Funding was made available by the Vermont Water Resources and Lake Studies Center through an agreement with the U.S. Geological Survey. Conclusions and opinions are those of the authors and not the Vermont Water Resources and Lake Studies Center or the USGS.

1. Sample collection
<ol>
	<li>Collect approximately 4 L of soil (or sediment) from desired sites. Collection areas should be relatively small to limit spatial variation in P and soil properties.</li>
	<li>Sieve samples through a coarse (20 mm) screen followed a 2 mm screen. Thoroughly hand-mix samples after sieving.</li>
	<li>Weigh out 100 g of field-moist soil or sediment. Dry in an oven at 105 &#176;C for 24 h and calculate gravimetric water content (soil water mass/dry soil mass).</li>
	<li>Take a 500 mL subsample for chemical analysis. 
	NOTE: Soil pH, organic matter content and labile inorganic P (Pi) concentration are recommended soil tests. Here, labile soil Pi availability was assessed by: 1) Pi extracted by 1.25 mol L-1 ammonium-acetate (pH = 4.8; hereafter referred to as modified Morgan extractable P) measured colorimetrically7,8, 2) Pi extracted by distilled water, and 3) P extracted by 1.25 mol L-1 ammonium-acetate (pH = 4.8) measured by inductively coupled plasma-optical emission spectroscopy (ICP)8.</li>
	<li>Use remaining sieved soil for microcosm studies or store in polyethylene bags at 5 &#176;C for later use. 
	NOTE: Soils dry out when refrigerated for long periods (&#62;30 days) and will require remoistening. Do not freeze soil samples as it affects microbial integrity and P release potential.</li>
</ol>
2. Microcosm construction
<ol>
	<li>Use one-liter (1 L) graduated polypropylene or other non-reactive plastic beakers as individual experimental units (microcosms). Wash beakers in 10% hydrochloric acid and triple rinse with distilled water.</li>
	<li>Measure 2 cm up from the bottom and place a mark next to beaker graduations. Drill a 1.25 cm diameter hole for drainage ports.</li>
	<li>Place a small bead of silicone around the inside edge of hose barb and outer circumference of the bore hole. Carefully insert the drainage port into the hole. 
	NOTE: Allow air-drying for at least 24 h before proceeding to step 2.4.</li>
	<li>Trace the outside circumference of hose barbs onto nylon mesh filter screen and cut out with scissors. Apply a thin bead of silicone around the circumference of each filter on the outside edge and press filters onto hose barb inlets. Allow at least 24 h of drying time before using. 
	NOTE: A 100 &#181;m pore size is recommended for most applications; however, finer-textured soils may require a larger filter pore size to avoid excessively long PW sample collection time.</li>
	<li>Fit a short piece of 0.625 cm diameter latex hose to hose barb ends. Attached a 3.3 cm wide paper binder clip to the hose to prevent flow.</li>
</ol>
3. Conducting a phosphorus release trial
<ol>
	<li>Place 500 mL of sample into duplicate microcosms and gently apply distilled water along beaker walls until FW reaches the 1 L mark. 
	NOTE: Allow microcosms to equilibrate for 24&#8722;48 h before taking initial samples.</li>
	<li>Unclip paper binders to induce PW flow through drainage port. Collect samples by placing clean 30 mL beakers directly beneath PW drainage ports. Allow several mL of PW to drain, discard and use the next 10 mL as a representative sample volume.</li>
	<li>Filter PW samples through 0.45 &#181;m membrane filters and immediately analyze for soluble reactive P (SRP). Record absorbance values and time of measurements. 
	NOTE: SRP is generally assumed to be orthophosphate; however, molybdate-reactive P can also form complexes with colloids and/or nanoparticles that pass through 0.45 &#181;m filters4.</li>
	<li>Take initial FW sample by inserting a 10 mL bulb syringe pipette halfway down the water column and withdraw a sample using a circular motion. Empty into beakers, filter and analyze immediately for SRP.</li>
	<li>Replace sampled water by refilling beakers to the 1 L level with distilled water. 
	NOTE: Evaporative losses will vary. The goal is to consistently maintain a total volume (flooded soil + water column) of 1 L in all microcosms. Replacing evaporative water losses has negligible dilution effects on SRP.</li>
	<li>Repeat steps 3.2 through 3.5 based on the desired number of P release time points for analysis. 
	NOTE: The number of samples taken over time depends on experimenter goals. Sampling one to three times per week is sufficient for many applications assuming incubations are close to 20 &#176;C. Incubating at higher temperatures increases SRP release rates and will require more frequent sampling. The intent here is to show the utility of the microcosm method rather than focusing on data analysis from experiments. Both kinetically-based and empirical models to fit P desorption/sorption data are presented elsewhere9,10. Since the microcosm method relies on a repeated measures design and accommodates replication and differing treatments, generalized linear mixed modeling approaches are also appropriate11.</li>
</ol>

