Take Two

1 02 2014

Recap: I am a Master of Science (and the Universe, according to my nephew when he wants the tickle attacks to stop), and my experiments in the intertidal were mostly successful, except for the live spawning attempt.  Again, to validate my project design and strengthen my simulated spawning results, we tried a second live spawning experiment at the end of last year before the permits expired… 

Me showing some abalone love (B. Blaud)

Me showing some abalone love (B. Blaud)

San Nicolas Island is one of my favorite places on earth, if you haven’t picked that up from my previous posts.  I feel incredibly blessed to have the unique opportunity to venture back, even after my schooling is complete.  I was plotting ways to get back to SNI, thinking of leveraging my experience on the island and in the intertidal to get a volunteer position with another group, when I got an email from Glenn that we were on for another live spawning experiment, and they would like me to be involved.  I didn’t even need to think about it before quickly replying, a hearty and enthusiastic “YES!  When do we leave?”

 

Withering syndrome is still a concern, even though it doesn’t appear to be as virulent as it was in the 1990s during its initial appearance on the island.  Although the disease presence has been documented on the island, the conservative approach to preventing further spread of withering syndrome dictates using only disease free animals.  The last spawning experiment involved naïve animals that were never exposed to the disease.  However, the difficulty we experienced spawning the larger animals and the learned knowledge that smaller animals are more gravid and easily spawned encouraged us to get younger red abalone.  To stay within the permit guidelines and only bring disease free animals to the island, the thirty red abalone we got for the experiment had to be treated with oxytetracycline (OTC), a broad spectrum antibiotic active against many types of bacteria, including rickettsiales.

 

The most ripe red abalone with bulging gonads were hand-picked by an abalone expert, Jim Moore, to improve our odds of a successful experiment.  Unfortunately, the OTC treatment is rather stressful, and involves the abalone sitting in a series of eight baths for 24 hours each over the course of three weeks.  During this time, the horny little abalone reabsorb their gametes, or may spawn in response to the stress.  To reduce their stress and hopefully maintain fuller gonads, the abalone were fed during the treatment.  They were allowed to recover for an additional three weeks after treatment concluded, and we hoped it would be enough.

 

Armed with a greater number of smaller abalone and knowledge from what led to failed experiments previously, we headed out to the island.  I think I inadvertently cursed myself after my last blog, Failure in the Field:

It is common for the first, and even the second field experiment to not work.  Unfortunately, I had everything riding on this one spawning release. There were many things I learned from this venture, the most obvious being to mix the iodine into the Lugol’s solution before adding it as a preservative to the eggs (duh).  I also later learned that the larger abalone are more difficult to spawn, and more successes are achieved with younger, smaller abalone.

In this second field experiment, I was in no way about to be in charge of the Lugol’s solution, and we brought formalin as a back-up preservative, just in case something else went wrong.  It’s science, and “things going wrong” is the name of the game. 

 

Red abalone in 3L of sexy spawning cocktail (Tris and hydrogen peroxide) (B. Blaud)

Red abalone in 3L of sexy spawning cocktail (Tris and hydrogen peroxide) (B. Blaud)

We set up the abalone in the temporary housing outside the archeological labs and storage, and less than a mile from a good release location at Site 2.  The plan was to spawn the abalone in the comfort of civilization, near electricity and running water, then take the gametes down to the experimental site, run the experiment at Site 2, and transport the canisters back to the lab where we would distribute eggs among the transportable tubes and add preservative.  That was the plan.

 

Now we just need the little buggers to spawn!  We added the sexy cocktail of Tris and hydrogen peroxide, ran down to quickly set everything up, and then waited… and waited… and waited…  When the tide started coming back in, and only one female had spawned a small amount of eggs in 6 hours, we decided to call it a day.  We put them all back in their temporary tank, and prepared to try again the next day.

 

Unfortunately, we were out of 6% hydrogen peroxide.  Luckily, someone thought ahead and brought back-up 3% hydrogen peroxide, purchased for $0.89 a bottle at the local pharmacy.  We doubled up the dose to try, try again.  Feeling slightly less optimistic, but still hopeful, we tried all the tricks we had learned in our combined spawning experiences and reading: temperature stress (exposure to sunlight to increase water temperature to 20°C in their 3L buckets, then water changes to bring the temperature back to 14°C); mechanical stress (EARTHQUAKE!  Shaking the buckets, tapping the buckets, mainly just agitating the poor abalone); musical stimulation (Barry Manilow, Madonna, Bruno Mars, Rhianna and Brittany – don’t judge!  It worked last time, and I was willing to try anything).  Our efforts were rewarded with some stressed out abalone that released several mucus plugs and poop, and appeared to want to spawn but had nothing to give.

