Potato Virus Y from Petunia can cause Symptoms of Potato Tuber Necrotic Ringspot Disease (PTNRD)

A potyvirus known to be an important agent involved in causing a disease of trailing petunias, was identified as being a member of the necrotic strain of potato virus Y (PVY) using a number of monoclonal antibodies. The sequence of the coat protein gene for the PVY isolate was determined and when compared with sequences for other PVY strains it was shown to cluster closely with isolates of PVY^NTN and to have a recombination point present within the coat protein common with other isolates of PVY^NTN. When inoculated onto potato tuber necrotic ringspot disease (PTNRD) susceptible potato cultivars the petunia isolate was found to be capable of causing necrotic tuber symptoms, consistent with those caused by other isolates of PVY^NTN. Due to the number of similarities it is thought the petunia isolate belongs to the PVY^NTN group of isolates. Out of 24 species of bedding and pot plant crops tested, 19 were shown by mechanical inoculation to be susceptible to PVY, highlighting not only a clear risk to a number of commercially important plant species from PVY^NTN infected trailing petunias, but also other susceptible crops grown in these areas.     

The effects of co‐infection by different Potato virus Y (PVY) isolates on virus concentration in solanaceous hosts and efficiency of transmission

The influence of co‐infection on concentration and accumulation of genetically different isolates of Potato virus Y (PVY) in potato and tobacco plants and the efficiency of transmission by Myzus persicae of PVY isolates from doubly versus singly infected plants were evaluated. The vector ability to simultaneously transmit two virus isolates was examined. Eight PVY isolates represented three strain groups: PVY^O (pathotype and serotype O), PVY^NW (pathotype N and serotype O), and PVY^NTN (pathotype and serotype N). Different diagnostic methods, including DAS‐ELISA, multiplex RT‐PCR, aphid transmission tests and bioassays, were applied to detect the presence of PVY isolates in source and assay plants. Significant reductions in concentrations of certain PVY isolates during co‐infection with other isolates were found both in potato and tobacco plants. The observed effects were both isolate‐ and host‐dependent in form. The highest rates of virus transmission by single aphids were recorded with PVY^NTN isolates, and the lowest ones with PVY^O isolates. Individual aphids of M. persicae were able to simultaneously transmit two PVY isolates. The frequency of transmission was generally low, but it reached as high as 20% for one of the isolate combinations. The findings presented in the work provide proof for antagonistic within‐plant interactions between isolates of PVY, with some implications of these interactions for virus transmission by aphid vectors. Consequently, this research contributes to a better understanding of the epidemiology of the disease caused by PVY.     

Occurrence of Potato virus Y strain PVYNTN in foundation seed potatoes in Japan,and screening for symptoms in Japanese potato cultivars

In 2008 and 2009 seasons, a sudden increase in Potato virus Y (PVY) incidence was recorded in foundation seed potatoes in Hokkaido, northern Japan. This increase was obvious during the field inspection and the postharvest indexing. Molecular typing revealed that besides the previously reported strains of PVY^O and PVY^NA‐N, the most common strain identified was the recombinant PVY^NTN, with three characteristic recombinant junctions at the HC‐Pro, VPg and CP regions. No potato tuber necrotic ringspot disease (PTNRD) was observed in foundation seed potatoes in correlation with the presence of PVY^NTN. Moreover, an isolate with a typical PVY^NTN recombinant genome, namely Eu‐12Jp, did not induce PTNRD in 62 Japanese potato cultivars tested in both primarily and secondarily infected plants. Two cultivars carrying the extreme resistance gene Ry_chc were resistant to the infection with Eu‐12Jp, which presents potential sources of resistance to PVY^NTN. Eu‐12Jp induced systemic mottle in potato cultivars Desiree and King Edward carrying resistance genes Ny and Nc, respectively, but induced a hypersensitive reaction in potato cultivar Maris Bard, with the Nz hypothetical resistance gene typical of the PVY^Z strain group. Therefore, based on the genome structure and the reaction of the potato N resistance genes, Eu‐12Jp should be classified as PVY^Z‐NTN, as described for isolates from Idaho, USA recently. This is the first report of PVY^Z‐NTN in Japan and the sudden and increased occurrence of PVY^NTN/PVY^Z‐NTN represents a potential risk of PTNRD developing and increases the significance of PVY in Japan.     

