Abstract
Outer membrane proteins, such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH), are considered immunodominant antigens for eliciting protective immunity against Vibrio harveyi, the main etiological agent of vibriosis in fish. Cationic antimicrobial peptides (AMPs), such as pleurocidin (PLE), play important roles in activating and recruiting immune cells, thereby contributing to subsequent innate and adaptive immune responses. In the present study, we aimed to use PLE peptide as a potent adjuvant to improve the immunogenicity of V. harveyi recombinant GAPDH (rGAPDH). In order to prepare a controlled-release vaccine, PLE peptide and rGAPDH protein were simultaneously encapsulated into polymeric microparticles made from the biodegradable poly(lactide-co-glycolide) (PLG) polymer. The resulting PLG-encapsulated PLE plus rGAPDH (PLG-PLE/rGAPDH) microparticles, 3.21–6.27 μm in diameter, showed 72%–83% entrapment efficiency and durably released both PLE and rGAPDH for a long 30-day period. Following peritoneal immunization in grouper (Epinephelus coioides), PLG-PLE/rGAPDH microparticles resulted in significantly higher (p < 0.05, nested design) long-lasting GAPDH-specific immunity (serum titers and lymphocyte proliferation) than PLG-encapsulated rGAPDH (PLG-rGAPDH) microparticles. After an experimental challenge of V. harveyi, PLG-PLE/rGAPDH microparticles conferred a high survival rate (85%), which was significantly higher (p < 0.05, chi-square test) than that induced by PLG-rGAPDH microparticles (67%). In conclusion, PLE peptide exhibits an efficacious adjuvant effect to elicit not only improved immunity, but also enhanced protection against V. harveyi in grouper induced by rGAPDH protein encapsulated in PLG microparticles.
Highlights
Grouper, a farmed fish species with high economic value, has rapidly become an important high-quality agricultural product in Southeast Asia
We examined the adjuvant effect of PLE peptide on improving protective immunity induced by recombinant GAPDH (rGAPDH) protein encapsulated into PLG microparticles
No substitution was found in the cloned glyceraldehyde-3-phosphate dehydrogenase (GAPDH) sequence in comparison with the previously reported GAPDH gene (GenBank Accession Number DQ184650.1)
Summary
A farmed fish species with high economic value, has rapidly become an important high-quality agricultural product in Southeast Asia. Current development efforts of subunit vaccines against V. harveyi have been focused mainly on the outer membrane proteins, such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH), since antigenic epitopes on the bacterial surface are favorably accessible to the host immune system [4,5,6,7]. GAPDH is an important enzyme in classical cytosolic glycolysis. It performs other functional activities in different cell compartments [8]. If alternative potent adjuvants that can effectively enhance the immunogenicity of a vaccine antigen [9,10,11], such as GAPDH, are used in the vaccine formulation, improved anti-V. harveyi protective immunity may be achieved in fish
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