Isolation of lytic bacteriophages against multidrug Resistant Escherichia coli using the double agar layer technique

Abstract

The rapid emergence of multidrug resistant (MDR) Escherichia coli necessitates reliable methods to recover lytic bacteriophages from environmental reservoirs for potential therapeutic and biocontrol applications. This article emphasis on Double Agar Layer (DAL) protocol used to isolate and purify lytic phages from animal waste disposal systems. Samples were collected from multiple farm waste strata clarified by differential centrifugation, filtered and enriched using log-phase MDR E. coli hosts. Phages were detected and enumerated by DAL plaque assays purified through successive single-plaque isolation cycles and stock lysates prepared by confluent lysis. The DAL method demonstrated high sensitivity and reproducibility for recovering lytic phages from complex matrices.