Metagenome and metatranscriptome sequences from deep-sea hydrothermal vent microbial communities collected on cruises AT42-22, TN405, and NA108 from May 2019 to Jun 2022
Project
| Contributors | Affiliation | Role |
|---|---|---|
| Hu, Sarah K. | Woods Hole Oceanographic Institution (WHOI) | Principal Investigator |
| Anderson, Rika | Carleton College | Co-Principal Investigator |
| Huber, Julie | Woods Hole Oceanographic Institution (WHOI) | Co-Principal Investigator |
| Mickle, Audrey | Woods Hole Oceanographic Institution (WHOI BCO-DMO) | BCO-DMO Data Manager |
Abstract
All samples were collected via ROVs, where an intake valve was positioned at the location of active diffuse flow (<100C) and collected into bags, bottles, or a filter directly. If collected into a bag or bottle, fluid was filtered immediately upon recovery. All filters had a pore size of 0.2µm and were composed of PES material.
Samples collected at the Mid-Cayman Rise and the Axial Seamount locations utilized the ROV Jason HOG sampler. Samples collected at the Gorda Ridge used the ROV Hercules SUPR sampler.
Metatranscriptomics
Messenger RNA (mRNA) was subset from previously extracted RNA from all sites and grazing experiments (Qiagen RNA mini kit). Then metatranscriptome libraries were prepared from the mRNA. Library was prepped with NEBNext Ultra II Directional RNA Library Prep Kit for Illumina #E7760S; with a Poly(A) mRNA magnetic isolation module first. For fragmentation, samples were incubated at 94C for 10 minutes. PCR cycles done until enough was isolated, range of 16-28. Mainly 20 or 22 cycles total. Input total RNA for metatranscriptome libraries was either 100 ng or 10 ng. However, with mRNA content ranging from 1-5% of the total RNA, the PCR amplification steps needed to be modified for the estimated input mRNA.
Libraries were sequenced with NovaSeq at the Northwest Genomics Sequence Center (Seattle, WA). Sequenced with NovaSeq S2 300 cycles. An average of 100 million sequences per sample were recovered from the metatranscriptome sequencing.
Metagenomics
Duplicate samples (other half of filter) from in situ sites at Mid-Cayman Rise were extracted for DNA (n = 12), then prepared as metagenome libraries, and sent for NovaSeq sequencing (data received January 2023). DNA extracted with MasterPure Complete DNA and RNA Purification Kit (Lucigen MC85200). Input DNA diluted to 50 ng total, or all DNA input (For low concentration samples) and sheared to target 400 bps with Covaris M220 focused-ultrasonicator, utilizing SonoLab 7.2. For 70 sections, 200 cycles (bursts) at 50 watts (Peak incident power), duty factor = 10%, and average incident power was 5 watts. Min temp: 18C, setpoint: 20C, and max: 22C.
Libraries were prepped with Ovation Ultralow System V2 (from Nugen). Library amplification was done at 15 cycles, and pooled. Sequencing was done with a NovaSeq S2 300 Cycle (UW Genomics NWGC). Over 80 million sequences per sample were recovered, to ensure sufficient sequencing depth.
- Imported submitted file "bco-dmo_Frenemies.csv" into the BCO-DMO system.
- Split lat_lon field into separate fields
- Converted lon to lon W, (West is negative)
- Created YEAR field in format YYYY by pulling the year from FIELDYR
- Added a CRUISE_ID field with corresponding cruise IDs given in submission by year
- Exported dataset as 936069_v1_frenemies_accession_metadata.csv
| Parameter | Description | Units |
| SAMPLE_ID | Complete sample ID associated with fastq files | unitless |
| SHORT_SAMPLE_ID | Short name sample identifier associated with cruise ID | unitless |
| SAMPLE_NAME | Related sample name associated with vent site | unitless |
| LAB_NUM | Internal number for sample inventory | unitless |
| CRUISE_ID | Identifer of cruise during which sample was collected | unitless |
| FIELD_REGION | Region of hydrothermal vent field | unitless |
| YEAR | Year of sample collection | unitless |
| FIELD_YEAR | Hydrothermal vent field with year of collection | unitless |
| VENT | Name of individual vent site or sample origin | unitless |
| LAT | Latitude of site where samples were collected. Negative values indicate South. | Decimal Degrees |
| LON | Longitude of site where samples were collected. Negative values indicate West. | Decimal Degrees |
| ORIGIN_TYPE | Origin of the sample, either in situ or grazing experiment | unitless |
| ORIGIN_DESCRIPTION | Collection method description | unitless |
| FRENEMIES_PROJ | Associated project | unitless |
| LIBRARY | Type of fastq sequence file | unitless |
| RUN | SRR number in NCBI | unitless |
| BIOSAMPLE | Biosample ID number in NCBI | unitless |
| BASES | Total number of bps in giga base pairs (Gb) | Giga base pairs (Gb) |
| BYTES | Total size of the sequence file in Gigabytes (GB) | Gigabytes |
| EXPERIMENT | Experiment ID for locating sequences in NCBI | unitless |
| LIBRARY_NAME | Sample identifer for fastq files | unitless |
| LIBRARY_SELECTION | Amplication used in library prep | unitless |
| geo_loc_name_country | Geo location name for SRA metadata | unitless |
| geo_loc_name_country_continent | Geo location associated with continent | unitless |
| geo_loc_name | Geo location name for site of vent field | unitless |
