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!

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Failure in the Field

3 11 2013

Recap: I have completed three successful experimental releases of surrogate egg and sperm in the tide pool habitat, and three successful experimental releases in the crevice habitat.  To support my project design and give plausibility to the experiments, I wanted to replicate the experimental release using live gametes from red abalone.  Last November, I received red abalone with which to practice the spawning protocols, and learned how to spawn pinto abalone up at the Mukilteo hatchery.  Now it’s time to do it in the field…

 

I realize that my last post alluded to the devastation I have felt with some of my experiments, and yeah, this is going to be the post that goes more in depth about that topic.  In truth, science makes me feel bipolar.  I reach such highs, such levels of excitement, triumph, and pure, unadulterated joy when things that shouldn’t possibly work do; but the higher the climb, the steeper the fall.

 

Camp at Site 8 (B. Blaud)

Camp at Site 8 (B. Blaud)

Once my 12 abalone (4 males and 8 females) arrived following the last experimental particle release in the crevice habitat, we set the abalone up in their temporary San Nicolas Site 8 hotel, and made our own camp.  Glenn and I would spend the next two nights camping at Site 8.  Camping was an unforgettable experience, both exciting and exhausting.  The abalone were set up in their temporary home mere feet from high tide (which happened in the middle of the night), and our tents were a short distance up the hill.  The wind was extremely high, reaching speeds of almost 35 mph, and made sleeping both uncomfortable and difficult.  That, combined with the required call-in checks every 4 hours to the Navy, and the water changes to keep the abalone clean, made solid sleep an unachievable luxury.  Did I forget to mention the sea lion serenades – it’s a thing of beauty.  These are not complaints though!  I loved every second of this – ok, I could have done without the cold, wet socks at 4am – but I loved everything else!

 

Finally, spawning day arrived.  It was a gorgeous day, like many days on San Nicolas Island.  No clouds, uncharacteristically low winds, air temperatures in the low-70s (in March – love it!!!), and I was ready to do this.  When everyone arrived, we scoped out the two areas for potential spawning (the crevice habitat and the tidepool habitat) and decided to aim for one spawning release in the tide pool habitat.  Each abalone was placed in a separate bucket, Tris and hydrogen peroxide cocktail, and then we waited.  To get the abalone in the reproducing mood, John played some Hot Chocolate, Barry Manilow, Madonna, Salt N’ Pepa, and other sexy tunes.

Spawning red abalone, marinating in a Tris and hydrogen peroxide cocktail (B. Blaud)

Spawning red abalone, marinating in a Tris and hydrogen peroxide cocktail (B. Blaud)

The spicy cocktail and sexy music worked for two abalone.  Of the 12 we brought to the island, only one male and one female released their gametes, but that’s all we needed!  Once egg and sperm were released, there was so much activity and we needed all sets of hands.  While someone attended to the abalone, getting them into clean, cool water, the gametes were taken to be washed of any residual hydrogen peroxide and be distributed between 21 canisters.  No water was added to the sperm to keep it as concentrated as possible.

 

Anchored canisters located  at -1 m, 0 m, 1 m, 2 m, 4 m, 8 m, and 12 m from the sperm release location (B. Blaud)

Anchored canisters located at -1 m, 0 m, 1 m, 2 m, 4 m, 8 m, and 12 m from the sperm release location (B. Blaud)

The live-gamete release experiment is set up much similar to the previous simulated spawning experiments.  Three canisters containing live eggs are anchored with fishing weights or sandbags at -1 m, 0 m, 1 m, 2 m, 4 m, 8 m, and 12 m from the live sperm release location.  The live sperm is released through an IV bag in the same method as the surrogate sperm in previous experiments.  There was only enough sperm for 13 minutes of release, and the canisters remained in the tide pool for an additional 30 minutes.  Once removed from the water, the eggs were preserved in falcon tubes with Lugol’s solution, and taped up for transport.  The rest of the short day was spent packing and hauling everything up the hill, everyone elated with the apparent success of the venture.

