Interaction of S. suis with endothelium/epithelium during invading Blood-Central Nervous System barriers and characterization of putative virulence factors

Streptococcus (S.) suis is a globally emerging zoonotic pathogen that can be found in the upper respiratory tract, vagina, and alimentary tract of pigs. S. suis is most abundant on the palatine tonsils, which are the main portal of entry in cases of systemic disease. S. suis is an important cause of invasive disease in pigs causing significant economic losses to the swine industry worldwide and increased use of antibiotics as prophylactic and metaphylactic interventions. S. suis also is a frequent cause of infectious disease outbreaks in humans exposed to infected pigs or contaminated pork products. In pigs and humans, S. suis invasive disease is commonly associated with acute sepsis, meningitis, and arthritis but other sequelae have been reported. To cause meningitis in pigs and humans, S. suis must cross epithelial barriers, persist in the bloodstream, disseminate to the brain through the blood vessels, and invade the blood-central nervous system barrier (BCNSB) comprising the blood-brain barrier (BBB) and blood-cerebrospinal fluid barrier (BCSFB). The invasion of S. suis into the BCNSB is regarded as the last and the most pivotal step to induce lesions to the brain and neuropathological symptoms. It is hypothesised that yet unidentified virulence factors contributing to the bacterial crossing of the BCNSB must be present on the exposed outer surface of S. suis. To investigate the translocation of S. suis and associated virulence factors across brain barriers, researchers have exploited transformed or cancer cell lines as in vitro models. These models facilitate investigation of the mechanism of pathogenesis and replace the use of animals but have some limitations. Cancer or transformed cells often exhibit aneuploidy or chromosomal rearrangements and are known to evolve in tissue culture. The main objectives of our study are: (i) to develop BCNSB models that more closely mimic the in vivo tissues for studying interactions of S. suis with the BBB and BCSFB, and (ii) to gain a better understanding of the mechanisms by which S. suis crosses the BBB and BCSFB. For (i) we aim to develop BBB models using brain microvascular endothelial cells derived from human and porcine induced pluripotent stem cells (iPSCs) and build choroid plexus (CP) BCSFB models using iPSC-derived CP epithelial cells, or CP organoids generated from human and porcine iPSCs or adult stem cells in the tissue. In (ii) we will identify crucial virulence factors that mediate S. suis invasion of the BCNSB by constructing knock-out mutants of candidate virulence factors. In addition, we will specifically test a hypothetical pathway for S. suis to invade the meninges through the BCSFB in the subarachnoid space (SAS). The results will contribute to our understanding of the molecular mechanisms of S. suis invasion of the BCNSB and have implications for the development of novel prevention and treatment strategies such as vaccine antigens critical to pathogenesis.