Harnessing lytic phages for biofilm control in carbapenem-resistant <i>Klebsiella pneumoniae</i> causing urinary tract infection

ABSTRACT Klebsiella pneumoniae is an opportunistic pathogen with rising multidrug resistance and biofilm-related infections. Molecular and phage characterization is crucial to understand resistance mechanisms and explore alternative therapies, such as phage therapy. Whole-genome sequencing and antibiotic susceptibility testing were performed on hospital-isolated K. pneumoniae (KP6697), followed by multilocus sequence typing (MLST), plasmid replicon analysis, and antimicrobial resistance gene (AMR) profiling using bioinformatics tools. Phages were isolated and characterized by electron microscopy, with assessments of anti-biofilm activity, lytic efficacy, stability, and host range. Phage genomes were sequenced to identify functional genes. The host K. pneumoniae (KP6697) was multidrug-resistant, exhibiting resistance to 18 of 22 tested antibiotics. Genome analysis identified it as sequence type 16 (ST16) with 8 plasmid replicons and 23 AMR genes, including bla CTX-M-15 , bla NDM-5 , and bla OXA-181 . Functional annotations revealed extensive metabolic versatility and a rich repertoire of genes for biofilm formation, quorum sensing, secretion systems, and stress response. A lytic phage, Phage_KP6697_Omshanti, was isolated and classified as a Caudoviricetes member with a 45.3-kb genome encoding lysis, replication, and structural genes. The phage demonstrated short latency, high burst size, acceptable thermal and pH stability, and moderate host range against multiple CRKP and other bacterial isolates. Importantly, microscopy confirmed its ability to inhibit and degrade biofilms at multiple stages, highlighting its strong therapeutic potential. Lytic Phage_KP6697_Omshanti, with depolymerase and endolysin activity, isolated from carbapenem-resistant KP6697, showed high burst size, biofilm disruption, and had essential genomic traits suggesting its potential use as an anti-CRKP agent. IMPORTANCE Klebsiella pneumoniae is increasing multidrug resistance and robust biofilm formation pose severe clinical challenges, limiting treatment options. Understanding the molecular basis of its resistance and exploiting bacteriophages with strong biofilm-disrupting properties provide promising alternative therapeutic strategies. This study highlights the isolation and genomic characterization of a lytic phage with potent anti-biofilm activity against carbapenem-resistant K. pneumoniae , underscoring its potential in combating resistant infections.