Multiplex real-time PCR for the detection and differentiation of equid gammaherpesvirus 2 and 5.
J Virol Methods
; 310: 114615, 2022 12.
Article
in En
| MEDLINE
| ID: mdl-36087793
ABSTRACT
Equid gammaherpesvirus 2 (EHV-2) and 5 (EHV-5) are widely distributed in the equines. Although their pathogenic potential is not yet fully understood, they appear to play a role in disease patterns like equine multinodular pulmonary fibrosis. In this study, a multiplex real-time PCR (rtPCR) was designed to detect DNA of the glycoprotein H (EHV-2) and E11 gene (EHV-5). Analytical specificity was determined by testing DNA of other herpesviruses by SYBR Green rtPCR and melting curve analysis, as well as Sanger sequencing of positive field samples. Analytical sensitivity was assessed by standard curve generation of serial plasmid dilutions containing the respective target gene. Melting curves and BLAST analysis of the sequences indicated specific detection of the viruses. The lower limit of detection of the singleplex rtPCR was 40 and 29 DNA copies per reaction for EHV-2 and EHV-5, respectively. Comparison of the Ct values of a selection of positive field samples showed only minimal differences between the singleplex and the multiplex assay. The here described multiplex rtPCR protocol allows sensitive and specific detection of EHV-2 and EHV-5. It represents a convenient and rapid tool for future studies to investigate the clinical relevance of EHV-2 and EHV-5 in more detail.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Herpesvirus 1, Equid
/
Herpesviridae Infections
/
Herpesvirus 4, Equid
/
Herpesviridae
/
Horse Diseases
Type of study:
Diagnostic_studies
/
Guideline
Limits:
Animals
Language:
En
Journal:
J Virol Methods
Year:
2022
Document type:
Article
Affiliation country:
Switzerland