Abstract:
Streptococcus pyogenes (group A Streptococcus, GAS) is an important human pathogen responsible for a wide range of diseases and high health burden worldwide. The ability of GAS to cause infections is attributed to the large arsenal of virulence factors expressed by this human exclusive pathogen. Pili are long filamentous protrusions that extend from the bacterial cell surface. After their recent discovery in GAS, multifaceted roles of pili in host-pathogen interactions have been reported, such as epithelial cell adhesion and biofilm formation. The immunogenic pilus proteins have also been shown to play a role in bacterial pathogenesis in several studies using animal models. The genes encoding the pilus proteins and assembly enzymes are clustered in the Fibronectin-binding, Collagen-binding, T antigen (FCT) region, of which 9 variants with diverse gene organisation and sequences have been identified so far, but only a few types have been studied. Pilus components of FCT type 6 produced by M2 serotype GAS are mutually distinctive from those of other better understood FCT types, as revealed by phylogenetic analyses of the FCT region nucleotide sequences and the amino acid sequences of pilus proteins. The uniqueness in FCT-region sequence is due to high inter- and intra-species gene recombination, and may indicate functional distinctiveness of this pilus. Using a combination of FCT-6 component GAS knockout mutants and L. lactis gain-of-function mutants, this study characterises the distinguishing features of this pilus, and illustrates the incredible complexity in host-pathogen interactions. The two ancillary subunits of FCT-6 pilus exhibit distinct roles on pilus assembly, which also agrees with their hypothetical locations at either the tip or the base of the pilus structure. The multiple sortases expressed by the FCT-6 region show functional redundancy on pilus processing, as deletion of a single sortase does not affect pilus formation. Despite the sequence similarity of certain pilus components between GAS FCT-6 pilus and group B Streptococcus (GBS) pilus island 1 (PI-1) pilus, the FCT-6 pilus does not share the same binding tropism with GBS. The epithelial cell adherence property of FCT-6 pilus is surprisingly attributed to the backbone pilin (BP, Spy0109), rather than the AP1 subunit commonly recognised as the pilus adhesin. The most striking feature of the FCT-6 pilus is the attributes of the backbone pilin. Unlike other backbone pilins described thus far, the backbone pilin of FCT-6 was observed to bind epithelial cells, human plasma proteins fibrinogen and fibronectin, and interact with peripheral blood monocytes. The interaction with these host components may be related to the transition from localised colonisation to invasive dissemination, and involved in immune evasion mechanisms. Pilus-mediated protection was seen in whole blood killing assays and macrophage bactericidal assays, and evidence of FCT-6 pilus-associated superior bacterial fitness and survival was also observed in a wax worm infection model. These observations collectively confirm the importance of FCT-6 pilus in GAS virulence, and provide new information on pili’s role in GAS pathogenesis. This study provides insights into one of the least studied GAS pilus. The unusual features revealed by the results not only confirm the uniqueness of the FCT-6 pilus, but also highlight the complicacy of the virulent strategies utilised by the pathogen. Certainly more information is needed for comprehensive understanding of this important bacterium, in order to develop advanced and successful preventive and therapeutic approaches to manage GAS infections in the future.