Tissue Culture Methods for the Screening and Analysis of Putative Virus-Resistant Transgenic Potato Plants

Following regeneration, putative virus-resistant transgenic plants are usually transferred from tissue culture to a greenhouse or growth chamber to screen for resistance to infection and disease development using mechanical, graft, or insect vector inoculation methods. To reduce initial screening costs and time, we developed mechanical and graft inoculation methods suitable for tissue culture use. The in vitro methods were validated by comparing them with similar greenhouse screens using putative potato virus Y strain o (PVY°) replicase-mediated resistant regenerants of the potato cultivar Atlantic. Five transgenic lines were tested, with similar results obtained from in vitro and greenhouse experiments. Two of the transgenic lines, A1 and A3, showed the greatest resistance to PVY°infection, as indicated by low enzyme-linked immunosorbent assay values and infection rates. In vitro mechanical inoculation methods were also used to infect wild-type tomato and tobacco plants with cucumber mosaic virus and potato virus Y. Potato plants were also infected with the phloem-restricted potato leafroll virus, a low-titer virus, using in vitro graft inoculation methods. These results suggest the potential usefulness of these simple, effective, and economical techniques for screening large numbers of putative virus-resistant plants.

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Extreme resistance to two Brazilian strains of Potato virus Y (PVY) in transgenic potato, cv. Achat, expressing the PVYº coat protein

Horticultura Brasileira ◽

10.1590/s0102-05362001000200004 ◽

2001 ◽

Vol 19 (2) ◽

pp. 118-122 ◽

Cited By ~ 3

Author(s):

Eduardo Romano ◽

Adriana T. Ferreira ◽

André N. Dusi ◽

Karina Proite ◽

Jose A. Buso ◽

...

Keyword(s):

Coat Protein ◽

Potato Virus ◽

Potato Virus Y ◽

Potato Cultivar ◽

Transgenic Potato ◽

Resistant Line ◽

Cp Gene ◽

Transgenic Lines ◽

The Stability

The coat protein (CP) gene of the potato virus Y strain "o" (PVY O) was introduced into potato, cultivar Achat, via Agrobacterium tumefaciens-mediated transformation. Sixty three putative transgenic lines were challenged against the Brazilian strains PVY-OBR and PVY-NBR. An extremely resistant phenotype, against the two strains, was observed in one line, denominated 1P. No symptoms or positive ELISA results were observed in 16 challenged plants from this line. Another clone, named as 63P, showed a lower level of resistance. Southern blot analysis showed five copies of the CP gene in the extremely resistant line and at least three copies in the other resistant line. The stability of the integrated transgenes in the extreme resistant line was examined during several in vitro multiplications over a period of three years, with no modification in the Southern pattern was observed. The stability of the transgenes, the absence of primary infections and the relatively broad spectrum of resistance suggest that the extremely resistant line obtained in this work can be useful for agricultural purposes.

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The Resistance Responses of Potato Plants to Potato Virus Y Are Associated with an Increased Cellular Methionine Content and an Altered SAM:SAH Methylation Index

Viruses ◽

10.3390/v13060955 ◽

2021 ◽

Vol 13 (6) ◽

pp. 955

Author(s):

Nadezhda Spechenkova ◽

Igor A. Fesenko ◽

Anna Mamaeva ◽

Tatyana P. Suprunova ◽

Natalia O. Kalinina ◽

...

Keyword(s):

Potato Virus ◽

Potato Virus Y ◽

Potato Cultivar ◽

Potato Plants ◽

Methylation Index ◽

Different Temperatures ◽

Adenosyl Methionine ◽

Methionine Cycle ◽

Virus Interactions ◽

Methionine Content

Plant-virus interactions are frequently influenced by elevated temperature, which often increases susceptibility to a virus, a scenario described for potato cultivar Chicago infected with potato virus Y (PVY). In contrast, other potato cultivars such as Gala may have similar resistances to PVY at both normal (22 °C) and high (28 °C) temperatures. To elucidate the mechanisms of temperature-independent antivirus resistance in potato, we analysed responses of Gala plants to PVY at different temperatures using proteomic, transcriptional and metabolic approaches. Here we show that in Gala, PVY infection generally upregulates the accumulation of major enzymes associated with the methionine cycle (MTC) independently of temperature, but that temperature (22° C or 28° C) may finely regulate what classes accumulate. The different sets of MTC-related enzymes that are up-regulated at 22 °C or 28 °C likely account for the significantly increased accumulation of S-adenosyl methionine (SAM), a key component of MTC which acts as a universal methyl donor in methylation reactions. In contrast to this, we found that in cultivar Chicago, SAM levels were significantly reduced which correlated with the enhanced susceptibility to PVY at high temperature. Collectively, these data suggest that MTC and its major transmethylation function determines resistance or susceptibility to PVY.