Characteristics of a resistance-breaking isolate of potato virus Y causing potato tuber necrotic ringspot disease

An Austrian isolate of potato virus Y^NTN, the causal agent of potato tuber necrotic ringspot disease (PTNRD), was serologically compared with seven Dutch PVY^N isolates. Using polyclonal and monoclonal antibodies, it was found indistinguishable from PVY^N. Determination of the nucleotide sequence of the coat protein cistron and comparison of the deduced amino acid sequence with coat protein sequences of other potyviruses revealed a high level of homology with PVY^N coat protein sequences. This confirmed the close taxonomic relationship of PVY^NTN with the PVY^N subgroup of potato virus Y. PVY^NTN is able to overcome all resistance genes known so far in commercial potato cultivars. Remarkably, transgenic PVY-protected tobacco plants are also resistant to PVY^NTN infection upon mechanical and aphid-mediated inoculation. These experiments indicate that genetically engineered resistance offers great potential in protection of potato to new aggressive strains of PVY^N.     

Characterization of potato potyvirus Y (PVY) isolates from seed potato batches. Situation of the NTN, Wilga and Z isolates

A collection of 38 PVY isolates from seed potato batches, originating from several Western European countries, was characterized by using current biological, serological and molecular tools differentiating PVY strains and groups. The correlation between the three kinds of tests was good but not absolute. No single serological or PCR method was able to discriminate among the five isolate groups found. Twenty-nine isolates belonged to the PVY^N strain and six to the PVY^O strain. No PVY^C was found. Two other isolates reacted serologically like PVY^O, but were unable to elicit a hypersensitive response from the Ny_tbr gene and probably represent the PVY^Z group. At the molecular level, these two isolates showed a combination of both PVY^O and PVY^N and could be recombinants of these strains. Another isolate reacted serologically like PVY^O, but induced vein necrosis in tobacco, like PVY^N-Wilga. Some PVY^N isolates caused tuber ring necrosis in glasshouse conditions. These might belong to the PVY^NTN group. The PVY^NTN, PVY^N-Wilga and PVY^Z groups probably represent pathotypes within strains PVY^N and PVY^O, respectively. The present study also confirms previous reports showing a high genetic variation at the 5 end within the PVYN strain.     

Systemic infection with potato virus YNTNalters the structure and activity of the shoot apical meristem in a susceptible potato cultivar

Stem nodes of the potato virus Y^NTN(PVY^NTN) susceptible potato cultivar (Solanum tuberosum cv. “Igor”) were studied to investigate whether alterations in shoot morphology caused by infection with PVY^NTNare related to changes in structure and activity of the shoot apical meristem. The PVY^NTNcauses severe disease symptoms known as the potato tuber necrotic ringspot disease (PTNRD) in susceptible potato plants grown in the field. Although in stem node culture the symptoms are mildly expressed, in systemically PVY^NTN-infected plants the shoot height stem diameter, number of leaves and the total leaf area were lower than in the healthy control plants at day 18 and 35 of culture. Histological analysis of the longitudinal sections of the shoot tip revealed that the infected plants had smaller shoot apical meristems due primarily to a lower cell number in different meristem zones. One of the most affected zones of the apical shoot meristem was the peripheral zone, the region in which leaf primordia are induced. The data is consistent in showing that the infection of potato plants with PVY^NTNleads to alterations in the shoot apical meristem structure and mitotic activity. These changes are, in turn, reflected in altered overall morphology of the infected plant.     

