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Evidence for nonstomatal uptake of Hg by aspen and translocation of Hg from foliage to tree rings in Austrian pine Jennifer Arnold, Mae Sexauer Gustin, and Peter Weisberg Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.7b04468 • Publication Date (Web): 12 Dec 2017 Downloaded from http://pubs.acs.org on December 21, 2017
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Evidence for nonstomatal uptake of Hg by aspen and translocation of Hg from foliage to
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tree rings in Austrian pine
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Jennifer Arnold, Mae Sexauer Gustin*, Peter J. Weisberg
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Department of Natural Resources and Environmental Science
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University of Nevada-Reno
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Reno, Nevada, USA 89557
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Corresponding author:
[email protected] 001-775-784-4203
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Abstract
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To determine whether trees are reliable biomonitors of air mercury (Hg) pollution
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concentrations were measured in bark, foliage, and tree rings. Data were developed using 4-year
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old Pinus and Populus trees grown from common genetic stock in Oregon and subsequently
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transferred to four air treatments differing in gaseous oxidized mercury (GOM) chemistry and
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total gaseous Hg (TGM) concentrations. Soil of a subset of trees was spiked with HgBr2 in
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solution to test for root uptake. Results indicate no significant effect of the soil spike or GOM
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compounds on tree tissue Hg concentrations. TGM treatment had a significant effect on Pinus
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and Populus foliage, and Pinus year 5 growth ring concentrations. Populus foliar Hg
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concentrations were highest in the exposure where 24 h TGM concentrations were highest,
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indicating the importance of the nonstomatal pathway for uptake. Pinus tree ring concentrations
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were correlated to day time TGM concentrations suggesting Hg accumulation into tree rings is
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by way of the stomata and subsequent translocation by way of phloem. Populus leaves and Pinus
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rings can be used as biomonitors for TGM concentrations over space. However, the use of trees
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as temporal proxies requires further investigation due to radial translocation observed in active
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sapwood tree rings.
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Keywords 1 ACS Paragon Plus Environment
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HgBr2; Pinus nigra; Populus tremuloides; gaseous oxidized mercury; total gaseous mercury
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Introduction Vegetation plays a significant role in the biogeochemical cycling of mercury (Hg) as
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studies have shown vegetation to be an important sink for atmospheric Hg. 1-3 Approximately 1/3
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of the Hg removed each year from the atmosphere is by vegetative uptake. Potential pathways
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for Hg to enter trees include uptake through the roots from Hg dissolved in soil and transport to
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the above ground tissues. 4-6 Previous studies have shown Hg concentration in foliage to be
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independent of or weakly correlated to soil Hg concentrations; thus, Hg uptake through tree roots
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is considered minor. 4, 7-10 A second pathway is passive transport through the bark from the
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atmosphere. Bark can absorb airborne pollutants because of its porosity, chemical inertness in
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the presence of inorganic and organic substances, and lack of metabolic processes. 10-11 The third
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pathway is assimilation from air into foliage through stomata and/ or cuticle, and translocation
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down by way of the phloem. 12-14 Previous studies have shown atmospheric Hg concentrations
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significantly influence foliage Hg concentrations of coniferous and deciduous trees by way of Hg
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assimilation through the stomata and/or cuticle. 3,7,8,15,16 In this study, we used controlled dose-response experiments to assess the influence of air
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Hg concentrations and soil Hg spikes on tree tissues (bark, rings, and foliage) of Pinus nigra
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(Austrian pine) and Populus tremuloides (quaking aspen) saplings. Four year (Y) old trees were
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placed in three separate greenhouse bays and one outdoor exposure for one growing season.
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Treatments included different concentrations of total gaseous mercury (TGM) and compounds of
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gaseous oxidized mercury (GOM). In addition, soils of a subset of trees were spiked with HgBr2
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prior to placement in greenhouse locations to determine the impact on Hg concentrations in tree
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tissues.
