Poly(lactic-co-glycolic acid) nanoparticle delivery enhances the immunogenicity and anticoccidial activity of recombinant multi-epitope 14EGT protein against multiple chicken Eimeria infections.

Chicken coccidiosis is frequently caused by single or multiple Eimeria species, whereas protective immunity is largely species-specific. Recombinant multi-epitope proteins derived from conserved Eimeria antigens represent promising candidates for cross-protection against multiple chicken Eimeria species, but their efficacy is often limited by insufficient antigen delivery. In this study, r14EGT, a recombinant multi-epitope protein containing concentrated T-cell epitopes from four conserved Eimeria antigens, including 14-3-3, EF-2, GAPDH, and transhydrogenase, was encapsulated into poly(lactic-co-glycolic acid) (PLGA) nanoparticles to enhance its immunogenicity and anticoccidial activity. The resulting PLGA-r14EGT nanoparticles were approximately 105 nm in diameter with an encapsulation efficiency of 62.46%. In an Eimeria tenella challenge model, PLGA-r14EGT achieved the highest anticoccidial index (ACI) among the tested r14EGT-based formulations and produced the greatest reduction in intestinal lesion scores. Immunization with PLGA-r14EGT also enhanced both cellular and humoral immune responses, as evidenced by increased splenic CD4+ and CD8+ T-cell proportions, elevated IFN-γ, IL-4, and IL-17 A transcription, and higher serum antigen-specific IgY levels. Further optimization identified ocular-nasal immunization with 50 μg PLGA-r14EGT at 0, 7, and 14 days of age as the most effective regimen. Under this schedule, PLGA-r14EGT achieved ACI values of 174.95, 171.75, and 169.72 against Eimeria tenella, Eimeria maxima, and Eimeria acervulina single-species infections, respectively, and 161.13 against mixed infection. These findings indicate that PLGA nanoparticle delivery enhances the biological activity of r14EGT protein and supports its potential for controlling single and mixed Eimeria infections in chickens.