLAUCH J M.HilD,HilC,and RtsA form homodimers and heterodimers to regulate expression of the Salmonella pathogenicity island Ⅰ type Ⅲ secretion system[J].Journal of Bacteriology，2020,202(9):e00012-20.
[16] CAI L L,XIE Y T,HU H J,et al.A small RNA,SaaS,promotes Salmonella pathogenicity by regulating invasion,intracellular growth,and virulence factors[J].Microbiology Spectrum,2023,11(1):e0293822.
[17] WAGNER C,HENSEL M.Adhesive mechanisms of Salmonella enterica[J]. Advances in Experimental Medicine and Biology,2011,715:17-34.
[18] DONG R,LIANG Y,HE S,et al.DsrA modulates central carbon metabolism and redox balance by directly repressing pflB expression in Salmonella Typhimurium[J].Microbiology Spectrum，2022,10(1):e0152221.
[19] KIM S,LEE Y H.Impact of small RNA RaoN on nitrosative-oxidative stress resistance and virulence of Salmonella enterica serovar Typhimurium[J].Journal of Microbiology,2020,58(6):499-506.
[20] LYU C,HU H,CAI L,et al.A trans-acting sRNA SaaS targeting hilD,cheA and csgA to inhibit biofilm formation of S. Enteritidis[J].Journal of Advanced Research,2024,S2090-1232(24)：00232-7.
[21] MENG X,HE M,XIA P,et al.Functions of small non-coding RNAs in Salmonella-host interactions[J].Biology (Basel),2022,11(9):1283.
[22] CHAN H,HO J,LIU X,et al.Potential and use of bacterial small RNAs to combat drug resistance:A systematic review[J].Infection and Drug Resistance,2017,10:521-532.
[23] BOYSEN A,MOLLER J J,KALLIPOLITIS B,et al.Translational regulation of gene expression by an anaerobically induced small non-coding RNA in Escherichia coli[J].Journal of Biological Chemistry,2010,285(14):10690-10702.
[24] DURAND S,STORZ G.Reprogramming of anaerobic metabolism by the FnrS small RNA[J].Molecular Microbiology，2010,75(5):1215-1231.
[25] CRAIG M,SADIK A Y,GOLUBEVA Y A,et al.Twin-arginine translocation system (tat) mutants of Salmonella are attenuated due to envelope defects,not respiratory defects[J].Molecular Microbiology，2013,89(5):887-902.
[26] FRIEDMAN E S,BITTINGER K,ESIPOVA T V,et al.Chemistry in the origin of the anaerobic gut lumen[J].Proceedings of the National Academy of Sciences of the United States of America，2018,115(16):4170-4175.