Mycological Progress

, 14:106 | Cite as

Identification of genes differentially expressed during the interaction between the plant symbiont Suillus luteus and two plant pathogenic allopatric Heterobasidion species

  • F. Sillo
  • E. Zampieri
  • L. Giordano
  • G. Lione
  • J. V. Colpaert
  • R. Balestrini
  • P. GonthierEmail author
Original Article


The effects of biological invasions by non-native species have been widely studied in terms of environmental, economic, and human health impacts. However, little is known about the consequences that non-native plant pathogens may determine on host plant symbionts, such as ectomycorrhizal (ECM) fungi. In this study, interactions between Suillus luteus, an ECM fungus of pine trees, and the allopatrically differentiated fungal pathogens of pines Heterobasidion irregulare and H. annosum were investigated in dual culture by morphological and gene expression analyses. Growth of S. luteus was inhibited by both Heterobasidion species, but based on statistical analysis, growth inhibition was due to the isolate rather than to the species. The expression analysis on genes related to cell wall hydrolytic enzymes and hydrophobins, putatively involved in the fungus–fungus interaction, allowed to identify significantly up- and down-regulated genes both in the symbiont and in the pathogens. Based on the transcript analysis, it was not possible to distinguish the impact of the two pathogenic species on the ECM fungus. The only exception was a S. luteus gene coding for a putative chitinase (SlGH18_8356) that was found to be differentially regulated during interaction with H. irregulare compared to H. annosum.


Ectomycorrhizal fungus Fungal pathogen Dual culture Gene expression Cell wall Phylogeny 



This work was supported by the Italian Ministry of Education, University and Research, within the FIRB program (grant number RBFRI280NN).

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

11557_2015_1130_MOESM1_ESM.doc (237 kb)
ESM 1 (DOC 237 kb)


