Advertisement

Protoplasma

, Volume 254, Issue 5, pp 1887–1901 | Cite as

Probing the contractile vacuole as Achilles’ heel of the biotrophic grapevine pathogen Plasmopara viticola

  • Viktoria Tröster
  • Tabea Setzer
  • Thomas Hirth
  • Anna Pecina
  • Andreas Kortekamp
  • Peter Nick
Original Article

Abstract

The causative agent of Grapevine Downy Mildew, the oomycete Plasmopara viticola, poses a serious threat to viticulture. In the current work, the contractile vacuole of the zoospore is analysed as potential target for novel plant protection strategies. Using a combination of electron microscopy, spinning disc confocal microscopy, and video differential interference contrast microscopy, we have followed the genesis and dynamics of this vacuole required during the search for the stomata, when the non-walled zoospore is exposed to hypotonic conditions. This subcellular description was combined with a pharmacological study, where the functionality of the contractile vacuole was blocked by manipulation of actin, by Na, Cu, and Al ions or by inhibition of the NADPH oxidase. We further observe that RGD peptides (mimicking binding sites for integrins at the extracellular matrix) can inhibit the function of the contractile vacuole as well. Finally, we show that an extract from Chinese liquorice (Glycyrrhiza uralensis) proposed as biocontrol for Downy Mildews can efficiently induce zoospore burst and that this activity depends on the activity of NADPH oxidase. The effect of the extract can be phenocopied by its major compound, glycyrrhizin, suggesting a mode of action for this biologically safe alternative to copper products.

Keywords

Contractile vacuole Downy Mildew Glycyrrhizin Plasmopara viticola RGD peptides 

Notes

Acknowledgements

We gratefully acknowledge Joachim Daumann and Kerstin Huber (Botanical Garden of the Karlsruhe Institute of Technology) for efficient support with the plant material. Also, we acknowledge Prof. Dr. Otmar Spring and Dr. Javier Goméz (University of Hohenheim) for kindly providing single-sporangia strains of P. viticola. This work was supported by the VITIFUTUR Interreg V Upper Rhine project co-financed by the European Union/European Regional Development Fund (ERDF) and the German Federal Agency for Agriculture (Programme for Sustainable Agriculture, BÖLN).

Compliance with ethical standards

Funding

This study was supported by funds from the BACCHUS Interreg IV Upper Rhine project co-financed by the European Union/European Regional Development Fund (ERDF) and the German Federal Agency for Agriculture (Programme for Sustainable Agriculture, BÖLN).

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

709_2017_1123_MOESM1_ESM.avi (92 kb)
Supplemental movie S1 (AVI 92 kb)
709_2017_1123_MOESM2_ESM.avi (1.2 mb)
Supplemental movie S2 (AVI 1278 kb)

