GreA蛋白对肠炎沙门菌生物学特性及致病力影响

doi:10.11843/j.issn.0366-6964.2024.11.032

摘要/Abstract

摘要：

旨在研究转录延长因子GreA蛋白对肠炎沙门菌生物学特性和致病力的影响。以ATCC13076为亲本株，通过λ-Red重组系统构建ΔgreA和ΔgreB单基因缺失株、ΔgreAΔgreB双基因缺失株，检测亲本株及缺失株的体外生长和运动力，对压力环境抵抗力，生物被膜形成能力，以及对细胞和小鼠的致病力，并利用RNA-seq检测差异表达基因。结果显示：GreA蛋白负调控肠炎沙门菌H₂O₂抵抗力、生长速度以及生物被膜形成能力，正调控肠炎沙门菌运动能力、细胞入侵和胞内增殖能力及其在小鼠脏器的定殖能力，但对高温和酸性环境抵抗力无显著影响。GreA蛋白影响107个基因表达，其中正向调控fim操纵子、flg操纵子、inv操纵子等，主要与细菌运动及入侵细胞相关；负向调控于cps操纵子、cys操纵子、leu操纵子等，主要与氨基酸合成及代谢相关。GreA蛋白通过调控鞭毛合成和细胞入侵相关代谢通路的表达，影响肠炎沙门菌环境适应性生存及致病力。

关键词: 肠炎沙门菌, GreA, 生物学特性, RNA-seq

Abstract:

The aim is to study the effect of GreA protein on the biological characteristics and pathogenicity of Salmonella Enteritidis. Based on the parent strain ATCC13076, ΔgreA and ΔgreB single gene deletion strains and ΔgreAΔgreB double gene deletion strains were constructed by λ-Red recombinant system. Their growth and swimming motility, resistance to stressful environment, biofilm formation ability, and pathogenicity to cells and mice of the parents and the deletion strains were detected. RNA-seq was used to detect differentially expressed genes (DEGs). GreA protein negatively regulated the H₂O₂ resistance, the growth in vitro and biofilm formation ability of Salmonella Enteritidis, and positively regulated the motor ability, cell invasion and intracellular proliferation ability of Salmonella Enteritidis and its colonizing ability in mouse organs, but have no significant effect on resistance of Salmonella Enteritidis to high temperature and acid environment. GreA protein affects the expression of 107 genes, including positive regulation of inv operon, fim operon and flg operon, which are mainly related to bacterial motility and cell invasion; and negative regulation of cps operon, cbp operon and leu operon, which are related to amino acid synthesis and metabolism. GreA protein affects the environmental adaptability and pathogenicity of Salmonella Enteritidis by regulating the expression of metabolic pathways related to flagellar synthesis and cell invasion.

Key words: Salmonella Enteritidis, biological characteristics, GreA, RNA-seq

中图分类号:

S852.612

蔡梦雷, 赵东旭, 张政钢, 刘东海, 姜婷婷, 苏士炫, 闫雪敏, 薛晓阳, 崔国林. GreA蛋白对肠炎沙门菌生物学特性及致病力影响[J]. 畜牧兽医学报, 2024, 55(11): 5173-5182.

Menglei CAI, Dongxu ZHAO, Zhenggang ZHANG, Donghai LIU, Tingting JIANG, Shixuan SU, Xuemin YAN, Xiaoyang XUE, Guolin CUI. The Effect of GreA Protein on the Biological Characteristics and Pathogenicity of Salmonella Enteritidis[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(11): 5173-5182.

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引物 Primers	序列(5′→3′) Sequence
greAF	AGCCGGGGTAGAGTTCATTGCCCCCTACAGGAATGTTCAAGAGGTATAACGTGTAGGCTGGAGCTGCTTC
greAR	ACAATACACCAACAATTTGCGTATTGAGTACTACTTAAAGGTATTCCACCATATGAATATCCTCCTTAG
greBF	ACGTTATACCCCGTGTGCGCAATATCGACAGCAAAGGTAAATCAACGAGGTGTAGGCTGGAGCTGCTTC
greBR	ATGCCAGCCATCAGCGGGGGCTTAGGATTCTTCTTGTCTTATTTGACGCATATGAATATCCTCCTTAG
greApF	GCGGATCCGATGCAAGCTATTCCGATGACCT
greApF	CGAAGCTTAAGGTATTCCACTTTAAGCACT
greBpF	GCGGATCCGATGAAAACGCCCCTGATCAC
greBpR	CGAAGCTTTTTGACGTATTCGATCGCA

差异类型 Difference type	操纵子 Operons	功能描述 Function description
上调 Upregulated	invA-invE-invG-invF	SPI-1
	ttrC-ttrA	SPI-2
	ssaL-ssaK	SPI-2
	ssaN-ssaV-ssaM	SPI-2
	flgJ-SEN_RS09710-flgH	Flagellar L-ring
	flgD-flgC-flgB	Flagellar bas al-body rod modification
	flhA-flhE	Flagellar biosynthesis
	fliR- fliQ	Flagellar biosynthesis
	fimA-fimI-fimC	Type 1 fimbrial
	cheR-cheB-cheY	Bacterial chemotaxis protein
	motA-motB-cheA-cheW	Bacterial chemotaxis protein
	dppD-dppC	Dipeptide transport system binding protein
	argC-argB	Arginine biosynthesis
	potE- speF	Arginine and proline metabolism
下调 Downregulated	leuD-leuC-leuB	Leucine synthesis
	cpsG-cpsB-wcaI- SEN_RS10955-SEN_RS10960-gmD-wcaF	Fructose and mannose metabolism
	yedF-yedE	Anaerobic metabolism
	cysC-cysN-cysD	Purine metabolism
	SEN_RS05075-wrbA	Ubiquinone and other terpenoid-quinone biosynthesis
	sigE-sopB	SPI-1
	cbpM-cbpA	Curved DNA-binding protein

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