  1. Adomas A, Eklund M, Johansson M, Asiegbu FO (2006) Identification and analysis of differentially expressed cDNAs during nonself-competitive interaction between Phlebiopsis gigantea and Heterobasidion parviporum. FEMS Microbiol Ecol 57:26–39CrossRefPubMedGoogle Scholar
  2. Baccelli I, Gonthier P, Bernardi R (2015) Gene expression analyses reveal a relationship between conidiation and cerato-platanin in homokaryotic and heterokaryotic strains of the fungal plant pathogen Heterobasidion irregulare. Mycol Progr. doi: 10.1007/s11557-015-1063-x Google Scholar
  3. Berg A, Meyer R, Yu J (2004) Deviance information criterion for comparing stochastic volatility models. J Bus Econ Stat 22:107–120CrossRefGoogle Scholar
  4. Bowman SM, Free SJ (2006) The structure and synthesis of the fungal cell wall. Bioessays 28:799–808CrossRefPubMedGoogle Scholar
  5. Branzanti MB, Rocca E, Pisi A (1999) Effect of ectomycorrhizal fungi on chestnut ink disease. Mycorrhiza 9:103–109CrossRefGoogle Scholar
  6. Červinková H (1990) Mycorrhizae and control of root pathogen Heterobasidion annosum. Agric Ecosyst Environ 28:55–58CrossRefGoogle Scholar
  7. Chaffin WL, López-Ribot JL, Casanova M, Gozalbo D, Martínez JP (1998) Cell wall and secreted proteins of Candida albicans: identification, function, and expression. Microbiol Mol Biol Rev 62:130–180PubMedCentralPubMedGoogle Scholar
  8. Crawley MJ (2013) The R book, 2nd edn. Wiley, ChichesterGoogle Scholar
  9. Dahlberg A, Finlay RD (1999) Suillus. In: Cairney JWG, Chambers SM (eds) Ectomycorrhizal fungi: key genera in profile. Springer-Verlag, Berlin, pp 33–64CrossRefGoogle Scholar
  10. Fries N (1978) Basidiospore germination in some mycorrhiza-forming Hymenomycetes. Trans Br Mycol Soc 70:319–324CrossRefGoogle Scholar
  11. Garbelotto M, Gonthier P (2013) Biology, epidemiology, and control of Heterobasidion species worldwide. Annu Rev Phytopathol 51:39–59CrossRefPubMedGoogle Scholar
  12. Garbelotto M, Linzer R, Nicolotti G, Gonthier P (2010) Comparing the influences of ecological and evolutionary factors on the successful invasion of a fungal forest pathogen. Biol Invasions 12:943–957CrossRefGoogle Scholar
  13. Garbelotto M, Guglielmo F, Mascheretti S, Croucher PJP, Gonthier P (2013) Population genetic analyses provide insights on the introduction pathway and spread patterns of the North American forest pathogen Heterobasidion irregulare in Italy. Mol Ecol 22:4855–4869CrossRefPubMedGoogle Scholar
  14. Giordano L, Gonthier P, Lione G, Capretti P, Garbelotto M (2014) The saprobic and fruiting abilities of the exotic forest pathogen Heterobasidion irregulare may explain its invasiveness. Biol Invasions 16:803–814CrossRefGoogle Scholar
  15. Gonthier P, Garbelotto M (2011) Amplified fragment length polymorphism and sequence analyses reveal massive gene introgression from the European fungal pathogen Heterobasidion annosum into its introduced congener H. irregulare. Mol Ecol 20:2756–2770CrossRefPubMedGoogle Scholar
  16. Gonthier P, Warner R, Nicolotti G, Mazzaglia A, Garbelotto MM (2004) Pathogen introduction as a collateral effect of military activity. Mycol Res 108:468–470CrossRefPubMedGoogle Scholar
  17. Gonthier P, Nicolotti G, Linzer R, Guglielmo F, Garbelotto M (2007) Invasion of European pine stands by a North American forest pathogen and its hybridization with a native. Mol Ecol 16:1389–1400CrossRefPubMedGoogle Scholar
  18. Gonthier P, Lione G, Giordano L, Garbelotto M (2012) The American forest pathogen Heterobasidion irregulare colonizes unexpected habitats after its introduction in Italy. Ecol Appl 22:2135–2143CrossRefPubMedGoogle Scholar
  19. Gruber S, Seidl-Seiboth V (2012) Self versus non-self: fungal cell wall degradation in Trichoderma. Microbiology 158:26–34CrossRefPubMedGoogle Scholar
  20. Gruber S, Vaaje-Kolstad G, Matarese F, López-Mondéjar R, Kubicek CP, Seidl-Seiboth V (2011) Analysis of subgroup C of fungal chitinases containing chitin-binding and LysM modules in the mycoparasite Trichoderma atroviride. Glycobiology 21:122–133CrossRefPubMedGoogle Scholar
  21. Hadfield JD (2010) MCMC methods for multi-response generalized linear mixed models: the MCMCglmm R package. J Stat Softw 33:1–22CrossRefGoogle Scholar
  22. Holdenrieder O (1984) Untersuchungen zur biologischen Bekämpfung von Heterobasidion annosum an Fichte (Picea abies) mit antagonistischen Pilzen. II. Interaktionstests auf Holz. Eur J Forest Pathol 14:137–153CrossRefGoogle Scholar
  23. Iakovlev A, Olson A, Elfstrand M, Stenlid J (2004) Differential gene expression during interactions between Heterobasidion annosum and Physisporinus sanguinolentus. FEMS Microbiol Lett 241:79–85CrossRefPubMedGoogle Scholar
