Grace’s Notebook: June 30, 2017

Re-submitted the Oly Population Structure Paper to Marine Ecology.

Did more Oly seed morphology measurements.

Link to image folder: here

Link to spreadsheet: here

To do:

Silo 9, Day 11

Silo 3 and 9 Day 13

Yaamini’s Notebook: Preliminary Spawning Plan

It’s time to think about crosses!

After my oysters are done conditioning, I will strip spawn them and rear larvae. Understanding which males and females to cross, and how many to use, are important to ensure genetic diversity in the F1 generation.

From each tank, I will pool eggs, but keep sperm separate. I will cross the pooled eggs from one tank with sperm from all males from all tanks. The next day, I will measure hatch success, then pool D-hinge larvae by treatment.

Table 1. Number of males available for crosses, assuming there is a 50/50 male-to-female ratio in each tank. Tanks 1-3 were exposed to low pH conditions, and tanks 4-6 had ambient water conditions. Number of oysters from each tank can be found here.

Tank Number of Males
1 9
2 9
3 7
4 8
5 9
6 8
Heat Shock 6
Total 56

With 56 males to cross and 7 different “tanks”, there are 392 possible crosses. If I were to limit the number of males used, I would have less crosses.

Table 2. Number of crosses for varying numbers of males used.

Number of Males Number of Crosses
1 49
2 98
3 147
4 196
5 245

After measuring hatch rate, I can pool my individual crosses into groups based on treatments.

Table 3. Treatment pool combinations. In total, there are nine possible pools.

Parent 1 Parent 2
Low Female Ambient Male
Low Male Ambient Female
Low Female Heat Shock Male
Low Male Heat Shock Female
Ambient Female Heat Shock Male
Ambient Male Heat Shock Female
Low Female Low Male
Ambient Female Ambient Male
Heat Shock Female Heat Shock Male

Each pool will be transferred to 28 micron silos, so I will need replicates. Assuming I can fit four totes on each level of the table at Manchester, and each tote can fit 8 silos, I can have a maximum of 64 silos. The number of replicates I need will depend on how many larvae hatch and my stocking density. Molly recommends having 15-25 larvae per mL of silo volume.

Table 4. Number of silos needed based on replicates used for each pool.

Replicates Total Silos Needed
2 18
3 27
4 36
5 45
6 54
7 63

Here’s what I need to do next:

  • Consult with Steven and Brent on the number of crosses, and where I can trim crosses
  • Obtain drippers (1 L/hr drippers for first few days, 1 gallon/hr drippers for remainder of larval period
  • Determine flow rate and pipe diameter needed for manifolds
  • Build manifolds
  • Ensure I have enough tubing to feed silos
  • Find a method to volumetrically count eggs and sperm

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Yaamini’s Notebook: Manchester Conditioning Update 3

Revised (final?) Conditioning Plan

Date Temperature Feeding
6/30/17 15 ºC 490 mL
7/1/17 16 ºC 700 mL
7/2/17 17 ºC 700 mL
7/3/17 18 ºC 750 mL
7/4/17 19 ºC 750 mL
7/5/17 20 ºC 800 mL
7/6/17 21 ºC TBD
7/7/17 22 ºC TBD
7/8/17 23 ºC TBD
7/9/17-7/28/17 23 ºC TBD

Overall notes:

  • Feeding needs to be increased to a point where the water is dark and the oysters are not clearing it
    • Start with 700 mL Reed’s paste in 200 L of water, and increase by 50 mL until water is the desired color
    • Laura will take care of feeding on 7/1/17 and 7/3/17, and I’ll be back on 7/5/17 to check on it
  • Dosing rate is 55%-60%
  • Temperatures indicated are desired, will need to adjust heaters and check AVTECH accordingly
  • Tanks should be vacuumed daily, and drained and cleaned at least once a week
  • If oysters start dying
    • Hold temperature for at least one day
    • Feed 100 mL more Reed’s
    • Resume temperature conditioning after 1-2 days (handler’s discretion)
  • After 7/15/17, oysters should no longer be drained and cleaned

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Yaamini’s Notebook: Manchester Conditioning Update 2

Sophie’s New Choice: Temperature, Flow or pH?

This week, I’ve been at Manchester trying to balance temperature, flow and pH in the conditioning set-up. On June 25, Laura told me that the pH in my tanks was reading very low (around 7.3). With all of the excess feeding, the oysters are respiring so much that they’re modifying the chemistry in the tanks.

Monday, June 26

I went ot Manchester to adjust the flow rate and raise the pH. First, I vacuumed the tanks and checked mortality. One oyster died recently from Tank 6A. I increased the flow to 1.5 L/min from 1.2 L/min, which raised the pH to 7.7. I then fed the oysters 490 mL of Reed’s Shellfish Diet in 200 L of freshwater, dosing at 55%. When I left, the pH was between 7.65 and 7.7.

Wednesday, June 28

After monitoring pH for a day, I found it was still dropping below 7.6 in my conditioning tanks. Additionally, the 800 W heater was unable to maintain my desired temperature with the increased flow. I drained and cleaned the oyster tanks, finding one dead oyster in Heat Shock A. I increased the flow to 2.4 L/min which stabilized the pH once more, but caused temperature to drop.

Thursday, June 29

First, I vacuumed the oyster tanks and bleached the algal line. No mortalities found!

Tag Label A B Total
1 8 9 17
2 9 8 17
3 7 6 13
4 9 7 16
5 9 8 17
6 7 8 15
Heat Shock 5 6 11
Full Amb 7 8 15
Spare 2 3 5
Total 63 63 126

Table 1. Current oyster count in conditioning set-up.

While feeding the oysters, I stabilized the pH at 7.7-7.8 by increasing flow to 2.7 L/min.

img_7820

Figure 1. pH and temperature reading after flow stabilization.

I took another 800 W heater from the lab and added it to my header tank. I set the original heater at 16 ºC. The second heating unit’s control box was in Farenheit, so I set it to 68 ºF to see what would happen.

img_7813

Figure 2. In order to plug in the second heater, Olivia and I had to move the AVTECH unit to a different outlet.

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Figures 3-4. Temperature settings on controllers. I wanted to the heated water temperature to be 16 ºC, so I set the old heater setpoint to 16 ºC and the new one to 68 ºF.

screen shot 2017-06-30 at 10 53 02 am

Figure 5. AVTECH computed pH values over the past 30 days. The pH dropped below 7.6 on June 17. It is now holding around 7.73.

screen shot 2017-06-30 at 10 54 31 am

Figure 6. AVTECH temperature readings for the past 10 days. The temperature held around 18ºC until June 25. Temperature reached a peak of 20ºC on June 26, dropping after an increase in flow. Temperature is now at 15.43ºC.

Now that I’ve stabilized pH, my next step is to revise my modified conditioning plan.

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