Isolation, characterization, and alginate hydrogel delivery of a Klebsiella aerogenes bacteriophage and its impact on biofilm degradation.

Multidrug-resistant Klebsiella aerogenes poses a significant clinical challenge due to its antimicrobial resistance and biofilm-forming capacity in chronic wounds and device-associated infections. Bacteriophage therapy offers a promising alternative to conventional antibiotics, though effective delivery systems remain critical for clinical translation. This study reports the isolation, genomic characterization, and alginate hydrogel-based formulation of a lytic bacteriophage, KA_SGEB_01, targeting multidrug-resistant K. aerogenes. The phage was isolated from hospital sewage using multidrug-resistant K. aerogenes as a host. Comprehensive characterizations included plaque morphology and host range determination, multiplicity of infection (MOI) optimization, one-step growth kinetics, antibiofilm activity assays, and environmental stability testing. Complete genome sequencing and annotation were performed. Phage-loaded alginate hydrogels were developed and evaluated for release kinetics, long-term viability, and antibiofilm efficacy. KA_SGEB_01 produced clear plaques (1 ± 0.1 mm) with halos indicative of depolymerase activity and exhibited strict host specificity. The optimal MOI was 0.01 with a 15-minute latent period and the phage remained stable between -20 °C and 50 °C; and across a pH range 6-9. Biofilm assays demonstrated around 60% Crystal-Violet-stained biomass reduction as well as significant biofilm disruption visualized by Scanning Electron Microscopy. Whole-genome sequencing revealed a 175,095 bp double-stranded DNA genome encoding 289 predicted coding sequences, classified within the family Straboviridae, genus Slopekvirus, and lacking any virulence or AMR genes. Encapsulation in alginate hydrogels preserved viability (>10⁹ PFU/mL) for 14 days and enabled sustained release (∼ 67% at 72 h), resulting in significant biofilm biomass inhibition. Collectively, KA_SGEB_01-loaded alginate hydrogels represent a promising platform for treating multidrug-resistant K. aerogenes in wound and device-associated infections caused by the bacterium.