Use of Trichoderma , Pseudomonas and Bacillus spp. in IPM Programs

1. Use of Trichoderma, Pseudomonas and Bacillus spp. in IPM Programs Barry J. Jacobsen Dept. of Plant Sciences and Plant Pathology Montana State University ESA, 2012

2. Typically used as seed, tuber, rhizome, root stock, and soil treatments Widely used in IPM CRSP activities in South Asia

3. Many products in both developed and developing world

4. Widespread acceptance by farmers after comparison to farmer practices that include pesticides- increase yield, quality, earlier harvest, profitability-100s of trials

5. Mechanisms for Biological Control of Plant Pathogens • Antibiosis-Biological Control Agent (BCA) produces antibiotic substance that suppresses pathogen. Pseudomonas, Bacillus, Trichoderma • Competition for nutrients-BCA competes for resource in short supply that pathogen needs. e.g. Fe+++ , sugars and other needed for spore germination or growth. Pseudomonas • Niche occupation-BCA occupies infection niche • Predation • Parasitism. Trichoderma, Bacillus penetrans-nematode • Alter plant physiology (transpiration, water relations, growth hormones, nutrient uptake, N fixation) • Induced resistance. Pseudomonas, Bacillus, Trichoderma • MOST BCAs HAVE MULTIPLE MECHANISMS

6. Rhizosphere • PGPR-Plant growth promoting rhizobacteria and Trichoderma colonize Rhizosphere/endosphere • Pseudomondas • Enterobacter • Bacillus • Azospirillum • Many others

7. Colonize rhizosphere occupy infection courts Tie up critical nutrients needed by pathogens, produce growth promoting substances Rhizosphere colonist produce antibiotics,etc Antagonistic to pathogens Induce plant defense genes systemically

8. Colonization (CFU/g ) of sugarbeet by 341-16-5 from treated seed 15C=1.13 e5 24C=7.59e 5 7.1 e5 1.3 e5 2.1 e5 1.5 e5 2.1 e5 5.4 e5 Many PGPR are endophytes

9. Growth promotion by PGPR Control of root pathogens, growth regulators, improved nutrition, induced resistance?

12. Healthy seedling protected by PGPR Trichoderma Damping-off Mixing Trichoderma product in potting medium Planting Trichoderma/PGPR colonized seedlings

13. Begonias were grown in the greenhouse and inoculated with Botrytis cinerea under conditions optimal for the development of disease. Treatments left to right : untreated (Un), CaCl2, chlorothalonil (Fung), and the biocontrol agent Trichoderma hamatum T382 inoculated into the potting mix (T382). Hoitink, et al

14. Competition for nutrients • Many rhizosphere colonizing Pseudomonads (PGPR), Serratia, Erwinia (Pantoea) provide disease suppression by competition for Fe+++ with pathogens via chelation by siderophores (pyroverdin, pseudobactin, pyochelin). • Fusarium wilts of flax, carnation, Take-all, Thielaviopsis, Rhizoctonia, Sclerotiumrolfsii, Erwiniacarotovora, several patch diseases of turf, dollar spot of turf(Sclerotinia), melting out of turf (Drechslera-Bipolaris), DRBs, Deliteriousrhizobacteria that produce HCN, - may also be SAR/ISR signaling agents • Fireblight- • Pantoeaagglomerans competes for nutrients and niche with Erwiniaamylovora, • Pseudomonas fluorescens A 506(Blight Ban)- Fe+++ allows production of antibiotic antagonistic against E. amylovora that allows competition for site-this is used commercially • PGPR competes for nutrients (root exudates) with slower growing pathogens-fungistasis, chlamydospores, macroconidia, oospores

15. Altered physiology • Trichodermaharzianum/viridae-improve water and nutrient uptake- many references • Bacillus subtilis (Kodiak, other), Bacillus- pumillus GB 34 (YieldShield), Azospirillum, Pseudomonas • Auxins- Asghar et.al.,2002, Idris et.al. 2007 • Gibberelins-Joo, et.al.2005 • Cytokinins-Garcia de Salmone, 2001, Castro, et.al. 2008, Dobbelaere,et.al. 1999. • P uptake-Ramirez and Kloepper, 2010 (phytase activity) • Improved N utilization-Shoebitz et.al. 2009-nitrogenase and IAA • improved water relations

16. Phyllosphere/Phylloplane Biological Control • Environment for BCA in this environment is relatively hostile compared to rhizosphere / endosphere. • Physical environment: great flux in moisture, relative humidity, UV/IR radiation, paucity of nutrients that change with leaf age and time • Biological environment: competition with phylloplane colonists and invaders, plant responses and exudates vary with physiological age, genetics, etc • Majority of products are oriented to greenhouse or controlled storage situations where environment is more stabile. • Even here BCA performance has greater variability than chemicals • Vast majority of research has focused on Botrytis, powdery mildew and fruit storage molds. • Significant markets-high value of vegetables, ornamentals and fruit • Fewer registered pesticides-fungicide resistance problems • Lower costs to register BCAs in many countries • BCAs considered more acceptable to greenhouse workers (reentry periods) and to consumers

