# Comparison of the inhibitory effects of Lactobacillus supernatant and coculture on Gardnerella vaginalis

## TL;DR

This study compares how Lactobacillus supernatant and direct coculture affect the growth of Gardnerella vaginalis, a bacterium linked to vaginal infections.

## Contribution

The study introduces a direct qPCR method to measure G. vaginalis inhibition in cocultures, revealing differences in inhibitory effects compared to supernatant assays.

## Key findings

- NYC-III broth significantly enhanced the growth of G. vaginalis and most Lactobacillus strains.
- A direct qPCR method was developed to measure G. vaginalis inhibition in cocultures without DNA extraction.
- Coculture experiments showed different inhibition levels than supernatant assays for some Lactobacillus strains.

## Abstract

Vaginal lactobacilli play a crucial role in inhibiting bacteria such as Gardnerella vaginalis (G. vaginalis), a key contributor to dysbiosis and bacterial vaginosis. We aimed to compare the inhibitory effects of Lactobacillus cell-free supernatants on G. vaginalis growth with those observed in Lactobacillus-G. vaginalis coculture, an experimental setup that more closely mimics in vivo conditions.

To identify the optimal medium for coculture experiments, MRS broth and NYC-III broth were compared. NYC-III significantly enhanced the growth of G. vaginalis and four of the five tested Lactobacillus strains. We then developed a direct quantitative PCR (qPCR) method that allowed us to specifically measure G. vaginalis genome concentrations in cocultures with Lactobacillus. This direct qPCR did not require DNA extraction and had a 4,096-fold dynamic range. We then assessed the inhibition of G. vaginalis growth by Lactobacillus cell-free supernatants in G. vaginalis cultures by measuring optical density and the Lactobacillus-mediated inhibition in cocultures by the G. vaginalis specific direct qPCR. The two measurement methods showed different levels of inhibitory activity for two of the five Lactobacillus strains tested. These findings suggest that coculture experiments should be conducted in place of, or in addition to, supernatant-based inhibitory assays.