METHODS

CURRENT ASSESSMENT AND MONITORING APPROACH

130 Iowa lakes were sampled three times over the 2011 monitoring season. Three sets of samples in a year have been shown to offer high adequate precision for the estimation of an annual mean for several important analytes (Downing et al., 2006a). Sampling times were arranged to represent spring-early summer, mid-summer, and late summer-fall. Monitoring samples were taken at one site in each lake basin, chosen to represent the open-water zone, since this has been shown to be an appropriate sampling method for most lakes (Bachmann et al., 1980; Bachmann et al., 1994; Downing et al., 2006b). Two field teams traveled throughout the state collecting water samples representing the upper mixed zone of each lake. Sampling was performed using a 2 m fixed upper mixed zone sampler since this method is more efficient and has been shown to be comparable to full-column upper mixed zone sampling. This sampling scheme has been chosen to ensure comparability with previous surveys (Bachmann et al., 1980; Bachmann et al., 1994; Downing et al., 2006a).

FIELD SAMPLE COLLECTION

Field technicians sampled at the deepest point in each lake basin, as determined by sonar and existing bathymetric maps, and recorded the spatial locations of sampling points using a Geographical Positioning System (Garmin 76CS). In-field deployment of YSI’s 6-Series, Multi-parameter Water Quality Monitors, provided profile data for temperature, dissolved oxygen, specific conductivity, pH, and turbidity.

YSI profile data were used to determine the depth or absence of the thermocline. Thermocline depth was defined as the region of maximum rate of temperature decrease (>1 ºC per meter) with respect to depth. If a thermocline was present and less than 2 meters, the water sample was taken from the surface to the depth of the thermocline. If no thermocline was present or was deeper than 2 meters, the water column was sampled from the surface to a depth of 2 meters.

Once these measurements were made and before collecting a water sample, the sampling equipment was triple rinsed with lake water on the opposite side of the boat from where the sample would be collected to avoid inter-lake sample contamination (Wedepohl et al., 1990). A mixed column sample was then collected using a column sampler, from the appropriate sample depth (as determined by measurements of thermocline depth or absence). The water was emptied from the column sampler into the pre-rinsed holding container and additional column samples were pulled and added to the container until enough water was collected to fill all required sample bottles.

Once a sufficient volume of water had been obtained, the holding container was agitated to mix the sample before transferring to the bottles. The water was then poured from the holding container into pre-cleaned sampling bottles using the triple washed funnel making sure not to touch the funnel to the inside of the sample bottles. This method was used to collect water samples for nutrient, chlorophyll, suspended solid, and phytoplankton analysis. Immediately after returning to shore, phytoplankton samples were preserved with 0.3 mL Lugol’s solution per 100 mL of sample collected (American Public Health Association, 1998).

Zooplankton samples were collected by vertically towing a Wisconsin net (63 µm mesh size) from the depth of the thermocline to the surface with a maximum depth of 9 m. If a thermocline did not exist a sample was taken from approximately 0.5 m off the bottom of the lake to the surface or from a depth of 9 m to the surface. To collect the sample, the Wisconsin net was lowered to the determined depth and then raised vertically at an even speed of approximately 0.5 m/s (American Public Health Association, 1998). The sample was rinsed down into the filter cup using a squirt bottle containing deionized water. The filter cup was removed and the sample is rinsed into a 125 mL bottle and filled approximately half full (about 60 mL). Immediately after returning to shore, zooplankton samples were preserved with approximately 60 ml of 10% Formalin solution containing sucrose (Haney and Hall, 1973) to create a 5% Formalin solution.

All sample bottles are labeled with sample location, unique site ID, sampler’s name(s), date and time of collection, and depth of column sampled (or vertical tow depth for zooplankton). All samples were kept cold until delivered to the laboratory for analysis.

LABORATORY METHODS

WATER CHEMISTRY

Soluble Reactive Phosphorus as P

Soluble reactive phosphorus samples are analyzed with an HP 8453 Spectrophotometer using the ascorbic acid method in Standard Methods (American Public Health Association, 1998). Analysis involves filtering the samples through a 0.45 µm syringe-tip filter to remove particulate matter, allowing color to develop for 10-30 minutes, creating standard curves (forced through origin) daily, running known samples and spikes with each group of samples and correcting for background turbidity using a 3-point drop correction.

