اثر عصاره های خام آبی و اتانولی گیاهان انجیر و زیتون تلخ بر بیماری ناشی از ویروس موزائیک خیار در گیاه خیار(Cucumis sativus) در شرایط گلخانه

نوع مقاله : مقاله پژوهشی

نویسندگان

گروه گیاهپزشکی، دانشکده علوم کشاورزی و صنایع غذایی، واحد علوم و تحقیقات دانشگاه آزاد اسلامی، تهران، ایران

چکیده

چکیده
زیتون تلخ (Melia azedarach) از خانواده Meliaceae و انجیر (Ficus carica)از خانواده Moraceae، گیاهان مهم دارویی بومی ایران هستند. در این تحقیق اثر عصاره­ های خام آبی و اتانولی گیاهان زیتون تلخ و انجیر در کاهش شدت بیماری ناشی از ویروس موزائیک خیار (Cucumber mosaic virus, CMV) در بوته­ های خیار با استفاده از آزمون ساندویچ دوطرفه الایزا (DAS-ELISA) و (Sq-RT-PCR) Semi quantitative-RT-PCR مورد سنجش قرار گرفت. همچنین از آزمون Sq-RT-PCR برای سنجش میزان نسخه برداری ژن PR-1 استفاده گردید و فعالیت اختصاصی آنزیم فنیل آلانین آمونیوم لیاز (PAL) سنجش شد. نتایج آزمون الایزا نشان داد میزان حضور CMV در گیاهچه­ های خیار، 1۵روز بعد از تیمار با غلظت­ های  ppm۱۰۰ عصاره اتانولی و ppm۱۰۰۰ عصاره آبی انجیر و زیتون تلخ در مقایسه با شاهد آلوده بدون تیمار به طور معنی­ داری کاهش پیدا کرده است. همچنین بیان ژن PR-1 پس از تیمار نمونه ­ها با عصاره اتانولی انجیر و زیتون تلخ در غلظت ppm 100 به طور معنی­ داری افزایش یافته و فعالیت اختصاصی آنزیم PAL در نمونه های تیمارشده با عصاره اتانولی انجیر و زیتون تلخ در غلظت ppm 100 تا 15 روز بعد از اضافه شدن عصاره در مقایسه با نمونه­ های آلوده تیمار نشده به طور معنی ­داری افزایش یافت و شدت بیماری ناشی از CMV در گیاهچه ­های خیار تیمار شده با عصاره اتانولی انجیر و زیتون تلخ در غلظت ppm 100 به طور معنی ­داری کاهش یافت. نتایج این بررسی در مجموع بیانگر فعال شدن احتمالی مقاومت سیستمیک اکتسابی در برابر CMV در گیاهچه­ های خیار از طریق سازوکارهای مختلف مولکولی و بیوشیمیایی توسط عصاره­ های الکلی گیاهان انجیر و زیتون تلخ مورد استفاده در این پژوهش می­ باشد.
 

کلیدواژه‌ها


عنوان مقاله [English]

The effect of Persian lilac and fig plant crude aquatic and ethanolic extracts on disease caused by Cucumber mosaic virus in cucumber (Cucumis sativus) plants under greenhouse condition

نویسندگان [English]

  • Maryam Sanjarian
  • Farshad Rakhshandehroo
  • Saeed Rezaee
Department of Plant Protection, College of Agricultural Sciences and Food Industries, Science and Research Branch, Islamic Azad University, Tehran, Iran.
چکیده [English]

