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The influence of inoculation techniques on induction of post-infection ethylene production in susceptible and resistant genotypes of bean (Phaseolus vulgaris) inoculated with the kappa race of Colletotrichum lindemuthianum was studied. Three inoculation methods (brushing, dipping and spraying) of excised leaves were compared. The brushing technique caused both the highest ethylene production at 120 h after inoculation, and better symptom development than dipping and spraying. It was, therefore, adopted to determine the post-infection ethylene production in four inoculated bean genotypes (GLP406 and Kaboon - resistant, GLP636 - moderately resistant; and MDRK - susceptible). Ethylene production increased slightly 24 h after inoculation in all four genotypes, followed by a rapid decline after 48 h. Ethylene production remained low until 120 h after which a sharp rise was observed in genotype MDRK. Increased production, though to a lesser extent, was observed in GLP636. This trend was repeatable and thus may offer an additional physiological marker to bean breeders to screen for resistance to C. lindemuthianum.

The nematode species Longidorus attenuatus, L. elongatus, L. macrosoma and Paralongidorus maximus are economically important pests to the viticulture industry due to their ability to vector two nepoviruses (Raspberry Ringspot Virus and Tomato Black Ring Virus) to grapevines. In Germany, these species occur in vineyard soil with other non-vector but morphologically similar longidorid species, L. helveticus, L.␣profundorum and L. sturhani. Species-specific primers were designed from ribosomal DNA for all seven species to facilitate taxonomic identification for non-specialists. Primers were assessed for their reliability by screening, where possible, a number of populations of each species. Furthermore, their selectivity and sensitivity were determined when challenged with closely related longidorid species and general nematode communities typical of vineyard soil. A multiplex approach using a common forward primer combined with species-specific reverse primers enabled three target nematode species to be detected in the same PCR reaction. All primers were highly specific, detecting all nematode developmental forms from disparate populations and were sufficiently sensitive to detect a single target nematode within a whole nematode community typical of a vineyard soil comprising of a range of non-target species. Given their specificity, sensitivity and reliability, these diagnostic primers should be of great benefit to both phytosanitary/quarantine services related to the viticulture industry and also as a decision management tool for growers. [ABSTRACT FROM AUTHOR]

Xiphinema diversicaudatum and X. index are vector nematode species of economic importance in viticulture regions as they can transmit Arabis Mosaic, Grapevine Fanleaf and Strawberry Latent Ringspot viruses to grapevine. Wang et al. (2003) designed species-specific diagnostic primers from ribosomal genes for both these vector species as well as a vector and a non-vector species X. italiae and X. vuittenezi, respectively. Our study aimed to confirm the specificity and determine the sensitivity and reliability of the primers for the two vector species, X. diversicaudatum and X. index when challenged with closely related longidorid species and general nematode communities typical of vineyard soil. With one exception, no PCR product was observed when the primers were tested against six Longidorus, one Paralongidorus and one Xiphinema non-target species. Occasionally (three out of eight replicate PCR reactions) a weak PCR product was noted when primers for X. index were tested with L. elongatus. Furthermore, when challenged with a range of non-target nematode species comprising the nematode community typical of viticulture soil, no PCR product was amplified. An experimental dilution series of extracted DNA rigorously demonstrated that DNA from an equivalent single specimen of the target virus-vector species, X. diversicaudatum and/or X. index, could be detected amongst 1000 equivalent non-target X. vuittenezi. Also, extracted DNA from an equivalent single target specimen was detected when added to DNA extracted from the overall soil nematode community. The primers were assessed further by using serial mixtures of actual nematodes rather than extracted DNA to simulate field soil. Using this method, a single target nematode could be detected amongst 200 non-target specimens. Given their specificity, sensitivity and reliability, it appears that these diagnostic primers will be of great benefit to phytosanitary/quarantine services related to the viticulture industry. [ABSTRACT FROM AUTHOR]