A novel type of Potato virus Y recombinant genome,determined for the genetic strain PVYE

Two Potato virus Y (PVY) isolates collected in Brazil, PVY‐AGA and PVY‐MON, were identified as recombinants between two parent genomes, PVY^NTN and PVY‐NE‐11, with a novel type of genomic pattern. The new recombinants had an ordinary PVY^NTN genome structure for approximately 6·7‐kb from the 5′‐end of the genome whereas the 3′‐terminal 3·0‐kb segment had two fragments of NE‐11‐like sequence separated by another small PVY^NTN‐like fragment. PVY strains are defined based on the hypersensitive resistance (HR) response in potato indicators. Both PVY‐AGA and PVY‐MON isolates did not induce the HR in potato cultivars carrying Ny, Nc, or (putative) Nz genes and thus were able to overcome all known resistance genes to PVY. Only one of the two isolates, PVY‐AGA, induced a vein necrosis reaction in tobacco. The biological responses of the potato indicators and tobacco defined PVY‐MON as an isolate of the PVY^E strain. To distinguish PVY‐AGA and PVY‐MON from other PVY^NTN isolates, an RT‐PCR test was developed utilizing new specific primers from the capsid protein gene area and producing a characteristic 955‐bp band. Serological profiling of these PVY isolates with three monoclonal antibodies revealed an unusual reactivity, where one of the two commercial PVY^N‐specific monoclonal antibodies did not recognize PVY‐AGA. The ability of these new PVY recombinants to overcome resistance genes in potato producing mild or no symptoms, combined with the lack of serological reactivity towards at least one PVY^N‐specific antibody may present a significant threat posed by these isolates to seed potato production areas.     

Effect of temperature on resistance to Potato virus Y in potato cultivars carrying the resistance gene Rychc

The effect of cultivation temperatures on the resistance reaction to three Potato virus Y strains (PVY^O, PVY^N and PVY^NTN) in potato cultivars carrying Ry_chc was examined. When potato plants carrying Ry_chc were cultivated at 22 °C, a few small necrotic spots developed on inoculated leaves by 5 days after mechanical inoculation (dpi), and systemic infection of a few symptomless plants was confirmed at 28 dpi by IC‐RT‐PCR. At 28 °C, distinct necrotic spots developed on inoculated leaves by 5 dpi, and systemic symptoms occasionally appeared at 28 dpi. Thus, high temperature weakens Ry_chc‐conferred resistance. However, the incidence of systemic infection and the titre of virus in resistant cultivars at 28 °C were lower than in a susceptible cultivar. In graft inoculation under high summer temperatures, some plants developed necrosis on the leaves and stem, but PVY was barely detected by RT‐PCR in leaves on potato carrying Ry_chc. When seedlings from progeny tubers of plants that were inoculated with PVY and grown in a greenhouse at >30 °C in the daytime were examined by ELISA and IC‐RT‐PCR, PVY was not detected in cultivars carrying Ry_chc. These results show that Ry_chc confers an extreme resistance to PVY strains occurring in Japan.     

Incidence of potato viruses and characterisation of <Emphasis Type="Italic">Potato virus Y</Emphasis> variability in late season planted potato crops in Northern Tunisia