Abstract
Persian lilac (Melia azedarach) of Meliaceae family and fig (Ficus carica) of Moraceae family are considered to be as important medicinal plants native to Iran. In this study, the antiviral effect of the crude ethanolic and aquatic extracts of the persian lilac and fig plants in decreasing disease severity caused by Cucumber mosaic virus (CMV) in cucumber plants was assessed using with the double antibody sandwich ELISA (DAS-ELISA) and Semi quantitative-RT-PCR (Sq-RT-PCR). In addition, the transcription level of pathogenesis related protein-1 (PR-1), resistance and specific activity of phenylalanine ammonia- lyase (PAL) enzyme were evaluated using Sq-RT-PCR assay. ELISA results showed that the presence level of CMV in cucumber seedlings was significantly decrease up to 15 days after their treating with the 100 ppm of ethanolic and 1000 ppm of aquatic extracts of fig and persian lilac plants in compare to the non-treated control plants. The PR-1 expression level and PAL enzyme specific activity were also significantly increased compared to the non-treated control infected plants in samples treated with the ethanolic extracts of persian lilac and fig plants at the concentration of 100 ppm up to 15 days after treatments. Disease severity was also significantly decreased in cucumber seedlings treated with the 100 ppm concentration of ethanolic extract of the persian lilac and fig plants. Totally, results of this study indicated that the systemic acquired resistance might be activated against CMV by applying ethanolic extracts of persian lilac and fig plant used in this study in treated cucumber seedlings through different molecular and biochemical mechanisms.
 

کلیدواژه‌ها [English]

  • Keywords: Antiviral activity
  • DSI
  • PR-1 gene
  • PAL enzyme
  • Systemic resistance
  • Sq-RT-PCR
References
 
