Isolation and characterization of a lytic bacteriophage against multidrug resistant Pseudomonas aeruginosa isolates and its effect on n-acetyl cysteine and biofilm

Objective: To isolate lytic bacteriophage effective against Pseudomonas aeruginosa isolates, the main source of infection in cystic fibrosis (CF) patients, and to evaluate the effect of this phage together with N-acetyl cysteine (NAC) on P. aeruginosa biofilm formation. Method: In this study, bacteriophages were isolated using traditional culture methods. Biological characterization included determination of lytic spectrum, pH stability, thermal stability, multiplicity of infection, one-step growth and burst size. Genomic characterization included phage DNA extraction, restriction fragment length polymorphism (RFLP) and phage genome analysis. In vitro biofilm formation of CF P. aeruginosa isolates was performed using microtiter plate assay with crystal violet staining. Results: MLPa10 phage was isolated from sewage water using P. aeruginosa isolated from CF patients, specifically P. aeruginosa 1Pa and P. aeruginosa 316Pa, as indicator hosts. The bacteriophage MLPa10 showed strong lytic activity against 94.11% of clinical P. aeruginosa isolates from CF patients. The latent and burst periods of the phage were 30 and 40 minutes, respectively, and the burst size was measured as 89 PFU/infected cell. MLPa10 phage was stable at pH 6 and pH 8 and at 40, 50, 60°C for 1 hour. Genome analysis of this phage revealed it to be taxonomically a lytic phage of the class Caudoviricetes. No virulence factors or antimicrobial resistance genes were detected in the phage genome. The study showed that MLPa10 phage effectively reduced mature biofilms in 76.9% of CF P. aeruginosa isolates and also inhibited biofilm formation in 65.38% of cases. In addition, NAC alone also showed efficacy against biofilms. However, the MLPa10 phage showed lower efficacy when combined with NAC. Conclusion: The extensive characterization of MLPa10 phage underlines its potential as a therapeutic agent against CF P. aeruginosa infections.