PVYN Isolates Research Articles

Dynamics of PVY strains in field grown potato: Impact of strain competition and ability to overcome host resistance mechanisms

Potato virus Y (PVY) is the most important viral pathogen affecting potato crops worldwide. PVY can be transmitted non-persistently by aphids that do not colonize the host plant, resulting in a rapid acquisition and transmission of the virus between plants. PVY exists as a complex of strains that can be distinguished according to their pathogenicity, serology and genomic analysis. While virus incidence remains low in Scottish seed potato crops, PVY has become the increasingly prevalent virus. The monitoring of PVYN and PVYO serotypes has revealed a recent shift towards PVYN which now accounts for more than 90% of all PVY cases. A survey of the molecular diversity of PVYN isolates indicated that 80%–90% belong to the recombinant European (EU)-NTN group, with North-American (NA)-NTN and non-recombinant EU-N variants accounting for the remainder. The shift from non-recombinant to recombinant PVY isolates is a common trend observed worldwide. Surveys of a range of PVY isolates representing the main strain and phylogenetic groups suggest that PVY has the ability to overcome hypersensitive response-mediated resistance with significant differences between isolates of the same strain group. Contrastingly, genes mediating extreme resistance (Ryadg, Rysto) provide efficient resistance to PVY transmission to progeny tubers. Transmission experiments in field conditions of PVY isolates representing the three main molecular groups (PVYO, PVYEU−NTN, PVYNA−NTN) indicate that PVYEU−NTN has the highest transmission rate. Our results suggest that PVYEU−NTN isolate has a competitive advantage over PVYO and PVYNA−NTN isolates which is likely to be an important factor in shaping the evolution of viruses and the population dynamics of PVY.

Recent Advances on Potato virus Y Research in Canada

Significant progress has been made in recent years in understanding pathological, serological and molecular properties of various strains of PVY and the aphid-mediated transmission. PVYO and PVYN appear to be the basic strain groups. Through genome recombination between these two groups, progeny groups whose genome possess one (e.g., PVYN:O or PVYN-Wi) to three (e.g., recombinant PVYNTN or European-PVYNTN) recombinant joints of PVYO and PVYN emerged. PVYN:O causes PVYN-like veinal necrosis in tobacco, but reacts to PVYO-specific antibody. PVYNTN causes potato tuber necrotic ringspot diseases in sensitive potato cultivars, and PVYN-like necrosis in tobacco plants, and reacts to PVYN-specific antibody. Through single nucleotide mutation(s), non-recombinant PVYNTN (or North American PVYNTN) also emerged from PVYN. It is also noteworthy that PVYN isolates originated from North America and Europe may have evolved separately; and to date most recombinant strains appear to be progenies of Eu-PVYN and PVYO. Several RT-PCR-based methodologies have been developed to characterize and detect various strains of PVY. A field survey revealed that PVYN:O has become a predominant strain in Manitoba and neighbouring states in USA. Moreover, three distinct variant groups inciting severe, intermediate and mild veinal/petiole/stem necrosis, respectively, on tobacco plants were observed within the PVYN:O isolates collected in Manitoba. Pathological and molecular diversity within PVYO strain group were revealed in New Brunswick, represented by PVYO-FL as a severe variant and by PVYO-RB as a mild variant. Studies on the transmission of PVY by various species of aphids revealed that aphid behavior plays an important role in the vector-mediated transmission. Application of mineral oil on the growing crop, especially in combination with use of crop borders, reduces aphid-mediated PVY transmission. Based on recent PVY research studies, Bartlett Superior 70 Oil was approved in 2011 for application to potato crops in Canada.

