A survey was conducted over several years in Italy and the Balkans in order to gain an understanding of the relationship between the Flavescence dorée (FD) phytoplasma isolates found in clematis and grapevine. A total of 399 clematis and 107 grapevine samples were analyzed. The results showed that 36% of the Clematis vitalba plant samples were infected by phytoplasmas which, in grapevine, are associated with FD, a quarantine disease in Europe. Infected clematis plants were also found in areas where FD phytoplasma had never previously been reported to infect grapevine, such as Macedonia, Croatia and some areas of Italy and Serbia. Molecular data from three phytoplasma genomic fragments showed the presence of different FD phytoplasma isolates, all belonging to the 16SrV-C subgroup, including the Italian FD-C isolate, the isolate found in Serbia, an isolate similar to the French FD2000 and a new isolate typical of central Italy. A few clematis plants were infected with single nucleotide polymorphism, insertion or deletion mutants of the FD-C isolate. Of all the potential Hemipteran vector species surveyed in Italy and Serbia, only 18 of 527 Dictyophara europaea individuals tested proved to be infected with the FD phytoplasma. Preliminary transmission experiments showed that this species is able to transmit the FD phytoplasma from clematis to grapevine. The presence of FD-infected clematis and of D. europaea could, therefore, constitute a risk for FD epidemics in the European viticultural regions.

An epidemiological study on a grapevine yellows disease called bois noir was carried out for 3 years in the Rhone valley (France). This yellows is caused by a stolbur type phytoplasma. Vectors and alternative host plants were searched, and the inoculation period was determined. Detection of stolbur phytoplasma in insects and plants was obtained using primers STOLl 1 f2,rl. In addition, a nested polymerase chain reaction (PCR) was used with primers P1/P7 and fU5/rU3 for detection in stolburinfected plants with low titre. Several thousand insects were captured and species of Hemiptera were listed. Fourteen wild or reared Hemiptera species were used in transmission trials. Thirty-four wild species were monitored for phytoplasma DNA by PCR. A planthopper.Hvalesthes obsoletus Sign., tested positive for stolbur at a level of 28% (98,/343) in 1995(98,/343) in and 38% (205,529) in 1996(98,/343) in . In 1995 leafhoppers, Mocydia crocea (1 78) and Euscelis tineolatus (2/309), were infected at a much lower ratio. Successful experimental transmission to grapevine, periwinkle and thorn apple was only obtained with H. obsoletus. Among wild plant species, hoary cress (Cardaria draba L.), bindweed (Convolvulus arvensis L.), sweet cherry {Prunus sp,), plum (Prunus domestica L), lilac (Syringa vulgaris L.), fig tree [Ficus carica L.) and elm {Ulmus sp.) were shown to be stolbur infected and hoary cress was identified as a new host plant for H. obsoletus in France. The role of some fallow lands close to vineyards as sites where the inoculum of stolbur phytoplasma was maintained in insect vector and reservoir plants, is discussed. The natural inoculation period to grapevine was shown to extend from June to August, corresponding to the adult activity of H. obsoletus. Together with the close relationships of the phytoplasmas involved in vergilbungskrankheit, a German grapevine disease, and bois noir. The results of this study suggest that the two yellows are very close. Zusammenfassung Bedeutung von Hyalesthes obsoletus (Hemiptera: Cixiidae) fur das Auftreten der Bois-noir-Krankheit der Weinrebe in Frankreich Im franzosischen Rhonetal wurde eine 3-jahrige epidemiologische Studie iiber die Bois-noir-Vergilbungskrankheit der Weinrebe durchgefuhrt. Diese Krankheit wird von einem Phytoplasma des Stolbur-Typs ausgelost. Es wurde nach Vektoren und alternativen Wirtspflanzen gesucht, und die Inokulationsperiode wurde bestimmt. Die Detektion von Stolbur-Phytoplasmen in insekten und Pftanzen erfolgte mit Hilfe der Primer STOL 1! f2/rl. Zudem wurde eine genestete PCR mit den Primem P1 /P7 und fU5 rU3 zur Detektion in stolburinfizierten Pflanzen mit niedrigem Titer durchgefuhrt. Mehrere tausend Insekten wurden gefangen, und die Hemiptera-Arten wurden aufgelistet. 14 wilde oder geziichtete HemipteraArten wurden in Ubertragungsversuchen verwendet, und 34 Wildarten wurden mittels PCR auf Phytoplasma-DNA untersucht. Der Fulgoride Hvalesthes obsoletus Sign, war 1995 mit 28% (98/343) und 1996 nut 38% (205/529) der getesteten Tiere stolburpositiv. ...

The syndrome “basses richesses” (SBR) is a disease of sugar beet in eastern France associated with two phloem-restricted, nonculturable plant pathogens: a stolbur phytoplasma and a γ-3 proteobacterium, here called SBR bacterium. Three planthopper (Hemiptera: Cixiidae) species were found to live near and within sugar beet fields in eastern France: Cixius wagneri, Hyalesthes obsoletus, and Pentastiridius leporinus. The role of these planthoppers in spreading the two pathogens to sugar beet was studied. Based on its abundance and high frequency of infection with the SBR bacterium, P. leporinus was considered to be the economic vector of SBR disease. C. wagneri, the primary vector of ‘Candidatus Phlomobacter fragariae’ to strawberry in western France, also was found to be infected by the SBR bacterium and to transmit the pathogen to sugar beet. Neither C. wagneri nor P. leporinus were infected by stolbur phytoplasma. Populations of H. obsoletus living on bindweed (Convolvulus arvensis) and nettle (Urtica dioica) collected near sugar beet fields did not carry the SBR bacterium, but were highly infected with two restriction fragment length polymorphism-differentiable stolbur phytoplasma isolates. In transmission assays, only the bindweed phytoplasma isolate was transmissible to and pathogenic on sugar beet. When compared with controls, the bindweed stolbur phytoplasma and SBR bacterium similarly reduced the biomass of sugar beet plants, but the phytoplasma caused greater reductions in taproot biomass and sugar content than the SBR bacterium.

A polymerase chain reaction (PCR)-based method was developed for the detection of phytoplasma in insect feeding medium (sucrose). A correlation was established between the transmissibility of Flavescence dorée phytoplasma in the experimental leafhopper vector Euscelidius variegatus and its detection by PCR in the insect feeding medium. However, phytoplasma were detected in the insects' bodies 3 weeks before they began to transmit. Hence, PCR assays of the sucrose medium reflected phytoplasma vectoring ability probably by detecting it in the insect saliva, whereas detection of phytoplasma in the insect's body did not identify it as a vector. The assay was applied to two field-collected leafhoppers suspected of being phytoplasma vectors in Israel (Orosius albicinctus and Anaceratagallia laevis). The presence of phytoplasma in the body of specimens of the latter species was assayed by PCR in 1999. Phytoplasmas were detected in insects' bodies throughout the year, with no specific seasonal pattern. In the saliva, however, no phytoplasma could be detected in the autumn. This seasonal pattern supported the validity of the feeding-medium tests and their correlation to the insect's ability to transmit phytoplasma. Transmission assays indicated, to our knowledge for the first time, that O. albicinctus and A. laevis are vectors of phytoplasma in Israel. A simple PCR-based assay is thus provided, circumventing the need for tedious biological assays and enabling epidemiological studies of phytoplasma transmissibility on a large scale.