Comparative genomics and high-resolution discrimination of Mycoplasma pneumoniae clinical isolates. Mycoplasmoides pneumoniae

Genomes of 16 Mycoplasma pneumoniae clinical isolates were sequenced and compared with 5 previously reported M. pneumoniae genome sequences of M129, M129-B7, FH, 309 and PO1 strains by comparative genomics approaches. Pangenomic analysis showed that 97% of pangenome sequence of 21 M. pneumoniae strains are shared by coregenome sequence of these strains, indicating highly closed pangenomic feature of this species, although there are deep genetic divergence between P1 ctyadhesin subtype 1 and 2 strain linages. From the comparison of coregenome sequences of 21 strains, we identified 1,370 single-nucleotide polymorphism (SNP) sites. Based on these SNPs, all of 21 strains were phylogenetically discriminated, demonstrating high discrimination power of whole-genome SNP analysis and its usability in M. pneumoniae strain classification. The results of whole-genome SNP analysis were largely consistent with MLVA typing, which is currently used as a standard classification method for M. pneumoniae strains. We also performed synonymous and nonsynonymous codon substitution rate (dN/dS) analysis using 21 M. pneumoniae genome sequences. Six ORFs, MPN049, MPN080, MPN278, MPN376, MPN496 and MPN571 were positively selected by this analysis. Five of these except for MPN278 were putative membrane-localized proteins and thought to be the candidates of proteins that undergo antigenic variation.

本研究通过比较基因组学（comparative genomics）手段，对16株肺炎支原体（Mycoplasma pneumoniae）临床分离株的基因组进行测序，并与已报道的5株肺炎支原体基因组（M129、M129-B7、FH、309及PO1菌株）开展比对分析。泛基因组（pangenome）分析结果显示，21株肺炎支原体的泛基因组序列中有97%可被这些菌株的核心基因组（coregenome）序列覆盖，表明该物种的泛基因组具有高度闭合的特征；尽管P1细胞黏附素（cytoadhesin）亚型1与亚型2的菌株谱系间存在显著遗传分化。通过对21株菌株的核心基因组序列进行比对，本研究共鉴定出1370个单核苷酸多态性（single-nucleotide polymorphism, SNP）位点。基于这些SNP位点，21株菌株均可通过系统发育分析实现区分，证明全基因组SNP分析具备较高的区分能力，可用于肺炎支原体的菌株分型。全基因组SNP分析结果与当前作为肺炎支原体菌株分型标准方法的多位点可变数目串联重复序列分型（Multilocus Variable-number tandem repeat analysis, MLVA）结果高度一致。本研究还利用21株肺炎支原体的基因组序列开展了同义密码子与非同义密码子替换速率（dN/dS）分析，经该分析鉴定出6个受到正向选择的开放阅读框（open reading frame, ORF），分别为MPN049、MPN080、MPN278、MPN376、MPN496及MPN571。其中除MPN278外的5个开放阅读框均被预测为膜定位蛋白，被认为是发生抗原变异的候选蛋白。