# Genetic methods for studying Gardnerella virulence

> **NIH NIH R21** · UNIVERSITY OF WISCONSIN-MADISON · 2024 · $194,375

## Abstract

PROJECT SUMMARY
Bacterial vaginosis (BV) occurs in nearly one third of women of reproductive age, causing significant problems
for sexual and reproductive health. Women with BV are predisposed to pregnancy complications including
amnionitis and preterm delivery. Gardnerella spp. are major players in the BV dysbiosis, displacing
Lactobacillus spp. and forming a robust biofilm supporting the growth of multiple anaerobes. The biofilm also
serves as the infectious form for transmitting bacterial vaginosis between people. Gardnerella make a number
of putative virulence factors including multiple pili, lipases, sialidases, and the pore-forming toxin vaginolysin. It
has been difficult to study Gardnerella virulence factors due to the inability to make targeted mutations or
introduce genetic constructs. We discovered a way to make targeted insertion mutations using suicide
plasmids in Gardnerella spp., and we propose to further develop these methods to produce a full set of tools
for Gardnerella genetic manipulation. First, we will improve methods for making insertional mutants by
determining factors affecting transformation frequency and insert stability, and by developing use of a counter-
selectable marker to allow for creation of selectable unmarked mutations such as deletions and point
mutations. Furthermore, we will test different methods of insert replacement to optimize creation of unmarked
mutations. Second, we will develop methods and constructs for gene expression from the Gardnerella
chromosome in order to produce constitutive or regulated complement gene expression, transcriptional fusions
to Gardnerella ‘lacZ, or translational fusions to the FLAG epitope. The usefulness of the methods will be tested
with mutations in the vaginolysin gene.

## Key facts

- **NIH application ID:** 10866243
- **Project number:** 1R21AI182550-01
- **Recipient organization:** UNIVERSITY OF WISCONSIN-MADISON
- **Principal Investigator:** Joseph P Dillard
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $194,375
- **Award type:** 1
- **Project period:** 2024-06-01 → 2026-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10866243

## Citation

> US National Institutes of Health, RePORTER application 10866243, Genetic methods for studying Gardnerella virulence (1R21AI182550-01). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10866243. Licensed CC0.

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