232
Exercise 15
7. Examine the filters under blue light with an epifluorescent microscope. Ciliates
will autofluoresce a light apple-green color. Shift to high power and enumerate
the number of micro spheres per ciliate. Count 50 to 100 cells on each filter.
8. Plot the number of micro spheres per ciliate versus time.
9. Calculate the feeding rate from the slope of the line for microspheres ingested
versus time multiplied by the ratio of bacterial cells to fluorescent microspheres.
Questions
1. The rate of ingestion of micro spheres is typically linear for the first several time
points and then declines. Why?
2. What is the purpose of the time zero sample? Were any microspheres "ingested"
at time zero?
3. How would you test the idea that the fluorescent micro spheres provide an accurate
estimate of feeding on bacteria?
4. Calculate the clearance rate from your estimate of ingestion. How does this rate
compare with that measured for the large zooplankton in the previous experiments?
Now divide the clearance rates ofthe zooplankter and the ciliate by their respective
weights or volumes. Weights can be estimated from size by assuming that the animals conform to simple geometric shapes (see Exercise 10) and then by converting
volume estimates to weight assuming a specific gravity of 1. The estimate of size
corrected feeding rate is called the specific rate. Which rate is greater and what
does this imply about the relative significance of a unit of protozoan biomass
versus a unit of zooplankton biomass?
Apparatus and Supplies
Zooplankton
1. Cultures of algal or bacterial food supplies.
a. Cultures of spherical algae ofa size between 10 and 20 Jim (e.g., Chlamydomonas)
and between 30 and 50 j.Lm.
b. Cultures of large algae, preferably a blue-green alga such as Anabaena.
c. If available, cultures of bacteria of large size (e.g., Pseudomonas) or yeast (e.g.,
Rhodotorula ).
2. Media for the culturing of algae [e.g., Stein (1973)] and micro flora [e.g., Rodina
(1972)].
3. Flasks or beakers, ca. 500 ml; graduated cylinders for volume measurements.
4. Zooplankton, preferably several species in pure or nearly pure cultures.
5. Counting cells to evaluate cell numbers of food organisms (see Exercise 10).
6. Algae or bacteria labeled with H 14 C0 3 or 32P04.
7. Safety apparatus for handling isotopes and labeled materials (e.g., waterproofed
absorbent pads, gloves, cleaning agents, and disposal facilities).
8. Scintillation vials. Accurate volumetric pipe (e.g., Eppendorf) for small volumes.
Appropriate biological solubilizers (e.g., Beckman NCS or BTS-450) and wateraccepting scintillation reagents (e.g., Packard Instagel). Scintillation radio assay
systems.
9. Membrane filtration apparatus for filtration oflabeled food sources for subsequent
radioassay (see Exercise 14).
10. Fine-mesh sieves (cylinders of plastic about 10 cm in diameter and 5 cm in length,
Exercise 15
7. Examine the filters under blue light with an epifluorescent microscope. Ciliates
will autofluoresce a light apple-green color. Shift to high power and enumerate
the number of micro spheres per ciliate. Count 50 to 100 cells on each filter.
8. Plot the number of micro spheres per ciliate versus time.
9. Calculate the feeding rate from the slope of the line for microspheres ingested
versus time multiplied by the ratio of bacterial cells to fluorescent microspheres.
Questions
1. The rate of ingestion of micro spheres is typically linear for the first several time
points and then declines. Why?
2. What is the purpose of the time zero sample? Were any microspheres "ingested"
at time zero?
3. How would you test the idea that the fluorescent micro spheres provide an accurate
estimate of feeding on bacteria?
4. Calculate the clearance rate from your estimate of ingestion. How does this rate
compare with that measured for the large zooplankton in the previous experiments?
Now divide the clearance rates ofthe zooplankter and the ciliate by their respective
weights or volumes. Weights can be estimated from size by assuming that the animals conform to simple geometric shapes (see Exercise 10) and then by converting
volume estimates to weight assuming a specific gravity of 1. The estimate of size
corrected feeding rate is called the specific rate. Which rate is greater and what
does this imply about the relative significance of a unit of protozoan biomass
versus a unit of zooplankton biomass?
Apparatus and Supplies
Zooplankton
1. Cultures of algal or bacterial food supplies.
a. Cultures of spherical algae ofa size between 10 and 20 Jim (e.g., Chlamydomonas)
and between 30 and 50 j.Lm.
b. Cultures of large algae, preferably a blue-green alga such as Anabaena.
c. If available, cultures of bacteria of large size (e.g., Pseudomonas) or yeast (e.g.,
Rhodotorula ).
2. Media for the culturing of algae [e.g., Stein (1973)] and micro flora [e.g., Rodina
(1972)].
3. Flasks or beakers, ca. 500 ml; graduated cylinders for volume measurements.
4. Zooplankton, preferably several species in pure or nearly pure cultures.
5. Counting cells to evaluate cell numbers of food organisms (see Exercise 10).
6. Algae or bacteria labeled with H 14 C0 3 or 32P04.
7. Safety apparatus for handling isotopes and labeled materials (e.g., waterproofed
absorbent pads, gloves, cleaning agents, and disposal facilities).
8. Scintillation vials. Accurate volumetric pipe (e.g., Eppendorf) for small volumes.
Appropriate biological solubilizers (e.g., Beckman NCS or BTS-450) and wateraccepting scintillation reagents (e.g., Packard Instagel). Scintillation radio assay
systems.
9. Membrane filtration apparatus for filtration oflabeled food sources for subsequent
radioassay (see Exercise 14).
10. Fine-mesh sieves (cylinders of plastic about 10 cm in diameter and 5 cm in length,