Theoretical and Applied Genetics

, Volume 111, Issue 4, pp 711–722 | Cite as

Genetic modification of potato against microbial diseases: in vitro and in planta activity of a dermaseptin B1 derivative, MsrA2

  • Milan Osusky
  • Lubica Osuska
  • William Kay
  • Santosh MisraEmail author
Original Paper


Dermaseptin B1 is a potent cationic antimicrobial peptide found in skin secretions of the arboreal frog Phyllomedusa bicolor. A synthetic derivative of dermaseptin B1, MsrA2 (N-Met-dermaseptin B1), elicited strong antimicrobial activities against various phytopathogenic fungi and bacteria in vitro. To assess its potential for plant protection, MsrA2 was expressed at low levels (1–5 μg/g of fresh tissue) in the transgenic potato (Solanum tuberosum L.) cv. Desiree. Stringent challenges of these transgenic potato plants with a variety of highly virulent fungal phytopathogens—Alternaria, Cercospora, Fusarium, Phytophthora, Pythium, Rhizoctonia and Verticillium species—and with the bacterial pathogen Erwinia carotovora demonstrated that the plants had an unusually broad-spectrum and powerful resistance to infection. MsrA2 profoundly protected both plants and tubers from diseases such as late blight, dry rot and pink rot and markedly extended the storage life of tubers. Due to these properties in planta, MsrA2 is proposed as an ideal antimicrobial peptide candidate to significantly increase resistance to phytopathogens and improve quality in a variety of crops worldwide with the potential to obviate fungicides and facilitate storage under difficult conditions.


Transgenic Plant Late Blight Phytophthora Transgenic Potato Transgenic Potato Plant 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



We thank X. Yu, B. Forward, T. Stevenson and B. Allen for expert technical assistance, Dr. Zamir Punja (Simon Fraser University, Burnaby, B.C., Canada) and Dr. Harold Platt (Agriculture & Agri-Food Canada) for providing fungal pathogens. This work was funded by a grant to SM from the Canadian Bacterial Diseases Network.


