Impact of phage therapy in post-weaning piglets challenged with ETEC strain in a controlled minitrial

Enterotoxigenic Escherichia coli (ETEC) is a pathogen responsible for post-weaning diarrhea (PWD) in piglets, which results in economic losses in pig production. The rise of antibiotic-resistant ETEC strains together with restrictions on addition of zinc oxide in pig feed require alternative management approaches. Our research examines bacteriophage therapy as a solution to control ETEC infections in newly weaned piglets. A cocktail of phages targeting a strain of ETEC F4LT1ST2 was identified and subsequently multiplicated in laboratory. We conducted a trial including nine piglets divided into three groups. The negative control group was exposed to the phage cocktail by administration with the bedding material (saw dust) on the floor of their pen. The treatment group was exposed to the ETEC strain and to the phage cocktail, and the positive control group was exposed to the ETEC strain only. Shotgun metagenomic sequencing was performed on fecal samples to characterize bacterial and phage dynamics. Throughout a 10-day period we monitored daily the rectal temperature and the diarrheal score of piglets. Subsequently we evaluated phage and bacterial counts in fecal samples to determine phage therapy effect on gut microbiota dynamics and piglet health. The PHAGE+ETEC group showed 19.2% lower cumulative diarrhea burden (p = 0.044) and 61.9% higher average daily gain (p = 0.065). Rectal temperature correlated significantly with diarrhea severity (per-piglet Spearman's ρ = 0.727, p = 0.027). Alpha diversity did not differ between treatment groups across timepoints, suggesting that phage administration did not cause major shifts in microbial diversity. Metagenomic analyses showed significant reduction of E. coli abundance in PHAGE+ETEC group compared to PHAGE groups (p = 0.009). Consistent with these observations, plaque assay results confirmed active phage-bacteria interactions: no plaque formation was detected in the feces of the ETEC-only group, whereas the PHAGE+ETEC group showed phage replication, reaching 10⁶ PFU/ml. This pilot study highlights the potential of phage therapy as an alternative to antibiotics for ETEC infections in piglets. Additional research with larger pig population and longer duration is required to confirm these findings and develop optimal phage application methods for swine production.