Development and validation of SYBR Green- and probe-based reverse-transcription real-time PCR assays for detection of the S and M segments of Schmallenberg virus.
J Vet Diagn Invest
; 32(5): 710-717, 2020 Sep.
Article
em En
| MEDLINE
| ID: mdl-32757829
ABSTRACT
Schmallenberg virus (SBV), discovered in Germany in 2011, causes congenital malformations in ruminants. Reverse-transcription real-time PCR (RT-rtPCR) assays based on various segments of SBV have been developed for molecular detection. We developed alternative RT-rtPCR assays for SBV detection to avoid earlier reported mutations and hypervariable regions of the S and M segments of the viral genome. For SYBR Green-based detection of the S segment, the R2 value and efficiency of the developed assay were 0.99 and 99%, respectively. For probe-based S segment detection, 2 assays were developed; the first had an R2 value of 0.99 and 102% efficiency, and the second had a R2 value of 0.98 and 86% efficiency. The probe-based M segment assay had an R2 value of 1.00 and 103% efficiency. Detection limits of the RT-rtPCR assays with new primer sets were 102 and 101 copies/µL for the S and M segments, respectively. Field samples from cattle and sheep were also used for primary validation of the developed assays. Our assays should be suitable for SBV detection in ruminants and for in vitro studies of various SBV strains.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Doenças dos Ovinos
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Doenças dos Bovinos
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Orthobunyavirus
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Infecções por Bunyaviridae
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Reação em Cadeia da Polimerase Via Transcriptase Reversa
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Reação em Cadeia da Polimerase em Tempo Real
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
Idioma:
En
Revista:
J Vet Diagn Invest
Ano de publicação:
2020
Tipo de documento:
Article