Living Life as a Sea Urchin Momma

Hailey Conrad, Rutgers University

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Me working hard to make my sea urchin babies

For my project I am using the same technique that the father of genetics, Gregor Mendel, used to establish his Laws of Heredity: cross breeding. So, I have to breed and raise a whole lot of sea urchins. For a refresher, I’m trying to determine if there is heritable genetic variation in how sea urchin (specifically an Arbacia punctulata population from Woods Hole, Massachusetts) larvae respond to ocean acidification. To do this, I’m rearing sea urchin larvae in low and high carbon dioxide conditions and measuring their skeletal growth. I’m breeding 3 sea urchin males with 3 sea urchin females at a time, for a total of 9 crosses. To tease apart the impact of genetic variation on just the larvae themselves, I will be fertilizing the sea urchin eggs in water aerated with either current atmospheric levels of carbon dioxide, about 410 parts per million, or 2.5 times current atmospheric carbon dioxide levels, about 1,023 parts per million. Then, I will be rearing the larvae in water aerated with either 409 ppm CO2 or 1,023 ppm CO2. This will give me four different treatments for each cross, giving me 36 samples in total. By fertilizing and rearing them in the same and different levels of carbon dioxide I will be able to see how much of an impact being fertilized in water with a higher carbon dioxide concentration has on larval growth versus just the larval growth itself. It’s important for me to make that distinction because I just want to identify genetic variation in larval skeletal growth, and separate out any extraneous “noise” clouding out the data. I’m rearing the larvae in a larval rearing apparatus. Each of the 36 samples will be placed in jar with water aerated with the correct CO2 treatment. Each jar will constantly have atmosphere with the correct CO2 concentration bubbled in. Each has a paddle in it that is hooked to a suspended frame that is swayed by a motor. This keeps the larvae suspended in the water column. The jars are chilled to 14 C by a water bath.

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My larval rearing apparatus

After a 6-day period the larvae are removed from the jars and their skeletal growth is measured. They are preserved with 23% methanol and seawater and frozen.

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An Arbacia punctulata pluteus

You’re probably curious how the heck I am able to measure the larva’s skeletons. They’re microscopic! Well, I use a microscope coupled to a rotary encoder with a digitizing pad and a camera lucida. Which, looks like this:

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A microscope coupled to a rotary encoder with a digitizing pad and a camera lucid hooked up to a computer

This complicated-sounding hodge-podge of different devices enables me to do something incredible. I can look through the microscope at the larva, and also see the digitizing pad next to the microscope, where I hold a stylus in my hand. When I tap the pad with the stylus and the coordinates of various points on the anatomy of the plutei that I am tapping at get instantly recorded on my computer! The rotary encoder is the piece attached to the left side of the microscope and it enables me to record coordinates in three dimensions. Then, I can use those coordinates to calculate the overall size of the skeleton. My favorite part of doing science is learning how scientists are able to do the seemingly impossible- like measuring something microscopic.

After I gather all of my data, I will do some statistical analysis to see the affect that the male parents have on the skeletal growth of their offspring. I will not be focusing on the impact that females have on the skeletal growth of their offspring. The quality of the egg itself could be an influencing factor on the size of the offspring, whereas sperm is purely genetic material. Like how I’m trying to isolate the influence of ocean acidification during larval rearing from during the act of fertilization, I am trying to isolate just genetic influences on larval skeletal growth from egg quality. Check back to see how it goes!

Special thanks to the National Science Foundation for funding this REU program, the College of Charleston and Grice Marine Laboratory for hosting me, and Dr. Bob Podolsky for mentoring me!

 

 

 

Catch of the Day(s)

Melanie Herrera, University of Maryland College Park

South Carolina is known for its iconic southern cuisine, including a staple of fresh seafood which fuels the buckets of shrimp & grits and “catch of the day”. In order to support this huge industry (and fill the bellies of every South Carolinian), I am conducting an experiment to figure out where this seafood is holing up prior to its demise. Dr. Harold, his graduate student, Mary Ann McBrayer, and I are out on Grice Beach collecting fishes, crabs, shrimps, and much more in order to figure what exactly is there… And what they are using to survive.

