West Coast Green Crab Experiment Part 8

Treatment Days 7-9

In which I troubleshoot a bunch of respirometry things.

7/5/2023:

  • When looking at the instability of the electrical set up, we noticed that T5, T6, and T8 aerators and T2 and T9 filters were all plugged into a TIMER OUTLET..idk when we started running an accidental deoxygenation experiment but we fixed it now.
  • Super glue was still drying on the respirometry chambers, so while it was wet we did TTR and sampled two crabs from all 15ºC and 25ºC tanks
  • Sarah and Julia cleaned all tanks, including cold room tanks
  • No mortalities, but two crabs sampled from 15ºC and 25ºC tanks
    • T1: 13
    • T2: 11
    • T3: 11
    • T4: 14
    • T5: 11
    • T6: 11
    • T7: 13
    • T8: 12
    • T9: 10
    • T10: 13
    • T11: 13
    • T12: 13
  • Sarah and Julia fixed the light set up by ziptying all of the power strips to the exterior frame of the tanks!
  • Cold room saga
    • The 5ºC tanks are now at 6.5ºC!! Since the room is at 9ºC, the chiller has to work less to get the water where it needs to be. I adjusted the room down to 7ºC to see what would happen.
    • The 10ºC tanks are at 14ºC………..so unless something drastically changes I think I’m going to scrap those tanks.
    • Emailed Rick about potential issues with the cold box but didn’t hear back (yet)
  • Respirometry saga
    • Once the chambers were dry and we were ready to roll, we encountered a few speed bumps
    • Only one probe was 0% oxygen calibrated. Chris showed me how to make a 1 g sodium sulfide in 1 L of water solution. Sodium sulfide scrubs oxygen from the water by forming sodium sulfates. This water will never retain oxygen as long as there is sodium sulfide. We used this water to 0% calibrate the chambers and probes.
    • There was no temperature logger, so I had to manually set a temperature.
    • After calibration, we were getting some really shitty data and some probes weren’t reading at all. Chris thought it was because the small tubing that held the probe in place had plastic at the end of it, so the probe wasn’t actually touching the sensor spot. We burned through the plastic with hot tweezers.
    • We also changed the intensity settings on the probe to 100%. Chris cautioned that this will be noisy, but he said that he uses this setting too because he has thick glass walls on his chambers (like us).
    • We recalibrated the probes and things looked good enough for us to run T2 respirometry! We’ll continue with respirometry tomorrow and Friday (which will be an easier lift since we’re definitely scrapping the 10ºC tanks).
  • Updated ammonia spreadsheet
  • Updated TTR spreadsheet
  • Labelled tubes for 5ºC tanks
  • Organized samples in the -80ºC.

7/6/23:

  • Got more acclimation chambers in the mail!! Sarah and Lauren covered them with trash bags and tape so they wouldn’t let light in
  • Cleaned T8 and T5 to set up acclimation chambers
  • Went to do T8 respirometry……..and noticed all of our calibration settings disappeared when we closed the program! I made more sodium sulfide solution and we recalibrated the probes. I clicked “Save Setup” to save the calibration settings for each temperature treatment afterwards.
  • During calibration, we noticed channel 1 on the USB was misbehaving regardless of which probe was being used so we used channel 4 instead
  • Completed ambient tank respirometry! After one set of acclimation chambers was done, we moved them to the next tank.
  • Did T6, T3, and T9 respirometry (in that order)
    • Probe 4 misbehaving for background measurements for T3 background
    • Probe 2 misbehaving for T9 background
    • Misbehaving: % oxygen saturation was increasing…need to talk to Chris
  • Bought more food since my food order didn’t come in
  • Cold room saga
    • 5ºC tanks ARE AT 5ºC!!!!!!
    • The 10ºC tanks are at 11.5-12ºC. I don’t think this is a meaningful difference from 14-15ºC for the ambient. And because I have less dedicated people power than I thought I did, I decided to scrap the 10ºC tanks. Julia will use these for her experiment.
  • Labelled remaining 5ºC tubes and filled with RNA later
  • TTR and sampling for 5ºC tanks
  • T7 is consisently cloudy and I don’t know why
  • No mortalities, but two crabs sampled from 5ºC and 5ºC tanks
    • T1: 11
    • T2: 11
    • T3: 11
    • T4: 12
    • T5: 11
    • T6: 11
    • T7: 11
    • T8: 12
    • T9: 10
    • T10: 13
    • T11: 13
    • T12: 13

7/7/23:

  • Lauren and I did ammonia testing and tank cleaning. Afterwards Lauren did water changes for all tanks except the 5ºC tanks (did not want to mess with water temperature prior to respirometry) and added ammonia conditioner to tanks with ammonia > 1.0
    • T1: 0
    • T4: 0.25
    • T7: 0
    • T9: 8.0
    • T3: 8.0
    • T6: 8.0
    • T2: 2.0
    • T8: 0.25
    • T5: 1.0
  • Crab escapees! Two from T2 (unknown) and 1 from T8 (15-155)
  • Performed cold respirometry
    • Tank order: T7, T4, T1
    • Saved 5ºC calibration settings
  • Reweighed 5-003 and 5-005
    • 5-003: 22.17, 5-005: 17.66
    • Updated yesterday’s TTR values
  • 5-060 recorded as used for TTR 7/3 and 7/5
    • There is no crab 5-060! It’s only 25-060
    • Inventoried T4. All but 5-066 and 5-075 accounted for. However, inventory was done in the dark during respirometry. Will re-check on Monday and relabel tubes accordingly.
  • Mortality in T10 (10-025) and another in T11 (eaten). Will inventory tanks on Monday.
    • T1: 11
    • T2: 11
    • T3: 11
    • T4: 12
    • T5: 11
    • T6: 11
    • T7: 11
    • T8: 12
    • T9: 10
    • T10: 12
    • T11: 12
    • T12: 13
  • Updated TTR spreadsheet
  • Updated survival spreadsheet
  • Updated ammonia spreadsheet

Going forward

  1. Continue doing daily checks and with experimental time points
  2. Test formatting and analysis for one set of respirometry data
  3. Respirometry analysis
  4. Update methods
  5. Update results
Written on July 7, 2023