ASITA 2017 post-conference thoughts

The triple helical structure of collagen, the stretchy material that makes up a third of our bone. Complicated, right?

Just got back from the University of Waterloo where the 2017 Advances in Stable Isotope Techniques and Applications (ASITA) workshop was held, and it was eye opening. I learned a few tips, some tricks, and received a whole lot of recommendations that are going to be very useful in our lab over the next few months. This conference was much smaller, had a different crowd and focus than other conferences I've been to. While previous conferences I've attended have really been about scientific theory this conference was really about the nuts and bolts about the methods, which are ultimately essential for advancing theory like you might see at some of the larger gatherings. Of the hundred plus people in attendance, most were lab managers, technicians, people running commercial labs, professors setting up their own labs and seeking expertise in their own institutions, and company sponsors, the manufacturers of instruments used to conduct stable isotope analyses (at times the conference was more like a trade show, with companies advertising the latest and greatest technology).

ASITA, as I learned in the opening presentation, was only formed in the early 90s with the advent of a now ubiquitous setup in many laboratories around the world: the continuous flow isotope ratio mass spectrometer with an elemental analyzer attached. While both those instruments existed in mature forms before they were hooked up to each other, it took around fifty years for people to use them in direct combination. By doing this one could increase the amount of samples to analyze in a single day from maybe ten to over one hundred; a truly revolutionary accomplishment that appeared in just a few years. As people got wind of such a development, ASITA was created to inform the community of stable isotope researchers on how to implement the technology in their own labs. Since then conference has continued to serve the purpose of informing the community of new technological leaps and developments, including new methodologies, incremental progress, and new instruments (such as laser based mass spectroscopy).

Though I was not the only graduate student in attendance we were certainly in the minority. However, it was certainly beneficial to see a different more technical side of the field. Below I'll just summarize some of the things I took notes on or otherwise found interesting:

• Use quarter inch tubing instead of eighth inch on pure oxygen cylinders, lest you experience an explosion due to adiabatic pressure effect leading to flash combustion of micro-fragments in the tube. 
• Though not necessarily useful for my own benefit, many people there were in attendance for a workshop on LIMS, a laboratory information management system. This seems almost necessary for the large commercial labs and government agencies which are handling thousands of samples and clients everyday.
• There is a much larger market for stable isotope analysis outside of academia than I realized. Several of the presentations discussed developments in techniques for identifying performance enhancing drugs in athletes, others in adulteration of foods and in environmental remediation efforts, not to mention forensics. 
• We now have the ability to analyze many of the lighter isotopes using the same instruments in a short amount of time; its possible to analyze a sample nitrogen, carbon, sulfur, oxygen and hydrogen in ten minutes. 
• Ramping the temperature in a GC column allows you to essentially erase the "memory" effects of carbon and sulfur, with sulfur being a particular nasty element that often sticks around long enough to influence the next sample that immediately follows.
• Hydrogen samples can be especially tricky. On many molecules (especially of interest to me, collagen from bones) there exists exchangeable and non-exchangeable hydrogen. We're interested in the non-exchangeable H as that was set in place during an animal's lifetime. After the molecule is exposed after death, the exchangeable H will freely equilibrate with whatever is around it. Therefore, every lab must treat the sample with a carefully controlled atmosphere of known composition so it can be corrected to an international standard. 
• Collagen must also be dealt with fast before analyzing for hydrogen, as the molecule happily absorbs water. Make sure to completely desiccate it before analysis.
• If you're interested in only carbon analysis, check out cavity ring down spectroscopy.
• CSIA (compound specific isotope analysis) via GC-C-IRMS is particularly sensitive to what standard you use (all samples you intend to run must be compared to a substance of known composition so the numbers you receive are comparable between labs) and should be as close to your sample in composition as possible. So if measuring hair, use a hair standard and not some random organic thing, and especially not your reference gas. 
• Splitless injectors for GCMS work seems to be more problematic than the more tradtional split injector. 

If many of these things seem confusing to you, do not worry! I plan to do a series on mass spectroscopy with pictures from our lab this summer. Stay tuned!

