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A novel indirect ELISA based on BP26 mAb to differentiate Brucella melitensis M5-90BPΔ26 vaccine immunization
Yujie Cui1, Shudan Liu2, Guohua Cai2
1College of Animal Science and Technology, Yangtze University, Jingzhou, Hubei, 434025, China.
Abstract:
Brucellosis, primarily caused by the highly invasive Brucella melitensis (B.melitensis), remains a major global zoonotic threat. Differentiating infected from vaccinated animals (DIVA) is critical for effective surveillance. In this study, we developed an indirect ELISA (iELISA) based on a novel monoclonal antibody (mAb) targeting the BP26 protein. Under optimized conditions, the assay established a cut-off value of 0.4, yielding a diagnostic sensitivity of 96.6% (29/30) and specificity of 100% (30/30). The iELISA demonstrated high analytical sensitivity with no cross-reactivity against common pathogens (E. coli, Salmonella, and Y. enterocolitica). Intra- and inter-assay coefficients of variation (CV) were all <10%, confirming robust reproducibility. Comparative analysis with a commercial kit showed high agreement rates across sera from goats immunized with M5-90BPΔ26 (90.18%), M5-90 (94.29%), and non-immune controls (96.02%). These results suggest that this iELISA is a reliable diagnostic tool for distinguishing wild-type infection from M5-90ΔBP26 vaccination in clinical practice.
Insights
A new indirect ELISA (iELISA) using a monoclonal antibody against BP26 protein accurately distinguishes Brucella melitensis infection from vaccination. This diagnostic tool offers high sensitivity and specificity for effective zoonotic disease surveillance.
Area of Science:
- Veterinary Immunology
- Zoonotic Disease Diagnostics
- Bacteriology
Background:
- Brucellosis, caused by Brucella melitensis, is a significant global zoonotic threat.
- Differentiating infected from vaccinated animals (DIVA) is crucial for controlling Brucellosis.
- Existing diagnostic methods require improvement for accurate DIVA strategies.
Purpose of the Study:
- To develop and validate a novel indirect ELISA (iELISA) for differentiating Brucella melitensis infection from vaccination.
- To utilize a monoclonal antibody targeting the BP26 protein for enhanced diagnostic accuracy.
Main Methods:
- Development of an indirect ELISA (iELISA) using a novel monoclonal antibody (mAb) against the BP26 protein.
- Optimization of assay conditions and determination of a cut-off value (0.4).
- Evaluation of diagnostic sensitivity, specificity, analytical sensitivity, and reproducibility (intra- and inter-assay CV <10%).
Main Results:
- The iELISA achieved a diagnostic sensitivity of 96.6% and specificity of 100%.
- No cross-reactivity was observed with common bacterial pathogens like E. coli, Salmonella, and Y. enterocolitica.
- High agreement rates were observed when compared to a commercial kit across vaccinated and non-immune goats.
Conclusions:
- The developed iELISA is a reliable and accurate tool for distinguishing Brucella melitensis infection from M5-90ΔBP26 vaccination.
- This assay supports effective DIVA strategies, crucial for Brucellosis surveillance and control.
- The high performance characteristics suggest its utility in clinical practice for animal health management.