Total Phosphorus

Total phosphorus samples are analyzed with an HP 8453 Spectrophotometer using the ascorbic acid method in Standard Methods (American Public Health Association, 1998). Analysis involves digesting the samples with persulfate, allowing color to develop for 10- 30 minutes, creating standard curves (forced through origin) daily, running known samples and spikes with each group of samples and correcting for background turbidity using a 3-point drop correction.

Ammonia+Ammonium as N

Ammonia nitrogen samples are analyzed with an HP 8453 Spectrophotometer using the phenate method in Standard Methods (American Public Health Association, 1998). Analysis involves filtering samples through a 0.45 µm syringe-tip filter to remove particulate matter, generating standard curves (forced through origin) daily and running known samples and spikes with each group of samples.

Nitrate+Nitrite as N

Nitrate + nitrite (NOx) samples are analyzed with an HP 8453 Spectrophotometer using the cadmium reduction method (4500-NO3, E) in Standard Methods (American Public Health Association, 1998). Analysis involves passing samples through a column of cadmium granules treated with copper sulfate, generating standard curves (forced through origin) daily and running known samples and spikes with each group of samples.

Total Kjeldahl Nitrogen

Total Kjeldahl Nitrogen (TKN) samples are analyzed with an HP 8453 Spectrophotometer using the colorimetric EPA method 351.2 v2 (EPA, 1993). Analysis involves digesting samples in a Technicon BD-40 block digestor, creating standard curves (forced through origin) daily, and running known samples and spikes with each group of samples.

Dissolved Organic Carbon

Dissolved organic carbon samples are analyzed with a Shimadzu TOC-V Analyzer using the high-temperature combustion method in Standard Methods (American Public Health Association, 1998). Analysis involves filtering samples through a 0.45 µm syringe-tip filter, acidifying and purging with CO2 to remove inorganic carbon, creating standard curves (forced through origin) daily, and running known samples with each group of samples.

Suspended Solids (Total, Volatile, & Inorganic)

Suspended solids samples are analyzed using the non-filterable residue method in Standard Methods (American Public Health Association, 1998) and the volatile residue EPA method 160.4 (Kopp and McKee, 1983). Analysis involves filtering samples through a pre-rinsed and pre-weighed 0.45 µm filter, drying at 105°C, weighing to determine TSS, combusting at 550°C, and weighing to determine VSS and ISS fractions. Known samples are run with each group of samples.

Chlorophyll a (corrected for pheophytin)

Chlorophyll a samples are analyzed with a TD-700 Fluorometer using the non-acidified fluorometry EPA method 445.0 (Arar & Collins, 1997). Analysis involves filtering samples through a 0.45 µm filter, extracting in 100% acetone using a probe sonicator (Jeffrey et al., 1997), creating standard curves (forced through origin) annually, and running known standards with each group of samples.

Total Alkalinity as CaCO3

Total alkalinity samples are analyzed with a Thermo Orion 950 Analytical Titrator using the titration method in Standard Methods (American Public Health Association, 1998). Analysis involves a temperature-compensated pH measurement and automated titration to pH = 4.5.

BIOLOGY ANALYSIS

Phytoplankton

Phytoplankton samples were concentrated, sub-sampled, and examined using a Leitz DM IL inverted microscope at 200x power (American Public Health Association, 1998) and analyzed using methods similar to those described by Lund et al. (1958), a variant of the Utermöhl (1958) method. Phytoplankton were identified to genus, counted, and measured. Simple geometric model formulae were used to calculate the biovolume per liter of six divisions and the total phytoplankton community (Findenegg, 1974; Hillebrand et al., 1999). The biovolume per liter was converted to wet mass per liter using a 1:1 ratio (Sournia, 1978).

Zooplankton

Zooplankton samples were concentrated, sub-sampled, and examined using a Nikon SMZ 1500 Stereoscopic Zoom Microscope and photographed using a Qimaging Retiga 2000R Digital Still Camera at 96x power (American Public Health Association, 1998). Zooplankton were identified to family, counted, and measured and length-weight regressions were used to calculate the dry mass per liter of the Cladocera and Copepoda (Dumont, et al., 1975). Rotifera dry mass per liter was determined by first calculating the biovolume per liter (Ruttner-Kolisko, 1977) then converting this measurement to dry mass per liter (Doohan, 1973 for all rotifers except Asplancha; Dumont et al., 1975 for Asplancha; McCauley, 1984).