Recombinants of PVY Strains Predominate Among Isolates from Potato Crop in Poland

Recombinants of PVY Strains Predominate Among Isolates from Potato Crop in Poland 282 Potato virus Y (PVY) isolates collected from potato crops in northern and central Poland from 1995 to 2009 were characterized by serological and biological assays. From these, 112 isolates collected from 2006 to 2009 were additionally analyzed by one-step triplex Reverse Transcription-Polymeraze Chain Reaction (RT-PCR). Recombinants of PVY strains predominate among the isolates tested. Using one-step triplex RT-PCR most PVYN-Wi isolates were classified as subgroup PVYNWi-P and most PVYN and/or PVYNTN isolates as the recombinant PVYNTN strain. A recombinant PVYNTN isolate (12/94) and two additional PVYN and/or PVYNTN isolates were not detected by one-step triplex RT-PCR. Twelve isolates were identified as the PVYO strain but PVYN, non-recombinant PVYNTN and PVYC strains were not found. Serological and biological assays of 144 isolates of PVYN-Wi strain showed that 100 isolates were the expected PVYO serotype with vein necrosis (VN) symptoms on tobacco. However, 10 isolates of the PVYNWi-P subgroup exhibited vein clearing (VCl) on tobacco and 2 isolates of the PVYNN242 subgroup unexpectedly exhibited as the PVYN serotype. All the isolates of PVYN-Wi strain induced severe local lesions (LL) on Chenopodium amaranticolor. Out of 126 isolates of PVYNTN strain tested, 76 were typical PVYN serotype with VN on tobacco, but their reactions on C. amaranticolor were different: 13 isolates did not show symptoms, 23 isolates induced weak, and 40 isolates induced severe LL. The remaining isolates of PVYN-Wi, PVYN and/or PVYNTN or PVYO were serologically PVYN and PVYO positive or exhibited unpredictable serological and biological reactions.

Identification of the Molecular Make-Up of the Potato virus Y Strain PVYZ: Genetic Typing of PVYZ-NTN

Potato virus Y (PVY) strains were originally defined by interactions with different resistance genes in standard potato cultivars. Five distinct strain groups are defined that cause local or systemic hypersensitive responses (HRs) in genetic background with a corresponding N gene: PVY(O), PVY(N), PVY(C), PVY(Z), and PVY(E). The nucleotide sequences of multiple isolates of PVY(O) and PVY(N) differ from each other by ≈8% along their genomes. Additionally, complete genome sequences of multiple recombinant isolates are composed of segments of parental PVY(O) and PVY(N) sequences. Here, we report that recombinant isolate PVY-L26 induces an HR in potato 'Maris Bard' carrying the putative Nz gene, and is not recognized by two other resistance genes, Nc and Ny(tbr). These genetic responses in potato, combined with the inability of PVY-L26 to induce vein necrosis in tobacco, clearly define it as an isolate from the PVY(Z) strain group and provide the first information on genome structure and sequence of PVY(Z). The genome of PVY-L26 displays typical features of European NTN-type isolates with three recombinant junctions (PVY(EU-NTN)), and the PVY-L26 is named PVY(Z)-NTN. Three typical PVY(NTN) isolates and two PVY(N) isolates, all inducing vein necrosis in tobacco, were compared with PVY-L26. One PVY(NTN) isolate elicited HR reactions in Maris Bard, similar to PVY-L26, while two induced a severe systemic HR-like reaction quite different from the quasi-symptomless reaction induced by two PVY(N) isolates. 'Yukon Gold' potato from North America produced HR against several PVY(NTN) isolates, including PVY-L26, but only late and limited systemic necrosis against one PVY(N) isolate. Consequently, according to symptoms in potato indicators, both PVY(Z) and PVY(NTN) isolates appeared biologically very close and clearly distinct from PVY(O) and PVY(N) strain groups.

Presence of necrotic strains of Potato virus Y in Mexican potatoes.

