European Journal of Forest Research

, Volume 133, Issue 4, pp 573–583 | Cite as

Low-density Ceratocystis polonica inoculation of Norway spruce (Picea abies) triggers accumulation of monoterpenes with antifungal properties

  • Metka Novak
  • Andreja Urbanek Krajnc
  • Ljerka Lah
  • Neja Zupanec
  • Nada Kraševec
  • Mitja Križman
  • Joerg Bohlmann
  • Radovan Komel
Original Paper

Abstract

Among the most devastating pests of Norway spruce (Picea abies) are the European spruce bark beetle (Ips typographus) and the associated pathogenic blue-stain fungus Ceratocystis polonica. Following attack and colonization, the beetle and the fungus must cope with induced host chemical defenses, such as monoterpenes that are generally thought to be toxic to both symbionts. The goal of this study was to better understand the response of Norway spruce following C. polonica inoculation at low density that does not overwhelm the tree and to identify monoterpenes mobilized toward the fungus. We inoculated healthy mature trees and monitored monoterpene profiles 2, 3, and 5 months post-inoculation. We also exposed three different C. polonica strains to the most abundant or significantly up-regulated monoterpenes to determine differences in monoterpene toxicity and resistance among strains. Total monoterpene levels, including limonene, were increased at 2 and 3 months after inoculation and had dropped after 5 months. In in vitro assays, all monoterpenes were inhibitory to C. polonica. Limonene and β-pinene were the most potent inhibitors of fungal growth. The extent of inhibition varied between the three strains tested. These results showed a defense response of Norway spruce to C. polonica, in which limonene may play a critical role in inhibiting the spread of the fungus. We also showed that differences between strains of C. polonica must be taken into account when assessing the role of the fungus in this bark beetle–symbiont system.

Keywords

Blue-stain fungi Monoterpene Growth inhibition Limonene 

Supplementary material

10342_2013_772_MOESM1_ESM.pdf (509 kb)
Supplementary material 1 (PDF 509 kb)

