Functional Characterization of Bacteriophage Resistant Mutants of Probiotic B. coagulans

Kavita R Pandey,Babu V Vakil

doi:10.4172/2476-2059.1000106

Abstract

Bacillus coagulans, a probiotic phage sensitive organism was subjected to random mutagenesis and several phage resistant mutants were isolated. Evaluation of mutants, in line with the WHO/FAO guidelines for probiotics, showed that Lactic acid production was not affected. Maximum sporulation efficiency was seen in mutant MIII (81% ± 2). Mutant MII showed least acid tolerance and tolerance to bile salts was lower in all the mutants compared to the parent (76% ± 0.5). All the cultures exhibited similar antibacterial activity (zones: 13-15 mm) against the pathogens used. For all the mutants bile salt hydrolase activity was positive and antibiotic susceptibility was similar to parental culture and all these mutants showed no hemolysis. Auto-aggregation potential of the mutants was somewhat higher than the parental strain except for mutant PIII (29.5% ± 1.2) whereas mutant PIII displayed highest co-aggregation potential (52% ± 0.7). MATS assay showed that mutant PIII had highest affinity for xylene (65% ± 2.1) and mutant MV for chloroform (66.1 ± 0.8%). Adhesion to Caco2 cell was highest for mutant MIII (20.4%). Comparison of the lipid profiles showed that Alpha-linoleic acid was produced only by the parent whereas PUFA like EPA was produced by only mutants PIII and MII. Mutants PIII also produced DHA. Parental strains as well as all the 7 mutants were found to be negative for siderophore production whereas Mutants MV showed highest anti-oxidative potential (32.3% ± 1.6). Thus, thorough functional characterization of the phage resistant mutants as probiotics coupled with evaluation for additional desirable attributes has helped in better understanding of these mutants in terms of their true potential.

Highlights

Probiotics have been defined jointly by FAO and WHO as “Live microorganisms which when administered in adequate amounts confer a health benefit to the host” [1]
The original phase sensitive parent culture as well as phage resistant mutants were maintained on glucose yeast extract agar (GYEA) slants and were preserved in 15% glycerol, at -20°C
The property of lactic acid production is the key to effectiveness of the probiotics in the local microflora environment of gut as it suppresses growth of many pathogenic or undesirable bacteria

Summary

Introduction

Probiotics have been defined jointly by FAO and WHO as “Live microorganisms which when administered in adequate amounts confer a health benefit to the host” [1]. The phage resistant mutants need to be characterized as per FAO/WHO guidelines for probiotic attributes like tolerance to acids and bile salt, lactic acid production, sporulation efficiency, adhesion and aggregation potentials, susceptibility to antibiotics, production of antimicrobial compounds, presence of bile salt hydrolase and hemolytic activity. Preliminary studies Lactic acid productivity, tolerance to acids and bile salts, sporulation efficiencies and aggregation potentials of the parental probiotic B. coagulans and its 7 phage resistant mutants showed variation as discussed below.

Results

Conclusion