Efficacy of Lactiplantibacillus plantarum PBS067, Bifidobacterium animalis subsp. lactis BL050, and Lacticaseibacillus rhamnosus LRH020 in the Amelioration of Vaginal Microbiota in Post-Menopausal Women: A Prospective Observational Clinical Trial

Abstract

The menopausal transition marks a significant physiological shift in women. Menopause-related symptoms can significantly affect a woman's quality of life and probiotics have emerged as a promising avenue. This study aims to investigate the benefits of probiotics in improving vaginal well-being and microbiota composition in post-menopausal women. A prospective observational clinical trial was carried out enrolling 50 post-menopausal healthy women, aged between 45 and 65 years old, taking a supplement containing Lactiplantibacillus plantarum PBS067, Bifidobacterium animalis subsp. lactis BL050, and Lacticaseibacillus rhamnosus LRH020 (3B CFU/day) for 28 days. Vaginal swabs were collected to evaluate microbiota fluctuation and the inflammatory pattern was recorded. A Vaginal Health Index was provided to evaluate vaginal well-being throughout the trial. Clinical outcomes revealed a decrease in menopausal symptoms. Significant improvements were observed across various parameters: a 50% enhancement in the VHI score (p < 0.0001), alongside substantial reductions in inflammatory cytokine levels. An 87.8% decrease in IL-6, 57.6% in IL-1β, and 40.8% in TNF-α was observed (p < 0.05). Moreover, the probiotic intervention facilitated the restoration of vaginal microbiota, evidenced by an increase in lactobacilli abundance. In conclusion, the combination of these specific probiotic strains, previously clinically tested in childbearing-age women, showed to be effective also for post-menopausal women.

Keywords: genitourinary syndrome of menopause; immunity; lactobacilli; menopause; pro-inflammatory cytokines; probiotics; vaginal atrophy; vaginal microbiota.

Figure 1

Timeline of the study.

Figure 2

Violin plots with box-and-whisker plots showing the comparison of alpha diversity measures between menopausal women at the beginning (T0, n = 43, in light blue) and at the end of the probiotic treatment (T1, n = 43, in blue). Observed = Observed Species; Phylogenetic_Diversity = Phylogenetic Diversity Whole Tree; Shannon = Shannon–Wiener index; InvSimpson = Inverse Simpson’s index. Median, first, third quartile and outliers are shown.

Figure 3

Principal Coordinate Analysis (PCoA) on unweighted and weighted UniFrac distance metric calculated on menopausal women at T0 (n = 43, light blue dots) and T1 (n = 43, blue dots). Each sample is represented by a dot. Axis 1 explained 17.2% and 43.3% of the variation observed, while Axis 2 explained 7.3% and 17.2% of the variation, in the left and right graph, respectively; dashed light blue ellipses, for T0 data, or blue ellipses, for T1 data, were calculated on the cluster of the sample data using the function ‘stat_ellipse’ and assuming a multivariate t–distribution; PERMANOVA on weighted and unweighted UniFrac Pr(>F) = n. s. (not significant).

Figure 4

Heat map of the core microbiome in the vaginal samples based on hierarchical clustering of bacterial species relative abundance (RA%) with an alluvial plot of the Community State Type changes in time. RA% color scale is indicated on the right with white corresponding to 0% and dark blue corresponding to 100%. (A) The vaginal microbiota at T0, on the right, can be divided into 4 clusters, corresponding to 4 different Community State Types (CSTs): CST I (n = 6), CST III (n = 7), CST IV (n = 25), and CST IV-C3 (n = 5). The vaginal microbiota can be divided into 5 clusters at T1, on the left: CST I (n = 7), CST III (n = 9), CST IV (n = 20), CST IV-C3 (n = 6), and CST V (n = 1). (B) Alluvial plot of the CST changes per subject of the study before (T0) and after treatment (T1).

Figure 5

LDA LEfSe barplot displaying the different associations of bacterial genera between untreated (T0, in red) and treated menopausal women (T1, in green); the latter is not visible in the panel since the comparison demonstrated a significant association only at T0 (LDA score > 2.0). Box-and-whisker plots with data points show the relative abundances of the Staphylococcus genus in the two groups. Mann–Whitney U Test result of the group comparison is shown in the figure. * p < 0.05.

Figure 6

(a) Total VHI score comparison between T0 and T1. (b) A single score of the parameters is included in the VHI evaluation. Results are expressed as mean ± SD. **** p < 0.0001.

Figure 7

pH ranges of the vaginal environment of women enrolled at the beginning of the study (T0) and after 4 weeks of probiotic supplementation (T1).

Figure 8

Vaginal concentrations of the interleukins were investigated in this study, comparing the conditions at the beginning of the study (T0) and after the probiotic treatment (T1). Results are expressed as mean ± SD. * p < 0.05.