Bacteriophage Sf14 survival under simulated gastrointestinal conditions

Abstract

Shigella flexneri is an enteric pathogen responsible for causing shigellosis, a type of bacillary dysentery of significant global health concern. The increasing prevalence of antibiotic resistance has driven the search for alternative treatments, including bacteriophage (phage) therapy. However, the effectiveness of orally administered phages for enteric pathogens depends on their ability to survive passage through the gastrointestinal tract. In this study, the ability of bacteriophage Sf14 to maintain infectivity was evaluated under simulated gastrointestinal conditions using a modified INFOGEST 2.0 in vitro digestion model. This model includes salivary, gastric, and intestinal fluids with physiologically relevant compositions of electrolytes, bile salts, and enzymes. Phages were incubated in each simulated phase at standard physiological temperature, then tested for infectivity using quantitative plaque assays. Results show that Sf14 remained stable during the short duration of the oral phase and for the duration of the intestinal phase. By contrast, survival in simulated gastric fluid required a pH of 2.75 or higher, with rapid and substantial loss of infectivity at pH 2.5 and below. These findings demonstrate that while Sf14 is stable in oral and intestinal environments, the gastric phase poses a critical barrier to phage survival. With in vivo fasting gastric pH ranging from 1.5 to 3, encapsulation would enable the most flexible oral phage therapy against enteric pathogens, but ingestion after eating or alongside antacids may be sufficient. Importance The bacterium Shigella flexneri causes shigellosis, a highly transmissible and potentially life-threatening intestinal infection in humans. It is becoming increasingly difficult to treat as antibiotic resistance rises. Lytic bacteriophages, viruses that specifically infect and destroy bacterial cells, offer a potential alternative to antibiotics. However, developing effective phage therapy requires not only identifying a phage that targets S. flexneri, but ensuring that sufficient viable phage particles survive the changing conditions of the gastrointestinal tract and reach the site of infection. This study identifies the acidic stomach environment as a major barrier to the oral delivery of bacteriophage Sf14 and demonstrates that small changes in gastric acidity can substantially alter its survival. Moreover, these findings emphasize the importance of considering physiological barriers during the preliminary stages of phage therapy development and may inform future dosing, formulation, and administration strategies for oral phage therapies for shigellosis.