The persistent and deadly cholera remains a significant burden on global health. Current countermeasures are inadequate against cholera. Toxin-coregulated pilin (TCP) has long been a target for understanding the biogenesis of Vibrio cholerae infection and for developing subunit vaccines against cholera. In this study, we examined TcpA B-cell epitopes of the O1 classical and El Tor biotypes, as well as the O139 serogroup, for functionality, potential biogenesis, and vaccine application. By empirically mapping TcpA functional epitopes across V. cholerae biotypes and serogroups, we found that antibodies raised against a shared epitope (EP #1, IQSQNMTKAAQ) provided partial protection against V. cholerae bacterial adherence in vitro . Data also showed that classical biotype TcpA epitope #6, ASKNLDLTNITHVE, is more effective in inducing functional antibodies against adherence of the classical biotype, but not against the El Tor biotype. Similarly, El Tor biotype TcpA epitopes #4 (FAAVADLGDFETSV) and #6 (GSANLNLTNITHVE) are effective in inducing functional antibodies against the adherence of El Tor biotype and O139 serogroup, but not against the classical biotype. This decodes early observations of a lack of cross-protection from TcpA-derived immunity. Interestingly, combined immunity raised against the classical epitope #6 and the El Tor epitope #6 (or #4) effectively protected against adherence from both biotypes. These results reveal the mechanism underlying the lack of protective immunity conferred by biotype-specific TcpA and indicate a combined epitope immunity for protection across biotypes and serogroups, providing a better understanding of TcpA immunity, perhaps its biogenesis, and guiding the development of cholera vaccines. IMPORTANCE: Current countermeasures are inadequate against cholera, a disease that remains a significant global health burden. Oral cholera vaccines (OCVs) prequalified by the World Health Organization (WHO) are serogroup-specific. In addition to the supply shortage, these whole-cell vaccines provide limited protection to children under 5 years of age in endemic countries, the most vulnerable population to cholera, urgently calling for more effective prevention strategies. By examining functional immunity raised against shared and biotype-specific TcpA epitopes, this study revealed the mechanism underlying why TCP-related immunity from the O1 classical biotype does not extend protection to the O1 El Tor biotype. The current study also showed that a combined biotype-epitope immunity provides cross-protection against O1 El Tor and classical biotypes, as well as against the O139 serogroup, potentially paving the way for the development of a cross-protective vaccine against Vibrio cholerae infection.

Canine brucellosis is an under-recognized zoonotic disease with important reproductive and public health implications. This study investigated molecular and serological evidence of Brucella infection in owned domestic dogs presented to veterinary clinics and referral facilities in Delhi, Bareilly (Uttar Pradesh), and Mannuthy (Kerala), and evaluated the performance of commonly used diagnostic tests. A total of 448 sampling units were included. Overall, 382 blood-derived DNA samples were screened by genus-specific PCR targeting the bcsp31 gene, 356 serum samples were tested by ELISA and agar gel immunodiffusion (AGID), and 22 vaginal swab-derived samples were screened by PCR. PCR detected Brucella DNA in 40/382 (10.47%) blood-derived samples, with all positive samples originating from Delhi (40/174; 22.99%), and in 1/22 (4.50%) vaginal swab-derived samples. ELISA detected antibodies in 34/356 (9.55%) samples, while AGID detected 13/356 (3.65%). Overall, 43/356 (12.08%) samples were positive by at least one serological test. Agreement between ELISA and AGID was 89.04%, with Cohen’s κ = 0.12; when ELISA was used as the comparator, AGID showed a relative sensitivity of 11.76% and relative specificity of 97.20%. Bayesian latent-class analysis estimated the model-derived latent Brucella-positive infection-related state at 17.84% (95% credible interval [CrI]: 11.99–25.13%). This estimate represents an underlying infection-related state inferred from the joint diagnostic-test results and should not be interpreted as confirmed active infection or clinical disease. Positivity was significantly higher among dogs older than two years (p < 0.05), while no significant association was observed with sex. Overall, the findings provide molecular and serological evidence of Brucella in the sampled owned domestic dogs and highlight the complementary information provided by molecular and serological testing. Further species-level characterization and structured surveillance are warranted to better define the epidemiology and public-health significance of Brucella infection detected in dogs in India.