Pseudomonas putida induces resistance to Fusarium oxysporum f. sp. radicis-lycopersici in tomato plants by activating expression of defense-related genes

Öz

Plant growth-promoting rhizobacteria (PGPR) may prevent attack from pathogenic microorganisms by eliciting induced systemic resistance (ISR). In the present work, Pseudomonas putida isolate TR21/1 showed significant biological control of tomato seedlings inoculated with Fusarium oxysporum f.sp. radicis-lycopersici (FORL). Here, the SA-responsive genes PR-1, PR-4, PR-6 and CH9 were downregulated upon induction of ISR by P. putida strain TR21/1 and induced when bacterized tomato roots were inoculated with FORL. This indicates that SAR involves the accumulation of SA-responsive genes but ISR does not. Similarly, expression of ET-regulated genes such as ACO1, ACO3, ACO4 were not induced in ISR-expressing tomato roots and P. putida treatment induced only ACO2 expression suggesting that ACO2 expression is involved in ISR-expressing tomato seedlings. In contrast, the infection of ISR expressing plants by FORL strongly induced ACO3, ACO2, and ACO1 indicating the transcriptional regulation of ACO genes in response to FORL attack which may be related to possible ethylene synthesis in response to pathogen. Here P. putida treatment increased ETR1 gene expression in roots and this induction was upregulated in presence of FORL indicating that ETR1 plays a role in the protection of plants against FORL by reducing ethylene sensitivity. Activation of SA-and ET- regulated genes in bacterized plants in the presence of FORL implies that not only SA but other signals as well, may play an important role in inducing resistance.

Anahtar Kelimeler

• Pseudomonas putida
• systemic acquired resistance (SAR)
• induced systemic resistance (ISR)
• Fusarium oxysporum f.sp. radicis-lycopersici (FORL)
• gene expression

Pseudomonas putida, domates bitkilerinde savunma ile ilgili genlerin ekspresyonunu aktive ederek Fusarium oxysporum f.sp. radicis-lycopersici'ye karşı direnci indüklemesi

Öz

Bitki büyümesini teşvik eden rizobakteriler (PGPR), indüklenmiş sistemik direnci (ISR) sağlayarak patojenik mikroorganizmaların saldırısını önleyebilir. Mevcut çalışmada, P. putida izolatı TR21/1, Fusarium oxysporum f.sp. radicis-lycopersici (FORL) ile enfekte olmuş domates fidelerinin önemli ölçüde biyolojik kontrolünü sağlamıştır. Burada SA-yanıt veren genler PR-1, PR-4, PR-6 ve CH9 ISR'nin P. putida streyn TR21/1 tarafından indüklenmesi üzerine ifadeleri baskılanmış ve bakterili domates kökleri Fusarium oxysporum f.sp. radicis-lycopersici ile enfekte edildiğinde bu genlerin ifadeleri indüklenmiştir. Bu sonuçlar, SAR'ın SA'ya yanıt veren genlerin birikimini içerdiğini ancak ISR'nin içermediğini göstermiştir. Benzer şekilde, ACO1, ACO3, ACO4 gibi ET tarafından düzenlenen genlerin ifadeleri, ISR gösteren domates köklerinde indüklenmemiş ve P. putida uygulaması, sadece ACO2 ekspresyonunu indüklemiştir. Buna karşılık, ISR gösteren bitkilerin FORL ile enfeksiyonu, ACO3, ACO2 ve ACO1 genlerinin ifadelerini güçlü bir şekilde indüklemesi FORL saldırısına yanıt olarak olası bir etilen sentezi ile ilgili ACO genlerinin transkripsiyonel düzenlemesini gösterir. Burada P. putida uygulaması köklerde ETR1 geninin ifadesini indüklemiş ve FORL ile inokülasyon bu genin ifadesinin indüksiyonunu daha da artırmıştır. Bu sonuçlar, ETR1'in etilen duyarlılığını azaltarak bitkilerin FORL'e karşı korunmasında rol oynadığını göstermiştir. FORL varlığında bakteri uygulanmış bitkilerde SA ve ET tarafından düzenlenen genlerin aktivasyonu, sadece SA'nın değil, diğer sinyallerin de direncin indüklenmesinde önemli bir rol oynayabileceği anlamına gelmektedir.

