Simple Identification of Natto Bacteria by MALDI-TOF MS Analysis with Ethanol-Formic Acid Pretreatment

Abstract

　In this study, to improve the reliability of identifying natto bacteria strains isolated from natto all over Japan, the effectiveness of a pretreatment, using an ethanol-formic acid treatment, in identifying and distinguishing the natto bacteria strains using MALDI-TOF MS was investigated. Samples pretreated with the direct coating-matrix treatment, which is the manufacturer's recommended pretreatment method, were subjected to MALDI-TOF MS and identified using an existing database. As a result, it was difficult to accurately identify the strains of the B. subtilis groups including the isolates from natto. Therefore, it was necessary to construct an original database to improve the reliability of identifying natto bacteria strains isolated from natto. To improve the accuracy of their analysis, the MALDI-TOF MS analysis was performed with the ethanol-formic acid treatment as the pretreatment. In a dendrogram that was constructed based on the mass spectral data obtained, strains isolated from natto formed independent clusters distinct from other species of the B. subtilis group. Furthermore, the mass spectra obtained for the strains isolated from natto were characteristic of "B. subtilis subsp. natto", which is different from those obtained for other B. subtilis groups.