Discovery and characterization of cross-reactive single-domain antibodies targeting key virulence factors of <i>Salmonella</i>and <i>Shigella</i>

Abstract Study Objective: Here, we utilize a nanobody yeast display library to identify nanobodies that target virulence factors in Shigellaspp. and Salmonella enterica typhimuriumand determine if these (Nbs) can inhibit host cell invasion vitro. Brief Statement of Methods: Using synthetic containing camelid Nbs with randomized CDR loops, selected for binders select Enterobacteriaceae proteins, including IpaD/SipD, IcsA, IcsP by MACS FACS. Lead were cloned expressed as monomeric VHH domains or Nb-Fc fusions. ELISA flow cytometry surface staining was used estimate affinity, functional activity assayed via assay Shigella flexneri, sonnei, typhimurium. Summary Results: Several high affinity isolated specificity the targe subset could bind whole bacterial cells. binned into epitopes competitive binding assays. The efficacy against specific proteins assays directly compared. Conclusions: A direct comparison high-affinity Nb various reveals only are accessible on bacteria able invasion. These results will inform development factor-based vaccines other immunotherapies. Wellcome Trust (REISTANT Consortium)