 

Our conclusion: they hadn’t had enough recovery time after the OTC treatment.  Next steps include renewing the permits, applying for grants, and praying that the third try does the trick.  On the bright side, I get to go to the island at least one more time.  Can’t complain about that!





Call Me Master

14 09 2013
img_pontos

Pontus the sea, Roman mosaic, Bardo Museum (http://www.theoi.com/Protogenos/Pontos.html)

I apologize for the long hiatus to blogging.  All my focus was devoted to working on my thesis, which I fondly nicknamed Pontus (after the pre-Olympian god of the sea that fathered all fishes and sea creatures).   I got bored of saying “I’m working on my thesis at Starbucks,” or “the thesis going is rough today.”  It was much more fun to say “I’m playing with Pontus at the park” or “Pontus is in a bad mood today, so I might spend a little more time with him than originally planned.”

 

All the sweat, tears, hard work, and long nights paid off though!  They finally gave me the certificate saying everyone must call me Master.  I read the degree and it says so in the fine print.  Now that the two chapter, 97-page thesis has been submitted AND accepted, I have spent the past three weeks catching up on sleep, working as many hours as Starbucks will allow to pay off a growing number of bills  I have accumulated in graduate school, and have scoured the local area for marine biologist positions.  I do miss blogging though, and want to fill you all in on the final chapter regarding my work towards an upper-graduate degree.

 

On par with the first part of this adventure, completing the project was not easy by any means.  There were several more failed experiments with varying degrees of devastation, frustrating hours spent in front of the microscope, and weeks of hair pulling while analyzing data.  (I’m lucky my hair is so long and thick that it covers the bald spots.)

 

When I last wrote, I had described the revised experimental design for three experimental releases in the tide pool habitat.  Most black abalone, however, are found in the more cryptic crevice habitat, formed by breaks and cracks in the sandstone and shale along the shoreline.  I expanded the project to include three additional releases in a crevice habitat and compare the results between the habitat types to see if it influences fertilization success.

 

By the time I started the revised releases in the tidepool habitat, I had worked out many of the kinks in the experimental design, all discovered through trial and error.  The three tide pool releases progressed smoothly, and for some reason, I didn’t anticipate many more problems with additional releases in a different habitat type.  Yeah, I’m a slow learner.  When choosing an appropriate release location, I needed a continuous trajectory of 13m with a main unidirectional flow, and happily found this habitat just a few hundred yards from the tide pool location.  Let the new releases begin!

 

The first release in the crevice habitat was an utter catastrophe.  The waves were slightly higher, so we tried to wait until there was enough flow to distribute the particles, but not too high that it would be dangerous.  We started the experiment when the waves were slightly higher than I was comfortable with, but I didn’t want to lose the tidal height.  Unfortunately, as soon as I started the experiment, the waves and water disappeared completely.  I was so worried about protecting my bucket of surrogate sperm (worth $2400) from the now non-existent waves that I forgot to release the surrogate eggs at 10 minutes, and instead released it after 20 minutes.  Additionally, the lack of water flow left literally no water for some samples at the 2m-collection location.  That was definitely NOT awesome.

 

Bundled up against the wind launching small projectiles at all the field workers during the last experimental release (J. Ugoretz).

Bundled up against the wind launching small projectiles at all the field workers during the last experimental release (J. Ugoretz).

The second crevice habitat release went incredibly smoothly, giving me a false sense of security, because disaster struck again on the third release.  One exciting aspect to work in the crevice habitat was doing work on bare rock one moment, then being up to your chest the next moment in rushing water that is trying to pull you out to sea while you scramble to hold on to the $2400 bucket of surrogate sperm and IV stand.  Thrilling, I’m going with thrilling to describe that, and will leave out terrifying, stressful, and intimidating.  Yup, not mentioning those words at all.  During the third crevice release, one of those waves hit, and although I was able to protect my bucket of surrogate sperm, the IV stand got knocked over.  We were able to recover the stand, replace the IV bag quickly, and there was minimal amount of solution lost, so we carried on with the experiment, but it was a nerve-wracking couple of minutes.  The only thing note worthy about the last crevice release was the high wind, making all field workers feel like they were being pelted with mini-missiles.  I like my field work that way – no excitement that effects the actual sample collection.

 

But that’s not all folks…