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.
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 nontarget 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.
The ectoparasitic nematode Xiphinema index transmits grapevine fanleaf virus (GFLV) during feeding on grapevine roots, causing fanleaf degeneration in the plant. Hence, resistance breeding is a key to develop novel rootstocks to overcome such threats. In the past years, various grapevine species were screened, and a few candidates with partial resistance were identified. Yet, they were hardly sufficient for viticulture due to many agronomical defects. To develop reliably resistant rootstocks applicable in viticulture, multiple Vitis spp. genotypes were analyzed using root inoculation with nematodes in glass vials as an early and easy evaluation test. Resistance levels were evaluated 35 days after inoculation based on nematode reproduction factors focusing on juveniles and eggs. Infection of grapevines with GFLV was analyzed after inoculation with viruliferous X. index. With this fast screening system, putative candidates with resistances against X. index have been identified for future breeding programs. Particularly, genotypes with the genetic background of V. aestivalis and V. labrusca were found to be nematode resistant.
Zusammenfassung Grapevine virus A (GVA) wird als der Erreger von Kober stem grooving, einer zu dem Komplex der Holzrunzeligkeit (Rugose Wood) gehörenden Krankheit, angesehen. Das Virus wird oft in blattrollkranken Reben nachgewiesen, ist aber nicht unmittelbar mit dieser Krankheit gekoppelt. GVA ist vermutlich weltweit verbreitet, es sind bisher aber kaum Informationen über die Verbreitung von GVA in Rheinland-Pfalz bekannt. Der Nachweis von GVA erfolgte im Indexing-Verfahren mit Kober 5BB als Indikator und mittels ELISA-Test. Bei der Holzveredlung mit infizierten Reben konnten frühestens nach zwei Jahren, an grünveredelten Pfropfreben erst fünf Jahre nach der Veredlung erste schwache Symptome von Kober stem grooving beobachtet werden. Die serologischen Untersuchungen ergaben, dass der GVA-Antigentiter am höchsten in Blattstielen von ausgereiften Blättern oder Phloemschabsel von verholzten Trieben war. Bei 46,9% der 209 untersuchten Reben wurde eine GVA-Infektion festgestellt. Davon wiesen 87,8% eine Mischinfektion mit GLRaV-1 auf, weitere 5,1% waren neben GVA mit GLRaV-3 infiziert. Nur bei 7,1% der insgesamt GVApositiv getesteten Reben konnte außer GVA keine weiteren Ampelo-oder Closteroviren nachgewiesen werden. Keine der positiv auf GVA getesteten Pflanzen fiel durch Symptome der Holzrunzeligkeit auf. Aus diesem Grund wird die Bedeutung einer GVA-Infektion in Deutschland als gering angesehen.U. Ipach (u) · L. Kling Dienstleistungszentrum Ländlicher Raum -Rheinpfalz,
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