Surveys were conducted of symptomatic potato plants in late season crops, from the major potato production regions in Northern Tunisia, for infection with six common potato viruses. The presence of Potato leafroll virus (PLRV), Potato virus Y (PVY), Potato virus X (PVX), Potato virus A (PVA), Potato virus S (PVS) and Potato virus M (PVM) was confirmed serologically with virus infection levels up to 5.4, 90.2, 4.3, 3.8, 7.1 and 4.8%, respectively. As PVY was prevalent in all seven surveyed regions, further biological, serological and molecular typing of 32 PVY isolates was undertaken. Only one isolate was shown to induce PVY^O-type symptoms following transmission to tobacco and to react only against anti-PVY^O-C antibodies. Typical vein necrosis symptoms were obtained from 31 samples, six of which reacted against both anti-PVY^N and anti-PVY^O-C antibodies showing they contained mixed isolates, while 25 of them reacted only with anti-PVY^N antibodies. An immunocapture RT-PCR molecular test using a PVY^NTN specific primer pair set in the 5’NTR/P1 genomic region and examination of recombinant points in three genomic regions (HC-Pro/P3, CI/NIa and CP/3’NTR) showed that all 25 serotype-N PVY isolates were PVY^NTN variants with similar recombinations to the standard PVYNTN-H isolate. This is the first report of the occurrence of the PVY^NTN variant and its high incidence in late season potatoes in Tunisia.     

Virus transmission by aphids in potato crops

This paper reviews the contribution of vector activity and plant age to virus spread in potato crops. Determining which aphid species are vectors is particularly important for timing haulm destruction to minimize tuber infection by potato virus Y (PVY). Alate aphids of more than 30 species transmit PVY, and aphids such asRhopalosiphum padi, that migrate in large numbers before flights of the more efficient vector,Myzus persicae, appear to be important vectors. Differences in methodology, aphid biotypes and virus strains prevent direct comparisons between estimates of vector efficiencies obtained for aphids in different countries in north western Europe. M. persicae is also the most efficient vector of potato leafroll virus (PLRV), but some clones ofMacrosiphum euphorbiae transmit PLRV efficiently toNicotiana clevelandii and potato test plants. The removal of infected plants early in the season prevents the spread of PLRV in cool regions with limited vector activity. The proportion of aphids acquiring PLRV from infected potato plants decreases with plant age, and healthy potato plants are more resistant to infection later in the season. Severe symptoms of secondary leafroll developed on progeny plants of cv. Maris Piper derived from mother plants inoculated with PLRV in June or July of the previous year. Progeny plants derived from mother plants inoculated in August showed only mild symptoms, but the concentration of PLRV in these plants was as high as that in the plants with severe symptoms.     

Aggressive and mild Potato virus Y isolates trigger different specific responses in susceptible potato plants

Differences in the early responses of two potato cultivars, Igor and Nadine, to two isolates of Potato virus Y (PVY), the aggressive PVY^NTN and the mild PVY^N, were monitored. Microarray and quantitative real‐time PCR analyses were carried out to identify differentially expressed genes after inoculation with each virus isolate. Additionally, symptom severity and development was observed and the amount of virus isolate accumulated in systemically infected leaves was evaluated, where a significantly higher amount of PVY^NTN was detected. Microarray analysis revealed 572, 1288 and 1706 differentially expressed genes at 0·5, 12 and 48 h post‐inoculation, respectively in cv. Igor, with a similar pattern observed in cv. Nadine. Microarray and quantitative real‐time PCR results implied an earlier accumulation of sugars and lower photosynthesis in leaves inoculated with the aggressive isolate than in leaves inoculated with the mild isolate. The PVY^NTN isolate did not activate early differential expression of the Fe‐superoxide dismutase and pectin methylesterase inhibitor (PMEI) genes, indicating a delay in plant response relative to that following PVY^N inoculation. Differences in the expression of the β‐glucanase‐I gene were also observed in early plant responses to inoculation with each virus isolate.     

