PlyNJ3: a three-domain endolysin with broad-spectrum anti-streptococcal activity and synergistic efficacy against multidrug-resistant Streptococcus suis.

Background: The global spread of multidrug-resistant (MDR) bacteria prompts the exploration of innovative antimicrobial strategies. Phage lysins-peptidoglycan hydrolases known for species-specific activity and low resistance potential-offer promising alternatives to conventional antibiotics. However, their typically narrow spectrum limits broad therapeutic application. To overcome this limitation, we leveraged the wide distribution of streptococcal mobilizable prophage (SMphage) families across Streptococcus species as a strategy to discover lysins with inherent broad-spectrum potential. Within these conserved prophages, we expressed PlyNJ3, an SMphage-derived endolysin homolog identified in S. suis, and evaluated its therapeutic efficacy in infection models.

Results: PlyNJ3 exhibited potent lytic activity against a broad panel of streptococci, including diverse S. suis clinical isolates (15 serotypes), 83.3% of S. uberis, and key strains of S. agalactiae, S. dysgalactiae, and S. pyogenes. Optimal bacteriolytic activity occurred at physiological temperature (37 °C) with dose-dependent efficacy. Electron microscopy revealed PlyNJ3-mediated cell wall perforation, cytoplasmic content release, and structural disintegration. In a murine invasive S. suis infection model, high-dose intraperitoneal PlyNJ3 (0.9 mg/head) conferred 87.5% survival, while synergism with ampicillin prolonged survival. Intramuscular administration in a murine thigh infection model enhanced bacterial clearance compared to intraperitoneal delivery. Phylogenetic and structural analysis confirmed conservation across SMphage-derived lysins, underpinning their broad streptococcal target potential.

Conclusions: Our findings establish SMphage-derived lysins as a conserved, therapeutically valuable class of antimicrobials with inherent broad-spectrum potential across diverse streptococcal pathogens. Building on the previously demonstrated in vivo efficacy of PlySK1249 against S. dysgalactiae and S. agalactiae, we show that its structural homolog, PlyNJ3, exhibits robust therapeutic activity against S. suis infections. Together, these results validate the SMphage lysin platform for generating precision-targeted agents against specific streptococcal infections.