Evaluation of antibacterial properties of Lactobacillus acidophilus and Lactobacillus gasseri on enterotoxigenic Escherichia coli associated with porcine post-weaning diarrhea (PWD) using Galleria mellonella model

Post-weaning diarrhea (PWD) in piglets, caused by enterotoxigenic Escherichia coli (ETEC), has been managed using antibiotics and zinc oxide. However, concerns regarding antibiotic resistance as well as the 2022 EU ban on zinc oxide, have prompted the development of sustainable alternatives. Growing evidence suggests that probiotics may play a promising role in controlling ETEC infections. In this study, we first evaluated the ability of three potential probiotics ( Lactobacillus gasseri, Lactobacillus acidophilus, and Limosilactobacillus reuteri ) to reduce ETEC infection in vitro . Subsequently, we used the Galleria mellonella model to further evaluate their impact in vivo , by testing two different approaches: co-infection (i.e. probiotic + ETEC) and prophylactic strategy (i.e. prior infection with probiotic for 4 h followed by ETEC infection). The survival rate and health index scores were assessed after 24, 48, and 72 h. In addition, quantitative reverse transcription polymerase chain reaction (RT-qPCR) was also performed to determine the transcript levels of genes encoding the G. mellonella antimicrobial peptides (AMPs), namely, lysozyme, inducible metalloproteinase inhibitor (IMPI), gallerimycin, and galliomycin, to infer the immune response to ETEC infection. Our results suggest that a co-infection strategy was ineffective in controlling ETEC infection under our experimental conditions. On the other hand, when using a prophylactic strategy, we observed significant differences between the treated larvae and the control. Overall, we observed that L. acidophilus significantly reduced infection by the ETEC strain SP11 ( p <0.05), while L. gasseri demonstrated anti-ETEC potential, specifically against the SP31 strain ( p <0.05). Differences in IMPI and galliomycin AMPs expression were also observed between treated and control conditions. However, these variations depended on the specific strains of both Lactobacillus and ETEC involved. In conclusion, selected Lactobacillus species showed promising potential as prophylactic alternatives for controlling ETEC infection, although efficacy was strain-dependent. • PWD in piglets is primarily caused by enterotoxigenic E. coli (ETEC). • Restrictions on zinc oxide and antibiotics raise the need for alternatives. • L. acidophilus and L. gasseri show strain-specific anti-ETEC effects. • Certain Lactobacillus strains improve survival and health in G. mellonella model. • Strain-specific probiotics may modulate immunity against ETEC infections.