Taphonomy and organic matter in bone

The bone bed found on a beach at Bahia Thetis

All that history in my last post leads to the reason for this travel in the first place: the collection of historic samples. Up until this current trip, I've looked at samples of fur seals and sea lions form archaeological sites. The majority of these are from the earliest known site with animal remains, Tunel, named after the Estancia Tunel not far from Ushuaia. I also have some samples that were collected in other sites along the Beagle Channel, Peninsula Mitre, and Isla de los Estados that extend my data set both geographically and temporally to the end of the marine hunting and gathering lifestyle employed by the indigenous people of Tierra del Fuego. With this new collection, I'll be able to see if the massive changes that happened culturally with the arrival of industrial marine exploitation also had an impact ecologically. Significant changes have been observed in ecosystems around the world, including in California with the near elimination of sea otters. The extent of ecological impacts of industrial exploitation in comparison to subsistence exploitation of marine animals in Tierra del Fuego remains a mystery...until I run these samples in a few weeks!

Sea lion and whale bone in an archaeological site

But the understanding of the process of what happens to a body after death (the field of taphonomy) is a pretty important for interpreting stable isotopic results from my bone samples. Much of this field was pioneered by Behrensmeyer in multidecadal studies in Kenya with observations of the breakdown of bodies, separation into different parts, and ultimately into individual bones that break apart themselves. There are many different things that influence this process, including biological: scavengers, which can rip apart the body, limbs and chew on bones; microbes, which can feast on soft tissues, fat and oils of the body and the bones, and produce byproducts such as acids that chew away at bone mineral. Physical processes can include transport, for example, a bone traveling down a stream, bumping and grinding against rocks; damage from sunlight, which bleaches the surface and wears down the mineral, leading to cracking and flaking in the bone surface; and chemical weathering, the dissolution of bone mineral from acids.

One of the reasons I've been able to analyze samples from here is because the climate and conditions here in Tierra del Fuego generally buffer or slow down these taphonomic processes. Bones from the even the earliest sites have nearly the amount of organic matter in them that you would find in modern samples. This is because they've been buried almost immediately after being processed by humans. As a sub-antarctic climate, its generally very cold here, like throwing your steak into a refrigerator, it'll last longer than sitting out in the open on the Kenyan savannah, In addition, they've been buried in piles of shells. These shells are largely calcium carbonate, which counteracts acids that could be introduced by rain, trees, and soil bacteria.

Bones from partially articulated sea lions 

In contrast, some of the bones I have were buried in sand, allowing more of these chemical processes to reduce the organic matter in my bones, noticeably reducing the amount of organic matter. The samples we recovered from the seal greasery at Bahia Thetis are in an even worse state. After the carcasses were thrown into the bay, biological agents surely ate away at the soft tissue, leaving behind bones. Tidal action moved the bones/carcasses along the floor of the bay, before they were washed up onto the beaches during extreme storm events. Despite these destructive forces, a large number of identifiable bones can be found up on the beaches. However, bones on the surface tend not to last very long at all, and almost all the bones we found were in an advanced state of deterioration despite being only 70 years old. It may end up being difficult to extract the organic matter from these, but we have a large volume to extract from, so even if a small fraction is there, we'll get it.

We did find a fully articulated skeleton buried in the beach, with just a piece of its skull sticking out on the surface. The buried bones were in a much better state that the buried elements. I imagine this carcass was buried before it could be disarticulated by external forces. It stands to reason that there are probably many more articulated skeletons and bones either buried in the beaches or still in the bay where they were originally deposited. Likewise, there are surely many seals and sea lions buried in shell middens which have yet to be excavated, of which there are thousands known in Tierra del Fuego.

Return from Bahia Thetis

Anchor from a shipwreck in Peninsula Mitre

After twelve days and much work, I've had a few days to rest and reflect on our mission to collect and sample sea lion, fur seals, kelp, water, birds, fish and shellfish from Bahia Thetis, one of the most remote parts of the island of Tierra del Fuego. And it was a absolute success: In total, we collected 101 left sea lion and fur seal jaws (the majority being adult female sea lions), around ~50 femurs, and 10 skin and fur samples from the 1940s factory site. In addition, we made three 50x50cm excavations of shell middens, containing numerous more bones, stone and glass artifacts and tools including the first harpoon point ever found in the peninsula.

I learned a lot about this part of the world, and Peninsula Mitre in particular, which I think is important for understanding the context of the work I'm doing here. Peninsula Mitre is different in a number of ways from the land surrounding the Beagle Channel. The Beagle Channel is characterized by steep mountains and sharp glacial features, drumlins, canyons and overall rocky with forest. The peninsula, and much of the Atlantic coast of Tierra del Fuego, has softer rolling hills, less forest, dominated by peat bogs, and shallow coastline with  a slowly descending continental slope that extends for hundreds of kilometers. This shallow coastline proved treacherous for navigating, resulting in many shipwrecks, many of which were scavenged to build ranch buildings.