  24. Ismail Y, McCormick S, Hijri M (2011) A fungal symbiont of plant-roots modulates mycotoxin gene expression in the pathogen Fusarium sambucinum. PLoS One 6:e17990PubMedCentralCrossRefPubMedGoogle Scholar
  25. Karlsson M, Stenlid J (2008) Comparative evolutionary histories of the fungal chitinase gene family reveal non-random size expansions and contractions due to adaptive natural selection. Evol Bioinform Online 4:47–60PubMedCentralPubMedGoogle Scholar
  26. Keller RP, Geist J, Jeschke JM, Kühn I (2011) Invasive species in Europe: ecology, status, and policy. Env Sci Eur 23:23CrossRefGoogle Scholar
  27. Kéry M (2010) Introduction to WinBUGS for ecologists: a Bayesian approach to regression, ANOVA, mixed models and related analysis. Academic Press, BurlingtonGoogle Scholar
  28. Kohler A, Kuo A, Nagy LG, Morin E, Barry KW, Buscot F, Canbäck B, Choi C, Cichocki N, Clum A, Colpaert J, Copeland A, Costa MD, Doré J, Floudas D, Gay G, Girlanda M, Henrissat B, Herrmann S, Hess J, Högberg N, Johansson T, Khouja HR, LaButti K, Lahrmann U, Levasseur A, Lindquist EA, Lipzen A, Marmeisse R, Martino E, Murat C, Ngan CY, Nehls U, Plett JM, Pringle A, Ohm RA, Perotto S, Peter M, Riley R, Rineau F, Ruytinx J, Salamov A, Shah F, Sun H, Tarkka M, Tritt A, Veneault-Fourrey C, Zuccaro A; Mycorrhizal Genomics Initiative Consortium, Tunlid A, Grigoriev IV, Hibbett DS, Martin F (2015) Convergent losses of decay mechanisms and rapid turnover of symbiosis genes in mycorrhizal mutualists. Nat Genet 47:410–415CrossRefPubMedGoogle Scholar
  29. Lalaymia I, Cranenbrouck S, Declerck S (2014) Maintenance and preservation of ectomycorrhizal and arbuscular mycorrhizal fungi. Mycorrhiza 24:323–337CrossRefPubMedGoogle Scholar
  30. Lass-Flörl C, Nagl M, Speth C, Ulmer H, Dierich MP, Würzner R (2001) Studies of in vitro activities of voriconazole and itraconazole against Aspergillus hyphae using viability staining. Antimicrob Agents Chemother 45:124–128PubMedCentralCrossRefPubMedGoogle Scholar
  31. Lei ZP, Jin JR, Wang CW (1995) Antagonism between ectomycorrhizal fungi and plant pathogens. In: Brundett M, Dell B, Malajczuk M (ed) Mycorrhizas for plantation forestry in Asia. Proceedings of an International Symposium and Workshop, 7–11 November 1994, Kaiping, China, proceedings no. 62. Australian Centre for International Agricultural Research, Canberra, Australia, pp 77–81Google Scholar
  32. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) Method. Methods 25:402–408CrossRefPubMedGoogle Scholar
  33. Lövei GL (1997) Biodiversity: global change through invasion. Nature 388:627–628CrossRefGoogle Scholar
  34. Millard PJ, Roth BL, Thi HP, Yue ST, Haugland RP (1997) Development of the FUN-1 family of fluorescent probes for vacuole labeling and viability testing of yeasts. Appl Environ Microbiol 63:2897–2905PubMedCentralPubMedGoogle Scholar
  35. Mohan V, Nivea R, Menon S (2015) Evaluation of ectomycorrhizal fungi as potential bio-control agents against selected plant pathogenic fungi. JAIR 3:408–412Google Scholar
  36. Mucha J, Dahm H, Strzelczyk E, Werner A (2006) Synthesis of enzymes connected with mycoparasitism by ectomycorrhizal fungi. Arch Microbiol 185:69–77CrossRefPubMedGoogle Scholar
  37. Mucha J, Zadworny M, Werner A (2009) Cytoskeleton and mitochondrial morphology of saprotrophs and the pathogen Heterobasidion annosum in the presence of Suillus bovinus metabolites. Mycol Res 113:981–990CrossRefPubMedGoogle Scholar
  38. Napierała-Filipiak A, Werner A (2000) Antagonism of higher fungi to Heterobasidion annosum (Fr.) Bref. in laboratory conditions. Dendrobiology 45:65–81Google Scholar
  39. Nicolotti G, Gonthier P, Varese GC (1999) Effectiveness of some biocontrol and chemical treatments against Heterobasidion annosum on Norway spruce stumps. Eur J For Pathol 29:339–346CrossRefGoogle Scholar
  40. Ohm RA, de Jong JF, Lugones LG, Aerts A, Kothe E, Stajich JE, de Vries RP, Record E, Levasseur A, Baker SE, Bartholomew KA, Coutinho PM, Erdmann S, Fowler TJ, Gathman AC, Lombard V, Henrissat B, Knabe N, Kües U, Lilly WW, Lindquist E, Lucas S, Magnuson JK, Piumi F, Raudaskoski M, Salamov A, Schmutz J, Schwarze FW, vanKuyk PA, Horton JS, Grigoriev IV, Wösten HA (2010) Genome sequence of the model mushroom Schizophyllum commune. Nat Biotechnol 28:957–963CrossRefPubMedGoogle Scholar
