Laboratory of Advanced Chemical Biology, Graduate School of Life Science, Hokkaido University, Sapporo, 001-0021, Japan.
Frontier Research Center for Advanced Material and Life Science, Faculty of Advanced Life Science, Hokkaido University, Sapporo, 001-0021, Japan.
Sci Rep. 2024 Jun 3;14(1):12719. doi: 10.1038/s41598-024-62729-1.
Polypeptide-targeted MALDI-TOF MS for microbial species identification has revolutionized microbiology. However, no practical MALDI-TOF MS identification method for O-antigen polysaccharides, a major indicator for epidemiological classification within a species of gram-negative bacteria, is available. We describe a simple MALDI glycotyping method for O-antigens that simultaneously identifies the molecular mass of the repeating units and the monosaccharide composition of the O-antigen. We analyzed the Escherichia coli O1, O6, and O157-type strains. Conventional species identification based on polypeptide patterns and O-antigen polysaccharide typing can be performed in parallel from a single colony using our MALDI-TOF MS workflow. Moreover, subtyping within the same O-antigen and parallel colony-specific O-antigen determination from mixed strains, including the simultaneous identification of multiple strains-derived O-antigens within selected colony, were performed. In MALDI glycotyping of two Enterobacteriaceae strains, a Citrobacter freundii strain serologically cross-reactive with E. coli O157 gave a MALDI spectral pattern identical to E. coli O157. On the other hand, an Edwardsiella tarda strain with no reported O-antigen cross-reactivity gave a MALDI spectral pattern of unknown O-antigen repeating units. The method described in this study allows the parallel and rapid identification of microbial genera, species, and serotypes of surface polysaccharides using a single MALDI-TOF MS instrument.
多肽靶向 MALDI-TOF MS 用于微生物物种鉴定已经彻底改变了微生物学。然而，对于革兰氏阴性菌种内流行病学分类的主要指标 O-抗原多糖，目前还没有实用的 MALDI-TOF MS 鉴定方法。我们描述了一种用于 O-抗原的简单 MALDI 糖型分析方法，该方法可以同时鉴定重复单元的分子量和 O-抗原的单糖组成。我们分析了大肠杆菌 O1、O6 和 O157 型菌株。使用我们的 MALDI-TOF MS 工作流程，可以从单个菌落中同时进行基于多肽模式和 O-抗原多糖分型的常规物种鉴定。此外，可以在同一 O-抗原内进行亚型分型，并从混合菌株中平行进行特定菌落的 O-抗原测定，包括同时鉴定选定菌落中多个菌株衍生的 O-抗原。在两种肠杆菌科菌株的 MALDI 糖型分析中，与大肠杆菌 O157 血清学交叉反应的弗氏柠檬酸杆菌菌株产生的 MALDI 光谱模式与大肠杆菌 O157 相同。另一方面，没有报道 O-抗原交叉反应的爱德华氏菌属菌株产生了未知 O-抗原重复单元的 MALDI 光谱模式。本研究中描述的方法允许使用单个 MALDI-TOF MS 仪器平行快速鉴定微生物属、种和表面多糖血清型。
