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  3. Identifying and determining the strains of Potato Y virus (PVY) of molecular hybrids in Golestan province

Identifying and determining the strains of Potato Y virus (PVY) of molecular hybrids in Golestan province

Number of pages: 67 File Format: word File Code: 32480
Year: 2012 University Degree: Master's degree Category: Agricultural Engineering
Tags/Keywords: PCR - Plant diseases - potato - Potato virus Y - Potato virus Y - Potato wow virus - PVY - virus
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  • Summary of Identifying and determining the strains of Potato Y virus (PVY) of molecular hybrids in Golestan province

    Dissertation for Master's Degree

    Plant Pathology Department

    Abstract
    One of the limiting factors for potato cultivation in Golestan province is plant diseases, the most important of which are viruses, and among them, PVY (Potato virus Y) It is the most destructive of them. This virus infects many species of the Solanaceae family, plus more than 60 other plant species. The symptoms of this virus are completely different depending on the strain and variety of potato. In general, PVY with three breeds PVYN, PVYC, PVYO was reported in Iran. In order to determine the strains of this virus, 170 samples were taken from 80 potato farms in Golestan province (Galikesh, Minodasht, Azadshahr, Ramian, Deland, Qaraq, Fazelabad, Aliabad, Gorgan and Kordkoi) in 2013. The samples were tested by indirect serological ELISA method with PVY antiserum, and the samples that were positive in this test were inoculated on the reference plant, and some of these positive samples were captured with the Capture mRNA kit. The RNA trapped in the reverse transcription reaction was converted into cDNA. In order to amplify the produced cDNA, the PCR reaction was performed. The final PCR product was sent to South Korea's Macrogen Company for synonymy determination. These synonyms were aligned and compared with the synonymy of other PVY isolates in the gene bank by Clustal X program, and the nucleotide and amino acid similarity between them was calculated in MegAlign program. The phylogenetic tree obtained by aligning synonyms by MEGA5 program was obtained by Neighbor-joining method with Kimura-2 method and evaluated by Treeview software. The results of this research showed that out of 170 samples, the number of samples were infected with the virus, and the three strains PVYN, PVYO, PVYWilga are reported for the first time in this province on the mentioned host. Also, the phylogenetic analysis of the CP region of the isolates investigated in this research showed that the PVYN race with 64.03% of distribution was prevalent as the dominant race in the potato fields of Golestan province, but the PVYC race was not observed in this province. Also, the results showed that Kurdkovi region with 54.8% of the frequency of contamination showed the highest amount of PVY contamination among other regions in the crop year 2019. The results of comparing the synonyms of the coat protein gene region showed that Gorgan and Azadshahr isolates were more similar to each other (99-97%), but Minodasht, Galikesh, Kordkoi isolates were included in the Aliabad subgroup. The isolates reported from Kerman are grouped with Golestan isolates, which indicates a close phylogenetic relationship between them. After that, isolates from Khorasan, Isfahan, Ardabil, and Shiraz (Bushehr race) were closer. The isolates from Zanjan and Ardabil are placed in a separate group that is more similar to the isolate from Ardabil. In the dendrogram related to the comparison of Iranian isolates with other countries of the world, the results showed that Golestan isolates were in the same group as Kerman isolates, which were similar to German, Canadian and American isolates. which indicates a high genetic relationship (94% genetic similarity). Also, the results showed that the genetic diversity between the isolates obtained by maximum likelihood method 1 in BioEdit software, the ratio of transition substitution to crossover substitution 2 is equal to 1.08. Therefore, it can be said that the mutations in the genome of the mentioned virus follow a certain process.

    Introduction:

    Viruses are very small organisms that cannot be seen with ordinary (light) microscopes that reproduce only in living cells. About 50 years ago, the first viruses were isolated and their chemical properties were determined. They were infectious plant viruses. At the same time, many debates were raised regarding the belonging of these factors to the living or non-living world. During the last 30 years, with the help of the application of molecular biology concepts and methods, it has become clear that viruses are part of the living world. In fact, viruses are very small obligate parasites that have one to several hundred genes. These genes, like cellular genes, can mutate and evolve (Pourrahim et al., 2018). The Potyviridae family is one of the most important plant pathogenic virus groups and one of the largest among them.Among the potyviruses[1], the potato virus (PVY)[2] is considered as one of the most destructive pathogens of tobacco, potato, tomato and pepper). So that in some cases, according to the virus strain, the number of potatoes and the weather conditions of the region, Y potato virus can cause between 40-70% damage (Beemster & De Bokx. 1987). Based on the reaction of the representative plants and the power of transmission by the aphid, potato virus Y has been divided into three racial groups (PVYo, PVYn, PVYc) (De bokx & Huttinga, 1981). In addition to the difference in the reaction of a number of representative hosts, aphids transmit PVYo and PVYn strains through nonpersistent, while PVYc group strains are not transmitted by aphids (Beemster & De Bokx, 1987). In recent years, the most research activities in the field of controlling viral diseases include breeding safe or resistant cultivars with classic genetic methods, controlling vectors using different strategies, producing virus-free seeds and vegetative organs, and producing tragenic plants that carry viral genes and cause resistance to the virus (Porrahim et al., 2012). The basis of any selection or breeding program is the existence of genetic diversity among plants of a species in response to a specific virus, and it can be used in the case of many plant viruses that are of economic importance (Jafarpour, 1382). Potato is one of the most important crops in the world, so that its production in 2000 was about 321 million tons of tubers from 6 million hectares. Potato is one of the most important crops among the world's crops. The nutritional, social and economic aspect of this important plant has been interesting and significant in recent years (1985). It has been cultivated in Iran for more than 200 years. The origin of the potato is in South and Central America. The heights of the Andes, Bolivia, Peru and Chile were the first areas where the potato was cultivated. The potato was first brought to Europe by the Spanish and through Spain and Ireland to other parts. Potatoes were introduced to Iran for the first time during the time of Feth Ali Shah Qajar. Potatoes are cultivated in all countries of the world. Today, potatoes are grown in all countries of the world. This plant is one of the tuber products that plays an important role in feeding the people of the world. It is protein and some vitamins and minerals, there are a lot of vitamins C, B1, B2 and A in potatoes. Another substance that is present in potatoes is solanine, which is a poisonous substance. The amount of the poisonous substance solanine is different compared to different cultivars. Today, in Iran, the provinces of Ardabil, Isfahan, Hamedan, East Azerbaijan, Khorasan and Fars are the most important producers of water potatoes, and the provinces of Mazandaran, Gilan, Golestan are the most important for rainfed cultivation. Solanaceae, which is cultivated to use its underground tuber. The genus Solanum has many species, but only the tuberosum species and a very limited number of others produce tubers. The aerial part of this plant dries up and disappears due to the cold of winter. But the remaining tubers in the soil can sprout and produce a new plant in the spring of the following year. Therefore, potato is a permanent plant in terms of underground stems at the end of which tubers are formed; But in terms of aerial stems and leaves, it is annual, the height of the herbaceous but standing potato plant reaches 60 to 150 cm, and its growth period is 3 to 6 months. Potatoes are characterized by the presence of an underground creeping stem 5 to 45 cm long, which originates from the lower nodes and underground of the aerial stems. These underground creeping stems have small, scaly leaves. And if it is exposed to light, it turns into aerial stems. The top of each creeping underground stem swells and turns into a potato tuber

  • Contents & References of Identifying and determining the strains of Potato Y virus (PVY) of molecular hybrids in Golestan province

    List:

