Abstract:
Background: The alarming rise of multidrug-resistant (MDR) Klebsiella pneumoniae poses a significant
threat to global public health and has a considerable economic impact, necessitating the development of
alternative therapeutic strategies. Bacteriophages are alternative therapeutic agents in the fight against such
bacteria. Phages are host-specific and can target specific bacteria precisely; thus, a detailed understanding
of phage-host interactions in the presence of antimicrobial drug is required for effective usage. This study
contributes to the growing body of research supporting phage therapy as an effective solution in the face of
rising antibiotic resistance.
Objective: This study was conducted to isolate and characterize virulent bacteriophages against MDR K.
pneumoniae clinical isolates and to assess the phages interaction with antibiotics and their effects on the
bacterial biofilm eradication.
Method: Laboratory based experimental study was used to conduct the study. Ten MDR K. pneumoniae
isolates were selected, and three of them were taken randomly for use in the isolation and enrichment
process of lytic phages. Four wastewater samples were collected at four different sites of Jimma medical
center compound through one time collection approach. Three lytic phages were isolated on spot assay and
further purified and quantified by plaque assay. The phages biological characteristics were studied using a
spot and plaque assay. Checkerboard assay was utilized for synergy and biofilm disruption testes.
Results: Three lytic phages; KpPhage57, KpPhage56, and KpPhage85 against MDR K. pneumoniae were
isolated and phenotypically characterized. The KpPhage57, KpPhage56, and KpPhage85 virus particles
yielded by the plaque assay were 1.80*109 PFU/ml, 1.60*108 PFU/ml, and 5.0*1010 PFU/ml, respectively.
These phages were stable between a temperature range of 4°C to 50°C and a pH value of 4 to 11, with
almost similar lytic activity and narrow host ranges, with KpPhage56 having a relatively broader host
range. KpPhage56 and KpPhage85 (at a titer of 106 PFU/ml) showed synergetic interaction with
Gentamicin (500µg/ml), having a fractional inhibitory concentration index (FICI) of 0.085 and 0.077,
respectively, indicating promising candidate for future phage therapy. All the three phages effectively
disrupted K. pneumoniae biofilm at multiplicity of infection (MOI) of 10.
Conclusion: The lytic and synergistic efficacy of the isolated phages, in conjunction with gentamicin,
demonstrates their significant potential as a compassionate treatment for patients afflicted with MDR K.
pneumoniae and biofilm-associated infections, particularly in wound and catheter-associated cases
especially in resource-limited settings like Ethiopia.
