Characterization of a novel
covS
SNP identified in Australian group A
Streptococcus
isolates derived from the M1
UK
lineage
Authors: Richter Johanna , Cork Amanda J. , Ong Yvette , Keller Nadia , Hayes Andrew J. , Schembri Mark A. , Jennison Amy V. , Davies Mark R. , Schroder Kate , Walker Mark J. , Brouwer Stephan 
Published: December 2024

Group A
Streptococcus
(GAS) is a human-adapted pathogen responsible for a variety of diseases. The GAS M1
UK
lineage has contributed significantly to the recently reported increases in scarlet fever and invasive infections. However, the basis for its evolutionary success is not yet fully understood. During the transition to systemic disease, the M1 serotype is known to give rise to spontaneous mutations in the control of virulence two-component regulatory system (CovRS) that confer a fitness advantage during invasive infections. Mutations that inactivate CovS function result in the de-repression of key GAS virulence factors such as streptolysin O (SLO), a pore-forming toxin and major trigger of inflammasome/interleukin-1β-dependent inflammation. Conversely, expression of the streptococcal cysteine protease SpeB, which is required during initial stages of colonization and onset of invasive disease, is typically lost in such mutants. In this study, we identified and characterized a novel
covS
single nucleotide polymorphism detected in three separate invasive M1
UK
isolates. The resulting CovS
Ala318Val
mutation caused a significant upregulation of SLO resulting in increased inflammasome activation in human THP-1 macrophages, indicating an enhanced inflammatory potential. Surprisingly, SpeB production was unaffected. Site-directed mutagenesis was performed to assess the impact of this mutation on virulence and global gene expression. We found that the CovS
Ala318Val
mutation led to subtle, virulence-specific changes of the CovRS regulon compared to previously characterized
covS
mutations, highlighting an unappreciated level of complexity in CovRS-dependent gene regulation. Continued longitudinal surveillance is warranted to determine whether this novel
covS
mutation will expand in the M1
UK
lineage.

IMPORTANCE

The M1
UK
lineage of GAS has contributed to a recent global upsurge in scarlet fever and invasive infections. Understanding how GAS can become more virulent is critical for infection control and identifying new treatment approaches. The two-component CovRS system, comprising the sensor kinase CovS and transcription factor CovR, is a central regulator of GAS virulence genes. In the M1 serotype,
covRS
mutations are associated with an invasive phenotype. Such mutations have not been fully characterized in the M1
UK
lineage. This study identified a novel
covS
mutation in invasive Australian M1
UK
isolates that resulted in a more nuanced virulence gene regulation compared to previously characterized
covS
mutations. A representative isolate displayed upregulated SLO production and triggered amplified interleukin-1β secretion in infected human macrophages, indicating an enhanced inflammatory potential. These findings underscore the need for comprehensive analyses of
covRS
mutants to fully elucidate their contribution to M1
UK
virulence and persistence.