    Abstract 1

    Introduction: 4

    1-1- Potato. 5

    1-1-1- The history of potatoes. 5

    1-1-2- Botany. 6

    1-1-3- Temperature: 8

    1-1-4- Soil and fertilizer 9

    1-1-5- The economic importance of potatoes. 10

    1-2-1- Symptoms of potato virus diseases. 11

    1-2-2- Epidemiology and disease cycle. 11

    1-2-3- control of potato virus diseases. 12

    1-3- Hypotheses 12

    1-4- Objectives. 12

    The second chapter. 13

    2-1- Features of the group of potyviruses 14

    2-1-1- Potato W virus. 15

    2-1-1-1- Host range and symptoms of potato blight virus. 16

    2-1-1-2- The status of potato blight virus in Iran. 17

    2-1-1-3- Potato blight virus carriers. 18

    2-1-1-4-genetic structure. 19

    2-1-1-5-races of potato virus. 19

    2-1-1-5-2-race N. 20

    2-1-1-5-3-race C. 21

    2-1-1-6-methods for investigating PVY races: 21

    2-1-1-7-recombination in the potato virus y genome. 22

    2-1-1-8-Comparison of two common methods of identifying PVY. 23

    The third chapter. 26

    3-1- Sampling of potato fields. 27

    3-2- Plant materials. 28

    3-2-1- Storage of samples in the laboratory 28

    3-3- Indirect ELISA test. 29

    3-5- Selection of positive isolates. 30

    3-6- Greenhouse studies. 30

    3-6-1- Preparation of greenhouse plants. 31

    3-6-2- Germination of greenhouse plants. 31

    3-7- RT-PCR test method. 33

    3-7-1 cDNA preparation method. 33

    3-7-3 stages of RNA virus trapping. 34

    3-9- Performing the PCR reaction. 35

    3-9-1- Primer design 35

    3-9-2 Polymerase chain reaction (PCR) test method 36

    3-10 Gel preparation method and performing electrophoresis to check PCR test results. 37

    3-11- Comparing the nucleotide sequence of PVY breeds with the same sequence in other isolates in the gene bank. 38

    Chapter Four: 42

    4-1- The source of the virus. 43

    4-2- Indirect ELISA test results. 44

    4-3 greenhouse studies. 46

    4-5- RT-PCR test results. 47

    4-6- Genetic diversity. 52

    4-7- The highest percentage of similarity of the studied isolates with the main PVY breeds in the world: 52

    4-7- Comparison of the results of indirect ELISA and RT-PCR tests. 54

    The fifth chapter. 55 5-1-Recombination investigation in Iranian isolates of Y potato virus: 58 5-2- Implementation and research suggestions. 59

    List of sources and references. 61

    English sources: 64

    Source:

     

    Ahmadikhah, A. 2017. Complementary genetics, publications of Gorgan University of Agricultural Sciences and Natural Resources. 350 pages

    Aizdpanah, K., Ashkan, M., Bani Hashemi, Z., Rahimian, H. and Minasian, V. 1389. In translation of plant pathology. (Agrios, George N). Tehran. Ayez. 1026 pages

    Binam. 2017. Statistical record of the General Department of Agricultural Affairs of Iran Tobacco Company. 63 pages.

    Pour Rahim, R., Farzadfar, Sh. and Glanraghi, A. 1381. In the translation of the basics of plant virology (Matheos, R. A. F.). Saman Pishehgar Co. 457 pages.

    Pourrahim, R., Shahrayin, N., Ahunmanesh, A. and Karimi, R. 2016. Report of Necrotic strain of Y potato virus from potato farms in Hamadan province. Journal of Plant Pathology. Volume 34 (2-1). Pages 37-42.

    Jafarpour, B., Jafarpour, B. 2012. Applied plant virology (translation). Publications. Ferdowsi University of Mashhad. 460 pages.

    Khakour, R., Shamsbakhsh, M. and Pourrahim, R. 2014. Status of six important pathogenic viruses in potato fields of Khuzestan province. Journal of Plant Pests and Diseases, Volume 73, Number 1, Pages 25-37.

    Khateri, H., Marafzadeh, N., Hosseini, A. and Mahtabi, R. A. 2015. Investigating the response of different tobacco cultivars to the potato virus under natural contamination conditions in Tirtash region of Mazandaran. Summary of articles of the 9th Iranian Genetics Congress, Tehran. 167 pages.

    Khodabandeh, N. 1368. Industrial plants. Tehran University Publications. 454 pages.

    Khajepour, M.R. 1383. Industrial plants. Academic Jihad of Isfahan Industrial Unit. 480 pages.

    Khoshkhati, N., Kohi Habibi, M. and Mosahibi Mohammadi, G.H. 2008. Occurrence of potato virus Y strain N (PVY) in celery fields in Tehran province. Summary of the fifth articles. Summary of the articles of the 5th Iranian Congress of Virology. 9 pages.

    Danesh, D., Soleimani, S., Philosopher, F. and Dehghan, M. 1370. Occurrence of 4 potato viruses in Isfahan province. Journal of Plant Pathology. Volume 28, pages 49-45.

    Danesh, M. 1379. Tobacco diseases (translation). Academic Publishing Center, Tehran. 235 pages.

