current research projects...


 

Beneath Yellowstone Lake at the hottest feature in YNP

 A map of our sampling sites on the lake floor off the coast of Stevenson Island in Yellowstone Lake. This is the hottest and deepest part of the lake. Map making credit goes to Pat Shanks of the USGS.

A map of our sampling sites on the lake floor off the coast of Stevenson Island in Yellowstone Lake. This is the hottest and deepest part of the lake. Map making credit goes to Pat Shanks of the USGS.

One of our current projects, "HD-YLAKE", focuses on Yellowstone Lake--but not the lake itself, rather the hydrothermal features raging quietly deep beneath the lake. We are using a variety of techniques to investigate the microbes living there, including a robotic vehicle called Yogi that can collect biomass from vents, and gravity cores that we drop from a boat to take samples of the mud near these hot spots. Back in the lab, we are measuring concentrations and isotopes of carbon dioxide and methane, and we are extracting DNA for analysis of microbial genes and genomes. 

With the help of Dave Lavalvo and his crew (Global Foundation for Ocean Exploration) we have collected microbial streamer samples from a few different hydrothermal vents. Streamers look like big long stringy pieces of snot that are attached to the lake floor right where a vent is spewing out of the mud. They're kind of like one of those crazy inflatable guys that whips around at a used car lots or mattress sales, but in this case super hot chemically energetic fluids are mixing with cold lake water and the microbes are swaying around in it. Here's a video of me picking some streamers apart with tweezers back in the lab.

We also have several sediment cores from sites all over Yellowstone Lake. Our methane data indicate that every single site has detectable methane at levels above atmospheric concentrations, from the surface sediments in contact with lake water to the deeper sediments down to almost 1 meter. My insanely talented undergraduate researcher, Kori Klingelsmith, has begun extracting DNA from these sediment cores and we are happy to say she has been able to get a lot! That translates to good news for sequencing the DNA, a technology which often requires high concentrations of high quality nucleic acids.

 

Methane, methane, methane.

 This sequence of photos captures much of our scientific process. We start with a question and then hike out to a hot spring to take a sample. Then we set up different types of enrichments in the lab, then we (Kori Klingelsmith) takes subsamples over time and measures them on an instrument, then we analyze the data and make a graph. The two duplicate samples in the graph are nicely agreeing with one another!

This sequence of photos captures much of our scientific process. We start with a question and then hike out to a hot spring to take a sample. Then we set up different types of enrichments in the lab, then we (Kori Klingelsmith) takes subsamples over time and measures them on an instrument, then we analyze the data and make a graph. The two duplicate samples in the graph are nicely agreeing with one another!

Throughout my career I always end up coming back to methane. Currently, the field of microbial methane cycling (methanogenesis and methanotrophy) is experiencing a paradigm shift in that we are learning that many more organisms may be involved in producing or utilizing methane than previously thought. Key words: MAY BE. We don't know this to be the case absolutely because we only have genetic evidence of it but no one has actually confirmed this in a laboratory by getting a new organism to do something with methane. But, we are trying very, very hard. We have set up all sorts of "fun" incubations with isotopically labeled substrates (that basically means food for microbes that is heavier than it should be), using methanol, methane, and carbon dioxide. In short, we are trying to catch these microbes with their hands in the cookie (methane) jar. 

An isotope, in this case, is an atom of carbon that has an extra neutron. By "extra" I mean that almost always there are only 6 neutrons in carbon, but we bought a special type of carbon that has 7 neutrons. This means that our special carbon weighs just baaaaarely more than the "normal" carbon, which has one less neutron. All living things need carbon, and when microbes get carbon from their environment, they use it to make biomolecules like proteins or DNA. But, not everyone can use every type of carbon. For example, humans cannot consume methane (which is one carbon atom and four hydrogen atoms, CH4). And many microbes out there also cannot use methane, but some can. So if we supply a community of microbes with "heavy carbon methane", and then some of the microbes use it, over time they will build biomolecules that are slightly heavier than what we would normally expect. Depending on whether this is the organism's sole source of carbon or whether it uses other carbon molecules, the percent incorporation of heavy carbon will change. Regardless, we can then detect these molecules with all sorts of special equipment (for example, using ultracentrifuges or secondary ion mass spectrometers). If we detect it, that means we pretty much proved that this particular organism is utilizing methane, and that's the goal!

 

The Hydrothermal Wonderland that is Guaymas Basin

 Temperatures measured in the mud from the seafloor down to 35cm depth, over the course of 8 days. Credit to Howard Mendlovitz (UNC) for building this temperature probe.

Temperatures measured in the mud from the seafloor down to 35cm depth, over the course of 8 days. Credit to Howard Mendlovitz (UNC) for building this temperature probe.

I spent most of graduate school thinking about a place called Guaymas Basin. 2000 meters beneath the ocean surface, there is a massive hydrothermal system where super hot fluids rise through thick, organic-rich mud and blast into the frigid deep water of the Gulf of California. Chemical and physical interactions between the hot fluids, the mud, and the deep ocean water make these sites oases of life. At the localized expressions of this amazingly complex thermal environment, microorganisms are thriving off of multiple sources of energy, from above and below.

From large, sulfur-oxidizing bacteria at the ocean floor to microscopic sulfate-reducing archaea and bacteria beneath the hot mud, there are tons of interesting metabolisms to investigate. In the past we have used genes to understand the "who's who" of the microbial world in this extreme environment, but we only scratched the surface with a taxonomic indicator gene that encodes small subunit ribosomal RNA (SSU rRNA). 

 Alvin dive 4563, taking a push core into a Beggiatoa mat, beneath which lives some juicy goodness.

Alvin dive 4563, taking a push core into a Beggiatoa mat, beneath which lives some juicy goodness.

I have worked on Guaymas mud many times because it was my graduate project, and recently I find myself at it again in a new collaboration to link viable microbial populations (as determined by RNA instead of DNA) to genomic content for some of the more elusive, novel, and unknown types of creatures. This time we focused on the hottest available sites from which we have core samples and we extracted RNA to look for microbes pushing the life's thermal limits--in other words, how hot is too hot? After reverse transcribing the 16S rRNA transcripts back into DNA, we sequenced them and found a bunch of sequences that had "unassigned" taxonomy. After building a phylogenetic tree to compare sequences we noticed that they were very distantly related to known lifeforms, meaning they may represent new major taxonomic groups. So now, we are comparing the 16S rRNA (from RNA) to a database of genomes in collaboration with Drs. Brett Baker and Nina Dombrowski to see if we can ground truth the single gene data with whole genome data. This would give us the entire genomes of unknown organisms that are living at the hottest temperatures in Guaymas Basin.