牛种布鲁氏菌MgtC蛋白在抵抗低Mg2+环境中的生物学功能研究

doi:10.11843/j.issn.0366-6964.2025.01.034

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (1): 365-377.doi: 10.11843/j.issn.0366-6964.2025.01.034

牛种布鲁氏菌MgtC蛋白在抵抗低Mg²⁺环境中的生物学功能研究

王恒泰¹(), 吕浪¹, 蒋卉¹, 程君生², 刘铭赫², 储岳峰³, 许健³, 李朋¹^,*(), 丁家波¹^,*()

1. 中国农业科学院北京畜牧兽医研究所, 北京 100193
2. 中国兽医药品监察所, 北京 100081
3. 中国农业科学院兰州兽医研究所兰州大学动物医学与生物安全学院动物疫病防控全国重点实验室, 兰州 730000

收稿日期:2024-02-28 出版日期:2025-01-23 发布日期:2025-01-18
通讯作者: 李朋,丁家波 E-mail:17861509833@163.com;lipeng01@caas.cn;dingjiabo@caas.cn
作者简介:王恒泰(1998-)，男，山东滕州人，硕士生，主要从事布鲁氏菌抵抗吞噬溶酶体不良环境的研究，E-mail: 17861509833@163.com
基金资助:
中国农业科学院北京畜牧兽医研究所中央级公益性科研院所基本科研业务费专项(2023-YWF-ZYSQ-05)；“十四五”国家重点研发计划(2023YFD1800703)；中国农业科学院科技创新工程项目(CAAS-CSLPDCP-202403)

Biological Function of MgtC Protein in Brucella abortus to Low-Mg²⁺ Resistant Environment

WANG Hengtai¹(), LU Lang¹, JIANG Hui¹, CHENG Junsheng², LIU Minghe², CHU Yuefeng³, XU Jian³, LI Peng¹^,*(), DING Jiabo¹^,*()

1. Institute of Animal Science and Veterinary Medicine, Chinese Academy of Agricultural Science, Beijing 100193, China
2. China Veterinary Drug Supervision Institute, Beijing 100081, China
3. State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730000, China

Received:2024-02-28 Online:2025-01-23 Published:2025-01-18
Contact: LI Peng, DING Jiabo E-mail:17861509833@163.com;lipeng01@caas.cn;dingjiabo@caas.cn

摘要/Abstract

摘要：

MgtC蛋白在多种胞内寄生菌中是一个重要的毒力因子并表现出抵抗低Mg²⁺环境的能力。布鲁氏菌是一种重要的胞内寄生菌，但是关于MgtC蛋白在布鲁氏菌中的生物学功能报道较少。为了全面阐述MgtC在布鲁氏菌中的生物学特性，本文研究了牛种布鲁氏菌MgtC蛋白对低Mg²⁺环境的适应性及其它可能的生物学功能。构建牛种布鲁氏菌(S2308株)的mgtC基因缺失突变株S2308ΔmgtC，同时采用质粒回补方式构建回补株S2308ΔmgtC(pBBR1MCS-mgtC)，随后对其开展低/高Mg²⁺环境中生长曲线测定，ATP含量测定，胞内存活，环境应激，生物被膜检测和小鼠攻毒试验等研究，全面分析MgtC在牛种布鲁氏菌中的生物学功能。结果显示：成功构建缺失株S2308ΔmgtC及其回补株S2308ΔmgtC(pBBR1MCS-mgtC)。牛种布鲁氏菌S2308在低Mg²⁺环境中，mgtC基因转录水平显著上调(P0.05)；与亲本株和回补菌株相比，缺失株S2308ΔmgtC在低Mg²⁺环境中的生长曲线(P < 0.001)、膜结构完整性(P < 0.05)、生物被膜形成能力(P < 0.01)均低于与亲本菌株和回补菌株，而菌体内ATP含量均高于亲本菌株和回补菌株(P < 0.001)。此外，mgtC基因的缺失不影响牛种布鲁氏菌在巨噬细胞内的存活以及对小鼠的致病能力。在低Mg²⁺环境下，牛种布鲁氏菌通过MgtC保证细菌ATP正常水解，维持自身正常生理活动和膜结构的完整性，但是MgtC对牛种布鲁氏菌的毒力和致病性无明显影响，本研究为深入了解牛种布鲁氏菌提供了数据支撑。