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The objectives of this study were to determine if Pinus and Populus trees can be used as
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monitors of spatial and temporal changes in air Hg concentrations, and to understand the
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pathways for Hg accumulation into tree rings. Trees were moved from areas of low air Hg
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concentrations to areas of higher air Hg concentrations to test whether foliage, bark, and tree
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rings record spatial changes in air Hg concentrations. Concentrations of Hg measured in the
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foliage and bark would be expected to change after being exposed to higher Hg air
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concentrations after one year of growth, because the source of Hg into foliage and bark is
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primarily of atmospheric origin.12 By quantifying the change in Hg concentrations after one year
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of growth in higher Hg air exposures, we investigated whether there is smearing or radial
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translocation of Hg in previously formed rings. This information is useful for determining
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whether tree rings can be used as reliable temporal monitors of environmental air Hg
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concentrations. Additionally, there is no conclusive evidence regarding the pathway for Hg
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accumulation into tree rings or on the relative importance of stomatal versus cuticular uptake of
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atmospheric Hg by foliage. This study provided an experimental framework for addressing these
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uncertainties.
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Materials and Methods
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Experimental Design
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Quaking aspen (Populus tremuloides) and Austrian pine (Pinus nigra) were purchased in
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March 2016 as 4-year-old saplings. Pinus saplings were grown at a tree farm in Canby, Oregon,
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and arrived in 38 L pots with the stems bundled with twine and roots contained in burlap sacks.
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Soil within the burlap sack was a clay soil, whereas soils outside the bundle contained more
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organic matter and were small in volume compared to soil within the sack. The clay soil within
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the burlap sack (where roots were initially present) had an Hg content of 38 ± 12 (n=5) ng g -1.
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Populus saplings were grown at a tree farm in Mt. Angel, Oregon, and arrived in 19 L pots with
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no foliage on the date of arrival. Soil of Populus pots was an organic mixture with a Hg content
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of 85 ± 4 (n=5) ng g-1.
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Plant treatments were conducted at four different sites located on the University of
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Nevada-Reno campus from April to September 2016. Three of the 4 experimental treatments
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were conducted within two environmentally controlled greenhouse facilities. One greenhouse
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facility was the Environmental Research Facility (ERF) that housed two treatments: an ambient
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air Hg exposure (ERF-A) and an increased Hg air exposure where ambient air was spiked with
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HgBr2 (ERF-S) (See Supplemental Information, SI). The other greenhouse facility used for
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experiments was the University of Nevada College of Agriculture, Biotechnology, and Natural
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Resources Valley Road Greenhouse Facility (VRG) that is a multi-bay greenhouse complex
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located on the southeast end of campus and parallels Interstate-80. One VRG bay was used that 3 ACS Paragon Plus Environment
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housed tubs of soil with Hg contaminated mine waste including heap-leached material, material
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removed as overburden, and tailings from Nevada gold mines 17. Hg emissions from these soil
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tubs into the air within the greenhouse bay exposed plants to an increased concentration of Hg in
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the air at night. Air was pulled W-E across the greenhouse bay using a large swamp cooler
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during the day, but circulation was shut off at night. The fourth setting was outside VRG and
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adjacent to Interstate-80 (HIWY) that has ~123,000 mobile sources pass by each day (NDOT,
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personal communication, Mark Wooster, 6/16/17). This setting was chosen because highway
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traffic is a source of oxidants that generates GOM compounds. 18 Populus were secured along a
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fence that parallels US Interstate 80. Pinus were spaced evenly between two greenhouse bays
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(see SI for details on these settings).
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For both ERF-S and ERF-A, and the VRG setting, soil of 5 trees of each species were
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spiked with a HgBr2 solution, prior to movement to the treatment location, to investigate how
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increased Hg in soil affected plant Hg concentrations. HgBr2 was used because other compounds
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HgCl2 and HgS have been used in previous experiments (see discussion below). Soil for each pot
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was “spiked” with 94 ng L-1 of a HgBr2 solution twice each week (every Monday and Friday; n=
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10 times). Soil spikes replaced regular watering that day for pots being treated and no water was
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allowed out of the pots. The HgBr2 solution of concentration18.8 µg L-1 was made by adding
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HgBr2 (Sigma Aldrich, 99.996%) to 18mΩ water. For each tree, 5 ml of the HgBr2 solution was
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added to 1 L of tap water to create the 94 ng/L solution, and then the 1 L was applied to the soil.
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During the experiment, aphids (Aphidoidea) were present on Populus leaves at all four
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sites and resulted in the loss of leaves and some trees. To control these insect infestations,
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pesticide application in the ERF bays included one application of M-PEDE® (76ml/gal, 2%,
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Gowan Company, AZ, USA), two applications of PyGanic (10ml/gallon, MGK, MN, USA), and
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the use of an organophosphate (1300 Orthene ® TR Micro Total Release Insecticide, BASF).
