• Vaccine Ontology ID: VO_0004113
• Type: Live, attenuated vaccine
• Preparation: The V3526 vaccine candidate was propagated by serial passage in MRC-5 cells to generate pilot-scale pre-master and master virus seed banks that were subsequently used to produce a bulk lot of virus vaccine. The VEEV TrD viruses used in the plaque reduction neutralizing antibody titer (PRNT) assays was produced by Southern Research Institute-Frederick (Frederick, MD) from a starting seed 15 passages from the original donkey virus isolate.
• Description: V3526 is a live-attenuated virus derived by site-directed mutagenesis from a virulent clone of the Venezuelan equine encephalitis virus (VEEV) IA/B Trinidad donkey (TrD) strain. It is intended for human use in protection against Venezuelan equine encephalitis (VEE). Vaccinations with V3526, at doses as low as 102 pfu, were safe and efficacious in protecting horses against a virulent TrD virus challenge.

Horse Response

• Vaccination Protocol: Horses were inoculated in the left shoulder with SC injection of V3526 vaccine or PCM administered SC. Clinical observations were recorded daily throughout the immunization phase and for 14 days PC. Blood and data were collected from all horses throughout the course of the study with Day 0 samples taken pre-vaccination and designated as the day of vaccination. Day 28 PV blood was collected prior to administration of the virus challenge. Blood samples for virus isolation were collected daily on Days 0–10 PV, Days 14 and 21 PV, and daily PC (Days 28–38 PV) (or until euthanasia) and on Day 42 PV for surviving horses.
• Challenge Protocol: Horses in all three trials were challenged on Day 28 PV with a 1 mL SC injection in the left shoulder with either 104 pfu TrD or 104 pfu of 64A99, a challenge dose comparable to that used in previous vaccine protection studies in mice [8]. Horses were monitored for signs of disease for 14 days PC.
  
  Blood was evaluated for viremia (via plaque assay) and sera for VEEV neutralizing antibodies. Blood for hematology analysis was collected once daily from Days −1 to 10 PV, and from Days 27 to 38 PV. Blood was analyzed using a QBC-V hematology analyzer (Clay Adams, Inc.) for hematocrit, red blood cells, platelet and total white blood cell (WBC) count, as well as percentage and absolute numbers of granulocytes and mononuclear (lymphocyte and monocyte) cells (PBMCs).
• Efficacy: None of the horses that received V3526 vaccine or PCM showed clinical signs of disease through the entire pre-challenge period. None of the horses developed a viremia after V3526 inoculation. All 25 horses vaccinated with V3526 vaccine, regardless of dose, survived challenge with either TrD or 64A99. In contrast, all five unvaccinated control horses developed severe disease, including anorexia, fever and malaise, beginning 1–2 days after TrD challenge.
• Description: These studies utilized 33 mixed breed horses, mostly quarter horse stock, with nearly equal numbers of males and females, and in the age range of 3–14 years (as estimated from dental examination). All horses tested negative for pre-existing virus neutralizing antibodies to EEEV, WEEV and VEEV by plaque reduction neutralization (PRN) assays.