  41. Olson A, Aerts A, Asiegbu F, Belbahri L, Bouzid O, Broberg A, Canbäck B, Coutinho PM, Cullen D, Dalman K, Deflorio G, van Diepen LT, Dunand C, Duplessis S, Durling M, Gonthier P, Grimwood J, Fossdal CG, Hansson D, Henrissat B, Hietala A, Himmelstrand K, Hoffmeister D, Högberg N, James TY, Karlsson M, Kohler A, Kües U, Lee YH, Lin YC, Lind M, Lindquist E, Lombard V, Lucas S, Lundén K, Morin E, Murat C, Park J, Raffaello T, Rouzé P, Salamov A, Schmutz J, Solheim H, Ståhlberg J, Vélëz H, de Vries RP, Wiebenga A, Woodward S, Yakovlev I, Garbelotto M, Martin F, Grigoriev IV, Stenlid J (2012) Insight into trade-off between wood decay and parasitism from the genome of a fungal forest pathogen. New Phytol 194:1001–1013CrossRefPubMedGoogle Scholar
  42. Perrin R (1990) Interactions between mycorrhizae and diseases caused by soil-borne fungi. Soil Use Manag 6:189–194CrossRefGoogle Scholar
  43. Petersen TN, Brunak S, von Heijne G, Nielsen H (2011) SignalP 4.0: discriminating signal peptides from transmembrane regions. Nat Methods 8:785–786CrossRefPubMedGoogle Scholar
  44. Pollastrini M, Luchi N, Michelozzi M, Gerosa G, Marzuoli R, Bussotti F, Capretti P (2015) Early physiological responses of Pinus pinea L. seedlings infected by Heterobasidion in an ozone-enriched atmospheric environment. Tree Physiol 35:331–340CrossRefPubMedGoogle Scholar
  45. Pyšek P, Richarson DM (2010) Invasive species, environmental change and management, and health. Ann Rev Environ Resour 35:25–55CrossRefGoogle Scholar
  46. Raffaello T, Asiegbu FO (2013) Evaluation of potential reference genes for use in gene expression studies in the conifer pathogen (Heterobasidion annosum). Mol Biol Rep 40:4605–4611CrossRefPubMedGoogle Scholar
  47. Raffaello T, Chen H, Kohler A, Asiegbu FO (2014) Transcriptomic profiles of Heterobasidion annosum under abiotic stresses and during saprotrophic growth in bark, sapwood and heartwood. Environ Microbiol 16:1654–1667CrossRefPubMedGoogle Scholar
  48. Rayner AD, Griffith GS, Wildman HG (1994) Induction of metabolic and morphogenetic changes during mycelial interactions among species of higher fungi. Biochem Soc Trans 22:389–394CrossRefPubMedGoogle Scholar
  49. Sivan A, Chet I (1989) Degradation of fungal cell walls by lytic enzymes of Trichoderma harzianum. J Gen Microbiol 135:675–682Google Scholar
  50. Slámová K, Bojarová P, Petrásková L, Křen V (2010) β-N-Acetylhexosaminidase: what’s in a name…? Biotechnol Adv 28:682–693CrossRefPubMedGoogle Scholar
  51. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Mol Biol Evol 30:2725–2729PubMedCentralCrossRefPubMedGoogle Scholar
  52. Thomson BD, Malajczuk N, Grove TS, Hardy GESJ (1993) Improving the colonization capacity and effectiveness of ectomycorrhizal fungal cultures by association with a host plant and re-isolation. Mycol Res 97:839–844CrossRefGoogle Scholar
  53. Umekawa M, Huang W, Li B, Fujita K, Ashida H, Wang LX, Yamamoto K (2008) Mutants of Mucor hiemalis endo-beta-N-acetylglucosaminidase show enhanced transglycosylation and glycosynthase-like activities. J Biol Chem 283:4469–4479CrossRefPubMedGoogle Scholar
  54. Werner A, Lakomy P (2002) Intraspecific variation in Heterobasidion annosum for mortality rate on Pinus sylvestris and Picea abies seedlings grown in pure culture. Mycologia 94:856–861CrossRefPubMedGoogle Scholar
  55. Werner A, Zadworny M (2003) In vitro evidence of mycoparasitism of the ectomycorrhizal fungus Laccaria laccata against Mucor hiemalis in the rhizosphere of Pinus sylvestris. Mycorrhiza 13:41–47CrossRefPubMedGoogle Scholar
  56. Werner A, Zadworny M, Idzikowska K (2002) Interaction between Laccaria laccata and Trichoderma virens in co-culture and in the rhizosphere of Pinus sylvestris grown in vitro. Mycorrhiza 12:139–145CrossRefPubMedGoogle Scholar
  57. Wösten HAB, Scholtmeijer K (2015) Applications of hydrophobins: current state and perspectives. Appl Microbiol Biotech 99:1587–1597CrossRefGoogle Scholar
  58. Yakovlev I, Vaaje-Kolstad G, Hietala AM, Stefańczyk E, Solheim H, Fossdal CG (2012) Substrate-specific transcription of the enigmatic GH61 family of the pathogenic white-rot fungus Heterobasidion irregulare during growth on lignocellulose. Appl Microbiol Biotechnol 95:979–990PubMedCentralCrossRefPubMedGoogle Scholar
  59. Zadworny M, Werner A, Idzikowska K (2004) Behaviour of the hyphae of Laccaria laccata in the presence of Trichoderma harzianum in vitro. Mycorrhiza 14:401–405CrossRefPubMedGoogle Scholar

Copyright information

© German Mycological Society and Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • F. Sillo
    • 1
  • E. Zampieri
    • 1
  • L. Giordano
    • 1
  • G. Lione
    • 1
  • J. V. Colpaert
    • 2
  • R. Balestrini
    • 3
  • P. Gonthier
    • 1
    Email author
  1. 1.Department of Agricultural, Forest and Food Sciences (DISAFA)University of TorinoGrugliascoItaly
  2. 2.Centre for Environmental Sciences (CMK)Hasselt UniversityDiepenbeekBelgium
  3. 3.Institute for Sustainable Plant ProtectionCNRTorinoItaly

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