关键词: 牛种布鲁氏菌S2308, mgtC, 低Mg²⁺环境, 生物学功能

Abstract:

MgtC protein is an important virulence factor in a variety of intracellular parasites and possesses ability of resistance to low Mg²⁺ environment. Brucella is an important intracellular parasite, and there are few reports on the biological function of MgtC protein in Brucella. To explore the biological characteristics of MgtC in Brucella abortus, the adaptability to low Mg²⁺ environment and other possible biological functions were studied in this study. Brucella abortus S2308 mgtC deletion mutant (S2308ΔmgtC) and the complementary strain S2308ΔmgtC (pBBR1MCS-mgtC) were constructed. Subsequently, studies on the three strains in low/high Mg²⁺ environment with growth curve determination, ATP content determination, intracellular survival, environmental stress, biofilm detection and mouse challenge test were conducted to comprehensively analyze the biological function of MgtC in Brucella abortus. The deletion strain S2308ΔmgtC and its complementary strain S2308ΔmgtC (pBBR1MCS-mgtC) were successfully constructed. In Brucella abortus S2308, expression of mgtC gene was significantly up-regulated in low Mg²⁺ environment(P < 0.05). The growth curve(P < 0.001), membrane structure integrity(P < 0.05)and biofilm formation ability(P < 0.01)of S2308ΔmgtC in low Mg²⁺ environment were lower than those of the parent strain and the complementary strain, while the ATP content in the strain was higher than that of the parent strain and the complementary strain(P < 0.001). In addition, the deletion of mgtC gene did not affect the survival of Brucella abortus to macrophages and the pathogenicity of mice. In low Mg²⁺ environment, Brucella abortus employed MgtC to ensure the normal hydrolysis of ATP and maintain its normal physiological activities and the integrity of membrane structure. However, MgtC had no significant effect on the virulence and pathogenicity of Brucella abortus. This study provides data support for further understanding of Brucella abortus.

Key words: Brucella abortus S2308, mgtC, low Mg²⁺ environment, biological function

中图分类号:

S852.614

王恒泰, 吕浪, 蒋卉, 程君生, 刘铭赫, 储岳峰, 许健, 李朋, 丁家波. 牛种布鲁氏菌MgtC蛋白在抵抗低Mg²⁺环境中的生物学功能研究[J]. 畜牧兽医学报, 2025, 56(1): 365-377.

WANG Hengtai, LU Lang, JIANG Hui, CHENG Junsheng, LIU Minghe, CHU Yuefeng, XU Jian, LI Peng, DING Jiabo. Biological Function of MgtC Protein in Brucella abortus to Low-Mg²⁺ Resistant Environment[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(1): 365-377.

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引物名称 Primer	引物序列(5′→3′) Sequence	长度/ bp Length	作用 Function
mgtC-RL-F	GCGTTTCAGCCTACTTAT	140	mgtC转录水平的检测
mgtC-RL-R	CGGTTCGTTCCTTCAATA
BAB_RS26555-UP-F	GAAGATTCCGTTCTTCCAGAGCACGGCTGC	1 000	扩增mgtC上游同源臂
BAB_RS26555-UP-R	CTCATTTACCGCCTGCTTGGGCACCC
BAB_RS26555-DOWN-F	GACTGCAATCTCATTTGCCGCCCGG	1 000	扩增mgtC下游同源臂
BAB_RS26555-DOWN-R	GTGGTCCAGAATGCGTCGAACAGCCGTTC
pBBR-mgtC-F	CACTTTCGTTTCTCCGGGGGCTAC	916	扩增带有自身启动子mgtC基因
pBBR-mgtC-R	TTATAATCCAGAAAGGCCACCGGAATC
2308-16S-F	GCTGCCCACTGTCACCACCA	153	mgtC转录水平的内参
2308-16S-R	CGCAACCCTCGCCCTTAGTT

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牛种布鲁氏菌MgtC蛋白在抵抗低Mg²⁺环境中的生物学功能研究

Biological Function of MgtC Protein in Brucella abortus to Low-Mg²⁺ Resistant Environment

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