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Journal of clinical microbiology1996; 34(6); 1481-1487; doi: 10.1128/jcm.34.6.1481-1487.1996

Detection of equine infectious anemia viral RNA in plasma samples from recently infected and long-term inapparent carrier animals by PCR.

Abstract: Control of equine infectious anemia (EIA) is currently based on detection of anti-EIA virus (EIAV) antibodies. However, serologic diagnostic methods may give false-negative results in infected horses that fail to respond adequately or are in the early stages of infection. We developed a reverse transcriptase nested PCR (RT-nPCR) assay for the detection of viral gag gene sequences in plasma from EIAV-infected horses. The ability of RT-nPCR to detect field strains of EIAV was investigated by assaying plasma samples from 71 horses stabled on EIA quarantine ranches. Positive PCR signals were detected in 63 of 63 horses with EIAV antibody test-positive histories on approved serologic tests, demonstrating that RT-nPCR was probably directed against highly conserved sequences in the viral genome. The RT-nPCR assay, agar gel immunodiffusion test, and conventional virus isolation were compared for detection of early infection in 12 experimentally infected ponies. Viral gag sequences were detected in all 12 animals by 3 days postinfection (p.i.) by RT-nPCR, whereas virus could not be routinely isolated on cell culture until 9 to 13 days p.i. and EIAV antibodies could not be detected by agar gel immunodiffusion until 20 to 23 days p.i. Finally, specificity of the RT-nPCR assay was examined by testing plasma from 43 horses with serologic test-negative histories and no known contact with EIAV-infected animals. Viral gag sequences were not detectable in this control group. These data suggest that the EIAV RT-nPCR assay effectively detects EIAV and is more sensitive than current standard methods for detection of early stages of infection.
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Publication Date: 1996-06-01 PubMed ID: 8735102PubMed Central: PMC229046DOI: 10.1128/jcm.34.6.1481-1487.1996Google Scholar: Lookup
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  • Comparative Study
  • Journal Article
  • Research Support
  • Non-U.S. Gov't
  • Research Support
  • U.S. Gov't
  • P.H.S.

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 discusses a new method for detecting equine infectious anemia (EIA) in horses by identifying viral RNA sequences. This new method, the reverse transcriptase nested PCR (RT-nPCR) assay, proved to be more sensitive and efficient than conventional methods in early detection of EIA.

Introduction

  • The study focusses on equine infectious anemia (EIA), currently detected via the presence of anti-EIA virus (EIAV) antibodies. However, traditional serologic methods may not identify infected horses that don’t respond adequately or are in the early infection stages.
  • The researchers developed a new method, a reverse transcriptase nested PCR (RT-nPCR) assay, to track the viral gag gene sequences in the plasma of EIAV-infected horses. This advanced method could revolutionize the early detection of EIAV and improve control measures over the disease.

Methodology

  • The applicability of RT-nPCR in detecting varying field strains of EIAV was tested by examining plasma samples from 71 horses on EIA quarantine ranches.
  • The team also compared RT-nPCR’s efficacy in early infection detection with the agar gel immunodiffusion test and traditional virus isolation in 12 experimentally infected ponies.
  • The researchers additionally corroborated the specificity of the RT-nPCR method by testing plasma from 43 horses – with negative serologic test histories and no known EIAV contact – to ensure no false-positives were registered.

Results

  • The RT-nPCR assay detected positive PCR signals in all 63 horses with prior EIAV antibody-positive test histories, suggesting the method targets highly conserved sequences in the viral genome.
  • In the experimental ponies, the RT-nPCR detected viral gag sequences by the third day post-infection, much earlier than the conventional method that took 9-13 days for virus isolation and 20-23 days to detect EIAV antibodies.
  • The RT-nPCR assay did not yield any false positives when tested on the control group of horses.

Conclusion

  • The data strongly support that the EIAV RT-nPCR assay is more efficient and sensitive in early infection detection than traditional techniques, implying its potential as an effective tool in controlling EIA.

Cite This Article

APA
Langemeier JL, Cook SJ, Cook RF, Rushlow KE, Montelaro RC, Issel CJ. (1996). Detection of equine infectious anemia viral RNA in plasma samples from recently infected and long-term inapparent carrier animals by PCR. J Clin Microbiol, 34(6), 1481-1487. https://doi.org/10.1128/jcm.34.6.1481-1487.1996

Publication

Journal of clinical microbiology
ISSN: 0095-1137
NlmUniqueID: 7505564
Country: United States
Language: English
Volume: 34
Issue: 6
Pages: 1481-1487

Researcher Affiliations

Langemeier, J L
  • Maxwell Gluck Equine Research Center, Department of Veterinary Science, University of Kentucky, Lexington 40546-0099, USA. JLLANGOO@UKCC.UKY.EDU
Cook, S J
    Cook, R F
      Rushlow, K E
        Montelaro, R C
          Issel, C J

            MeSH Terms

            • Animals
            • Base Sequence
            • Carrier State / diagnosis
            • Carrier State / veterinary
            • Carrier State / virology
            • DNA Primers / genetics
            • Diagnostic Errors
            • Equine Infectious Anemia / diagnosis
            • Equine Infectious Anemia / virology
            • Horses
            • Immunodiffusion / methods
            • Immunodiffusion / statistics & numerical data
            • Infectious Anemia Virus, Equine / genetics
            • Infectious Anemia Virus, Equine / isolation & purification
            • Molecular Sequence Data
            • Polymerase Chain Reaction / standards
            • Polymerase Chain Reaction / statistics & numerical data
            • Polymerase Chain Reaction / veterinary
            • Prospective Studies
            • RNA, Viral / blood
            • RNA, Viral / genetics
            • Reference Standards
            • Sensitivity and Specificity
            • Time Factors
            • Virology / methods
            • Virology / statistics & numerical data

            Grant Funding

            • R01-AI-25850 / NIAID NIH HHS
            • R01-CA-49296 / NCI NIH HHS

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            Citations

            This article has been cited 8 times.
            1. Issel CJ, Scicluna MT, Cook SJ, Cook RF, Caprioli A, Ricci I, Rosone F, Craigo JK, Montelaro RC, Autorino GL. Challenges and proposed solutions for more accurate serological diagnosis of equine infectious anaemia. Vet Rec 2013 Feb 23;172(8):210.
              doi: 10.1136/vr-2012-100735pubmed: 23161812google scholar: lookup
            2. Alvarez I, Gutierrez G, Ostlund E, Barrandeguy M, Trono K. Western blot assay using recombinant p26 antigen for detection of equine infectious anemia virus-specific antibodies. Clin Vaccine Immunol 2007 Dec;14(12):1646-8.
              doi: 10.1128/CVI.00293-07pubmed: 17959820google scholar: lookup
            3. Harrold SM, Cook SJ, Cook RF, Rushlow KE, Issel CJ, Montelaro RC. Tissue sites of persistent infection and active replication of equine infectious anemia virus during acute disease and asymptomatic infection in experimentally infected equids. J Virol 2000 Apr;74(7):3112-21.
              doi: 10.1128/jvi.74.7.3112-3121.2000pubmed: 10708426google scholar: lookup
            4. Lonning SM, Zhang W, Leib SR, McGuire TC. Detection and induction of equine infectious anemia virus-specific cytotoxic T-lymphocyte responses by use of recombinant retroviral vectors. J Virol 1999 Apr;73(4):2762-9.
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