Evaluation of repetitive-PCR and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for rapid strain typing of Bacillus coagulans.

Jun Sato,Takahisa Miyamoto,Motokazu Nakayama,Takumi Sonoda,Motomitsu Hasumi,Ayumi Tomita,Kalimuthusamy Natarajaseenivasan

doi:10.1371/journal.pone.0186327

Jun Sato, Takahisa Miyamoto + Show 5 more

Open Access

https://doi.org/10.1371/journal.pone.0186327

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Journal: PloS one	Publication Date: Oct 11, 2017
Citations: 7	License type: CC BY 4.0

Affiliation: Kao Corporation (Japan), Kyushu University

Abstract

In order to establish rapid and accurate typing method for Bacillus coagulans strains which is important for controlling in some canned foods and tea-based beverages manufacturing because of the high-heat resistance of the spores and high tolerance of the vegetative cells to catechins and chemicals, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and repetitive-PCR (rep-PCR) were evaluated. For this purpose, 28 strains of B. coagulans obtained from various culture collections were tested. DNA sequence analyses of the genes encoding 16S rRNA and DNA gyrase classified the test strains into two and three groups, respectively, regardless of their phenotypes. Both MALDI-TOF MS and rep-PCR methods classified the test strains in great detail. Strains classified in each group showed similar phenotypes, such as carbohydrate utilization determined using API 50CH. In particular, the respective two pairs of strains which showed the same metabolic characteristic were classified into the same group by both MALDI-TOF MS and rep-PCR methods separating from the other strains. On the other hand, the other strains which have the different profiles of carbohydrate utilization were separated into different groups by these methods. These results suggested that the combination of MALDI-TOF MS and rep-PCR analyses was advantageous for the rapid and detailed typing of bacterial strains in respect to both phenotype and genotype.

Highlights

Rapid identification of bacterial species and typing of strains are important for the assessment of clonal relationships among environmental isolates in food and beverage processing industries, especially when bacterial spoilage occurred on their products
The discriminatory power of strain typing of B. coagulans was compared among rep-polymerase chain reaction (PCR), MALDI-TOF MS, ribotyping, and carbohydrate utilization in the point of view of the precision and rapidity
It was difficult to distinguish B. coagulans strains by DNA sequencing of housekeeping genes and the PCR band patterns of ribosomal internal transcribed spacer (ITS) region because of their small difference in DNA sequence

Summary

Introduction

Rapid identification of bacterial species and typing of strains are important for the assessment of clonal relationships among environmental isolates in food and beverage processing industries, especially when bacterial spoilage occurred on their products. As a result of the development of the polymerase chain reaction (PCR) method in the 1990’s, molecular biological methods for identification of prokaryotes were established on the basis of the nucleotide sequences of 16S ribosomal RNA gene [1]. This bacterial identification technique spread widely in order to improve the bacterial control in food and beverage manufacturing. An automated microbial typing system with repetitive sequence-based PCR (rep-PCR) [9,10,11] and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) [12, 13] have been used for identification of bacterial species and may provide potential alternative methods for strain typing. We evaluated the reliability of rep-PCR and MALDI-TOF MS as strain typing of Bacillus coagulans by the comparison of ribotyping and phenotype of carbohydrate utilization

Materials and methods

28 DSM2385

Results

Discussion