References

  1. Baier P, Führer E, Kirisits T, Rosner S (2002) Defence reactions of Norway spruce against bark beetles and the associated fungus Ceratocystis polonica in secondary pure and mixed species stands. For Ecol Manage 159:73–86CrossRefGoogle Scholar
  2. Bohlmann J (2012) Pine terpenoid defences in the mountain pine beetle epidemic and in other conifer pest interactions: specialized enemies are eating holes into a diverse, dynamic and durable defence system. Tree Physiol 32:943–945PubMedCrossRefGoogle Scholar
  3. Boone CK, Aukema BH, Bohlmann J, Carroll AL, Raffa KF (2011) Efficacy of tree defense physiology varies with bark beetle population density: a basis for positive feedback in eruptive species. Can J For Res 41:1174–1188CrossRefGoogle Scholar
  4. Christiansen E (1985) Ceratocysts polonica inoculated in Norway spruce: blue-staining in relation to inoculum density, resinosis and tree growth. For Pathol 15:160–167CrossRefGoogle Scholar
  5. Christiansen E, Bakke A (1988) The spruce bark beetle of Eurasia. In: Dynamics of forest insect populations: patterns, causes, implications. Plenum Press, New YorkGoogle Scholar
  6. Croteau R, Gurkewitz S, Johnson MA, Fisk HJ (1987) Biochemistry of Oleoresinosis: monoterpene and diterpene biosynthesis in lodgepole pine saplings infected with Ceratocystis clavigera or treated with carbohydrate elicitors. Plant Physiol 85:1123–1128PubMedCentralPubMedCrossRefGoogle Scholar
  7. DiGuistini S, Wang Y, Liao NY, Taylor G, Tanguay P, Feau N, Henrissat B, Chan SK, Hesse-Orce U, Alamouti SM, Tsui CKM, Docking RT, Levasseur A, Haridas S, Robertson G, Birol I, Holt RA, Marra MA, Hamelin RC, Hirst M, Jones SJM, Bohlmann J, Breuil C (2011) Genome and transcriptome analyses of the mountain pine beetle-fungal symbiont Grosmannia clavigera, a lodgepole pine pathogen. Proc Natl Acad Sci USA 108:2504–2509PubMedCentralPubMedCrossRefGoogle Scholar
  8. Eckhardt LG, Menard RD, Gray ED (2009) Effects of oleoresins and monoterpenes on in vitro growth of fungi associated with pine decline in the Southern United States. For Pathol 39:157–167CrossRefGoogle Scholar
  9. Erbilgin N, Colgan LJ (2012) Differential effects of plant ontogeny and damage type on phloem and foliage monoterpenes in jack pine (Pinus banksiana). Tree Physiol 32:946–957PubMedCrossRefGoogle Scholar
  10. Erbilgin N, Krokene P, Christiansen E, Zeneli G, Gershenzon J (2006) Exogenous application of methyl jasmonate elicits defenses in Norway spruce (Picea abies) and reduces host colonization by the bark beetle Ips typographus. Oecologia 148:426–436PubMedCrossRefGoogle Scholar
  11. Evensen PC, Solheim H, Høiland K, Stenersen J (2000) Induced resistance of Norway spruce, variation of phenolic compounds and their effects on fungal pathogens. For Pathol 30:97–108CrossRefGoogle Scholar
  12. Faccoli M (2009) Effect of weather on Ips typographus (Coleoptera Curculionidae) phenology, voltinism, and associated spruce mortality in the southeastern Alps. Environ Entomol 38:307–316PubMedCrossRefGoogle Scholar
  13. Fäldt J, Solheim H (2006) Influence of fungal infection and wounding on contents and enantiomeric compositions of monoterpenes in phloem of Pinus sylvestris. J Chem Ecol 32:1779–1795PubMedCrossRefGoogle Scholar
  14. Franceschi VR, Krokene P, Christiansen E, Krekling T (2005) Anatomical and chemical defenses of conifer bark against bark beetles and other pests. New Phytol 167:353–375PubMedCrossRefGoogle Scholar
  15. Hall DE, Robert JA, Keeling CI, Domanski D, Quesada AL, Jancsik S, Kuzyk MA, Hamberger B, Borchers CH, Bohlmann J (2011) An integrated genomic, proteomic and biochemical analysis of (+)-3-carene biosynthesis in Sitka spruce (Picea sitchensis) genotypes that are resistant or susceptible to white pine weevil. Plant J Cell Mol Biol 65:936–948CrossRefGoogle Scholar
  16. Hammerbacher A, Schmidt A, Wadke N, Wright LP, Schneider B, Bohlmann J, Brand WA, Fenning TM, Gershenzon J, Paetz C (2013) A common fungal associate of the spruce bark beetle metabolizes the stilbene defenses of Norway spruce. Plant Physiol 162(3):1324–1336PubMedCentralPubMedCrossRefGoogle Scholar
  17. Horntvedt R, Christiansen E, Solheim H, Wang S (1983) Artificial inoculation with Ips typographus-associated blue-stain fungi can kill healthy Norway spruce trees (Ceratocystis polonica, Ceratocystis penicillata, water stress). Rep Nor For Res Inst 38:1–20Google Scholar