Ahmad J, Khan I, Khan S, Iqbal D, 2013. Evaluation of antioxidant and antimicrobial activity of Ficus Carica leaves: an in vitro approach. Journal of Plant Pathology and Microbiology 4: 2–4.
Alche LE, Ferek GA, Meo M, Coto CE, Maier MS, 2003. An antiviral meliacarpin from leaves of Melia azedarach L. Zeitschrift fur Naturforschung 58: 215–219.
Ali S, Ganaib BA, Kamilib AN, Bhatc AA, Mira ZA, et al., 2018. Pathogenesis-related proteins and peptides as promising tools for engineering plants with multiple stress tolerance. Microbiological Research 212–213: 29–37.
Andrei G, Couto A.S, De Lederkramer RM, Coto CE, 1994. Purification and partial characterization of an antiviral active peptide from Melia Azedarach L. Antiviral Chemistry and Chemotherapy 5(2): 105–110.
Arafati N, Farzadfar Sh, Pourrahim R, 2013.  Characterization of coat protein gene of Cucumber Mosaic Virus isolates in Iran. Iranian Journal of Biotechnology 11: 109–114.
Asl-Najjari AH, Rajabi Z, Vasfi-Marandi M, Dehghan Gh, 2015.The effect of the hexanic extracts of fig (Ficus carica) and olive (Olea europaea) fruit and nanoparticles of selenium on the immunogenicity of the inactivated avian influenza virus subtype H9N2. Veterinary Research Forum 6: 227– 231.
Bradford MM, 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Annals of Biochemistry 72: 248–254.
Bashir NS, Nematollahi S, Torabi E, 2008. Cucumber mosaic virus subgroup IA frequently occurs in the northwest IRAN. Acta Virologica 52: 237–242.
Bashir NS, Kolhar MR, Zarghani SN, 2006. Detection, differentiation and phylogenetic analysis of cucumber mosaic virus isolates from cucurbits in the northwest region of Iran. Virus Genes 32: 277–288.
Boller TV, 1986. Roles of proteolytic enzymes in interaction of plants with other organisms. In: Dalling, M.J. (Ed.), Plant Proteolytic Enzymes, 1. CRC Press, Boca Raton, FL.  Pp: 67–96.
Clark MF, Adams AN, 1977. Characteristics of the microplate method of enzyme-linked immunosorbent assay for the detection of plant viruses. Journal of General Virology 34: 475–483.
Collins P, 1983. The spectrum of antiviral activities of acyclovir in vitro and in vivo. The Journal of Antimicrobial Chemotherapy 12: 19–27.
Dehghan S, Sadeghi M, Poppel A, Fischer R, Lakes-Harlan R, et al., 2014. Differential inductions of phenylalanine ammonia-lyase and chalcone synthase during wounding, salicylic acid treatment, and salinity stress in safflower, Carthamus tinctorius. Bioscience Reports 34: 273–82.
Del Toro FJ, Aguilar E, Hernández-Walias FJ, Tenllado F, Chung BN, et al., 2015.  High temperature, high ambient CO2 affect the interactions between three positive-sense RNA viruses and a compatible host differentially, but not their silencing suppression efficiencies.  PLoS One, 10:e0136062. DOI:10.1371/journal. pone. 0136062.
Edwaron JR, Christie RG, 1991.  CRC Handbook of Viruses Infecting Legumes. CRC Press, Boca Raton, University of Florida, USA. 293 pp.
Helal IM, 2019. Use of biocides for controlling viral diseases that attack common bean and cucumber plants. Folia Horticulturae31: 159–170.
Elsharkawy MM, Shimizu M, TakahashiH, Ozaki K, Hyakumachi M, 2013. Induction of systemic resistance against cucumber mosaic virus in Arabidopsis thaliana by Trichoderma asperellum SKT-1. Plant Pathology Journal 29: 193–200.
Eyvazi A, Dizaji A, Rastgou M, Koohi-Habibi M, 2015. Bioassay and phylogeny of five Iranian isolates of Cucumber mosaic virus from different hosts based on CP gene sequence. Plant Protection Science 51: 200–207.
Goel N, Anukrati K, Paul PK, 2016. Anti-phytopathogenic and SAR inducing properties of Neem: a review. Journal of Chemical and Pharmaceutical Sciences 9: 2544–2555.
Goodman RN, Kiraly Z. Wood KR, 1986. The Biochemistry and Physiology of Plant Disease. USA, University of Missouri Press. Pp. 433–438.
Hernández-Herrera RM, Santacruz-Ruvalcaba F, Ruiz-Lopez MA, Norrie J, Hernández-Carmona G, 2014. Effect of liquid seaweed extracts on growth of tomato seedlings (Solanum lycopersicum L.). Journal of Applied Phycology 26: 619–628.
Jing B, Ma Z, Feng J, Liang H, Li C, Zhang X, 2012. Evaluation of the antiviral activity of extracts from plants grown in the Qinling region of china against infection by tobacco mosaic virus (TMV). Journal of Phytopathology 160: 181–186.
 Jones RCA, Naidu R, 2020. Global dimensions of plant virus diseases: current status and future perspectives. Annual Review of Virology 6: 387-409.
Jones IK, Glazzer AN, 1970. Comparative studies on four sulfhydryl endopeptidases (Ficins) of Ficus glabrata latex. Journal of Biological Chemistry 245: 2765– 2772.
Mahmoud SYM, Gad-Rab SMF, Hussein N, Shoreit AAM, 2010.  Antiviral Activity of Latex from Ficus nitida Against Plant Viruses. Global Journal of Biotechnology and Biochemistry 5: 198–205.
Marone M, Mozzetti S, Ritis D, Pierelli L, Scambia G, 2001. Semiquantitative RT-PCR analysis to assess the expression levels of multiple transcripts from the same sample. Biological Procedures Online 3: 19–25.
Maisaro M, Sugiharto B, Dewanti P, 2017. The effect of concentration and exposure time Acyclovir for elimination sugarcane mosaic virus (SCMV) on the apical bud culture of sugarcane PS 881. Journal ILMU Dasar 18: 31–38.
Mawa S, Husain K, Jantan I, 2013. Ficus carica L. (Moraceae): Phytochemistry, Traditional Uses and Biological Activities. Evidence-Based Complementary and Alternative Medicine 3: 1–8. doi: 10.1155/2013/974256.
Medini F, Fellah H, Ksouri R, Abdelly C, 2014. Total phenolic, flavonoid and tannin contents and antioxidant and antimicrobial activities of organic extracts of shoots of the plant Limonium delicatulum. Journal of Taibah University for Science 8: 216–224.

