Juvenile check

  • Algae feed was not flowing and had been off since the night before (8pm?) because Sam turned the pump off to stop the flow to his feed silo. We got it turned back on by 11am (so animals in heath stacks had ~15 hours without food).

  • Shuffled heath trays:
    Old order:
      1. T1
      2. T5
      3. T7
      4. T3
        New order:
      5. T7
      6. T3
      7. T5
      8. T1
  • Didn’t measure or count any animals but took pictures of each tray. Maybe can get measurements using imageJ and using the tube diameter for reference
  • T1
  • T5
  • T7
  • T3

Water chem for Sam’s experiment

  • measured discrete pH, salinity, and temperature for each heath tray in Sam’s experiment, and also for H1T1, H1T2, and H2T6. Measurements were as follows:

    • pictures of measurements are here

  • did TA for all trays as well as H1T1 and H1T2. Didn’t do any TA for H2 because we haven’t been keeping track of that

    • H2T6 was -30.1 mV where H1T1 was -35.4 mV and both are being fed by the same head tank. These measurements are close enough to indicate that there is no difference in the trays

Juvenile Respirometry for Sam’s experiment

  • incubator set at 17C
  • Laptop on, presens software open, set file path (save file in folder) 101429
  • Plate reader in incubator on rotator. Sam set’s incubator 1 degree below what the treatment water is.The vials are stored in the cabinet because they have a light sensitive sensor on the bottom 101615
  • Sam filters the treatment water through a 0.2um filter so that he can remove microbial influence on respiration. 101800
  • This is Sam’s plate map to track what animal/treatment each vial contains. 101825
  • He fills the vials half full with treatment water before adding animals 101833 101853
  • He adds 3 animals to each vial using chopsticks, then tops vials off with treatment water in excess so that water forms a convex surface 103515
  • He caps the vials and inverts them to check for air bubbles. If air bubbles exist, he uncaps, adds more water, wipes the vial top dry, taps out the cap, recaps, and inverts again to check for bubbles again. 104906

  • Each vial gets placed back into it’s respective well in the 96-well plate and a foam top gets taped on over the vials to hold them in place during the whole trial. This prevents the vials from shifting around in the plate during readings. The whole thing gets taped onto the plate reader on the rotator and switched on before starting the run.

  • Controls and animal vials should read the same initially (~200umol), then animal vials should show a decrease in O2 over time.

  • Trial runs ~20 min taking readings every 15 sec.

standardization metrics

  • shell length: take pictures of animals with red olympus camera on tripod and reference key
  • weigh water inside vials to account for differences in water volume between vials
    • sam has tried seeing if respiration rate correlates with shell length, but it’s pretty fuzzy data. Biovolume (measured by total vial volume - water volume (inferred from weight) correlates even more fuzzy with respiration rate. It’s not clear what the best way to normalize is.

possible optimizations:
Seems like more 24 well plates would help:

  1. for keeping vials organized
  2. for taking pictures faster and taking 1 picture of all animals as opposed to individual pictures

preserving juveniles for Sam’s experiment

  • Preserved 3 individuals in one tube and 1 individual in one tube (3x) for each “tank” (tanks are small plastic petry dish style)
    • I did this by taking out each individual tank and using chopsticks as Sam demonstrated to place animals into screwcap tubes (labeled with numbers up to 1150). Started with tanks in the back of the tray and worked my way forward
    • I only did this for the heath trays on the left and the bottom middle tray. I did the last row of the top middle tray, but didn’t have time to complete the rest of that tray.
    • In all tanks many animals with broken shells (at least half). Sam said this was worse for the animals conditioned in low pH.

Odds and ends

  • new thermometer blinks read when turned on. Nothing in manual about this. Sam said he hasn’t experienced any problems with it except one time that he plugged the probe into the device port incorrectly.
  • brought back Sam’s -80C samples in dewer that Kaitlyn brought out last week. They are in the -80C rack 2 columns 3, 4, and 5
  • Power supply cord for 2nd Apex that isn’t properly functioning is still missing. It was last plugged in in the hatchery when Sam had tried to troubleshoot it. After that, he and Kaitlyn unplugged everything and brought it all back to the dry lab. It’s not certain where the cord went. We looked in all the dry lab cabinets and in the hatchery without luck.
  • brought back mesh tubes to prepare for outplanting at the end of August