  1. Asai T, Tena G, Plotnikova J, Willmann MR, Chiu W-L, Gomez-Gomez L, Boller T, Ausubel FM, Sheen J (2002) MAP kinase signaling cascade in Arabidopsis innate immunity. Nature 415:977–983Google Scholar
  2. Baghian A, Jaynes J, Enright F, Kousolas KG (1997) An amphipathic alpha-helical synthetic peptide analogue of melittin inhibits herpes simplex virus-1 (HSV-1)-induced cell fusion and virus spread. Peptides 18:177–183CrossRefPubMedGoogle Scholar
  3. Banzet N, Latorse M-P, Bulet P, Francois E, Derpierre C, Dubald M (2002) Expression of insect cysteine-rich antifungal peptides in transgenic tobacco enhances resistance to a fungal disease. Plant Sci 162:995–1006CrossRefGoogle Scholar
  4. Belaid A, Aouni M, Khelifa R, Trabelsi A, Jemmali M, Hani K (2002) In vitro antiviral activity of dermaseptins against herpes simplex virus type 1. J Med Virol 66:229–234CrossRefPubMedGoogle Scholar
  5. Biezen E van der (2001) Quest for antimicrobial genes to engineer disease-resistant crops. Trends Plant Sci 6:89–91CrossRefPubMedGoogle Scholar
  6. Boyd AEW (1972) Potato storage diseases. Rev Plant Pathol 51:297–321Google Scholar
  7. Campbell MA, Fitzgerald HA, Ronald PC (2002) Engineering pathogen resistance in crop plants. Transgenic Res 10:1–15Google Scholar
  8. Chakrabarti A, Ganapathi TR, Mukherjee PK, Bapat VA (2003) MSI-99, a magainin analogue, imparts enhanced disease resistance in transgenic tobacco and banana. Planta 216:587–596Google Scholar
  9. Chernysh S, Kim SI, Bekker G, Pleskach VA, Filatova NA, Anikin VB, Platonov VG, Bulet P (2002) Antiviral and antitumor peptides from insects. Proc Natl Acad Sci USA 99:12628–12632CrossRefPubMedGoogle Scholar
  10. Cohn J, Sessa G, Martin GB (2001) Innate immunity in plants. Curr Opin Immunol 13:55–62CrossRefPubMedGoogle Scholar
  11. Coote PJ, Holyoak CD, Bracey D, Ferdinando DP, Pearce JA (1998) Inhibitory action of the amphibian skin peptide dermaseptin S3 on S. cerevisiae. Antimicrob Agents Chemother 42:2160–2170PubMedGoogle Scholar
  12. Dagan A, Efron L, Gaidukov M, Mor A, Ginsburg H (2002) In vitro antiplasmodium effects of dermaseptin S4 derivatives. Antimicrob Agents Chemother 46:1059–1066CrossRefPubMedGoogle Scholar
  13. Dangl JL, Jones JDG (2001) Plant pathogens and integrated defence responses to infection. Nature 411:826–833CrossRefPubMedGoogle Scholar
  14. Datla RSS, Bekkaoui F, Hammerlindl JK, Pilate G, Dunstan DI, Crosby WL (1993) Improved high-level constitutive foreign gene expression in plants using an AMV RNA4 untranslated leader sequence. Plant Sci 94:139–149CrossRefGoogle Scholar
  15. De Block M (1988) Genotype-independent leaf disc transformation of potato (Solanum tuberosum) using Agrobacterium tumefaciens. Theor Appl Genet 76:767–774CrossRefGoogle Scholar
  16. De Gray G, Rajasekaran K, Smith F, Sanford J, Daniell H (2001) Expression of an antimicrobial peptide via the chloroplast genome to control phytopathogenic bacteria and fungi. Plant Physiol 127:852–862CrossRefPubMedGoogle Scholar
  17. De Lucca AJ, Bland JM, Jacks TJ, Grimm C, Walsh TJ (1998) Fungicidal and binding properties of the natural peptides cecropin B and dermaseptin. Med Mycol 36:291–298CrossRefPubMedGoogle Scholar
  18. Epple P, Apel K, Bohlmann H (1997) Overexpression of an endogenous thionin enhances resistance of Arabidopsis against Fusarium oxysporum. Plant Cell 9:509–520CrossRefPubMedGoogle Scholar
  19. Fagoaga C, Rodrigo I, Conejero V, Hinarejos C, Tuset JJ, Arnau J, Pina JA, Navarro L, Pena L (2001) Increased tolerance to Phytophthora citrophthora in transgenic orange plants expressing a tomato pathogenesis related protein PR-5. Mol Breed 7:175–185CrossRefGoogle Scholar
  20. Fischer KS, Barton J, Khush GS, Leung H, Cantrell R (2000) Collaboration in rice. Science 290:279–280CrossRefPubMedGoogle Scholar
  21. Fleury Y, Vouille V, Beven L, Amiche M, Wroblewski H, Delfour A, Nicolas P (1998) Synthesis, antimicrobial activity and gene structure of a novel member of the dermaseptin B family. Biochim Biophys Acta 1396:228–236PubMedGoogle Scholar
  22. Frye CA, Tang D, Innes RW (2001) Negative regulation of defense responses in plants by a conserved MAPKK kinase. Proc Natl Acad Sci USA 98:373–378CrossRefPubMedGoogle Scholar
  23. Gao AG, Hakimi SM, Mittanck CA, Wu Y, Woerner BM, Stark DM, Shah DM, Liang JH, Rommens CMT (2000) Fungal pathogen protection in potato by expression of a plant defensin peptide. Nat Biotechnol 18:1307–1310CrossRefPubMedGoogle Scholar
  24. Guillemot D (1999) Antibiotic use in human and bacterial resistance. Curr Opin Microbiol 2:494–498Google Scholar