Naserinasab F, Heydari R, Sanjarian F, Rakhshandehroo F. 2018. Investigation of the defense genes expression of Phenylalanine Ammoniumase and Peroxidase in interaction with Neem extract of and Meloidogyne javanica nematodes in tomato. Journal of Cell &Tissue 9: 360–377.

Palukaitis P, García-Arenal F, 2018. Cucumber Mosaic Virus. APS Press; St. Paul, MN, USA.
Palukaitis P, Garcia- Arenal F, 2003. Cucumoviruses. Advances in Virus Research 62: 241–323.
Petrov N, Stoyanova M, Valkova M, 2016. Antiviral activity of plant extract from Tanacetum vulgare against Cucumber Mosaic Virus and Potato Virus Y. Journal of Bioscience and Biotechnolology 5: 189–194.
Rakhshandehroo F, Takeshita M, Squires J, Palukaitis P, 2009. The Influence of RNA-Dependent RNA Polymerase 1 on Potato virus Y Infection and on Other Antiviral Response Genes. Molecular Plant-Microbe Interactions 22: 1312–1318.
Sels J, Mathys J, De Coninck BMA, Cammue BPA, De Bolle MFC, 2008. Plant pathogenesis-related (PR) proteins: A focus on PR peptides. Plant Physiology and Biochemistry 46: 941–950.
Sharma S, Singh B, Rani G, Zaidi AA, Hallan V, et al., 2007. Production of Indian citrus ringspot virus-free plants of Kinnow mandarin (Citrus nobilis Lour ×C. deliciosa Tenora) employing chemotherapy coupled with shoot tip grafting. Journal of Central European Agriculture 8: 1–8
Singh B, 2015. Effect of antiviral chemicals on in vitro regeneration response and production of PLRV-free plants of potato. Crop Science and Biotechnology 18: 341–348.
Spencer WE, Christensen MJ, 1999. Multiplex relative RT-PCR method for verification of differential gene expression. BioTechniques 27:1044–1052.
Sung KD, Kook-Hwang B, 2014. An important role of the pepper phenylalanine ammonia-lyase gene (PAL1) in salicylic acid-dependent signaling of the defence response to microbial pathogens. Journal of Experimental Botany 65: 2295– 2306.
Van Loon LC, Geraats BPJ, Linthorst HJM, 2006. Ethylene as a modulator of disease resistance in plants. TrenDSI in Plant Science 11:184–191.

Vijayanand S, Wesely EG, 2014.Antimicrobial and antioxidant activity of Melia azadirachta and Murray koenigil. International Journal of Research in Pharmacy and Science 5: 1022–1028.

Vuko E, Rusak G, Dunki´ V, Kremer D, Kosalec I, et al., 2019. Inhibition of Satellite RNA Associated Cucumber Mosaic Virus Infection by Essential Oil of Micromeria croatica (Pers.) Schott. Molecules. 24(7):1342.
Yan W, Zhao M, Ma Y, Pan YH, Yuan WX, 2011.   Primary purification of two antifungal proteins from leaves of the fig (Ficus carica L.). Africican Journal of  Biotechnology 10: 375–379.
Yang XM, Yu W, Ou ZP, Ma H, Liu WM, et al., 2009.Antioxidant and Immunity Activity of Water Extract and Crude Polysaccharide from Ficus carica L. Fruit Plant. Plant FooDSI for Human Nutrition 64: 167–173.
Zeng R, Liao Q, Feng J, Li D, Chen J, 2007. Synergy between Cucumber Mosaic Virus and Zucchini Yellow Mosaic Virus on Cucurbitaceae hosts tested by Real-time Reverse Transcription-Polymerase chain reaction. Acta Biochimica et Biophysica Sinica 39: 431–437.