Using a seine net, we encircle marine animals in dense and sparse patches of an invasive sea grass, Gracilaria, for collection. We hypothesize that Gracilaria is helping the local economy (a surprising contribution from an invasive species) by creating refuge for young animals. On the beach, we submerge separate samples of animals (from dense versus sparse areas of Gracilaria) into a euthanizing solution to bring them up to the lab for preservation and analysis (Figure 1).

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Figure 1: An example of animals caught in separate habitats at Grice Cove. The left exhibits animals caught in a dense area of Gracilaria and the right exhibits animals caught from a sparse area of Gracialaria. Credit: Melanie Herrera

In the lab, separate samples (dense versus sparse) undergo a few transfers into different fixatives (10% seawater formalin, 25% isopropyl, and 50% isopropyl consecutively) to keep the fish from decaying. After this preserving process, fish and other animals are separated and categorized by family, genus, and species. This categorization enables us to identify and analyze what types of animals and how many of each are using different habitat. Our analysis will give us insight on what type of habitat, either patches dominated by Gracilaria or areas with more open water, benefits fish. Specifically, we will be able to identify if Gracilaria is more advantageous to young fish or if their survivorship is independent from their habitat.

So far, we have collected lots of pipefish, narrow skinny fish that resemble a hair strand-size snake, Atlantic Silversides, a fish that looks exactly like it sounds, and more shrimp than anyone needs (Figure 2). Although some of these animals do not directly contribute to the seafood industry, its presence in the Charleston Harbor can tell us a lot of things. For example, we have seen some fishes that usually stay in warmer waters in the Southern U.S. Their expanding habitat can lead us to some more hypotheses on climate change and warm weather moving northward. In addition, we can find out if Gracilaria has a stake in rearing economically important fish in the future.

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Figure 2: (From left to right) Pipefish, Atlantic Silversides, and Grass Shrimp caught for analysis.Credit: Melanie Herrera

Thank you so much to my mentor Dr. Tony Harold and his lab for his advice and guidance. Thank you to Mary Ann McBrayer for helping me facilitate this project. This research is funded through the National Science Foundation and College of Charleston’s Grice Marine Lab.

 

The Problem with PFCs- Seeking Answers in Plasma

Kady Palmer, Eckerd College

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I previously outlined the problem of perfluorinated chemicals (PFCs) in the environment and their unknown health effects.  In order to gain this knowledge, it is essential to determine what types of PFCs are frequently used and the mechanisms by which an individual would be exposed to them. Here, we are measuring the presence or absence of 15 PFCs that are commonly associated with non-stick cookware, firefighting foam, and water-resistant materials.

This compiled list of PFCs is the basis of my research procedure. From here, I must learn how these compounds interact with biological components in organisms in order to understand their subsequent health effects. With that being said, the type of samples I am analyzing is a topic worth explaining. PFCs are known to be “proteinophilic” or, attracted to proteins in the bloodstream of organisms like humans and, in the case of my study, manatees. Therefore, I am using manatee plasma to test for the total individual burden of PFCs. 

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Fig 1. 69 collection tubes containing manatee plasma samples (left). Aliquots of 22 samples of manatee plasma for future studies (right). Photos taken by me!

With 69 different plasma samples, I am performing a series of procedures that allow me to extract the PFCs. After completing multiple chemical processes (methodology proposed by Reiner et al., 2012), I am left with a liquid (containing the PFCs), measuring no more than 1 mL to be placed into a small vial. From here the vials are inserted into a liquid chromatography tandem mass spectrometer (LC-MS/MS), a machine that reads each of the 15 unique chemical structures of the outlined PFCs of interest and determines their abundance in each vial. This system isolates the concentration of each perfluorinated chemical for every one of the 69 manatee samples.

