Boosting plant defenses using silicon fertilization

Boosting plant defenses using silicon fertilization

Summary

The purpose of this project is to study the effect of silicon supplementation on the enhancement of plant defenses against insect herbivores. Plants have developed a variety of defense mechanisms to protect themselves against herbivores. Physical defenses include trichomes, thorns, spines, tough leaves, lacticifers, and silica deposition. Chemical defenses include secondary metabolites, proteins and enzymes that have a negative effect on plant feeding insects. Silicon (Si) fertilization has been known to be beneficial for plants and enhance defenses against environmental and biotic stresses such as herbivores. This project integrates field and laboratory experiments to test the protective effect of Si against herbivorous insects on different crops. Our initial experiments indicate that, upon damage, Si supplementation enhances the activity of protein defensive enzymes, total phenolics and the production of glandular trichomes in tomato plants. Our bioassays indicate that these plant defenses reduced the weight gain of the tomato fruit worm (Helicoverpa zea) caterpillars. Although not conclusive, our initial experiments are promising and suggest a beneficial effect of Si supplementation in the enhancement of plant immunity.

Objectives/Performance Targets

1) To determine the effect of Si fertilization on yield and plant resistance to herbivores under greenhouse and field conditions.

1a). To test the effect of Si supplementation in the enhancement of plant defense responses upon herbivory in greenhouse conditions.

Tomato plants supplemented with Si (2 mM and 5 mM potassium silicate) and not Si-supplemented controls (2 mM and 5 mM potassium chloride) were grown in greenhouse conditions and exposed to Helicoverpa zea caterpillars three weeks after transplanted. 10 days later, tissue from the newer leaves was harvested to test for the activity of defensive proteins (polyphenol oxidase (PPO), peroxidase and trypsin protease inhibitor activity) and total phenolics. For all plant treatments, the activity of PPO, peroxidase, trypsin inhibitor, and the total phenolics content was significantly higher on plants exposed to herbivory compared with undamaged controls (Fig.1-3). When the activity of these proteins was compared only among plants treated with caterpillars, there were not significant differences among treatments. A significantly higher content of total phenolics was found on plants supplemented with 5 mM Si compared with plants supplemented with 5 mM KCl (t = -2.72, p-value = 0.026; N= 5) (Fig. 4).

1b). To test the effect of Si supplementation on plant resistance to herbivores and yield in field conditions.

Tomato, corn and soybean plants were planted at the Russell E. Larson Agricultural Research Center (Rock springs, PA), and fertilized at the moment of transplanting with either two different doses of calcium silicate (3gr and 1gr) or their respective controls (3gr and 1gr of calcium carbonate). The presence of insect damage was monitored every week during 3 months. Insects did not affect tomato or corn plants during the whole season, only a few aphids were observed in tomato by the end of August. The presence of aphids was not affected by the treatments. Three different insect species were found in soybean plants: first, the silver spotted skipper caterpillar (Epargyreus clarus) was found by the end of June; second, the soybean looper (Pseudoplusia includens) was found by mid July; lastly, Japanese beetles (Popillia japonica) were the most predominant insect found during July and August. The distribution and damage of these insects was not affected by the plant treatments.

The effect of Si treatments on yield was measured by harvesting and weighing mature tomato fruits, corn ears and soybean pots. No significant differences were found between treatments for the total weigh (gr) of tomatoes (ANOVA, F_3,76 = 0.25 p value = 0.861; N=20 plants per treatment) or soybeans (ANOVA, F_3,12 = 0.66 p value = 0.595; N=4 plots per treatment of 6 plants each) per plant (Fig. 5 & 6). No data was obtained for corn due to extensive raccoon damage.

2) To determine the effect of silicon fertilization and insect damage on the density of trichomes and mineral composition of plant leaves.

