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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.)

Three monocot-infecting mastreviruses from Australia, all found primarily in pasture and naturalised grasses, have been characterised at the molecular level. Here, we present the full genome sequence of a fourth, Paspalum striate mosaic virus (PSMV), isolated from Paspalum dilatatum from south-east Queensland. The genome was 2816 nt long and had an organisation typical of other monocot-infecting mastreviruses. Its nearest relative is Bromus cartharticus striate mosaic virus (BCSMV), with which it shares an overall genome identity of 75%. Phylogenetic analysis of the complete genome and each of the putative viral proteins places PSMV in a group with the other three Australian striate mosaic viruses. PSMV, BCSMV and Digitaria didactyla striate mosaic virus all contain a similar, small recombinant sequence in the small intergenic region.

In 1999, banana streak disease outbreaks occurred at two locations in Australia in new banana hybrids that were being screened for fusarium wilt resistance. Two different badnaviruses, banana streak GF virus and a newly discovered virus called banana streak IM virus (BSIMV), were detected in these plants. The complete nucleotide sequence of the BSIMV genome was determined and comprised 7768 nt. Three open reading frames were detected, the first beginning with a non-conventional start codon (CUG). A 55-nt repetition in the putative pregenomic RNA promoter was also identified. Phylogenetic analysis suggests that BSIMV is most closely related to banana streak VN virus.

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.