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

Plant Disease ◽

10.1094/pd-90-0559 ◽

2006 ◽

Vol 90 (5) ◽

pp. 559-566 ◽

Cited By ~ 45

Author(s):

P. M. Baldauf ◽

S. M. Gray ◽

K. L. Perry

Keyword(s):

United States ◽

Resistance Gene ◽

Potato Virus ◽

Potato Virus Y ◽

Potato Cultivar ◽

Potato Virus X ◽

Enzyme Linked Immunosorbent Assay ◽

Northeastern United States ◽

Potato Virus S ◽

Tuber Necrosis

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.

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Monitoring Current-Season Spread of Potato virus Y in Potato Fields Using ELISA and Real-Time RT-PCR

Plant Disease ◽

10.1094/pdis-03-12-0283-re ◽

2013 ◽

Vol 97 (5) ◽

pp. 641-644 ◽

Cited By ~ 17

Author(s):

Manphool S. Fageria ◽

Mathuresh Singh ◽

Upeksha Nanayakkara ◽

Yvan Pelletier ◽

Xianzhou Nie ◽

...

Keyword(s):

Real Time ◽

Potato Virus ◽

Potato Virus Y ◽

Enzyme Linked Immunosorbent Assay ◽

Rt Pcr ◽

Pcr Assay ◽

Current Season ◽

Seed Market ◽

Polymerase Chain ◽

Time Of Harvest

The current-season spread of Potato virus Y (PVY) was investigated in New Brunswick, Canada, in 11 potato fields planted with six different cultivars in 2009 and 2010. In all, 100 plants selected from each field were monitored for current-season PVY infections using enzyme-linked immunosorbent assay (ELISA) and real-time reverse-transcription polymerase chain reaction (RT-PCR) assay. Average PVY incidence in fields increased from 0.6% in 2009 and 2% in 2010 in the leaves to 20.3% in 2009 and 21.9% in 2010 in the tubers at the time of harvest. In individual fields, PVY incidence in tubers reached as high as 37% in 2009 and 39% in 2010 at the time of harvest. Real-time RT-PCR assay detected more samples with PVY from leaves than did ELISA. A higher number of positive samples was also detected with real-time RT-PCR from growing tubers compared with the leaves collected from the same plant at the same sampling time. PVY incidence determined from the growing tubers showed a significant positive correlation with the PVY incidence of tubers after harvest. Preharvest testing provides another option to growers to either top-kill the crop immediately to secure the seed market when the PVY incidence is low or leave the tubers to develop further for table or processing purposes when incidence of PVY is high.

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Cytological responses of susceptible and extremely resistant potato plants to inoculation with potato virus Y

Physiological and Molecular Plant Pathology ◽

10.1006/pmpp.1999.0216 ◽

1999 ◽

Vol 55 (3) ◽

pp. 143-150 ◽

Cited By ~ 32

Author(s):

J. HINRICHS-BERGER ◽

M. HARFOLD ◽

S. BERGER ◽

H. BUCHENAUER

Keyword(s):

Potato Virus ◽

Potato Virus Y ◽

Potato Plants

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Arbuscular Mycorrhiza Changes the Impact of Potato Virus Y on Growth and Stress Tolerance of Solanum tuberosum L. in vitro

Frontiers in Microbiology ◽

10.3389/fmicb.2019.02971 ◽

2020 ◽

Vol 10 ◽

Cited By ~ 1

Author(s):

Edyta Deja-Sikora ◽

Anita Kowalczyk ◽

Alina Trejgell ◽

Adriana Szmidt-Jaworska ◽

Christel Baum ◽

...

