Student research team studies microbes from unusual cave system

A team of six Henderson State University biology students are on a mission to examine and analyze mysterious microbes found deep underground in a unique Tennessee cave. For the past two years, they have been using metagenomic DNA sequencing to identify microbes from a “petroleum pond” found in the cave.

In 2019, Michael Ray Taylor, chair of Henderson’s Department of Communication and Theatre Arts, explored the “highly unusual” cave system, known as “Secret Squirrel Cave” to protect its location.

Taylor has explored caves in the southern United States for 35 years and written about his adventures for a variety of national publications.

Taylor had heard about an unusual “petroleum pond” in the cave, and he discovered some unusual features during his exploration. He collected nine samples and brought them back to Henderson for analysis.

Dr. James Engman, a professor of biology at Henderson, had led a study at Blanchard Springs Caverns in Arkansas. He adapted the research techniques from that study to begin a new research effort focusing on the samples collected by Taylor.

Students Kaylie Wheeless, Lauren Camp, Aspen Huseman, and Maya Robles are the original members of the team. With Camp and Wheeless graduating this year, Rocio Alferez and Mitti Fairchild have joined the research group.

When explorers initially entered Secret Squirrel Cave more than 10 years ago, Engman said they considered it a “typical” cavern. But recent exploration and mapping has revealed previously overlooked passages that have now been mapped to more than 15 miles with many unusual features and formations.

Henderson’s research team has focused on the samples from the section known as the “Petroleum Passage” for its strong odor of petroleum.

Engman said unusual dark patches surrounded by colored rings of sediment were discovered on the sandy bottom of a large pond in the passage. He said the patches, called “mini-vents,” occasionally release bubbles of an oily material that rise to the surface and burst, releasing the petroleum odor.

At times, this passage has dense populations of larval cave salamanders and cave-adapted millipedes that are not expected so deep within a cave where the products of photosynthesis are “scarce to absent.”

The cave appears to have been at least partly formed by the action of sulfuric acid, rising from below and dissolving the limestone, rather than the typical means of cave formation involving mildly acidic rainwater from the surface, Engman said.

“This process, which leaves behind impressive gypsum formations, is known from some of the most famous caves in the western U.S., but is not well-documented in the eastern U.S.,” he said. “Such caves have been proposed as possible models for life in subsurface environments on Mars.

“The science in this project is really fascinating, and we are expanding our understanding of extreme environments,” Engman said. “For me, though, the most exciting part is getting undergraduate students involved in discovery and fieldwork that they get so excited about.”

To date, the students have identified 593 different microbes. While some are common, and would be expected in most caves, others are unique and have the ability to metabolize methane, sulfur compounds, ammonia, and hydrocarbons, Engman said.

The student team now seeks to conduct its own exploration of Secret Squirrel Cave to continue and enhance its research. To provide the group with caving experience before actually exploring and conducting research in Secret Squirrel Cave, Taylor took them on a three-day excursion in August to a large cave within Fall Creek Falls State Park in Tennessee to learn caving techniques.

The team hopes to explore Secret Squirrel Cave and collect more samples later this year.

Maya Robles, a sophomore from Katy, Texas, said she had “no idea” that she would become involved in such an exciting project.

“Our recent caving expedition jump-started my desire to seek adventure,” Robles said. “As we were walking through the tight passages, enormous rooms, and sketchy climbs, I never wanted to turn around.

“After my experience in the cave and time spent with my team and the other cavers, I hope to continue having adventures like these. I am so excited for these years to come.”

Kaylie Wheeless, a senior from Cedar Hill, Texas, said, “It felt like time had paused when we entered the cave.

“As biology students, we couldn’t help but wonder what microorganisms we would find if we were to take a swab from each fragile cave formation we passed,” Wheeless said. “I thought it was interesting that as we climbed, hours would pass and I’d have no idea how it could possibly have been that long.”

Aspen Huseman, a junior from Cabot, said she was “at a loss for words” in trying to describe her caving experience.

“There is no true way to depict the experience of caving until you get to actually do it,” Huseman said. “Coming to Henderson, I never expected to get opportunities like this. It was one of the scariest, most adventurous, thrilling, and most amazing things I have done.”

Mattison Fairchild, a junior from Conway, described the adventure as “surreal.”

“I have been in a cave before, but this was my first experience in true caving,” Fairchild said. “I enjoyed the stunning sights inside, but I also enjoyed the physical challenges that the cave presented. This was one of the most amazing experiences.”

Lauren Camp, a senior from Maumelle, said she was “extremely nervous” at first.

“Once I got inside the cave, I was completely taken away,” Camp said. “It was beautiful, and nothing like I had ever seen before. It was truly amazing to see what most don’t get to see.”

Rocio Alferez is an international student from El Salvador. She was impressed by the cave formations, the size of the room, and the cave crickets.