A more recent shipwreck from the 1980s

Geographic differences also played out ethnographically between indigenous groups. The Yamana occupied the Beagle Channel and southern archipelago, the Selk'nam in the interior and northern parts of Tierra del Fuego, the Alakuf on the western archipelago in Chile and the Haush in Peninsula Mitre. It is important to note that boundaries were not absolute for these groups, language and culture mixed and created gradients. The Haush and Selk'nam commonly incorporated more terrestrial animals into their diets, especially Guanaco, but they also hunted sea lions and seals like the more marine oriented Yamana.

Bahia Policarpo, site of the Policarpo family sheep ranching operation

When European settlers arrived, such as the Anglican missionary Thomas Bridges, they first set up in the Beagle Channel. However, it didn't take long for others to settle in other parts of Tierra del Fuego. Estancia Policarpo was established in the late 1800s by a Chilean family southeastern Atlantic coast of Tierra del Fuego, extending from the easternmost part of the Island at Cabo San Diego to its northern border with Estancia Maria Luisa some 50 km away. Like the ranch at Harberton, Estancia Policarpo exploited sheep for their wool. Estancia Policarpo operated until the mid 20th century, succumbing to falling wool prices from technological innovations in synthetic fabrics and wool that was cheaper to produce elsewhere. A caretaker maintained the what was left of the ranch until the 1980s, but most of the ranch buildings fell into disrepair throughout the late 20th century. Today the estancia is operated as a cattle ranch.

Blubber boiling vats

One way to subsidize the cost of the operation of the failing ranch was the exploitation of pinnipeds. In the 1940s, over one hundred Chileans, experienced in seal in sea lion hunting more common on the western coast of South America, set up a seal oil production facility in Bahia Thetis. The company would raid the multiple sea lion colonies that existed along the shores of Estancia Policarpo, killing over 30,000 seals and sea lions over a decade. The pinnipeds would be killed at the site of their colonies, loaded onto boats and taken to Bahia Thetis, were they were skinned, stripped of blubber and thrown into the bay. The blubber was then boiled in large vats, producing oil, much in the same way as whale blubber. In 1952, sealing was banned and the operation abandoned, leaving behind ruins of both the seal greasery and the ranch.

Bahia Thetis, site of the 1940s seal greasery facilities

That's I've got for today, stay tuned for more!

Estancia Harberton

Estancia Harberton, home of missionary Thomas Bridges and family in 1886. Imported brick by brick from England to Tierra del Fuego. Located east of Ushuaia by 50 km or so.

Yesterday I joined the archaeologist crew to collect some of my own samples and help carry bags of rocks. They were very heavy and my arms are quite tired still. But it was quite a productive day trip! We managed to carry 145 rocks over several kilometers, which will eventually help my fellow graduate student Carmen determine the source of lithic tools that were used by the Yamana. For my own sampling, I took some samples of kelp and some water samples, one of each from a bay exposed to a river mouth and one from the bay facing the Estancia Harberton (pictured above) which does not have a direct freshwater input. I doubt there will be significant differences between the two areas but good to measure extra just in case. The waters of the Beagle Channel are known to be exceptionally fresh compared to ocean waters from other parts of the world, due to the high amount of precipitation and freshwater inputs (from snowmelt and glaciers), but I will find this out and verify this claim personally!

Loading a large bag of rocks to be carried back to the truck.

The purpose of sampling water here is two fold. First I'm interested in differences in the waters of the Beagle Channel and the southern Atlantic/Argentine Sea, specifically to see what differences in water chemistry there might be, especially when it comes to nutrients and trace elements. Different nutrient loads can have an impact on the productivity of food webs, which would in turn have effects on species high in the food chain, like top predators such as fur seals, sea lions, and cetaceans (whales and dolphins). Trace elemental analysis can help me not only differentiate bodies of water, but serve as an independent method that would potentially verify where my fur seals and sea lions spend their time. Trace elements are incorporated into your tissues just like stable isotopes are and can (hopefully) be used a tracer.

A figure showing the interaction of different bodies of water in South America (from  Acha, E. M., et al. Marine fronts at the continental shelves of austral South America: Physical and ecological processes. J. Mar. Syst. 44, 83–105 (2004).) 