References

  1. Allen RN, Newhook FJ (1973) Chemotaxis of zoospores of Phytophthora cinnamomi to ethanol in capillaries of soil pore dimensions. Transact Brit Mycol Soc 61:287–302CrossRefGoogle Scholar
  2. Baluška F, Šamaj J, Wojtaszek P, Volkmann D, Menzel D (2003) Cytoskeleton plasma membrane-cell wall continuum in plants. Emerging links revisited. Plant Physiol 133:482–491CrossRefPubMedPubMedCentralGoogle Scholar
  3. Bartlett DW, McClough J, Godwin JR, Hall AA, Hamer M, Parr-Dobrzanski B (2002) Thestrobilurin fungicides. Pest Manag Sci 58:649–662CrossRefPubMedGoogle Scholar
  4. Beakes GW, Glocklin SL, Sekimoto S (2012) The evolutionary phylogeny of the oomycete “fungi”. Protoplasma 249:3–19CrossRefPubMedGoogle Scholar
  5. Becker B, Doan JM, Wustman B, Carpenter EJ, Chen L, Zhang Y, Wong GK-S, Melkonian M (2015) The origin and evolution of the plant cell surface: algal integrin-associated proteins and a new family of integrin-like cytoskeleton-ECM linker proteins. Genome Biol Evolution 7:1580–1589CrossRefGoogle Scholar
  6. Bleyer G, Kassemeyer H-H, Breuer M, Krause R, Viret O, Dubuis P. H, Fabre A.L, Bloesch B, Siegfried W, Naef A, Huber M (2011) “VitiMeteo”—a future-oriented forecasting system for viticulture. IOBC/Wprs Bulletin 67:69–77Google Scholar
  7. Brun LA, Maillet J, Richarte J, Herrmann P, Remy JC (1998) Relationships between extractable copper, soil properties and copper uptake by wild plants in vineyard soils. Environ Pollut 102:151–161CrossRefGoogle Scholar
  8. Canut H, Carrasco A, Galaud JP, Cassan C, Bouyssou H, Vita N, Ferrara P, Pont-Lezica R (1998) High affinity RGD-binding sites at the plasma membrane of Arabidopsis thaliana links the cell wall. Plant J 16:63–71CrossRefPubMedGoogle Scholar
  9. Chen HR, Sheu SJ (1993) Determination of glycyrrhizin and glycyrrhetinic acid in traditional Chinese medicinal preparations by capillary electrophoresis. J Chromatogr A 29:184–188CrossRefGoogle Scholar
  10. Chen J, Dai G, Gu Z, Miao Y (2002) Inhibition effect of 58 plant extracts against grape downy mildew (Plasmopara viticola). Natural Product Res Development 14:9–13Google Scholar
  11. Chen WJ, Delmotte F, Richard-Cervera S, Douence L, Greif C, Corio-Costet MF (2007) At least two origins of fungicide resistance in grapevine downy mildew populations. Appl Environm Microbiol 73:5162–5172CrossRefGoogle Scholar
  12. Chitcholtan K, Garrill A (2005) A beta4 integrin-like protein co-localises with a phosphotyrosine containing protein in the oomycete Achlya bisexualis: inhibition of tyrosine phosphorylation slows tip growth. Fung Genet Biol 42:534–545CrossRefGoogle Scholar
  13. Cho CW, Fuller MS (1989) Observations of the water expulsion vacuole of Phytophthora palmivora. Protoplasma 149:47–55CrossRefGoogle Scholar
  14. Dagostin S, Formolo T, Giovannini O, Pertot I, Schmitt A (2010) Salvia officinalis extract can protect grapevine against Plasmopara viticola. Plant Dis 94:575–580CrossRefGoogle Scholar
  15. Dagostin S, Schärer HJ, Pertot I, Tamm L (2011) Are there alternatives to copper for controlling grapevine downy mildew in organic viticulture? Crop Prot 30:776–788CrossRefGoogle Scholar
  16. Dercks W, Buchenauer H (1987) Comparative studies on the mode of action of aluminium ethyl phosphite in four Phytophthora species. Crop Prot 6:82–89CrossRefGoogle Scholar
  17. Eggenberger K, Sanyal P, Hundt S, Wadhwani P, Ulrich AS, Nick P (2017) Challenge integrity: the cell-permeating peptide BP100 interferes with the actin-auxin oscillator. Plant Cell Physiol 58:71–85PubMedGoogle Scholar
  18. Eibach R, Zyprian E, Welter L, Töpfer R (2007) The use of molecular markers for pyramiding resistance genes in grapevine breeding. Vitis 46:120–124Google Scholar
  19. Gay JL, Greenwood AD, Heath IB (1971) The Formation and Behaviour of Vacuoles (Vesicles) during Oosphere Development and Zoospore Germination in Saprolegnia. Microbiology 65:233–241Google Scholar