  18. Jost R, Rice A (2008) Monoterpene emissions from lodgepole and jack pine bark inoculated with mountain pine beetle associated fungi. J Wood Chem 28:37–46CrossRefGoogle Scholar
  19. Keeling CI, Bohlmann J (2006a) Diterpene resin acids in conifers. Phytochemistry 67:2415–2423PubMedCrossRefGoogle Scholar
  20. Keeling CI, Bohlmann J (2006b) Genes, enzymes and chemicals of terpenoid diversity in the constitutive and induced defence of conifers against insects and pathogens. New Phytol 170:657–675PubMedCrossRefGoogle Scholar
  21. Klepzig KD, Robison DJ, Fowler G, Minchin PR, Hain FP, Allen HL (2005) Effects of mass inoculation on induced oleoresin response in intensively managed loblolly pine. Tree Physiol 25:681–688PubMedCrossRefGoogle Scholar
  22. Klepzig KD, Adams AS, Handelsman J, Raffa KF (2009) Symbioses: a key driver of insect physiological processes, ecological interactions, evolutionary diversification, and impacts on humans. Environ Entomol 38:67–77PubMedCrossRefGoogle Scholar
  23. Klepzig KD, Smalley EB, Raffa KF (1996) Combined chemical defenses against an insect-fungal complex. J Chem Ecol 22:1367–1388Google Scholar
  24. Krajnc AU (2009) A temporal analysis of antioxidative defense responses in the phloem of Picea abies after attack by Ips typographus. Tree Physiol 29:1059–1068CrossRefGoogle Scholar
  25. Krokene P, Solheim H (1996) Fungal associates of five bark beetle species colonizing Norway spruce. Can J For Res 2115–2122Google Scholar
  26. Krokene P, Solheim H (1997) Growth of four bark-beetle-associated blue-stain fungi in relation to the induced wound response in Norway spruce. Can J Bot 75:618–625CrossRefGoogle Scholar
  27. Krokene P, Solheim H (1998) Pathogenicity of four blue-stain fungi associated with aggressive and nonaggressive bark beetles. Phytopathology 88:39–44PubMedCrossRefGoogle Scholar
  28. Krokene P, Solheim H (1999) What do low-density inoculations with fungus tell us about fungal virulence and tree resistance? In: Lieutier F, Mattson WJ, Wagner MR (eds) Physiology and genetics of tree-phytophage interactions INRA editions. Versailles, France, pp 353–362Google Scholar
  29. Krokene P, Christiansen E, Solheim H, Franceschi V, Berryman A (1999) Induced resistance to pathogenic fungi in Norway spruce. Plant Physiol 121:565–570PubMedCentralPubMedCrossRefGoogle Scholar
  30. Krokene P, Solheim H, Christiansen E (2001) Induction of disease resistance in Norway spruce (Picea abies) by necrotizing fungi. Plant Pathol 50:230–233CrossRefGoogle Scholar
  31. Krokene P, Nagy NE, Krekling T (2008) Traumatic resin ducts and polyphenolic parenchyma cells in conifers. In: Schaller A (ed) Induced plant resistance to herbivory. Springer, Netherlands, Dordrecht, pp 147–169CrossRefGoogle Scholar
  32. Krupa S, Fries N (1971) Studies on ectomycorrhizae of pine. Production of volatile organic compounds. Can J Bot 49:1425–1431CrossRefGoogle Scholar
  33. Lah L, Haridas S, Bohlmann J, Breuil C (2012a) The cytochromes P450 of Grosmannia clavigera: genome organization, phylogeny, and expression in response to pine host chemicals. Fungal Genet Biol 50:72–81PubMedCrossRefGoogle Scholar
  34. Lah L, Haridas S, Bohlmann J, Breuil C (2012) The cytochromes P450 of Grosmannia clavigera: genome organization, phylogeny, and expression in response to pine host chemicals. Fungal Genet BiolGoogle Scholar
  35. Lewinsohn E, Savage T (1993) Simultaneous analysis of monoterpenes and diterpenoids of conifer oleoresin. Phytochem Anal 4:220–225CrossRefGoogle Scholar
  36. Li SH, Nagy NE, Hammerbacher A, Krokene P, Niu XM, Gershenzon J, Schneider B (2012) Localization of phenolics in phloem parenchyma cells of Norway spruce (Picea abies). Chembiochem: A Eur J Chem Biol 18:2707–2713CrossRefGoogle Scholar
  37. Lieutier F, Yart A, Salle A (2009) Stimulation of tree defenses by ophiostomatoid fungi can explain attack success of bark beetles on conifers. Ann For Sci 66:1–22CrossRefGoogle Scholar
  38. Linnakoski R, De Beer ZW, Niemelä P, Wingfield MJ (2012) Associations of conifer-infesting bark beetles and fungi in Fennoscandia. Insects 3:200–227CrossRefGoogle Scholar
  39. Lu M, Wingfield MJ, Gillette NE, Mori SR, Sun JH (2010a) Complex interactions among host pines and fungi vectored by an invasive bark beetle. New Phytol 187:8CrossRefGoogle Scholar