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Fig 2. The basic process a mass spectrometer performs in order to provide the concentration of chemicals being studied. Photo from: http://www.emdmillipore.com/US/en/water-purification/learning-centers/Anwendungen/organic-analysis/lc-ms/lWib.qB.vb4AAAFA5fIBvVBh,nav?ReferrerURL=https%3A%2F%2Fwww.google.com%2F&bd=1

The concentrations of these chemicals is the ultimate goal of my research study. This data will be compared to manatee location, morphometrics, body condition, sex, and more, in order to gain a better understanding of the overall PFC burden on these animals. These factors, or variables, may also provide insight into what may be influencing the burden intensity an individual may face. Once this knowledge is gathered, potential links to the health effects of PFC accumulation can be investigated in both manatees and humans.

I’d like to thank the National Science Foundation for funding this research opportunity and the College of Charleston’s Grice Marine Laboratory REU program for making this experience possible. A special thanks to the NIST team who has been teaching and supporting me throughout this process, specifically, Dr. Jessica Reiner, Jacqueline Bangma, and my mentor, Dr. John Bowden.

Note: These samples were collected as part of a health assessment of manatees by the USGS Sirenia Project. No manatees were harmed in the process of obtaining them.

References

Reiner, Jessica, Karen Phinney, and Jennifer Keller. “Determination of Perfluorinated Compounds in Human Plasma and Serum Standard Reference Materials Using Independent Analytical Methods.” Analytical & Bioanalytical Chemistry 401, no. 9 (January 15, 2012): 2899–2907. doi:10.1007/s00216-011-5380-x.z

Playing with Plutei

Hailey Conrad, Rutgers University

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Me! Photo Credit: Kady Palmer

Ocean acidification is known as climate change’s evil twin. When the pH of ocean water drops, carbonate ions in the water form carbonic acid instead of calcium carbonate. Calcium carbonate is the form of calcium that marine animals that have calcium-based skeletons (like us!) and shells use to build their bones and shells. Having smaller and weaker skeletons or shells impacts their ability to survive. However, some individuals within certain species or populations of species have genes that make them more resistant to ocean acidification. If these individuals are able to pass on these genes to their offspring, then the species has the ability to evolve in response to ocean acidification instead of going extinct. This summer I’m working with Dr. Bob Podolsky in College of Charleston’s Grice Marine Field Station to study the extent to which ocean acidification affects Atlantic purple sea urchins, Arbacia punctulata. We are specifically trying to see if any individuals within a population from Woods Hole, Massachusetts, have any heritable genetic resistance to the negative impacts of ocean acidification. We hypothesize that there will be genetic resistance given that the northern Atlantic coast naturally has lower levels of saturated calcium carbonate, so a population that has evolved to live in that type of environment should have some resistance to lower calcium carbonate levels already (Wang et al 2013). We’re using a basic cross breeding technique to rear Arbacia punctulata larvae to their plutei stage, when they have four main body rods. At this stage they look less like sea urchins than they do like Sputnik!

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A sea urchin pluteus larvae with four body rods

Then, we will look to see if any of the male parents consistently produce male offspring that are more resistant to ocean acidification.  If males like these exist within this population, then the species has the capacity to evolve in response to ocean acidification, instead of going extinct! This is a very big deal, and could potentially be very hopeful. Even if we don’t get the results that we are hoping for, the results of this research could inform policy and management decisions.

Literature Cited:

Wang, Z. A., Wanninkhof, R., Cai, W., Byrne, R. H., Hu, X., Peng, T., & Huang, W. (2013). The marine inorganic carbon system along the Gulf of Mexico and Atlantic coasts of the United States: Insights from a transregional coastal carbon study. Limnology and Oceanography, 58(1), 325-342. doi:10.4319/lo.2013.58.1.0325

Thank you to the National Science Foundation and College of Charleston’s Grice Marine Laboratory for funding my project. And, special thanks to Dr. Bob Podolsky for being a wonderful and supportive mentor!

 

Are Manatees the Key?