    Shams Bakhsh, N., Khakur, R. and Pourrahim, R. 1381. Purification and preparation of antiserum and serological identification of Y and X viruses isolated from potato fields in Khuzestan province. Scientific journal of agriculture. Volume 24: Pages 35-47.

    Shahrayin, N. and Elahina, S.A. 2008. Serological and immuno-electron microscopic studies of potato virus Y (PVY) isolated from tobacco plant cultivar Koker 347 in Gilan region. Journal of Plant Pests and Diseases, Volume 67, Number 1 and 2. Pages 17-27.

    Tusi, N., Ahunmanesh, A., Pourahim, R. and Bahar, M. 2013. Identification of C and N strains of potato Y virus using Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) and Restriction Fragment Length Polymorphism (RFLP). Summary of articles of the 16th Iran Herbal Medicine Congress. 224 pages.

    Tusi, N. 2016. Determining potato W virus strains using serological and molecular methods, master's thesis in plant pathology, Isfahan University of Technology. 100 pages.

    Mujdabadi Farahani, S., Jafarpour, B., Falahati Rastgar, M., and Sabek Khiz, M. A. 2018. Tracking and identification of the NTN strain of potato virus from the farms of Razavi Khorasan province. Summary of the articles of the 5th Iranian Congress of Virology. 160 pages.

    Marafzadeh, N., Khatiri, H., Kohi Habibi, M., Mosahebi Mohammadi, G. H., Hamzeh, N., Ghazanfari, Kivan., and Hosseini, A. 2015. The status of contamination of commercial tobacco cultivars with potato y virus and identification of the necrotic strain of the virus in tobacco fields in Mazandaran and Golestan provinces. Summary of articles of the 17th Congress of Iranian Medicinal Plants. 225 pages.

    Marafzadeh, N., Khatiri, H., Kohi Habibi, M., Mosahebi Mohammadi, G. H. and Hamza, N. 2017. Identification of the dominant strain of potato y virus in commercial tobacco cultivars of Golestan, Mazandaran, Gilan and West Azarbaijan provinces. Summary of articles of the 18th Iran Herbal Medicine Congress. 474 pages.

    Maqsoudi, R., Jafarpour, B. and Falahati Rostgar, M. 2013. Occurrence of N strain of potato Y virus (PVYN) in Khorasan province and O strain of this virus (PVYO) in Khorasan, Gilan and Mazandaran provinces. Summary of articles of the 16th Iran Herbal Medicine Congress. 223 pages.

    Mousavi, L., Mozafari, J., Rakhshandaro, F., Gadmiari, S. and Sokhdan, N. 2018. Molecular diagnosis of potato Y virus strains using RT-PCR in East Azarbaijan province. Summary of articles of the 5th Congress of Virology of Iran. 188 pages.

     

     

    English sources:

    Abedi, H. 1998. Appearance of PVY in tobacco fields of Iran (Regions of Mazandaran and Gorgan). Bull. Sp. Coresta, Brighton Congress. p.24.

     

    Bagnall, R.H. 1992. Epidemiology Potato virus Y in New Brunswick, Canada- A biennial rhythm and 9 plus year cycle. Can. J. Plant Pathology. 14: 137-146.

     

    Bapat, V. A., Ghosh, S. B., Nagi, L.H.S., Ganapathi, T. R., Khurana, S.M.P. 2002. Cloning and sequencing of potato virus Y coat protein gene from an Indian isolate and development of transgenic tobacco for PVY resistance. Current. Science (Indian). v.82 (7) P.855-859.

     

    Barnett, O. W. 1992, A summary of potyvirus taxonomy and definitions. Arch. Virology. Supplementum. 5: 435-444.

    Bawden, F.C. 1936. The viruses causing top necrosis (acronecrosis) of the Potato. Ann. Appl. Biol 23: 487-497.

     

    Blanco-Urgoiti, B., Tribodet, M., Leclere, S., Ponz, F., Perez de San Roman, C., Legorburu, F.J., and Kerlan, C. 1998. Characterization of Potato Y (PVY) isolates from potato seed batches. Situation of the NTN, Wilga and Z isolates. Eur. J. Plant Pathol. 104: 811-819.      

     

    Boonham, N., Walsh, K., Hims, M., Preston, S., North, J. and Barker, I.

Identifying and determining the strains of Potato Y virus (PVY) of molecular hybrids in Golestan province
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