By Diana Setterberg, MSU News Service
BOZEMAN – It's been known for nearly a century that swarms of single-celled organisms thrive by consuming chemicals from their environments and expelling methane gas as a byproduct. In 2024, researchers in the laboratory of Roland Hatzenpichler, associate professor in the Department of Chemistry and Biochemistry in Montana State University's College of Letters and Science , published the first-ever descriptions of methane-producing microbes outside the lineage Euryarchaeota, which – in a soon-to-be-published study – they have confirmed to be ubiquitous in the environment.
These methanogens are members of the lineage Thermoproteota within a part of the tree of life called Archaea. Until the journal Nature published the findings from Hatzenpichler's group last year, scientists believed all methanogens existed in the Euryarchaeota.
Hatzenpichler said scientists are eager to know how much methane thermoproteotal microbes produce because methanogens produce approximately 60% of the world's methane – a gas 28 times more potent than carbon dioxide in trapping heat in the atmosphere, according to the U.S. Environmental Protection Agency . After the thermoproteotal group was discovered, scientists began preparing genomic models to predict the organisms' environmental impact.
But new evidence from Hatzenpichler's lab, published Dec. 12 in the journal Science Advances , indicates that another newly identified methanogen in the Thermoproteota defies the predictions of those models. MSU researchers hypothesized that the new microbe, a member of the group Methanonezhaarchaeia, would convert carbon dioxide to methane. However, through experimentation on cultures of samples harvested from a Yellowstone National Park hot spring, Hatzenpichler's former graduate student Anthony Kohtz and current graduate student Sylvia Nupp discovered that the organism grows and produces methane not by feeding on carbon dioxide but on methylated compounds, which are omnipresent in the environment. Both Thermoproteota previously identified at MSU also use compounds such as methanol for growth and survival.
Hatzenpichler said the results of the research cast doubt on the validity of the genomic models and demonstrate the necessity of further experimental study to validate or disprove modeling predictions.
"Our genome predictions say a huge chunk of CO2 is converted to methane, based on a few experiments and the assumption that the results are representative of all of those genomes," Hatzenpichler said. "But we have historically ignored one group of organisms (Thermoproteota), so we're ignorant about what is making methane and about the substrates that are converted in the first place. And most researchers don't actually measure which compounds are converted to methane in the environment. A lot is simply based on assumptions."
To further bolster foundational knowledge, Hatzenpichler's group is continuing its research into thermoproteotal methanogens under a four-year, $1 million research grant awarded in September 2024 by the U.S. Department of Energy's Biological and Environmental Research program. In collaboration with researchers at the DOE's Joint Genome Institute and Environmental Molecular Sciences Laboratory , the goal is to determine how much methane is generated by a widely distributed group of methanogens within the Thermoproteota – the Methanosuratincola – and to better understand their physiology and biological processes.
Another paper from Hatzenpichler's lab, recently accepted for publication by the journal Current Opinion in Microbiology, reveals that methanogens belonging to the Thermoproteota live in any environment devoid of oxygen, including landfills, oil reservoirs, rice paddies, wastewater treatment plants and wetlands – not to mention in Yellowstone's hot springs, where the three lineages identified at MSU were found.
"Experimentally, it's very demanding, and we probably can't do it for all the environments we want to study," said Hatzenpichler, who is also director of MSU's Thermal Biology Institute . "On a global level, hot springs are completely irrelevant to methane emission, so now we need to go to ecosystems that are more important than hot springs to methane production."
He said graduate student Nicole Matos Vega, a recent recipient of a Graduate Research Fellowship awarded by the National Science Foundation, has collected and cultured microbes from mangrove swamps in Puerto Rico. She discovered, for still poorly understood reasons, that they are "making a lot of methane." Hatzenpichler said Matos Vega also will study the methanogens in their native swamps to ensure that they react the same way in the lab as they do in their natural habitat.
Another target ecosystem is wastewater, studies of which have already begun on samples provided by collaborators in Illinois. Graduate student Joelie Van Beek, a new recipient of a NSF EPSCoR Graduate Fellowship, found thermoproteotal organisms living in Bozeman's wastewater treatment plant, and those may be brought into culture, as well. Hatzenpichler said having the plant nearby will make it easier for his lab to test a number of variables in that ecosystem.
One problem with genomic modeling is it relies on measured levels of compounds found in various environments to predict whether methanogenesis is taking place, Hatzenpichler said. However, he noted that just because the level of a compound is low in a particular environment doesn't mean that it isn't important there. Instead, it may be low because the methanogens there are consuming it.
"Genome analysis is very, very powerful, though limited, and experimentation is very demanding," he said. "The idea with our research is that we will try to understand these ecosystems from all different kinds of angles."
