Study of bovine brucellosis in continental Ecuador

BACKGROUND Brucellosis is a zoonotic infectious disease that is a serious health problem for livestock and causes high economic losses in many developing countries around the world, hampering trade and exports. Although many efforts have been made to control brucellosis, the disease continues to spread, especially in endemic countries, and represents one of the most serious health threats to animals and humans. In Ecuador, Brucella abortus is the species most commonly involved in bovine brucellosis, which is characterized by spontaneous abortions in the last third of pregnancy, retained fetal membranes and reduced milk production. Infected animals can carry the bacteria asymptomatically and serve as a source of infection for other animals and humans. The objective of this thesis was to review the bovine brucellosis status in continental Ecuador, with special attention to seroprevalence, risk factors, evaluation of vaccine quality control and vaccination procedures, development of diagnostic methods, isolation and typing of strains, and to provide recommendations to the Brucellosis Program of the Animal and Plant Health Regulatory and Control Agency (AGROCALIDAD). METHODOLOGY In the first study, a stratified cross-sectional sampling plan was used to evaluate the seroprevalence of bovine brucellosis and associated risk or protective factors in 290 herds originating from 23 provinces of continental Ecuador, representing a total of 3737 cows. cELISA was used to detect antibodies against Brucella and an Epidemiologic survey was conducted to evaluate risk factors in these 290 farms. The second study conducted in Ecuador evaluated the quality control of Brucella vaccines supplied and registered in the country. A quality control analysis of the vaccines was performed at the national reference laboratory, taking into account the World Organization for Animal Health control guidelines. In addition, a survey of 95 veterinarians and 30 commercial companies was conducted to determine vaccination and vaccine management practices in Ecuador. The third study focused on the isolation of Brucella strains from 25 lymph node samples and 50 milk samples that were positive in serological tests by ELISA and Rose Bengal for the years 2022, 2023 and 2024 within the framework of the National Brucellosis Control Program of Ecuador. Bacterial cultures were performed in specific media (CITA and Farrel) and molecular identification was performed by PCR techniques and MLVA (Multi-Locus Variable Number Tandem Repeat Analysis). In the study of Bayesian estimation of the true prevalence of bovine brucellosis in Ecuador, the sensitivity and specificity of a cELISA and an iELISA based on a new synthetic antigen were evaluated. A stratified random sample was selected from the continental bovine population of Ecuador, consisting of 3299 cattle over 24 months of age from 223 herds. Estimation of true prevalence and test performance (sensitivity and specificity) was performed in all animals (n=3299; model 1) and in three subgroups: unvaccinated animals (n=2506; model 2), vaccinated with the S19 strain (n=383; model 3) and vaccinated with the RB51 strain (n=392; model 4). RESULTS The results of the first study showed that the apparent prevalence was 21.3% at the herd level and 6.2% at the animal level. Multivariate analysis revealed that herd size (>70 ha) and number of calves per animal were risk factors, while location in the eastern region and absence of clinical signs were identified as protective factors. The results of the second study showed that two of the three companies registered in Ecuador did not meet the quality control parameters established by the World Organization for Animal Health in terms of bacterial count. In addition, vaccination practices in the field were evaluated through surveys of veterinarians and technicians, as well as in veterinary stores, showing 62.29% and 56.86% of compliance, respectively. The third study confirmed the presence of Brucella abortus strains of biovars 1, 2 and 4 in Ecuador, the first detection of biovar 2 in the country. Accurate molecular identification is essential for epidemiologic studies and control strategies. In study 4, the performance of two ELISA tests (cELISA and iELISA) to diagnose bovine brucellosis in cattle in Ecuador was analyzed. Both tests showed similar sensitivity: 94.02% for cELISA and 94.05% for iELISA. However, the specificity of iELISA was higher (98.10%) compared to cELISA (95.85%). The actual prevalence of brucellosis was 1.63% in unvaccinated cattle, 0.97% in those vaccinated with S19 and 2.75% in those vaccinated with RB51. The findings indicate that iELISA, due to its higher specificity, could be the good alternative to detect bovine brucellosis in Ecuador, underlining the importance of optimizing the control program, particularly on the adequate use of the vaccination protocol. CONCLUSION The failure to control this disease is due to several reasons, such as the lack of highly effective vaccines, the lack of commitment of farmers to certify farms free of brucellosis, the limited importance of government investment at national and regional level, and the main factor was the lack of budget to deal with this disease. A "One Health" approach between the Ministry of Agriculture and the Ministry Of Public Health is needed at the country level to control this disease and improve the sustainability of the livestock sector at the regional and national levels. These findings provide a solid basis for improving the national bovine brucellosis control program in Ecuador, including education on good veterinary practices, strengthening vaccine quality control, and management of identified risk factors.