 

From my foreshadowing early on, you, my intelligent reader, may have surmised that something went horribly wrong.  Once the eggs made it back to Seattle, I opened the tubes to discover they have all degraded beyond identification.  I had not properly mixed the Lugol’s solution with the iodine, and therefore no preservative was added to the eggs.  It’s still difficult for me to think back on these moments when I discovered that all the work that everyone put in to getting abalone to the island, the nights camping on location, all the gear carried to the location, the sweat, the tears, the highs of perceived success on spawning day were all for nothing when the eggs became unidentifiable.  I won’t go into detail about the tears shed over this loss.  I had amazing support, and was heartened as several other researchers shared their tales of woe and failures 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.  While this experiment would have strengthened my project and supported my results, it is not my only experiment.  I still have a Master’s project without it.  I would like to try again.  It can still be done and may support my project once published.  I promise, I will keep you updated on any more ventures.  This part of the story is not over…





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…





Allee Effect

22 01 2013
A "crackle" of black abalone on San Nicolas Island (B. Blaud).  Crackle: a group of black abalone.

A “crackle” of black abalone on San Nicolas Island (B. Blaud). Crackle: a group of black abalone.

The Allee effect was first described by its namesake, Warder Clyde Allee in the 1930s in an article on goldfish.  His article (which I cannot access but have a summary through other sources) investigates the effect aggregation has on overall fitness on the individual goldfish, in this case.  He found that goldfish grow more rapidly in a tank that holds more individuals.  Although he never comes right out and says, “the Allee effect is defined as…”, the generally accepted definition is that higher densities within a population lead to improved individual fitness.  This basically means that larger group sizes provide individuals with more opportunities to mate, defend themselves, feed themselves, and/or can work together to alter their environment in a beneficial manner to the whole group.

 

The term, Allee effect, was often misused and erroneously applied, however, so Stephens et al. (1999) decided to take it upon themselves and define the principle in clear terms laden with scientific jargon and information.  I have a secret to admit: when I see scientific papers, my eyes gloss over.  If the paper has equations, game over.  I check out completely.  If you talk to researchers, many admit to only reading the abstract (a roughly 200 word summary of the paper at the very beginning of the paper), looking at the figures, and skimming the conclusion.  With the overwhelming amount of publications out there, that really is the only way to see if the paper is relevant to what we’re studying and saves a considerable amount of time.  Between the jargons, big words scientists use to justify the tens to hundreds of thousands of dollars spent on earning advanced degrees; you can tease out the basic information of what they’re trying to get across.

 

Black abalone on San Nicolas Island (B. Blaud)

Black abalone on San Nicolas Island (B. Blaud)

Stephens et al. (1999) paper, titled “What is the Allee effect” has large amounts of jargon as well as confusing equations.  I was able to bear down and wade through it.  I need to justify the exorbitant amount of money being spent on my education as well, and prove that I understand demographic stochasticity as much as the next overly educated person.  Side note: I think that if my spelling autocorrect in Word doesn’t know that it’s a word, I should either.  But then again, the autocorrect doesn’t know my last name, so this opinion doesn’t have strong support.  But as usual, I digress; back to the point.  After exploring the numerous examples Allee lays out to support his principle, Stephens et al. (1999) is able lay out a clear definition:

 

…we thus define the Allee effect as: a positive relationship between any component of individual fitness and either numbers or density of conspecifics.

 

Basically, the more of you there are, the better you all will be in terms of certain basic needs (reproduction, food, protection, etc).  There are limitations with this.  Too many individuals and you become competitors for resources, such as food, so oftentimes there is a threshold.  A population size below 50, for example, is bad; a population between 50 and 100 is good; and, a population over 100 is bad.

 

Stephens et al. (1999) continue with their definition to explore the distinction between the component Allee effect and the demographic Allee effect.  The distinction lies basically in the scale, where the component Allee effect is the positive relationship between any measurable component of individual fitness (breeding, feeding, or defense) and population density; the demographic Allee effect is the positive relationship between the overall individual fitness and population density.  So adding component Allee effects together produces an overall demographic Allee effect.  This is where a lot of the fun equations come into existence, but I’m going to avoid all that because I don’t want to get away from the whole point of this post, which is: how does the Allee effect apply to my project.

 

Black abalone on San Nicolas Island (B. Blaud)

Black abalone on San Nicolas Island (B. Blaud)

This leads back to the basic core question I’m trying to answer dealing with recovering black abalone populations on San Nicolas Island.  How close together do two black abalone need to be in order to successfully reproduce?  My question delves into a component Allee effect, just one aspect of individual fitness that’s investigated as a result of population density.  The more dense abalone are, the closer they are located, the more their gametes will mix and more babies will be made.  When 99% of the population was eliminated, numbers drastically decreased and density was also affected.  As they are recovering, numbers are slowly rebounding, but more interesting is the rate the population density is increasing.  Black abalone are clustering closer together as they recover, which leads us to another interesting story about the Nearest Neighbor Data.  But that is a story for another time…

 

Stephens, P.A., W.J. Sutherland, and R.P. Freckleton.  1999.  What is the Allee effect?  Oikos, Vol. 87(1): 185-190.





