Molecular Routes to Specific Identification of the Lactobacillus Casei Group at the Species, Subspecies and Strain Level.

Piotr Jarocki,Agnieszka Glibowska,Anna Wzorek,Agnieszka Korzeniowska-Kowal,Monika Kordowska-Wiater,Michał Dworniczak,Monika Pytka,Elwira Komoń-Janczara

doi:10.3390/ijms21082694

Abstract

The genus Lactobacillus includes, among others, Lactobacillus casei, Lactobacillus paracasei and Lactobacillus rhamnosus, species that are collectively referred to as the Lactobacillus casei group. Many studies have shown that strains belonging to this group may decrease lactose intolerance, the effects of inflammatory bowel disease, diarrhea, constipation, food allergies and even colon cancer. Moreover, evidences exists of positive effects of these bacteria on mucosal immunity and blood cholesterol level. Because of their beneficial influence on human health, many of them are used as food additives and probiotic pharmaceuticals. It should be stressed that health-promoting properties are not attributed at the species level, but to specific strains. Therefore, procedures are necessary to allow specific identification at each phylogenetic level—genus, species and strain. In this paper we present a practical overview of molecular methods for the identification and differentiation of L. casei bacteria. The research included 30 bacterial strains belonging to three species: L.casei, L. paracasei and L. rhamnosus. Among the tested procedures were genus- and species-specific PCR, multiplex-PCR, Real-Time HRM analysis, RFLP-PCR, rep-PCR, RAPD-PCR, AFLP-PCR, and proteomic methods such as MALDI-TOF MS typing and SDS-PAGE fingerprinting. The obtained results showed that multiplex-PCR and MALDI-TOF MS turned out to be the most useful methods to identify the tested bacteria at the species level. At the strain level, the AFLP-PCR method showed the highest discriminatory power. We hope that the presented results will allow for the easy selection of an appropriate procedure, depending on the experiment conducted and the equipment capabilities of any given laboratory.

Highlights

The Lactobacillus genus belongs to a very large, heterogeneous group of lactic acid bacteria (LAB), that are commonly applied in the food and pharmaceutical industries and in agriculture [1,2]
L. casei specific primers (Table S1) [17] were used in the PCR reaction. Such an experimental system seems to be useful when isolating new bacterial strains, and when it is necessary to quickly and confirm that they belong to the L. casei group
No positive result was obtained for strain Japan Collection of Microorganisms (JCM) 8677, which may suggest that the isolate was misclassified as L. casei

Summary

Introduction

The Lactobacillus genus belongs to a very large, heterogeneous group of lactic acid bacteria (LAB), that are commonly applied in the food and pharmaceutical industries and in agriculture [1,2]. The LAB includes, among others, Lactobacillus casei, Lactobacillus paracasei and Lactobacillus rhamnosus species, that are collectively referred to as the Lactobacillus casei group. These microorganisms are associated with habitats that are rich in nutrients such as fermented dairy products. They are common inhabitants of the human gastrointestinal and urogenital tracts. Many studies show that strains belonging to L. casei group may induce physical health benefits in humans and livestock. The use of probiotics in animal feed improves the effectiveness of agricultural production, as it results in the increased digestibility of feedstuffs and synthesis of some vitamins, thereby increasing body weight gain, as well as improving the health of animals, increasing their resistance to stress conditions, and ensuring faster recovery from disease [7,8]

Methods

Results

Conclusion