Biostimulant Research in Turfgrass: Fertilizer or Other?

Biostimulant Research in Turfgrass: Fertilizer or Other? Erik H. Ervin, Ph.D. Professor, Turfgrass Culture & Physiology Crop and Soil Environmental Sciences Department; eervin@vt.edu VIRGINIA TECH August 6, 2015: Denver

North America Biostimulant Coalition Definition, 2013 Substances (derived from natural or biological sources), including microorganisms, that are applied to plants, seeds, soil or other growing media that may enhance the plant s ability to assimilate applied nutrients or provide benefits to plant growth and development.

Schmidt and Ervin definition, 2002 A biostimulant is an organic material that, when applied in small quantities, enhances plant growth and development such that the response cannot be attributed to application of traditional plant nutrients

Ag biostimulant review paper, 4 types Calvo et al., 2014. Plant and Soil 383:3-41. 1) Microbial inoculants 2) Humic & fulvic acids 3) Protein hydrolysates & amino acids 4) Seaweed extracts

Labels: Turf Biostimulants 100% Kelp Extract Assume 0.5% N label rate 6 oz/1000 ft 2 = 0.10 lb N/A Standard rates: 10 to 50 lb N/A

Labels: Turf Biostimulants A 3-0-20 liquid fertilizer, with micronutrients (7% Fe), plus 22 million cfu/g of Bacillus spp. Humic acid, 3% Seaweed extract, 4.1% At label rate 0.22 lb N/A applied; 1.5 lb K 2 O/A

Labels: Turf Biostimulants Contains no humate, or seaweed extract

Composition of Ascophyllum nodosum alkaline extract Item Value ph ~10 Carbohydrates ~50% Amino Acids ~5% Nitrogen 0.8 to 1.5% Phosphorus 0.5 to 1.0% Potassium 14 to 18% Calcium 0.3 to 0.6% Sodium 3.0 to 5.0% Micronutrients 1 to 250 ppm (Fe) Source: Acadian Seaplants Limited, Dartmouth, Nova Scotia, 2012

Reported Cytokinins and Auxin Contents in Seaweed Extracts (SWE) Author, year Cytokinins level, DW Method Tay, 1985 7 µg/l GC/MS Sanderson, 1986 1.3 µg/l GC/MS Tay, 1987 13 µg/l GC/MS Stirk, 2004 50 µg/l Plant bioassay Ervin, 2004, 2008 70 & 45 µg/l ELISA Ervin, 2009 12 µg/l LC/MS/MS Wally, 2013 10 µg/l LC/MS/MS Auxin (IAA) level Ervin, 2009 12 µg/l LC/MS/MS Wally, 2013 3 to 47 µg/l LC/MS/MS Cytokinins level found in bentgrass leaf tissue 5 to 60 ng/g FW* *Zhang and Ervin. 2004, 2008, 2010. Crop Science

Other compounds in SWE with biological activity Complex polysaccharides not present in plants Laminaran, fucoidan, alginate: shown to induce plant defense responses Betaines: natural plant dehydration avoiders Sterols: may play a role in freezing tolerance As reviewed in Calvo et al., 2014. Plant and Soil 383:3-41.

Scientific Consensus on SWE The chemical compositions of several seaweed extracts are known, and because they can maintain plant-promoting bioactivity at relatively low concentrations (<0.01% w/v) (Crouch van Staden, 1993), it is unlikely that the growth-promoting ability is due to nutrient composition alone (Wally, 2013). Similar statements in Blunden 1972, 1991 and Khan et al., 2009. Blunden, G. 1972. Proc Int Seaweed Symp 7:584-589. Blunden, G. 1991. Seaweed Resources in Europe: uses and potential, pp. 65-81. Crouch, I.J. and J. van Staden. 1993. Plant Growth Regulation. 13:21-29. Khan W., et al., 2009. J Plant Growth Regulation 28:386-399. Wally, O.S.D., et al., 2013. J Plant Growth Regulation 32:324-339.

