Campylobacter is a leading cause of human gastroenteritis, with poultry serving as the primary reservoir host. Effective preharvest control strategies are crucial for preventing or reducing Campylobacter contamination on meat surfaces. As concerns grow regarding the use of antimicrobials in animal agriculture, the importance of non-antimicrobial preharvest strategies in poultry production has become increasingly significant. This comprehensive review focuses on the biology of Campylobacter, its impact on public health, and current and emerging preharvest strategies, with a special emphasis on vaccination. Preharvest strategies are broadly classified into biosecurity measures, gut microbiota modifications using prebiotics, probiotics, postbiotics, feed additives, and vaccination. While some vaccines have proven to be effective in research settings, no commercial vaccines are currently available. Because no single strategy can effectively combat Campylobacter, integrating multiple approaches, such as improved biosecurity measures, immunization, and dietary modifications, may provide a solution for reducing Campylobacter loads in poultry. Embracing a “One Health” approach, gaining a deeper understanding of Campylobacter pathophysiology, advancing vaccine technology, and implementing holistic farm management practices will be essential for the sustainable control of Campylobacter and for reducing the risk of human campylobacteriosis.

This systematic review aimed to (i) determine the risk of antimicrobial resistance (AMR) development associated with antimicrobial use (AMU) and other exposure factors in broilers, and (ii) identify best management practices to mitigate preharvest AMR development of enteric bacteria alongside broiler production. Study selection criteria comprised the population, exposure or intervention, comparator, and outcome framework and included broiler (population), AMU or other management practices (exposure or intervention), organic or antibiotic-free production (comparator), and the presence of AMR-enteric bacteria/genes (outcome). Peer-reviewed primary research studies were searched in PubMed on 19 December 2022, and AGRICOLA, Embase, Scopus, and Web of Science on 10 February 2023. The risk of bias in studies was assessed using the modified ROBIS-E risk of bias assessment tool. The results were synthesised and presented narratively according to PRISMA 2020 guidelines. In total, 205/2699 studies were subjected to full-text review, with 15 included in the final synthesis. Enteric bacteria Escherichia coli, Salmonella^, and Campylobacter exhibited AMR and multidrug resistance against several critically important antimicrobials (aminoglycoside, cephalosporin, chloramphenicol, macrolide, penicillin, quinolone, tetracycline, and sulfonamide) for human health. The risk of AMR development in bacteria was shown to be potentially higher with AMU in broiler production. Substandard farm management practices, poor biosecurity measures, and conventional production systems have also been associated with the dissemination of AMR in bacteria. Our findings indicate that AMU exposure is associated with considerably higher risk of AMR development in enteric bacteria. Antimicrobial stewardship, organic/antibiotic-free broiler production, good farm management practices, and high-level biosecurity measures are able to substantially mitigate preharvest AMR development in enteric bacteria. However, most of studies were cross-sectional, and therefore causal inference cannot be established.

Campylobacter is a leading cause of human gastroenteritis, with poultry serving as the primary reservoir host. Effective preharvest control strategies are crucial for preventing or reducing Campylobacter contamination on meat surfaces. As concerns grow regarding the use of antimicrobials in animal agriculture, the importance of non-antimicrobial preharvest strategies in poultry production has become increasingly significant. This comprehensive review focuses on the biology of Campylobacter, its impact on public health, and current and emerging preharvest strategies with a special emphasis on vaccination. Preharvest strategies are broadly classified into biosecurity measures, gut microbiota modifications using prebiotics, probiotics, postbiotics, feed additives, and vaccination. While many live, attenuated, and subunit vaccines have proven effective in research settings, there are currently no commercial vaccines available. Because no single strategy can effectively combat Campylobacter, integrating multiple approaches, such as improved biosecurity measures, immunization, and dietary modifications, may provide a solution for reducing Campylobacter loads in poultry. Embracing a “One Health” approach, gaining a deeper understanding of Campylobacter pathophysiology, making advancements in vaccine technology, and implementing holistic farm management practices will be essential for sustainable control of Campylobacter and for reducing the risk of human campylobacteriosis.

Background: Non-typhoidal Salmonella (NTS) is a major foodborne pathogen, with poultry products, especially eggs, being the primary source of human infections. Current serovar-specific poultry vaccines effectively reduce targeted Salmonella serovars but may inadvertently promote the emergence of untargeted serovars within poultry flocks. Therefore, novel vaccine candidates providing broad cross-serovar protection are needed to improve overall effectiveness of Salmonella control programs. Objectives: This study evaluated the immunogenicity of the novel subunit vaccine candidate InvG and assessed its ability to reduce Salmonella colonization in vaccinated laying hens and their progeny through maternally derived antibodies transferred via egg yolk. Methodology: Three experiments were performed. Experiment I evaluated the immunogenicity of purified recombinant InvG by (a) measuring anti-InvG antibodies using an enzyme-linked immunosorbent assay (ELISA) and (b) completing transcriptomic profiling of immune responses in vaccinated chickens. Vaccinated chickens were subsequently challenged with Salmonella Enteritidis to assess the efficacy of anti InvG antibodies in reducing intestinal colonization of Salmonella. Experiment II involved immunizing hens with InvG, to evaluate passive transfer of antibodies via egg yolk and the protective efficacy of maternally derived antibodies against Salmonella challenge. Passive transfer was assessed by measuring IgY antibodies in hen serum, egg yolk, and progeny serum, as well as secretory IgA (sIgA) antibodies in progeny intestinal washings using ELISA. Protective efficacy was evaluated by orally challenging one-day-old chicks with three different Salmonella serovars. Experiment III assessed the persistence of anti-InvG antibodies in the serum of vaccinated hens and their transfer into eggs following two doses of InvG. Results: InvG vaccination induced robust IgY antibody responses in hens, with efficient maternal antibody transfer to progeny via egg yolk. A statistically significant reduction in Salmonella colonization was observed in both vaccinated hens and their progeny. Conclusions: These findings demonstrate that InvG represents a promising subunit vaccine candidate for Salmonella control in poultry and warrants further investigation towards development as a broadly protective commercial poultry vaccine against Salmonella.