<ol>
	<li>Patrick, W. H., Khalid, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphate+release+and+sorption+by+soils+and+sediments:+Effect+of+aerobic+and+anaerobic+conditions.">Phosphate release and sorption by soils and sediments: Effect of aerobic and anaerobic conditions.</a> Science. 186 (4158), 53-55 (1974).</li><li>Moore, P. A., Reddy, K. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+Eh+and+pH+on+phosphorus+geochemistry+in+sediments+of+Lake+Okeechobee,+Florida.">Role of Eh and pH on phosphorus geochemistry in sediments of Lake Okeechobee, Florida.</a> Journal of Environmental Quality. 23, 955-964 (1994).</li><li>Young, E. O., Ross, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphate+release+from+seasonally+flooded+soils:+a+laboratory+microcosm+study.">Phosphate release from seasonally flooded soils: a laboratory microcosm study.</a> Journal of Environmental Quality. 30 (1), 91-101 (2001).</li><li>Henderson, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anoxia-induced+release+of+colloid-+and+nanoparticle-bound+phosphorus+in+grassland+soils.">Anoxia-induced release of colloid- and nanoparticle-bound phosphorus in grassland soils.</a> Environmental Science &amp; Technology. 46 (21), 11727-11734 (2012).</li><li>Vidon, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hot+spots+and+hot+moments+in+riparian+zones:+potential+for+improved+water+quality+management.">Hot spots and hot moments in riparian zones: potential for improved water quality management.</a> Journal of the American Water Resources Association. 46 (2), 278-298 (2010).</li><li>Young, E. O., Ross, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphorus+mobilization+in+flooded+riparian+soils+from+the+Lake+Champlain+Basin,+VT,+USA.">Phosphorus mobilization in flooded riparian soils from the Lake Champlain Basin, VT, USA.</a> Frontiers in Environmental Science. 6 (120), 1-12 (2018).</li><li>McIntosh, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bray+and+Morgan+soil+extractants+modified+for+testing+acid+soils+from+different+parent+materials.">Bray and Morgan soil extractants modified for testing acid soils from different parent materials.</a> Agronomy Journal. 61 (2), 259-265 (1969).</li><li>Young, E. O., Ross, D. S., Cade-Menun, B. J., Liu, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphorus+speciation+in+riparian+soils:+a+phosphorus-31+nuclear+magnetic+resonance+and+enzyme+hydrolysis+study.">Phosphorus speciation in riparian soils: a phosphorus-31 nuclear magnetic resonance and enzyme hydrolysis study.</a> Soil Science Society of America Journal. 77 (5), 1636-1647 (2013).</li><li>McGechan, M. B., Lewis, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sorption+of+phosphorus+by+soil:+Part+1.+Principles,+equations,+and+models.">Sorption of phosphorus by soil: Part 1. Principles, equations, and models.</a> Biosystems Engineering. 82 (1), 1-24 (2002).</li><li>Cabrera, M. L., Radcliffe, D. E., Cabrera, M. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modeling+phosphorus+in+runoff:+Basic+approaches.">Modeling phosphorus in runoff: Basic approaches.</a> Modeling Phosphorus in the Environment. , 65-81 (2007).</li><li>Gbur, E. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Analysis of Generalized Linear Mixed Models in the Agricultural and Natural Resources Sciences. , (2012).</li><li>Moore, P. A., Reddy, K. R., Fisher, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphorus+flux+between+sediment+and+overlying+water+in+Lake+Okeechobee,+Florida:+Spatial+and+temporal+variations.">Phosphorus flux between sediment and overlying water in Lake Okeechobee, Florida: Spatial and temporal variations.</a> Journal of Environmental Quality. 27 (6), 1428-1439 (1998).</li><li>Young, E. O., Briggs, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphorus+concentrations+in+soil+and+subsurface+water:+A+field+study+among+cropland+and+riparian+Buffers.">Phosphorus concentrations in soil and subsurface water: A field study among cropland and riparian Buffers.</a> Journal of Environmental Quality. 37 (1), 69-78 (2008).</li><li>Hoffmann, C. C., Kjaergaard, C., Uusi-K&#228;mpp&#228;, J., Hansen, H. C. B., Kronvang, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphorus+retention+in+riparian+buffers:+review+of+their+efficiency.">Phosphorus retention in riparian buffers: review of their efficiency.</a> Journal of Environmental Quality. 38 (5), 1942-1955 (2009).</li><li>Bartlett, R. J., Ross, D. S., Tabatabai, M. A., Sparks, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemistry+of+Redox+Processes+in+Soils.">Chemistry of Redox Processes in Soils.</a> Chemical Processes in Soils, SSSA Book Ser. 8. , 461-487 (2005).</li><li>Radcliffe, D. E., Freer, J., Schoumans, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diffuse+phosphorus+models+in+the+United+States+and+Europe:+Their+usages,+scales,+and+uncertainties.">Diffuse phosphorus models in the United States and Europe: Their usages, scales, and uncertainties.</a> Journal of Environmental Quality. 38 (5), 1956-1967 (2009).</li><li>Bartlett, R. J., James, B. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studying+dried,+stored+soil+samples&#8212;some+pitfalls.">Studying dried, stored soil samples&#8212;some pitfalls.</a> Soil Science Society of America Journal. 44 (4), 721-724 (1980).</li><li>Turner, B. L., McKelvie, I. D., Haygarth, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+water+extractable+soil+organic+phosphorus+by+phosphatase+hydrolysis.">Characterization of water extractable soil organic phosphorus by phosphatase hydrolysis.</a> Soil Biology &amp; Biochemistry. 34 (1), 27-35 (2002).</li><li>Turner, B. L., Haygarth, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phosphorus+solubilization+in+rewetted+soils.">Phosphorus solubilization in rewetted soils.</a> Nature. 411, 258 (2001).</li><li>Sparks, D. L., Sparks, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinetics+of+reactions+in+pure+and+mixed+systems.">Kinetics of reactions in pure and mixed systems.</a> Soil Physical Chemistry. , (1986).</li></ol>