“You see huge mountains of breakdown, followed by pitch darkness, and wonder, ‘can I really climb this?’” Alferez said. “The curiosity and intrigue of seeing what’s on the other side makes you keep going further.”

Huseman added, “I’m hoping the team and I will soon have the opportunity to use our new caving skills to explore the Secret Squirrel Cave and collect more samples and data from our research site.”

The research team’s work has been presented at several professional meetings, including the Council on Undergraduate Research “Posters on the Hill” Symposium, and the Arkansas INBRE conference.

Wheeles presented at the annual convention of the National Speleological Society in July and received the award for Best Student Paper.

Engman described the project as the “most exciting” he’s been involved with during his tenure at Henderson. It has a growing list of collaborators, including: an analytical chemist from the stable isotope lab at the University of Arkansas; a salamander expert from the University of Alabama; and a karst geologist from Tennessee.

“Mike’s (Taylor) role in this project cannot be underestimated,” Engman said. “A caver for 42 years, he is closely connected with not only the caving community, but with the groundbreaking microbiologists who have studied the most amazing cave systems we have.

“His many books on caving have received much critical acclaim and awards. He may be the most prominent caving journalist there is.”

Engman emphasized the opportunities undergraduate research provides for students.

“I believe that undergraduate research is one of the most important things to get students involved in,” Engman said. “It increases their competitiveness for admission to graduate and professional programs, hones critical thinking skills, and can give them experiences they never dreamed of.”

The project is funded from several sources.

Robles and Wheeless are McNair Scholars and receive financial support for their work from the TRIO program. Wheeless received a State Undergraduate Research Fund grant last spring.

Camp received a NASA/Arkansas Space Grant Consortium, and Huseman was awarded a Scholars Fellowship grant from the National Cave and Karst Research Institute. Robles is waiting to hear the decision on her grant submitted to the Explorer’s Club.

The team members described their recent caving experience in detail:

Maya Robles

When I came to Henderson, I had no idea I’d get involved in such an exciting and eventful research project. This caving trip was not my first time in a cave. I’ve been lucky enough to see show caves and have been on a wild cave tour before our trip to Tennessee. As a prelude to this cave adventure and being on the research team, I visited Mammoth Cave National Park in Kentucky this past summer to fully immerse myself in speleology. I found myself fascinated by the geology, biology, and history of it all. The most recent caving trip opened my eyes to a new world: the exciting subterranean adventures of spelunking.

The caving expedition jump-started my desire to seek adventure. As we were walking through the tight passages, enormous rooms, and sketchy climbs, I never wanted to turn around. For me, the two things keeping me going were the thrill of the physical challenge of technical climbs, as well as my curiosity and wanting to see as much of the cave as I could. Everything leading up to that point had my anticipation building and seeing it through was incredibly rewarding- at the end of the path was this breathtaking underground river. Something Kaylie and I found hilarious during this otherwise serene moment was finding abandoned inflatable boats by the water. I guess past cavers had left them in case of a future impromptu cave river pool party!

Sitting by the slope of the river and looking off into the distance, I felt like a kid again, small but eager to discover the world around me. The team that guided us through the cave was amazing. They all were energetic and willing to lend a helping hand, arm, or leg, to help all of us get through. The going was tough but never felt rushed or dangerous, and their cheerful banter and encouraging words were something I was happy to be a part of.

After my experience in the cave and time spent with my team and the other cavers, I hope to continue having adventures like these. I am so excited for these years to come, I’m so glad I get to spend it working with my incredible team, and I hope to contribute as much as possible to the research on Secret Squirrel Cave.

Kaylie Wheeless

This was not my first time in a cave, but my previous experience could not have prepared me for the cave we explored last weekend. My first cave was more geared toward tourists, and the most recent one was much more intense. Our trip into the cave began with a mile hike. It was a hot, humid summer day, but as we approached the cave entrance it became noticeably chilly. The cavers we were with called it “cave conditioning”. The cool air from the cave was drifting out into the surrounding area and cooling the air around us before we even got to the entrance.

The entrance was a gaping hole in a massive wall of limestone surrounded by brilliantly green ferns. I remember looking back at the ferns as we walked into the cave entrance and how much I appreciated them when we came back out. They were the last green, living item we would see for the next seven hours and eleven minutes. The first five minutes were easy, but it didn’t last long. We were quickly climbing up massive piles of broken rock and squeezing through tiny passages. It was exciting to come out of those tiny cracks and see massive rooms. Each room had a new pile of clay-coated rocks to climb up or slide down to get to the next passage. The size of the rooms and the mountains of broken rock within them were overwhelming at times, but every so often we would come upon a fragile cave formation that reminded us how careful we had to be to not only avoid hurting ourselves, but also to avoid damaging beautiful formations that could take thousands of years to reform. I remember how excited we got each time we saw them.