Campylobacter hepaticus is the etiological agent of Spotty Liver Disease (SLD), a newly emerging bacterial disease of laying hens resulting in significant mortality and production losses primarily in free-range systems. Although its economic impact continues to grow, the molecular basis of C. hepaticus pathogenesis remains poorly understood. In this study, we conducted transcriptomic profiling of C. hepaticus in three host-relevant conditions, exposure to chicken bile, infection of a chicken liver hepatocellular carcinoma (LMH) cell line, and isolation from liver lesions of naturally infected chickens. Through RNA-seq analysis, we found unique gene expression signatures in each environment. In the bile, C. hepaticus exhibited differential expression of 412 genes, with upregulation of genes related to motility, cell envelope remodeling, glycosylation, nitrate respiration, and multidrug efflux systems, indicating a stress-adaptive, metabolically active lifestyle. In LMH, on the other hand, 125 genes were differentially expressed, primarily reflecting downregulation of motility, oxidative stress response, chaperones, and core metabolic processes, suggesting that these cells adopt a less active, intracellular dormant lifestyle. Transcriptomic analysis of C. hepaticus isolated from the liver identified 26 differentially expressed genes, featuring selective upregulation of genes associated with nitrate respiration, sulfur metabolism, and pyridoxal 5’ phosphate homeostasis, alongside downregulation of the major outer membrane porin (momp), stress response chaperones (dnaK, groL), and genes involved in oxidative stress defense and energy production. Furthermore, the immune evasion-related gene cmeA and a glycosyltransferase gene were found to be highly upregulated. This study presents the first in-depth transcriptomic exploration of C. hepaticus in multiple host relevant niches. Our findings reveal niche-specific gene expression profiles and highlight metabolic and structural adaptations that enable C. hepaticus to survive during bile exposure, persist within host cells, and contribute to liver pathology. These insights provide a basis for identifying novel virulence determinants and may inform the development of targeted interventions, including vaccines or antimicrobial therapy, to control SLD in commercial poultry operations.

Background/Objectives: Spotty liver disease (SLD), caused by Campylobacter hepaticus, is an emerging disease that leads to substantial production losses in the egg industry. The shift toward antibiotic-free and cage-free production systems has further intensified the impact of SLD. The current control measures largely rely on autogenous killed vaccines; however, their use is constrained by the slow and fastidious growth of C. hepaticus and inconsistent efficacy. To overcome these limitations, this study aimed to identify immunogenic mimotopes as vaccine candidates and express them on the surface of an avian pathogenic Escherichia coli (APEC) vector. Methods: To identify immunogenic mimotopes, Ph.D.-12 phage display peptide library was screened using the hyperimmune serum raised against killed whole-cell C. hepaticus in specific pathogen-free chickens. Subsequently, the outer membrane protein C (OmpC) of E. coli was used as a scaffold for constructing a surface display library. A single restriction site, PstI, located in the seventh external loop of OmpC, was strategically utilized to insert each 12-amino-acid mimotope with a six-histidine (6xHis) tag sequence at its N-terminus, generating ompC + mimotope fusion constructs. These constructs were cloned into the inducible expression vector pTrc and electroporated into an E. coli DH5α ∆ompC strain, which lacked ompC. The surface expression of the mimotopes was confirmed in vitro. The verified ompC + mimotope constructs were subsequently subcloned into the pYA3422 constitutive expression vector and electroporated into the APEC PSUO78 ∆aroA ∆asd vaccine vector strain. A chicken vaccination–challenge trial was conducted using nine groups of chickens, including an unvaccinated challenged control and an unvaccinated–unchallenged negative control. Each experimental group received a mixture of two recombinant E. coli strains carrying different mimotopes at a dose of 1 × 10⁹ CFU, which were administered orally twice at 16 and 18 weeks of age. Results: Fourteen immunogenic mimotopes corresponding to 13 different C. hepaticus proteins were identified as potential vaccine candidates. The expression of these mimotopes on the surface of the E. coli was successfully demonstrated using the OmpC-mediated surface display system. Of the 14 mimotopes tested, two flagellar-related peptides and one major outer membrane protein (MOMP)-derived peptide elicited significant immune responses and conferred protection against the C. hepaticus challenge. Conclusions: We successfully developed a functional E. coli surface display system that was capable of expressing 12-amino-acid mimotopes of C. hepaticus, providing a robust platform for evaluating vaccine candidates against SLD. Immunogenicity and efficacy studies in chickens demonstrated that three identified mimotopes conferred protection against C. hepaticus colonization of the bile and liver. Future in vivo investigations are necessary to develop and evaluate the immunogenicity and protective efficacy of a multivalent mimotope vaccine consisting of three identified mimotopes against both C. hepaticus and APEC, utilizing the ΔaroA Δasd APEC PSU078 strain as the vaccine vector.