The authors declare that this work was conducted in the absence of any commercial or financial relationships that could be interpreted as a potential conflict of interest.

A main technical advantage of the microcosm approach is its ability to simulate in-situ conditions whereby saturated soil or sediment is immediately overlain by FW that may substantially differ in redox and P status. Landscapes with variable source area hydrology such as drainage ditches, flooded cropland, wetlands, and riparian/near-stream zones are all examples of where reduced PW is periodically overlain by more oxidized water with lower Pi concentrations. These redox gradients can strongly affect Pi sorption...

Phosphorus (P) is a critical limiting nutrient for both crop and aquatic biomass productivity. Surface water hydrology is a main driver of P fate and transport, as it controls the physical transport of sediment and P while also affecting remobilization potential during runoff and flooding/ponding events. Various laboratory-based extraction methods are typically used to estimate P release at the field scale under oxidizing conditions. While different mechanisms can contribute to P release, reductive dissolution of iron-phosphates is a well-established reaction mechanism that can lead to large orthophosphate-P fluxes to water1,2,3,4. In a review of mechanisms controlling P biogeochemistry in wetlands, redox status was hypothesized to be the main variable controlling P release to soils and shallow groundwater5. As such, traditional P tests may not be reliable predictors of P release under prolonged saturation.
Given the importance of water residence time and redox status on P fate and transport, laboratory approaches designed to better simulate in situ conditions could lead to improved P transport risk indices for agricultural and wetland ecosystems subject to variable saturation. Since orthophosphate is immediately bioavailable, the rate and extent of desorption during saturation can be used as an index of nonpoint source P pollution risk. Our method was designed to quantify P desorption to porewater (PW) and mobilization to overlying floodwater (FW), a typical condition in areas with variable source area hydrology (e.g., flooded agricultural fields, wetlands, drainage ditches, and riparian/near-stream zones). The method was originally developed to characterize P release potential in seasonally flooded soils from northern New York (USA) and recently applied to quantify P desorption potential of riparian soils from northwestern Vermont&#8217;s Lake Champlain Basin6. Here, we provide a protocol for the laboratory microcosm method and highlight results from a recently published study demonstrating its ability to quantify P desorption potential. We also demonstrate the relationship between P release potential and the reliability of routine soil tests (labile extractable P, pH) to predict release across sites.
Carrying out the method requires access to an analytical laboratory with adequate climate control, ventilation, water, and a proper acid waste disposal system. The method presumes access to routine chemical reagents and laboratory equipment (sinks, hoods, glassware, etc.). Beyond routine laboratory needs, a membrane filtration (&#8804; 0.45 &#181;m) system is required and a UV spectrophotometer to measure P. A pH meter or multiparameter water quality probe are also recommended but not required. Laboratory temperature is an important factor and should be kept constant unless temperature itself is being investigated as an experimental factor (20 &#176;C is recommended). Unhindered access to an adequate analytical laboratory with proper equipment is a prerequisite to perform the method properly and generate meaningful results.