Tomato plants supplemented with Si (2 mM and 5 mM potassium silicate) and not Si-supplemented controls (2 mM and 5 mM potassium chloride) were grown in greenhouse conditions and exposed to Helicoverpa zea caterpillars three weeks after transplanted. 10 days later the newer fully expanded leaf was harvested to count the number of glandular trichomes in a 0.24 mm² area and the remaining leaves dried for mineral analysis. For all treatments, the number of trichomes was higher on plants exposed to herbivory than in the corresponding undamaged controls. Among the plants treated with caterpillars, the number of trichomes was significantly higher on plants supplemented with 5 mM Si compared with plants supplemented with 5 mM KCl (t = -5.66, p-value < 0.0001; N=6), but there were not significant differences between plants treated with 2 mM Si, 2 mM KCl and 5 mM Si (Fig. 7). Tomato plants supplemented with Si accumulated significantly more Si on the leaves than the not supplemented controls (ANOVA, F_2,18 = 40.87 p value < 0.0001: N=7 plants per treatment) (Fig. 8), which is surprising since tomato is not a Si accumulator plant.

 3) To determine the effect of Si-fertilized plants on caterpillar growth, mandible wear and integrity of the insect’s gut protective membrane.

 3a). To test the effect of Si-fertilized plants on caterpillar weight gain.

Tomato plants supplemented with Si (2 mM and 5 mM potassium silicate) and not Si-supplemented controls (2 mM and 5 mM potassium chloride) were grown in greenhouse conditions and exposed to Helicoverpa zea caterpillars three weeks after transplanted. 10 days later the new leaves were detached and used to feed 3-day old H. zea caterpillars. These caterpillars were weighed 6 days later and the weight gain calculated. For all treatments, the caterpillar weight gain was significantly lower when fed on plants previously exposed to herbivory than when fed on the corresponding undamaged controls. Interestingly, caterpillars gained more weight when fed on control plants treated with Si than when fed on control plants treated with KCl (ANOVA, F_3,23 = 8.24 p value < 0.0001: N= 6-8). However, when young caterpillars where fed on plants previously exposed to herbivory, they gained less weight when fed on plants supplemented with 5 mM Si compared with their corresponding 5 mM KCl treated controls (t=2.26, p-value = 0.041; N= 7,8). There were not significant differences in weight gain by caterpillars fed on 5 mM Si compared with 2 mM Si and 2 mM KCl (Fig. 9).

3b). To test the effect of Si-fertilized plants on caterpillar mandible wear and integrity of their gut membrane.

The proposed activities to test this objective will be carried out in spring 2016.

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Accomplishments/Milestones

May –Sep. 2015: first field season: planting, weekly assessment of insect damage, harvest and yield measurements (objective 1b). The second field season will take place in summer 2016.

Oct –Dec. 2015: Greenhouse experiments with tomato plants, defensive protein assays, trichome counting and bioassays (objectives 1a and 2). Two complete replications in tomato have been carried out, the results presented on this report correspond to the first replicate, the samples of the second replicate are stored at -80 for analyses. The corn and soybean experiments will be carried out in spring 2016.

Dec. 2015. Total phenolics analysis, tissue grinding, Si measurements (objective 2), statistical analyses, and preparation of the first report.

In general, most of the activities originally programmed for this semester were accomplished. It wasn’t possible to do the greenhouse experiments with corn and soybean due to greenhouse reparations.

Impacts and Contributions/Outcomes

Si is known to accumulate in plant tissues where it plays an important role in maintaining plant structure and rigidity. It also enhances physical protection against pathogens and insect herbivores. This project is exploring the use of Si as an enhancer of plant physical and biochemical defenses in response to herbivory. So far, our results show that Si supplemented tomato plants accumulate 2.5 fold more Si than not supplemented controls. Upon herbivory, Si supplemented plants seems to produce higher number of glandular trichomes and total phenolics than their corresponding controls. Furthermore, young caterpillars gained less weight when fed on plants supplemented with Si that were previously exposed to herbivory compared with plants supplemented with KCl. So far, our results indicate that Si supplementation may enhance plant immunity in non Si-accumulator plants like tomato. The outcomes of this project have the potential to incorporate an additional practice for IPM programs reducing the use of pesticides.

Collaborators: