Recombinant LAB vector-based multicomponent vaccine against Campylobacter jejuni potentially promoting a healthier microbial balance in the poultry gut

Background Diarrheal diseases remain the second leading cause of preventable death globally, particularly among children under the age of 5 in developing countries, accounting for an estimated 2–3 million deaths annually. Among bacterial pathogens causing diarrheal illness, Campylobacter jejuni ( C. jejuni ) remains a major contributor, particularly in low- and middle-income countries (LMICs). As a common gut pathogen, C. jejuni expresses several secretory or surface-expressed colonization proteins (SECPs), namely haemolysin co-regulated protein (Hcp), valine glycine repeats G (VgrG), Campylobacter adhesion to fibronectin (CadF), fibronectin-like protein A (FlpA), and jejuni lipoprotein A (JlpA). Most of these proteins play pivotal roles in bacterial self-survival, host-cell adhesion, and invasion of avian and non-avian hosts. To minimize C. jejuni adhesion and subsequent colonization in the avian gut , we explored the potential of a multicomponent mucosal vaccine composed of CadF, Hcp, and JlpA protein of C. jejuni. Results For this purpose, we bioengineered a food-grade Lactic Acid-producing Bacterium, Lactococcus lactis ( L . lactis ), to express three key immunogenic subunits of C. jejuni , CadF, Hcp, and JlpA. Utilizing this live vector-based multicomponent mucosal vaccine platform, we investigated the immunoprotective potential of these antigens in chickens. Since the particular strain of L. lactis is non-colonizing, we used chitosan, a natural mucoadhesive, biodegradable polymer, to microencapsulate the engineered bacteria and increase their gut retention time for optimal interaction with local immune cells . Our in vivo immunization study demonstrated that oral administration of this multicomponent vaccine formulation elicited a strong local antibody response (sIgA) ( p < 0.0001) and upregulated key pro-inflammatory cytokines, leading to robust mucosal immune protection (~ 1.54 log 10 reduction) against the cecal colonization of C. jejuni . Beyond targeting C. jejuni , we hypothesized that the vaccine may influence the overall gut microbiota, potentially promoting a healthier microbial balance in the poultry gut. To this end, gut metagenomic analysis of vaccinated birds revealed a marked reduction in the phylum Campylobacterota (~ 2-fold), accompanied by increased abundance of the phyla Bacteroidota, as part of a beneficial microbial community. Conclusions Together, this study underscores the potential of a live vector-based, multicomponent mucosal vaccine as a promising, cost-effective strategy to reduce the cecal load of C. jejuni , potentially limiting the risk of foodborne transmission in poultry production systems.