Virus transmission by aphids in potato crops

This paper reviews the contribution of vector activity and plant age to virus spread in potato crops. Determining which aphid species are vectors is particularly important for timing haulm destruction to minimize tuber infection by potato virus Y (PVY). Alate aphids of more than 30 species transmit PVY, and aphids such asRhopalosiphum padi, that migrate in large numbers before flights of the more efficient vector,Myzus persicae, appear to be important vectors. Differences in methodology, aphid biotypes and virus strains prevent direct comparisons between estimates of vector efficiencies obtained for aphids in different countries in north western Europe.M. persicae is also the most efficient vector of potato leafroll virus (PLRV), but some clones ofMacrosiphum euphorbiae transmit PLRV efficiently toNicotiana clevelandii and potato test plants. The removal of infected plants early in the season prevents the spread of PLRV in cool regions with limited vector activity. The proportion of aphids acquiring PLRV from infected potato plants decreases with plant age, and healthy potato plants are more resistant to infection later in the season. Severe symptoms of secondary leafroll developed on progeny plants of cv. Maris Piper derived from mother plants inoculated with PLRV in June or July of the previous year. Progeny plants derived from mother plants inoculated in August showed only mild symptoms, but the concentration of PLRV in these plants was as high as that in the plants with severe symptoms.     

The impact of potato cysteine proteinases in plant growth and development

The role of cysteine proteinases in the growth and development of healthy and PVY^NTN infected potato plants (Solanum tuberosum L. cv. ‘Désirée’) and the subcellular localization of a particular potato leaf cysteine proteinase PLCP-2 were studied in tissue culture. The immunolocalization of PLCP-2 on the ultrathin sections of potato plants was examined by electron microscopy. PLCP-2 was found in protein bodies in vacuoles, in cytoplasm and in cell walls of shoot tips, leaves, stems and root tips. PLCP-2 was observed at all locations where its endogenous inhibitors PCPI 6·6 and multicystatin were localized, which indicates the possible regulation of PLCP-2 by these inhibitorsin vivo . In order to study the physiological role of cysteine proteinases in potato plants, their total activity was diminished by introducing Ep-475 into the growth media and further into the plant through root absorption. The morphological characteristics of healthy and PVY^NTN infected plants grown on media with added Ep-475 were followed. The results suggest that cysteine proteinases are involved in the synthesis and transport of plant metabolites and in processes that lead to organogenesis. Observations also indicate that the ratio between cysteine proteinases and their endogenous inhibitors in potato plants is altered after infection with PVY^NTN.     

Problems associated with potyviruses in potato certification-field inspection and serological testing1

In UK, the tobacco veinal necrosis strain of potato virus Y (PVY^N), potato virus A (PVA) and potato virus V (PVV) each occur in the field only in limited ranges of potato cultivars in which they mostly cause mild symptoms or even symptomless infection; little is known about incidence of strain C of PVY (PVY^C). The ordinary strain of PVY (PVY°), however, is widespread causing symptoms ranging in severity from very severe through to very mild, depending on cultivar sensitivity/tolerance. During field inspections, very mild potyvirus symptoms may be missed, so inspectors are trained to be particularly vigilant when examining problem cultivars which react in this way. PVA is almost invariably treated, along with PVX, as a mild kind of virus infection, but infections with PVY°, PVY^N and PVV are treated as severe with stricter tolerances being applied for them (especially for PVY^N) regardless of symptom severity. Wide variation within the same cultivar in the behaviour of variants within the PVY° strain group also sometimes causes difficulties in interpretation at inspection. To detect PVY, PVA and PVV in routine serological testing on potato certification samples, it is necessary to employ specific antisera to each of them. PVY^N-specific monoclonal antibodies can be used in ELISA to distinguish PVY^N from PVY°.     

Direct tuber testing for Potato Y potyvirus by real‐time RT‐PCR and ELISA: reliable options for post‐harvest testing?*

The method currently used for testing potato tubers for viruses following harvest involves a growing‐on test. This takes up to 6 weeks to complete, and there is therefore a demand for more rapid test results. The sensitivity and reliability of direct tuber testing by DAS‐ELISA and real‐time RT‐PCR (TaqMan) were compared with the growing‐on test. In addition, the reliability of all three methods for the detection of Potato Y potyvirus (PVY) in tubers was compared over post‐harvest intervals of 6, 10, 14 and 18 weeks. The test material came from plots of tubers (cv. ‘Maris Piper’) containing a primary infection of strains PVY^N and PVY^O, following aphid transmission from marked infector plants grown during the 2003 season. Sample material was homogenized and divided, to provide comparative test material for detection of PVY by ELISA and real‐time RT‐PCR. Tuber eye‐plugs were then taken and subjected to the growing‐on test. The remainder of the tuber was also grown on and tested, to ensure infection was not missed as a consequence of an uneven distribution of virus throughout the tuber material. The results obtained using the two methods for direct testing of the tubers, and those results obtained from the traditional growing‐on test, are compared. The advantages and disadvantages of each method are discussed.     

