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Bead-based suspension array systems enable simultaneous fluorescence-based identification of multiple nucleic acid targets in a single reaction. This study describes the development of a novel approach to plant virus and vector diagnostics, a multiplexed 7-plex array that comprises a hierarchical set of assays for the simultaneous detection of begomoviruses and Bemisia tabaci, from both plant and whitefly samples. The multiplexed array incorporates genus, species and strain-specific assays, offering a unique approach for identifying both known and unknown viruses and B. tabaci species. When tested against a large panel of sequence-characterized begomovirus and whitefly samples, the array was shown to be 100% specific to the homologous target. Additionally, the multiplexed array was highly sensitive, efficiently and concurrently determining both virus and whitefly identity from single viruliferous whitefly samples. The detection limit for one assay within the multiplexed array that specifically detects Tomato yellow leaf curl virus-Israel (TYLCV-IL) was quantified as 200fg of TYLCV-IL DNA, directly equivalent to that of TYLCV-specific qPCR. Highly reproducible results were obtained over multiple tests. The flexible multiplexed array described in this study has great potential for use in plant quarantine, biosecurity and disease management programs worldwide., (Copyright © 2014 Elsevier B.V. All rights reserved.)

A semi-automated, immunomagnetic capture-reverse transcription PCR (IMC-RT-PCR) assay for the detection of three pineapple-infecting ampeloviruses, Pineapple mealybug wilt-associated virus-1, -2 and -3, is described. The assay was equivalent in sensitivity but more rapid than conventional immunocapture RT-PCR. The assay can be used either as a one- or two-step RT-PCR and allows detection of the viruses separately or together in a triplex assay from fresh, frozen or freeze-dried pineapple leaf tissue. This IMC-RT-PCR assay could be used for high throughput screening of pineapple planting propagules and could easily be modified for the detection of other RNA viruses in a range of plant species, provided suitable antibodies are available.