Detection and differentiation of low virulence and virulent Orthoavulavirus javaense using a molecular beacon with RT-LAMP.

Autor: Mears MC; Exotic and Emerging Avian Viral Disease Research Unit, Southeast Poultry Research Laboratories, US National Poultry Research Center, 934 College Station Road, Athens, GA, 30605, USA., Olivier TL; Exotic and Emerging Avian Viral Disease Research Unit, Southeast Poultry Research Laboratories, US National Poultry Research Center, 934 College Station Road, Athens, GA, 30605, USA., Williams-Coplin D; Exotic and Emerging Avian Viral Disease Research Unit, Southeast Poultry Research Laboratories, US National Poultry Research Center, 934 College Station Road, Athens, GA, 30605, USA., Espinoza E; Exotic and Emerging Avian Viral Disease Research Unit, Southeast Poultry Research Laboratories, US National Poultry Research Center, 934 College Station Road, Athens, GA, 30605, USA., Bakre A; Exotic and Emerging Avian Viral Disease Research Unit, Southeast Poultry Research Laboratories, US National Poultry Research Center, 934 College Station Road, Athens, GA, 30605, USA. Abhijeet.Bakre@usda.gov.
Jazyk: angličtina
Zdroj: Scientific reports [Sci Rep] 2024 Aug 05; Vol. 14 (1), pp. 18047. Date of Electronic Publication: 2024 Aug 05.
DOI: 10.1038/s41598-024-68816-7
Abstrakt: Newcastle disease (ND), an economically important disease in poultry, is caused by virulent strains of the genetically diverse Orthoavulavirus javaense (OAVJ). Laboratories rely on quantitative real-time reverse transcription PCR (qRT-PCR) to detect OAVJ and differentiate between OAVJ pathotypes. This study demonstrates that a fusion cleavage site based molecular beacon with reverse transcription loop mediated isothermal amplification (MB-RT-LAMP) assay can detect and differentiate OAVJ pathotypes in a single assay. Data show that the assay can rapidly identify diverse OAVJ genotypes with sensitivity only one log-fold lower than the current fusion qRT-PCR assay (10 4 copies), exhibits a high degree of specificity for OAVJ, and the molecular beacon can differentiate mesogenic/velogenic sequences from lentogenic sequences. Further, data show that a two-minute rapid lysis protocol preceding MB-RT-LAMP can detect and differentiate OAVJ RNA from both spiked samples and oropharyngeal swabs without the need for RNA isolation. As the MB-RT-LAMP assay can rapidly detect and discriminate between lentogenic and mesogenic/velogenic sequences of OAVJ within one assay, without the need for RNA isolation, and is adaptable to existing veterinary diagnostic laboratory workflow without additional equipment, this assay could be a rapid primary screening tool before qRT-PCR based validation in resource limited settings.
(© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)
Databáze: MEDLINE
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