Development, evaluation, and laboratory validation of immunoassays for the diagnosis of equine infectious anemia (EIA) using recombinant protein produced from a synthetic p26 gene of EIA virus.
Abstract: Equine infectious anemia (EIA)-a retroviral disease caused by equine infectious anemia virus (EIAV)-is a chronic, debilitating disease of horses, mules, and donkeys. EIAV infection has been reported worldwide and is recognized as pathogen of significant economic importance to the horse industry. This disease falls under regulatory control program in many countries including India. Control of EIA is based on identification of inapparent carriers by detection of antibodies to EIAV in serologic tests and "Stamping Out" policy. The current internationally accepted test for diagnosis of EIA is the agar gel immune-diffusion test (AGID), which detects antibodies to the major gag gene (p26) product. The objective of this study was to develop recombinant p26 based in-house immunoassays [enzyme linked immunosorbent assays (ELISA), and AGID] for EIA diagnosis. The synthetic p26 gene of EIAV was expressed in Escherichia coli and diagnostic potential of recombinant p26 protein were evaluated in ELISA and AGID on 7,150 and 1,200 equine serum samples, respectively, and compared with commercial standard AGID kit. The relative sensitivity and specificity of the newly developed ELISA were 100 and 98.6 %, respectively. Whereas, relative sensitivity and specificity of the newly developed AGID were in complete agreement in respect to commercial AGID kit. Here, we have reported the validation of an ELISA and AGID on large number of equine serum samples using recombinant p26 protein produced from synthetic gene which does not require handling of pathogenic EIAV. Since the indigenously developed reagents would be economical than commercial diagnostic kit, the rp26 based-immunoassays could be adopted for the sero-diagnosis and control of EIA in India.
Publication Date: 2013-08-08 PubMed ID: 24426297PubMed Central: PMC3832691DOI: 10.1007/s13337-013-0149-9Google Scholar: Lookup
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- Journal Article
- Antibodies
- Diagnosis
- Diagnostic Technique
- Disease
- Disease Diagnosis
- Enzyme-Linked Immunosorbent Assay (ELISA)
- Epidemiology
- Equine Health
- Equine Infectious Anemia
- Horses
- Immune Response
- Immunology
- Infection
- Infectious Disease
- Laboratory Methods
- Pathogens
- Recombinant Proteins
- Veterinary Care
- Veterinary Medicine
- Veterinary Research
- Virus
Summary
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This research focused on the development, assessment, and validation of diagnostic tests for equine infectious anemia (EIA), a disease affecting horses and related animals, using a recombinant protein produced from a synthetic gene of the virus causing EIA. The newly developed tests demonstrated high sensitivity and specificity, proving their potential for EIA diagnosis without having to handle the pathogenic virus, thus, making them a safer and cost-effective alternative, particularly in India.
Study Objective and Background
- The goal of this study was to fabricate immunoassays based on a recombinant p26 protein, produced from a synthetic gene unique to the equine infectious anemia virus (EIAV).
- This research was driven by the need for efficient diagnosis of EIA, a prevalent and economically costly disease affecting horses, mules, and donkeys worldwide, including India, where the disease is under regulatory control.
- Conventional diagnosis of EIA is performed using the agar gel immune-diffusion test (AGID), which detects antibodies to the major gag gene (p26) product. However, this method requires handling the live EIAV, posing significant safety risks.
Methodology and Testing
- The researchers created a synthetic p26 gene of EIAV and used it to generate a recombinant p26 protein in Escherichia coli.
- The diagnostic potential of this recombinant p26 protein was then evaluated in two types of immunoassays: enzyme linked immunosorbent assays (ELISA), and AGID. These assays were performed on a large number of equine serum samples (7,150 for ELISA and 1,200 for AGID).
- Performance of these newly developed tests was compared against a commercially available standard AGID kit to validate their accuracy and reliability.
Outcomes and Implications
- The newly developed ELISA and AGID tests demonstrated high sensitivity (100% and similar to AGID kit, respectively) and specificity (98.6% and equivalent to AGID kit, respectively), indicating their effectiveness for EIA diagnosis.
- Being based on a synthetic EIAV gene, these tests do not require handling the harmful EIAV, thereby enhancing safety for lab personnel.
- As these diagnostic tests use indigenously developed reagents, they are likely to be more cost-effective than the commercially available kits, making them a feasible option for controlling EIA in India.
Cite This Article
APA
Singha H, Goyal SK, Malik P, Khurana SK, Singh RK.
(2013).
Development, evaluation, and laboratory validation of immunoassays for the diagnosis of equine infectious anemia (EIA) using recombinant protein produced from a synthetic p26 gene of EIA virus.
Indian J Virol, 24(3), 349-356.
https://doi.org/10.1007/s13337-013-0149-9 Publication
Researcher Affiliations
- National Research Centre on Equines, Sirsa Road, Hisar, 125 001 Haryana India.
- National Research Centre on Equines, Sirsa Road, Hisar, 125 001 Haryana India.
- National Research Centre on Equines, Sirsa Road, Hisar, 125 001 Haryana India.
- National Research Centre on Equines, Sirsa Road, Hisar, 125 001 Haryana India.
- National Research Centre on Equines, Sirsa Road, Hisar, 125 001 Haryana India.
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Citations
This article has been cited 2 times.- Hu Z, Guo K, Du C, Sun J, Naletoski I, Chu X, Lin Y, Wang X, Barrandeguy M, Samuel M, Wang W, Lau PI, Wernery U, Raghavan R, Wang X. Development and evaluation of a blocking ELISA for serological diagnosis of equine infectious anemia.. Appl Microbiol Biotechnol 2023 May;107(10):3305-3317.
- Singha H, Goyal SK, Malik P, Singh RK. Use of heavy water (D2O) in developing thermostable recombinant p26 protein based enzyme-linked immunosorbent assay for serodiagnosis of equine infectious anemia virus infection.. ScientificWorldJournal 2014;2014:620906.
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