猪链球菌2型唾液酸合成酶neu B基因敲除突变株的构建及其生物学特性

Construction and Function Study of the neuB Gene Encoding Sialic Acid Synthase Knock-out Mutant of Streptococcus suis Serotype 2

摘要:

利用同源重组基因敲除方法构建猪链球菌2型强毒株05ZYH33唾液酸合成酶neuB基因敲除突变株。PCR和Southern杂交结果均显示neuB基因在1株转化重组体中完全被壮观霉素抗性基因替代, 表明neuB基因敲除突变体构建成功。生物学特性鉴定显示, 突变体与强毒株在菌落形态、溶血活性以及染色特性方面均无明显差异; 电镜检查发现突变体表面结构组分与强毒株有显著差异, 荚膜明显变薄, 质地更加紧密; 小鼠致病性试验结果显示, 突变体毒力显著减弱。研究结果提示菌体荚膜中的唾液酸对于猪链球菌2型侵袭和致病具有重要作用。

关键词:猪链球菌2型, 唾液酸合成酶, 基因敲除, 生物学特性, 致病性

Abstract:

Based on the principle of homologous recombination, we construct gene knock-out mutant of neuB gene encoding sialic acid synthase in Streptococcus suis serotype 2 (SS2) virulent strain 05ZYH33. PCR analysis and Southern hybridization confirmed that the coding gene of neuB was replaced completely by spcr cassette in one mutant, the mutant of 05ZYH33 neuB gene was successfully constructed. Analysis of biological characteristics showed that there were no differences in mycelial morphology, hemolytic activity and dyeing properties between the mutant and 05ZYH33; but the capsule of mutant was thinner and more compacted than that of the wild type strain 05ZYH33. Virulence assays with murine model confirmed that the mutant is avirulent. These observations indicate that sialic acid plays an important role in the pathogenesis and invasiveness of SS2.

Key words:Streptococcus suis serotype 2, Sialic acid synthase, Gene knock-out, Biological characteristics, Virulence assays