Anahtar Kelimeler

• Pseudomonas putida
• Fusarium oxysporum f.sp. radicis-lycopersici (FORL)
• Sistemik kazanılmış direnç (SAR)
• indüklenmiş sistemik direnç (ISR)
• gen ekspresyonu

Kaynakça

1. Akhgar, A., M. Arzanlou, P. Bakker, and M. Hamidpour. 2014. Characterization of 1- amino cyclopropane-1-carboxylate (ACC) deaminase-containing Pseudomonas spp. in the rhizosphere of salt-stressed canola. Pedosphere 24(4): 461–8.
2. Anandham, R., P.I. Gandhi, M. Madhaiyan, and T. Sa. 2008. Potential plant growth promoting traits and bioacidulation of rock phosphate by thiosulfate oxidizing bacteria isolated from crop plants. J. Basic Microbiol. 48(6): 439-447. doi:10.1002/jobm.200700380
3. Ardakani, S. S., A. Heydari, L. Tayebi, and M. Mohammadi. 2010. Promotion of cotton seedings growth characteristics by development and use of new bioformulations. Int. J. Botany 6: 95-100.
4. Arora, N. K., S. C. Kang, and D. K. Maheshwari. 2001. Isolation of sidero-phore-producing strains of Rhizobium meliloti and their biocon-trol potential against Macrophomina phaseolina that causescharcoal rot of groundnut. Curr Sci 81(6):673–677.
5. Arshad M., and W.T. Frnakenberger. 1992. Microbial production of plant growth regulators. pp 307-347. In: Jr. MFB (Ed) Soil Microbial Ecology. Marcel Dekker New York.
6. Baysal, Z., F. Uyar, M. Doğru, and H. Alkan. 2008. Production of extracellular alkaline α-amylase by solid state fermentation with a newly isolated Bacillus sp. Prep. Biochem. Biotechnol. 38(2): 184-190. doi:10.1080/10826060701885167.
7. Bolwerk, A., A. L. Lagopodi, A. H. M. Wijfjes, G. E. M. Lamers, T. F. C. Chin-A-Woeng, B. J. J. Lugtenberg, and G. V. Bloemberg. 2003. Interactions in the tomato rhizosphere of two pseudomonas biocontrol strains with the phytopathogenic fungus Fusarium oxysporum f. sp. radicis-lycopersici. Mol. Plant-Microbe Interact. 16(11): 983-993. doi:10.1094/ mpmi.2003.16.11.983
8. Bray E. 1988. Drought- and ABA-Induced changes in polypeptide and mRNA accumulation in tomato leaves. Plant Physiol. 88: 1210-1214.