  20. Gessler C, Pertot I, Perazzolli M (2011) Plasmopara viticola: a review of knowledge on downy mildew of grapevine and effective disease management. Phytopathol Mediterr 50:3–44Google Scholar
  21. Giraud F, Molitor D, Bleunven M, Evers D (2013) Fungicide sensitivity profiles of the Plasmopara viticola populations in the Luxembourgian grape-growing region. J Plant Pathol S1:55–62Google Scholar
  22. Gómez-Zeledón J, Zipper R, Spring O (2013) Assessment of phenotypic diversity of Plasmopara viticola on Vitis genotypes with different resistance. Crop Prot 54:221–228CrossRefGoogle Scholar
  23. Hardham AR (2005) Phytophthora cinnamomi. Mol Plant Pathol 6:589–604CrossRefPubMedGoogle Scholar
  24. Heath IB, Harold RL (1992) Actin has multiple roles in the formation and architecture of zoospores of the oomycetes, Saprolegnia ferax and Achlya bisexualis. J Cell Science 102:611–627Google Scholar
  25. Heumann HG (1992) Microwave-stimulated glutaraldehyde and osmium tetroxide fixation of plant tissue: ultrastructural preservation in seconds. Histochemistry 97:341–347CrossRefPubMedGoogle Scholar
  26. Hostetter MK (2000) RGD-mediated adhesion in fungal pathogens of humans, plants and insects. Current Op Microbiol 3:344–348CrossRefGoogle Scholar
  27. Hyde GJ, Hardham AR (1993) Microtubules regulate the generation of polarity in zoospores of Phytophthora cinnamomi. Eur J Cell Biol 62:75–85PubMedGoogle Scholar
  28. Hyde GJ, Lancelle S, Hepler PK, Hardham AR (1991) Freeze substitution reveals a new model for sporangial cleavage in Phytophthora, a result with implications for cytokinesis in other eukaryotes. J Cell Sci 100:735–746PubMedGoogle Scholar
  29. Ismail A, Takeda S, Nick P (2014) Life and death under salt stress: same players, different timing? J Exp Bot 65:2963–2979CrossRefPubMedGoogle Scholar
  30. Jürges G, Kassemeyer H-H, Dürrenberger M, Düggelin M, Nick P (2009) The mode of interaction between Vitis and Plasmopara viticola Berk. & Curt. Ex de Bary depends on the host species. Plant Biol 11:886–898CrossRefPubMedGoogle Scholar
  31. Kaminskyj SG, Heath IB (1995) Integrin and spectrin homologues, and cytoplasm-wall adhesion in tip growth. J Cell Sci 108:849–856PubMedGoogle Scholar
  32. Kiefer B, Riemann M, Büche C, Kassemeyer HH, Nick P (2002) The host guides morphogenesis and stomatal targeting in the grapevine pathogen Plasmopara viticola. Planta 215:387–393CrossRefPubMedGoogle Scholar
  33. Kleinig H, Sitte P (1986) Zellbiologie—ein Lehrbuch. Gustav-Fischer, Stuttgart-New YorkGoogle Scholar
  34. Koch E, Enders M, Ullrich C, Molitor D, Berkelmann-Löhnertz B (2013) Effect of Primula root and other plant extracts in infection structure formation of Phyllosticta ampelicida (asexual stage of Guignardia bidwellii) and on black rot disease of grapevine in the greenhouse. J Plant Diseases Protection 120:26–33CrossRefGoogle Scholar
  35. Koning AJ, Lum PY, Williams JM, Wright R (1993) DiOC6 staining reveals organelle structure and dynamics in living yeast cells. Cell Mot Cytoskelet 25:111–128CrossRefGoogle Scholar
  36. Kortekamp A (2003) Leaf surface topography does not mediate tactic response of Plasmopara-zoospores to stomata. J Applied Bot 77:41–46Google Scholar
  37. Kutschera U, Hossfeld U (2012) Physiological phytopathology-origin and evolution of a scientific discipline. J Appl Bot 85:1–5Google Scholar
  38. Liu Q, Qiao F, Ismail A, Chang X, Nick P (2013) The plant cytoskeleton controls regulatory volume increase. BBA Membranes 1828:2111–2120CrossRefGoogle Scholar
  39. Mitchell HJ, Hardham AR (1999) Characterisation of the water expulsion vacuole in Phytophthora nicotianae zoospores. Protoplasma 206:118–130CrossRefGoogle Scholar
  40. Mitchell HJ, Kovac KA, Hardham AR (2002) Characterisation of Phytophthora nicotianae zoospore and cyst membrane proteins. Mycol Res 106:1211–1223CrossRefGoogle Scholar
