The development of a multiplexed Soluble Phage Array (SPAr) for the detection of zoonotic pathogens

Serosurveillance is a valuable approach to disease monitoring in animal populations to facilitate effective intervention and containment strategies. It has been suggested that effective serosurveillance of zoonotic diseases could provide an early-warning system for the potential emergence of a zoonotic pathogen in human populations. However, to date, serosurveillance is somewhat limited by a single-pathogen focus and lacks the ability to distinguish between very closely related pathogens. Therefore, there is a need for an affordable, accessible, multiplexed approach with very high specificity to monitor for a wide range of established and emerging pathogens. The huge diversity of phage display peptide libraries combined with the screening power of next generation sequencing (NGS) in a process termed next generation phage display (NGPD) can be applied to discover large panels of peptide mimotopes recognised by antibody responses to disease. We propose that a phage-peptide sub-library enriched to contain multiple infection-specific mimotopes can itself be used as the assay 'antigen' to diagnose infection with a particular pathogen; an assay format we have termed Soluble Phage Array (SPAr). This assay would measure recognition of the panel of infection-specific phage-displayed mimotopes by antibodies in serum samples by NGS analysis of phage that are bound by the antibodies. We hypothesise that NGPD can be used to develop SPAr assays that can accurately identify infection with zoonotic pathogens and that these can be multiplexed with high accuracy and reproducibility. This proof-of-concept study will focus on four emergent zoonotic viruses that affect sheep or horses. These are louping ill (LIV), tick-borne encephalitis (TBEV) and Rift Valley fever (RVFV) viruses in sheep, and West Nile virus (WNV) in horses. The selected viruses represent the genera Flavivirus (LIV, TBEV and WNV) and Phlebovirus (RVFV). These viruses have been carefully chosen to represent the complexity of serosurveillance of emerging viral zoonoses in order to demonstrate the potential of the SPAr assay. A multiplexed SPAr assay would have considerable advantages over existing serosurveillance approaches. It would not be limited to a single-pathogen approach and would have the ability to distinguish very closely related pathogens, a property vital to inform effective control strategies for zoonotic pathogens.