Abstract / Summary
Abstract Background Madariaga virus (MADV) is an understudied mosquito-borne alphavirus that circulates widely in Latin America. Following spillover from its enzootic cycle, MADV can infect both equids and humans, with equine cases often preceding human infections. Despite the high equine morbidity and case fatality rates reported during epizootics, it remains unclear whether infected horses can contribute to virus amplification by serving as a source of infection for mosquitoes. We sought to establish a large-animal experimental system for studying MADV infection and equine-to-mosquito transmission under high-containment conditions. Methods Two pilot trials were conducted. Trial 1 evaluated the feasibility of housing horses and conducting direct mosquito-feeding under containment. In Trial 2, two horses were inoculated subcutaneously with MADV and monitored for 14 days to characterize clinical, laboratory, virological, and serological responses, with histopathological analysis performed after euthanasia. Two mosquito species were allowed to feed directly on the horses during the first 8 days post-inoculation and were subsequently tested for MADV RNA. Results Trial 1 demonstrated the feasibility of horse housing and direct mosquito-feeding under containment. In Trial 2, conducted in the high-containment spaces, neither horse developed overt clinical disease, although one exhibited delayed hindlimb proprioception for multiple days. MADV RNA was not detected in horse plasma or mosquitoes that fed on inoculated horses. Both horses seroconverted, with one developing a stronger neutralizing antibody response. Hematological and biochemical parameters remained within reference intervals. No gross or histopathological central nervous system lesions were observed. Conclusions These pilot studies establish the feasibility of a large-animal BSL-3 platform for experimental MADV infection and direct equine-to-mosquito transmission studies and provide an initial experimental evaluation of whether infected horses can serve as a source of MADV for mosquito infection. Importantly, this system can be expanded to include mosquito-to-horse transmission. This would allow both directions of the mosquito-equid transmission cycle to be reproduced experimentally, enabling studies of transmission determinants and pathogenesis, as well as evaluation of vaccine efficacy, durability of immunity, and therapeutic interventions. The platform could be applied to MADV and other mosquito-borne arboviruses of human and animal health importance that infect equids.