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Pesticide application in the Valley Road greenhouse included one application of M-PEDE® (76
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ml/gal, 2%, Gowan Company, AZ, USA), and two applications of PyGanic (10 ml/gallon, MGK,
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MN,USA). The PyGanic and M-PEDE were tested for Hg content, which was found to be
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negligible ( ERF-A (5 ng m-3). The
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Highway site (HIWY) mean 24 h TGM (2 ng m-3) was from spring and summer of 2015 as
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reported by Gustin et al. 18 GOM concentrations were higher at the HIWY location due to
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mobile source pollution (Table 1).
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Thermal desorption profiles for specific GOM compounds in ERF bays were the same.
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There was no clear indication of the HgBr2 spike in the ERF-S bay (e.g. Figure SI 3). This
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suggests a reduction or loss of Hg once permeated into the bay. Based on profiles, there was a
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mixture of HgBr2, HgCl2, Hg-nitrogen, and Hg-sulfate compounds present that were desorbed
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between 100° and 200° C (Figure SI 3). Desorption profiles for VRG showed a prominent peak
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between 170° to 190° that likely indicated a Hg sulfate/sulfur-based compound given the mine
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tailings in the greenhouse. Desorption temperature was slightly higher than the HgSO4 peak
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previously reported 22; In addition, clear evidence for HgO, HgCl2, HgBr2, and nitrogen
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compounds were sometimes observed (data not shown). In general, in Reno halogenated
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compounds may be derived from the free troposphere and from the marine boundary layer; and
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N, O, and S compounds were derived from mobile source activity adjacent to the highway,
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similar to what was reported in Gustin et al. 18 and Pierce et al.23
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Spike Results Two-way ANOVA analyses indicated foliar Hg concentrations from Pinus sampled in
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June and July were not significantly different between trees with and without the soil Hg spike
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(Table SI 1). Populus foliage after 30 days of first leaf out also showed no significant effect of
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the soil spike on foliar Hg concentrations (Table SI 1). Bark concentrations were not
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significantly different between trees with and without the soil Hg spike (Table SI 1).
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Additionally, there was no significant effect of the soil spike on the most recent growth ring (Y
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5) that grew during this experiment (Table SI 2). Because there were no significant differences
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between the root spiked and non-root spiked trees, data for each treatment were aggregated for
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further statistical tests.
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Impact of Air Exposures on Tree Tissue Hg Concentrations
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Pinus TGM exposures had a significant effect on Pinus foliage concentrations (Table SI 1).
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Highest foliar Hg concentrations (27.9 + 5.2 ng g-1) were measured in the VRG where the
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greatest 24 h TGM concentrations were recorded (Figure 1). Foliage Hg concentrations 8 ACS Paragon Plus Environment
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measured in June and July were significantly higher relative to foliage as initially received (19.6
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+ 4.0 ng g-1) in VRG (p HIWY (Figure SI 5) suggesting that nonstomatal uptake for this species
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could also be important.
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Pinus trees in this study had lower foliar Hg concentrations than Populus, which could be
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due to the influence of nonstomatal/stomatal processes of Populus. Differences in stomatal
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structures and stomatal density can influence stomatal conductance between the species as well
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as a difference in rate of gas exchange through the stomata. 9 Populus species have stomata that
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are on the underside of the leaf surface whereas Pinus species have stomata that are submerged
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in crevices on the needles. 27 Conifers have been suggested to intercept a larger amount of
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atmospheric Hg than deciduous trees resulting in higher Hg concentrations.2 Conifers receive a
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greater amount of direct Hg deposition all year because they retain their needles, but they may
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assimilate Hg in different amounts depending on the rate of stomatal conductance during the
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growing season.28 Frescholtz demonstrated that under similar conditions stomatal conductance
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was higher for aspen than pine. 26
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Stamenkovic and Gustin 3 pointed out the influence of atmospheric relative humidity
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(RH) on stomatal conductance of Populus. It is also well documented in plant physiology
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literature that stomatal conductance is reduced at higher relative humidity. 29 Stamenkovic and
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Gustin 3 found that low RH (
