两株母乳源植物乳杆菌的全基因组测序分析

Whole genome sequencing of two Lactobacillus plantarum strains isolated from human milk

摘要:

【背景】母乳是一个重要的益生菌筛选库，其中植物乳杆菌是一种用途广泛、适应性强的益生菌。然而不同菌株具有不同的功能，现有的生理生化方法对其潜在益生特性研究十分有限，有必要采用高通量的方法寻找具有种群特异性的优质益生菌。【目的】结合菌株生化特征在全基因组的测序与分析的基础上对两株植物乳杆菌的潜在功能进行预测，并重点找寻与肠液耐受性及细菌素的合成相关的基因，即在基因组的结构上对菌株的表型进行探究。【方法】分离筛选出两株母乳源植物乳杆菌MP55、MP37，并利用Illumina genome analyzer对菌株的全基因组进行测序，采用Prokka软件对细菌基因组进行注释，采用Carbohydrate-active enzymes (CAZy)、Koyto encyclopedia of genes and genomes (KEGG)和Clusters of orthologous genes (COG)数据库对基因组进行功能注释；同时采用Prodigal、RNAmmer等工具对编码序列、核糖体RNA进行预测，并用CGView软件绘制菌株的基因组环形图谱。【结果】通过基因组装得到了两株植物乳杆菌的全基因组信息，植物乳杆菌MP37、MP55基因组大小分别为3 204 421 bp和3 299 180 bp；(G+C)mol%含量分别为44.36%和44.46%；分别包含3 012个和3 101个DNA编码序列，结合菌株生化特征在基因组上找到4个与肠液耐受相关的基因及一段细菌素合成相关基因簇。基因组序列原始数据和拼接结果已提交至“gcMeta”平台。【结论】通过高通量测序分析在基因组水平上揭示了植物乳杆菌MP55、MP37在肠道存活性与抑制病原菌相关的可能机理。植物乳杆菌MP55、MP37是两株潜在的益生菌候选菌株，实验结果为进一步阐明其益生菌特性的功能机制提供了遗传学基础。

Abstract:

[Background] Breast milk is an important screening database for probiotics, among which Lactobacillus plantarum is a one that is adaptable and widely used. Different strains of Lactobacillus plantarum show differences in functions, but the existing physiological and biochemical methods have limited research on its potential probiotic characteristics. Identify probiotics with good specificity by high-throughput method has become a hot topic in probiotics, research. [Objective] Combining with the biochemical characteristics of the bacterial strains, the potential functions of the two strains of Lactobacillus plantarum were predicted based on the whole genome sequencing and analysis. We explored the phenotype of the bacterial strains based on the genomic structure. In particular, we searched for genes related to intestinal fluid tolerance and bacteriocin synthesis. [Methods] We separated two strains of Lactobacillus plantarum (MP55, MP37) from human milk, used the Illumina genome analyzer to sequence whole genomes of the two strains, annotated the genomes by Prokka software, adopt carbohydrate-active enzymes (CAZy), and performed functional annotation using the koyto encyclopedia of genes and genomes (KEGG) and clusters of orthologous genes (COG) databases. Coding sequences and ribosomal RNA were predicted using Prodigal, RNAmmer and other tools, and the genome ring map of the bacterial strain was plotted using CGView software. [Results] The whole genome information of two strains of Lactobacillus plantarum was obtained through gene assembly. The genome sizes of Lactobacillus plantarum MP37 and MP55 were 3 204 421 bp and 3 299 180 bp, respectively. (G+C)mol% contents were 44.36% and 44.46%, respectively. The genome contained 3 012 and 3 101 DNA coding sequences, and four genes related to intestinal fluid tolerance and a cluster of genes related to bacteriocin synthesis were found by combining the biochemical characteristics of the bacterial strain. Raw genomic sequence data and splicing results have been submitted to the “gcMeta” platform. [Conclusion] Through high-throughput sequencing analysis, this study revealed the possible mechanism of MP55 and MP37 activity of Lactobacillus plantarum in the intestinal tract and inhibition of pathogenic bacteria. Lactobacillus plantarum MP55 and MP37 are two potential candidate strains of probiotics. The results of this experiment provide genome information for further elucidation of the functional mechanism of the probiotics.