Translation of a laboratory-validated equine herpesvirus-1 specific real-time PCR assay into an insulated isothermal polymerase chain reaction (iiPCR) assay for point-of-need diagnosis using POCKIT™ nucleic acid analyzer

Udeni B.R. Balasuriya 1 Pei-Yu Alison Lee 2 Yun-Long Tsai 3 Chuan-Fu Tsai 3 Yu-Han Shen 3 Hsiao-Fen Grace Chang 3 Ashley Skillman 1 Hwa-Tang Thomas Wang 3 Stéphane Pronost 4, 5 Yan Zhang 6
1 Maxwell H. Gluck Equine Research Center, Department of Veterinary Science
2 GeneReach USA
3 GeneReach USA
4 LABÉO - LABÉO, Pôle d’analyses et de recherche de Normandie
5 U2RM - Unité de Recherche Risques Microbiens
6 Animal Disease Diagnostic Laboratory
Abstract : Equine herpesvirus myeloencephalopathy (EHM), a major problem for the equine industry in the United States, is caused by equine herpesvirus-1 (EHV-1). In addition, EHV-1 is associated with upper respiratory disease, abortion, and chorioretinal lesions in horses. Here we describe the development and evaluation of an inexpensive, user-friendly insulated isothermal PCR (iiPCR) method targeting open reading 30 (ORF30) to detect both neuropathogenic and non-neuropathogenic strains on the field-deployable POCKIT™ device for point-of-need detection of EHV-1. The analytical sensitivity of the EHV-1 iiPCR assay was 13 genome equivalents per reaction. The assay did not cross react with ten non-target equine viral pathogens. Performance of the EHV-1 iiPCR assay was compared to two previously described real-time PCR (qPCR) assays in two laboratories by using 104 archived clinical samples. All 53 qPCR-positive and 46 of the 51 qPCR-negative samples tested positive and negative, respectively, by the iiPCR. The agreement between the two assays was 95.19% (confidence interval 90.48-99.90%) with a kappa value of 0.90. In conclusion, the newly developed EHV-1 iiPCR assay is robust to provide specificity and sensitivity comparable to qPCR assays for the detection of EHV-1 nucleic acid in clinical specimens.
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https://hal-normandie-univ.archives-ouvertes.fr/hal-02167941
Contributeur : Erika Hue <>
Soumis le : vendredi 28 juin 2019 - 12:09:11
Dernière modification le : vendredi 13 septembre 2019 - 12:03:30

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COMUE-NORMANDIE | UNICAEN | BIOTARGEN | BIOTARGEN-MECIIR

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Udeni B.R. Balasuriya, Pei-Yu Alison Lee, Yun-Long Tsai, Chuan-Fu Tsai, Yu-Han Shen, et al.. Translation of a laboratory-validated equine herpesvirus-1 specific real-time PCR assay into an insulated isothermal polymerase chain reaction (iiPCR) assay for point-of-need diagnosis using POCKIT™ nucleic acid analyzer. Journal of Virological Methods, Elsevier, 2017, 241, pp.58-63. ⟨10.1016/j.jviromet.2016.12.010⟩. ⟨hal-02167941⟩

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