17. Pseudomonas syringae ESC 10/11 antibiosis and niche occupation

18. Induced Resistance=SAR,SIR and ISRCommon mechanism for Pseudomonas, Bacillus, Trichoderma • SAR-Systemic Acquired Resistance-SIR-Systemic Induced Resistance • Activation of master switch via salicyclic acid pathway signal –Classical PR-Proteins-Chitinases, βglucanases, proteinases, etc • ISR-Induced Systemic Resistance • Activation via jasmonic acid/ ethylene pathway -no classic PR-proteins but the defense compounds • Usually associated with PGPR(plant growth promoting rhizobacteria( Pseudomonas sp.)-insects Induced Resistance now we know that many biological inducers induce via salicylic acid, NPR-1 gene, jasmonic acid, ethylene or combination of these pathways

19. Induced resistance • Seed Treatments: Pseudomonas, Bacillus, Trichoderma-root diseases caused by fungi, nematodes-foliar diseases caused by bacteria, fungi, viruses • Foliar treatments: Bacillus mycoides-foliar diseases caused by bacteria, fungi, viruses-Root diseases caused by Pythium

20. PGPR induced resistance is a state of enhanced defensive capacity developed by a plant reacting to specific biotic or chemical stimuli Stimulus from PGPR/Trichoderma Stimulus Stimulus Stimulus

21. PGPR induced resistance is a state of enhanced defensive capacity developed by a plant reacting to specific biotic or chemical stimuli potentiated induction of stress-related genes enhanced resistance

22. SAR/ISR-Foliar Induction • Protective effects of SAR extend to all plant parts • Resistance is detectable 2-3 days post induction • Peaks 5-7 days post induction • Effective for ~14-20 days or longer • Suppresses many pathogens: fungi, bacteria, viruses Point of induction

23. Trichodermaharzianum/viridae Fungal Parasite Antibiotic producer Improved water and nutrient uptake Induced Systemic Resistance Inducer

24. Trichodermaviridae and antibiotic deficient mutants Trichoderma antibiotic deficient mutants still retain biocontrol activity

25. Mycoparasitism deficent mutant Trichodermamycoparasitismdeficient mutants still produce biocontrol

26. Phase 2: low MW Antibiotics receptors? Trichoderma host CWDEs The pre-contact events of mycoparasitim may also activate ISR in the plant Mycoparasitism and ISR Phase 1: high MW Antibiotics host Trichoderma CWDEs Cell wall degrading enzymes Lorito

27. PGPR- Viruses • 1996- Raupach et al. Two strains of PGPR induce ISR in cucumber and tomato against CMV • Some strains of Pseudomonas fluorescens, Bacillus pumilis, B. amyloliquefaciens, B subtilis, Kluyveracryocrescensrhizobacteria reduced CMV and Tomato Mottle geminivirus infection (50-70%), reduced symptoms and lengthened period from infection to symptom development-Zehender et al, 1999 • Bacillus globisporus, Pseudomonas fluorescens, Streptomycesgibsonii-30-60% reduction of tobacco necrosis virus local lesions in bean. Shoman, et al 2003 • Pseudomonas fluorescens- Barley Yellow Dwarf Mosaic- Mysusavenae-Wheat and Barley~50% reduced disease severity. Al Ani et al.2011

28. BmJ Virus Disease Controlmechanical transmission

29. PVY Greenhouse-mechanical transmission

30. 2010 Greenhouse PVY Aphid Transmission March-May Transferred 10 green peach Aphid/ plant from PVY infected potato- 20 replications summary of 3 experiments % infection ELISA

31. Hermiston, OR- Integrated PVY Management Plots Red flags- Russet Norkotah-Mazzama Borders

33. How does induced resistance reduce virus • Direct effect on insect vector-JA? • Love, et al., 2007 showed salicylic acid pathway involved in delayed symptoms and severity and alternative oxidase. • Ethylene/Jasmonic acid deficient mutants implicate ISR in reduced long distance spread in plant . • Lewsey et al., 2009 showed RNA silencing and salicylic acid mediated defense to restrict virus replication and movement. Jasmonic acid may have direct effect on aphid vector. • Data using salicylic acid, Acibenzolar-s-methyl-(Actigard,Bion)/ CMV TMV in tobacco, squash, Arabidopsis show reduced virus movement-cell to cell (delay symptom development) and systemic movement. IR involves mitochondrial enzyme alternate oxidase and RNA dependent RNA polymerase. • Mayers, et al 2005 : Madhusudham, et al., 2008

34. Conclusion • IR shown to delay symptom onset and reduce infection, disease severity, virus titer, virus movement or symptom severity for a wide range of viruses .. • Control levels are generally in the range of 30-80% and that the mode of action or efficacy differs remarkably by biological control agent and plant species. • May have direct effect on aphid vectors

35. Thank You &Happy Trails