<table>
	<tbody>
		<tr>
			<td>1.25 cm plastic hose barbs</td>
			<td>numerous</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Chemical reagents for phosphorus determination</td>
			<td>numerous</td>
			<td>NA</td>
			<td>P analysis capability is assumed; refer to cited references for details on method</td>
		</tr>
		<tr>
			<td>Chordless or electric drill with 1.25 cm bit</td>
			<td>numerous</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Graduated plastic beakers (1L)</td>
			<td>numerous</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Laboratory with fume hoods, temperature control, and acid waste disposal system</td>
			<td>NA</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Nylon mesh filter screen (100um)</td>
			<td>numerous</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>Silicone</td>
			<td>numerous</td>
			<td>NA</td>
			<td></td>
		</tr>
		<tr>
			<td>UV Spectrophotometer</td>
			<td>numerous</td>
			<td>NA</td>
			<td></td>
		</tr>
	</tbody>
</table>

measuring phosphorus release in laboratory microcosms for water quality assessment

Phosphorus (P) is a critical limiting nutrient in agroecosystems requiring careful management to reduce transport risk to aquatic environments. Routine laboratory measures of P bioavailability are based on chemical extractions performed on dried samples under oxidizing conditions. While useful, these tests are limited with respect to characterizing P release under prolonged water saturation. Labile orthophosphate bound to oxidized iron and other metals can rapidly desorb to solution in reducing environments, increasing P mobilization risk to surface runoff and groundwater. To better quantify P desorption potential and mobility during extended saturation, a laboratory microcosm method was developed based on repeated sampling of porewater and overlying floodwater over time. The method is useful for quantifying P release potential from soils and sediments varying in physicochemical properties and can improve site-specific P mitigation efforts by better characterizing P release risk in hydrologically active areas. Advantages of the method include its ability to simulate in situ dynamics, simplicity, low cost, and flexibility.

Results from a recent study focused on P release potential of riparian areas are highlighted to demonstrate the method&#8217;s ability to characterize site-level P release dynamics6. While some soils showed minimal changes in SRP over time, others had large increases in PW- and FW-SRP concentrations (Figure 1). Two sites with contrasting trends are shown in Figure 1. Soil 7 is a riparian site with low soil pH and characterized by nearly c...

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Measuring Phosphorus Release in Laboratory Microcosms for Water Quality Assessment

Accurate quantification of phosphorus (P) desorption potential in saturated soils and sediments is important for P modeling and transport mitigation efforts. To better account for in situ soil-water redox dynamics and P mobilization under prolonged saturation, a simple approach was developed based on repeated sampling of laboratory microcosms.

Phosphorus (P) is a critical limiting nutrient in agroecosystems requiring careful management to reduce transport risk to aquatic environments. Routine ...

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6.4K Views.  United States Department of Agriculture, Agricultural Research Service (USDA-ARS), Institute for Environmentally Integrated Dairy Management. Accurate quantification of phosphorus (P) desorption potential in saturated soils and sediments is important for P modeling and transport mitigation efforts. To better account for in situ soil-water redox dynamics and P mobilization under prolonged saturation, a simple approach was developed based on repeated sampling of laboratory microcosms.