Kady Palmer, Eckerd College

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Contaminants. One word, countless different connotations. Therefore, the exposure to contaminants is a constant concern to both the public and the scientific community. The study I will be performing this summer focuses on perfluorinated chemicals, or PFCs. PFCs are a class of contaminants that are utilized in many commercially available products (ex: non-stick pans, stain resistant sprays, and water-resistant materials) and have been classified as highly abundant and persistent chemicals of concern, in relation to overall environmental and, subsequently, human health.

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Photo from: “Should You Ban Your Teflon Pan? California.” Savvy California, January 1, 2015. https://savvycalifornia.com/teflon-pan-toxic-or-not/. 

Through various mechanisms, PFCs have been noted to integrate into the environment and end up in the air, soil, and water. As this is happening, the organisms living in these areas become exposed and are put into a precarious situation. Little research has been performed on examining exactly what the effect these compounds have on organisms in these types of environments. Although it would be just as interesting to scoop water samples from different places to determine a basis for this environmental change, my project will be delving a bit deeper. Because previous studies have shown data supporting PFC accumulation in the bloodstream of different marine animals and their subsequent health consequences, I will be expanding this research by analyzing the types and abundance of PFCs in the Florida manatee.

The Florida manatee (Trichechus manatus latirostris) inhabits areas of warm water, close to the shoreline. Unfortunately, manatees have a history of endangerment, as a result of human impacts (boat strikes, entanglements, drowning due to drainages) and environmental changes. Perfluorinated chemicals, as described above, could very well be impacting manatees in ways currently unknown. This study aims to isolate the types and abundance of PFCs in Florida manatees and potential health concerns associated with this exposure. While the health of manatees is undoubtedly important, the results of this research could provide insight as to the overall health of the ecosystems examined. Manatees could function as a model for other organisms, demonstrating the possible repurcussions of PFC exposure. If that is the case, the knowledge gained from this organism, living so close to the shoreline of human inhabited areas, may be applicable in aiding future human research.

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Photo from: “West Indian Manatee.” Southeast Region of the U.S. Fish and Wildlife Service. Accessed June 23, 2017. https://www.fws.gov/southeast/wildlife/mammals/manatee/.

I’d like to sincerely thank everyone involved in the National Institute of Standards and Technology laboratories who have been a wealth of information and guidance, specifically Dr. Jessica Reiner, Jackie Bangma, and my mentor, Dr. John Bowden. This project would not be possible without samples and information provided by Robert Bonde with USGS, funding from the National Science Foundation, and the College of Charleston’s Grice Marine Laboratory.

References:

Bangma, Jacqueline T., John A. Bowden, Arnold M. Brunell, Ian Christie, Brendan Finnell, Matthew P. Guillette, Martin Jones, et al. “Perfluorinated Alkyl Acids in Plasma of American Alligators (Alligator Mississippiensis) from Florida and South Carolina.” Environmental Toxicology and Chemistry, no. 4 (2017): 917. doi:10.1002/etc.3600.

“CDC – NBP – Biomonitoring Summaries – PFCs.” Accessed June 19, 2017. https://www.cdc.gov/biomonitoring/pfcs_biomonitoringsummary.html.

West Indian Manatee”. Southeast Region of the U.S. Fish and Wildlife Service. Accessed June 23, 2017. https://www.fws.gov/southeast/wildlife/mammals/manatee/.

Stressing Out My Algae

Emily Spiegel, Bryn Mawr College

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One intern’s perspective on lab work, South Carolina, and the coolest organisms in and out of water: phytoplankton.

 

The lab itself is large, packed to bursting with equipment, boxes, cabinets, monitors, and glassware. An antechamber acts as a sterile room for the most delicate of procedures, demanding precision and care. Many things reside in this room, but never quiet. The constant whirling of a machine’s fan, the hum of a freezer housing samples from a time beyond easy recollection, the typing of a research assistant hunched over innumerable data sheets…all these and more cut through the quiet throughout all hours of the day and night.

 

And at the heart of it all is the algae.