Invincible Black Abalone

29 12 2012
Pre-disease black abalone population on San Nicolas Island (G. VanBlaricom)

Pre-disease black abalone population on San Nicolas Island (G. VanBlaricom)

When I first began this blog, I introduced withering syndrome and it’s devastating effect on black abalone populations in Southern California.  The effect of the disease on the populations on San Nicolas Island is the equivalent of wiping out all the people in the world except for the populations in the US, Indonesia, and Brazil.  Only three of the most densely populated countries out of the total 242 countries in the world manage to survive.  It’s a little sobering when you think of it that way.  If you break it down so populations are evenly lost among each country, only the populations in Washington and Idaho would survive in the United States (I’m in Washington, so I’m happy I survive the fake apocalypse).  I’m not stating the statistics in the manner to convince everyone to move to the northwest, even though it’s the best place to live, I’m just trying to paint a picture of how the US would look if only 1% of the population survived.  Out of the 315,077,987 people in the US, only approximately 3 million make it.

 

Then, imagine among the people dodging the zombies in the apocalypse (I’ve been watching a lot of Walking Dead, so that’s the only apocalypse scenario I can picture right now), there is a group of individuals resistant to the brain-eating bacteria!  And they’re found on San Nicolas Island.

 

Disease-resistant black abalone discovered on US island

Biologists have discovered that black abalone on San Nicolas Island in the Santa Barbara Channel are more resistant to the deadly bacterial disease known as withering syndrome than abalone on the mainland. The discovery may help save these now rare intertidal molluscs from extinction, as scientists hope to soon breed these animals in captivity for release in the wild.

According to Carolyn Friedman, from the School of Aquatic and Fishery Sciences at the University of Washington in Seattle, San Nicolas Island has been the site of several severe outbreaks of withering syndrome. The present population appears to be the descendants of that 1% of the population that survived the onslaught.

Friedman and her colleague on the California Sea Grant project, Steven Roberts, also at the University of Washington, are now trying to identify which genes are responsible for resistance and the mechanisms by which this resistance is conferred. This work includes studying differences in gene expression between island black abalone and those from Carmel in Monterey, as the animals are subjected to high loads of the withering syndrome pathogen.

Withering syndrome, which causes severe atrophy of the animal’s foot muscle and is caused by a water-borne pathogen excreted in abalone faeces, occurs in relatively warm water, such as those found in the Santa Barbara Channel. Until recently, waters off the more northerly Carmel have been too cold to trigger outbreaks. As a result, abalone in Carmel have little natural protection against the disease.

Source: California Sea Grant

Anonymous.  2008.  Disease-resistant black abalone discovered on US island.  Marine Pollution Bulletin, 56(1): 7

 

Super Black Abalone (B. Blaud)

Super Black Abalone (B. Blaud)

Beginning in 2001, isolated populations on San Nicolas Island began to increase, so Dr. Friedman hypothesized that there may be disease resistant animals.  My very first trip to the island in 2004 was to collect animals and bring them back to UW to test this hypothesis.  The exposure of naïve (uncontaminated, un-diseased) black abalone from Carmel and comparing the rate of mortality to infected animals from San Nicolas Island indicated that San Nicolas Island black abalone are, in fact, resistant to the disease.  It’s a snail…it’s a rock…it’s SUPER BLACK ABALONE!  Yeah, I’m a dork.

 

But the story doesn’t end here.  Continuing this work, Dr. Friedman and PhD student Lisa Crosson investigated which genes were potentially responsible for the disease resistance in black abalone populations, and in doing so, made a surprising discovery.  But that is a story for another time…

 





Run #7

24 12 2012

Recap: My preliminary runs were exciting, although it was difficult to get used to releasing a thousand dollars in the form of small particles and watching it drift away in the ocean.  In my experiment, as designed, a single male abalone was incapable of fertilizing an egg, even if a female was spawning right next to him.  I wasn’t convince that my design was perfectly awesome since it only showed the results of a single male spawning for 10-minutes when often a spawning event takes place for up to an hour.  After revising my project design, I was ready for a second take…

My tide pool at Site 8 (B. Blaud)

My tide pool at Site 8 (B. Blaud

 

It was the perfect day for a tide-pool experiment.  The tide was low, but not too low, the sky was clear, the temperature a comfortable 72 degrees, and the waves were not too high.  By all reasoning, I should have been in a great mood, but I couldn’t relax.  The experiment has tripled in size, and we are now releasing three times as many particles into the water, which means that over $2500 is now being released into the tide pool to float away.  I couldn’t deal with any thought of failure and wanted this run to be successful so bad, I could taste it.  It tastes salty, like a combination of ocean, sweat, and tears. 

