Sequence analyses of S10-spcoperon among 16SrV group phytoplasmas: phylogenetic relationships and identification of discriminating single nucleotide polymorphisms

Phytoplasmas are cell-wall less, uncultivable bacteria, belonging to the class Mollicutes. They parasitise both plant hosts and insect vectors, but knowledge of their biological and molecular properties is limited. In particular, analyses of 16S rDNA nucleotide sequences revealed that 16SrV group phytoplasma strains identified in grapevine plants are closely related, but associated with distinct ecologies. In this work, we investigated the evolutionary relationships of eight representative 16SrV phytoplasma strains by sequencing 15 ribosomal protein (rp) genes positioned within the S10-spc operon. Phylogenetic analyses of concatenated nucleotide and amino acid sequences underscored that 16S…

Le syndrome des basses richesses de la betterave sucrière en Bourgogne, un modèle de maladies infectieuses émergentes - Bilan des recherches 1997-2007

National audience; Une maladie inconnue de la betterave sucrière est apparue en Côte d'Or au début des années 1990 et a gagné le reste du bassin sucrier du Centre-est (Bourgogne et Jura). Les programmes d'étude financés par la filière betterave-sucre et Conseil régional de Bourgogne de 1997 à 2007, ont permis d'en élucider entièrement l'étiologie et la transmission, de développer des méthodes de diagnostic moléculaire, et de parvenir à une compréhension globale de son épidémiologie. Les méthodes de lutte directes et agronomiques préconisées ont montré leur efficacité pour limiter les pertes de rendement en sucre associées à cette maladie. Par ailleurs, la biologie de bactéries pathogènes ha…

Phylogenetic relationships among Flavescence doree strains and related phytoplasmas determined by heteroduplex mobility assay and sequence of ribosomal and Nonribosomal DNA

Heteroduplex mobility assay (HMA) and DNA sequencing were performed on Flavescence doree (FD) phytoplasma strains and related phytoplasmas belonging to the elm yellows group. Part of the ribosomal RNA gene operon and a nonribosomal DNA region were utilized for phylogenetic analyses. Two FD strains, FD92 and FD-D, detected in France and Italy, respectively, were identical in both DNA fragments, confirming previous results. Other FD strains were all very similar and most closely resembled ALY, an Italian alder phytoplasma. Phytoplasmas associated with German Palatinate grapevine yellows were shown to form a distinct subcluster, also different from the elm yellows phytoplasma subcluster. Strai…

Vector Transmission of a Plant-Pathogenic Bacterium in the Arsenophonus Clade Sharing Ecological Traits with Facultative Insect Endosymbionts

Bressan, A., Semetey, O., Arneodo, J., Lherminier, J., and BoudonPadieu, E. 2009. Vector transmission of a plant-pathogenic bacterium in the Arsenophonus clade sharing ecological traits with facultative insect endosymbionts. Phytopathology 99:1289-1296. The planthopper Pentastiridius leporinus (Hemiptera: Cixiidae) is the major vector of a nonculturable plant-pathogenic γ-3 proteobacterium associated with a disease of sugar beet called syndrome “basses richesses” (SBR). The bacterium, here called SBR bacterium, belongs to the Arsenophonous clade, which includes mostly insect-associated facultative symbionts. Assays using field-collected planthopper nymphs and adults were carried out to inve…

Vector activity of Hyalesthes obsoletus living on nettles and transmitting a stolbur phytoplasma to grapevines: a case study

International audience; We report a case study on the vector activity of a Hyalesthes obsoletus (Hemiptera: Cixiidae) population living on nettle plants (Urtica dioica) and transmitting a stolbur phytoplasma (Sp) to grapevines (Vitis vinifera). The research was conducted in a site that included a vineyard bordered with a large fallow area where nettles were the predominant plant species together with sparse old grapevines. Nettles hosted a high population of H. obsoletus. By using transparent sticky traps to sample adults, we observed that the daily flight activity of males and females to grapevines in the fallow was unimodal peaking between 15 and 21 h in the day. Adults were unable of gre…

Characterization and evolution of two bacteriome-inhabiting symbionts in cixiid planthoppers (Hemiptera: Fulgoromorpha: Pentastirini).

International audience; Like other plant sap-sucking insects, planthoppers within the family Cixiidae (Insecta: Hemiptera: Fulgoromorpha) host a diversified microbiota. We report the identification and first molecular characterization of symbiotic bacteria in cixiid planthoppers (tribe: Pentastirini). Using universal eubacterial primers we first screened the eubacterial 16S rRNA sequences in Pentastiridius leporinus (Linnaeus) with PCR amplification, cloning, and restriction fragment analysis. We identified three main 16S rRNA sequences that corresponded to a Wolbachia bacterium, a plant pathogenic bacterium, and a novel gammaproteobacterial symbiont. A fourth bacterial species affiliated w…

Development of a specific assay using RISA for detection of the bacterial agent of 'basses richesses' syndrome of sugar beet and confirmation of a Pentastiridius sp. (Fulgoromopha, Cixiidae) as the economic vector