  41. Müller K, Sleumer H (1934) Biologische Untersuchungen über die Peronosporakrankheit des Weinstockes mit besonderer Berücksichtigung ihrer Bekämpfung nach der Inkubationskalendermethode. Landwirtschaftl Jahrb Z Wissenschaftl Landwirtschaft 79:509–576Google Scholar
  42. Nick P (2011) Mechanics of the cytoskeleton. In: Wojtaszek P (ed) Mechanical integration of plant cells and plants. Springer, Berlin-Heidelberg, pp 53–90CrossRefGoogle Scholar
  43. Patterson DJ (1980) Contractile vacuoles and associated structures: their organization and function. Biol Rev 55:1–46CrossRefGoogle Scholar
  44. Pickard BG (2008) “Second extrinsic organizational mechanism” for orienting cellulose: modeling a role for the plasmalemmal reticulum. Protoplasma 233:7–29CrossRefPubMedGoogle Scholar
  45. Riemann M, Büche C, Kassemeyer HH, Nick P (2002) Microtubules and actin microfilaments guide the establishment of cell polarity during early development of the wine pathogen Plasmopara viticola. Protoplasma 219:13–22CrossRefPubMedGoogle Scholar
  46. Rouxel M, Mestre P, Comont G, Lehman BL, Schilder A, Delmotte F (2013) Phylogenetic and experimental evidence for host-specialized cryptic species in a biotrophic oomycete. New Phytol 197:251–263CrossRefPubMedGoogle Scholar
  47. Ruoslahti E (1996) RGD and other recognition sequences for integrins. Annu Rev Cell Develop Biol 12:697–715CrossRefGoogle Scholar
  48. Schaubschläger WM, Becker G, Mazur G, Gödde M (1994) Occupational sensitization to Plasmopara viticola. J Allergy Clinical Immunol 93:457–463CrossRefGoogle Scholar
  49. Scherf A, Treutwein J, Kleeberg H, Schmitt A (2012) Efficacy of leaf extract fractions of Glycyrrhiza glabra L. against downy mildew of cucumber (Pseudoperonospora cubensis). Eur J Plant Pathol 134:55–762CrossRefGoogle Scholar
  50. Schuster C, Konstantinidou-Doltsinis S, Schmitt A (2010) Glycyrrhiza glabra extract protects plants against important phytopathogenic fungi. Commun Agric Appl Biol Sci 75:531–540PubMedGoogle Scholar
  51. Schützendübel A, Polle A (2002) Plant responses to abiotic stresses: heavy metal-induced oxidative stress and protection by mycorrhization. J Exp Bot 53:1351–1365PubMedGoogle Scholar
  52. Van Zwieten M, Stovold G, Van Zwieten L (2004) Literature review and inventory of alternatives to copper for disease control in the Australian organic industry. A report for the Rural Industries Research and Development Corporation. RIRDC Project DAN-208A. ISBN 0 7347 1590 0Google Scholar
  53. Williams MG, Magarey PA, Sivasithamparam K (2007) Effect of temperature and light intensity on early infection behaviour of a Western Australian isolate of Plasmopara viticola, the downy mildew pathogen of grapevine. Austral Plant Pathol 36:325–331CrossRefGoogle Scholar
  54. Yokozawa T, Liu ZW, Chen CP (2000) Protective effects of Glycyrrhizae radix extract and its compounds in a renal hypoxia (ischemia)-reoxygenation (reperfusion) model. Phytomedicine 6:439–445CrossRefPubMedGoogle Scholar
  55. Yu ZL, Zhang JG, Wang XC, Chen J (2008) Excessive copper induces the production of reactive oxygen species, which is mediated by phospholipase D, nicotinamide adenine dinucleotide phosphate oxidase and antioxidant systems. J Integr Plant Biol 50:157–167CrossRefPubMedGoogle Scholar
  56. Zaban B, Maisch J, Nick P (2013) Dynamic actin controls polarity induction de novo in protoplasts. J Integr Plant Biol 55:142–159CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Wien 2017

Authors and Affiliations

  • Viktoria Tröster
    • 1
  • Tabea Setzer
    • 1
  • Thomas Hirth
    • 1
  • Anna Pecina
    • 1
  • Andreas Kortekamp
    • 2
  • Peter Nick
    • 1
  1. 1.Molecular Cell BiologyBotanical Institute Karlsruhe Institute of Technology (KIT)KarlsruheGermany
  2. 2.Institute of Plant Protection State Education and Research Center (DLR) RheinpfalzNeustadtGermany

Personalised recommendations