Enzyme-linked immunosorbent assay (ELISA) for the detection of potato viruses A and Y in potato leaves and sprouts

With the enzyme-linked immunosorbent assay (ELISA) potato virus A (PVA) could be detected reliably in potato sprouts, especially when these were young and sappy. The detection of this virus in leaves of glasshouse-grown potato plants was less reliable. The tobacco veinal necrosis strain of potato virus Y (PVY^N) was readily demonstrated in foliage of glass-house-grown potato plants using an antiserum to this strain. Plants infected with the common strain (PVY^O) did not react in ELISA with this antiserum. In young sappy sprouts, using the PVY^N antiserum, PVY^N could be detected reliably when samples with PVY^O were excluded, as the reaction of samples infected with the latter virus was intermediate between PVY^N-diseased and PVY-free samples. PVY was also detected in plants inadvertently infected during the experiments.     

Spread of potato virus Y in potato cultivars (Solanum tuberosum L.) with different levels of sensitivity

The aim of this work was to correlate the appearance of the symptoms, multiplication and spread of virus after mechanical inoculation of potato (Solanum tuberosum L.) cultivars showing different levels of susceptibility and sensitivity to Potato virus Y^NTN (PVY^NTN). The potato cultivars used were the resistant cultivar Sante and susceptible cultivars Igor, Pentland squire and Désirée. The spread of the virus PVY^NTN in infected plants was monitored using different methods: DAS-ELISA, tissue printing, immuno-serological electron microscopy and real-time PCR. In all three susceptible cultivars, the virus was detected in the inoculated leaves 4–5 days after inoculation. From there virus spread rapidly, first into the stem, then more or less simultaneously to the upper leaves and roots. Real-time PCR was shown to be very sensitive and enabled viral RNA to be detected in non-inoculated leaves of susceptible cultivar Igor earlier than other methods. Therefore, for exact studies of plant–virus interaction, a combination of methods which detect viruses on the basis of their different properties (coat protein, morphology or RNA) should be used to monitor the spread of viruses.     

两个马铃薯Y病毒黑龙江马铃薯分离物株系鉴定

为明确分离自黑龙江省克山县马铃薯上的2个病毒分离物KS4和KS7的分类地位,通过RTPCR扩增、克隆获得其基因组序列,利用重组分析程序包和最大似然法分别进行重组分析和系统发育分析。结果显示,分离物KS4和KS7的开放阅读框均有9 186个核苷酸,编码3 061个氨基酸,分离物KS4的核苷酸和氨基酸序列均与马铃薯Y病毒(potato virus Y,PVY)分离物Mb112一致率最高,分别为96.9%和98.4%;分离物KS7的核苷酸序列与PVY分离物12-94一致率最高,为97.4%,其氨基酸序列与PVY分离物SYR-Ⅱ-Be1一致率最高,为97.8%。重组分析表明,分离物KS4和KS7均为分离物N-605和Oz的重组体,其中KS4基因组5′-端的2 392个核苷酸来自分离物N-605,其余核苷酸来自分离物Oz;KS7基因组的第800～2 227个核苷酸和第5 637～8 950个核苷酸来自分离物N-605,其余核苷酸来自分离物Oz。系统发育分析发现,分离物KS4被聚类到N:O株系(PVY~(N:O)),分离物KS7被聚类到NTN株系(PVY~(NTN))b型。