As part of a routine screening for the possible presence of the necrotic strains of potato virus Y affecting potatoes in Mexico, five PVY isolates were submitted to biological and molecular analysis. Considering their serological properties, two belong to the common strain (O) and three to the necrotic strain (N). All the isolates induced vein necrosis in Nicotiana tabacum. To characterize the isolates, 5' NTR and P1 genes were sequenced and compared with sequences from GenBank database. Nucleotide sequence similarity ranged from 47–100% in the 5' NTR and from 63–100% in the P1 coding region. The lowest amino acid similarity between sequences of P1 gene was 55%. In phylogenetic trees of 5'NTR analysis, two PVYO Mexican isolates clustered with other PVYO isolates. In turn, the three PVYN isolates grouped with PVYN-NTN isolates. The phylogenetic analysis of P1 sequences (nucleotide and amino acid) showed two PVYO isolates grouping next to N-NTN cluster. A detailed analysis of the PVYO isolates showed two potential recombination regions in the P1 gene, in contrast to 5'NTR where no recombination was detected.

Biological and Serological Properties of Potato virus Y Isolates in Northeastern United States Potato.

A survey of six potato viruses, Potato virus A (PVA), Potato virus M (PVM), Potato virus S(PVS), Potato virus X (PVX), Potato virus Y (PVY), and Potato leafroll virus (PLRV), was conducted in New York and Maine during 2002 and 2003. Leaf samples were tested by enzyme-linked immunosorbent assay and PVY-positive samples were further tested to determine whether a necrotic strain of PVY (PVYN) or a strain able to induce necrosis in tobacco and in potato tubers (PVYNTN) were present. In both years, PVY and PVS were identified in a majority of the samples, and mixed infections predominated in 83% of the symptomatic leaves in 2002. Of the total 394 PVY-positive samples, 3 reacted with monoclonal antibody (MAb) 1F5 and caused veinal necrosis (VN) in tobacco. Two of these isolates caused tuber necrosis in the potato cv. Yukon Gold. Three PVY isolates reacted with MAb 1F5 but did not cause VN in tobacco, and two caused VN but did not react with MAb 1F5. None of these eight isolates were able to overcome the Ry resistance gene in the potato cultivar Eva, but several were able to overcome the Ny resistance gene found in Allegany. PVYN isolates were not widespread in the northeastern United States; however, several PVY isolates differed from both PVYN and the ordinary strain of PVY and may represent strain recombinants.

Differentiation and molecular detection of Canadian necrotic strains of Potato virus Y

Isolates of Potato virus Y (PVY) from potatoes grown in Canada were identified as tuber necrotic strains on the basis of biological, serological, and molecular characteristics. Only one isolate, however, caused symptoms of potato tuber necrotic ringspot disease (PTNRD) in progeny tubers of inoculated potato plants Solanum tuberosum 'Nadine' under greenhouse conditions. None of the isolates caused symptoms of PTNRD in potato plants 'Russet Burbank' or 'Saco'. A primer set based on the NIb gene for detection of PVYO by RT-PCR was modified to include detection of the Canadian necrotic isolates of PVY. Restriction analysis of amplicons generated from suspected PVYNTN isolates with primers based on the P1 protein gene differentiated two sets of isolates. Subsequent comparison of the P1 gene sequence of the Canadian isolates with published sequences confirmed that the isolates corresponded to North American and European types of PVYNTN. A new primer pair, based on the NIb gene sequence amplified by RT-PCR, allowed the amplification by RT-PCR of a gene fragment from all PVYN and PVYNTN isolates but not from PVYO strains. These primers proved to be useful for detecting the virus in composite samples of potato leaves, sprouts, and tubers.

