Optimization and validation of indirect ELISA using truncated TssB protein for the serodiagnosis of glanders amongst equines.
Abstract: To express truncated TssB protein of Burkholderia mallei and to evaluate its diagnostic efficacy for serological detection of glanders among equines. Methods: In an attempt to develop recombinant protein based enzyme-linked immunosorbent assay (ELISA), N-terminal 200 amino acid sequences of B. mallei TssB protein-a type 6 secretory effector protein--were expressed in prokaryotic expression system. Diagnostic potential of recombinant TssB protein was evaluated in indirect ELISA using a panel of glanders positive (n = 49), negative (n = 30), and field serum samples (n = 1811). Cross-reactivity of the assay was assessed with equine disease control serum and human melioidosis positive serum. Results: In comparison to CFT, diagnostic sensitivity and specificity of ELISA were 99.7% and 100%, respectively. Conclusions: The indirect ELISA method using the truncated TssB offered safer and more rapid and efficient means of serodiagnosis of glanders in equines. These data highlight the use of TssB as potential diagnostic antigen for serological diagnosis of glanders.
Publication Date: 2014-02-03 PubMed ID: 24672321PubMed Central: PMC3932216DOI: 10.1155/2014/469407Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
- Validation Study
Summary
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This study successfully developed an optimized and validated indirect ELISA method that uses a truncated TssB protein for glanders diagnosis amongst equines, exhibiting an impressive sensitivity and specificity.
Introduction and Objective
- The motivation of this research is to diagnose glanders, a disease that affects equines, using serological detection.
- Specifically, the researchers aimed to express the truncated TssB protein of Burkholderia mallei and evaluate its usability for diagnostic purposes.
Methods
- In their attempt to develop a recombinant protein-based enzyme-linked immunosorbent assay (ELISA), the scientists expressed the N-terminal 200 amino acid sequences of B. mallei’s TssB protein. This protein is known as a type 6 secretory effector protein.
- Next, they evaluated the diagnostic potential of the recombinant TssB protein via indirect ELISA. This was performed using a panel of glanders positive (n = 49), negative (n = 30), and field serum samples (n = 1811).
- The researchers also checked the cross-reactivity of the assay with equine disease control serum and human melioidosis positive serum to ensure that the ELISA does not mistake other diseases for glanders.
Results
- When compared to the complement fixation test (CFT), which is another method of diagnosing glanders, the diagnostic sensitivity and specificity of the ELISA were 99.7% and 100%, respectively.
- Demonstrating such high sensitivity and specificity means that the new ELISA method can very accurately detect true positive cases and true negative cases of glanders.
Conclusions
- Using the truncated TssB protein, the indirect ELISA method offers a safer and more rapid and efficient means of diagnosing glanders amongst equines compared to other current methods.
- These data highlight that the TssB could serve as a powerful diagnostic antigen for the serological diagnosis of glanders.
Cite This Article
APA
Singha H, Malik P, Goyal SK, Khurana SK, Mukhopadhyay C, Eshwara VK, Singh RK.
(2014).
Optimization and validation of indirect ELISA using truncated TssB protein for the serodiagnosis of glanders amongst equines.
ScientificWorldJournal, 2014, 469407.
https://doi.org/10.1155/2014/469407 Publication
Researcher Affiliations
- National Research Centre on Equines, Sirsa Road, Hisar, Haryana 125 001, India.
- National Research Centre on Equines, Sirsa Road, Hisar, Haryana 125 001, India ; Veterinary Type Culture Collection, National Research Centre on Equines, Sirsa Road, Hisar, Haryana 125 001, India.
- National Research Centre on Equines, Sirsa Road, Hisar, Haryana 125 001, India.
- National Research Centre on Equines, Sirsa Road, Hisar, Haryana 125 001, India.
- Department of Microbiology, Kasturba Medical College, Manipal University, Manipal, Karnataka 576104, India.
- Department of Microbiology, Kasturba Medical College, Manipal University, Manipal, Karnataka 576104, India.
- National Research Centre on Equines, Sirsa Road, Hisar, Haryana 125 001, India.
MeSH Terms
- Animals
- Base Sequence
- Blotting, Western
- Burkholderia mallei / genetics
- Enzyme-Linked Immunosorbent Assay / methods
- Glanders / diagnosis
- Horse Diseases / diagnosis
- Horses
- Molecular Sequence Data
- Sequence Homology, Nucleic Acid
- Serologic Tests
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Citations
This article has been cited 9 times.- Ichikawa Y, Iinuma Y, Okagawa T, Shimbo R, Enkhtuul B, Khurtsbaatar O, Kinoshita Y, Niwa H, Aoshima K, Kobayashi A, Batbaatar V, Ohashi K, Kimura T. Comparison of immunogenicity of 17 Burkholderia mallei antigens and whole cell lysate using indirect ELISA. J Vet Med Sci 2025 Apr 1;87(4):394-401.
- Grause JF, Elschner MC, Ledesma NA, Murphy G. Development and validation of a chemiluminescent western blot assay for glanders (Burkholderia mallei) serodetection. J Vet Diagn Invest 2024 Mar;36(2):283-286.
- Charron P, Gao R, Chmara J, Hoover E, Nadin-Davis S, Chauvin D, Hazelwood J, Makondo K, Duceppe MO, Kang M. Influence of genomic variations on glanders serodiagnostic antigens using integrative genomic and transcriptomic approaches. Front Vet Sci 2023;10:1217135.
- Elschner MC, Melzer F, Singha H, Muhammad S, Gardner I, Neubauer H. Validation of a Commercial Glanders ELISA as an Alternative to the CFT in International Trade of Equidae. Front Vet Sci 2021;8:628389.
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