Characterization of Salmonella Enteritidis Bacteriophages and Evaluation of Phage Delivery Systems to Increase phage Survival in Simulated Digestive System and Chickens

Abstract

Salmonella enterica serovar enteritidis, a multidrug resistant strain, is one of the leading causes of foodborne illness in the globe. Humans mostly contract this non-typhoidal Salmonella serovar by eating tainted poultry flesh and other poultry items. For lowering the prevalence of multi-drug resistant non-typhoidal Salmonella in chicken farms, bacteriophages are an alternative to antibiotics. Phages with a stronger prophylactic or therapeutic potential may be able to survive the harsh conditions of the gastrointestinal tract, which have a low pH, high temperatures, and several digestive enzymes. Using various pH-adjusted medium, incubation temperatures, and simulated gastric and intestinal fluids, this study examined the host range, identification, and stability of 10 distinct Salmonella enteritidis phages isolated from Kenyan chicken farms. Additionally, their capacity to survive in Kenya's water sources—including rivers, boreholes, rainwater, and tap water—was evaluated. Additionally tested was the capacity of silica vesicles to adsorb/encapsulate, release, and safeguard phages in artificial stomach juice. Finally, 3-day old broiler chicks were used to assess their capacity for survival in vivo (24). On seven different strains of Salmonella enteritidis, all phages showed a wider host range and were relatively stable for 12 hours at pH values between 5 and 9 and temperatures between 25 °C and 42 °C. After 3 hours of incubation at pH 3, a viral titre decreases of up to 3 logs was seen. Phages remained stable in simulated stomach fluid for 20 minutes before losing their ability to infect. For up to two hours, phages remained largely stable in simulated intestinal fluid. Salmonella growth was significantly inhibited by phages in pH 2 and pH 3-adjusted media as well as in simulated gastric fluid at pH 2.5, but this effect was less pronounced in simulated intestinal fluid at pH 8. The other studied waters had just a minor impact on the phages, but river water had the greatest negative impact. The adsorption/encapsulation efficiencies of the three silica vesicles (SV 100, SV 140, and SV 140-C18) were 57.4%, 60%, and 90%, respectively. They were able to shield phages in stomach fluid for an hour and had modest, steady phage release rates up until day 4. SV 140-C18 had the lowest log reduction of 4 logs PFU/ml. Both SVs 100 and 140 lost six logs of PFU/ml decrease. Up until day 8 following inoculation, silica vesicle-encapsulated phages in the chickens displayed larger phage titres than non-encapsulated phages; however, until day 28 there was no discernible difference. On day 28, SV-encapsulated phages K¬28 and K11 had the highest titres. These findings imply that some of these phages may have a chance of surviving in living organisms and may be given orally by drinking water and survive the digestive system to avoid salmonellosis. The 10 Salmonella Enteritidis phages can be investigated for phage release and protection in people and other hosts, including chickens, where non-typhoidal Salmonella can be decolonized in vivo. SV 140-C18 should also be evaluated for phage release and protection in humans.