Gardnerella vaginalis and Neisseria gonorrhoeae Are Effectively Inhibited by Lactobacilli with Probiotic Properties Isolated from Brazilian Cupuaçu (Theobroma grandiflorum) Fruit

Abstract

In recent years, certain Lactobacillus sp. have emerged in health care as an alternative therapy for various diseases. Based on this, this study is aimed at evaluating in vitro the potential probiotics of five lactobacilli strains isolated from pulp of cupuaçu fruit fermentation against Gardnerella vaginalis and Neisseria gonorrhoeae. Our lactobacilli strains were classified as safe for use in humans, and they were tolerant to heat and pH. Our strains were biofilm producers, while hydrophobicity and autoaggregation varied from 13% to 86% and 13% to 25%, respectively. The coaggregation of lactobacilli used in this study with G. vaginalis and N. gonorrhoeae ranged from 15% to 36% and 32% to 52%, respectively. Antimicrobial activity was present in all tested Lactobacillus strains against both pathogens, and the growth of pathogens in coculture was reduced by the presence of our lactobacilli. Also, all tested lactobacilli reduced the pH of the culture, even in incubation with pathogens after 24 hours. The cell-free culture supernatants (CFCS) of all five lactobacilli demonstrated activity against the two pathogens with a halo presence and CFCS characterization assay together with gas chromatography revealed that lactic acid was the most abundant organic acid in the samples (50% to 62%). Our results demonstrated that the organic acid production profile is strain-specific. This study revealed that cupuaçu is a promising source of microorganisms with probiotic properties against genital pathogens. We demonstrated by in vitro tests that our Lactobacillus strains have probiotic properties. However, the absence of in vivo tests is a limitation of our work due to the need to evaluate the interaction of our lactobacilli with pathogens in the vaginal mucosa. We believe that these findings may be useful in developing a product containing our lactobacilli and their supernatants in order to support with vaginal health.

Figure 1

Heat resistance standard of lactobacilli isolated from cupuaçu fermentation. Lc24: L. casei 24; Lf38: L. fermentum 38; Lf47: L. fermentum; Lp81: L. plantarum 81; Lp90: L. plantarum 90. ^∗∗∗Statistically significant differences compared to control (P < 0.001). Presented values represent the mean and the standard deviation from triplicate determinations.

Figure 2

Growth of lactobacilli in different pH ranges: (a) L. casei 24; (b) L. fermentum 38; (c) L. fermentum; (d) L. plantarum 81; (e) L. plantarum 90. Presented values represent the mean and the standard deviation from triplicate determinations.

Figure 3

Growth inhibition of pathogens by lactobacilli after 24 h of culture. The growth of pathogenic microorganism is expressed as log₁₀ CFU.mL⁻¹. Each value corresponds to the mean ± standard deviation of triplicate determinations. Control: (Gv) G. vaginalis or (Ng) N. gonorrhoeae. The different lactobacilli isolates are represented by their respective numbers: (a) represents coculture of G. vaginalis with lactobacilli; (b) represents coculture of N. gonorrhoeae with lactobacilli. ^∗Statistically significant differences compared to control (P < 0.05). ^∗∗∗Statistically significant differences compared to control (P < 0.001). Presented values represent the mean and the standard deviation from triplicate determinations.

Figure 4

Hydrogen peroxide levels in CFCS of lactobacilli isolated from cupuaçu fermentation. The standard curve (R² = 0.9927) was performed together with the experimental samples in a controlled environment protected from light. Presented values represent the mean and the standard deviation from triplicate determinations.