Characterization of the bacteriophage vB_KpnP_Henu1_3 lytic for K1 <i>Klebsiella pneumoniae</i> and its therapeutic efficacy in <i>Galleria mellonella</i> larvae and mice

New anti-infective therapies are urgently needed for the treatment of drug-resistant bacterial infections in the context of the rapid spread of drug resistance. Phages, the natural enemies of bacteria, have irreplaceable advantages in the treatment of bacterial infections. Here, we report a novel phage, vB_KpnP_Henu1_3, that specifically lyses capsule-type K1 <i>Klebsiella pneumoniae</i>. The phage vB_KpnP_Henu1_3 shows relatively favorable thermal stability (4°C-55°C) and pH tolerance (pH = 4-11). In addition, the optimal multiplicity of infection is 0.01, with a burst size of approximately 253 ± 54 PFU/cell. Genomic analysis reveals that phage vB_KpnP_Henu1_3 contains double-stranded DNA (total length of 49,808 bp) with a G + C content of 50.76%, and the genome comprises 75 open reading frames with no virulence- or antibiotic resistance-related genes. Transmission electron microscopy observations revealed that phage vB_KpnP_Henu1_3 possessed an icosahedral head and siphovirus morphology. Based on the genome sequence, phage vB_KpnP_Henu1_3 could be assigned to a new species in the genus <i>Webervirus</i> of the family <i>Drexlerviridae</i>. Furthermore, phage vB_KpnP_Henu1_3 rapidly inhibits the growth of <i>K. pneumoniae</i> within 4 h, prevents biofilm formation, and disrupts mature biofilms <i>in vitro</i>. In infection animal models, phage vB_KpnP_Henu1_3 significantly increases the survival rate of <i>K. pneumoniae</i>-infected <i>Galleria mellonella</i> larvae and mice while reducing the bacterial loads. These findings demonstrate that phage vB_KpnP_Henu1_3 has promising potential as a safe alternative for controlling and treating multidrug-resistant K1-type <i>K. pneumoniae</i> infections.IMPORTANCEThe widespread use of antibiotics has led to increasing antibiotic resistance, which is a growing global health concern. Therefore, the development of novel antimicrobial therapy that can cure drug-resistant bacteria-induced infections is imperative. Phages are of increasing interest as natural enemies of bacteria with clear advantages in antibacterial.