  40. Lu M, Wingfield MJ, Gillette NE, Mori SR, Sun J-H (2010b) Complex interactions among host pines and fungi vectored by an invasive bark beetle. New Phytol 187:859–866PubMedCrossRefGoogle Scholar
  41. Marei GIK, Abdel Rasoul MA, Abdelgaleil SAM (2012) Comparative antifungal activities and biochemical effects of monoterpenes on plant pathogenic fungi. Pestic Biochem Physiol 103:56–61CrossRefGoogle Scholar
  42. Martin D, Tholl D, Gershenzon J, Bohlmann J (2002) Methyl jasmonate induces traumatic resin ducts, terpenoid resin biosynthesis, and terpenoid accumulation in developing xylem of Norway spruce stems. Plant Physiol 129:1003–1018PubMedCentralPubMedCrossRefGoogle Scholar
  43. Paine TD, Raffa KF, Harrington TC (1997) Interactions among scolytid bark beetles, their associated fungi, and live host conifers. Annu Rev Entomol 42:179–206PubMedCrossRefGoogle Scholar
  44. Pasquier-Barre F, Palasse C, Goussard F, Auger-Rozenberg M-A, Géri C (2001) Relationship of Scots pine clone characteristics and water stress to hatching and larval performance of the sawfly Diprion pini (Hymenoptera: Diprionidae). Environ Entomol 30:1–6CrossRefGoogle Scholar
  45. Persson M, Sjödin K, Borg-Karlson A-K, Norin T, Ekberg I (1996) Relative amounts and enantiomeric compositions of monoterpene hydrocarbons in xylem and needles of Picea abies. Phytochemistry 42:1289–1297CrossRefGoogle Scholar
  46. Phillips MA, Croteau RB (1999) Resin-based defenses in conifers. Trends Plant Sci 4:184–190PubMedCrossRefGoogle Scholar
  47. Raffa KF, Berryman AA (1982) Accumulation of monoterpenes and associated volatiles following inoculation of grand fir with a fungus transmitted by the fir engraver, Scolytus ventralis (Coleoptera: Scolytidae). Can Entomol 114:797–810CrossRefGoogle Scholar
  48. Raffa KF, Berryman AA (1983a) Physiological aspects of lodgepole pine wound responses to a fungal symbiont of the mountain pine beetle, Dendroctonus ponderosae (Coleoptera: Scolytidae). Can Entomol 115:723–734CrossRefGoogle Scholar
  49. Raffa KF, Berryman AA (1983b) The role of host plant resistance in the colonization behavior and ecology of bark beetles (Coleoptera: Scolytidae). Ecol Monogr 53:27CrossRefGoogle Scholar
  50. Raffa KF, Smalley EB (1995) Interaction of pre-attack and induced monoterpene concentrations in host conifer defense against bark beetle-fungal complexes. Oecologia 102:285–295CrossRefGoogle Scholar
  51. Raffa KF, Berryman AA, Simasko J, Teal W, Wong BL (1985) Effects of grand fir monoterpenes on the fir engraver, Scolytus ventralis (Coleoptera: Scolytidae), and its symbiotic fungus. Environ Entomol 14:5Google Scholar
  52. Raffa KF, Aukema BH, Bentz BJ, Carroll AL, Hicke JA, Turner MG, Romme WH (2008) Cross-scale drivers of natural disturbances prone to anthropogenic amplification: the dynamics of bark beetle eruptions. Bioscience 58:501CrossRefGoogle Scholar
  53. Reed AN, Hanover JW, Furniss MM (1986) Douglas-fir and western larch: chemical and physical properties in relation to Douglas-fir bark beetle attack. Tree Physiol 1:277–287PubMedCrossRefGoogle Scholar
  54. Robert JA, Madilao LL, White R, Yanchuk A, King J, Bohlmann J (2010) Terpenoid metabolite profiling in Sitka spruce identifies association of dehydroabietic acid, (+)-3-carene, and terpinolene with resistance against white pine weevil. Botany 88:810–820CrossRefGoogle Scholar
  55. Rocchini LA, Lindgren BS, Bennett RG (2000) Effects of resin flow and monoterpene composition on susceptibility of lodgepole pine to attack by the Douglas-fir pitch moth, Synanthedon novaroensis (Lep., Sesiidae). J Appl Entomol 124:87–92CrossRefGoogle Scholar
  56. Sallé A, Monclus R, Yart A, Garcia J, Romary P, Lieutier F (2005) Fungal flora associated with Ips typographus: frequency, virulence, and ability to stimulate the host defence reaction in relation to insect population levels. Can J For Res 35:365–373CrossRefGoogle Scholar
  57. Schiebe C, Hammerbacher A, Birgersson G, Witzell J, Brodelius PE, Gershenzon J, Hansson BS, Krokene P, Schlyter F (2012) Inducibility of chemical defenses in Norway spruce bark is correlated with unsuccessful mass attacks by the spruce bark beetle. Oecologia 170:183–198PubMedCrossRefGoogle Scholar
  58. Silvestrini E, Michelozzi M, Skroppa T, Brancaleoni E, Ciccioli P (2004) Characterisation of different clones of Picea abies (L.) Karst using head-space sampling of cortical tissues combined with enantioselective capillary gas chromatography for the separation of chiral and non-chiral monoterpenes. J Chromatogr A 1034:183–189PubMedCrossRefGoogle Scholar
