Article Navigation
Article Contents
-
Abstract
-
Acknowledgements
-
Author contributions
-
Conflicts of interest
-
Funding
-
Data availability
-
References
- < Previous
- Next >
Journal Article
, Shengjie Li Department of Biogeochemistry, Max Planck Institute for Marine Microbiology , Bremen 28359 , Germany Search for other works by this author on: Oxford Academic Damon Mosier Department of Earth , Energy and Environment, University of Calgary , Calgary, AB T2N 1N4 , Canada Search for other works by this author on: Oxford Academic Angela Kouris Department of Earth , Energy and Environment, University of Calgary , Calgary, AB T2N 1N4 , Canada Search for other works by this author on: Oxford Academic Pauline Humez Department of Earth , Energy and Environment, University of Calgary , Calgary, AB T2N 1N4 , Canada Search for other works by this author on: Oxford Academic Bernhard Mayer Department of Earth , Energy and Environment, University of Calgary , Calgary, AB T2N 1N4 , Canada Search for other works by this author on: Oxford Academic Marc Strous Department of Earth , Energy and Environment, University of Calgary , Calgary, AB T2N 1N4 , Canada Search for other works by this author on: Oxford Academic Muhe Diao Department of Earth , Energy and Environment, University of Calgary , Calgary, AB T2N 1N4 , Canada Corresponding author: Muhe Diao, EEEL509, University of Calgary, 2500 University Drive NW, Calgary, AB T2N 1N4, Canada. Email: muhe.diao@ucalgary.ca Search for other works by this author on: Oxford Academic
ISME Communications, Volume 4, Issue 1, January 2024, ycae023, https://doi.org/10.1093/ismeco/ycae023
Published:
12 February 2024
Article history
Received:
08 December 2023
Revision received:
02 February 2024
Accepted:
09 February 2024
Published:
12 February 2024
Corrected and typeset:
17 March 2024
- Split View
- Views
- Article contents
- Figures & tables
- Video
- Audio
- Supplementary Data
-
Cite
Cite
Shengjie Li, Damon Mosier, Angela Kouris, Pauline Humez, Bernhard Mayer, Marc Strous, Muhe Diao, High diversity, abundance, and expression of hydrogenases in groundwater, ISME Communications, Volume 4, Issue 1, January 2024, ycae023, https://doi.org/10.1093/ismeco/ycae023
Close
Search
Close
Search
Advanced Search
Search Menu
Abstract
Hydrogen may be the most important electron donor available in the subsurface. Here we analyse the diversity, abundance and expression of hydrogenases in 5 proteomes, 25 metagenomes, and 265 amplicon datasets of groundwaters with diverse geochemistry. A total of 1545 new [NiFe]-hydrogenase gene sequences were recovered, which considerably increased the number of sequences (1999) in a widely used database. [NiFe]-hydrogenases were highly abundant, as abundant as the DNA-directed RNA polymerase. The abundance of hydrogenase genes increased with depth from 0 to 129m. Hydrogenases were present in 481 out of 1245 metagenome-assembled genomes. The relative abundance of microbes with hydrogenases accounted for ~50% of the entire community. Hydrogenases were actively expressed, making up as much as 5.9% of methanogen proteomes. Most of the newly discovered diversity of hydrogenases was in “Group 3b”, which has been associated with sulfur metabolism. “Group 3d”, facilitating the interconversion of electrons between hydrogen and NAD, was the most abundant and mainly observed in methanotrophs and chemoautotrophs. “Group 3a”, associated with methanogenesis, was the most abundant in proteomes. Two newly discovered groups of [NiFe]-hydrogenases, observed in Methanobacteriaceae and Anaerolineaceae, further expanded diversity. Our results highlight the vast diversity, abundance and expression of hydrogenases in groundwaters, suggesting a high potential for hydrogen oxidation in subsurface habitats.
