# The Role of Cas9 in Group B Streptococcal Colonization and Disease

> **NIH NIH F32** · UNIVERSITY OF COLORADO DENVER · 2021 · $68,562

## Abstract

PROJECT SUMMARY
Group B Streptococcus (GBS) remains the leading etiologic agent of neonatal bacterial meningitis and a major
opportunistic pathogen in certain adult populations, including pregnant women. During pregnancy, GBS
asymptomatically colonizes the vaginal tract of 20-30% of healthy women, but can be transmitted to the
neonate in utero or during birth resulting in neonatal meningitis upon GBS disruption of the blood-brain barrier
(BBB) and 10-15% mortality, regardless of antibiotic treatment. Despite this major public health concern, the
specific
mechanisms
and
regulation
of
Group
B
streptococcal
effectors
that
mediate
successful colonization
and development of meningitis remain unknown. Recently, Cas9, an endonuclease traditionally known for its
role in CRISPR/Cas bacterial adaptive immunity, was shown to attenuate virulence and the interaction of
pathogens with host cells through endogenous gene regulation. However, the role of Cas9 in the colonization
and pathogenesis of other pathogens, such as GBS, has not been well-studied. My preliminary work shows
that GBS cas9 mutants are attenuated in vivo using models of GBS vaginal colonization and hematogenous
meningitis and invade brain endothelium and vaginal epithelium significantly less than WT GBS, in vitro.
Further, RNA-Seq analysis indicated that Cas9 globally regulates gene expression in GBS, including
dysregulation of putative virulence factors, metabolic and cell wall formation factors, and riboflavin synthesis
machinery. Based on my preliminary data, this proposal seeks to elucidate the molecular mechanisms by
which Cas9 globally regulates GBS factors, such as the riboflavin pathway, and to determine how dysregulated
riboflavins might affect GBS evasion of host immunity during vaginal colonization. These questions will be
addressed with both in vitro and in vivo models of GBS vaginal colonization and BBB penetration in the
following specific aims: AIM 1: Identify components of the CRISPR/cas operon and Cas9 residues that
contribute to Cas9-mediated gene regulation of GBS factors; AIM 2: Determine if Cas9 regulates global GBS
gene expression at the genomic or transcriptomic level; AIM 3: Characterize the role of Cas9-regulated
riboflavin synthesis in GBS immune evasion. This proposal is the first to examine both the mechanism of Cas9-
regulation of GBS factors and the role of these factors in host immune response to GBS, which may ultimately
afford novel targets and alternative therapeutic strategies.

## Key facts

- **NIH application ID:** 10220672
- **Project number:** 5F32AI143203-02
- **Recipient organization:** UNIVERSITY OF COLORADO DENVER
- **Principal Investigator:** Brady L Spencer
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $68,562
- **Award type:** 5
- **Project period:** 2020-07-01 → 2022-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10220672

## Citation

> US National Institutes of Health, RePORTER application 10220672, The Role of Cas9 in Group B Streptococcal Colonization and Disease (5F32AI143203-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10220672. Licensed CC0.

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