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John Freeman

Publications and source records attributed to John Freeman.

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A combined compost, dolomite, and endophyte addition is more effective than single amendments for improving phytorestoration of metal contaminated mine tailings

Background and aims Re-vegetation of mining-impacted landscapes reduces transport of toxic elements while improving soil fertility. This study evaluated whether the planting of a native perennial grass with a consortium of diazotrophic microbial endophytes and municipal waste compost—alone and in combination—enhanced plant growth while stabilizing metal(loids) in dolomite-amended tailings from a historically mined polymetallic mineral deposit. Methods We grew Bouteloua curtipendula seedlings in tailings with hazardous concentrations of As, Cd, Pb, Mn, and Zn. We evaluated how plant growth, organic matter accumulation, and major, minor, and trace element mobilization and phytostabilization responded to microbial endophyte and/or compost amendments after the 45-day growth experiment. Results Although most of the added endophytes were not uniquely identified, the best plant growth and fertility outcomes were achieved with a combination of amendments: dolomite to reduce acidity, compost to increase nitrogen, and a mixed consortium endophyte seed coating to synergistically increase organic carbon and grass biomass yields. Compost reduced shoot and root concentrations—but not yields—of contaminant metals. Endophytes increased foliar Cd, Co, Mn, and Pb yields but mobilized Pb and Zn from the tailings. Root stabilization of Cd, Co, Mn did not require amendments. Conclusion The most effective means of revegetating these acidic, polymetallic tailings with the native B. curtipendula is with a simultaneous dolomite, compost, and endophyte seed treatment. Due to potential phosphate solubilization and siderophore production by this consortium of endophytes, strategies to capture solubilized metal(loids) may be needed for sulfidic tailings with metal(loids) associated with mobile mineral phases.

Plant and Soil

Compost, plants and endophytes versus metal contamination: Choice of a restoration strategy steers the microbiome in polymetallic mine waste

Finding solutions for the remediation and restoration of abandoned mining areas is of great environmental importance as they pose a risk to ecosystem health. In this study, our aim was to determine how remediation strategies with (i) compost amendment, (ii) planting a metal-tolerant grass Bouteloua curtipendula , and (iii) its inoculation with beneficial endophytes influenced the microbiome of metal-contaminated tailings originating from the abandoned Blue Nose Mine, SE Arizona, near Patagonia (USA). We conducted an indoor microcosm experiment followed by a metataxonomic analysis of the mine tailings, compost, and root samples. Our results showed that each remediation strategy promoted a distinct pattern of microbial community structure in the mine tailings, which correlated with changes in their chemical properties. The combination of compost amendment and endophyte inoculation led to the highest prokaryotic diversity and total nitrogen and organic carbon, but also induced shifts in microbial community structure that significantly correlated with an enhanced potential for mobilization of Cu and Sb. Our findings show that soil health metrics (total nitrogen, organic carbon and pH) improved, and microbial community changed, due to organic matter input and endophyte inoculation, which enhanced metal leaching from the mine waste and potentially increased environmental risks posed by Cu and Sb. We further emphasize that because the initial choice of remediation strategy can significantly impact trace element mobility via modulation of both soil chemistry and microbial communities, site specific, bench-scale preliminary tests, as reported here, can help determine the potential risk of a chosen strategy.

Environmental Microbiome

Microbial endophytes and compost improve plant growth in two contrasting types of hard rock mining waste

The re-vegetation of mining wastes with native plants is a comparatively low-cost solution for mine reclamation. However, re-vegetation fails when extreme pH values, low organic matter, or high concentrations of phytotoxic elements inhibit plant establishment and growth. Our aim was to determine whether the combined addition of municipal waste compost and diazotrophic endophytes (i.e., microorganisms that fix atmospheric N 2 and live within plants) could improve plant growth, organic matter accumulation, and phytostabilization of trace element contaminants in two types of hard rock mine waste. We grew a widespread native perennial grass, Bouteloua curtipendula , for one month in alkaline waste rock (porphyry copper mine) and tailings (Ag-Pb-Au mine, amended with dolomite) sourced from southeastern Arizona, United States. B. curtipendula tolerated elevated concentrations of multiple phytotoxic trace elements in the tailings (Mn, Pb, Zn), stabilizing them in roots without foliar translocation. Adding compost and endophyte seed coats improved plant growth, microbial biomass, and organic matter accumulation despite stark differences in the geochemical and physical characteristics of the mining wastes. The widespread grass B. curtipendula is a potential candidate for re-vegetating mine wastes when seeded with soil additives to increase pH and with microbial and organic amendments to increase plant growth.

