Basic HTML Version
SOP7 – Live Organisms >50 μm
54
8.2.2. Challenge Condition (Control)
Transfer sample to large beaker/small-pail (6 L minimum) and make up volume to 3 L. Agitate
appropriately (via random motion stirring with stirring plate) and subsample 2 ml with Stempel
pipette. This will result in a 1/1500 split; note split and subsample volumes. Other splits/beaker-
volumes may be used if too few or too many animals are present; however, a 1/1500 split tends
to be a good place to start. Aim for >200 but <400 animals/tray. Place subsample in labyrinth
sorting tray. Note all taxa present. Quantify dead organisms only, but do so by taxa. Perform 3-
replicates of this assay. Do not fail to note overall conditions such as blooms, high debris,
seasonal assembly shifts, or anything else that might be of later interest. The control (challenge-
condition) live-analysis procedure is the same for fill and discharge. Later offsite analysis of the
fixed samples gives total concentration and proportion dead at time of live-analysis.
When control sample analysis is complete, sieve/rinse with pre-filtered treatment water into pre-
labeled 500 ml wide-mouth Nalgene HDPE sample bottle. Continue to maintain separation of
equipment and only use filtered, control water in processing. Adjust volume in bottle to about
400 ml (the pre-filled 10 L carboy of filtered control water is used to refill squirt bottle) and fix
with 45 ml full-strength formaldehyde to obtain 10% formalin. Rinse and wash all equipment
that has been in contact with sample or ambient water with freshwater prior to placing in storage.
If any questions of taxonomy have arisen, they may be addressed at leisure using the fixed
samples, which will be kept on archive at least until reports have been finalized.
8.3. Fixed Sample Analysis
Materials and Supplies
Amount
Dissecting scope, such as Leica Wild MZ8 or
equivalent
1
Controllable-intensify fiber-optic light source
1
Labyrinth sorting trays, such as Bogorov or
equivalent
1
3” diameter sieves, 35 µm max mesh size
1
Squirt bottle filled with Tap Water
1
Small funnel
1
Probe
1
Data sheets and pen
As needed
DC3-SD camera
1
Computer w MicroCap image capture software 1
Total number of organisms or to more accurately identify organisms can be done from the fixed
samples collected above. Once the samples have been preserved for >24 hours a sample can
then be processed for total organism (dead + live) counts employing time-tested methods (Omori
& Ikeda 1984, UNESCO 1968).
Sieve and wash sample with tap water to remove formaldehyde, into an approved formaldehyde
waste container. Rinse bottle several times into sieve to clean Transfer sample to large
beaker/small-pail (6 L minimum) and make up volume to 5 L. Agitate appropriately (via random
motion stirring) and subsample 2 ml with Stempel pipette. This will result in a 1/2500 split; note