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Title:   Author  item
                                            Vaughan, Marthaitem
                                            THOLL, DOROTHEA - Virginia Polytechnic Institution & State Universityitem
                                            TOKUHISA, JAMES - Virginia Polytechnic Institution & State University
  Submitted to: Plant Methods
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 1/17/2011
                                    Publication  Date: 4/1/2011
                                    Citation: Vaughan, M.M., Tholl, D., Tokuhisa, J.G. 2011. An aeroponic culture system for the study of root herbivory on Arabidopsis thaliana. Plant Methods. 7(5):1-10.Interpretive Summary: Roots constitute 50% to 90% of plant biomass, and consequently a large number of diverse organisms exploit belowground plant tissues for their carbon and nutrient supply. However, research investigating plant defense has mainly focused on visible aboveground interactions between plants and herbivores because investigation of belowground defense has been impeded by accessibility of root tissue and the lack of developed techniques.  As a result, little is known about how plants defend their roots from belowground insect herbivores. Scientists at the USDA-ARS Center for Medical, Agricultural and Veterinary Entomology in Gainesville, Florida and the University of Virginia Tech have developed an aeroponic culture system based on clay pebbles that allows the growth of the model system plant Arabidopsis while providing easy access to the root tissue and a more natural subterranean environment for root herbivores. Using this system, the feeding performance and plant tissue damage caused by the dark-winged fungus gnat larvae, a generalist root herbivore was evaluated. The aeroponic culture system enabled direct observation of fungus gnat larvae feeding and behavior on the in tacked root tissue.  Larval feeding on plants resulted in reduced root mass and impaired water absorption.  Herbivore damage caused to mature roots grown under aeroponic conditions was compared to that of seedlings grown in potting soil.  Soil grown seedlings infested with fungus gnat larvae were more susceptible had increased mortality and surviving plants grew much slower. This system has helped overcome the methodological difficulties associated with research investigating belowground root- herbivore interactions and plant defense. This bioassay can easily be performed to assess the effect of herbivory on the formation of chemical defense in roots and provides a basis for future investigations on the biochemistry, molecular regulation and defensive function of root chemicals in interaction with belowground herbivores. Knowledge of the natural defense potential of plants against root herbivores can then be transferred to agricultural applications in an effort to control belowground herbivore damage.
                                Technical Abstract: Abstract
Background:  Plants have evolved complex defense mechanisms to combat herbivore attack. While plant defense against herbivory has been studied primarily in aerial tissues, research investigating defense responses belowground has been hindered by experimental difficulties associated with the accessibility and quality of root tissue and the lack of bioassays using model plants with altered defense profiles. 
Results: We have developed an aeroponic culture system based on a calcined clay substrate that allows insect herbivores to feed on plant roots while providing easy access to the root tissue. The culture method was validated by a root-herbivore system developed for Arabidopsis thaliana and the root herbivore Bradysia spp. (fungus gnat). Arabidopsis root mass obtained from aeroponically grown plants was comparable to that in other culture systems, and roots were morphologically normal. Bradysia larvae caused considerable root damage resulting in reduced root biomass and water absorption. Larvae pupated and emerged as adults under these culture conditions. Root damage of mature plants cultivated in aeroponic substrate was compared to that of Arabidopsis seedlings grown in potting mix. Seedlings were notably more susceptible to Bradysia feeding than mature plants? and showed decreased overall growth and survival rates.
Conclusions:  A root-herbivore system consisting of Arabidopsis thaliana and larvae of the opportunistic herbivore Bradysia has been established that more closely mimics herbivory in the rhizosphere. Bradysia infestation of Arabidopsis grown in this culture system significantly affects plant performance. The culture method will allow simple profiling and in vivo functional analysis of root defenses such as chemical defense metabolites that are released in response to belowground insect attack.


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