Tracking de novo protein synthesis in the activated sludge microbiome using BONCAT-FACS

Zhe Du; Sebastian F. Behrens

doi:10.1016/j.watres.2021.117696

Tracking de novo protein synthesis in the activated sludge microbiome using BONCAT-FACS

Zhe Du, Sebastian F. Behrens

Civil, Environmental, and Geo- Engineering

Research output: Contribution to journal › Article › peer-review

4 Scopus citations

Abstract

In order to ensure stable performance of engineered biotechnologies that rely on mixed microbial community systems, it is important to identify process-specific microbial traits and study their in-situ activity and responses to changing environmental conditions and system operational parameters. We used BioOrthogonal Non-Canonical Amino acid Tagging (BONCAT) in combination with Fluorescence-Activated Cell Sorting (FACS) and 16S rRNA gene amplicon sequencing to identify translationally active cells in activated sludge. We found that only a subset of the activated sludge microbiome is translationally active during the aerobic treatment phase of a full-scale sequencing batch reactor designed to enhance biological phosphorus removal from municipal wastewater. Relative abundance of amplicon sequence variants was not a reliable predictor of species activity. BONCAT-positive and -negative cells revealed a broad range of population-wide and taxa-specific translational heterogeneity. BONCAT-FACS in combination with amplicon sequencing can provide new insights into the ecophysiology of highly dynamic microbiomes in activated sludge systems.

Original language	English (US)
Article number	117696
Journal	Water Research
Volume	205
Early online date	Sep 24 2021
DOIs	https://doi.org/10.1016/j.watres.2021.117696
State	Published - Oct 15 2021

Bibliographical note

Funding Information:
We would like to thank the staff at Brainerd Wastewater Treatment Facilities for plant access and help with sample collection. We highly appreciate the assistance of Julie Johnston and Aaron Pauleon on field trips and during samples processing for FACS in the lab. We thank Mariah Dorner for critically reading and editing of the manuscript. Dr. Zhe Du was supported by a MnDRIVE Environment Seed Grant for postdoctoral scholars. Minnesota's Discovery, Research, and InnoVation Economy (MnDRIVE) is a partnership between the University of Minnesota and the State of Minnesota. The research was enabled by the Legislative-Citizen Commission on Minnesota Resources (LCCMR) through a grant entitled “Wastewater Treatment Process Improvements” funded by the Environment and Natural Resources Trust Fund ( ENRTF ) under legal citation M.L. 2016, Chp. 186, Section 2 , Subd. 04 k.

Funding Information:
We would like to thank the staff at Brainerd Wastewater Treatment Facilities for plant access and help with sample collection. We highly appreciate the assistance of Julie Johnston and Aaron Pauleon on field trips and during samples processing for FACS in the lab. We thank Mariah Dorner for critically reading and editing of the manuscript. Dr. Zhe Du was supported by a MnDRIVE Environment Seed Grant for postdoctoral scholars. Minnesota's Discovery, Research, and InnoVation Economy (MnDRIVE) is a partnership between the University of Minnesota and the State of Minnesota. The research was enabled by the Legislative-Citizen Commission on Minnesota Resources (LCCMR) through a grant entitled ?Wastewater Treatment Process Improvements? funded by the Environment and Natural Resources Trust Fund (ENRTF) under legal citation M.L. 2016, Chp. 186, Section 2, Subd. 04 k.

Publisher Copyright:
© 2021

Keywords

Activated sludge
Bioorthogonal non-canonical amino acid tagging (BONCAT)
Fluorescence-activated cell sorting (FACS)
Microbial activity
Sequencing batch reactor
Wastewater
Microbiota
Flow Cytometry
Sewage
Bioreactors
RNA, Ribosomal, 16S/genetics
Amino Acids

PubMed: MeSH publication types

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10.1016/j.watres.2021.117696

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title = "Tracking de novo protein synthesis in the activated sludge microbiome using BONCAT-FACS",

abstract = "In order to ensure stable performance of engineered biotechnologies that rely on mixed microbial community systems, it is important to identify process-specific microbial traits and study their in-situ activity and responses to changing environmental conditions and system operational parameters. We used BioOrthogonal Non-Canonical Amino acid Tagging (BONCAT) in combination with Fluorescence-Activated Cell Sorting (FACS) and 16S rRNA gene amplicon sequencing to identify translationally active cells in activated sludge. We found that only a subset of the activated sludge microbiome is translationally active during the aerobic treatment phase of a full-scale sequencing batch reactor designed to enhance biological phosphorus removal from municipal wastewater. Relative abundance of amplicon sequence variants was not a reliable predictor of species activity. BONCAT-positive and -negative cells revealed a broad range of population-wide and taxa-specific translational heterogeneity. BONCAT-FACS in combination with amplicon sequencing can provide new insights into the ecophysiology of highly dynamic microbiomes in activated sludge systems.",

keywords = "Activated sludge, Bioorthogonal non-canonical amino acid tagging (BONCAT), Fluorescence-activated cell sorting (FACS), Microbial activity, Sequencing batch reactor, Wastewater, Microbiota, Flow Cytometry, Sewage, Bioreactors, RNA, Ribosomal, 16S/genetics, Amino Acids",

