Characterization of 29 newly isolated bacteriophages as a potential therapeutic agent against IMP-6-producing Klebsiella pneumoniae from clinical specimens

Abstract

Carbapenemase-producing Enterobacteriaceae (CPE) are one of the most detrimental species of antibiotic-resistant bacteria globally. Phage therapy has emerged as an effective strategy for the treatment of CPE infections. In western Japan, the rise of Klebsiella pneumoniae strains harboring the pKPI-6 plasmid encoding bla IMP-6 is of increasing concern. To address this challenge, we isolated 29 phages from Japanese sewage, specifically targeting 31 K . pneumoniae strains and one Escherichia coli strain harboring the pKPI-6 plasmid. Electron microscopy analysis revealed that among the 29 isolated phages, 21 (72.4%), 5 (17.2%), and 3 (10.3%) phages belonged to myovirus, siphovirus, and podovirus morphotypes, respectively. Host range analysis showed that 18 Slopekvirus strains within the isolated phages infected 25–26 K . pneumoniae strains, indicating that most of the isolated phages have a broad host range. Notably, K. pneumoniae strain Kp21 was exclusively susceptible to phage øKp_21, whereas Kp22 exhibited susceptibility to over 20 phages. Upon administering a phage cocktail composed of 10 phages, we observed delayed emergence of phage-resistant bacteria in Kp21 but not in Kp22. Intriguingly, phage-resistant Kp21 exhibited heightened sensitivity to other bacteriophages, indicating a “trade-off” for resistance to phage øKp_21. Our proposed phage set has an adequate number of phages to combat the K. pneumoniae strain prevalent in Japan, underscoring the potential of a well-designed phage cocktail in mitigating the occurrence of phage-resistant bacteria. IMPORTANCE The emergence of Klebsiella pneumoniae harboring the bla IMP-6 plasmid poses an escalating threat in Japan. In this study, we found 29 newly isolated bacteriophages that infect K. pneumoniae strains carrying the pKPI-6 plasmid from clinical settings in western Japan. Our phages exhibited a broad host range. We applied a phage cocktail treatment composed of 10 phages against two host strains, Kp21 and Kp22, which displayed varying phage susceptibility patterns. Although the phage cocktail delayed the emergence of phage-resistant Kp21, it was unable to hinder the emergence of phage-resistant Kp22. Moreover, the phage-resistant Kp21 became sensitive to other phages that were originally non-infective to the wild-type Kp21 strains. Our study highlights the potential of a well-tailored phage cocktail in reducing the occurrence of phage-resistant bacteria.

Potential of an Isolated Bacteriophage to Inactivate Klebsiella pneumoniae: Preliminary Studies to Control Urinary Tract Infections

“…In another study, phage HS106 presented higher burst size (183 PFU/host cell) and low latent period (10 min) [43]. However, the phage's burst size is also affected by the host, as observed by Kondo (2023) [49]. Although phage KP1LMA presents a small burst size (7.9 ± 0.3 PFU/host cell) and a long latency period (100 min) (Figure 4), phages replicate efficiently in the host, causing a high reduction in K. pneumoniae growth, suggesting that other factors regulate the phage-bacteria interaction.…”

Characterizing the gut phageome and phage-borne antimicrobial resistance genes in pigs

“…The PhaBOX strategy comprises four tools, including PhaMer for phage identification [ 34 ], PhaTYP for lifestyle prediction [ 35 ], PhaGCN for taxonomy classification [ 36 ], and CHERRY for host prediction [ 37 ]. These tools combine the strength of alignment-based strategies and deep learning models to learn different sequence-based features [ 27 ] and have been used in recent studies [ 38 – 44 ]. In our study, a total of 174,897 non-redundant gut phage genomes were assembled and a total of 33,487 gut phage genomes were classified from 112 pig gut metagenomes using the PhaBOX strategy.…”

Analysis of a new phage, KZag1, infecting biofilm of Klebsiella pneumoniae: genome sequence and characterization

“…Bacteriophages, or phages, represent a promising alternative to conventional antibiotics due to their specific targeting of bacterial pathogens [ 1 – 9 ]. These viruses have gained attention for their ability to specifically target and kill bacterial pathogens, while leaving beneficial bacteria and human cells unharmed [ 2 , 7 – 10 ]. Phages are naturally occurring entities that infect and replicate within bacteria.…”

“…Phages are naturally occurring entities that infect and replicate within bacteria. They possess a high degree of specificity, with different phages targeting specific strains or species of bacteria [ 3 – 10 ]. This specificity is attributed to the recognition and binding of phage tail structures to specific receptors on the surface of bacterial cells.One of the key advantages of phages over traditional antibiotics is their ability to evolve alongside bacteria.…”

“…They have been extensively studied and used in certain regions for decades, particularly in Eastern Europe, as a therapeutic option for bacterial infections. Phages are generally well-tolerated by the human body and have minimal impact on the normal microbiota [ 10 – 12 ]. Phage therapy, the use of phages to treat bacterial infections, has shown promising results in both in vitro and in vivo studies.…”