Rioux S, Galarneau C, Harel J, Frey J, Nicolet J, Kobisch M, Dubreuil J D, Jacques M
Département de pathologie et microbiologie, Faculté de médecine vétérinaire, Université de Montréal, St-Hyacinthe, QC, Canada.
Can J Microbiol. 1999 Dec;45(12):1017-26. doi: 10.1139/w99-107.
Lipopolysaccharide (LPS) has previously been identified as the major adhesin of Actinobacillus pleuropneumoniae involved in adherence to porcine respiratory tract cells. The purpose of the present study was to isolate and characterize mutants in LPS biosynthesis by using a mini-Tn10 transposon mutagenesis system. Seven mutants appeared to possess a rough LPS (among which two had similar Southern blot profiles) while one mutant (#5.1) expressed the high-molecular-mass LPS, but as visualized by Tricine SDS-PAGE, showed an additional band in the core-lipid A region. The LPS mutants showed sensitivity to pig serum to various degrees, while the parent strain was serum-resistant. Use of piglet frozen tracheal sections indicated that, surprisingly, the rough LPS mutants adhered similarly or in greater numbers than the parent strain. However, the LPS mutant #5.1 adhered significantly less than the parent strain and was also less virulent in pigs. The gene affected by mini-Tn10 in LPS mutant #5.1 is galU, the structural gene for UTP-alpha-D-glucose-1-phosphate uridylyltransferase, involved in LPS core biosynthesis. Complementation analysis confirmed that the phenotypic characteristics of LPS mutant #5.1 are the result of the inactivation of the galU gene. Our data suggest that although the presence of O-antigen does not seem to be essential, an intact core-lipid A region might be required for adherence of A. pleuropneumoniae to porcine respiratory tract cells. To the best of our knowledge, these mutants represent the first isogenic mutants of A. pleuropneumoniae defective in LPS biosynthetic genes.
脂多糖（LPS）先前已被确定为胸膜肺炎放线杆菌的主要黏附素，参与对猪呼吸道细胞的黏附。本研究的目的是利用mini-Tn10转座子诱变系统分离和鉴定LPS生物合成中的突变体。七个突变体似乎具有粗糙型LPS（其中两个具有相似的Southern杂交图谱），而一个突变体（#5.1）表达高分子量LPS，但经Tricine SDS-PAGE可视化显示，在核心脂质A区域有一条额外的条带。LPS突变体对猪血清表现出不同程度的敏感性，而亲本菌株具有血清抗性。使用仔猪冷冻气管切片表明，令人惊讶的是，粗糙型LPS突变体的黏附情况与亲本菌株相似或数量更多。然而，LPS突变体#5.1的黏附显著少于亲本菌株，并且在猪体内的毒力也较低。LPS突变体#5.1中受mini-Tn10影响的基因是galU，它是参与LPS核心生物合成的UTP-α-D-葡萄糖-1-磷酸尿苷酰转移酶的结构基因。互补分析证实，LPS突变体#5.1的表型特征是galU基因失活的结果。我们的数据表明，虽然O抗原的存在似乎不是必需的，但完整的核心脂质A区域可能是胸膜肺炎放线杆菌黏附猪呼吸道细胞所必需的。据我们所知，这些突变体代表了胸膜肺炎放线杆菌中首个LPS生物合成基因缺陷型的同基因突变体。
