Lactobacillus-vectored nanobodies improve broiler productivity under sub-clinical necrotic enteritis with associated microbiome and transcriptome changes.

Sub-Clinical Necrotic Enteritis (SCNE), caused by toxin-producing Clostridium perfringens, is a major challenge in poultry production. SCNE has traditionally been managed with in-feed antibiotics; however, increasing concerns about the spread of antimicrobial resistance call for antibiotic-free strategies for its control. We recently described an NE control strategy leveraging Limosilactobacillus reuteri probiotic strains genetically engineered to deliver nanobodies against alpha toxin and NetB from C. perfringens in the poultry gut. Here, in a controlled study under SCNE conditions, we found that the engineered strains significantly improved feed conversion ratios and weight gain of broilers, outperforming treatment with either a prophylactic antibiotic or the wild-type probiotic strains. To investigate the systemic factors contributing to these performance differences, we analyzed histomorphometrics of the small intestine, microbial metatranscriptomics of jejunal contents, and gene expression from the jejunum and liver tissues. Our results confirmed the in situ expression of the nanobodies and provided evidence that nanobody delivery mitigates SCNE-associated inflammation in the jejunum and toxin-induced damage in the liver, leading to a more quiescent immune state, lower oxidative stress, and improved growth performance. Our findings demonstrate the potential of probiotic-vectored nanobody delivery as an effective strategy for targeting gut antigens across a range of diseases.