author = "Zhe Du and Behrens, {Sebastian F.}",

note = "Funding Information: We would like to thank the staff at Brainerd Wastewater Treatment Facilities for plant access and help with sample collection. We highly appreciate the assistance of Julie Johnston and Aaron Pauleon on field trips and during samples processing for FACS in the lab. We thank Mariah Dorner for critically reading and editing of the manuscript. Dr. Zhe Du was supported by a MnDRIVE Environment Seed Grant for postdoctoral scholars. Minnesota's Discovery, Research, and InnoVation Economy (MnDRIVE) is a partnership between the University of Minnesota and the State of Minnesota. The research was enabled by the Legislative-Citizen Commission on Minnesota Resources (LCCMR) through a grant entitled “Wastewater Treatment Process Improvements” funded by the Environment and Natural Resources Trust Fund ( ENRTF ) under legal citation M.L. 2016, Chp. 186, Section 2 , Subd. 04 k. Funding Information: We would like to thank the staff at Brainerd Wastewater Treatment Facilities for plant access and help with sample collection. We highly appreciate the assistance of Julie Johnston and Aaron Pauleon on field trips and during samples processing for FACS in the lab. We thank Mariah Dorner for critically reading and editing of the manuscript. Dr. Zhe Du was supported by a MnDRIVE Environment Seed Grant for postdoctoral scholars. Minnesota's Discovery, Research, and InnoVation Economy (MnDRIVE) is a partnership between the University of Minnesota and the State of Minnesota. The research was enabled by the Legislative-Citizen Commission on Minnesota Resources (LCCMR) through a grant entitled ?Wastewater Treatment Process Improvements? funded by the Environment and Natural Resources Trust Fund (ENRTF) under legal citation M.L. 2016, Chp. 186, Section 2, Subd. 04 k. Publisher Copyright: {\textcopyright} 2021",

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AU - Behrens, Sebastian F.

N1 - Funding Information: We would like to thank the staff at Brainerd Wastewater Treatment Facilities for plant access and help with sample collection. We highly appreciate the assistance of Julie Johnston and Aaron Pauleon on field trips and during samples processing for FACS in the lab. We thank Mariah Dorner for critically reading and editing of the manuscript. Dr. Zhe Du was supported by a MnDRIVE Environment Seed Grant for postdoctoral scholars. Minnesota's Discovery, Research, and InnoVation Economy (MnDRIVE) is a partnership between the University of Minnesota and the State of Minnesota. The research was enabled by the Legislative-Citizen Commission on Minnesota Resources (LCCMR) through a grant entitled “Wastewater Treatment Process Improvements” funded by the Environment and Natural Resources Trust Fund ( ENRTF ) under legal citation M.L. 2016, Chp. 186, Section 2 , Subd. 04 k. Funding Information: We would like to thank the staff at Brainerd Wastewater Treatment Facilities for plant access and help with sample collection. We highly appreciate the assistance of Julie Johnston and Aaron Pauleon on field trips and during samples processing for FACS in the lab. We thank Mariah Dorner for critically reading and editing of the manuscript. Dr. Zhe Du was supported by a MnDRIVE Environment Seed Grant for postdoctoral scholars. Minnesota's Discovery, Research, and InnoVation Economy (MnDRIVE) is a partnership between the University of Minnesota and the State of Minnesota. The research was enabled by the Legislative-Citizen Commission on Minnesota Resources (LCCMR) through a grant entitled ?Wastewater Treatment Process Improvements? funded by the Environment and Natural Resources Trust Fund (ENRTF) under legal citation M.L. 2016, Chp. 186, Section 2, Subd. 04 k. Publisher Copyright: © 2021

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N2 - In order to ensure stable performance of engineered biotechnologies that rely on mixed microbial community systems, it is important to identify process-specific microbial traits and study their in-situ activity and responses to changing environmental conditions and system operational parameters. We used BioOrthogonal Non-Canonical Amino acid Tagging (BONCAT) in combination with Fluorescence-Activated Cell Sorting (FACS) and 16S rRNA gene amplicon sequencing to identify translationally active cells in activated sludge. We found that only a subset of the activated sludge microbiome is translationally active during the aerobic treatment phase of a full-scale sequencing batch reactor designed to enhance biological phosphorus removal from municipal wastewater. Relative abundance of amplicon sequence variants was not a reliable predictor of species activity. BONCAT-positive and -negative cells revealed a broad range of population-wide and taxa-specific translational heterogeneity. BONCAT-FACS in combination with amplicon sequencing can provide new insights into the ecophysiology of highly dynamic microbiomes in activated sludge systems.

AB - In order to ensure stable performance of engineered biotechnologies that rely on mixed microbial community systems, it is important to identify process-specific microbial traits and study their in-situ activity and responses to changing environmental conditions and system operational parameters. We used BioOrthogonal Non-Canonical Amino acid Tagging (BONCAT) in combination with Fluorescence-Activated Cell Sorting (FACS) and 16S rRNA gene amplicon sequencing to identify translationally active cells in activated sludge. We found that only a subset of the activated sludge microbiome is translationally active during the aerobic treatment phase of a full-scale sequencing batch reactor designed to enhance biological phosphorus removal from municipal wastewater. Relative abundance of amplicon sequence variants was not a reliable predictor of species activity. BONCAT-positive and -negative cells revealed a broad range of population-wide and taxa-specific translational heterogeneity. BONCAT-FACS in combination with amplicon sequencing can provide new insights into the ecophysiology of highly dynamic microbiomes in activated sludge systems.

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KW - Wastewater

KW - Microbiota

KW - Flow Cytometry

KW - Sewage

KW - Bioreactors

KW - RNA, Ribosomal, 16S/genetics

KW - Amino Acids

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Tracking de novo protein synthesis in the activated sludge microbiome using BONCAT-FACS

Abstract

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Keywords

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