Video: Measuring Phosphorus Release in Laboratory Microcosms for Water Quality Assessment

Laboratory Estimation of Net Trophic Transfer Efficiencies of PCB Congeners to Lake Trout (Salvelinus namaycush) from Its Prey

A technique for laboratory estimation of net trophic transfer efficiency (&#947;) of polychlorinated biphenyl (PCB) congeners to piscivorous fish from their prey is described herein. During a 135-day laboratory experiment, we fed bloater (Coregonus hoyi) that had been caught in Lake Michigan to lake trout (Salvelinus namaycush) kept in eight laboratory tanks. Bloater is a natural prey for lake trout. In four of the tanks, a relatively high flow rate was used to ensure relatively high activity by the lake trout, whereas a low flow rate was used in the other four tanks, allowing for low lake trout activity. On a tank-by-tank basis, the amount of food eaten by the lake trout on each day of the experiment was recorded. Each lake trout was weighed at the start and end of the experiment. Four to nine lake trout from each of the eight tanks were sacrificed at the start of the experiment, and all 10 lake trout remaining in each of the tanks were euthanized at the end of the experiment. We determined concentrations of 75 PCB congeners in the lake trout at the start of the experiment, in the lake trout at the end of the experiment, and in bloaters fed to the lake trout during the experiment. Based on these measurements, &#947; was calculated for each of 75 PCB congeners in each of the eight tanks. Mean &#947; was calculated for each of the 75 PCB congeners for both active and inactive lake trout. Because the experiment was replicated in eight tanks, the standard error about mean &#947; could be estimated. Results from this type of experiment are useful in risk assessment models to predict future risk to humans and wildlife eating contaminated fish under various scenarios of environmental contamination.

Laboratory Estimation of Net Trophic Transfer Efficiencies of PCB Congeners to Lake Trout Salvelinus namaycush from Its Prey

A technique for laboratory estimation of net trophic transfer efficiency of polychlorinated biphenyl (PCB) congeners to piscivorous fish from their prey is presented. To maximize applicability of the laboratory results to the field, the piscivorous fish should be fed prey fish that are typically eaten in the field.

A technique for laboratory estimation of net trophic transfer efficiency (&#947;) of polychlorinated biphenyl (PCB) congeners to piscivorous fish from ...

Laboratory-determined Phosphorus Flux from Lake Sediments as a Measure of Internal Phosphorus Loading

Eutrophication is a water quality issue in lakes worldwide, and there is a critical need to identify and control nutrient sources. Internal phosphorus (P) loading from lake sediments can account for a substantial portion of the total P load in eutrophic, and some mesotrophic, lakes. Laboratory determination of P release rates from sediment cores is one approach for determining the role of internal P loading and guiding management decisions. Two principal alternatives to experimental determination of sediment P release exist for estimating internal load: in situ measurements of changes in hypolimnetic P over time and P mass balance. The experimental approach using laboratory-based sediment incubations to quantify internal P load is a direct method, making it a valuable tool for lake management and restoration.
Laboratory incubations of sediment cores can help determine the relative importance of internal vs. external P loads, as well as be used to answer a variety of lake management and research questions. We illustrate the use of sediment core incubations to assess the effectiveness of an aluminum sulfate (alum) treatment for reducing sediment P release. Other research questions that can be investigated using this approach include the effects of sediment resuspension and bioturbation on P release.
The approach also has limitations. Assumptions must be made with respect to: extrapolating results from sediment cores to the entire lake; deciding over what time periods to measure nutrient release; and addressing possible core tube artifacts. A comprehensive dissolved oxygen monitoring strategy to assess temporal and spatial redox status in the lake provides greater confidence in annual P loads estimated from sediment core incubations.

Lake eutrophication is a water quality issue worldwide, making the need to identify and control nutrient sources critical. Laboratory determination of phosphorus release rates from sediment cores is a valuable approach for determining the role of internal phosphorus loading and guiding management decisions.

Eutrophication is a water quality issue in lakes worldwide, and there is a critical need to identify and control nutrient sources. Internal phosphorus (P) ...