Generating hydrogen (H2) from solar and wind energy, and subsequently storing it on a terawatt scale in the subsurface is currently considered a key aspect of the energy transition [1-3]. One of the potential challenges of this approach is the microbial oxidation of hydrogen, which could induce hydrogen loss [4-6]. Our recent work suggested a high potential for microbial hydrogen turnover in groundwaters, based on dissolved hydrogen concentrations, as well as detection and activity of hydrogenotrophic methanogens [7]. Here we explored the diversity and potential functions of hydrogenases with an expanded sample set encompassing 265 groundwater samples (geochemically characterised and amplicon sequenced, with additional 25 metagenomes and 5 proteomes) from 138 wells in Alberta (Canada), with sampling depths between 0 and 157m (Fig. S1, Tables S1, S2). The groundwaters displayed a range of oxidation states from oxic to completely reduced, accompanied with a wide range of sulfate (>10g/L to below detection) and methane concentrations (74mg/L to below detection).
The abundance and expression of different types of hydrogenases were estimated based on unassembled reads, assembled contigs, metagenome-assembled genomes (MAGs), and proteins in 25 groundwaters. Few [Fe]- and [FeFe]-hydrogenases were present in our data. Notably, the catalytic subunit of [NiFe]-hydrogenase was as abundant as the DNA-directed RNA polymerase (rpoB) (Fig. 1A, Table S3). In 12 out of 25 metagenomes, hydrogenase genes were more abundant than rpoB genes, indicating multiple copies of same subtypes or various subtypes of hydrogenases in a single genome. The ratio of hydrogenase over rpoB correlated positively with depth (P = 0.009, Fig. 1B). From metagenomes, 616 high-quality and 629 medium-quality MAGs were obtained. Hydrogenases were present in 481 out of 1245 MAGs, which together accounted on average for 50% of the relative abundance of all MAGs (Fig. 1C, Table S4). In eight samples, MAGs with hydrogenases accounted for >70% of all MAGs. Although conducting proteomics with groundwater samples is challenging due to low cell counts, we obtained proteomes of five groundwaters, showing hydrogenases accounted for 0.016–1.0% of all proteins (Tables S5–S9). In proteomes of individual species associated with our MAGs, the relative abundance of hydrogenases ranged from 0.0026% (Methylomonadaceae) to 5.9% (Methanobacteriaceae) (Fig. 1D). Interestingly, hydrogenase was more abundant in the three methanogen proteomes (>1.3%) than the 21 bacterial proteomes (<0.19%) (Tables S5–S9). Thus, hydrogenase genes might be one of the most prevalent genes in the subsurface and active expression indicated these genes were functional.
Figure 1
Abundance and expression of hydrogenases. (A) Ratio of reads mapped to hydrogenase genes over reads mapped to rpoB genes in metagenome of 25 groundwater samples. The arrow indicates the subgroup of hydrogenases with the highest abundance in sampled groundwaters. (B) Relationship between depth and total abundance of hydrogenases. Spearman’s rank correlation coefficient and the P-value are shown. The line shows the linear regression. (C) Relative abundance of 1245 MAGs (13–194 per sample) with and without hydrogenases. MQ: medium-quality. HQ: high- quality. Relative abundances were based on reads mapped to an MAG divided by total reads mapped. (D) Abundance of hydrogenases in proteomes. Relative abundance within a sample was calculated as % of all peptide spectral matches of the sample. Relative abundance for individual MAGs was calculated as % of all peptide spectral matches associated with the MAG.
Open in new tabDownload slide
From the assembled contigs, 1545 [NiFe]-hydrogenase gene sequences were recovered (Supplementary Result 1), which displayed vast diversity (Table S10). These groundwater hydrogenase sequences considerably increased the number (1999) of [NiFe]-hydrogenase sequences present in a widely used database (Fig. 2) [8]. The newly discovered diversity, abundance, and expression were concentrated among a few specific subtypes of [NiFe]-hydrogenases, groups 1e, 3a, 3b, and 3d. Most of the new diversity was observed in group 3b, while group 3d was the most abundant in metagenomes and group 3a was the most abundant in proteomes.