  59. Six DL, Wingfield MJ (2011) The role of phytopathogenicity in bark beetle-fungus symbioses: a challenge to the classic paradigm. Annu Rev Entomol 56:255–272PubMedCrossRefGoogle Scholar
  60. Sjödin K, Persson M, Faldt J, Ekberg I, Borg-Karlson A-K (2001) Occurrence and correlations of monoterpene hydrocarbon enantiomers in Pinus sylvestris and Picea abies. J Chem Ecol 26:1701–1720CrossRefGoogle Scholar
  61. Smith RH (1965) Effect of monoterpene vapors on the western pine beetle. J Econ Entomol 58:2Google Scholar
  62. Solheim H (1986) Species of Ophiostomataceae isolated from Picea abies infested by the bark beetle Ips typographic. Nordic J Bot 6:199–207CrossRefGoogle Scholar
  63. Solheim H (1991) Oxygen deficiency and spruce resin inhibition of growth of blue stain fungi associated with Ips typographus. Mycol Res 95:1387–1392CrossRefGoogle Scholar
  64. Tausz M, Wonisch A, Grill D, Morales D, Jiménez MS (2003) Measuring antioxidants in tree species in the natural environment: from sampling to data evaluation. J Exp Bot 54:1505–1510PubMedCrossRefGoogle Scholar
  65. Viiri H, Annila E, Kitunen V, Niemelä P (2001) Induced responses in stilbenes and terpenes in fertilized Norway spruce after inoculation with blue-stain fungus, Ceratocystis polonica. Trees 15:112–122CrossRefGoogle Scholar
  66. Wallin KF, Raffa KF (1999) Altered constitutive and inducible phloem monoterpenes following natural defoliation of jack pine: implications to host mediated interguild interactions and plant defense theories. J Chem Ecol 25:861–880CrossRefGoogle Scholar
  67. Wang Y, Lim L, Diguistini S, Robertson G, Bohlmann J, Breuil C (2013) A specialized ABC efflux transporter GcABC-G1 confers monoterpene resistance to Grosmannia clavigera, a bark beetle-associated fungal pathogen of pine trees. New Phytol 197:886–898PubMedCrossRefGoogle Scholar
  68. Wermelinger B (2004) Ecology and management of the spruce bark beetle Ips typographus—a review of recent research. For Ecol Manage 202:67–82CrossRefGoogle Scholar
  69. Wright E (1933) A cork-borer method for inoculating trees. Phytopathology 23:487–488Google Scholar
  70. Zamponi L, Michelozzi M, Capretti P (2007) Terpene response of Picea abies and Abies alba to infection with Heterobasidion sl. For Pathol 37:243–250CrossRefGoogle Scholar
  71. Zeneli G, Krokene P, Christiansen E, Krekling T, Gershenzon J (2006) Methyl jasmonate treatment of mature Norway spruce (Picea abies) trees increases the accumulation of terpenoid resin components and protects against infection by Ceratocystis polonica, a bark beetle-associated fungus. Tree Physiol 26:977–988PubMedCrossRefGoogle Scholar
  72. Zhao T, Krokene P, Björklund N, Långström B, Solheim H, Christiansen E, Borg-Karlson AK (2010) The influence of Ceratocystis polonica inoculation and methyl jasmonate application on terpene chemistry of Norway spruce, Picea abies. Phytochemistry 71:1332–1341PubMedCrossRefGoogle Scholar
  73. Zhao T, Krokene P, Hu J, Christiansen E, Björklund N, Långström B, Solheim H, Borg-Karlson AK (2011) Induced terpene accumulation in Norway spruce inhibits bark beetle colonization in a dose-dependent manner. PLoS ONE 6:e26649PubMedCentralPubMedCrossRefGoogle Scholar
  74. Zulak KG, Bohlmann J (2010) Terpenoid biosynthesis and specialized vascular cells of conifer defense. J Integr Plant Biol 52:86–97PubMedCrossRefGoogle Scholar
  75. Zulak KG, Lippert DN, Kuzyk MA, Domanski D, Chou T, Borchers CH, Bohlmann J (2009) Targeted proteomics using selected reaction monitoring reveals the induction of specific terpene synthases in a multi-level study of methyl jasmonate-treated Norway spruce (Picea abies). Plant J 60:1015–1030PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Metka Novak
    • 1
  • Andreja Urbanek Krajnc
    • 2
  • Ljerka Lah
    • 1
  • Neja Zupanec
    • 1
  • Nada Kraševec
    • 1
  • Mitja Križman
    • 1
  • Joerg Bohlmann
    • 4
  • Radovan Komel
    • 1
    • 3
  1. 1.National Institute of ChemistryLjubljanaSlovenia
  2. 2.Faculty of Agriculture and Life SciencesUniversity of MariborHočeSlovenia
  3. 3.Faculty of Medicine, Institute of BiochemistryUniversity of LjubljanaLjubljanaSlovenia
  4. 4.Michael Smith LaboratoriesUniversity of British ColumbiaVancouverCanada

Personalised recommendations