Bacillus subtilis with a 1% population enhances the activity of Enterococcus faecalis with a 99% population

Abstract

Microbes are present as communities in the environment. However, the importance of minor populations has not been well studied experimentally. In this study, we evaluated the role of Bacillus subtilis with a 1% population and its effect on co-incubated Enterococcus faecalis with a 99% population. Here we used an azo dye-decolorizing Enterococcus faecalis strain T6a1 and non-dye-decolorizing Bacillus subtilis strain S4ga. The dye decolorization assay enabled the investigation of the effects of B. subtilis S4ga on the activity of E. faecalis T6a1, even when B. subtilis S4ga was present at only 1% relative abundance or lower. We found that non-decolorizing B. subtilis S4ga enhanced the dye decolorization activity of E. faecalis T6a1, shortened the lag time of E. faecalis T6a1 to start decreasing the dye concentration, and increased the time for E. faecalis T6a1 to continue dye decolorization. These effects were correlated with redox potential values. We compared the extracellular amino acids between each incubation culture of E. faecalis T6a1 and B. subtilis S4ga, which revealed their mutual relationship by cross-feeding of specific amino acids. We also compared the intracellular primary metabolites between co-incubation and sole incubation of E. faecalis T6a1. The arginine deiminase (ADI) pathway in the co-incubated E. faecalis T6a1 was activated compared to that of E. faecalis T6a1 incubated solely. These findings explained that co-incubation with B. subtilis S4ga promoted ATP production in E. faecalis T6a1 cells to a greater extent and enhanced dye-decolorization activity. IMPORTANCEThis study highlights the importance of minor bacterial populations and their effects on major populations. We used Enterococcus faecalis as the major population and Bacillus subtilis as the minor population. Both species are becoming increasingly important. Some strains of E. faecalis are antibiotic-resistant pathogens, show probiotic effects, or are applicable for textile wastewater treatment. Some strains of B. subtilis are known to produce antimicrobial agents, reduce intestinal inflammation, or restore gut microbiota. We demonstrated that a low abundance of B. subtilis with 1% population increased the amount of energy produced by E. faecalis with 99% population, which appeared as enhanced dye decolorization activity of dye-decolorizing E. faecalis. Metabolomic analysis suggested that E. faecalis and B. subtilis had a mutual relationship by feeding specific amino acids to each other. These results provide new insights into co-existing minor populations in microbial communities and will improve our understanding of bacterial control. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=155 SRC="FIGDIR/small/462685v2_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@950ce8org.highwire.dtl.DTLVardef@1826200org.highwire.dtl.DTLVardef@f02e5forg.highwire.dtl.DTLVardef@749e07_HPS_FORMAT_FIGEXP M_FIG C_FIG