EPA Method 1615. Measurement of Enterovirus and Norovirus Occurrence in Water by Culture and RT-qPCR. Part III. Virus Detection by RT-qPCR

EPA Method 1615 measures enteroviruses and noroviruses present in environmental and drinking waters. This method was developed with the goal of having a standardized method for use in multiple analytical laboratories during monitoring period 3 of the Unregulated Contaminant Monitoring Rule. Herein we present the protocol for extraction of viral ribonucleic acid (RNA) from water sample concentrates and for quantitatively measuring enterovirus and norovirus concentrations using reverse transcription-quantitative PCR (RT-qPCR). Virus concentrations for the molecular assay are calculated in terms of genomic copies of viral RNA per liter based upon a standard curve. The method uses a number of quality controls to increase data quality and to reduce interlaboratory and intralaboratory variation. The method has been evaluated by examining virus recovery from ground and reagent grade waters seeded with poliovirus type 3 and murine norovirus as a surrogate for human noroviruses. Mean poliovirus recoveries were 20% in groundwaters and 44% in reagent grade water. Mean murine norovirus recoveries with the RT-qPCR assay were 30% in groundwaters and 4% in reagent grade water.

Here we present a procedure to quantify enterovirus and norovirus in environmental and drinking waters using reverse transcription-quantitative PCR. Mean virus recovery from groundwater with this standardized procedure from EPA Method 1615 was 20% for poliovirus and 30% for murine norovirus.

EPA Method 1615 measures enteroviruses and noroviruses present in environmental and drinking waters. This method was developed with the goal of having a ...

Automated Gel Size Selection to Improve the Quality of Next-generation Sequencing Libraries Prepared from Environmental Water Samples

Next-generation sequencing of environmental samples can be challenging because of the variable DNA quantity and quality in these samples. High quality DNA libraries are needed for optimal results from next-generation sequencing. Environmental samples such as water may have low quality and quantities of DNA as well as contaminants that co-precipitate with DNA. The mechanical and enzymatic processes involved in extraction and library preparation may further damage the DNA. Gel size selection enables purification and recovery of DNA fragments of a defined size for sequencing applications. Nevertheless, this task is one of the most time-consuming steps in the DNA library preparation workflow. The protocol described here enables complete automation of agarose gel loading, electrophoretic analysis, and recovery of targeted DNA fragments. 
In this study, we describe a high-throughput approach to prepare high quality DNA libraries from freshwater samples that can be applied also to other environmental samples. We used an indirect approach to concentrate bacterial cells from environmental freshwater samples; DNA was extracted using a commercially available DNA extraction kit, and DNA libraries were prepared using a commercial transposon-based protocol. DNA fragments of 500 to 800 bp were gel size selected using Ranger Technology, an automated electrophoresis workstation. Sequencing of the size-selected DNA libraries demonstrated significant improvements to read length and quality of the sequencing reads.

This manuscript describes an automated gel size selection approach for purifying DNA fragments for next-generation sequencing. The Ranger Technology provides complete automation of the entire process of agarose gel loading, electrophoretic analysis, and recovery of targeted DNA fragments allowing for high-throughput and high quality next-generation sequencing libraries.

Next-generation sequencing of environmental samples can be challenging because of the variable DNA quantity and quality in these samples. High quality DNA ...

A CO2 Concentration Gradient Facility for Testing CO2 Enrichment and Soil Effects on Grassland Ecosystem Function

Continuing increases in atmospheric carbon dioxide concentrations (CA) mandate techniques for examining impacts on terrestrial ecosystems. Most experiments examine only two or a few levels of CA concentration and a single soil type, but if CA can be varied as a gradient from subambient to superambient concentrations on multiple soils, we can discern whether past ecosystem responses may continue linearly in the future and whether responses may vary across the landscape. The Lysimeter Carbon Dioxide Gradient Facility applies a 250 to 500 &#181;l L-1 CA gradient to Blackland prairie plant communities established on lysimeters containing clay, silty clay, and sandy soils. The gradient is created as photosynthesis by vegetation enclosed in in temperature-controlled chambers progressively depletes carbon dioxide from air flowing directionally through the chambers. Maintaining proper air flow rate, adequate photosynthetic capacity, and temperature control are critical to overcome the main limitations of the system, which are declining photosynthetic rates and increased water stress during summer. The facility is an economical alternative to other techniques of CA enrichment, successfully discerns the shape of ecosystem responses to subambient to superambient CA enrichment, and can be adapted to test for interactions of carbon dioxide with other greenhouse gases such as methane or ozone.