Figure 2
Phylogenetic tree of the catalytic subunit of [NiFe]-hydrogenases. The tree is midpoint-rooted. An arrow inside indicates the place of [NiFe]-hydrogenases with high diversity, abundance, or expression discovered in sampled groundwaters. Any sequences with total read counts over 1000 in the 25 samples are marked with a star. From inside to outside, the three rings around the tree indicate (1) source, (2) phylum-level taxonomy, and (3) subgroups based on HydDB [5].
Open in new tabDownload slide
Hydrogenases of groups 1e and 3b are associated with sulfur reduction [8, 9]. In our data, the abundance of groups 1e and 3b both positively correlated with sulfate concentration (P = 0.025 and 0.007, respectively, Fig. S2). Group 3b hydrogenases were occasionally observed in close proximity to sulfhydrogenase subunit delta and sulfite reductase subunit A (Tables S12, S13). Among high-quality MAGs, group 1e was exclusively present in members of Burkholderiales, and sometimes co-existed with group 3b. Group 3b was commonly detected in MAGs of sulfate-reducing microorganisms, particularly thirteen Desulfobacterota and three Thermodesulfovibrionia. These MAGs also encoded sulfate adenylyltransferase, adenylylsulfate reductase, and dissimilatory sulfite reductase (Table S11). Some group 3b hydrogenases were detected in MAGs of sulfur-oxidising microorganisms such as three Gallionellaceae that encoded sulfide:quinone oxidoreductase and sulfite dehydrogenase, and four Thiobacillaceae that contained the thiosulfate oxidation sox complex. These hydrogenases might also function alongside sulfur oxidation, coupled to oxygen or nitrate reduction. Many other group 3b were detected in genomes of microbial “dark matter” clades, such as Patescibacteria (14) and Omnitrophota (5), consistent with previous findings [10, 11].
Group 3a hydrogenases are associated with methanogenesis [8, 9] and were most abundant in our proteomes (Fig. 1D, Tables S5–S9). They were exclusively observed in MAGs of hydrogenotrophic methanogens, six Methanomicrobiales and four Methanobacteriales [12]. All Methanobacteriales also encoded tetrahydromethanopterin-reducing [Fe]-hydrogenases. Nine of them possessed tetrahydromethanopterin S-methyltransferase (Mtr), the key enzyme for hydrogenotrophic methanogenesis [13]. Two of them contained Methylamine:Coenzyme M Methyltransferase (mtbA), suggesting that they might produce methane from methylamine [14]. These results were consistent with previous research showing active conversion of CO2 into methane in hydrocarbon reservoirs [15].
Group 3d is associated with fermentative metabolism and chemoautotrophy, interconverting electrons between hydrogen and NADH depending on cellular redox state [8, 9]. Group 3d was the most abundant subgroup in 15 out of 25 groundwater samples (Fig. 1A). Most group 3d hydrogenase genes were close to an NADP oxidoreductase gene (Tables S13, S14). 3d hydrogenase genes were present in 89 high-quality MAGs, with 12 of them encoding formate C-acetyltransferases or lactate dehydrogenases, both signature genes of fermentative metabolism. Of these MAGs, 21 were associated with methanotrophic Methylomonadaceae. For the other 68 MAGs, 40 of them contained both RuBisCO and phosphoribulokinase, indicating a functional Calvin cycle. For instance, MAGs associated with Rhodocyclaceae (12), Hydrogenophaga (7), Nitrosomonas (7), and Rhodoferax (5) fall into this category. Thus, it is likely that these chemolithoautotrophs can use hydrogen as an additional energy source, with the hydrogenase transferring electrons from H2 to NAD+ to drive their Calvin cycles.
Two newly discovered groups of [NiFe]-hydrogenases further expanded diversity. The first was positioned near the root of the tree (Fig. 2). This group consisted of three sequences, exclusively found in Methanobacteriaceae. The other newly discovered group was near the root of group 3b, composed of six sequences, including five sequences affiliated with Anaerolineaceae and one affiliated with Bathyarchaeia.
