Characterization of mAbs against Klebsiella pneumoniae type 3 fimbriae isolated in a target-independent phage display campaign

• Published in: Microbiology spectrum 2024-08, Vol.12 (8), p.e0040024
• Main Authors: Berry, Sophia K, Rust, Steven, Irving, Lorraine, Bartholdson Scott, Josefin, Weinert, Lucy A, Dougan, Gordon, Christie, Graham, Warrener, Paul, Minter, Ralph, Grant, Andrew J
• Format: Article
• Language: English
• Subjects: Adhesins, Bacterial; Animals; Antibodies, Bacterial - immunology; Antibodies, Monoclonal - immunology; Bacteriology; Cell Surface Display Techniques; Fimbriae Proteins - immunology; Fimbriae, Bacterial - immunology; Humans; Klebsiella Infections - immunology; Klebsiella Infections - microbiology; Klebsiella pneumoniae; Klebsiella pneumoniae - immunology; Macrophages - immunology; Macrophages - microbiology; Mice; monoclonal antibody; MrkA; Peptide Library; Phagocytosis; Research Article; scFv; type 3 fimbriae
• ISSN: 2165-0497; 2165-0497
• DOI: 10.1128/spectrum.00400-24

We used phage display, antibody engineering, and high-throughput assays to identify antibody-accessible targets of . We report the discovery of monoclonal antibodies (mAbs) binding to type 3 fimbrial proteins, including MrkA. We found that anti-MrkA mAbs were cross-reactive to a diverse panel of clinical isolates, representing different O-serotypes. mAbs binding to MrkA have previously been described and have been shown to provide prophylactic protection, although only modest protection when dosed therapeutically in a murine lung infection model. Here, we used a combination of binding and opsonophagocytic killing studies using a high-content imaging platform to provide a possible explanation for the modest therapeutic efficacy reported in that model. Our work shows that expression of type 3 fimbriae in culture is not homogenous within a bacterial population. Instead, sub-populations of bacteria that do, and do not, express type 3 fimbriae exist. In a high-content opsonophagocytic killing assay, we showed that MrkA-targeting antibodies initially promote killing by macrophages; however, over time, this effect is diminished. We hypothesize the reason for this is that bacteria not expressing MrkA can evade opsonophagocytosis. Our data support the fact that MrkA is a conserved, immunodominant protein that is antibody accessible on the surface of and suggest that additional studies should evaluate the potential of using anti-MrkA antibodies in different stages of infection (different sites in the body) as well as against biofilms in the body during infection and associated with medical devices.IMPORTANCEThere is an unmet, urgent need for the development of novel antimicrobial therapies for the treatment of infections. We describe the use of phage display, antibody engineering, and high-throughput assays to identify antibody-accessible targets of . We discovered monoclonal antibodies (mAbs) binding to the type 3 fimbrial protein MrkA. The anti-MrkA mAbs were found to be highly cross-reactive, binding to all strains tested from a diverse panel of clinical isolates, and were active in an opsonophagocytic killing assay at pM concentrations. MrkA is important for biofilm formation; thus, our data support further exploration of the use of anti-MrkA antibodies for preventing and/or controlling in biofilms and during infection.