Brucellosis Abattoir Surveillance Using Serology and Molecular Tests Among Livestock in Free State and Limpopo Provinces, South Africa

Brucellosis remains a significant zoonotic disease globally, causing considerable economic and public health consequences, particularly in high-risk areas of South Africa (SA) with extensive animal production, including the Limpopo and Free State provinces. This study used a combination of serological, microbiological, and molecular assays to investigate the prevalence and characteristics of Brucella in 580 slaughtered livestock (384 cattle and 196 sheep) in Limpopo and the Free State (192 cattle and 98 sheep in each province) and to compare the diagnostic findings obtained using these assays. Tissue samples collected from each animal included liver, lung, spleen, and lymph nodes. Using the standard diagnostic rose Bengal test (RBT) and complement fixation test (CFT) in series (RBT-CFT), only 2.6% (10/384) of cattle were positive, with 2.1% (95% CI: 0.7–4.9) and 3.1% (95% CI: 1.3–6.3) in the Free State and Limpopo, respectively, while 4.7% (18/384) were positive by RBT-indirect enzyme-linked immunosorbent assay (iELISA), with 5.2% (95% CI: 2.7–9.0) and 4.2% (95% CI: 2.0–7.7) in the Free State and Limpopo, respectively. The molecular prevalence was 18.5% for Brucella deoxyribonucleic acid (DNA), while 97% of PCR-positive cattle tested seronegative in both provinces. Ovine samples were largely seronegative despite showing higher polymerase chain reaction (PCR) positivity (28.6%). Brucella abortus predominated in both species, with B. melitensis and occasional mixed infections also detected. Mixed infections in this study were defined as samples producing multiple species-specific bands within the same sample, consistent with the presence of more than one Brucella species during multiplex PCR analysis. While liver and spleen tissue provided reliable PCR detection, bacterial culture yielded a 0% isolation rate. These findings, together with reports from studies in brucellosis-endemic areas in sub-Saharan Africa, demonstrate discrepancies between traditional serological methods and molecular methods, including detection of Brucella DNA in seronegative animals. This may indicate that some infected animals (chronic or latent carriers) are not identified by conventional serological assays alone, particularly in endemic settings. To strengthen surveillance and enhance the detection of exposed animals, the incorporation of iELISA as a complementary tool into South Africa’s routine surveillance programmes may be beneficial, particularly when combined with regular testing of all animals.