Bacteriocin production of the probiotic Lactobacillus acidophilus KS400

Abstract

In the last years, the use of probiotics, including Lactobacillus species, has received much attention to prevent and treat vaginal disorders. These species have been described as having the ability to colonize the epithelial surface and produce antimicrobial metabolites that are able to control the remaining vaginal microflora. This study aimed to identify and characterize, for the first time, a bacteriocin natively produced by Lactobacillus acidophilus KS400 (probiotic strain from Gynoflor^®-Medinova AG, Switzerland) and its antimicrobial activity against relevant urogenital pathogens. After organic acids and hydrogen peroxide neutralization in the fermented Lactobacillus acidophilus KS400 culture medium, bacteriocin activity was tested against the indicator microorganism Lactobacillus delbrueckii ATCC9649. The fermentation of Lactobacillus acidophilus KS400 for bacteriocin production was carried out in batch mode, and its antimicrobial activity, optical density and pH were monitored. After production and extraction, the bacteriocin molecular weight was estimated by electrophoresis and tested against vaginal pathogenic microorganisms. As described for other bacteriocins, batch fermentation profiles indicated that bacteriocin production occurs during the exponential growth phase of the lactobacilli, and declines during their stationary growth phase. The molecular weight of the bacteriocin is approximately 7.5 kDa. The bacteriocin containing protein extract was shown to inhibit the growth of Gardnerella vaginalis, Streptococcus agalactiae, Pseudomonas aeruginosa and the indicator strain Lactobacillus delbrueckii ATCC9649. We conclude that L. acidophilus KS400 produces bacteriocin with antimicrobial activity against relevant urogenital pathogens.

Keywords: Bacteriocins; Lactobacillus acidophilus KS400; Vaginal infections; Vaginal lactobacilli; Vaginal probiotics.

Fig. 1

Antimicrobial activity of Lactobacillus acidophilus KS400 bacteriocin against Lactobacillus delbrueckii ATCC9649. Culture extracts of Lactobacillus acidophilus KS400 tested by the agar well diffusion method described (Tagg and McGiven 1971). S1—culture medium from Lactobacillus acidophilus KS400 medium; PBS 1X—negative control corresponding to ×1 phosphate buffered saline

Fig. 2

a Batch fermentation of Lactobacillus acidophilus KS400 strain in MRSB with non-controlled pH (initial value of 6.5) at 37 °C. Left axis: absorvance (Abs) at 600 nm; Primary right axis: pH values; Secondary right axis: Relative bacteriocin activity (%); Bottom axis: Time (hours). Symbols: Optical density at 600 nm; Changes in pH values; (-grey bars-) Bacteriocin activity production. b Antimicrobial activity of Lactobacillus acidophilus KS400 strain during fermentation; Activity was tested against Lactobacillus delbrueckii ATCC9649 and plates were incubated at 37 °C; 10% CO₂

Fig. 3

Direct detection of antimicrobial activity of Lactobacillus acidophilus KS400 bacteriocin (A) Tricine-SDS-PAGE gel was observed in ChemiDoc™ MP System (Biorad) using the stain free function for protein gels: (MW) molecular mass protein standards (NZYTECH, Portugal); − β/+ β—absence and presence of β-mercaptoethanol (B) Zone of growth inhibition corresponding to the position of the antimicrobial protein (bacteriocin)