Recombinant Limosilactobacillus reuteri strains expressing nanobodies against Clostridium perfringens toxins improve broiler performance during subclinical necrotic enteritis challenge

Subclinical Necrotic Enteritis (SNE) caused by Clostridium perfringens (Cp) toxins is estimated to affect up to 40% of global commercial broiler flocks, resulting in substantial economic losses due to poor production performance. Engineered gut-derived probiotic bacteria, such as lactobacilli expressing nanobodies (Nbs) against pathogens or pathogenic toxins, are emerging as promising alternatives for the prevention and treatment of human and animal diseases. This study evaluated the effects of the investigational veterinary product BE-101, which combines two recombinant Limosilactobacillus reuteri (LR) vectors individually expressing nanobodies against the Cp toxins alpha-toxin and NetB, on body weight gain (BWG), feed intake (FI), feed conversion ratio (FCR), and mortality across feeding phases in an SNE challenge model conducted with 2,600 male Ross 708 broilers under commercial-like management conditions. A 2-dose regimen of BE-101, consisting of a day-of-hatch (DOH) coarse spray and day 14 application through drinking water, significantly reduced FCR by 4 points, numerically increased BWG by 2.4%, and numerically reduced total mortality by 25% compared with challenge control birds over the 43-day study. The 1-dose regimen, given only at day-of-hatch, provided partial benefit to BWG (+1%) and FCR (-1 point) and numerically reduced mortality by 33%. The backbone vector strains not expressing nanobodies provided only limited benefit. Overall, these results indicate that engineering probiotic lactobacilli to express toxin-neutralizing nanobodies can enhance the modest benefits of probiotics alone and may provide a practical approach for reducing SNE-associated production losses in commercial broiler systems.