Natural Variant of Collagen-Like Protein A in Serotype M3 Group A Streptococcus Increases Adherence and Decreases Invasive Potential

Abstract

c Group A Streptococcus (GAS) predominantly exists as a colonizer of the human oropharynx that occasionally breaches epithelial barriers to cause invasive diseases. Despite the frequency of GAS carriage, few investigations into the contributory molecular mechanisms exist. To this end, we identified a naturally occurring polymorphism in the gene encoding the streptococcal collagen-like protein A (SclA) in GAS carrier strains. All previously sequenced invasive serotype M3 GAS possess a premature stop codon in the sclA gene truncating the protein. The carrier polymorphism is predicted to restore SclA function and was infrequently identified by targeted DNA sequencing in invasive strains of the same serotype. We demonstrate that a strain with the carrier sclA allele expressed a full-length SclA protein, while the strain with the invasive sclA allele expressed a truncated variant. An isoallelic mutant invasive strain with the carrier sclA allele exhibited decreased virulence in a mouse model of invasive disease and decreased multiplication in human blood. Further, the isoallelic invasive strain with the carrier sclA allele persisted in the mouse nasopharynx and had increased adherence to cultured epithelial cells. Repair of the premature stop codon in the invasive sclA allele restored the ability to bind the extracellular matrix proteins laminin and cellular fibronectin. These data demonstrate that a mutation in GAS carrier strains increases adherence and decreases virulence and suggest selection against increased adherence in GAS invasive isolates. T he bacterial pathogen Streptococcus pyogenes (group A Streptococcus [GAS]) causes a wide range of disease in humans. GAS causes severe invasive infections such as toxic shock syndrome and necrotizing fasciitis but is also the cause of milder, more benign infections (e.g., pharyngitis). In addition to causing disease, GAS colonizes the throats of humans in the absence of symptoms. Asymptomatic colonization rates range between 5 and 15% in healthy children (1), a rate that far exceeds that of GAS invasive disease (2). However, despite the high prevalence of GAS carriage, little is known about the molecular factors that contribute to asymptomatic colonization.Colonization of the host epithelium is a key first step to establishing GAS carriage or disease. GAS elaborates several key surface proteins that contribute to this process, including the M protein, fibronectin-binding proteins, the GAS pilus, and streptococcal collagen-like (Scl) proteins. The collagen-like protein SclA (also known as Scl1) is found in all GAS serotypes examined to date (3) and is positively regulated by the well-characterized regulator Mga (4). The SclA protein extends from the GAS cell surface in a homotrimeric, "lollipop-like" fashion (5). The outermost region consists of a globular head that varies considerably between GAS serotypes, followed by a repeating Gly-X-Y (GXY) collagen-like sequence that is in turn linked to the cell wall through a prolinerich linker region (see Fig. 1A). SclA contribu...

A Single Amino Acid Replacement in the Sensor Kinase LiaS Contributes to a Carrier Phenotype in Group A Streptococcus

“…Despite relatively modest decreases in bacterial burden, the carrier mutations resulted in large decreases in virulence. Similar patterns have been observed in GAS carrier strains with mutations affecting expression of the stand-alone regulator Mga (21) and the surface protein SclA (22). Taken together, these data indicate that it is undoubtedly the case that multiple genetic pathways contribute to GAS carriage, making individual mutations rare events.…”

Increased Pilus Production Conferred by a Naturally Occurring Mutation Alters Host-Pathogen Interaction in Favor of Carriage in Streptococcus pyogenes

“…Few studies have directly examined the bacterial molecular genetic factors that lead to the persistence of GAS in humans (15,16,18). Our findings substantially enhance understanding of the molecular mechanism underlying the carrier mutation (liaS R135G ) and its contribution to the asymptomatic carrier phenotype in GAS.…”

“…We reported a mutation unique to carrier strains in the promoter of the stand-alone regulator Mga that led to decreased transcription of mga and Mgaregulated genes, including emm, encoding M protein (18). We also identified a carrier mutation that restored the function of the GAS surface protein SclA (streptococcal collagen-like protein A) (16). In addition, we discovered that mutations eliminating GAS capsule production were the most frequently identified mutations in GAS carrier strains (15).…”

“…We have defined the carrier phenotype to be one characterized by an increased ability to adhere to epithelial surfaces and a decreased ability to cause disease, a definition supported by investigation of independent carrierspecific mutations and their contribution to the carrier phenotype (15)(16)(17)(18). The liaS R135G carrier mutation causes an increased frequency of mouse colonization in a model of nasopharyngeal infection (17).…”

“…Comparison of the whole genomes of carriage and disease-causing strains of GAS has identified mutations unique to carrier strains that contribute to GAS carriage (15)(16)(17)(18). A recurring theme that has emerged from GAS comparative genomic studies is that mutations affecting bacterial cell surface molecules alter the host-pathogen interaction and are critical to the development of carriage and a carrier phenotype.…”