 

 

I scrambled to get everything ready: all the sample bottles clean and present, all the tubes labeled and secured, triple checked that the particles were packed (there’s nothing like hiking for an hour to realize the guest of honor is back in the room), and hoped I wasn’t forgetting anything major – I knew I would forget something, I could feel it, but was hoping it was something that could be fixed in the field.  While setting up for the run down at the site, I realized I forgot the snap-shackles, which are used to firmly anchor the nose of the IV tube in the water.  I knew I forgot something!!!  Annoying, but not major, and something that can be improvised with some well-placed rocks.  Catastrophe averted, but I still couldn’t relax.  There were too many possibilities for things to go wrong along the way.

 

Black abalone on San Nicolas Island (B. Blaud)

Black abalone on San Nicolas Island (B. Blaud)

I went over the choreography for the experiment, because in my mind, the whole thing is like one big dance.  It needs to be timed, everyone’s movements synchronized, and when it goes well, it’s quite pretty to watch.  Dave, to clarify after I described the process, asks if he is releasing his sperm at 0, 1, 5, 10, 20, 30, 45, and 60 minutes after the initial release.  Incredulous, and rather impressed that anyone would be able to release their sperm in timely intervals like that, I suggested that I be the only one to release sperm, as mine is in the form of surrogate, sperm-sized particles and not real sperm.  Everyone agreed that that would be best, and would collect samples at those intervals instead.

 

Everyone took their place, and I had enough amazing volunteers that each person was responsible for only one distance.  I was the bucket lady, running around rinsing sample bottles between collections to avoid left-over particles, making sure the sperm supply didn’t run out, keeping track of the time and orchestrating the sample collections.  Someone later suggested I should teach an aerobics class, because my voice barked orders in a no-nonsense way.  I took it as a compliment.

 

It all went rather smoothly.  I glared at Glenn when he suggested as much halfway through the experiment.  “Don’t jinx it!”  I know science and superstition don’t exactly go hand in hand, but science is about exploring the unknown.  I’m not about to tempt fate and will do anything to ensure a successful run, including not mentioning how well it’s going until I’m sure nothing can go wrong!  It’s along the same line as not saying the number seven at the craps table when the button is on.  If someone had suggested that I can also improve my odds of success by clicking my shiny, red heels together while exclaiming, “this WILL work” three times, I would have done it without a second thought, totally comfortable looking like an idiot as long as all is right in my world.  Essentially, even though it was going amazingly well, I still couldn’t relax until the last sample was collected and safely packed away.

Mel, Eric, and Glenn enjoying an E.O.D.B. at the top of Heartbreak Hill (B. Blaud)

Mel, Eric, and Glenn enjoying an E.O.D.B. at the top of Heartbreak Hill (B. Blaud)

 

Despite my fears and insecurities, it was perfect.  There are always things that can be corrected and perfected, but nothing major went wrong.  Once all my samples were stowed, I could finally smile and laugh at the sperm-related jokes.  At the end of the day, for the first time ever, I was able to hike up Heart Break Hill without stopping to “look for whales,” while carrying a 50lb backpack.  Although my backpack was heavier, weighed down with water samples, my burden was lighter now that the run was successfully completed. 

 

Since we were only able to find funds to pay for one run, the rest of the week was spent in the kitchen counting particles.  Planning for this, I brought a dissecting scope (I’m no longer going to even try to deal with a coulter counter) and all the supplies needed for counting the egg and sperm-sized particles.  With few distractions that life on SNI offers (mainly, no cell reception and limited internet access), I was able to buckle down and power through the counting, completing over half my samples.  It’s a good start!