 

Small, marine microorganisms constituting a larger class known as phytoplankton, algae are the unsung heros of the environmental world. Energy, or the basic ability to do work, is the key to survival, growth, and reproduction. Without it you (and your genes) aren’t going anywhere. Algae harness the energy readily available from sunlight and convert it into a useable currency in a process known as primary production. This energy is then distributed to the many higher animals that eat them. They are the foundation of the marine food web and of the world’s energy supply, contributing to 45% of the planet’s primary production (Brierley 2017). In short, algae are cool.

So cool in fact, I’ve decided to spend my entire summer studying them. More specifically, I’ll be studying patterns of their reproduction and growth. A grad student running an experiment in this lab last year got unexpected results when she raised algae in 24 hours of continuous light instead of the normal 12 hours of light:12 hours of darkness she had followed previously.  Despite a limitation in the nitrogen added to these samples, which typically inhibits growth, the populations grown in 24 hours of light were able to grow successfully. So researchers went looking for answers.

One potential explanation is that the continuous light conditions caused the induction of sexual reproduction in the algae samples. Algae, like the rest of us, don’t like to be stressed. And being constantly exposed to light, which they automatically begin to utilize for primary production, is very stressful. It’s kind of like giving a kid a bunch of candy bars. A little is nice, a lot induces a sugar high and headaches for anyone within a 20m radius. The algae have too much energy and so they start to adjust their behavior to accommodate for the stressful conditions. One accommodation is sex. That’s right, stress out your algae and they might just turn on the Marvin Gaye and set the mood. Normally the species I’m studying (Fragilariopsis cylindrus, or just Frag for anyone without a PhD) reproduces asexually allowing high growth rates within the population. My lab is also curious as to whether low light conditions (a cycle of 6 hours of light and 18 hours of darkness) might be equally stressful to the algae and cause a similar response.

This is where I come in. This summer I’ll be exposing algae to conditions of varying light and nutrient stress in order to determine if stress actually does cause them to start reproducing sexually. Along the way, we’ll keep track of growth rates by measuring biomass, or the amount of live material within a sample. This can be measured by a variety of cool devices which tell me the number of cells in a particular volume of sample and the amount of chlorophyll being utilized in that sample. Chlorophyll is a component of the cycle of photosynthesis and is therefore a measure of the primary producers (i.e. the algae) in the sample. Eventually I’ll also run genetic analyses, tracking the utilization of genes involved in sexual reproduction as a way to determine if the algae are reproducing sexually instead of asexually.

All in all, it’s bound to be an interesting summer. Full of days at the beach, early mornings with a culture counter, and lots and lots of algae.

 

I’d like to acknowledge the entire DiTullio/Lee lab at the National Oceanographic and Atmospheric Administration as well as the National Science Foundation’s Research Experiences for Undergraduates program organized by the College of Charleston Grice Marine Laboratory. This project would not be possible without the support and guidance from these institutions and individuals. 

 

Works Cited

Brierley, Andrew. “Plankton.” Current Biology Magazine 27 (2017): 478-83.

New Philadelphia to Charleston

Aaron Baumgardner, The University of Akron

Coming from the landlocked small town of New Philadelphia in the Midwest, I feel like I’m dreaming when I realize I’m spending my summer researching in Charleston, SC. I’m thankful for the opportunity that my mentor, the College of Charleston, and the National Science Foundation has given me to learn and grow in my scientific ability.

However, I do not believe I would be where I am today if it weren’t for my Aunt Jane. She is the only member of my family with a background in science, and even though she is hundreds of miles away at UPenn, she is always an email or phone call away. She has always shown an interest in my academics and will always be there for any advice I may ask. She has helped me develop my professionalism and offered insight on which graduate schools are worth going to.   Because of her, I can finally realize it’s not a dream. It’s reality that I’m spending my summer in Charleston, SC. It’s because I’ve worked hard in school and reached out for opportunities for me to mature as scientist. And I owe her so much for pushing me to succeed.

Thank you Aunt Jane!