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Multiplex Real Time PCR Assay Targeting the eqbE for Rapid Detection and Differentiation of Streptococcus equi from Other Streptococcus Species in Indian Equines.
Abstract: Strangles is a tremendously contagious and chronic infectious disease caused by . The disease compromises the respiratory health of horses and other equines, leading to decreased productivity. The time-consuming culture methods and similarity between and , which can cause opportunistic infections in horses and severe infections in humans, complicate its diagnosis In this study, we applied a multiplex real-time PCR assay on 45 isolates from India, aiming to detect and distinguish it from other species, including . Multiplex real-time PCR using gene of accurately identified all culture-positive samples, yielding 100% sensitivity and specificity. In contrast, conventional PCR revealed a lower sensitivity of 94.1%. Both tests showed no false positives, indicating high specificity The ability to swiftly identify and differentiate from other species facilitates targeted intervention strategies, ultimately aiding in the prevention and mitigation of this debilitating disease. The distribution underscores the necessity of strengthening surveillance, implementing early diagnosis protocols, and adopting targeted control strategies in the affected regions. This is the first report in India of a multiplex real-time PCR for comparative analysis of molecular diagnostic assays for the detection and differentiation of in Indian equines. It established a valuable precedent for future epidemiological surveillance and molecular diagnosis.
© Association of Microbiologists of India 2026. Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
Publication Date: 2026-03-15 PubMed ID: 42860975PubMed Central: PMC13653900DOI: 10.1007/s12088-026-01548-6Google Scholar: Lookup The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
- Journal Article
Summary
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Overview
- This study developed and tested a multiplex real-time PCR assay targeting the eqbE gene for rapid and accurate detection of Streptococcus equi, the causative agent of strangles, from other Streptococcus species in Indian horses.
- The assay showed high sensitivity and specificity compared to conventional PCR, representing an important advance for quick diagnosis and better control of strangles in Indian equine populations.
Background
- Strangles is a highly contagious and chronic respiratory disease predominantly affecting horses and other equines.
- The disease impacts respiratory health and reduces productivity in affected animals.
- Diagnosis is complicated by:
- Time-consuming traditional culture methods.
- The close genetic and phenotypic similarity between Streptococcus equi and other related Streptococcus species such as Streptococcus zooepidemicus.
- Other Streptococcus species can also cause opportunistic infections in horses and severe infections in humans, making accurate differentiation critical.
Objectives
- To apply and evaluate a multiplex real-time PCR assay targeting the eqbE gene for quick detection of Streptococcus equi isolates in Indian horses.
- To distinguish Streptococcus equi from other Streptococcus species found in Indian equines, including Streptococcus zooepidemicus, ensuring diagnostic accuracy.
Methodology
- A total of 45 Streptococcus isolates from Indian equines were tested.
- Multiplex real-time PCR assay targeted the eqbE gene, a marker specific to Streptococcus equi.
- Results from the multiplex real-time PCR assay were compared to conventional PCR methods.
Key Findings
- The multiplex real-time PCR assay successfully identified all culture-positive Streptococcus equi samples, achieving 100% sensitivity and specificity.
- In comparison, conventional PCR showed a slightly lower sensitivity of 94.1% but maintained high specificity with no false positives.
- Both assays showed no false positives, indicating highly reliable differentiation between Streptococcus equi and other Streptococcus species.
Significance
- The rapid and accurate identification of Streptococcus equi enables prompt and targeted interventions such as quarantine or treatment, which can limit the spread of strangles.
- Improved diagnostic protocols inform better surveillance and control measures essential for affected regions in India.
- This study represents the first reported use of multiplex real-time PCR for Streptococcus equi detection in Indian equines, setting a precedent for future molecular diagnostics and epidemiological studies.
- The assay supports early diagnosis and effective management strategies, potentially reducing morbidity and economic losses in the equine industry.
Conclusion
- The multiplex real-time PCR assay targeting the eqbE gene is a reliable, sensitive, and specific diagnostic tool for detecting Streptococcus equi in Indian horses.
- Its ability to differentiate Streptococcus equi from other closely related Streptococcus species aids in better disease control and epidemiological surveillance.
- Adoption of this assay could improve diagnostic turnaround time and help establish early intervention protocols in India.
Cite This Article
APA
Chhabra D, Nagra J, Manuja A, Dahiya R, Prasad M, Goutam U, Kumar B.
(2026).
Multiplex Real Time PCR Assay Targeting the eqbE for Rapid Detection and Differentiation of Streptococcus equi from Other Streptococcus Species in Indian Equines.
Indian J Microbiol, 66(5), 1177-1185.
https://doi.org/10.1007/s12088-026-01548-6 Publication
Researcher Affiliations
- ICAR-National Research Centre on Equines, Hisar, 125001 India.
- Department of Molecular Biology & Genetic Engineering, School of Bioengineering and Biosciences, Lovely Professional University, Phagwara, Punjab India.
- ICAR-National Research Centre on Equines, Hisar, 125001 India.
- ICAR-National Research Centre on Equines, Hisar, 125001 India.
- ICAR-National Research Centre on Equines, Hisar, 125001 India.
- ICAR-National Research Centre on Equines, Hisar, 125001 India.
- Department of Molecular Biology & Genetic Engineering, School of Bioengineering and Biosciences, Lovely Professional University, Phagwara, Punjab India.
- ICAR-National Research Centre on Equines, Hisar, 125001 India.
Conflict of Interest Statement
Conflicts of interestThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
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