We designed a long-term water diversion experiment introducing two frequencies of water additions from a natural pond to trenches excavated to the depth of ice-cement. In Dec. 2011 we established three sets of permanent plots (7.5 X 15m) on the south-facing hillslope above Many Glaciers Pond in Taylor Valley. High-resolution LIDAR imaging, instrumentation, and comprehensive pre-treatment sampling of all plots was conducted in Dec. of 2011, and in Jan. 2013, prior to an additional round of pre-treatment sampling and then initiation of the experiment anticipated in Jan. 2014. Instrumentation consists of thermocouples, delta-T moisture probes and water activity probes buried at multiple active-layer depths every 2 m down-gradient from the water-addition trenches. The three different plots will receive the following different treatments:

1. PRESS Treatment (H): Annual water additions sufficient to raise soil water to 10% water content (as observed during the melt event in Jan. 2002). This is achieved by pumping water from Many Glaciers Pond to a holding tank above the experimental plots, from which we will apply water by gravity feed to a trench during the warmest week of the year (typically the 1st or 2nd week of Jan.).
2. PULSE Treatment (L): Same as above, but water diversions added only in alternate years to simulate a lower frequency of thawing permafrost
3. Positive Control (C): Trenched and instrumented, but no water additions

Sampling campaigns consist of 84 soil samples (28 per treatment) collected from permanent plots for quantification of soil biota (invertebrates determined by microscopy and bacteria by 16S rRNA sequencing) and intensive geochemical analyses. This experiment will address the overarching hypothesis: Climate warming in the McMurdo Dry Valleys will amplify connectivity among landscape units leading to enhanced coupling of nutrient cycles across landscapes, and increased biodiversity and productivity. Soil samples were taken for organism enumeration and moisture content analysis as follows: Sampling bags were prepared with one sterile Whirlpak bag and clean plastic scoop per sample. Samples were taken from within the 1m2 area of each plot. The location of the sampling was recorded each year so that areas were not re-sampled. Using the plastic scoop, soil was collected to 10 cm depth. Very large rocks (more than 20 mm diameter) were excluded from the sample. The soil was shoveled into the Whirlpak bag until three quarters full (about 1.5 kg soil). The soil was mixed well in the bag, then the bag was closed tightly, expelling as much air as possible. The soil samples were stored in a cooler for transportation. On return to the laboratory (within 8 hours of sampling), the soils were stored at +4C until further processing. In the laboratory, soil samples were handled in a laminar flow hood to prevent contamination. The Whirlpak bags of soil were mixed thoroughly prior to opening. A sub-sample of approximately 50g was removed and placed in a pre-weighed aluminum soil can, and weighed on a balance accurate to 0.01g. This sample was dried at 105C for 24 hours. The sample was removed, placed in a desiccator to cool down, and re-weighed. These data were used to calculate water content of the soil. The remaining soil in the plastic beaker was weighed. Cold tap water was added up to 650 mL. The soil suspension was stirred carefully (star stir or figure of 8) for 30 seconds, using a spatula. Immediately the liquid was poured into wet screens - a stack of 40 mesh on top of a 400 mesh. The screens were rinsed gently with ice cold tap water (from a wash bottle) through the top of the stack, keeping the screens at an angle as the water filtered through. The water was kept on ice at all times. The top screen was removed, and the lower screen rinsed top down, never directly on top of the soil, but at the top of the screen and from behind. The water was allowed to cascade down and carry the particles into the bottom wedge of the angled screen. The side of the screen was tapped gently to filter all the water through. The suspension was rinsed from the front and the back, keeping the screen at an angle and not allowing the water to overflow the edge of the screen. The soil particles were backwashed into a 50mL plastic centrifuge tube, tipping the screen into the funnel above the tube and rinsing the funnel gently. The suspension was centrifuged for five minutes at 1744 RPM. The liquid was decanted, leaving a few mL on top of the soil particles. The tube was filled with sucrose solution (454g sucrose per liter of tap water, kept refrigerated) up to 45mL. This was stirred gently with a spatula until the pellet was broken up and suspended. The suspension was centrifuged for one minute at 1744 RPM, decanted into a wet 500 mesh screen, rinsed well with ice cold tap water and backwashed into a centrifuge tube. Samples were refrigerated at 4C until counted. Samples were washed in to a counting dish and examined under a microscope at x10 or x2 0 magnification. Rotifers and tardigrades were identified and counted. Nematodes were identified to species and sex, and counted. Total numbers in each sample were recorded on data sheets. All species of nematode, and all rotifers and tardigrades found in the sample were recorded. Data were entered in to Excel files, printed, and checked for errors. Extraction of chlorophyll from the soil. All procedures were carried out in the dark or very low irradiance to avoid degradation of the chlorophyll. The soil samples were mixed thoroughly in the vials, and a sample of approximately 5 g was weighed out in to a 50 mL plastic centrifuge tube with a screw-top cap. 10 mL of a 50:50 DMSO/90percent acetone solution was added to each sample and they were mixed thoroughly on a bench-top Vortex mixer for about 5 seconds. The vials were placed in a -4C constant temperature room, in the dark, and left for 12-18 hours. Determination of chlorophyll a concentration. This was determined fluorometrically using a Turner model 111 fluorometer. A calibration using a known concentration of chlorophyll was carried out prior to sample analysis. The machine was blanked using a 50:50 DMSO/90percent acetone solution. Each vial was mixed thoroughly, then centrifuged for 5 minutes at about 1800 RPM. A sample of approximately 4mL of the DMSO/acetone solution was taken from the top of the sample with a pipette, being careful not to get any soil particles in the solution. The sample was placed in a cuvette, in to the fluorometer and the fluorescence was recorded. This was done fairly quickly in order to prevent light from breaking down the chlorophyll. This measurement is called Fo, the initial fluorescence. After taking this reading, 0.1 mL of 1N HCl was added directly to the cuvette and the cuvette was gently agitated. After 20 seconds, the fluorescence was re-measured. (During this step, the acid converts the chlorophyll to phaeophytin by releasing a magnesium ion in an acidic environment). This measurement is called Fa, the fluorescence after acidification. The solution was discarded in to a waste container, and the cuvette rinsed 3 times with DMSO/90percent acetone solution before proceeding with the next sample. Data were entered in to Excel files, printed, and checked for errors. For measurements of pH, 40 of DI water was added to 20g of soil in a clean, DI- rinsed glass beaker (coarse fragments greater than 2 mm were removed). The samples were stirred until thoroughly mixed (about 5-10 sec). After sitting to equilibrate for 10 minutes the samples were stirred again and a reading was taken with a Beckman 0265 pH meter. For measurements of electrical conductivity, an additional 60 ml of DI water was then added (totaling 100 ml water). The samples were stirred until thoroughly mixed (about 5-10 sec). After sitting to equilibrate for 10 minutes the samples were stirred again and a reading was taken with a YSI 30 conductivity meter.