Synergistic activity of a KL51-depolymerase and a <i>Sugarlandvirus</i> bacteriophage against ST16 <i>Klebsiella pneumoniae</i>

The rising prevalence of multidrug-resistant <i>Klebsiella pneumoniae</i> strains, particularly the emerging high-risk ST16 lineage, highlights the urgent need for alternative therapeutic strategies. A major barrier to effective bacteriophage (phage) therapy against <i>K. pneumoniae</i> is its polysaccharide capsule, which hinders phage access to surface receptors. In this study, we identified and characterized an anti-KL51 capsule depolymerase derived from the anti-ST16 phage PWKp9B. The recombinant DpoK51-9B enzyme was expressed in <i>Escherichia coli</i>, purified, and shown to degrade the KL51 capsule using Percoll density gradient, halo formation on double-layered-agar, and FITC-dextran exclusion assays. The depolymerase exhibited strong activity at nanomolar concentrations, retained function up to 50°C, and was active only against KL51-positive strains, including ST16 and ST231 isolates, but not against strains with other capsule types. Importantly, the depolymerase enabled the lytic activity of the <i>Sugarlandvirus</i> phage GPH82, normally inactive against encapsulated ST16, demonstrating potent phage-depolymerase synergy. These findings highlight the potential of recombinant depolymerases to expand phage host range by unmasking the phage receptors on the surface of the bacteria and therefore enhance the efficacy of phages against encapsulated bacteria. IMPORTANCE Bacteriophage therapy and phage-derived therapeutics are attracting increasing attention as promising alternatives or adjuvants to antibiotics in the treatment of multidrug-resistant bacterial infections. Among phage-encoded enzymes, depolymerases are of particular interest due to their ability to degrade the bacterial polysaccharide capsule that prevents phage adsorption. Our results highlight the utility of combining recombinant depolymerases with phages to broaden their host range and enhance their activity against encapsulated, drug-resistant pathogens.