International Journal of Phytoremediation

Acetylenotrophic and diazotrophic Bradyrhizobium sp. strain I71 from TCE-contaminated soils

Abstract Acetylene (C 2 H 2 ) is a molecule rarely found in nature, with very few known natural sources, but acetylenotrophic microorganisms can use acetylene as their primary carbon and energy source. As of 2018 there were 15 known strains of aerobic and anaerobic acetylenotrophs; however, we hypothesize there may yet be unrecognized diversity of acetylenotrophs in nature. This study expands the known diversity of acetylenotrophs by isolating the aerobic acetylenotroph, Bradyrhizobium sp. strain I71, from trichloroethylene (TCE)-contaminated soils. Strain I71 is a member of the class Alphaproteobacteria and exhibits acetylenotrophic and diazotrophic activities, the only two enzymatic reactions known to transform acetylene. This unique capability in the isolated strain may increase the genus’ economic impact beyond agriculture as acetylenotrophy is closely linked to bioremediation of chlorinated contaminants. Computational analyses indicate that the Bradyrhizobium sp. strain I71 genome contains 522 unique genes compared to close relatives. Moreover, applying a novel hidden Markov model of known acetylene hydratase (AH) enzymes identified a putative AH enzyme. Protein annotation with I-TASSER software predicted the AH from the microbe Syntrophotalea acetylenica as the closest structural and functional analog. Furthermore, the putative AH was flanked by horizontal gene transfer (HGT) elements, like that of AH in anaerobic acetylenotrophs, suggesting an unknown source of acetylene or acetylenic substrate in the environment that is selecting for the presence of AH. Importance The isolation of Bradyrhizobium strain I71 expands the distribution of acetylene-consuming microbes to include a group of economically important microorganisms. Members of Bradyrhizobium are well studied for their abilities to improve plant health and increase crop yields by providing bioavailable nitrogen. Additionally, acetylene-consuming microbes have been shown to work in tandem with other microbes to degrade soil contaminants. Based on genome, cultivation, and protein prediction analysis, the ability to consume acetylene is likely not widespread within the genus Bradyrhizobium . These findings suggest that the suite of phenotypic capabilities of strain I71 may be unique and make it a good candidate for further study in several research avenues.

California

Phytoremediation of slightly brackish, polycyclic aromatic hydrocarbon‐contaminated groundwater from 250 ft below land surface: A pilot‐scale study using salt‐tolerant, endophyte‐enhanced hybrid poplar trees at a Superfund site in the Central Valley of California, April‒November 2019

Slightly brackish groundwater contaminated by polycyclic aromatic hydrocarbons (PAHs) at a Superfund site in the Central Valley of California was pumped from 250 feet below land surface to a water storage tank using solar power and then gravity‐fed into 18, 330‐gallon intermediate bulk containers (totes) as follows: (1) Five totes contained planting medium with three salt‐tolerant hybrid poplar trees per tote ( n = 15); (2) Seven totes contained planting medium with three salt‐tolerant hybrid poplar trees per tote and inoculated with the naturally occurring, PAH‐degrading endophyte Pseudomonas putida PD1 ( n = 21); (3) Three totes contained planting medium only ( n = 0); (4) One tote contained groundwater with three PD1‐inoculated trees ( n = 3) and one tote contained groundwater with three regular trees ( n = 3); and (5) One tote contained groundwater only ( n = 0). All trees grew well during the 7‐month growing season in spite of the area's hot, dry air temperature, little precipitation, tote‐influent chloride concentrations of 290 mg/L, and tote‐influent naphthalene concentrations that ranged from 650 to 5100 mg/L. PD1‐inoculated trees initially had 56% larger tree area (tree height × tree width) than regular trees and up to 69% larger tree area by the end of the growing season, indicating some conferred phytoprotection to the PAH contamination. All trees had similar trunk caliper (diameter) and leaf chlorophyll content by the end of the growing season. Total naphthalene removal ranged from 88% to 100% across all totes. The lowest naphthalene removal of 88% was observed in a tote that contained only planting medium and indicates substantial adsorption of naphthalene onto the high organic content of the planting medium. Contaminant removal due to uptake by the hybrid poplar trees was confirmed by the detection of naphthalene in in vivo passive samplers placed in tree trunks. Benzene, toluene, ethylbenzene, total xylenes, 2‐methylnaphthalene, 1,2,4‐trimethylbenzene, and isopropylbenzene were also detected. These results from the pilot‐scale study indicate that a full‐scale application of using salt‐tolerant hybrid poplar trees at this site could effectively decrease naphthalene concentrations in groundwater pumped from the deep aquifer. These initial results provide hope for similar application at other contaminated sites characterized by groundwater at considerable depths, especially at Superfund sites where costly pump‐and‐treat systems have been used long term to treat low levels of groundwater contamination.

California