  25. Hancock REW, Diamond D (2000) The role of cationic antimicrobial peptides in innate host defences. Trends Microbiol 8:402–410CrossRefPubMedGoogle Scholar
  26. Hancock REW, Scott MG (2000) The role of antimicrobial peptides in animal defense. Proc Natl Acad Sci USA 97:8856–8861CrossRefPubMedGoogle Scholar
  27. Hancock REW, Falla T, Brown M (1995) Cationic bactericidal peptides. Adv Microbiol Physiol 37:135–175Google Scholar
  28. Hernandez C, Mor A, Dagger F, Nicolas P, Hernandez A, Bernadetti EL, Dunia I (1992) Functional and structural damages in Leishmania mexicana exposed to the cationic peptide dermaseptin. Eur J Cell Biol 59:414–424PubMedGoogle Scholar
  29. Hoffman T, Schmidt JS, Zheng X, Bent AF (1999) Isolation of ethylene-insensitive soybean mutants that are altered in pathogen susceptibility and gene-for-gene disease resistance. Plant Physiol 119:935–950CrossRefPubMedGoogle Scholar
  30. Holsters M, de Waele D, Depicker A, Messens E, van Montagu M, Schell J (1978) Transfection and transformation of A. tumefaciens. Mol Gen Genet 163:181–187CrossRefPubMedGoogle Scholar
  31. Hwang PM, Vogel HJ (1998) Structure–function relationship of antimicrobial peptides. Biochem Cell Biol 76:235–246CrossRefPubMedGoogle Scholar
  32. James WC, Teng PS, Nutter WF (1990) Estimated losses of crops from plant pathogens. In: Pimentel D (ed) CRC handbook of pest management 1. CRC Press, Boca Raton, pp 5–50Google Scholar
  33. Jutglar L, Borrell JI, Ausio J (1991) Primary, secondary, and tertiary structure of the core of a histone H1-like protein from the sperm of Mytilus. J Biol Chem 266:8184–8191PubMedGoogle Scholar
  34. Kadish D, Cohen Y (1992) Overseasoning of metalaxyl-sensitive and metalaxyl-resistant isolates of Phytophthora infestans in potato tubers. Phytopathology 82:887–889Google Scholar
  35. Kanzaki H, Nirasawa S, Saitoh H, Ito M, Nishihara M, Terauchi R, Nakamura I (2002) Overexpression of the wasabi defensin gene confers enhanced resistance to blast fungus (Magnaporthe grisea) in transgenic rice. Theor Appl Genet 105:809–814CrossRefPubMedGoogle Scholar
  36. Krugliak M, Feder R, Zolotarev VY, Gaidukov L, Dagan A, Ginsburg H, Mor A (2000) Antimalarial activity of dermaseptin S4 derivatives. Antimicrob Agents Chemother 44:2442–2451CrossRefPubMedGoogle Scholar
  37. Marcos JF, Beachy RN, Houghten RA, Blondelle SE, Perez-Paya E (1995) Inhibition of plant virus infection by melittin. Proc Natl Acad Sci USA 92:12466–12469PubMedGoogle Scholar
  38. Melchers LS, Stuiver MH (2000) Novel genes for disease resistant breeding. Curr Opin Plant Biol 3:147–152Google Scholar
  39. Mitsuhara I, Matsufuru H, Ohshima M, Kaku H, Nakajima Y, Murai N, Natori S, Ohashi Y (2000) Induced expression of sarcotoxin IA enhanced host resistance against both bacterial and fungal pathogens in transgenic tobacco. Mol Plant Microbe Interact 13:860–868PubMedGoogle Scholar
  40. Mor A, Nicolas P (1994a) Isolation and structure of novel defensive peptides from frog skin. Eur J Biochem 219:145–154Google Scholar
  41. Mor A, Nicolas P (1994b) The NH2-terminal α-helical domain 1–18 of dermaseptins is responsible for antimicrobial activity. J Biol Chem 269:1934–1939Google Scholar
  42. Mor A, Nguyen VH, Delfour A, Migliore-Samour D, Nicolas P (1991) Isolation, amino acid sequence, and synthesis of dermaseptin, a novel antimicrobial peptide of amphibian skin. Biochemistry 30:8824–8830Google Scholar
  43. Mor A, Amiche M, Nicolas P (1994a) Structure, synthesis, and activity of dermaseptin B, a novel vertebrate defensive peptide from frog skin: relationship with adenoregulin. Biochemistry 33:6642–6650CrossRefPubMedGoogle Scholar
  44. Mor A, Hani K, Nicolas P (1994b) The vertebrate peptide antibiotics dermaseptins have overlapping structural features but target specific microorganisms. J Biol Chem 269:31635–31641Google Scholar
  45. Nanon-Venezia S, Feder R, Gaidukov L, Carmeli Y, Mor A (2002) Antibacterial properties of dermaseptin S4 derivatives with in vivo activity. Antimicrob Agents Chemother 46:689–694CrossRefPubMedGoogle Scholar
  46. Nir-Paz R, Prevost M-C, Nicolas P, Blanchard A, Wroblewski H (2002) Susceptibilities of Mycoplasma fermentas and Mycoplasma hyorhinis to membrane-active peptides and enrofloxacin in human tissue cell cultures. Antimicrob Agents Chemother 46:1218–1225CrossRefPubMedGoogle Scholar
  47. Oh H, Hedberg M, Wade D, Edlund C (2000) Activities of synthetic hybrid peptides against anaerobic bacteria: aspects of methodology and stability. Antimicrob Agents Chemother 44:68–72PubMedGoogle Scholar
