Rabies, a zoonotic disease, causes > 55,000 human deaths globally and results in at least 500 million dollars in losses every year. The currently available rabies vaccines are mainly inactivated and attenuated vaccines, which have been linked with clinical diseases in animals. Thus, a rabies vaccine with high safety and efficacy is urgently needed. Peptide vaccines are known for their low cost, simple production procedures and high safety. Therefore, in this study, we examined the efficacy of multi-epitope-based vaccine candidates against rabies virus. The ability of various peptides to induce epitope-specific responses was examined, and the two peptides that possessed the highest antigenicity and conservation, i.e., AR16 and hPAB, were coated with adjuvant canineGp96 and used to prepare vaccines. The peptides were prepared as an emulsion of oil in water (O/W) to create three batches of bivalent vaccine products. The vaccine candidates possessed high safety. Virus neutralizing antibodies were detected on the day 14 after the first immunization in mice and beagles, reaching 5–6 IU/mL in mice and 7–9 IU/mL in beagles by day 28. The protective efficacy of the vaccine candidates was about 70%–80% in mice challenged by a virulent strain of rabies virus. Thus, a novel multi-epitope-based rabies vaccine with Gp96 as an adjuvant was developed and validated in mice and dogs. Our results suggest that synthetic peptides hold promise for the development of novel vaccines against rabies.
Here, we introduce an additional method that we believe has merits with respect to reducing labor, decreasing wait times, improving objectivity, and decreasing long-term costs. This new method, called fluorescently labeled infected cell inoculum titration (FLICIT), utilizes high-throughput fluorescent microscopy instrumentation such as Cytation 3 from Biotek, and is therefore only applicable to transgenic viruses that cause host cells to express fluorescent proteins. For the purpose of developing this method, we used the recombinant virus HSV-1 strain 17-Syn+, which expresses green fluorescent protein (GFP) (Figure 1A) and exhibits the same replication pattern as the wild type counterpart (Foster et al., 1998). This new method relies on titrating the inoculum so that dilution ensures that the rate of infection is less than 50%, with cells infected by nearly one viral particle each; thus, each fluorescent signal can be attributed to a single viral particle.