| Dataset-specific Instrument Name | Illumina NovaSeq 6000 |
| Generic Instrument Name | Automated DNA Sequencer |
| Dataset-specific Description | Libraries were sequenced with NovaSeq at the Northwest Genomics Sequence Center (Seattle, WA). |
| Generic Instrument Description | General term for a laboratory instrument used for deciphering the order of bases in a strand of DNA. Sanger sequencers detect fluorescence from different dyes that are used to identify the A, C, G, and T extension reactions. Contemporary or Pyrosequencer methods are based on detecting the activity of DNA polymerase (a DNA synthesizing enzyme) with another chemoluminescent enzyme. Essentially, the method allows sequencing of a single strand of DNA by synthesizing the complementary strand along it, one base pair at a time, and detecting which base was actually added at each step. |
| Dataset-specific Instrument Name | ROV Jason, HOG sampler |
| Generic Instrument Name | Hydrothermal Organic Geochemistry Sampler |
| Dataset-specific Description | All samples were collected via ROVs, where an intake valve was positioned at the location of active diffuse flow ( |
| Generic Instrument Description | The Hydrothermal Organic Geochemistry (HOG) sampler is designed to collect large volume (2–9 L) fluid samples with minimal introduction of organic or microbial contamination, and to be powered and deployed in real time from a submersible. Additional design constraints include utilizing materials appropriate for sampling fluids with elevated temperatures, fitting the sampler into the space available on the submersible, and minimizing the time needed to remove samples and prepare the sampler for re-deployment between dives. |
| Dataset-specific Instrument Name | Covaris M220 focused-ultrasonicator |
| Generic Instrument Name | ultrasonic cell disrupter (sonicator) |
| Dataset-specific Description | Input DNA diluted to 50 ng total, or all DNA input (For low concentration samples) and sheared to target 400 bps with Covaris M220 focused-ultrasonicator |
| Generic Instrument Description | Instrument that applies sound energy to agitate particles in a sample. |
AT42-22
| Website | |
| Platform | R/V Atlantis |
| Start Date | 2020-01-14 |
| End Date | 2020-02-06 |
TN405
| Website | |
| Platform | R/V Thomas G. Thompson |
| Start Date | 2022-07-08 |
| End Date | 2022-07-12 |
NA108
| Website | |
| Platform | E/V Nautilus |
| Start Date | 2019-05-24 |
| End Date | 2019-06-09 |
| Description |
Collaborative Research: Microbes need frenemies: unveiling microbial relationships with protists and viruses that support deep-sea hydrothermal vent food webs (frenemies)
Non-technical abstract:
Ecological interactions among microbes (bacteria and archaea), viruses, and eukaryotic microorganisms are critical junctions in marine food webs. These interactions range from mutually beneficial relationships to sources of microbial mortality. Interactions between viruses-microbes and eukaryotes-microbes at deep-sea hydrothermal vents impact local carbon cycling. This project aims to identify these microbial interactions, specifically those related to cell death by protistan grazing or viral lysis, and explore how they vary across different hydrothermal vent habitats. By providing a better understanding of the composition and nature of these relationships, the investigators aim to build a better food web model of deep-sea hydrothermal vents and improve our understanding of how climate change and other human activities impact the ecosystem. Outcomes from this project include the generation of new microbiology, oceanography, and computer science curricula targeted at community college students. In addition, it involves research with undergraduate students at all stages of the research process and provides opportunities for professional development and peer-to-peer mentoring.
Technical abstract:
This project examines trophic interactions among microbial eukaryotes, viruses, bacteria, and archaea at deep-sea hydrothermal vents using metagenomics and metatranscriptomics and characterizes these ecologically-significant interactions, such as mutualism, predator-prey, or virus-host. The investigators are sequencing samples collected to target archaea/bacteria, viruses, and eukaryotic grazers during a 2020 expedition to the Mid-Cayman Rise hydrothermal vent field to accomplish these goals. Specific aims of this project are to 1) Investigate the microbial, viral, and protistan assemblages and determine how lifestyle, community composition, and metabolism vary across venting fluids of the Mid-Cayman Rise; and 2) Identify ecologically-significant interactions among protists, viruses, bacteria, and archaea and incorporate these interactions into a model for turnover and exchange of carbon in the vent-associated food web. By modeling how trophic interactions influence microbial mortality, the proposed project substantially contributes to our understanding of the fate of carbon in one of the most productive ecosystems of the deep sea.
This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.
| Funding Source | Award |
|---|---|
| NSF Division of Ocean Sciences (NSF OCE) |
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