Abstract
Metaproteomics is an increasingly popular methodology that provides information regarding the metabolic functions of specific microbial taxa and has potential for contributing to ocean ecology and biogeochemical studies. A blinded multi-laboratory intercomparison was conducted to assess comparability and reproducibility of taxonomic and functional results and their sensitivity to methodological variables. Euphotic zone samples from the Bermuda Atlantic Time-series Study (BATS) in the North Atlantic Ocean collected by in situ pumps and the autonomous underwater vehicle (AUV) Clio were distributed with a paired metagenome, and one-dimensional (1D) liquid chromatographic data-dependent acquisition mass spectrometry analysis was stipulated. Analysis of mass spectra from seven laboratories through a common bioinformatic pipeline identified a shared set of 1056 proteins from 1395 shared peptide constituents. Quantitative analyses showed good reproducibility: pairwise regressions of spectral counts between laboratories yielded R2 values averaged 0.62 ± 0.11, and a Sørensen similarity analysis of the top 1000 proteins revealed 70 %–80 % similarity between laboratory groups. Taxonomic and functional assignments showed good coherence between technical replicates and different laboratories. A bioinformatic intercomparison study, involving 10 laboratories using eight software packages, successfully identified thousands of peptides within the complex metaproteomic datasets, demonstrating the utility of these software tools for ocean metaproteomic research. Lessons learned and potential improvements in methods were described. Future efforts could examine reproducibility in deeper metaproteomes, examine accuracy in targeted absolute quantitation analyses, and develop standards for data output formats to improve data interoperability. Together, these results demonstrate the reproducibility of metaproteomic analyses and their suitability for microbial oceanography research, including integration into global-scale ocean surveys and ocean biogeochemical models.
| Original language | English |
|---|---|
| Pages (from-to) | 4889-4908 |
| Number of pages | 20 |
| Journal | Biogeosciences |
| Volume | 21 |
| Issue number | 21 |
| DOIs | |
| State | Published - Nov 8 2024 |
Funding
This article is a product of the sustained efforts of a small group activity supported by the Ocean Carbon and Biogeochemistry (OCB) Project Office (NSF OCE-1850983 and NASA NNX17AB17G), based on a proposal written by Mak A. Saito and Matthew R. McIlvin. The research expedition where samples were collected was supported by the NSF Biological Oceanography and Chemical Oceanography. AUV Clio sample collection was supported by NSF OCE 1658030 and 1924554. Analyses by participating laboratories acknowledge support from the following: NSERC Discovery Grant RGPIN-2015-05009 and Simons Foundation grant no. 504183 to Erin M. Bertrand; the Austrian Science Fund (FWF) DEPOCA (project no. AP3558721) to Gerhard J. Herndl; Simons Foundation grant no. 402971 to Jacob R. Waldbauer; National Institute of Health grant no. 1R21ES034337-01 to Brook L. Nunn; the Norwegian Centennial Chair Program at the University of Minnesota for funding to Pratik D. Jagtap, Subina Mehta, and Tim J. Griffin (grant nos. NIH R01 GM135709, NSF OCE-1924554, and OCE-2019589); and Simons Foundation grant no. 1038971 to Mak A. Saito We thank the R/V Atlantic Explorer and the Bermuda Atlantic Time-series Study team for assistance at sea. We thank Mary Zawoysky for editorial and study anonymization assistance. We thank Magnus Palmblad, John Kucklick, and an anonymous reviewer for comments on the pre-submission version of the article. We also thank the two anonymous reviewers for their constructive comments during the review of this article. This article is a product of the sustained efforts of a small group activity supported by the Ocean Carbon and Biogeochemistry (OCB) Project Office (NSF OCE-1850983 and NASA NNX17AB17G), based on a proposal written by Mak A. Saito and Matthew R. McIlvin. The research expedition where samples were collected was supported by the NSF Biological Oceanography and Chemical Oceanography. AUV Clio sample collection was supported by NSF OCE 1658030 and 1924554. Analyses by participating laboratories acknowledge support from the following: NSERC Discovery Grant RGPIN-2015-05009 and Simons Foundation grant no. 504183 to Erin M. Bertrand; the Austrian Science Fund (FWF) DEPOCA (project no. AP3558721) to Gerhard J. Herndl; Simons Foundation grant no. 402971 to Jacob R. Waldbauer; National Institute of Health grant no. 1R21ES034337-01 to Brook L. Nunn; the Norwegian Centennial Chair Program at the University of Minnesota for funding to Pratik D. Jagtap, Subina Mehta, and Tim J. Griffin (grant nos. NIH R01 GM135709, NSF OCE-1924554, and OCE-2019589); and Simons Foundation grant no. 1038971 to Mak A. Saito
| Funders | Funder number |
|---|---|
| University of Minnesota | |
| Ocean Carbon and Biogeochemistry | |
| Bermuda Atlantic Time-series Study | |
| Austrian Science Fund | |
| Magnus Palmblad | |
| John Kucklick | |
| DEPOCA | AP3558721, 402971 |
| Natural Sciences and Engineering Research Council of Canada | RGPIN-2015-05009 |
| Natural Sciences and Engineering Research Council of Canada | |
| National Institutes of Health | 1R21ES034337-01, OCE-2019589, R01 GM135709, 1038971, OCE-1924554 |
| National Institutes of Health | |
| Simons Foundation | 504183 |
| Simons Foundation | |
| National Science Foundation | OCE-1850983, 1924554, NASA NNX17AB17G, OCE 1658030 |
| National Science Foundation |