9. Çakır, B., A. Gül, L. Yolageldi, and H. Özaktan. 2014. Response to Fusarium oxysporum f.sp. radicis-lycopersici in tomato roots involves regulation of SA- and ET-responsive gene expressions. Eur. J. Plant Pathol. 139: 379–391. doi:DOI 10.1007/s10658-014-0394-9.
10. Callan, N. W., D. E. Mathre, and J. B. Miller. 1990. Bio-priming seed treatment for biological control of Pythium ultimum preemergence damping-off in sh2 sweet corn. Plant Dis. 74(5): 368-372.
11. Ciardi, J. A., D. M. Tieman, S. T. Lund, J. B. Jones, R.E. Stall, and H.J. Klee. 2000. Response to Xanthomonas campestris pv.vesicatoria in tomato involves regulation of ethylene receptor gene expression. Plant Physiol. 123(1): 81-92.
12. Cohn, J. R., and G. B. Martin. 2005. Pseudomonas syringae pv. tomato type III effectors AvrPto and AvrPtoB promote ethylene-dependent cell death in tomato. The Plant J. 44 (1): 139-154. doi:10.1111/j.1365-313X.2005.02516.x.
13. Cummings, S., P. Gyaneshwar, P. Vinuesa, F. T. Farruggia, M. Andrews, D. Humphry, G. Elliott, A. Nelson, C. Orr, D. Pettitt, G. Shah, S. Santos, H. Krishnan, D. Odee, F. Moreira, J. I. Sprent, J. Young, and E. James. 2009. Nodulation of Sesbania species by Rhizobium (Agrobacterium) strain IRBG74 and other rhizobia. Environ. Microbiol. 11: 2510-2525.
14. Datnoff, L. E., S. Nemec, and K. Pernezny. 1995. Biological Control of Fusarium crown and root rot of tomato in Florida using Trichoderma harzianum and Glomus intraradices. Biol. Contr. 5(3): 427-431. doi:http:// dx.doi.org/10.1006/bcon.1995.1051.
15. de Laat, A.M.M., and L.C.van Loon. 1983. The relationship between stimulated ethylene production and symptom expression in virus-infected tobacco leaves. Physiological Plant Pathol. 22(2): 261-273. doi:http://dx.doi.org/10.1016/S0048-4059(83)81014-5.
16. Delaney, T. P., S. Uknes, B. Vernooij, L. Friedrich, K. Weymann, D. Negrotto, T. Gaffney, M. Gut-Rella, H. Kessmann, E. Ward, and J. Ryals. 1994. A central role of salicylic acid in plant disease resistance. Science 266(5188): 1247-1250.
17. Freeman, B. C., and G. A. Beattie. 2008. An overview of plant defenses against pathogens and herbivores. J. Plant Pathol. Microbiol. 94.
18. Galindo-Gonzalez, L., and M. K. Deyholos. 2016. RNA-seq transcriptome response of flax (Linum usitatissimum L.) to the pathogenic fungus Fusarium oxysporum f. sp. lini. Frontier in Plant Sci. 7: 1766.
19. Gul, A., Ozaktan, H., Yolageldi, L., Cakir, B., Sahin, and S. M. Akat. 2012. Effect of rhizobacteria on yield of hydroponically grown tomato plants. Acta Hortic. (ISHS) 952: 777-784.
20. Haas, D., and G. Defago. 2005. Biological control of soil-borne pathogens by fluorescent pseudomonads. Nat Rev Microbiol. 3(4): 307-319.
21. Horinouchi, H., N. Katsuyama, Y. Taguchi, and M. Hyakumachi. 2008. Control of Fusarium crown and root rot of tomato in a soil system by combination of a plant growth-promoting fungus, Fusarium equiseti, and biodegradable pots. Crop Protec. 27(35): 859-864. doi:http://dx.doi.org/10.1016/j.cropro.2007. 08.009.
22. Iavicoli A., E. Boutet, A. Buchala, and J. P. Metraux. 2003. Induced systemic resistance in Arabidopsis thaliana in response to root inoculation with Pseudomonas fluorescens CHA0. Mol Plant–Microbe Interac. 16: 851–858.
23. Janssen, J., N. Weyens, S. Croes, B. Beckers, L. Meiresonne, and P. Van Peteghem. 2015. Phytoremediation of metal contaminated soil using willow: exploiting plant-associated bacteria to improve biomass production and metal uptake. Int. J. of Phytoremediation 17: 1123–1136. doi: 10.1080/ 15226514.2015.1045129.
24. Jones, J. B., J. P. Jones, R.E. Stall, and T. A. Zitter 1991. Compendium of tomato diseases. APS Press (The American Pathological Society Press) 73.
