屎肠球菌F11.1G比较基因组、代谢组学分析及代谢产物挖掘

Comparative genomics analysis, metabolomics analysis, and metabolite mining of Enterococcus faecium F11.1G

摘要:

【背景】屎肠球菌(Enterococcus faecium)具有抑制常见致病菌的作用，在禁抗的大环境下，逐渐成为常规抗生素生长促进剂的有效替代品。但随着屎肠球菌应用范围的扩大，其安全性问题日益受到关注。【目的】从基因水平阐明其细菌素编码基因簇，评估其安全性并探究屎肠球菌F11.1G基因功能和潜在代谢产物，代谢组学分析菌株胞内外的代谢物差异，并对其活性代谢产物进行挖掘。【方法】全基因组测序对其基因信息进行注释和功能预测，采用功能基因组和比较基因组方法，分析菌株F11.1G的益生性和安全性特征，揭示其与益生屎肠球菌T110之间的差异，代谢组学对菌株中的代谢物进行鉴定，随后通过结构鉴定挖掘出活性代谢产物。【结果】菌株F11.1G注释到4个毒力相关基因和3个耐药基因，antiSMASH数据库预测到了3种次级代谢产物基因簇；比较基因组分析结果表明，菌株F11.1G与菌株T110具有较高相似性，菌株T110未预测到毒力基因、耐药基因和病原菌基因。非靶向代谢组学结果表明，胞内外差异代谢产物主要集中在萜类和聚酮类、氨基酸、有机杂环化合物等，可能与菌株F11.1G的抑菌作用相关。从菌株F11.1G的活性代谢产物分离鉴定出α-d-型葡萄糖、4-氧化戊酸、毛蕊花糖、水苏糖、二聚体白屈菜默碱、1-羟基蒽醌和极少量酮类活性物质。【结论】菌株F11.1G具有合成抗生素或新型次级代谢产物的潜力，但还携带毒力因子和毒力基因，后期可通过分离纯化肠球菌素发挥其益生菌特性。

Abstract:

[Background] Enterococcus faecium with the ability to inhibit common pathogenic bacteria has gradually become an alternative to conventional antibiotic growth promoters in the context of the antibiotic ban. However, the safety of E. faecium has aroused growing concern with the expansion of its application. [Objective] To elucidate the bacteriocin-encoding gene clusters, evaluate the safety, investigate the gene functions and potential metabolites, compare the intracellular and extracellular metabolites by metabolomics, and mine the active metabolites of E. faecium F11.1G. [Methods] Whole genome sequencing was performed to annotate the gene information and predict gene functions of E. faecium F11.1G. The functional genomic and comparative genomic approaches were used to analyze the probiotic and safety characteristics of strain F11.1G and reveal the differences between this strain and the probiotic Enterococcus faecium T110. Metabolomics was employed to identify the metabolites in the strain, and then active metabolites were mined out through structural identification. [Results] Four virulence genes and three antibiotic resistance genes were annotated for strain F11.1G. Three secondary metabolite gene clusters were predicted by antiSMASH. The comparative genomic analysis showed high similarity between F11.1G and T110, and no virulence genes, resistance genes or pathogenicity genes were predicted for T110. The results of non-targeted metabolomics showed that the differential metabolites were mainly terpenoids, polyketides, amino acids, and organic heterocyclic compounds, which might be associated with the antibacterial effect of F11.1G. Among the active metabolites of F11.1G, α-d-glucose, 4-oxopentanoic acid, verbascose, stachyose, dimeric chelidonine, 1-hydroxy anthraquinone, and a very small amount of active ketones were identified. [Conclusion] Strain F11.1G has the potential to synthesize antibiotics or novel secondary metabolites and carries virulence factors and virulence genes. Its probiotic properties can be exploited later by separation and purification of enterocin.