Dataset: Montastraea annularis white plague viromes and 16S amplicons

Name: Montastraea annularis white plague coral viromes and 16S amplicon libraries analyzed in the Vega Thurber lab at Florida International University, North Miami, FL (Coral Virus project)
Published: 2015-05-12
Keywords: oceans

View Data: Data not available yet

Cite This Dataset

Project:

Effects of Viruses on Coral Fitness (Coral Virus)

Principal Investigator:

Rebecca Vega Thurber (Oregon State University, OSU)

BCO-DMO Data Manager:

Shannon Rauch (Woods Hole Oceanographic Institution, WHOI BCO-DMO)

Version:

12 May 2015

Version Date:

2015-05-12

Validated:

Current State:

Preliminary and in progress

Montastraea annularis white plague coral viromes and 16S amplicon libraries analyzed in the Vega Thurber lab at Florida International University, North Miami, FL (Coral Virus project)

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Description

This collection of 23 viral metagenomes and their associated 16S amplicon data were derived from seawater (n=2) and tissues of 1) apparently healthy Montastraea annularis corals (n=2), 2) diseased M. annularis corals (n=7) exhibiting signs of white plague disease, 3) bleached tissues of M. annularis corals exhibiting signs of white plague disease (n=7), and 4) bleached tissues of apparently healthy but bleached M. annularis (n=5).

Data access: These data are stored at Metavir a web server designed to annotate viral metagenomic sequences (raw reads or assembled contigs). http://metavir-meb.univ-bpclermont.fr/index.php?page=Welcome. This dataset on the published viromes, can be identified under "public projects", under the project entitled “WP reads – Soffer et al 2013.” One must "launch" each library and download the associated FASTA files individually: A=bleached from diseased samples, H=healthy bleached samples, D=diseased samples, UB=unbleached healthy samples, SW= seawater, SWD=seawater diseased.

Methods & Sampling

Sampling of the scleractinian coral, M. annularis, was conducted at Brewers Bay, in St. Thomas, US Virgin Islands, during a concurrent WP outbreak and bleaching event in September 2010. Collections took place over 2 days at depths of 5.5–7.6 m. Temperature at depth was 29.0 degrees C. All M. annularis colonies used in this study were located within ~75 m of each other. Coral samples were collected using SCUBA, where two to three cores of tissue attached to skeleton were removed from each M. annularis colony using a 2 cm diameter corer and hammer (USVI Department of Planning and Natural Resources permit #STT-050-10). Samples were collected at depths of 5.5–7.6 m and at temperatures 29 degrees C. Colonies were within 5–7 m of each other and 75 m from shore.

Viromes: Phage and eukaryovirus particles were isolated and sequenced separately from bacterial cells (using CsCl density gradient ultracentrifugation) before sequencing (Vega Thurber et al., 2009; more details in Soffer et al., 2014). DNA was then extracted with a phenol-chloroform extraction protocol (Vega Thurber et al., 2009; Soffer et al., 2014) and amplified using non-specific MDA according to manufacturer’s protocol (GenomPhi, GE Healthcare, Pittsburgh, PA, USA). The coral virome libraries (21 coral samples) were barcoded and pyrosequenced at EnGencore (University of South Carolina) on a Roche Titanium 454 platform. The final numbers of replicate libraries for each coral health state were: H (n = 2), B (n = 5), BD (n = 7) and D (n = 7).

Bacterial Amplicons: 500 ul of ethanol/tissue slurry was pipetted for DNA extractions. A modified organic extraction protocol was used to purify DNA (for details see Soffer, Zaneveld, and Vega Thurber (2015) Phage–bacteria network analysis and its implication for the understanding of coral disease. Environmental Microbiology 17:1203-1218). Isolated nucleic acids were amplified using barcoded primers 515F and 806R, which were chosen due to their ability to amplify both bacteria and archaea (Caporaso et al., 2011; Walters et al., 2011). Triplicate amplicon libraries were prepared using GoTaq Flexi reagents from Promega (Madison, WI, USA) using standard protocols and the following PCR cycle: 1 cycle of 94 degrees C for 3 min, 35 cycles of 94 degrees C for 45 s, 50 degrees C for 60 s, and 72 degrees C for 90 s, and then 1 cycle of 72 degrees C for 10 minutes. PCR products were run on a 1.5 agarose gel, triplicates pooled, cleaned using AMPure magnetic beads from Agencourt (Brea, CA, USA) and quantified with a Qubit HS dsDNA kit (Invitrogen,Eugene, OR, USA) into equimolar ratios. Quality of amplicons were determine on an Agilent Bioanalyser 2100 before being pyrosequenced on a 454 GS Junior Roche at the Oregon State University Center for Genome Research and Biocomputing Core Laboratories.

Related Publications:
Soffer,N., Brandt, M.E., Correa, A.M., Smith, T.B. and Vega Thurber, R.L. 2014. Potential role of viruses in white plague coral disease. ISME J., 8(2): 271-283. doi:10.1038/ismej.2013.137

Soffer, N., Zaneveld, J., and Vega Thurber, R. 2015. Phage–bacteria network analysis and its implication for the understanding of coral disease. Environmental Microbiology, 17:1203-1218. doi:10.1111/1462-2920.12553

Data Processing Description

Sequence reads underwent several preliminary bioinformatic steps.

Viromes: SFF files were converted to FASTA/FASTQ files and de-replicated using the program GALAXY (Goecks et al., 2010). Low quality reads (that is, thoseo100 bp in length and/or with quality scoresoQ20) were removed. To eliminate any potential non-viral sequences from the data sets, the program DeconSeq was used to identify and remove reads with nucleic acid homology (based on 60% identity and 94% similarity) to eukaryotes (mouse, fish, human and mosquito), bacteria and/or archaea (Schmieder and Edwards, 2011. The coral viromes (24 samples total, 1 plate) were barcoded and pyrosequenced on a Titanium 454 platform from Roche at EnGencore (San Francisco, CA, USA) (University of South Carolina).

These data were stored at Metavir a web server designed to annotate viral metagenomic sequences (raw reads or assembled contigs). http://metavir-meb.univ-bpclermont.fr/index.php?page=Welcome. This dataset on the published viromes, can be identified under "public projects", under the project entitled “WP reads – Soffer et al 2013.” One must “launch” each library and download the associated FASTA files individually: A=bleached from diseased samples, H=healthy bleached samples, D=diseased samples, UB=unbleached healthy samples, SW= seawater, SWD=seawater diseased.

Bacterial amplicons: 16S rRNA gene sequence libraries were analysed in QIIME and PRIMER 6 (Clarke and Gorley, 2006; Caporaso et al., 2010). First, reads were de-multiplexed based on their error correcting barcodes and filtered for quality using default parameters (quality score greater than or eaqual to 25, min length = 200, max length = 1000, and no ambiguous bases and mismatches allowed). Next OTUs were assigned at a 97% similarity threshold UCLUST (Edgar, 2010), and OTU tables constructed from the assignments. Lastly, taxonomic annotations were made to the RDP database using the RDP classifier (Wang et al., 2007). OTUs identified as chloroplasts were removed from the analysis (mean 23% +/- 0.2). Amplicon libraries were paired with all virome samples with the exception that there was an additional bleached sample was processed for bacterial analysis.

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Funding Sources