A polymerase chain reaction for detection of equine herpesvirus-1 in routine diagnostic submissions of tissues from aborted foetuses.
Abstract: Equine herpesvirus 1 (EHV-1) is the causative agent of abortion, perinatal foal mortality, neurological and acute respiratory diseases in horses. Conventional laboratory diagnosis involving viral isolation from aborted foetuses is laborious and lengthy and requires processing of samples within 24 h of collection, which is problematic for samples that come from long distances. The aim of this study was to develop a polymerase chain reaction (PCR) assay useful in Argentina to detect DNA sequences of EHV-1 in different tissues from aborted equine foetuses with variable quality of preservation and without the use of conventional DNA fenolic extraction. Several DNA extraction protocols and primers were evaluated. The amplification method was standardized and its specificity was analysed using 38 foetal samples of variable quality of preservation. Of the 38 different foetal tissues, nine livers, six spleens and two lungs in good preservation and eight livers, one spleen and four lungs in a poor state of preservation were positive for PCR. EHV-1 was recovered only from the nine livers, five spleens and two lungs in good preservation. No virus was isolated from the samples that were poorly preserved. Viral isolation was confirmed by cytopathic effect and indirect immunofluorescence. The specificity of the PCR results was confirmed by the restriction endonuclease digestion of PCR products and hybridization.
Publication Date: 2001-07-27 PubMed ID: 11471844DOI: 10.1046/j.1439-0450.2001.00455.xGoogle Scholar: Lookup
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- Journal Article
Summary
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The researchers have developed an efficient method to detect Equine Herpesvirus 1 (EHV-1), a virus causing abortion and mortality in foals, in aborted horse fetuses using a polymerase chain reaction (PCR) technique. The PCR developed can work with preserved tissues, overcoming previous hurdles in diagnosing this disease.
Objective of the Study
- The aim of the research was to establish a polymerase chain reaction (PCR) technique that could detect sequences of EHV-1 in various tissue samples from aborted equine foetuses irrespective of their preservation quality.
- This research aimed to overcome the issue of the traditional viral isolation method from aborted foetuses, which is labor-intensive, time-consuming, and demands samples within 24 hours of collection. This is a significant issue for samples arriving from large distances.
Methodology
- In this study, the researchers evaluated numerous DNA extraction protocols and primers to standardize an efficient PCR amplification method.
- The researchers examined the assay’s specificity using 38 tissue samples from foetuses that varied in preservation quality.
- The samples included several organs such as the liver, spleen, and lungs, some in a good state of preservation while others were in poor condition.
- The PCR results for each sample were confirmed by restriction endonuclease digestion and hybridization.
- Viral isolation results were verified by cytopathic effect and indirect immunofluorescence.
Results
- Out of the 38 various foetal tissues, positive PCR results were seen in nine livers, six spleens, and two lungs in good preservation, along with eight livers, one spleen, and four lungs in a poor state of preservation.
- EHV-1 was recovered primarily from the nine livers, five spleens, and two lungs, which were in good preservation.
- No virus was isolated from samples in poor preservation, illustrating the efficacy of the developed PCR method in detecting the virus across samples of different preservation states.
- The study successfully demonstrated the specificity of the PCR, making it a reliable detection tool.
Cite This Article
APA
Galosi CM, Vila Roza MV, Oliva GA, Pecoraro MR, Echeverría MG, Corva S, Etcheverrigaray ME.
(2001).
A polymerase chain reaction for detection of equine herpesvirus-1 in routine diagnostic submissions of tissues from aborted foetuses.
J Vet Med B Infect Dis Vet Public Health, 48(5), 341-346.
https://doi.org/10.1046/j.1439-0450.2001.00455.x Publication
Researcher Affiliations
- Department of Virology, Faculty of Veterinary Sciences, National University of La Plata, Argentina. cmgalosi@fcv.medvet.unlp.edu.ar
MeSH Terms
- Abortion, Veterinary / embryology
- Abortion, Veterinary / virology
- Animals
- DNA, Viral / analysis
- Fetus / virology
- Herpesviridae Infections / embryology
- Herpesviridae Infections / veterinary
- Herpesviridae Infections / virology
- Herpesvirus 1, Equid / genetics
- Herpesvirus 1, Equid / isolation & purification
- Horse Diseases / embryology
- Horse Diseases / virology
- Horses
- Polymerase Chain Reaction / methods
- Polymerase Chain Reaction / veterinary
- Sensitivity and Specificity
Citations
This article has been cited 3 times.- Taktaz Hafshejani T, Nekoei S, Vazirian B, Doosti A, Khamesipour F, Anyanwu MU. Molecular Detection of Equine Herpesvirus Types 1 and 4 Infection in Healthy Horses in Isfahan Central and Shahrekord Southwest Regions, Iran. Biomed Res Int 2015;2015:917854.
- Mori E, Lara Mdo C, Cunha EM, Villalobos EM, Mori CM, Soares RM, Brandão PE, Fernandes WR, Richtzenhain LJ. Molecular characterization of Brazilian equid herpesvirus type 1 strains based on neuropathogenicity markers. Braz J Microbiol 2015 Jun;46(2):565-70.
- Smith KL, Li Y, Breheny P, Cook RF, Henney PJ, Sells S, Pronost S, Lu Z, Crossley BM, Timoney PJ, Balasuriya UB. New real-time PCR assay using allelic discrimination for detection and differentiation of equine herpesvirus-1 strains with A2254 and G2254 polymorphisms. J Clin Microbiol 2012 Jun;50(6):1981-8.
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