Ervin SWE Research Products used primarily on putting greens Equalized nutrient inputs, Hoaglands Few commercial products used, just generic alkaline seaweed extracts at 0.15% w/v Challenged bentgrass with drought or heat stress

Drought in Greenhouse, 2004 0.15% w/v SWE solution (estimated 3.5 µg/l cytokinins) or ashed SWE applied 7 d prior to 28-d dry down; complete fertilizer solution supplied evenly Leaf Cytokinin Relative Levels at Soil Moisture Wilting Point (5%): Exp. 1 then Exp. 2 1. Control = 100% 100% 2. Ashed SWE = 101% ns 3. SWE = 123% sig 158% sig Zhang and Ervin, 2004. Crop Science, 44:1737-1745

Drought in Greenhouse, 2004 We also tested Humic Acid (HA at 0.43% w/v) from leonardite, alone and in combination with SWE Treatment PE* Vit. E Shoot Wt Root Wt CK HA 0.33b 17.3b 1.28ab 0.80a 19.3b SWE 0.30b 14.3b 1.46a 0.69b 28.9a HA + SWE 0.40a 23.7a 1.49a 0.83a 26.6a Control 0.23c 10.7c 0.96b 0.59b 18.3b *PE = Photochemical Efficiency (0.7 = healthy); Vitamin E; CK = Zeatin riboside (cytokinin) level

Roots left after 28-d dry down

Ascophyllum nodosum extracts tested, 2008 Acadian Seaplants (Dartmouth, Nova Scotia) ASL liquid concentrate: KOH extracted, 14.4% solids Applied to foliage at 3.5 mg ZR/L = 28 ml SWE/L = 10 µmol ZR = 1.3 lb solids/acre = 3.3 oz/m Ocean Organics (Waldoboro, Maine) Liquid concentrate: KOH extracted, 8% solids Applied to foliage at 3.5 mg ZR/L = 28.2 ml SWE/L = 10 µmol ZR = 0.75 lb solids/acre = 3.3 oz/m Zhang and Ervin, 2008. Crop Sci. 48(1):364-370.

Procedures L-93 creeping bentgrass grown from seed to maturity in conetainers in a 75/68 F greenhouse. 1 st SWE treatment foliar-applied 7 d prior to moving to 95/77 F growth chamber Conetainers suspended in ¼-strength Hoagland s solution (aerated and changed weekly) 2 nd SWE treatment applied 14 d after heat stress began Whole conetainer subsamples were taken destructively every 7 days (for 49 days) to quantify responses over time

Leaf zeatin riboside content due to treatment during heat stress Control Aca SWE Oce SWE ZR SWE ash 60 50 40 Zeatin Riboside 30 (ng/g FW) 20 NS b 2 nd app a a a ab b a a b b b a a b b 10 0 0 14 28 42 Days of Heat Stress Bars with the same letter are not diff at P=0.05

Cytokinin-containing compound effects on bentgrass lipid peroxidation (MDA content) under heat stress Control Aca SWE Oce SWE ZR SWE ash MDA content (nmol/g FW) 200 180 160 140 120 100 80 60 40 20 0 a a b c c b d c NS a bc ab d c 0 21 35 49 b a Days of Heat Stress Bars with the same letter are not diff at P=0.05

Root FW (g) Cytokinin-containing compound effects on bentgrass Root Fresh Weight under heat stress Control Aca SWE Oce SWE ZR SWE ash 1.2 1 NS a 0.8 0.6 NS b b b b b a ab ab b 0.4 0.2 0 0 21 35 49 Days of Heat Stress Bars with the same letter are not diff at P=0.05

Check Aca SWE OO SWE CK-check ashed-swe End of 2006 trial, 49 days of heat stress

Bentgrass Heat Stress Study: 2007 We investigated 5 rates of SWE against 5 equivalent rates of pure cytokinin (zeatin riboside, ZR) against 1 True Foliar program at 95-100 F day/ 77-82 F night and 70-80% RH Treatments (4 apps @ 2 wk frequency) 1: Fert Control = 0.05 lb N/M solution of 20-20-20 2-6: SWE at 0.1 1, 10, 100, 200 um ZR (or 0.028, 0.28, 2.8, 28, and 56% solutions) 7-11: Pure ZR at 0.1, 1, 10, 100, 1000 um ZR Treatments 2-11 also mixed with 0.05 lb N/M 20-20-20 12: CPR (3 oz) + True Foliar K (1 oz), T-F NK (4 oz) + T-F Ca (1 oz) = same SWE and fert inputs as 2-4

RESULTS (Zhang and Ervin. 2010. Crop Sci. 50:316-320) The 2.8% (10 um=3.5 ppm CK) SWE and ZR solutions worked better than the weaker ones but the same as the 28% solutions The 200 and 1000 um solutions caused phytotoxicity The True Foliar program mix (= 2.1% SWE solution) was in top group

Fert Control 10 um ZR 10 um ZR from SWE True Foliar mix 8 weeks at 95 F day/77 F night, treated every 2 wk with 0.05 lb N + SWE or CK

Photochemical Efficiency 0.8 0.75 0.7 0.65 0.6 0.55 0.5 0.45 0.4 Photochemical Efficiency 2 4 6 8 Run 2: Weeks at 100/82 F Check SWE 10 ZR 10 True Foliar

