Thursday 2016-09-01

• Checked for mortalities
  • CA1T (2), CA4B (2), CA2T (1), 011 from OR1T
• I bought 150 white shellfish tags (1/8″ x 1/4″) labelled 000-149 in order to label some of the oysters to be used in the OA experiment. This is to help determine which population an oyster belongs to in case it gets mixed up during sampling.
  • Sean used seawater epoxy to attach tags to some of the oysters that will be used in the adult OA experiment. Before attaching tags, he recorded weight for the oyster in grams. He tried to randomize the oysters chosen for the experiment by size.
  • Since some oysters already had orange shellfish tags (labelled 001-074) from the summer larval experiment, all subsequent labels will have a “W” to denote the ID of an oyster with a white tag. To cure the epoxy, these oysters were left in a single layer on the bottom of a bucket with an airstone and flow-through of algae and seawater.

• I took pictures and counted the number of live oysters on some of the tiles from the larval experiment, as well as emptied out some silos that still had larvae in them. If any larvae had set on the side of the silos, I counted them and scraped them off into a 1.5 mL tube with RNALater. Tiles with oysters on them were left hanging in a bucket fed by the oyster broodstock manifold.
  • Tiles w/oysters: B61, B89, B67, A48
  • Tiles w/out oysters: A82, A71
  • Silos: A26, A48, A27
• I dissected adductor muscle from some California and Oregon oysters that were not going to be used in the adult OA experiment.
  • CA4B_A, CA4B_B, CA4B_C (tiny oyster attached to CA4B_B),CA4B_D, CA4B_E, CA4B_F, CA4B_G, CA4B_H, CA4B_I, CA4B_J, CA4B_K
  • OR2T_E, OR2T_F, OR2T_G, OR2T_H, OR2T_I, OR2T_J, OR2T_K, OR2T_L, OR2T_M, OR2T_N, OR2T_O, OR2T_P, OR2T_Q, OR2T_R
  • OR5B_A, OR5B_B
  • OR2B_A, OR2B_B (attached to C), OR2B_C (attached to B), OR2B_D, OR2B_E, OR2B_F

Wednesday 2016-08-31

• Screened out silos from the new new experiment started ??. Day 7

• Counted and took pics of tiles that had already been removed from silos. For tiles that had at least 20 oysters I sampled ~10 oysters and stored in RNALater.

Monday 2016-08-29

• Screened out silos from new new experiment started ?? for Day 7

• Counted newly released larvae
  • over 10,000: CA2_T, OR5_B, CA4_B (mostly dead)
• Screen out silos with tiles in them. Counted planktonic larvae in silo, spat on sides of silo, and spat on tiles. Rinsed off tiles gently with freshwater.

Out with the old…

In a classic fieldwork moment, I had just finished setting up one system for my broodstock, when a newly published paper and a phone chat changed my mind and I had to almost completely redo it. Initially, I had decided to split my adults into 4 or 5 buckets per population of ~25 oysters each in order to maximize genetic diversity by minimizing the chance of one male fertilizing all females. I built a manifold for this system (new term I learned meaning “pipe branching into several openings”) and had 0.5 gal/hour drippers delivering 16degC seawater and algae to the buckets. The broodstock were taken out of their combined tank on Tues. May 31 and separated into a single 5 gal bucket per population. They were split up into ~25 per bucket on Wed. June 15, with 4 buckets for California and 5 buckets each for BC and OR. My cleaning schedule for buckets was to rinse them and wash with Vortexx 3 times a week, and change out the drippers every day I was at the hatchery with ones that had been sitting overnight in Vortexx.

Then on Friday June 17 I read this new paper out of George Waldbusser’s lab at OSU. They conducted a carbonate chemistry manipulation experiment with fertilized eggs extracted from brooding Olympia oysters. A quick reminder- male Olympia oysters release sperm into the water column which is then filtered out by females for fertilization. The females then keep the fertilized eggs in their mantle cavity for up to 12 days before releasing the now weakly swimming larvae. In Waldbusser et al. 2016,they found that larvae extracted ~36 hours after fertilization developed at a similar rate as larvae left in the brood chamber. As far as I know, this was one of the first experiments where Ostrea larvae had been raised outside of the brood. In order to identify a newly fertilized female, they looked for eggs that had been essentially spit out by an adult oyster. Chatting with Matthew Gray from the project was really helpful in figuring out how I can adapt my system to identify brooding females and possibly try to run some experiments of my own on newly fertilized larvae.

…in with the new

On Thursday June 23, I did away with most of the buckets and laid out oysters 2″ apart in black plastic 9 gal tubs (originally meant for cement mixing), with 40-50 oysters per tub and 3 tubs for BC and 2 tubs for CA and OR making 7 tubs total. This number was primarily dictated by space, or else I would’ve split them up into more trays. Some oysters were stuck together in a way that made it impossible to separate without killing them, and so were put in 3gal buckets with small 100 um banjo filters. Each tray has an airstone, either two 1/2 gal/hour drippers or a single 1 gal/hour dripper, and a banjo filter.

Cleaning these trays is a pain as they are too heavy and awkward to pick up and pour out into a bucket, so we use a hose to drain the tray into a bucket and keep the broodstock in a clam bag and separate bucket of seawater during cleaning and filtering for larvae. As we screened out the first batch of larvae on Thursday, we will be cleaning all trays/buckets every day to check for larvae. The official start of the experiment will be when all three populations are releasing enough larvae.

Friday May 13 was my first day back out at Manchester. This summer I will be conducting an acidification stress experiment on Olympia oyster larvae from San Francisco Bay, CA, Coos Bay, OR, and Ladysmith Harbor, BC. Wild adults were collected from each site in December and January and shipped in coolers to a quarantine room at the Manchester Research Station so that the parents may experience the same over-wintering conditions together. They were kept in the same large tank in separate, triply labelled clam bags and fed a mix of live algae from the hatchery and algae paste.

Since arriving in Washington, I’ve mostly been getting my experiment system ready and conditioning my broodstock for spawning. There have been some logistical issues, as a rock scallop project also housed in the quarantine room hasn’t completed yet so we’ve had to adapt the OA system for the possibility of both experiments running at the same time. With many Home Depot/Harrington’s Plastics runs and hours of fitting PVC pipe together, it has felt like slow going. At the very least, if science doesn’t work out I think I’m now qualified to be an assistant plumber!

Attached below is a todo list document I made at one point to help keep track of all the things that needed to be accomplished.

Exp_5_27_16

One great improvement for this summer over last is the help of a part-time intern, Sean, who recently graduated from the University of Washington. As someone who has extensive experience building and running shellfish experiments through his work with the Friedman lab, I’m sure I couldn’t complete this project without his help.