Therapeutic efficacy of phage vB_SalP_NW15 and cinnamaldehyde against drug-resistant Salmonella Enteritidis in a chicken infection model.

Abstract

Salmonella is a major Gram-negative pathogen causing infections in human and poultry. Currently, clinical treatment primarily relies on antibiotics, but their overuse has led to the widespread prevalence of multidrug-resistant (MDR) strains. The combination of phages and natural bioactive substances offers a novel approach to combat MDR Salmonella. In this study, a lytic Salmonella phage vB_SalP_NW15 was isolated from the wastewater of a livestock farm and identified as a member of the Siphoviridae family through transmission electron microscopy. The phage demonstrated a broad-spectrum, effectively lysing 90 % (18/20) of the tested Salmonella isolates. Phage vB_SalP_NW15 exhibited considerable stability across a wide pH range (4-13) and demonstrated thermal tolerance up to 60°C. The latent period of phage was 10 min, with a burst size of 96 PFU/cell. Genomic analysis confirmed the absence of genes associated with virulence, antibiotic resistance, or lysogeny, underscoring its safety profile for biocontrol applications. Crucially, a checkerboard assay demonstrated a potent synergistic effect between phage vB_SalP_NW15 and cinnamaldehyde (CA), with a Fractional Inhibitory Concentration Index (FICI) of 0.5. In vitro, the combination of the phage (10-10PFU/mL) and a sub-inhibitory concentration (0.3125 mg/mL) of CA resulted in significantly enhanced bacterial growth inhibition. The therapeutic efficacy of this combination was validated in vivo using a chicken infection model, where co-administration significantly improved survival rates, alleviated histopathological damage in the heart and liver, and reduced bacterial loads in tissues, with outcomes superior to either monotherapy. Collectively, these findings establish that the synergistic combination of phage vB_SalP_NW15 and CA represents a promising therapeutic strategy against MDR Salmonella infections in poultry.