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CRISPR interference in a Streptococcus agalactiae multi-locus sequence type 17 strain

William D Cutts1, Aidan W Flanagan2, Brice K Gorman2

  • 1Department of Molecular and Cell Biology, University of Texas at Dallas, Dallas, Texas, USA.

Journal of Bacteriology
|January 14, 2026
PubMed

Insights

Researchers developed a CRISPR interference (CRISPRi) system to study Group B Streptococcus (GBS) pathogenesis. This tool allows for targeted gene knockdown in the hypervirulent ST-17 strain COH1, aiding research into neonatal meningitis.

Area of Science:

  • Microbiology
  • Genetics
  • Infectious Diseases

Background:

  • Group B Streptococcus (GBS) is a primary cause of neonatal bacterial meningitis.
  • The hypervirulent serotype III, sequence type 17 (ST-17) strain COH1 is linked to severe disease but is genetically challenging to manipulate.
  • Understanding GBS virulence factors at the blood-brain barrier (BBB) is crucial for combating neonatal meningitis.

Purpose of the Study:

  • To develop a novel CRISPR interference (CRISPRi) system for targeted gene knockdown in the GBS ST-17 COH1 strain.
  • To enable functional genomics and high-throughput screening of GBS virulence factors.
  • To investigate GBS interactions at the BBB.

Main Methods:

  • Development of a CRISPR interference (CRISPRi) system using catalytically inactivated Cas9 (dCas9) in the COH1 strain.
  • Confirmation of system efficacy via hemolysis assays and qPCR transcriptional analysis.
  • Assessment of gene knockdown effects in vitro using human brain endothelial cell infection models.

Main Results:

  • The CRISPRi system successfully achieved tunable gene expression knockdown in ST-17 GBS.
  • Phenotypic knockdowns of key virulence genes (PI-2b, srr2, iagA) were observed.
  • Reduced bacterial adhesion, invasion, and inflammatory responses at the BBB were demonstrated.

Conclusions:

  • The developed CRISPRi platform provides a versatile tool for genetic manipulation in ST-17 GBS.
  • This system facilitates rapid functional genomics and pathogenesis research for GBS.
  • The findings contribute to a better understanding of GBS virulence and potential therapeutic targets.
Keywords:
CRISPRGroup B Streptococcus

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