Module 4 Stream Ecology Laboratory

1. Module 4 Stream Ecology Laboratory Measuring Periphyton Biomass

2. Periphyton Biomass Measurements • Ash-Free Dry Weight • Pigment Analysis • Biovolume Measurement

3. Periphyton Biomass Measurements • Many methods have been developed to collect periphyton from natural substrates. All involve quanitatively removing material from a known surface area. • Different methods may be needed to collect periphyton from rocks, woody debris, aquatic macrophytes, or other substrates. • Substrate size will also determine how samples are collected. Large boulders require the use of a device that prevents the sample from being lost downstream. If the substrate is a fist-sized cobble the rock can be paced in a pan, scraped, then washed into a bottle with stream water.

4. One way to scrape a known area is to lay a plastic 35 mm slide (film removed) over the rock and scrape off the material within the slide area (scrub area = 2.3cmX3.5cm=8cm2).

5. Rocks don’t always look like they have much on them Nearly all the stuff scrubbed off this one was organic matter –most of it living algae S.Loeb and J.Reuter images Periphyton (attached algae) sampling

6. Resulting material from a rock scrub contains macroinvertebrates, detritus, fungi, bacteria, as well as algae.

7. Here’s a portion of the previous sample after being pulled on to a GF/C filter is preparation for chlorophyll extraction or AFDW determination.

8. Pigment Analysis The filter is then placed in 90% acetone to extract the chlorophyll.

9. Ash-Free Dry Weight Determination The filter is dried to a constant weight then ashed at 555 ºC to burn off all of the organic matter. The difference in weight is called ‘loss upon ignition” and is a measure of the % organic content of the material Muffle furnace

10. Once you have a measure of chlorophyll or AFDW you’ll need to calculate per unit area.

11. Periphyton Biovolume • Involves the examination of the algal cells under the microscope • Then measuring the cell dimensions with an ocular or stage micrometer and calculating cell volume.

12. Periphyton Biovolume • Counting and measuring individual algae • Identification to genus and species level • Calculate cell volume by approximation to nearest geometrical shape • Count cells over a known area of the slide so cells per unit volume can be determined Some formulas to estimate biovolume from cell dimensions (also see Wetzel & Likens 2000) B B A A A Rod Sphere Ellipsoid

13. Taxonomic keys often include questions about size. Determining size is basically like using a ruler. The standard ruler for a microscope is called an "ocular micrometer", which is fitted into the eyepiece of your microscope. If you don’t have a micrometer, you can use the diameter of the field. Each of these methods requires that you first standardize your microscope against a ruler of known length; at low magnification, this standard could be a transparent office ruler, but at higher magnifications a stage micrometer is needed. Be aware that different microscopes are not exactly the same and the size goes down with increased magnification.