As biology students, we couldn’t help but wonder what microorganisms we would find if we were to take a swab from each formation we passed. I thought it was interesting that as we climbed, hours would pass and I’d have no idea how it could possibly have been that long. It felt like time had paused when we entered the cave. Exploring the cave was exciting, exhilarating, and terrifying all at the same time. I have spent two years working on our cave research project and enjoyed it so much that I was considering continuing to study microbiology in caves in graduate school. My experience on this trip helped me confirm that it is a valid path to continue on should I get the opportunity. I hope that I will have the opportunity to explore many more caves in the future.

Aspen Huseman

I found myself at a loss for words, simply because there is no true way to depict the experience of caving, until you get to actually do it. Before going last weekend, I thought I had a true idea of what it would be like in the cave, but it was not at all what I expected. The experience was more spectacular than my imagination and preparation had led me to believe. When I walked into the cave, I instantly felt a drop of temperature and a sense of astonishment as I looked around at the mud-covered ground and articulated walls that lined the cave. I followed behind experienced TAG cavers who led the excursion with passion and excitement. This was when I knew the experience would be awesome, and unique; I was surrounded by people who absolutely loved caving.

As we ventured further into the cave, we reached the first big challenge — climbing up a steep incline of slippery rubble and rock (known by cavers as “breakdown”). Honestly, I was really afraid of falling, of heights, of the unknown; but as I slowly crawled and climbed up the breakdown, everyone there was reassuring and encouraging, helping me figure out where good footholds were to pull myself up through these massive tunnels of rock. It was there that I conquered my first big challenge in the cave and where I learned firsthand one of the most important rules of caving — trusting yourself and teammates. I was so proud of myself when we made it up the breakdown and was truly amazed at the complexity of the cave system. This feeling continued throughout the entire 71/2-hour cave trip.

This complex system sort of reminded me of a jigsaw puzzle. In the cave, there were three magnificent rooms. When we arrived at each one, the team would turn off their lights and wait for one of the TAG cavers to turn on their bright light. This light would brighten the whole space. At every big room, we did this and saw the intricate designs in a different light. I think the most amazing part of the cave to me was looking at the beautiful formations within it that took thousands of years to form. These are just a few of the many examples of amazing things I got to see and conquer while on my first caving adventure. I will definitely be doing this again. Hopefully, the team and I will get to use our new caving skills to go to our research site and collect more samples and data on the “Secret Squirrel Cave.”

Coming to Henderson, I never expected to get opportunities like this, and I am forever thankful for this team and the wonderful Biology Department at Henderson for giving me a challenging, unique, and practical education. My weekend trip was one of the scariest, most adventurous, thrilling, and most amazing things I’ve done, and I will always cherish the experience.

Lauren Camp

My experience caving for the first time was incredible. I was extremely nervous before we began. However, once I got inside the cave, I was completely taken away. It was beautiful and nothing like I had ever seen before. This was also my first time getting to socialize with “cavers.” I had always heard that cavers are such good people, and that statement couldn’t be more true. I was encouraged the whole way. Since my trip, I have recommended caving to everyone I have spoken to. It was truly amazing to see what most don’t get to see.

Mattison Fairchild

It felt surreal to be in a cave that large. I have been in a cave before, but this was my first experience in “true caving”. I am definitely looking forward to going caving again because I really enjoyed it. I enjoyed the stunning sights inside, but I also enjoyed the physical challenges that the cave posed in getting there. The most interesting part of exploring the cave was seeing the three big “rooms” that we encountered. The rooms are huge, dome-like spaces where the ceiling of the cave has fallen over the years. They were breath-taking, especially when they were lit up with the large spotlights that the other cavers had brought.

Along with the large rooms, we also had to squeeze through some pretty tight spaces, some being so tight that we had to take our backpacks off to fit through. Something that I thought was unusual was that I could see my breath in the cave even though it was 60 degrees inside. This was due to the extreme humidity in the cave. We also saw intriguing rock formations in the cave that were formed due to water dripping on them for a long period of time. Overall, this was one of the most amazing experiences, and I hope to do it again someday.

Rocio Alferez

This was my first time inside a cave, I cannot really find words to describe the exact feeling of it, but it was amazing, exciting, scary and challenging. You see huge mountains of breakdown followed by pitch black darkness and wonder, “Can I really climb this?” And the curiosity and intrigue of seeing what’s on the other side makes you keep going further.

The most interesting aspect of exploring the cave for me was seeing the cave formations, the size of the rooms of the cave, and cave crickets that were in there. It’s so incredible knowing you’re standing in the middle of a room that was formed thousands of years ago, and that maybe I am the first Salvadoran person or one of the few to have ever been there. I am definitely looking forward to doing this again. It was a very rewarding experience, and I am extremely thankful for Dr. Engman, Mr. Michael Taylor, and everyone who helped make this trip possible for our research team.