Quantification of Lactobacillus spp. of interest for the study of the vaginal microbiota

The optimal vaginal microbiota is dominated by Lactobacillus spp., particularly Lactobacillus crispatus or Lactobacillus gasseri. Lactic acid production by Lactobacillus inhibits the growth of other bacterial species. Bacterial vaginosis (BV) is characterized by depletion of vaginal Lactobacillus and treatment with current choice antibiotics often fails urging novel alternatives including probiotics or prebiotics. Testing new components for BV treatment often utilize in vitro quantification of Lactobacillus. Thus, this study aimed to compare methods for L. crispatus and L. gasseri quantification. Type strains were cultured in Yeast extract/peptone/tryptone/Tween 80/glucose (LAPTg) and New York City III (NYCIII) broth media, and their glucose-restricted preparations (LAPTgr, NYCIIIgr). Bacterial growth was assessed by measurements of optical density at 540 nm (OD₅₄₀) and plate counting (PC) (CFU/mL) at 0 h, 8 h, 12 h, and 24 h. Measurements of bacterial growth at all timepoints were compared with ANOVA at 5 % significance level. Higher OD₅₄₀ reads were observed in LAPTg (p < 0.05) and NYCIII (p < 0.05) with L. crispatus and L. gasseri when compared to LAPTgr and NYCIIIgr, respectively. However, PC did not differ between LAPTg and LAPTgr (p = 0.628 for L. crispatus; p = 0.161 for L. gasseri). In NYCIII, only L. crispatus exhibited higher PC in NYCIII when compared to NYCIIIgr (p = 0.016). These findings suggest that PC does not reflect Lactobacillus spp. counts under the tested conditions. Future studies exploring BV treatments should combine OD measurements with methods for Lactobacillus quantification other than PC, e. g. molecular methods.