The measurement of kelp will serve a similar purpose. Giant kelp, Macrocystis pyrifera, grows around the world, from the coasts of Alaska, New Zealand, Japan, all the way down to Tierra del Fuego, It's a very successful species, one of the fasting growing on the planet (a meter a day in the right conditions) and forms the basis of many highly productive food webs around the world. Constraining the variation between southern Atlantic kelp and Beagle Channel kelp will give me an understanding of what to expect when I compare the stable isotopic values between giant kelp, kelp based fish and ultimately fur seals and sea lions. Fur seals and sea lions might be moving back and forth from open ocean to kelp ecosystems or changing their degree of residency in kelp ecosystems over time so it's important to understand what the kelp ecosystem looks like versus the open ocean.

Soon I leave for the Atlantic Coast, Peninsula Mitre and Bahia Thetis, site of the abandoned sealing operation that ran from 1946 to 1952. Bahia Thetis was formerly known as the Bay of Good Success, I hope it lives up to its name!

Wild horses seen while sampling near Estancia Harberton

Ushuaia 2017

The recreational airport club overlooking Ushuaia Bay and downtown Ushuaia

 After a bit of a hiatus I return to describe what may be my last trip to Tierra del Fuego. Since I've last updated this blog, quite a bit has happened:

• I've measured more samples (including fish!)
• I've passed my qualifying exam (hooray!)
• I've moved universities from Environmental Systems at UC Merced to Earth Sciences at UC Riverside (possibly invalidating my qualifying exam)
• and, finally, I've returned to Ushuaia for (maybe) the last time

But what opportunity awaits while I'm here! I've got some big plans. Next week I'll be traveling to Harberton Ranch to collect water and kelp samples, which eventually should help me do some forensics on where my fur seals, sea lions, fish and other members of the marine food web spent their time (open ocean or kelp forest? Beagle Channel, Atlantic, or Southern Ocean?) At the same time I'll be helping a fellow grad student who has been collecting rocks in a radial fashion around an archaeological site to identify their availability for making tools. 

In about a week and a half, I'll be leaving for Peninsula Mitre, the easternmost point of the Island of Tierra del Fuego. There are no roads to this place, and my colleagues normally reach this place by helicopter. However, we'll be taking the scenic route by horse which will take 3-4 days each way. Very few scientists use horses in their fieldwork these days, so it'll be harkening back to the days of old adventuring. Some French tourists may be accompanying us for the ride, so it'll surely be an interesting experience. I'll be journaling the experience and sharing with you all when I return!

The primary objective of traveling to this place is to collect fur seal and sea lion remains from industrial seal hunting that occurred during the 20th century. So far I've previously looked at fur seal and sea lion remains produced at indigenous archaeological sites, dating from roughly 6000 years ago to the 1700s. I expect to see significant changes in the diets of these animals as human influences changed from subsistence hunting to wholesale slaughter, but I'll find out when we return with the samples to the lab. I'll also be collecting more water and kelp samples, while my archaeologist colleagues will be digging some test pits to compare cultural differences between ancient sites on the peninsula versus sites to the west in the Beagle Channel. 

An inset historic map from James Cook's expedition, one of the first European explorers to document the area. Note that north is pointing to the right on this map and, that the Beagle Channel has not yet been identified. We'll be riding to the far western edge to the Strait of Marie (Detroit de le Maire). I've analyzed some samples from the area labeled "Baye de Valentin" and "Terre des Etats" already (Valentine's Bay, or Bahia Valentin and Staaten Island, or Isla de los Estados today).

Finally, I've spent a bit of time thinking about my PhD journey over the past several weeks and thought I'd share a bit about the process. I came in starting this degree with an idea for my PhD thesis fairly well formed within the first few months of starting at Merced. Those original ideas have shifted as logistics and preliminary data came into view. The questions I originally set out to ask changed and become more focused since the broader questions turned out to be more difficult to answer than expected. This realization required me to think about different types of methods to be used, the learning of new tools to help answer those questions and the number, and all the while scale and scope of the project increased. But it's this process that I think is probably the core of what a PhD is about: to come up with good questions and find a way to answer them, given limited time and resources. Of course, this sort of thing takes some planning so I made a timeline to illustrate it. Just in case you might considering a PhD, maybe you will find this helpful!