# Preparation of sodium alginate-coated microencapsulated phage vB_SalP_SE29 and evaluation of its therapeutic effect on Salmonella enteritidis infection

## TL;DR

This study shows that microencapsulating phage vB_SalP_SE29 improves its stability and effectiveness in treating Salmonella infections in rats.

## Contribution

The study introduces a novel microencapsulation method using gelatin, sodium alginate, and calcium chloride to enhance phage stability and therapeutic efficacy.

## Key findings

- Microencapsulated phages showed 86.52% encapsulation efficiency and 1 × 10⁷ PFU/mL titer.
- The microencapsulated phages improved rat survival to 70% and reduced inflammation and organ lesions.
- Storage at 4°C for 6 weeks caused less titer loss in microencapsulated phages compared to free phages.

## Abstract

Phages have demonstrated significant therapeutic efficacy in treating infections caused by antibiotic-resistant bacteria, yet their poor stability in the gastrointestinal tract limits oral application. This study aimed to address this barrier by preparing microencapsulated phage vB_SalP_SE29 (a lytic phage against Salmonella) using the droplet method, with gelatin, sodium alginate, and calcium chloride as wall materials. We optimized the microencapsulation process via single-factor experiments and orthogonal design, then evaluated the microspheres’ biological properties and in vivo therapeutic efficacy in a rat model of Salmonella enteritis. The optimal formulation (2% sodium alginate, 8% gelatin, and 1.65% calcium chloride) achieved an encapsulation efficiency of 86.52% and a titer of 1 × 107 PFU/mL. The potency remained stable in simulated gastric fluid at pH 2, was stable in 1% and 2% bile hydrochloric acid, and displayed improved release in simulated intestinal fluid. Storage at 4°C for 6 weeks reduced microencapsulated phage titer by 0.146 lg PFU/g (vs. 0.591 lg PFU/mL for free phages). Furthermore, in vivo, microencapsulated phages increased the survival rate of infected rats to 70% (vs. 40% for free phages). Microencapsulated phage also significantly reduced inflammatory factors compared to free phage and notably improved organ lesions in infected rats. These findings demonstrated that gelatin-sodium alginate-calcium chloride microencapsulation enhanced phage stability and therapeutic efficacy, supporting the potential of microencapsulated vB_SalP_SE29 as a safe bio-antimicrobial for combating Salmonella infections.

Salmonella is a common gram-negative bacterium widely distributed in nature, as well as in the intestines of humans and animals, and serves as a typical representative of foodborne pathogens. Due to the increasingly prominent issue of its multidrug resistance, phage therapy has garnered extensive attention as a potential alternative. However, when administered orally, phages are readily inactivated by gastric acid, leading to a significant reduction in phage titer. As a biopolymer with excellent biocompatibility, low cost, and low toxicity, sodium alginate can form a gel through cross-linking with Ca²⁺ ions at room temperature. The preparation of microencapsulated phages effectively protects phages from gastric acid damage, thereby enhancing their antibacterial efficacy. Evaluation of relevant biological characteristics has demonstrated that microencapsulated phages significantly improve their survival ability in the gastrointestinal environment. In vivo studies have further confirmed the good efficacy and safety of these microencapsulated phages in the treatment of rat enteritis.