Differences in aphid transmissibility and translocation between PVYN and PVYO isolates

The aphid transmissibility and within-plant movement of two PVY isolates belonging to PVYO and PVYN strains were studied. Potato plants were infected by Myzus persicae either 1 week after emergence or 30 or 45 days after emergence. Infected leaves, petioles of infected leaves, and different segments of the stems of plants infected 1 week after emergence were ELISA-tested 7, 11 and 14 days after infection. Segments of the plants infected 30 or 45 days after emergence were ELISA tested 30 days after infection. Normalised ELISA values were used for statistical analyses. Virus infection of progeny tubers was assessed by post-harvest inspection. The ability of 15 aphid species to transmit the virus isolates was also examined. There was no difference in the aphid transmissibility of the two isolates 7 or 11 days after exposure to M. persicae. However, normalised ELISA values were significantly affected by both isolate and sampling date 11 and 14 days after M. persicae transmission. The level of infection of different stem segments was significantly affected by both isolate and growth stage of the plant at infection. The proportions of infected leaves, stem segments and progeny tubers were all significantly higher for PVYN than for PVYO. PVYN was more effectively transmitted by a number of different aphid species than was PVYO. Newly described vectors of PVYN include Schizaphis graminum, Aphis fabae cirsiacanthoides, Aphis spiraecola, Myzus ligustri, Aphis fabae, Aphis spiraephaga, Myzus cerasi, Macrosiphum rosae, Diuraphis noxia, Aphis pomi and Rhopalosiphum padi, while those of PVYO include Aphis fabae cirsiacanthoides, Myzus cerasi and Myzus ligustri.

Potato Tuber Necrotic Ringspot Disease Occurring in Japan: Its Association with Potato virus Y Necrotic Strain.

Unfamiliar necrotic symptoms on or within potato tubers of cultivars Nishiyutaka and Dejima were observed in Nagasaki prefecture, Japan, in 1992. Symptoms were typically on the surface of the tuber, which either protruded at first and then became sunken, or showed necrotic spots, with necrosis within the tubers. Symptoms sometimes appeared at harvesting but more often appeared after storage for several months. Investigations revealed that the causal agents of the disease were isolates of Potato virus Y necrotic strain (PVYNTN) and the disease was potato tuber necrotic ringspot disease (PTNRD), previously reported in Europe and Lebanon. Five potato cultivars were inoculated with an isolate from necrotic tubers. The highest percentage of progeny tubers showing PTNRD was found in cv. Nishiyutaka (23.3%). In contrast, cvs. Shima-bara, Mayqueen, and Danshaku showed a low percentage of PTNRD. Additional potato tubers with PTNRD were also observed after storage of the tubers. To investigate the relatedness between isolates of PVYNTN and of necrotic strain PVYN, previously isolated in Japan, Nishiyu-taka was inoculated with an isolate of PVYN, which also induced PTNRD. Nucleotide sequences of coat protein (CP) genes of six PVYNTN isolates were determined. The CPs were 267 amino acids in length, with a substitution of one or no amino acid among each of the six isolates. The phylogenetic relationship based on nucleotide sequences of CP genes showed that these six PVYNTN isolates clustered together with PVYN isolates. This is the first report of PTNRD caused by PVYNTN isolates in Asia.

Screening by a 3-primer PCR of North American PVYN isolates for European-type members of the tuber necrosis-inducing PVYNTN subgroup

Oligonucleotide primers for a 3-primer polymerase chain reaction (PCR) were optimized by adjusting their Tm values to identify European-type members of the potato tuber necrotic ringspot disease (PTNRD) inducing PVYNTN sub-group of PVYN. Fiftyseven North American isolates of PVYN were screened using this assay and one, originating from garden potato tubers in Ontario, was found to be positive. This isolate was confirmed to induce the diagnostic PTNRD, using a panel of 20 potato cultivars. AC Chaleur, AC Novachip, Century Russet, Cherokee, and Frontier Russet reacted severely and developed PTNRD either at the time of harvest or during storage. About 800 tubers collected from New Brunswick and Prince Edward Island fields, from plants exhibiting mosaic and leafroll symptoms were also screened for various strains of PVY using PCR primers. Results indicate that primer pairs developed are specific for PVY spp., PVYN, and PVYNTN.

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.

Occurrence of Potato Tuber Necrotic Ringspot Disease (PTNRD) in Italy.

