Development of a novel real-time PCR multiplex assay for detection of Streptococcus equi subspecies equi and Streptococcus equi subspecies zooepidemicus.
Abstract: Strangles is a contagious bacterial disease of horses caused by Streptococcus equi subspecies equi (SEE) that occurs globally. Rapid and accurate identification of infected horses is essential for controlling strangles. Because of limitations of existing PCR assays for SEE, we sought to identify novel primers and probes that enable simultaneous detection and differentiation of infection with SEE and S. equi subsp. zooepidemicus (SEZ). Comparative genomics of U.S. strains of SEE and SEZ (n = 50 each) identified SE00768 from SEE and comB from SEZ as target genes. Primers and probes for real-time PCR (rtPCR) were designed for these genes and then aligned in silico with the genomes of strains of SEE (n = 725) and SEZ (n = 343). Additionally, the sensitivity and specificity relative to microbiologic culture were compared between 85 samples submitted to an accredited veterinary medical diagnostic laboratory. The respective primer and probe sets aligned with 99.7 % (723/725) isolates of SEE and 97.1 % (333/343) of SEZ. Of 85 diagnostic samples, 20 of 21 (95.2 %) SEE and 22 of 23 SEZ (95.6 %) culture-positive samples were positive by rtPCR for SEE and SEZ, respectively. Both SEE (n = 2) and SEZ (n = 3) were identified by rtPCR among 32 culture-negative samples. Results were rtPCR-positive for both SEE and SEZ in 21 of 44 (47.7 %) samples that were culture-positive for SEE or SEZ. The primers and probe sets reported here reliably detect SEE and SEZ from Europe and the U.S., and permit detection of concurrent infection with both subspecies.
Copyright © 2023 Elsevier B.V. All rights reserved.
Publication Date: 2023-06-03 PubMed ID: 37290208DOI: 10.1016/j.vetmic.2023.109797Google 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
This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.
The research article is about the development of a new real-time PCR multiplex assay that is capable of accurately detecting and differentiating infections caused by Streptococcus equi subspecies equi (SEE) and Streptococcus equi subspecies zooepidemicus (SEZ) in horses.
Objective of the Research
- The main purpose of the study was to create an efficient diagnostic tool for detecting and differentiating between infections caused by Streptococcus equi subspecies equi (SEE) and Streptococcus equi subspecies zooepidemicus (SEZ). These infectious bacteria are known to cause the contagious bacterial disease, strangles, in horses worldwide.
Methodology
- The researchers began with comparative genomics of 50 strains each of SEE and SEZ from the US. The target genes identified were SE00768 from SEE and comB from SEZ.
- Primers and probes were then designed for real-time PCR (rtPCR) for these genes.
- In silico alignment with genomes of strains of SEE (725 in number) and SEZ (343 in number) was done.
- Lastly, sensitivity and specificity compared to microbiological culture were evaluated with 85 samples submitted to a recognized veterinary diagnostic laboratory.
Findings
- The primer and probe sets were observed aligning with 99.7% (723 out of 725) isolates of SEE and 97.1% (333 out of 343) of SEZ.
- Among the 85 diagnostic samples analysed, rtPCR identified 95.2% SEE and 95.6% SEZ culture-positive samples, signifying rtPCR’s high sensitivity and specificity compared to conventional microbiological culture techniques.
- RtPCR identified SEE and SEZ infections among culture-negative samples indicating superior detection capability of this technique.
- RtPCR detected concurrent infections of SEE and SEZ in around 47.7% samples that were culture positive, illustrating this tool’s capacity to simultaneously detect and differentiate both subspecies.
Conclusion
- The developed primer and probe sets aid in reliable detection of SEE and SEZ in both Europe and the U.S., and also allow for the detection of concurrent infection with both these subspecies.
- This newly developed rtPCR assay can significantly enhance the detection and management of Strangles in horses, helping to prevent the disease’s transmission and escalation.
Cite This Article
APA
Morris ERA, Schroeder ME, Ferro PJ, Waller AS, McGlennon AA, Bustos CP, Gressler LT, Wu J, Lawhon SD, Boyle AG, Lingsweiler S, Paul N, Dimitrov K, Swinford AK, Bordin AI, Cohen ND.
(2023).
Development of a novel real-time PCR multiplex assay for detection of Streptococcus equi subspecies equi and Streptococcus equi subspecies zooepidemicus.
Vet Microbiol, 284, 109797.
https://doi.org/10.1016/j.vetmic.2023.109797 Publication
Researcher Affiliations
- Department of Large Animal Clinical Sciences, School of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, TX, USA.
- Texas A&M Veterinary Medical Diagnostic Laboratory, College Station, TX, USA.
- Texas A&M Veterinary Medical Diagnostic Laboratory, College Station, TX, USA. Electronic address: pferro@tvmdl.tamu.edu.
- Intervacc AB, Hägersten, Sweden; Department of Biomedical Science and Veterinary Public Health, Swedish University of Agricultural Sciences, Uppsala, Sweden.
- Department of Pathobiology and Population Sciences, Royal Veterinary College, University of London, Hatfield, United Kingdom.
- Universidad de Buenos Aires, Facultad de Ciencias Veterinarias, Cátedra de Enfermedades Infecciosas, Ciudad Autónoma de Buenos Aires, Argentina; Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Argentina.
- Laboratório de Microbiologia e Imunologia Veterinária, Medicina Veterinária, Instituto Federal Farroupilha (IFFar), Frederico Westphalen, Rio Grande do Sul, Brazil.
- Department of Veterinary Pathobiology, School of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, TX, USA.
- Department of Veterinary Pathobiology, School of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, TX, USA.
- Department of Clinical Studies, New Bolton Center, University of Pennsylvania, School of Veterinary Medicine, Kennett Square, PA, USA.
- Texas A&M Veterinary Medical Diagnostic Laboratory, College Station, TX, USA.
- Texas A&M Veterinary Medical Diagnostic Laboratory, College Station, TX, USA.
- Texas A&M Veterinary Medical Diagnostic Laboratory, College Station, TX, USA.
- Texas A&M Veterinary Medical Diagnostic Laboratory, College Station, TX, USA.
- Department of Large Animal Clinical Sciences, School of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, TX, USA.
- Department of Large Animal Clinical Sciences, School of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, TX, USA. Electronic address: ncohen@cvm.tamu.edu.
MeSH Terms
- Animals
- Horses
- Streptococcus equi / genetics
- Real-Time Polymerase Chain Reaction / veterinary
- Horse Diseases / diagnosis
- Horse Diseases / microbiology
- Streptococcus / genetics
- Streptococcal Infections / diagnosis
- Streptococcal Infections / veterinary
- Streptococcal Infections / microbiology
Conflict of Interest Statement
Declaration of Competing Interest The 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.
Citations
This article has been cited 0 times.Use Nutrition Calculator
Check if your horse's diet meets their nutrition requirements with our easy-to-use tool Check your horse's diet with our easy-to-use tool
Talk to a Nutritionist
Discuss your horse's feeding plan with our experts over a free phone consultation Discuss your horse's diet over a phone consultation
Submit Diet Evaluation
Get a customized feeding plan for your horse formulated by our equine nutritionists Get a custom feeding plan formulated by our nutritionists