Consistency in the types/subgroups of hydrogenases and metabolisms among MAGs with the same taxonomic identity was observed for common groundwater residents, which helped to extrapolate metagenomic findings to 265 amplicon-sequenced groundwater samples. For example, the total relative abundance of Methylomonadaceae bacteria (all 21 MAGs had hydrogenases) could reach 88.6% (Table S15). Members of Hydrogenophaga (8 out of 14 MAGs had hydrogenases) could be as abundant as 71.2%. These findings suggest a high potential for hydrogen consumption in sampled subsurface habitats.
While the subsurface ecosystems analysed here would not be suitable for hydrogen storage, our study adds to growing evidence that hydrogenases are diverse, functional and ubiquitous in subsurface environments [16-19]. However, as hydrogenases were most abundant in methanogen proteomes, this need not always be a barrier to hydrogen storage, since recovery of methane could still be a desirable outcome. Likely, any subsurface environment at a temperature conducive to life would harbor microorganisms that thrive on hydrogen.
Acknowledgements
The authors thank the Groundwater Observation Well Network team members of Alberta Environment and Protected Areas (https://www.alberta.ca/groundwater-observation-well-network.aspx) for providing access to groundwater monitoring wells, sampling, and providing highest quality groundwater samples, and for sharing measurement results and expertise. We would like to thank the University of Calgary’s Center for Health Genomics and Informatics for sequencing and informatics services. We thank Carmen Li for help with MiSeq sequencing. We also thank Daan R. Speth for help with phylogenetic analysis.
Author contributions
SL, MS, and MD designed the study. SL, DM, AK, PH, BM, and MD performed lab research. SL, DM, MS, and MD analyzed the data. PH and BM assisted with data interpretation. SL, MS, and MD wrote the manuscript.
Conflicts of interest
None declared.
Funding
This study was supported by the Natural Sciences and Engineering Research Council (NSERC) through a Discovery Grant and Canada Research Chair (CRC-2020-00257 to M.S.), the Canada Foundation for Innovation (CFI), the Digital Research Alliance of Canada, the Canada First Research Excellence Fund (CFREF), the Government of Alberta, and the University of Calgary.
Data availability
Amplicons in this study are under the Bioproject PRJNA861683 and PRJNA700657. Metagenomes and metagenome-assembled genomes are under the Bioproject PRJNA700657 (NCBI). The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository [20] with the dataset identifier PXD044305.
References
1.
Krevor S de Coninck H Gasda SE
Subsurface carbon dioxide and hydrogen storage for a sustainable energy future
.
Nat Rev Earth Environ
2023
;
4
:
102
–
18
2.
Tarkowski R
Underground hydrogen storage: characteristics and prospects
.
Renew Sust Energ Rev
2019
;
105
:
86
–
94
3.
Zivar D Kumar S Foroozesh J
Underground hydrogen storage: a comprehensive review
.
Int J Hydrogen Energ
2021
;
46
:
23436
–
62
4.
Momper L Jungbluth SP Lee MD
Energy and carbon metabolisms in a deep terrestrial subsurface fluid microbial community
.
ISME J
2017
;
11
:
2319
–
33
5.
Stevens TO McKinley JP
Lithoautotrophic microbial ecosystems in deep basalt aquifers
.
1995
;
270
:
450
–
5
6.
Lappan R Shelley G Islam ZF
Molecular hydrogen in seawater supports growth of diverse marine bacteria
.
Nat Microbiol
2023
;
8
:
581
–
95
7.
Ruff SE Humez P de Angelis IH
Hydrogen and dark oxygen drive microbial productivity in diverse groundwater ecosystems
.
Nat Commun
2023
;
14
:
3194
8.
Sondergaard D Pedersen CN Greening C
HydDB: a web tool for hydrogenase classification and analysis
.