25. Kamilova, F., S. Validov, T. Azarova, I. Mulders, and B. Lugtenberg. 2005. Enrichment for enhanced competitive plant root tip colonizers selects for a new class of biocontrol bacteria. Environ. Microbiol. 7(11): 1809-1817. doi:10.1111/j.1462-2920.2005. 00889.x.
26. Katagiri, F., and K. Tsuda. 2010. Understanding the plant immune system. Mol plant-microbe interac. MPMI. 23: 1531-1536. doi:10.1094/MPMI-04-10-0099.
27. Kavroulakis, N., K. K. Papadopoulou, S. Ntougias, G. I. Zervakis, and C. Ehaliotis. 2006. Cytological and other aspects of pathogenesis-related gene expression in tomato plants grown on a suppressive compost. Ann. Bot. 98(3): 555-564. doi:10.1093/aob/mcl149.
28. Kloepper, J. W. 1980. Effects of rhizosphere colonization by plant growthpromoting rhizobacteria on potato plant development and yield. Phytopathol. 70:1078–1082.
29. Knoester, M., C. M. J. Pieterse, J. F. Bol, and L. C. Van Loon. 1999. Systemic resistance in Arabidopsis induced by rhizobacteria requires ethylene-dependent signaling at the site of application. Mol Plant-Microbe Interac. 12(8): 720-727. doi:10.1094/mpmi.1999.12.8.720.
30. Lawton, K., K. Weymann, L. Friedrich, B. Vernooij, S. Uknes, and J. Ryals. 1996. Systemic acquired resistance in Arabidopsis requires salicylic acid but not ethylene. Mol. Plant Microbe Interact. 8(6): 863-870.
31. Leeman, M., J. A. Vanpelt, F. M. Denouden, M. Heinsbroek, P. Bakker, and B. Schippers. 1995. Induction of systemic resistance against fusarium-wilt of radish by lipopolysaccharides of Pseudomonas-fluorescens. Phytopathology 85(9): 1021-1027. doi:10.1094/ Phyto-85-1021.
32. Lelliott, R. A., and D. E. Stead. 1987. Methods for the diagnosis of bacterial diseases of plants. pp.11. In: T. F. Preece (Ed). Methods in Plant Pathology. Blackwell Scientific Publications Oxford.
33. Lund, S. T., R. E. Stall, and H. J. Klee. 1998. Ethylene regulates the susceptible response to pathogen infection in tomato. The Plant Cell Online 10(3): 371-382.
34. Malamy, J., J. P. Carr, D. F. Klessig, and I. Raskin. 1990. Salicylic acid: A likely endogenous signal in the resistance response of tobacco to viral infection. Science 250 (4983): 1002-1004.
35. Menzies, J., C. Koch, and F. Seywerd. 1990. Additions to the host range of Fusarium oxysporum f.sp. radicis-lycopersici. Plant Dis. 74: 569-572.
36. Minuto, A,. D. Spadaro, A. Garibaldi, and M. L. Gullino. 2006. Control of soilborne pathogens of tomato using a commercial formulation of Streptomyces griseoviridis and solarization. Crop Prot. 25(5): 468-475. doi:http://dx.doi.org/10.1016/j.cropro.2005.08.001.
37. Murphy, J.F., M. S. Reddy, C-M. Ryu, J. W. Kloepper, and R. Li. 2003. Rhizobacteria-mediated growth promotion of tomato leads to protection against cucumber mosaic virus. Phytopathology 93(10): 1301-1307. doi:10.1094/phyto.2003.93.10.1301
38. Nutter, F. W., C.G.Warren, O. S. Wells, and W.E. MAchardy. 1978. Fusarium foot and root rot of tomato in New Hampshire. Plant Dis Rep. 62: 976-978.
39. Ortiz-Castro, R., H. A. Contreras-Cornejo, L. Macias-Rodriguez, and J. Lopez-Bucio. 2009. The role of microbial signals in plant growth and development. Plant Signal Behav. 4(8): 701-712.
40. Pieterse, C. M. J., S. C. M. van Wees, J. A. van Pelt, M. Knoester, R. Laan, H. Gerrits, P. J. Weisbeek, and L.C. van Loon. 1998. A novel signaling pathway controlling induced systemic resistance in Arabidopsis. The Plant Cell Online 10(9): 1571-1580.
41. Pieterse, C. M., C. Zamioudis, R. L. Berendsen, D. M. Weller, S. C. Van Wees, and P.A. Bakker. 2014. Induced systemic resistance by beneficial microbes. Ann Rev Phytopathol. 52: 347–375.
