Isolation, Characterization, and Host-Range Evaluation of Bacteriophages Against Multidrug-Resistant Escherichia coli Clinical Isolates

BackgroundThe increasing prevalence of multidrug-resistant (MDR) Escherichia coli has created a need for alternative antibacterial approaches.Bacteriophages are being investigated because of their ability to specifically infect and lyse bacterial hosts.However, the host range of individual phages may be limited.The present study evaluated bacteriophages isolated from environmental water sources against clinical MDR E. coli isolates and assessed the observed lytic coverage of a three-phage cocktail. MethodsAn in vitro experimental study was conducted using 375 MDR E. coli clinical isolates recovered from clinical specimens.Pond water, sewage water, and Ganga water were screened for bacteriophages using a spot assay, followed by plaque assay and plaque purification.Purified phages were propagated, and their titers were determined by plaque assay.The host-range activity of three purified phages was evaluated against all 375 MDR E. coli isolates.The cumulative coverage of a three-phage combination was then derived from the individual host-range results.Lytic activity among the three individual phages was compared using Cochran's Q test, followed by exact McNemar tests with Bonferroni correction. ResultsAmong the 375 MDR E. coli isolates, the highest resistance was observed against ciprofloxacin (95.2%), while the highest susceptibility was observed to amikacin (72.3%).Preliminary spot screening identified phageassociated lysis in 72 (19.2%) isolates, while plaque formation was observed in 36 (9.6%) isolates.Three purified bacteriophages were recovered from pond water, sewage water, and Ganga water and designated Phage-1, Phage-2, and Phage-3, respectively.Their estimated titers were 1.6 × 10¹², 1.7 × 10¹⁶, and 1.9 × 10¹⁵ PFU/mL, respectively.Phage-1 produced detectable lysis in 146 (38.9%) isolates, Phage-2 in 90 (24.0%), and Phage-3 in 169 (45.1%).Cochran's Q test demonstrated a significant overall difference in lytic activity among the three phages (p < 0.001).The difference between Phage-1 and Phage-3 was not statistically significant (p = 0.259).The cumulative coverage of the three phages combined, derived from the individual host-range results, was 260 (69.3%) isolates, while 115 (30.7%) showed no detectable lysis with any of the three phages. ConclusionThe study demonstrated that environmental water sources yielded bacteriophages with detectable lytic activity against clinical MDR E. coli isolates.The individual phages showed variable host-range activity, and their combined (cumulative) coverage, derived from the individual host-range results, was broader than that of any individual phage.This represents cumulative phenotypic coverage and should not be interpreted as the result of a separately tested cocktail or as evidence of phage synergy.Further biological, genomic, safety, and efficacy studies are required to evaluate the potential of these bacteriophages for subsequent phagedevelopment research.