猪肺炎支原体强弱毒株的生物学特性及比较基因组学分析

doi:10.11843/j.issn.0366-6964.2025.02.033

摘要/Abstract

摘要：

旨在为更好地理解猪肺炎支原体(Mycoplasma hyopneumoniae，Mhp)强弱毒株在物质代谢、毒力变化的差异，本研究基于Mhp强毒株ES-2株及其传代致弱株ES-2L株，开展二者的生长特性、黏附细胞能力、代谢产生过氧化氢水平、生物被膜形成能力等生物学特性及比较基因组学研究。在生长特性方面，ES-2L株的生长速度和滴度要高于ES-2株，其颜色改变单位(color change unit, CCU)为1.0×10¹⁰ CCU·mL^-1，在培养基上具有更好的适应性。在与毒力变化相关的特性方面，ES-2和ES-2L株均可以形成生物被膜，但ES-2L株的生物被膜形成能力要弱于ES-2株。ES-2L株代谢甘油产生过氧化氢的水平要低于ES-2株，且其对支气管上皮细胞的黏附能力要弱于ES-2株。ES-2和ES-2L基因组共线性比对分析结果表明，二者的总体共线性良好，但ES-2L株存在几处基因的缺失、倒位现象，对ES-2L株中的缺失基因进行定位分析，共检测到9个大片段(>500 bp)的基因缺失，这9个片段共包含18个缺失基因。SNP(单核苷酸多态性)和Indel(插入与缺失突变)分析表明，共检测到348个SNVs(单核苷酸突变)，这些SNVs主要分布于99个基因上，并且有22个dels(缺失突变)主要分布于19个基因上。基因岛分析结果表明，ES-2L株在395~425 kb有一个基因岛编码序列，ES-2株在354~364、920~945 kb有两个基因岛编码序列。这些缺失或发生SNP的基因、基因岛的变化可能与ES-2L毒力下降及生长特性改变有关。综上，本研究阐明了Mhp强弱毒株在物质代谢、毒力变化的差异，并采用比较基因组学的方法，分析了二者在基因水平的变化，为进一步理解Mhp致病机制和毒力因子挖掘提供一种视角。

关键词: 猪肺炎支原体, 毒力, 比较基因组学, 生物被膜

Abstract:

In order to better understand the differences in material metabolism and virulence changes between virulent and attenuated strains of Mycoplasma hyopneumoniae (Mhp), this study was based on the virulent strain ES-2 and the attenuated strain ES-2L induced by passaging of strain ES-2.The growth characteristics, cell adhesion ability, metabolic production of hydrogen peroxide level, biofilm formation ability and other biological characteristics and comparative genomics of the two strains were studied. In terms of growth characteristics, the growth rate and titer of ES-2L strain were higher than that of ES-2 strain, and its color change unit (CCU) was 1.0×10¹⁰ CCU·mL^-1, which had better adaptability on medium. In terms of virulence changes, both ES-2 and ES-2L strains could form biofilm, but the biofilm formation capacity of ES-2L strain was weaker than that of ES-2 strain. The level of metabolizing glycerol to produce hydrogen peroxide of ES-2L strain was lower than that of ES-2 strain, and its adhesion ability to bronchial epithelial cells was weaker than that of ES-2 strain. The results of genome collinearity comparison between ES-2 and ES-2L showed that the overall collinearity of the two strains was good, but there were several gene deletion and inversion in ES-2L strains. The localization analysis of the deletion genes in ES-2L strains showed that a total of 9 large fragments (>500 bp) of gene deletion were detected, and these 9 fragments contained a total of 18 deletion genes. Analysis of SNP (single nucleotide polymorphism) and Indel (Insertion and deletion variation) showed that 348 SNVs (single nucleotide variations) were detected, which were mainly distributed in 99 genes, and 22 dels (deletion variations) were mainly distributed in 19 genes. The results of gene island analysis showed that ES-2L had a gene island coding sequence ranging from 395 to 425 kb, and ES-2 had two gene island coding sequences ranging from 354 to 364 kb, and 920 to 945 kb.These deletion or SNP genes and gene island changes may be related to the decrease of virulence and the change of growth characteristics of ES-2L. In summary, this study clarified the differences in material metabolism and virulence changes between virulent and attenuated strains of Mhp, and analyzed the changes of the two strains at the gene level by using comparative genomics method, providing a perspective for further understanding the pathogenic mechanism of Mhp and mining virulence factors.

Key words: Mycoplasma hyopneumoniae, virulence, comparative genomics, biofilm

中图分类号:

S852.62

李真亚, 刘洁, 李允, 王飞, 孔嫄嫄, 李泳, 贾荣玲. 猪肺炎支原体强弱毒株的生物学特性及比较基因组学分析[J]. 畜牧兽医学报, 2025, 56(2): 851-859.

LI Zhenya, LIU Jie, LI Yun, WANG Fei, KONG Yuanyuan, LI Yong, JIA Rongling. Biological Characteristics and Comparative Genomic Analysis of Virulent and Attenuated Strains of Mycoplasma hyopneumoniae[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(2): 851-859.

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突变类型 Mutation type	起始位点 Starting site	终止位点 Termination site	大小/bp Size	包含基因 Contain genes	编码蛋白 Coding protein
缺失 Deletion	113 237	114 699	1 462	E5E95_00505	丝氨酸蛋白酶
缺失 Deletion	159 989	161 778	1 789	E5E95_00670	假定蛋白
				E5E95_00665	假定蛋白
				E5E95_00675	假定蛋白
缺失 Deletion	203 386	208 867	5 481	E5E95_00935	假定蛋白
				E5E95_00940	假定蛋白
缺失 Deletion	261 435	262 128	693	E5E95_01210	假定蛋白
缺失 Deletion	262 717	263 417	700	E5E95_01215	假定蛋白
缺失 Deletion	417 189	420 421	3 232	E5E95_01850	假定蛋白
				E5E95_01845	假定蛋白
				E5E95_01855	假定蛋白
				E5E95_01860	乙醇脱氢酶
缺失 Deletion	429 321	433 928	4 607	E5E95_01905	DNA促旋酶
				E5E95_01910	假定蛋白
				E5E95_01915	丝氨酸蛋白酶
缺失 Deletion	558 155	560 135	1 980	E5E95_02435	IS1634家族转座酶
				E5E95_02440	DNA甲基转移酶
缺失 Deletion	925 798	927 220	1 422	E5E95_03755	DDE结构域蛋白

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