Keyword(s):

Solanum Tuberosum ◽

Arbuscular Mycorrhiza ◽

Stress Tolerance ◽

Potato Virus ◽

Potato Virus Y ◽

Solanum Tuberosum L ◽

The Impact

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Epidemic of Potato virus Y and Cucumber mosaic virus in Henan Province Tobacco

Plant Disease ◽

10.1094/pdis.2001.85.4.447c ◽

2001 ◽

Vol 85 (4) ◽

pp. 447-447 ◽

Cited By ~ 1

Author(s):

X. D. Li ◽

Y. Q. Li ◽

H. G. Wang

Keyword(s):

Cucumber Mosaic Virus ◽

Mosaic Virus ◽

Potato Virus ◽

Potato Virus Y ◽

Early Stage ◽

Potato Virus X ◽

Henan Province ◽

Enzyme Linked Immunosorbent Assay ◽

Resistant Varieties ◽

Vein Necrosis

Flue-cured tobacco is an important crop in Henan Province, China. During the 2000 growing season, many tobacco plants showed various degrees of mottling, mosaic, vein clearing, or vein necrosis in most of the counties. Some plants even died at an early stage of growth. A survey was conducted in May-June in several tobacco-growing counties, and the incidence of symptomatic plants in individual fields ranged from 10 to 85%. The most widely planted tobacco varieties, NC89, K326, and K346, were highly susceptible. Symptomatic plants were collected from Jiaxian and Xiangcheng counties and samples were tested by enzyme-linked immunosorbent assay for Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), Potato virus Y (PVY), and Potato virus X (PVX). Of 65 samples tested, 21 were positive for only PVY, 16 positive for only CMV, one each was positive for only TMV or PVX. Nineteen samples were doubly infected with various combinations of these viruses and six were infected with combinations of three viruses. The causal agent(s) in the remaining sample could not be determined. In total, CMV was detected in 40 samples, PVY in 38, PVX in 10, and TMV in 7 samples. TMV and CMV used to be the most important viruses and PVY occurred only rarely. But PVY has become prevalent in Henan and in neighboring Shandong province (2). CMV and TMV were reported to be the most prevalent viruses in Shanxi (1) and Fujian Provinces (3). Because resistant varieties are not available, and mixed infections are more common, the results presented here explain why huge damage is occurring in tobacco crops in recent years. Some varieties are partially resistant to TMV and CMV but the varieties commonly grown are highly susceptible to PVY. Therefore, breeding for resistance to viruses, especially to PVY, is urgent to control the occurrence of tobacco viral diseases. References: (1) J. L. Cheng et al. Acta Tabacaria Sin. 4:43, 1998. (2) J. B. Wang et al. Chinese Tobacco Sci. 1:26, 1998. (3) L. H. Xie et al. Acta Tabacaria Sin. 2:25, 1994.

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Detection and genome characterization of Potato virus Y isolates infecting potato (Solanum tuberosum L.) in La Union (Antioquia, Colombia)

Agronomía Colombiana ◽

10.15446/agron.colomb.v34n3.59014 ◽

2016 ◽

Vol 34 (3) ◽

pp. 317-328 ◽

Cited By ~ 4

Author(s):

Pablo Gutiérrez S. ◽

Mauricio Marín M. ◽

Daniel Muñoz E.

Keyword(s):

Solanum Tuberosum ◽

Potato Virus ◽

Potato Virus Y ◽

Seed Tuber ◽

Rt Pcr ◽

Potato Plants ◽

Nextgeneration Sequencing ◽

Prevalent Strain ◽

Detailed Molecular Analysis

Potato virus Y (PVY) is one of the most severe viruses affecting the production of potato (Solanum tuberosum) in the world. This study presents a detailed molecular analysis using nextgeneration sequencing (NGS), IC-RT-qPCR and RT-PCR on the PVY isolates infecting seed-tubers and foliage of potato plants cv. Diacol-Capiro in La Union (Antioquia, Colombia). Analysis of incidence by IC-RT-qPCR in 15 random leaf samples of three cultivation plots and fifteen sprouting tuber eye-buds reveal infection levels between 13.4 and 80%; a higher incidence of 86.7% was observed in seed-tuber samples with threshold cycle (Ct) values as low as 24.3. Genome assembly from a bulk of foliage samples resulted in a consensus PVY genome (PVY_LaUnionF) of 9,702 nt and 399 polymorphic sites within the polyprotein ORF; while the assembled genome from sprouts of tubers has 9,704 nt (PVY_LaUnionT) and contained only six polymorphic nucleotide sites. Phylogenetic analysis demonstrates that the PVY isolates from leaf samples are in the recombinant PVYNTN group (sequence identity >99%); while those from tuber sprouts are in the PVYN/NTN group with identities above 95%. Sanger sequencing of viral capsid suggests the presence of a third variant related to PVYO, a prevalent strain reported in potato fields worldwide.

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