Potato tuber necrotic ringspot disease, caused by a tuber necrotic isolate of potato virus YN named PVYNTN, was originally described in Hungary in 1980; later on, the disease spread throughout northern and eastern Europe (1). Recently, the virus was reported in Portugal (2) on fresh-market potatoes. During the summer of 1997, the disease appeared in the north of Italy, causing serious damage in processing and fresh-market potatoes. After harvesting, we observed in most of the potato cultivars tubers showing the typical superficial necrotic rings and areas. These viral lesions were predisposed to secondary infection by fungi and bacteria. Moreover, PVYNTN causes degradation of tuber starch into simple carbohydrates. This metabolic activity resulted in undesirable browning in chips. In different areas of northern Italy, samples were collected from 48 stocks of cv. Hermes, the most important processing cultivar in Italy, and from 12 stocks of other potato cultivars (Ernterstolz, Liseta, Monalisa, Primura, Kennebec). The original seed of the stocks came from five different European countries: Austria, Italy, Scotland, Switzerland, and The Netherlands. Samples were serologically tested by indirect enzyme-linked immunosorbent assay with specific monoclonal antibodies to tobacco venial necrosis strain group of PVY. The results showed that 37 out of 60 analyzed stocks were infected by an isolate belonging to PVYN group, 7 stocks were infected by a common isolate of PVY, and 16 were PVY free. To distinguish the tuber-necrosis isolate (PVYNTN) from the PVYN isolates, reverse transcription-polymerase chain reaction (RT-PCR) and immunocapture-RT-PCR were carried out with specific primers for PVYNTN (3). Both tuber and leaf sap were tested. RT-PCR was performed on tuber sap and immunocapture-RT-PCR on leaf sap. Leaf and tuber sap of healthy, PVY-infected, and PVYNTN-infected tobacco was used in each experiment. At least two tubers from each of 32 PVYN-infected stocks were included in the molecular assay. All samples showed an amplified product of the characteristic size for PVYNTN (835 bp), whereas neither the healthy control nor the PVY sample gave a product. All tested cultivars were infected and all countries from which tuber seed came revealed infected stocks. The high proportion of samples in which PVYNTN was detected suggests that the virus is now endemic to potato-growing regions of northern Italy. The weather in these regions during 1997 was mild and drought in winter-spring and very warm in late summer; these conditions were ideal for disease diffusion by vectors and for symptom development. This is the first report of the occurrence of PVYNTN in Italy.

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

An Austrian isolate of potato virus YNTN, the causal agent of potato tuber necrotic ringspot disease (PTNRD), was serologically compared with seven Dutch PVYN isolates. Using polyclonal and monoclonal antibodies, it was found indistinguishable from PVYN. 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 PVYN coat protein sequences. This confirmed the close taxonomic relationship of PVYNTN with the PVYN subgroup of potato virus Y. PVYNTN is able to overcome all resistance genes known so far in commercial potato cultivars. Remarkably, transgenic PVY-protected tobacco plants are also resistant to PVYNTN 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 PVYN.

Molecular cloning and sequencing of the capsid and the nuclear inclusion protein genes of a North American PVYN isolate

The sequence of the 3'-terminal 3258 nucleotides of a tobacco veinal necrosis strain of a potato virus Y (PVYN) isolate from North America was determined. The sequence revealed an open reading frame of 2931 nucleotides, of which the start codon was not identified. The nontranslated region contains 327 nucleotides upstream of a poly(A) tract. The open reading frame encodes a large polyprotein containing 976 amino acids. The data indicate that the coat protein (CP) and the nuclear inclusion protein (NIb) are derived from the large polypeptide by proteolytic cleavage similar to many other potyviruses. The putative cleavage sites of NIa/NIb and NIb/CP correspond to Q/A and Q/G sequences, respectively, as in other PVYN isolates. The CP and NIb contain 267 and 519 amino acid residues, respectively. Higher sequence homology of CP was observed with other PVYN isolates than with the PVYO isolates.