  48. Ok SL, Boyoung L, Nammi P, Ja CK, Young HK, Theerta PD, Chandrakant K, Hyun JC, Boyoung RJ, Doh HK, Jaesung N, Jae-Gil Y, Sang-Soo K, Moo JC, Dae-Jin Y (2003) Pn-AMPs, the hevein-like proteins from Pharbitis nil confers disease resistance against phytopathogenic fungi in tomato, Lycopersicum esculentum. Phytochemistry 6:1073–1079Google Scholar
  49. Ortiz R (2001) The state of the use of potato genetic diversity. In: Cooper HD, Spillane C, Hodgkin T (eds) Broadening the genetic base of crops production. CABI/FAO, New York, pp 81–200Google Scholar
  50. Osusky M, Zhou G, Osuska L, Hancock REW, Kay WW, Misra S (2000) Transgenic plants expressing cationic peptide chimeras exhibit broad-spectrum resistance to phytopathogens. Nat Biotechnol 18:1162–1166CrossRefPubMedGoogle Scholar
  51. Osusky M, Osuska L, Hancock REW, Kay WW, Misra S (2004) Transgenic potatoes expressing a novel cationic peptide are resistant to late blight and pink rot. Transgenic Res 13:181–190CrossRefPubMedGoogle Scholar
  52. Ponti D, Mangoni ML, Mignogna G, Simmaco M, Barra D (2003) An amphibian antimicrobial peptide variant expressed in Nicotiniana tabacum confers resistance to phytopathogens. Biochem J 370:121–127CrossRefPubMedGoogle Scholar
  53. Punja ZK (2001) Genetic engineering of plants to enhance resistance to fungal pathogens—a review of progress and future prospects. Can J Plant Pathol 23:216–235Google Scholar
  54. Rich AE (1991) Plant diseases. In: Pimentel D (ed) CRC handbook of pest management 3. CRC Press, Boca Raton, pp 623–675Google Scholar
  55. Rommens CM, Kishore GM (2000) Exploiting the full potential of disease-resistance genes for agricultural use. Curr Opin Biotechnol 11:120–125CrossRefPubMedGoogle Scholar
  56. Salas B, Stack RV, Secor GA, Gudmestad NC (2000) The effect of wounding, temperature, and inoculum on the development of pink rot of potatoes caused by Phytophthora erythroseptica. Plant Dis 84:1327–1333Google Scholar
  57. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring HarborGoogle Scholar
  58. Sharma A, Sharma R, Imamura M, Yamakawa M, Machii H (2000) Transgenic expression of cecropin B, an antibacterial peptide from Bombyx mori, confers enhanced resistance to bacterial leaf blight in rice. FEBS Lett 484:7–11CrossRefPubMedGoogle Scholar
  59. Shattock RC (2002) Phytophthora infestans: populations, pathogenicity and phenylamides. Pest Manage Sci:58:944–950CrossRefGoogle Scholar
  60. Staub T (1991) Fungicide resistance: practical experience with anti-resistance strategies and the role of integrated use. Annu Rev Phytopathol 29:421–432CrossRefGoogle Scholar
  61. Strahilevitz J, Mor A, Nicolas P, Shai Y (1994) Spectrum of antimicrobial activity and assembly of dermaseptin-b and its precursor form in phospholipid membranes. Biochemistry 33:10951–10960CrossRefPubMedGoogle Scholar
  62. Thomzik JE, Stenzel K, Stocker R, Schreier PH, Hain R, Stahl DJ (1997) Synthesis of a grapevine phytoalexin in transgenic tomatoes (Lycopersicon esculentum Mill.) conditions resistance against Phytophthora infestans. Physiol Mol Plant Pathol 51:265–278CrossRefGoogle Scholar
  63. Vanhoye D, Bruston F, Nicolas P, Amiche M (2003) Antimicrobial peptides from hylid and ranin frogs originated from a 150-million-old ancestral precursor with a conserved signal peptide but a hypermutable antimicrobial domain. Eur J Biochem 270:2068–2081Google Scholar
  64. Wachinger M, Kleinschmidt A, Winder D, von Pechmann N, Ludvigsen A, Neumann M, Holle R, Salmons B, Erfle V, Brack-Werner R (1998) Antimicrobial peptides melittin and cecropin inhibit replication of human immunodeficiency virus 1 by suppressing viral gene expression. J Gen Virol 79:731–740PubMedGoogle Scholar
  65. Yang K-Y, Liu Y, Zhang S (2001) Activation of a mitogen-activated protein kinase pathway is involved in disease resistance in tobacco. Proc Natl Acad Sci USA 98:741–746CrossRefPubMedGoogle Scholar
  66. Yaron S, Rydlo T, Schachar D, Mor A (2003) Activity of dermaseptin K4-S4 against foodborne pathogens. Peptides 24:1815–1821CrossRefPubMedGoogle Scholar
  67. Zasloff M (2002) Antimicrobial peptides of multicellular organisms. Nature 415(6870):389–395CrossRefPubMedGoogle Scholar
  68. Zhu B, Chen THH, Li PH (1996) Analysis of late-blight disease resistance and freezing tolerance in transgenic potato plants expressing sense and antisense genes for an osmotin-like protein. Planta 189:70–77Google Scholar

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Milan Osusky
    • 1
  • Lubica Osuska
    • 1
  • William Kay
    • 1
  • Santosh Misra
    • 1
    Email author
  1. 1.Department of Biochemistry and MicrobiologyUniversity of VictoriaVictoriaCanada

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