[Potato virus Y (PVY): from its discovery to the latest outbreaks]. / Le virus Y de la pomme de terre (PVY) : de la première description aux derniers épisodes d'émergence.

Potato virus Y (PVY, family Potyviridae, genus Potyvirus) is one of the most economically important viruses infecting potato. This plant virus is transmitted by aphids and is present in all potato growing areas across the world. Thanks to the steady set-up of biological, serological and molecular detection/characterization tools, PVY potato strain isolates have been classified in groups (PVYN, PVYO, PVYC, PVYZ and PVYE) or as sub-groups (PVYNTN and PVYN-W). Epidemiological data available for PVY show the recent modification of PVY group and sub-group proportions in PVY populations. This modification has led to the current prevalence of necrotic recombinant PVY isolates. In order to identify factors involved in this evolution of PVY populations, characterization of i) the molecular determinants of necrotic properties, ii) the impact of the increase of PVY virulence and aggressiveness on fitness, iii) the role of recombination in PVY evolution and iv) the genetic variability of viral populations have been carried out. The main results of this research have been combined with data already published to write the present review.

Discussion paper: The naming of Potato virus Y strains infecting potato.

Potato virus Y (PVY) strain groups are based on host response and resistance gene interactions. The strain groups PVY(O), PVY(C) and PVY(N) are well established for the isolates infecting potato in the field. A switch in the emphasis from host response to nucleotide sequence differences in the virus genomes, detection of isolates recombining sequences of different strains, and the need to recognize isolates that cause necrotic symptoms in potato tubers have led to the assignment of new acronyms, especially to isolates of the PVY(N) strain group. This discussion paper proposes that any newly found isolates should be described within the context of the original strain groups based on the original methods of distinguishing strains (i.e., tobacco and potato assays involving use of 'differential' potato cultivars). Additionally, sequence characterization of the complete genomes of isolates is highly recommended. However, it is acceptable to amend the names of PVY isolates with additional, specific codes to show that the isolate differs at the molecular, serological or phenotypic level from the typical strains within a strain group. The new isolates should preferably not be named using geographical, cultivar, or place-association designations. Since many new variants of PVY are being discovered, any new static classification system will be meaningless for the time being. A more systematic investigation and characterization of PVY from potato at the biological and molecular levels should eventually result in a biologically meaningful genetic strain concept.

Specific detection of the PVY(N)-W variant of Potato virus Y.

PVY(N)-W is one of the variant populations of Potato virus Y (PVY). This variant is of concern in seed potato production and requires a specific diagnosis since it induces more or less symptomless infections and is not detectable easily in field inspections. Moreover, this variant is serologically indistinguishable from the common strain PVY(O). This study describes a simple and specific molecular detection test for the PVY(N)-W variant using a PCR protocol based on the recombinant point within the HC-Pro/P3 region of PVY(N) variants (PVY(NTN), PVY(N)-W). To avoid both detection of recombinant PVY(NTN) and PVY(N)-W isolates, a forward PVY(N)-like primer located in the HC-Pro region coupled to a reverse PVY(O)-like primer located in the NIa region was designed to amplify a specific PCR product of 4114 nt from PVY(N)-W isolates. This technique was assessed on 41 PVY reference and field isolates. Only isolates referenced as PVY(N)-W were amplified and gave the expected PCR product of 4114 nt, whereas no band was obtained from PVY(N), PVY(NTN) or PVY(O) isolates. In conclusion, this PVY(N)-W diagnosis tool is rapid, easy-to-use and suitable for large-scale testing in laboratories of seed potato certification.

Genomic variability in Potato potyvirus Y (PVY): evidence that PVY(N)W and PVY(NTN) variants are single to multiple recombinants between PVY(O) and PVY(N) isolates.

Fourteen Potato virus Y (PVY) isolates representative of PVY(O), PVY(N), PVY(NTN) and PVY(N)W groups were characterised at genomic level. Restriction fragment length polymorphism study (RFLP) of each gene of these isolates and sequencing of the first 2700 nucleotides of two PVY(N)W isolates were performed. A mosaic structure was revealed in PVY(N)W and PVY(NTN) genomes, which showed either PVY(O) or PVY(N)-like sequences, depending on the particular gene. Indeed, starting from the 5'-end, these isolates showed a switching, from PVY(N)- to PVY(O)-like sequence, in the HC-Pro C-terminal region. Reversion to PVY(N)-like sequence was also revealed in the NIa N-terminal area of PVY(NTN) isolates, followed by a switching back to a PVY(O)-like sequence in the CP gene. Lastly, some PVY(N)W isolates showed a switching from PVY(O)- to PVY(N)-like sequence in the P1 N-terminal part, thus separating our PVY(N)W isolates into two subgroups. All these apparent recombination events were shown by statistical analysis. Comparison of molecular traits with pathogenic properties of our isolates suggested that the HC-Pro protein is involved in induction of necrosis in tobacco leaves, and the NIa, NIb and/or CP protein in necrosis in potato tubers. Nevertheless, multiple recombination events observed in the PVY(NTN) genome may play a role in the latter phenomenon.

RFLP mapping of the whole genome of ten viral isolates representative of different biological groups of potato virus Y.

