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Panel of real-time PCRs for the multiplexed detection of two tomato-infecting begomoviruses and their cognate whitefly vector species

van Brunschot, S. L. and Gambley, C. F. and De Barro, P. J. and Grams, R. and Thomas, J. E. and Henderson, J. and Drenth, A. and Geering, A. D. W. (2013) Panel of real-time PCRs for the multiplexed detection of two tomato-infecting begomoviruses and their cognate whitefly vector species. Plant Pathology, 62 (5). pp. 1132-1146. ISSN 0032-0862

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Article Link(s): http://dx.doi.org/10.1111/ppa.12033

Abstract

A new approach for the simultaneous identification of the viruses and vectors responsible for tomato yellow leaf curl disease (TYLCD) epidemics is presented. A panel of quantitative multiplexed real-time PCR assays was developed for the sensitive and reliable detection of Tomato yellow leaf curl virus-Israel (TYLCV-IL), Tomato leaf curl virus (ToLCV), Bemisia tabaci Middle East Asia Minor 1 species (MEAM1, B biotype) and B.tabaci Mediterranean species (MED, Q biotype) from either plant or whitefly samples. For quality-assurance purposes, two internal control assays were included in the assay panel for the co-amplification of solanaceous plant DNA or B.tabaci DNA. All assays were shown to be specific and reproducible. The multiplexed assays were able to reliably detect as few as 10 plasmid copies of TYLCV-IL, 100 plasmid copies of ToLCV, 500fg B.tabaci MEAM1 and 300fg B.tabaci MED DNA. Evaluated methods for routine testing of field-collected whiteflies are presented, including protocols for processing B.tabaci captured on yellow sticky traps and for bulking of multiple B.tabaci individuals prior to DNA extraction. This work assembles all of the essential features of a validated and quality-assured diagnostic method for the identification and discrimination of tomato-infecting begomovirus and B.tabaci vector species in Australia. This flexible panel of assays will facilitate improved quarantine, biosecurity and disease-management programmes both in Australia and worldwide.

Item Type:Article
Business groups:Horticulture and Forestry Science
Additional Information:van Brunschot, S. L. Gambley, C. F. De Barro, P. J. Grams, R. Thomas, J. E. Henderson, J. Drenth, A. Geering, A. D. W. Cooperative Research Centre for National Plant Biosecurity; Australian Government The authors would like to acknowledge the support of the Cooperative Research Centre for National Plant Biosecurity, established and supported under the Australian Government's Cooperative Research Centres Program. We thank the following researchers for their kind provision of samples: Z. Hall, B. Conde, C. Pearce, P. Campbell, E. Moriones, P. Stephens, P. Chiemsombat, S. K. Green, K. Zanic, M. Cahill, A. Hanafi, M. Muniz, S. Abd-Rab and A. Hulthen. We thank D. Pagendam (CSIRO Mathematics, Informatics and Statistics) for statistical advice, and the anonymous reviewers for valuable comments and suggestions. Wiley-blackwell Hoboken
Keywords:Bemisia tabaci diagnostics internal control multiplex Tomato leaf curl virus Tomato yellow leaf curl virus bemisia-tabaci hemiptera leaf-curl-virus polymerase-chain-reaction transmitted geminiviruses gennadius hemiptera mediterranean basin host-range q biotypes b-biotype aleyrodidae
Subjects:Plant pests and diseases > Pest control and treatment of diseases. Plant protection
Science > Statistics
Science > Invasive Species > Plants
Plant culture > Food crops
Deposited On:28 Aug 2014 03:49
Last Modified:28 Aug 2014 03:49

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