Grams/pot Run 1: Check SWE 10 ZR 10 True Foliar 1.4 1.2 1 0.8 NS Check SWE 10 0.6 ZR 10 0.4 True Foliar 0.2 0 Root Mass Run 2: 8 weeks at 100/82 F

Seaweed Extract Summary Consistent results showing that low-rate foliar applications provide increased cytokinin leaf tissue levels that correlate closely with improved physiological adjustments to abiotic stress Effects not associated with mineral component of SWE Cytokinins most likely not solely responsible for improved plant function

Humic Substances From the decomposition (or humification) of carbon containing compounds (plant or animal) Nonhumic substances Are less stable in soil but may be bio-active: Polysaccharides, proteins/enzymes, organic acids (IAA) Source: Brady & Weil, Soil Science

Humic Substances defined Humic substances comprise 60-80% of soil OM and are the components of OM most resistant to microbial decomposition Classified into: Fulvic acid: smallest & soluble in acid & alkali Humic acid: medium & soluble in alkali Humin: largest, most persistent, dark, insoluble Sources Leonardite (mined soft coal, lignite): 30-80% HA, 150 CEC Peats: 5-25% HA, Biosolids/Compost: 3-15% HA

Liquid-applied Humic Substances At 0.05 to 0.10% HA concentrations have been shown to mimic auxin in promoting root growth. * Are non-humic components (saccharides, organic acids) the biologically active fraction? HA and FA also chelate Fe, Mg, Zn, Mn in exchangeable forms for more efficient root availability No soil moisture or structural effects predicted due to very low mass applied VT Research rate is 15 g leonardite/m = ~1.5 lb/a = 0.4% concentration applied in our trials *O Donnell, 1973. Soil Sci. 116(2):106-112

Humic Acid has been shown to increase photosynthesis and root growth In a greenhouse experiment, Crenshaw bent was grown in solution culture supplied with non-limiting nutrients and 3 concentrations of HA. Photosynthesis and root growth was increased significantly from 1 to 4 weeks after treatment by HA at 400 ppm (0.0004%) VT foliar spray rate ~4000 ppm Liu, Cooper, Bowman. 1998. HortScience 33:1023-1025

Auxin coleoptile elongation bioassay Leonardite-HA assayed to have 198 ppb auxin activity or about 10 times the free auxin content found in plant tissues; in comparison HA extracted from compost only had 73 ppb auxin activity (Zhang and Ervin, unpublished data, 2004)

Auxin or Indole Acetic Acid Produced by plants and soil bacteria Promote cell division & stem elongation Induction of rooting In combination with cytokinins they regulate ratio of tillering to root initiation and growth IAA overproducing mutants of Arabidopsis are known as rooty and superroot Rogg & Bartel. 2001. Developmental Cell. 1(5):595

Drought in Greenhouse, 2004 We also tested Humic Acid (HA at 0.43% w/v) from leonardite, alone and in combination with SWE Treatment PE* Vit. E Shoot Wt Root Wt CK HA 0.33b 17.3b 1.28ab 0.80a 19.3b SWE 0.30b 14.3b 1.46a 0.69b 28.9a HA + SWE 0.40a 23.7a 1.49a 0.83a 26.6a Control 0.23c 10.7c 0.96b 0.59b 18.3b *PE = Photochemical Efficiency (0.7 = healthy); Vitamin E; CK = Zeatin riboside (cytokinin) level

KBG sod establishment on sand Two humic sources compared against fertilizer only; fertilizer inputs equalized Peat based versus leonardite, both applied at 3 oz/m every 2 wks for 6 applications Table 4. Humic acid (HA) effects on monthly post-transplant root strength measurements of Kentucky bluegrass Treatment Root Strength 22 May 2002 21 June 2002 22 July 2002 ------------------------------ kg m -2 ------------------------------ Control 367.4a 483.7a 470.1a Leonardite HA 401.1a 532.6a 587.6b Peat HA 430.7a 600.5b 593.0b Root mass Treatment --------mg cm -3 -------- Control 0.60a Leonardite 0.85b HA Peat HA 1.06b 34 and 73% transplant rooting increase

KBG sod establishment on sand Ervin et al., 2005. Acta Hort

SWE + HA Summary Humic acid consistently improves grass rooting Correlated with auxin activity; we have not directly studied question of improved macro or micro-nutrient efficiency Seaweed Extract consistently improves shoot responses to stress Correlated with cytokinin content, not mineral content Other bio-active compounds may play role in stress tolerance Both together give best grass stress response

We have tested amino acids, some bacterial products but in a much less rigorous way QUESTIONS?