Award: OCE-1948720

The primary goal of the research is to investigate biogeochemical processes controlling microbial aerobic methane oxidation rates (AMOR) in bottom waters surrounding gas seep sites in the deep ocean. In this project, we have been able to answer key questions about deep-sea methane dynamics by using novel in situ methods to investigate AMOR and the microbial communities that mediate it.

 

Intellectual merit: The delays in ship-time due to covid-19 mean that we did the first part of this project in a freshwater lake to demonstrate the principle that in situ incubations of aerobic methane oxidation can be linked to changes in the microbial community. Experiments were run in Jordan Lake from March 2020 through October 2021 to develop and test an in situ closed loop system capable of continuous monitoring of oxygen and methane concentrations while maintaining pressure and temperature. We demonstrated methane consumption at different temperatures as well as methane, oxygen, dissolved inorganic nitrogen (DIN), and organic matter concentrations. We found that the relative quantities of methanotrophs (which oxidize only methane) and methylotrophs (which demethylate organic matter) were consistent between replicates, but otherwise did not follow any patterns with respect to temperature, oxygen, or nitrogen concentrations. Methanotrophs had a robust  negative correlation with oxygen concentrations. Microaerophilic activity has often been observed in methanotrophs, but it is usually assumed to be an adaptation for getting closer to high methane, which is biologically produced in anoxic conditions. However, our results show this is not the case. The relative abundance of methanotrophs correlates tightly with oxygen, but not methane concentrations, suggesting that oxygen aversion is the primary motivator for microaerophily during aerobic methane oxidation. We gained robust evidence that methanotrophs are responsible for the observed AMOR because the rate constants of methanotrophy correlated positively with the relative abundance of methanotrophs. This was not true of methylotrophs or any other microbial group in these natural communities. This demonstrates the observed variation in AMOR between different experiments is driven by the density of methanotrophs, which are in turn determined by an aversion to oxygen. Our work implies that the amount of methanotrophs, rather than temperature or methane concentration is the primary determinant of the ability of the natural community to absorb shifting methane concentrations. It also suggests that oxygen aversion by methanotrophs may be the ultimate determinant of where they are found.

We completed three cruises to the T1 seep field site, ~350 samples were taken from the water column, sediment, and in situ incubations for microbiological analysis. 16SrRNA gene amplicon analysis has been performed on the water column and in situ incubation samples from the R/V Sharp and R/V Savannah cruises have revealed the presence of Methylomonadaceae (aerobic methanotrophs),Methylophilaceae (nonmethanotrophic methylotrophs), and Nitrosopumilaceae (ammonia-oxidizing archaea), among others. The methane-oxidizing community has low diversity, with the uncultured genus Milano-WF1B-03 being essentially the only aerobic methanotroph present. The relative abundances of Methylomonadaceae and Methylophilaceae exhibit a strong positive correlation across most depths which suggests possible syntrophy between aerobic methanotrophs and nonmethanotrophic methylotrophs in natural deep-sea waters. Further, depth appears to be an important driver of overall community diversity, with the beta diversity of water column samples aligning following a neat gradient across depth.

In summary, we have made the first in situ rate measurements for methane oxidation. They are robust and repeatable in both a freshwater lake and in the deep-sea water column over methane seeps off the coast of Delaware. We have now found strong correlations between geochemical parameters and the natural community that suggest what limits methanotrophy in the open ocean. Our work alsosuggests tight syntrophy between methanotrophs and methylotrophs may help alleviate oxygen toxicity in the open ocean. This work has supported five peer-reviewed manuscripts, with more currently being considered at journals and being prepared for future submissions.

Broader impacts: This work has been the primary support for one UT PhD student who has been trained in field and laboratory techniques, as well as computational analysis, and presentation (both written and oral) completely focused on this project. She has also submitted and revised a first-author paper from this work, and is preparing two more. This work has also supported an undergraduate student from rural Tennessee who published two first-authored peer-reviewed papers, won the NSF graduate research fellowship, and is now a PhD student at MIT. Furthermore, this work partially supported the work of another PhD student, who successfully defended his PhD and is now doing microbiological research in the US Army, as well as another undergraduate student who published a first-author peer-reviewed paper and is now a PhD student at Colorado State University. In addition, Lloyd published a short animated film for Scientific American, translating information about deep subsurface biogeochemistry and completed a book on this subject for popular audiences that will be published by Princeton University Press in May 2025.

Last Modified: 01/22/2025
Modified by: Karen G Lloyd