Detection and characterization of tobacco mild green mosaic virus (TMGMV) large type isolate from trailing petunia in France.

During 2003 and 2004, unusual viral symptoms were observed on Surfinia trailing petunias in protected cultivations of Southern France. Symptoms consisted in yellow mosaic and distortion of the leaves accompanied by vein necrosis in some samples. The flowers were deformed and showed light colour break of the petals. Electron microscope observation of negatively stained leaf-dip from symptomatic leaves showed straight rod-shaped virus particles of about 300 nm in length. Sap extracts reacted in double-immunodiffusion tests by forming weak precipitin bands with antisera against Tomato mosaic virus (ToMV) and Tobacco mild green mosaic virus (TMGMV). However, symptoms developed on host range after mechanical inoculation suggested that ToMV was not involved in the disease. By using specific primer pairs designed to amplify the coat protein (CP) genes of ToMV and TMGMV in reverse transcriptase-polymerase chain reaction (RT-PCR), expected amplicon was obtained only with TMGMV primer pair. The identity of the virus was also confirmed by using a specific TMGMV riboprobe in dot-blot hybridization assays of symptomatic leaf extracts. The nucleotide sequence of TMGMV CP of the isolate from trailing petunia, named TMGMV-Pt, was determined and compared with those available from EMBL. The percentage of nucleotide identity was 97-98% compared with those of other isolates. Further molecular and biological characterization revealed that TMGMV-Pt belonging to the large type group of TMGMV isolates. In fact, the 3' UTR region of TMGMV-Pt consisted of 360 nucleotides, comprising of a 147 base repeat, as reported only for TMGMV large type isolates. Moreover, symptoms development observed on a differentially host range, used to distinguish between large type and small type isolates, confirmed that TMGMV-Pt belonging to the large type group of isolates. Only one commercial variety of trailing petunia out of 12 tested remained symptomless after mechanical inoculation with TMGMV-Pt. This highlights the potential risk that TMGMV could represent to petunia cultivations. To our knowledge this is the first report of a natural infection by TMGMV in trailing petunia.

First Report of an Isolate of Pelargonium zonate spot virus in Commercial Glasshouse Tomato Crops in Southeastern France.

In summer 2000, symptoms similar to Pelargonium zonate spot virus (PZSV) were observed for the first time on tomato plants in southeastern France. The plants were from commercial glasshouse fresh-market crops. Symptoms observed were chlorotic mottling with bright yellow distinct rings on leaves and curved line patterns on stems. Fruit symptoms included chlorotic and necrotic spotting, marked concentric ring patterns, and distortions. Diagnosis was made from symptomatic leaves and fruits by mechanical inoculation on a set of host plants. Local chlorotic and necrotic lesions were observed on Chenopodium amaranticolor, C. quinoa, Cucumis sativus cv. Marketer, Cucumis melo cv. Vedrantais, Phaseolus vulgaris cv. Pinto, Vicia faba cv. D'Aguadulce, Vigna unguiculata cv. Black Eye, and systemic symptoms were observed on Capsicum annuum cvs. Yolo Wonder, Yolo Y, Florida VR2, and Criollo de Morelos 334, Datura stramonium, Lycopersicon esculentum cvs. Momor and Stevens, L. hirsutum (PI 134417 and PI 247087), Nicotiana benthamiana, N. clevelandii, N. tabacum cv. Xanthi nc, Ocimum basilicum cv. Latino, Petunia hybrida cv. Rose du ciel, and Physalis floridana. No reaction was observed on Pisum sativum cv. Douce Provence, Salvia splendens cv. Etna, or Zinnia elegans cv. Liliput. Symptoms on tomato of PZSV, Parietaria mottle virus (PMoV), and Tomato spotted wilt virus (TSWV) are similar, particularly those elicited in fruits. Therefore, the field samples were checked using double-antibody sandwich enzyme-linked immunosorbent assay against antisera of the type-strain of PZSV and tomato strain of PMoV and their homologous antigenes, which were supplied by D. Gallitelli and P. Roggero respectively, and our antiserum of TSWV. Electron microscopy of negatively stained preparations from leaves of tomato and D. stramonium showed that the sap contained very few paraspheric shaped particles, 26 to 29 nm in diameter. Three isolates collected from two different regions (Vaucluse and Bouches du Rhône) showed a very close serological relationship with the Italian type-strain of PZSV and tested negative against antisera of PMoV and TSWV. The French isolates were biologically different from the type-strain, but were similar to the Spanish strain of PZSV because they infected D. stramonium, N. benthamiana, O. basilicum, and V. unguiculata (2). Moreover, in transverse tissue sections, virions were not observed in the nucleus and tubular structures, unlike the Italian isolates, (1) but were present in the cytoplasm and particularly in the mesophyll cells. There are only a few records of the occurrence and distribution of PZSV in Mediterranean countries. References: (1) M. A Castellano and G. P Martelli. Phytopathol. Mediterr. 20:64, 1981. (2) M. Luis-Arteaga. Plant Dis. 84:807, 2000.

Occurrence of Chilli veinal mottle virus in Solanum aethiopicum in Tanzania.