Spawning By Myself…

20 12 2012

Recap:  To bring more credibility to my project and determine how realistic my experimental design with surrogate particles is, I will release live gametes and measure the amount of fertilized eggs in a mesh container as a function of distance from live sperm, released from a fixed distance using an IV bag.  To learn how to spawn abalone, I participated in a pinto spawning up in Mukilteo several weeks ago.  I received 16 red abalone from the Cayucos Hatchery, and have been holding them for six weeks with the intention of further honing my spawning skills…

 

My red abalone (B. Blaud)

My red abalone (B. Blaud)

Spawning by myself sounds like something completely dirty, but I assure you, it’s not.  Since November 6th, I have been holding red abalone in the basement at the University of Washington with the intention of practicing the spawning protocol I learned with pinto abalone up in Mukilteo and honing the methods to something I can use in the field as part of my experimentBy the time I was ready to practice spawning them, I had 13 red abalone, 9 females and 4 males.  I was preparing to go to San Nicolas Island (SNI) for one of my experimental runs with surrogate particles, but wanted to have one solo spawning under my belt beforehand.

 

I headed down to the basement fairly early on the Friday morning to set up for the spawning experience.  The first steps are measuring up the participants, their gonads to be exact.  I measured the gamete index for each abalone, to identify which abalone was ripe, and separated them into buckets based on gender – boys in one bucket, girls in another.  Determining ripeness was a little more difficult than I anticipated, as none of my abalone were all that ready to spawn.  I looked under each of their skirts, but had a hard time identifying males from females in many cases.  It’s supposed to be fairly straightforward.  I hold the abalone, shell down in my right hand with the base of the swirl in the shell, their head near my fingertips and the apex in my palm.  After the abalone calms down for a second, I’m able to move the foot to the side to see the gonad at the base of the shell.  The gonad index identifies ripeness based on color and the amount of swelling in the gonad.  Yellowish-creamy white means male (makes sense – sperm is white), and a greenish-purple indicates female (the eggs come out an olive green).  An index of 0 is unripe, where the gonad is indistinguishable as either male or female; an index of 1 indicates when there is slight coloration and swelling of the gonad; an index of 2 has swelling of the gonad up to the mantle, the edge of the shell; and the highest index of 3 is visibly swelled over the edge of the shell, where you can see it with barely moving the foot to the side.  All of my abalone were at my un-expert opinion a gonad index of 0 or 1. 

 

Diagram of abalone gonad (Rogers-Bennett et al. 2004 J Shellf Res 23: 553).

Diagram of abalone gonad (Rogers-Bennett et al. 2004 J Shellf Res 23: 553).

I separated the abalone I rated as a gonad index of 1 into separate buckets, then put a couple abalone rated as 0 into their own buckets, since I couldn’t conclusively determine their gender, but wanted to spawn more than a handful of my abalone.  I added the appropriate amount of Tris(hydroxymethyl) aminomethane (or Tris, for short) and 6% hydrogen peroxide, covered them in a tarp to provide darkness (and a little privacy), and then left them alone with only short, periodic checks for spawners.  I mentioned before that these are all unripe abalone with a gonad index of 0 or 1, so it goes without saying that I wasn’t overly optimistic about the possibility of getting any of them to spawn.  So, imagine my surprise when I went down after only 90 minutes of them sitting in the dark to find one male and one female spawning in separate buckets!  I was so excited, I barely remembered that I was supposed to rinse them off and put them in a bucket of clean seawater (with no chemical additions), so they could finish spawning and have usable gametes.  As soon as that was done, I ran up a couple flights of stairs to find Glenn, who would share my astonishment and simple joy at these unripe, unready abalone suddenly releasing their gametes.

 

Hatched abalone larvae (http://www.lib.noaa.gov/)

Hatched abalone larvae (http://www.lib.noaa.gov/)

Eventually, after about 3 hours, all the red abalone had been rinsed and placed in buckets of clean seawater.  Due to the unripe nature of the abalone, they didn’t spawn a significant amount of gametes.  It was a great exercise going through the protocol, and learned a lot about what would work in the field, what needs to be modified, and ponder what it would be like to be an abalone parent, with millions of children, somewhere out there… but I’m weird.

 

After spawning my red abalone, a good practice run, I felt oddly optimistic about future experiments.  I should say, cautiously optimistic, because when you say the word “experiment,” I get an ominous chill and view of everything that could potentially go wrong.  I am starting to get more and more excited about the results that experiments with live gametes will yield.  But that, my faithful readers, is a story for another day.

 

And next, I will tell you all about Run 7, and my latest stay on the island…I can’t wait to go back!