Sci Rep
2016
;
6
:
34212
9.
Greening C Biswas A Carere CR
Genomic and metagenomic surveys of hydrogenase distribution indicate H2 is a widely utilised energy source for microbial growth and survival
.
ISME J
2016
;
10
:
761
–
77
10.
Alvarado LEV Fakra SC Probst AJ
Autotrophic biofilms sustained by deeply-sourced groundwater host diverse CPR bacteria implicated in sulfur and hydrogen metabolism
.
Microbiome
2024
;
12
:
15
11.
Williams TJ Allen MA Berengut JF
Shedding light on microbial "dark matter": insights into novel Cloacimonadota and Omnitrophota from an Antarctic lake
.
Front Microbiol
2021
;
12
:741077. https://doi.org/10.3389/fmicb.2021.741077.
OpenURL Placeholder Text
12.
Thauer RK Kaster AK Goenrich M
Hydrogenases from methanogenic archaea, nickel, a novel cofactor, and H2 storage
.
Annu Rev Biochem
2010
;
79
:
507
–
36
13.
Berghuis BA Yu FB Schulz F
Hydrogenotrophic methanogenesis in archaeal phylum Verstraetearchaeota reveals the shared ancestry of all methanogens
.
Proc Natl Acad Sci U S A
2019
;
116
:
5037
–
44
14.
Vanwonterghem I Evans PN Parks DH
Methylotrophic methanogenesis discovered in the archaeal phylum Verstraetearchaeota
.
Nat Microbiol
2016
;
1
:
16170
15.
Tyne RL Barry PH Lawson M
Rapid microbial methanogenesis during CO2 storage in hydrocarbon reservoirs
.
Nature
2021
;
600
:
670
–
4
16.
Hernsdorf AW Amano Y Miyakawa K
Potential for microbial H2 and metal transformations associated with novel bacteria and archaea in deep terrestrial subsurface sediments
.
ISME J
2017
;
11
:
1915
–
29
17.
Adhikari RR Glombitza C Nickel JC
Hydrogen utilization potential in subsurface sediments
.
Front Microbiol
2016
;
7
:
8
18.
Hidalgo KJ Sierra-Garcia IN Zafra G
Genome-resolved meta-analysis of the microbiome in oil reservoirs worldwide
.
Microorganisms
2021
;
9
:
1812
19.
Vigneron A Alsop EB Lomans BP
Succession in the petroleum reservoir microbiome through an oil field production lifecycle
.
ISME J
2017
;
11
:
2141
–
54
20.
Perez-Riverol Y Csordas A Bai J
The PRIDE database and related tools and resources in 2019: improving support for quantification data
.
Nucleic Acids Res
2019
;
47
:
D442
–
50
© The Author(s) 2024. Published by Oxford University Press on behalf of the International Society for Microbial Ecology.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
Issue Section:
Brief Communication
Download all slides
Supplementary data
Supplementary data
SupplementaryInformation_ycae023 - docx file
SupplementaryTables_ycae023 - xlsx file
Advertisem*nt
Citations
Views
667
Altmetric
More metrics information
Metrics
Total Views 667
404 Pageviews
263 PDF Downloads
Since 2/1/2024
Month: | Total Views: |
---|---|
February 2024 | 77 |
March 2024 | 266 |
April 2024 | 157 |
May 2024 | 93 |
June 2024 | 74 |
Citations
Powered by Dimensions
Altmetrics
Email alerts
Article activity alert
Advance article alerts
New issue alert
In progress issue alert
Receive exclusive offers and updates from Oxford Academic
Email alerts
Advance article alerts
New issue alert
In progress issue alert
Receive exclusive offers and updates from Oxford Academic
Related articles in
- Google Scholar
Citing articles via
Google Scholar
-
Latest
-
Most Read
-
Most Cited
More from Oxford Academic
Biological Sciences
Microbial Ecology
Microbiology
Science and Mathematics
Books
Journals
Advertisem*nt