African eggplant, or garden egg (Solanum aethiopicum) is an important vegetable in most sub-Saharan African countries. Since June 1997, viral symptoms, including mosaic, vein clearing, and stunting, have been observed on several crops of African eggplant cv. Tengeru White at a number of sites in the Arusha region of northern Tanzania. Field inspections revealed disease incidence ranging from 50 to 90%. During the same period, high populations of the green peach aphid Myzus persicae were observed in affected crops of African eggplant. These aphids were also found to reproduce in African eggplants. Flexuous, rodshaped virus-like particles, approximately 750 nm long and 12 nm wide, were found in electron microscope leaf dips from field samples of naturally affected African eggplants. The particle size suggested a species of Potyviridae. Thus, 20 field-infected samples of S. aethiopicum (randomly collected from four farms) were assayed in double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) for the presence of Potato virus Y (PVY) and Pepper veinal mottle virus (PVMV), known to infect tomato and other solanaceous crops in the region (2). However, all samples gave negative results. Further DAS-ELISA were performed with the same extracts from naturally infected plants of S. aethiopicum with antisera directed against Tobacco etch virus, Tobacco vein mottling virus, Pepper mottle virus, and Chilli veinal mottle virus (ChiVMV). All 20 samples were positive only for ChiVMV. ChiVMV, a single-stranded RNA virus transmitted in a nonpersistent manner by several aphid species, is one of the most important viruses of pepper in Asia (1). To confirm DAS-ELISA results, an isolate of ChiVMV from African eggplant was transmitted by mechanical inoculations, resulting in disease on tobacco (Nicotiana tobacco cv. Xanthi nc), pepper (Capsicum annuum cv. Yolo Wonder), tomato (Lycopersicon esculentum cv. Tengeru 97), and African eggplant (S. aethiopicum cv. Tengeru White). Extracts from the inoculated plants tested positive for the presence of ChiVMV in DAS-ELISA. This mechanically transmitted isolate did not infect melon (Cucumis melo), cucumber (C. sativus), or cowpea (Vigna unguiculata), which are nonhosts of ChiVMV. To our knowledge, this is the first report of the natural occurrence of ChiVMV in African eggplant. References: (1) S. K. Green et al. PETRIA 9:332, 1999. (2) R. Nono-Womdim et al. J. S. Afr. Soc. Hort. Sci. 6:41-44, 1996.

Evidence for two distinct subgroups of alfalfa mosaic virus (AMV) from france and italy and their relationships with other AMV strains Brief report.

The nucleotide sequence of the putative coat protein open reading frame of seven previously uncharacterized AMV strains from Italy and France was determined and aligned with comparable sequences of other AMV strains (425 L, 425 M, YSMV, S, VRU, 15/64 and Da). The data set of AMV sequences was used to determine phylogenetic relationships by both a stochastic (stationary Markov model) and a deterministic method (maximum-parsimony) of analysis. The topology of the trees obtained with the two methods was essentially the same showing that all AMV strains clustered in two monophyletic groups. Close clustering of Italian strains in subgroup I and of French strains in subgroup II seems to suggests the effect of geographic distinctiveness of evolutionary dynamics of these AMV strains. This separation did not correlate with differences in host range or symptoms (necrotic or non necrotic) induced in tomato but rather it reflected variations in the amino acid sequence of their CP, which might be related to structural properties of virus particles. A simple and rapid procedure based on the reverse transcriptase-polymerase chain reaction (RT-PCR) followed by ezymatic digestion (RFLP) was developed to identify and classify AMV isolates into the two subgroups. The method applied to a number of other AMV isolates from Italy and France supported their division in two distinct subgroups. This RT-PCR RFLP method may be useful way to investigate the dynamics of AMV populations in nature.

[Optimization of gold immunolabeling techniques on ultrathin slides obtained from samples fixed with glutaraldehyde and osmium tetroxide and enclosed in epoxy resin]. / Optimisation de la technique de l'immunomarquage à l'or sur sections ultrafines réalisées à partir d'échantillons fixés à la glutaraldéhyde et au tétroxyde d'osmium et inclus dans une époxy-résine.

An immunogold labeling technique was carried out on plants infected with CMV, fixed with glutaraldehyde and osmium tetroxide and embedded in araldite CY 212. The effect of the type of support-film used, the resin and manipulations of the grids during immunogold steps, were studied and are discussed. The antigenic activity of virus was restored by treating the sections with sodium metaperiodate. The very high non-specific reactions observed with the support-film or with the resin were eliminated by adding powdered skimmed milk or non-purified albumin into the buffers. Purified bovine serum albumin (grade V) or chicken albumin (grade III to V) were inefficient in reducing this non-specific background.

[Differential thermal behavior of various strains of cucumber mosaic virus. Hypothesis of a pleiotropic mechanism connecting various properties]. / Comportement thermique différentiel de certaines souches du virus de la mosaïque du concombre. Hypothèse d'un mécanisme pléiotropique reliant plusieurs propriétés

Pseudorecombinat viruses are constructed by mixing RNA coming from two CMV strains differentiated by their thermosensitivity. The pseudorecombinant shows thermal properties analogous to the parental strain diving RNA number 3. In order to explain the numerous properties coded by RNA 3 a pleiotropic effect is considered.