A CO2 Concentration Gradient Facility for Testing CO2 Enrichment and Soil Effects on Grassland Ecosystem Function

The Lysimeter Carbon Dioxide Gradient Facility creates a 250 to 500 &#181;l L-1 linear carbon dioxide gradient in temperature-controlled chambers housing grassland plant communities on clay, silty clay, and sandy soil monoliths. The facility is used to determine how past and future carbon dioxide levels affect grassland carbon cycling.

Continuing increases in atmospheric carbon dioxide concentrations (CA) mandate techniques for examining impacts on terrestrial ecosystems. Most ...

Characterization and Application of Passive Samplers for Monitoring of Pesticides in Water

Five different water passive samplers were calibrated under laboratory conditions for measurement of 124 legacy and current used pesticides. This study provides a protocol for the passive sampler preparation, calibration, extraction method and instrumental analysis. Sampling rates (RS) and passive sampler-water partition coefficients (KPW) were calculated for silicone rubber, polar organic chemical integrative sampler POCIS-A, POCIS-B, SDB-RPS and C18 disk. The uptake of the selected compounds depended on their physicochemical properties, i.e., silicone rubber showed a better uptake for more hydrophobic compounds (log octanol-water partition coefficient (KOW) &#62; 5.3), whereas POCIS-A, POCIS-B and SDB-RPS disk were more suitable for hydrophilic compounds (log KOW &#60; 0.70).

A protocol about the characterization and application of five different passive sampling devices is presented.

Five different water passive samplers were calibrated under laboratory conditions for measurement of 124 legacy and current used pesticides. This study ...

Controlled-release of Chlorine Dioxide in a Perforated Packaging System to Extend the Storage Life and Improve the Safety of Grape Tomatoes

A controlled-release chlorine dioxide (ClO2) pouch was developed by sealing a slurry form of ClO2 into semipermeable polymer film; the release properties of the pouch were monitored in containers with or without fruit. The pouch was affixed to the inside of a perforated clamshell containing grape tomatoes, and the effect on microbial population, firmness, and weight loss was evaluated during a 14 day storage period at 20 &#176;C. Within 3 days, the ClO2 concentration in the clamshells reached 3.5 ppm and remained constant until day 10. Thereafter, it decreased to 2 ppm by day 14. The ClO2 pouch exhibited strong antimicrobial activity, reducing Escherichia coli populations by 3.08 log CFU/g and Alternaria alternata populations by 2.85 log CFU/g after 14 days of storage. The ClO2 treatment also reduced softening and weight loss and extended the overall shelf life of the tomatoes. Our results suggest that ClO2 treatment is useful for extending the shelf life and improving the microbial safety of tomatoes during storage without impairing their quality.

Here, we describe a protocol for the application of a novel, slow-release ClO2 product that reduces spoilage and extends the shelf life of fresh fruit. The slow-release ClO2 product was added to standard commercial grape tomato packaging and tested against Escherichia coli and Alternaria alternata.

A controlled-release chlorine dioxide (ClO2) pouch was developed by sealing a slurry form of ClO2 into semipermeable polymer film; the release properties ...

Measuring and Mapping Patterns of Soil Erosion and Deposition Related to Soil Carbonate Concentrations Under Agricultural Management

Spatial patterns of soil erosion and deposition can be inferred from differences in ground elevation mapped at appropriate time increments. Such changes in elevation are related to changes in near-surface soil carbonate (CaCO3) profiles. The objective is to describe a simple conceptual model and detailed protocol for repeatable field and laboratory measurements of these quantities. Here, accurate elevation is measured using a ground-based differential global positioning system (GPS); other data acquisition methods could be applied to the same basic method. Soil samples are collected from prescribed depth intervals and analyzed in the lab using an efficient and precise modified pressure-calcimeter method for quantitative analysis of inorganic carbon concentration. Standard statistical methods are applied to point data, and representative results show significant correlations between changes in soil surface layer CaCO3 and changes in elevation consistent with the conceptual model; CaCO3 generally decreased in depositional areas and increased in erosional areas. Maps are derived from point measurements of elevation and soil CaCO3 to aid analyses. A map of erosional and depositional patterns at the study site, a rain-fed winter wheat field cropped in alternating wheat-fallow strips, shows the interacting effects of water and wind erosion affected by management and topography. Alternative sampling methods and depth intervals are discussed and recommended for future work relating soil erosion and deposition to soil CaCO3.