International audience; A technique for the specific diagnosis in insects of SBRp (the γ-3 proteobacterium associated with the syndrome ‘basses richesses’ (SBR) of sugar beet crops in eastern France), using the RISA (rDNA intergenic spacer analysis) technique, was developed. PCR using the Alb1/Oliv1 primer pair specifically amplified a 16S-ITS region of SBRp and produced a characteristic DNA fingerprint. This PCR assay did not detect other closely related organisms, including the Arsenophonus endosymbiont of Diaphorina citri, the secondary endosymbiont of Glycaspis brimblecombei, or ‘Candidatus Phlomobacter fragariae’, a related phytopathogenic γ-3 proteobacterium. Six different ribosomal o…

Multilocus sequence typing confirms the close genetic inter-relatedness between three distinct flavescence doree phytoplasma strain clusters and group 16SrV phytoplasmas infecting grapevine and alder in Europe

ABSTRACT Vineyards of southern France and northern Italy are affected by the flavescence dorée (FD) phytoplasma, a quarantine pathogen transmitted by the leafhopper of Nearctic origin Scaphoideus titanus . To better trace propagation of FD strains and identify possible passage between the vineyard and wild plant compartments, molecular typing of phytoplasma strains was applied. The sequences of the two genetic loci map and uvrB - degV , along with the sequence of the secY gene, were determined among a collection of FD and FD-related phytoplasmas infecting grapevine, alder, elm, blackberry, and Spanish broom in Europe. Sequence comparisons and phylogenetic analyses consistently indicated the…

A chromosome map of the Flavescence dorée phytoplasma

International audience; The Flavescence dorée phytoplasma (FD-P), a non-cultivable, plant-pathogenic bacterium of the class Mollicutes, is the causal agent of a quarantine disease affecting vineyards of southern Europe, mainly in southern France and northern Italy. To investigate FD-P diversity and phytoplasma genetic determinants governing the FD-P life cycle, a genome project has been initiated. A physical map of the chromosome of FD-P strain FD92, purified from infected broad beans, was constructed by performing restriction digests of the chromosome and resolving the fragments by PFGE. Single and double digestions of the chromosome with the enzymes SalI, BssHII, MluI and EagI were perfor…

Identification and biological traits of a planthopper from the genus Pentastiridius (Hemiptera: Cixiidae) adapted to an annual cropping rotation

International audience; Cixiid planthoppers have been shown to vector phloem-limited prokaryotes associated with plant diseases world-wide. In eastern France, an emerging disease of sugar beet called syndrome basses richesses has been associated with phloem-restricted bacteria transmitted by a cixiid planthopper within the genus Pentastiridius. Early investigation suggested the species being Pentastiridius beieri. On the basis of a morphological and phylogenetic study we report the identification of the planthopper as Pentastiridius leporinus. Furthermore we report some biological traits of the species, which shows a surprising ecological adaptation to an annual cropping rotation sugar beet…

Une relation trophique originale : la vection entomophile d'agents pathogènes

Bgpi : équipe 6 chapitre 34; Une relation trophique originale : la vection entomophile d'agents pathogènes

Insect Vectors (Hemiptera: Cixiidae) and Pathogens Associated with the Disease Syndrome “Basses Richesses” of Sugar Beet in France

International audience; 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 prim…

Abstracts of papers presented at the 9th meeting of the international council for the study of viruses and virus diseases of the grapevine

Monitoring of grapevine leafroll-associated virus 1 (GLRaV-1) dispersion by the mealybug phenaccocus aceris

International audience; A four-year monitoring in a vineyard affected with leafroll and first transmission experiments were performed in order to understand leafroll dispersion by the mealybug Phenacoccus aceris. Our data showed for the first time that GLRaV-1 progression was due to the colonisation of the monitored vineyard by infective P. aceris spreading from bordering infected vineyards. Additionally, GLRaV-3 and Grapevine virus A (GVA) were transmitted by first instar nymphs of P. aceris.

Spatio-temporal pattern of Pentastiridius leporinus migration in an ephemeral cropping system

International audience; * 1 Cixiid planthoppers (Hemiptera: Fulgoromorpha: Cixiidae) are considered to be important economic pests because of their ability to transmit phloem-restricted prokaryotes causing emerging plant diseases worldwide. However, little information is available on the biology and ecology of such species. This is the case for Pentastiridius leporinus (Linnaeus), a cixiid planthopper reported to live on common reed across Countries of Central and Northern Europe. However, in the east of France, the same planthopper species appears to complete its life cycle in the sugar beet-wheat cropping system and has been repeatedly shown to transmit prokaryotic plant pathogens that ar…

Association with the Syndrome "Basses Richesses" of Sugar Beet of a Phytoplasma and a Bacterium-Like Organism Transmitted by a Pentastiridius sp.

The syndrome “basses richesses” of sugar beet (SBR) was first observed in 1991 in Burgundy, France. A cixiid planthopper, Pentastiridius beieri, has been proved to be involved in the transmission to sugar beet of a stolbur phytoplasma, which could be detected in some affected plants. In 2000, periwinkle and sugar beet exposed to field-collected cixiids developed symptoms similar to SBR on sugar beet. Use of 4′-6-diamidino-2-phenylindole (DAPI) staining and transmission electron microscopy confirmed the presence of phytoplasma in some of the plants, which were also positive for this pathogen in a polymerase chain reaction (PCR) analysis. A phloem-restricted gram-negative bacteria was seen i…