Project : USDA ARS

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Research Project: Chemical Communications of Plants, Insects, Microbes, and Nematodes

Location: Chemistry Research

2022 Annual Report

Objectives
Objective 1: Investigate semiochemicals of agricultural plant-insect-microbe interactions, with emphasis on mites, weevils, leaffooted bugs, and fruit flies; and, volatile biomarkers from insect or microbe infestations of agricultural products. Objective 2: Collect, analyze, and identify semiochemicals of pollinator pest-host and nematode-plant-microbe interactions, including varroa mites, hive beetles, entomopathogenic nematodes, or associated soil/root microbes. Objective 3: Survey floral nectar microorganisms occurring in flowering crops and determine their impacts on nectar chemistry, pathogen loads, and pollinator affinity.

Approach
Investigate the chemical communications of agricultural plant-insect-microbe systems, such as those involving mites, weevils, leaffooted bugs, and fruit flies, as well as entomopathogenic nematodes and their associated soil/root microbes. This work will also include investigating how pheromones and multitrophic signaling can be utilized for effective control of pest organisms. Provide new or improved biological, behavioral, or cultural control methods to reduce agricultural reliance on broad spectrum chemistries. To address the ever-changing abiotic stressors, will provide flexible pest management tools for agricultural ecosystems, including those that protect plant-pollinator habitats or hosts. Will address these pollinator issues by identifying semiochemicals of pollinator pest-hosts, including varroa mites and hive beetles, and providing cost-effective, environmentally safe pest management strategies. This research will utilize numerous interactive laboratory- and field-based bioassays with insects, mites, microbes, nematodes, and plants, as well as purified biochemicals and other organisms. Isolation and identification of new bioactive chemicals that mediate arthropod and nematode behaviors and other inter-organismal interactions will be achieved using a combination of approaches including HPLC, LC-MS, preparative flash chromatography, GC-MS, FT-IR, NMR, micro-degradation, and synthesis where applicable. Major target insects for this research will include those listed above, with other target insects selected as needed during progression of the project.

Progress Report
Progress has been made on Research Project 6036-22430-001-000D by Gainesville, Florida Scientists. In Objective 1, progress included the following: Nematodes – by utilizing a soil-filled dual choice olfactometer and very sensitive CO2 measurements, we established that although an attractant, CO2 is not a major factor used by Steinernema diaprepesi entomopathogenic nematodes (EPNs) host selection and recognition; synthetic kairomone assays confirmed that EPNs locate hosts by the help of host and root-produced kairomones; we discovered that some plant-produced (terpenoid) kairomones, in addition to being possible nematode attractants, also increases insects ability to defend themselves against the nematodes (manuscript submitted to journal); with collaborators, we established the “follow the leader” aggregative host searching and host insect mass infection behaviors; isolation of kairomones is in progress; and probes for collections of herbivory-induced root volatile were modified for sand- and organic material-rich soil. Sampling of corn and blueberry plant root volatiles is ongoing. This work also includes a study of how some defensive responses in roots are expressed and regulated, e.g., using lipoxygenase mutant corn plants; Maize weevil – results of behavioral assays of candidate attractants indicated that both male and female weevils were attracted to the compound hexanal, a compound associated with the smell of fresh cut grass. Ongoing research will investigate the potential use of hexanal as an inexpensive method for monitoring and control of this insect pest; Fruit Flies – volatile collection media cartridges were sent to collaborators in Kenya for the investigation of fruit fly (Bactrocera dorsalis) infestation of mangoes. Eleven odors were identified as potential biomarkers. Manuscript is in preparation and Animal and Plant Health Inspection Service (APHIS) collaborators have been notified; in collaboration with a University of Florida researcher, odors from Drosophila suzukii-foraged strawberries were collected and analyzed. Manuscript is in preparation; leaffooted bug (LFB) – odors from a plant that is highly attractive to LFB have been analyzed via GC-MS and candidate odors are undergoing electrophysiological assays. A colony of LFB has been successfully established. Under Objective 2, progress included the following: Nematodes – we established that host selective EPN rearing results show a clear “innate” preference for that host using sand-filled dual choice olfactometers. We have also established that for training on synthetic versions of root volatiles that typically are released in response to root herbivory results in a clear preference for the trained root volatile; with Steinernema diaprepesi, we found no clear and simple hierarchy when combining selective rearing versus trained preference (manuscript in preparation); when trained on the known EPN attractant pregeijerene, infective juvenile (IJ) EPNs showed increased infection rates when paired with a non-preferred host but no synergistic effect when paired with a preferred host. Furthermore, IJs on ß-caryophyllene or a-pinene showed increased infection rates when paired with a non-preferred host but decreased infection rates when paired with a preferred host. These results demonstrate that care will have to be taken when preparing EPNs for field release; Varroa mites – in collaboration with researchers at the University of Florida, we developed novel methods to rear and maintain populations of Varro destructor in the laboratory. Honey bee populations continue to suffer significant gross losses yearly due to the Varroa mite. Research on Varroa has been limited by an inability to study it independent of honey bee colonies. Our advancements in laboratory reared Varroa will further the understanding of the host-parasite relationship and develop better control strategies. A manuscript is in preparation. Under Objective 3, progress included the completion of three research priorities that facilitate microbial antagonism assays of nectar microbes. First, key agricultural pathogens that infect flowers and fruit, including 18 strains representing four distinct microbial species, were acquired. Second, a quantitative PCR method was developed to selectively measure the target pathogen cell density. Third, the sugar and amino acid concentrations of blueberry floral nectar were measured to inform synthetic nectar media preparation. Other tangential projects included the discovery of nine pathogen-produced odor repellents of Drosophila suzukii emanating from pathogen-infected blueberry fruit. An invention disclosure has been submitted and a manuscript describing the microbiome of wild and cultivated blueberry nectar is in preparation. ARS scientists at Gainesville, Florida, experienced possible complications of switching carrier gas for gas chromatography/mass spectrometry instrumentation from helium to nitrogen or hydrogen. We distributed reports/procedures to several ARS laboratories. Following the established procedures, research can be continued even with limited helium supply and rising price.