Spatial patterns of soil erosion and deposition can be inferred from differences in ground elevation mapped at appropriate time increments. Such changes in elevation are related to changes in near-surface soil carbonates. Repeatable methods for field and laboratory measurements of these quantities and data analysis methods are described here.

Spatial patterns of soil erosion and deposition can be inferred from differences in ground elevation mapped at appropriate time increments. Such changes ...

Experimental Column Setup for Studying Anaerobic Biogeochemical Interactions Between Iron (Oxy)Hydroxides, Trace Elements, and Bacteria

Fate and speciation of trace elements (TEs), such as arsenic (As) and mercury (Hg), in aquifers are closely related to physio-chemical conditions, such as redox potential (Eh) and pH, but also to microbial activities that can play a direct or indirect role on speciation and/or mobility. Indeed, some bacteria can directly oxidize As(III) to As(V) or reduce As(V) to As(III). Likewise, bacteria are strongly involved in Hg cycling, either through its methylation, forming the neurotoxin monomethyl mercury, or through its reduction to elemental Hg&#176;. The fates of both As and Hg are also strongly linked to soil or aquifer composition; indeed, As and Hg can bind to organic compounds or (oxy)hydroxides, which will influence their mobility. In turn, bacterial activities such as iron (oxy)hydroxide reduction or organic matter mineralization can indirectly influence As and Hg sequestration. The presence of sulfate/sulfide can also strongly impact these particular elements through the formation of complexes such as thio-arsenates with As or metacinnabar with Hg.
Consequently, many important questions have been raised on the fate and speciation of As and Hg in the environment and how to limit their toxicity. However, due to their reactivity towards aquifer components, it is difficult to clearly dissociate the biogeochemical processes that occur and their different impacts on the fate of these TE.
To do so, we developed an original, experimental, column setup that mimics an aquifer with As- or Hg-iron-oxide rich areas versus iron depleted areas, enabling a better understanding of TE biogeochemistry in anoxic conditions. The following protocol gives step by step instructions for the column set-up either for As or Hg, as well as an example with As under iron and sulfate reducing conditions.

Experimental Column Setup for Studying Anaerobic Biogeochemical Interactions Between Iron OxyHydroxides, Trace Elements, and Bacteria

Fate and speciation of arsenic and mercury in aquifers are closely related to physio-chemical conditions and microbial activity. Here, we present an original experimental column setup that mimics an aquifer and enables a better understanding of trace element biogeochemistry in anoxic conditions. Two examples are presented, combining geochemical and microbiological approaches.

Fate and speciation of trace elements (TEs), such as arsenic (As) and mercury (Hg), in aquifers are closely related to physio-chemical conditions, such as ...

Continuous Hydrologic and Water Quality Monitoring of Vernal Ponds

Vernal ponds, also referred to as vernal pools, provide critical ecosystem services and habitat for a variety of threatened and endangered species. However, they are vulnerable parts of the landscapes that are often poorly understood and understudied. Land use and management practices, as well as climate change are thought to be a contribution to the global amphibian decline. However, more research is needed to understand the extent of these impacts. Here, we present methodology for characterizing a vernal pond's morphology and detail a monitoring station that can be used to collect water quantity and quality data over the duration of a vernal pond's hydroperiod. We provide methodology for how to conduct field surveys to characterize the morphology and develop stage-storage curves for a vernal pond. Additionally, we provide methodology for monitoring the water level, temperature, pH, oxidation-reduction potential, dissolved oxygen, and electrical conductivity of water in a vernal pond, as well as monitoring rainfall data. This information can be used to better quantify the ecosystem services that vernal ponds provide and the impacts of anthropogenic activities on their ability to provide these services.

Understanding the ecosystem services and processes provided by vernal ponds and the impacts of anthropogenic activities on their ability to provide these services requires intensive hydrologic monitoring. This sampling protocol using in-situ monitoring equipment was developed to understand the impact of anthropogenic activities on water levels and quality.

Vernal ponds, also referred to as vernal pools, provide critical ecosystem services and habitat for a variety of threatened and endangered species. ...