Isolation and characterization of bacteriophages with lytic activity against multidrug-resistant non-typhoidal Salmonella from Nairobi City county, Kenya.

Abstract

Background: Non-typhoidal Salmonella (NTS) typically cause self-limiting enterocolitis, but can lead to life-threatening invasive diseases, particularly in sub-Saharan Africa. In Kenya, multidrug-resistant (MDR) NTS strains with increasing non-susceptibility to third-generation cephalosporins pose a growing public health threat. As traditional antimicrobial treatments become less effective, bacteriophages are emerging as a potential alternative. This study aimed to isolate and characterize bacteriophages targeting MDR and extended spectrum-β-lactamase (ESBL)-producing non-typhoidal Salmonella (NTS).

Methods: Environmental samples were collected from seven sites in Nairobi City County, Kenya. Four NTS bacterial strains were used for phage enrichment, screening, and purification via spot tests and plaque assays. Phage efficacy was assessed in vitro by testing host range and efficiency of plating (EOP) against 12 Salmonella strains isolated in Kenya over different years. Ten selected broad-host-range phages were evaluated for thermal and pH stability and their ability to disrupt pre-formed NTS biofilms. Phage genomes were sequenced using the Illumina sequencing platform, and analyzed with bioinformatics tools to screen for antimicrobial resistance (AMR), lysogeny, virulence, and allergenic genes. The morphological characteristics of four representative phages were examined using Transmission Electron Microscopy.

Results: Thirty-one phages were isolated, with host ranges varying from lysing one strains to all 12 strains. Ten phages lysed more than 80% of the Salmonella strains and were selected for further characterization. Most phages exhibited high production EOP on at least one bacterial strain, except KE26 and KE28. All phages were stable from - 80 °C to 40 °C and pH 5 to 11, with noticeable but statistically insignificant biofilm disruption. Genome sizes ranged from 23,215 bp to 159,981 bp, and were free of known AMR, lysogeny, or virulence genes. Allergenicity screening identified no allergenic hits across most phages, with exception of KE23, which showed potential allergenic regions in its tail fiber and endolysin proteins. All phages belonged to class Caudoviricetes, with KE23, KE26, and KE28 exhibiting a myovirus-like morphotype, and KE15 displaying a siphovirus morphotype.

Conclusion: This study identified phages with desirable safety and stability profiles for potential usage against MDR and ESBL-producing NTS infections. Further in vivo studies are recommended to evaluate their therapeutic potential.

Importance: Non-typhoidal Salmonella (NTS) typically cause self-limiting enterocolitis but can lead to life-threatening invasive diseases. In Kenya, multidrug-resistant (MDR) NTS strains with increasing nonsusceptibility to third-generation cephalosporins have been reported, posing a significant public health concern that requires urgent attention. Bacteriophages are increasingly being considered as an alternative treatment for MDR bacterial infections because of the growing ineffectiveness of conventional antibiotics. Our study reports the isolation and characterization of lytic Salmonella phages devoid of detectable antimicrobial resistance (AMR) genes, lysogeny potential, allergens or virulence factors. These attributes position them as promising candidates for therapeutic interventions against MDR NTS infections. These findings highlight the potential of our study phages as a therapy for drug-resistant NTS and underscore the need for further investigation into their clinical application against MDR strains.