Accomplishments
1. Discovered several potential biomarkers for infestation of fruit fly eggs and larvae in mangoes. Fruit flies are a critical insect pest to many agricultural products and incur serious commodity and economic damage. Part of the economic damage to the agricultural industry is international shipping restrictions due to fruit fly infestation of the products. ARS researchers in Gainesville, Florida, in collaboration with USDA-APHIS and researchers in Kenya have identified biomarker odors, chemical compounds unique to insect infestation, of important agricultural commerce products and the infestation of their associated insect pests. In this study, the identification of biomarkers denoting fruit fly infestation of mangoes provide USDA-APHIS with important information for the detection and removal of infested fruits at commerce points, thus helping to control the spread of invasive insect species as well as helping to protect the safety of food products.

2. Discovered nine potential repellents for the invasive pest, spotted wing Drosophila. In the U.S., the invasive fruit fly spotted wing Drosophila is a serious pest of fruits, causing over $300 million in damages annually. Because their eggs and larvae are enclosed within infested fruit, they are shielded from insecticide sprays making them difficult to control using conventional methods. As an alternative or supplement, odors can be used to manipulate the pest’s behavior in the field. ARS researchers in Gainesville, Florida, in collaboration with Rutgers University discovered that flies avoided fruit infected with a specific pathogen. By studying the odors emitted from pathogen-infected fruit and examining pest responses to these odors in the laboratory, they identified nine previously unreported chemical repellents that can potentially be used to prevent flies from damaging crops. This developing technology can improve crop protection and help reduce insecticide reliance.

3. Development of a novel blend of beer volatiles for attracting small hive beetle. There are few trapping methods used in the management of the small hive beetle, a predatory pest of the western honey bee, Apis mellifera. Comparative studies demonstrated that the beetles have a strong attraction for beer when combined with existing oil traps in both laboratory and field trapping assays. ARS researchers in Gainesville, Florida, investigated the potential attractive substances in beer to determine why it was preferred by the beetles. The odors produced by beer were collected, analyzed, and evaluated as a potential attractant. Researchers identified four compounds from beer that were tested to see if the beetles were attracted to these compounds. The beetles responded to all four compounds in multiple experiments, and the blend captured significantly more beetles than all the other treatments. These results advance efforts to produce new attractants for controlling the small hive beetle and can be easily incorporated into existing beetle control practices.

4. Use of companion crops to reduce infections of tomato by Meloidogyne incognita. Plant parasitic nematodes infections of cash crops cause an excess of $125 billion annual crop damage worldwide, and plant parasitic nematodes are widely regarded as the most damaging biotic crop stressor to which there is no real viable management option. In collaboration with scientists from Nairobi, Kenya, ARS researchers in Gainesville, Florida, reported a novel and sustainable approach to an intractable pathogen, the root knot nematode, Meloidogyne incognita. We used the wild growing, readily available vegetable black-jack (Bidens pilosa) as a companion crop to deliver root exudate masking/defensive chemical compounds during plant growth to interfere with the fundamental biology of the root knot nematode-tomato interaction. Achieving safe and effective nematode control is a key part of the movement toward environmentally sustainable agriculture.

U.S. DEPARTMENT OF AGRICULTURE

Chemistry Research: Gainesville, FL