Location: Characterization and Interventions for Foodborne Pathogens
2020 Annual Report
Objectives
1: Increase efficiency of AM fungus inoculum production and utilization to enhance vegetable production and reduce inputs.
1A. On-farm production and utilization of AM fungus inoculum to enhance vegetable crop yields and reduce inputs.
o Increase propagule density of inoculum produced when the indigenous community is used as starter inoculum. (Douds)
o Demonstrate use of AM fungus inoculum produced on-farm for eggplant production.
1B. Examine the use of black plastic mulch upon indigenous AM fungus activity and evaluate relative to opportunities to utilize AM fungus inoculum. (Douds)
2: Determine the effects of arbuscular mycorrhizal fungus species on improving crop physiological responses that enhance nutrient uptake, increase production of polyphenols, and enhance plant resistance to insect pests and diseases.
2A. Quantify the impact of AM fungus colonization upon production of polyphenols of known health benefit in different crops.
2B. Enhance the utilization of the AM symbiosis through studies of basic physiology and nutrient uptake.
o Measure the flow of C, N, and P between plants of varying photosynthetic rates via common, interconnected AM fungus networks and correlate to expression of plant P and NH4 transporter genes.
o Determine the impact of newly-identified root exudate signal molecules (abietic acid and dehydroabietic acid) on carbon uptake and lipid synthesis by germinating spores of AM fungi via NMR spectroscopy.
o Determine the impact of semi-purified root exudate signals in combination with low oxygen concentrations upon AM fungus germination and growth.
3: Develop management practices that minimize the potential negative effects of off-site transport of pathogenic bacteria in integrated crop/livestock production systems.
3A. Inactivate pathogenic bacteria, originating from animal manure, moving in surface water as a result of heavy rainfalls, before they can enter farm irrigation ponds by using erosion control socks amended with fast pyrolysis biochar.
Approach
Arbuscular mycorrhizal [AM] fungi are obligate symbiotic soil fungi that form a mutualistic symbiosis with the majority of crop plants. Better utilization of this symbiosis will enable farmers to increase or maintain yields while reducing synthetic chemical inputs. Earlier, we developed a method for the on-farm production of inoculum of AM fungi using bahiagrass as the host plant. This method will be refined for the production of AM fungi indigenous to the farm and its usefulness demonstrated for the production of eggplant. The obligate symbiotic nature of these fungi require that a host plant be present during the production of inoculum for use by farmers. Large scale production of pure inoculum of these fungi (i.e. growth of the fungi by themselves, in the absence of a plant) is possible if the physiological limitations that require colonization of a host plant could be overcome. We will attempt to overcome these constraints by growing the fungi in the presence of a combination of environmental factors (root exudate signals, high CO2, and low oxygen) more representative of the environment in which they naturally grow rather than typical laboratory conditions. Other studies will examine the impact of AM fungus colonization upon mineral nutrient uptake and the production of human health-promoting compounds in plants. Farms with integrated livestock and crop production are faced with regulatory pressures designed to minimize the risk of contamination of produce with bacteria originating in animal manure. We will explore the idea that erosion control socks containing biochar, previously shown to reduce the population of pathogenic bacteria in soil by 100 fold, could be used to intercept rainwater runoff from pastures and inactivate bacteria before they contaminate the water in an irrigation pond.
Progress Report
Related to Objective 1 [Increase efficiency of arbuscular mycorrhizal (AM) fungus inoculum production and utilization to enhance vegetable production and reduce inputs] quantitative PCR (qPCR) assays were developed for the identification and quantification of 11 different species of AM fungi. Previously, it has been shown that mixed-species arbuscular mycorrhizal fungi (AMF) inoculum results in beneficial yield increases of multiple crops, but the measurement of impact of individual species was not possible. Identification and quantification of AM fungi from soil samples previously required isolation of spores from the soil, morphological identifications, and time-consuming plant-based colonization assays. Measurement of the colonization efficiency of individual species of AM fungi from a mixed inoculum was not possible.
To develop the qPCR assays, amplicon sequencing of the internal transcribed spacer (ITS) and glomalin related protein gene was completed from >40 accessions of AM fungi in the international culture collection of (vesicular) arbuscular mycorrhizal (INVAM) repository and several cultures from the Eastern Regional Research Center (ERRC) greenhouse. This sequence data was utilized to identify species-specific targets within the genomes of the 11 different species represented. Specificity testing confirmed the assays do not cross react with DNA from non-target AM fungi. qPCR enables the rapid detection and quantification of AM fungi directly from soil DNA extracts, and enables species-specific quantification of AM fungi colonization of plant roots. Samples can be extracted and analyzed within a single day, and future work will involve multiplexing the assays for increased throughput. The availability of qPCR will enable more precise field and laboratory studies of the relationship between individual species of AM fungi and crops, and guide the selection of AM species for development as biofertilizers to improve nutrient acquisition and water uptake and reduce susceptibility to pathogens and disease.
Related to Objective 1.B (Examine the use of black plastic mulch upon indigenous AM fungus activity and evaluate relative to opportunities to utilize AM fungus inoculum), experiments were delayed due to critical vacancies, but an opportunity to pursue this objective with at an alternate site has arisen. A Pennsylvania Specialty Crop Block Grant was awarded to the Rodale Institute to study the impact of black plastic mulch in Rodale Vegetable Systems Trial. ARS is a collaborator on this 3-year grant. The crops to be tested include sweet corn and butternut squash; the impact of AMF on yield and nutritional content of the produce will be evaluated. On-farm propagation of AMF inoculum has begun during 2020 for use in the 2021 growing season.
Related to Objective 2.B.3 (Enhance the utilization of the AM symbiosis through studies of basic physiology and nutrient uptake, evaluation of the combination of low O2 tensions and root exudate signals on growth and carbon utilization by germinating spores of AM fungi), preparations for these experiments have been made. Past vacancies have caused delays in initiating these experiments, but it is anticipated that much of the original proposed work can be completed before the end of the project cycle. Root exudates have been prepared for use and stored, AM fungi have been propagated for these experiments, and equipment (new CO2 incubator, hypoxia chambers) are on site.
Accomplishments