Ten PVY isolates representative of four PVY groups (YN, YNTN, YN-W, YO), differing by their ability to induce reactions of vein necrosis on tobacco and tuber necrosis on potato, were studied in order to research the regions of the viral genome involved in these necrosis phenomena. The whole genome of these isolates was amplified in two fragments (4,063 and 5,670 nucleotides) and was subjected to a restriction fragment length polymorphism (RFLP) study. In the first 4,063 nucleotides of the PVY genome, a phenetic analysis of RFLP data resulted in a clustering of our PVY isolates into three groups: PVYN isolates (group A); PVYNTN and PVYN-W isolates (group B) and PVYO isolates (group C). In the last 5,670 nucleotides, two groups were found: PVYN and PVYNTN isolates (group D) and PVYO and PVYN-W isolates (group E). From this clustering and the necrosing properties known for these isolates, the tobacco necrosis determinants seem more likely located in the 5' than in the 3' half part of the viral RNA, whereas it would be the opposite situation for the determinants of the necrosis on potato tubers. Moreover a recombination event seemed to have occurred in the genome of the PVYN-W isolates.

Ultrastructural study of acquisition and retention of potato leafroll luteovirus in the alimentary canal of its aphid vector, Myzus persicae Sulz.

The chronology of PLRV acquisition and retention by Myzus persicae was investigated using electron microscopy. Examination demonstrated a rapid translocation of the virus through the intestine into the haemocoel. Indeed, viral particles could be observed in the intestinal epithelial cells, then in the haemocoel, 4 and 8 h, respectively, after their arrival in the lumen of the alimentary canal. However, the virus accumulated in the intestinal epithelial cells. In these cells, the first viral particles were seen enclosed in isometric or tubular isolated vesicles; a few hours later, they were present in tubular aggregated vesicles and also in lysosomes or multivesicular bodies. After a 40 h acquisition period, all studied intestinal epithelial cells exhibited high numbers of viral particles which were consistently distributed throughout these cell structures. When aphids were removed from viral source, viral particles were detected in intestinal lumen for a further three days and in intestinal epithelial cells for a total of eight days. Virus content in these cells began to decrease from the second day. Areas with tubular aggregated vesicles were maintained for seven days following aphid removal from viral source, but progressively became smaller and fewer. The accumulation and the persistence of PLRV in the intestine are discussed.

The intestine is a site of passage for potato leafroll virus from the gut lumen into the haemocoel in the aphid vector, Myzus persicae Sulz.

Four detection techniques, three of which gave reliable identification of the virus particles, were used to locate potato leafroll virus (PLRV) in the alimentary canal of its main aphid vector, Myzus persicae Sulz: immunofluorescence on cryostat sections, conventional transmission electron microscopy on ultrathin sections and immune electron microscopy with gold labeling, either prior to or after fixation-embedding. Each method clearly showed the presence of the virus in the intestine epithelium and its absence in cells of the other parts of the alimentary canal. Under the experimental conditions used, the intestinal cells seemed to be the pathway for PLRV transport from the gut lumen into the haemocoel. Electron microscopy examinations showed many virus particles close to the apical plasmalemma of the epithelial cells in the gut lumen of the intestine. Other particles were seen in shallow pit-like regions or surrounded by coated vesicles in the apical part of these cells. Thus the virus particles seemed to enter the epithelial cells of the intestine by a mechanism of endocytosis. In the cytoplasm of these cells, virions were also frequently observed in isolated--or more often aggregated--tubular vesicles. The latter could be involved in PLRV transport through the cell since they were observed fusing with different cell organelles. A few viral particles were also detected in lysosomes as well as in multivesicular bodies. Virus particles were observed between the plasmalemma and basal lamina of the intestine cells but not in the haemocoel, where probably they were quickly dispersed. Our results are discussed in relation to other reports which have shown hindgut and stomach as sites of passage from the gut lumen into the aphid's body cavity for PLRV and other circulative viruses.

Localization of cauliflower mosaic virus in the cell nucleus of Brassica pekinensis L.

Cauliflower mosaic virus (CaMV) particles were observed in the nuclei of xylem parenchyma cells in Brassica pekinensis L. doubly infected by CaMV and turnip mosaic virus (TuMV). CaMV particles were aggregated in the nucleoplasm but not embedded in viroplasms. This phenomenon was not detected in cell nuclei of mesophyll tissue. Typical features associated with infection by either CaMV or TuMV normally occurred in the cytoplasm of cells of both tissues: two types of viroplasms with embedded CaMV particles and cylindrical inclusions induced by TuMV. Among the main hypotheses which could explain this particular CaMV localization, we checked that it was not the result of the coinfection with TuMV, since we also found CaMV particles in the same place in single infected plants. Other explanations are discussed, including a possible particularity of the infection in xylem tissue or a specific property of an unusual CaMV isolate.

Visualization of a luteovirus in the vector aphid's body by two gold immunolabelling techniques: a comparative study.

Two gold immunolabelling techniques using electron microscopy were compared to examine the in situ localization of a luteovirus, potato leafroll virus (PLRV), inside its main aphid vector, Myzus persicae SULZ. With Gildow's technique, virus particles were labelled prior to fixation, embedding by injecting PLRV-specific IgGs into the living aphids. This facilitated the detection of extracellular particles located between the basal lamina and plasmalemma by trapping them in aggregates. The heavy coating of particles by antibodies and gold indicated good labelling sensitivity. Isometric virus-like particles were also observed inside the cytoplasm, but they were non decorated because the cell membrane prevented labelling reagents from entering the cell. With the second technique, ultrathin sections were immunolabelled after fixation-embedding. Since PLRV lost its antigenicity when aphid tissues were normally treated for electron microscopy, the successful application of this technique required fixation in 4% formaldehyde before embedding in Lowicryl at low temperature; it was also necessary to use PLRV-specific monoclonal antibodies to eliminate non-specific reactions. In these conditions, all intra- and extra-cytoplasmic virions present on the surface of sections were surrounded by gold particles, but the antibody coating was not discernible, and, because the resin limited the access of markers to antigens, the inner virus particles were not labelled. In conclusion, both techniques must be applied on the same material to give complementary information.