42. Ryu, C-M., J. Kim, O. Choi, S. H. Kim, and C.S. Park. 2006. Improvement of biological control capacity of Paenibacillus polymyxa E681 by seed pelleting on sesame. Biol Control 39(3): 282-289. doi:http://dx. doi.org/10.1016/j.biocontrol.2006.04.014.
43. Ryu, C-M., M. A. Farag, C-H. Hu, M. S. Reddy, J. W. Kloepper, and P.W. Par. 2004. Bacterial volatiles induce systemic resistance in Arabidopsis. Plant Physiol. 134(3): 1017-1026.
44. Schaad, N., J. Jones, and W. Chun. 2001. Laboratory guide for identification of plant pathogenic bacteria. (APS Press ):St. Paul, Minnesota, USA.
45. Sivan, A., O. Ucko, and I. Chet. 1987. Biological control of Fusarium crown rot of tomato Trichoderma harzianum undr field contidions. Plant Dis.71: 587-592.
46. Stockwell, V., M. Kawalek, L. Moore, and J. Loper. 1996. Transfer to pAgK84 from the biocontrol agent Agrobacterium radiobacter K84 to A. tumefaciens under field conditions. Phytopathology 86: 31-37.
47. Suslow, T. 1982. Rhizobacteria of sugar beets: Effects of seed Application and root colonization on yield. Phytopathology 72: 199-206. 10.1094/Phyto-72-199.
48. Timmusk, S., I. A. AbdEl-Daim, L. Copolovici, T. Tanilas, A. Kännaste, and Ü. Niinemets. 2014. Drought-tolerance of wheat improved by rhizosphere bacteria from harsh environments: enhanced biomass production and reduced emissions of stress volatiles. PLoS ONE 9. e96086. doi: 10.1371/journal. pone. 0096086
49. Ton, J., J. A. Van Pelt, L. C. Van Loon, and C. M. J. Pieterse. 2002. Differential effectiveness of salicylate-dependent and jasmonate/ethylene-dependent induced resistance in Arabidopsis. Mol Plant Microbe Interac. 15(1): 27-34. doi:10.1094/mpmi.2002.15.1.27.
50. Validov, S. Z., F. Kamilova, and B. J. J. Lugtenberg. 2009. Pseudomonas putida strain PCL1760 controls tomato foot and root rot in stonewool under industrial conditions in a certified greenhouse. Biol Control. 48(1): 6-11. doi:http://dx.doi.org/10.1016/j.biocontrol. 2008.09.010.
51. Validov, S., F. Kamilova, S. Qi, D. Stephan, J. J. Wang, N. Makarova, and B. Lugtenberg. 2007. Selection of bacteria able to control Fusarium oxysporum f. sp. radicis-lycopersici in stonewool substrate. J Appl. Microbiol. 102(2): 461-471. doi:10.1111/j.1365-2672.2006.03083.x.
52. van Loon, L. C., P. A. H. M. Bakker, and C. M. J. Pieterse. 1998. Systemic resistance induced by rhizosphere bacteria. Ann Rev Phytopathol. 36(1): 453-483. doi:10.1146/annurev.phyto.36.1.453.
53. van Wees, S. C. M., E. A. M. de Swart, J. A. van Pelt, van L. C. Loon, and C. M. J. Pieterse. 2000. Enhancement of induced disease resistance by simultaneous activation of salicylate- and jasmonate-dependent defense pathways in Arabidopsis thaliana. Proc. Natl. Acad. Sci. U.S.A. 97(15): 8711-8716.
54. Van Wees, S. C. M., S. Van der Ent, and C. M. J. Pieterse. 2008. Plant immune responses triggered by beneficial microbes. Curr. Opin. Plant Biol. 11(4): 443-448. doi:http://dx.doi.org/10.1016/j.pbi.2008.05.005.
55. Wojtasik, W., A. Kulma, A. Boba, and J. Szopa. 2014. Oligonucleotide treatment causes flax β-glucanase up-regulation via changes in gene-body methylation. BMC Plant Biol 14: 261. https://doi.org/10.1186/ s12870-014-0261-z.
56. Yang, J., J. W. Kloepper, and C. M. Ryu. 2009. Rhizosphere bacteria help plants tolerate abiotic stress. Trends in Plant Sci. 14: 1-4.
57. Zahir, Z. A., M. Arshad, and Jr W.T. Frankenberger. 2003. Plant growth promoting rhizobacteria: applications and perspectives in agriculture. Advances in Agronomy 81: 97-168.
58. Zhang, S., A-L. Moyne, M. Reddy, and J. Kloepper. 2002. The role of salicylic acid in induced systemic resistance elicited by plant growth-promoting rhizobacteria against blue mold of tobacco. Biol. Control. 25: 288-296.