Chromogenic detection of yam mosaic virus by closed-tube reverse transcription loop-mediated isothermal amplification (CT-RT-LAMP)

Abstract

A closed-tube reverse transcription loop-mediated isothermal amplification (CT-RT-LAMP) assay was developed for the detection of yam mosaic virus (YMV, genus Potyvirus) infecting yam (Dioscorea spp.). The assay uses a set of six oligonucleotide primers targeting the YMV coat protein region, and the amplification products in YMV-positive samples are visualized by chromogenic detection with SYBR Green I dye. The CT-RT-LAMP assay detected YMV in leaf and tuber tissues of infected plants. The assay is 100 times more sensitive in detecting YMV than standard RT-PCR, while maintaining the same specificity.Electronic supplementary materialThe online version of this article (10.1007/s00705-018-3706-0) contains supplementary material, which is available to authorized users.

Assessment of Yam mild mosaic virus coat protein gene sequence diversity reveals the prevalence of cosmopolitan and African group of isolates in Ghana and Nigeria

“…The quarantine monitoring of germplasm needs to be strengthened to mitigate this risk, coupled with improvements to the production and distribution of clean planting for seed and ware yam production. The sequences generated in the study will facilitate the development of field-based diagnostic tools necessary for virus indexing and seed certification as it was achieved with YMV [ [25] , [26] , [27] ].…”

Tissue culture and next-generation sequencing: A combined approach for detecting yam (Dioscorea spp.) viruses

“…The use of virus-free, clonally propagated planting materials is the most effective method to control the spread of viruses infecting yam [3]. Molecular diagnostic tools such as RT-RPA [51] and CP-RT-LAMP [53] have been developed for routine detection of one such virus, YMV, which is endemic in the West African ‘yam belt’ [1]. These and similar tools need to be adopted and used to verify the infection status of planting material in West Africa, where efforts to boost production of virus-free seed yam and establish sustainable seed systems are ongoing [3,53].…”

“…Molecular diagnostic tools such as RT-RPA [51] and CP-RT-LAMP [53] have been developed for routine detection of one such virus, YMV, which is endemic in the West African ‘yam belt’ [1]. These and similar tools need to be adopted and used to verify the infection status of planting material in West Africa, where efforts to boost production of virus-free seed yam and establish sustainable seed systems are ongoing [3,53]. Research into modern yam seed production methods, including vine cutting, tissue culture, aeroponics, and TIBs, highlights the importance of an integrated multiplication scheme that combines two or more methods of seed yam production [39].…”

“…Several serological and nucleic acid-based methods, such as enzyme-linked immunosorbent assay (ELISA), immunocapture reverse transcription-PCR (IC-RT-PCR), RT-PCR, reverse-transcription recombinase polymerase amplification (RT-RPA), closed-tube reverse transcription loop-mediated isothermal (CT-RT-LAMP), and rolling circle amplification (RCA), have been used in indexing known yam viruses and also to characterise new yam potyviruses and badnaviruses [[48], [49], [50], [51], [52], [53]]. Next-generation sequencing (NGS) methods are increasingly being employed in the discovery and sequencing of new plant viral genomes [54,55].…”

A quick and sensitive diagnostic tool for detection of Maize streak virus

“…Although the real-time detection used in our study avoided any risk of contamination associated with post-amplification processes there are several low-cost portable fluorescence-reading instruments (e.g. Genie II (Optigene Ltd., West Sussex, UK), Bioranger (Diagenetix, Inc., Honolulu, HI, USA) and Genelyzer FIII (Canon Medical Systems, Tochigi, Japan)) available in the market that can be used to measure the fluorescence signal and thereby reducing the cost of the assay 18,30,31 . These simple battery-powered instruments offer a major advantage of the LAMP assay in places experiencing repeated power failures as reactions could proceed even without electricity.…”