Encapsulation of Bacteriophages in Alginate Beads: Improved Viability Under Harsh Simulated Gastric and Intestinal Conditions for Phage Therapy Applications

<b>Background/Objectives</b>: Bacteriophages offer a promising alternative to conventional antibiotics. However, their therapeutic efficacy is often limited by instability in harsh environmental conditions, particularly within the gastrointestinal tract. This study aimed to isolate lytic bacteriophages from wastewater and evaluate the protective capacity of sodium alginate encapsulation against various stressors to enable effective oral delivery. <b>Methods</b>: Four distinct lytic phages (As, Ec, Pa, Gc) were isolated from wastewater and characterized by Transmission Electron Microscopy (TEM) and PCR, confirming their families (Siphoviridae, Podoviridae, Myoviridae). These phages demonstrated potent lytic activity against diverse bacterial pathogens, including <i>Aeromonas hydrophila</i>, <i>Escherichia coli</i>, <i>Pseudomonas aeruginosa</i>, and <i>Glutamicbacter creatinolyticus</i>. The phages were encapsulated in 5% sodium alginate via an extrusion method. Stability was assessed under extreme pH (2.0 and 13), at elevated temperature (up to 80 °C), and in simulated gastrointestinal transit. <b>Results</b>: Encapsulation efficiency exceeded 95%. Unencapsulated phages were completely inactivated at pH 2.0 within 10 min, whereas encapsulated phages maintained significant viability (3.06-3.43 log PFU/mL). Encapsulation also significantly enhanced phage survival under extreme alkaline conditions and elevated temperatures. In simulated gastrointestinal transit, encapsulated phages exhibited superior recovery (2.50 log PFU/mL) compared to their free counterparts (≤1 log PFU/mL). Long-term storage evaluations over three months further confirmed the robust stability of the encapsulated formulations at both 4 °C and 21 °C. <b>Conclusions</b>: Sodium alginate encapsulation effectively shields bacteriophages from severe environmental degradation, particularly acidic gastric stress, enhancing their potential for oral delivery. These findings support the development of stable, formulated phage products for diverse practical applications in phage therapy to combat antimicrobial resistance.