Here, we present two high-quality, draft metagenome-assembled genomes of deltaproteobacterial OalgDelta3 endosymbionts from the gutless marine worm Olavius algarvensis. Their 16S rRNA gene sequences share 98% identity with Delta3 endosymbionts of related host species Olavius ilvae (GenBank accession no. AJ620501) and Inanidrilus exumae (GenBank accession no. FM202060), for which no symbiont genomes are available.
ABSTRACT
Here, we present two high-quality, draft metagenome-assembled genomes of deltaproteobacterial OalgDelta3 endosymbionts from the gutless marine worm Olavius algarvensis. Their 16S rRNA gene sequences share 98% identity with Delta3 endosymbionts of related host species Olavius ilvae (GenBank accession no. AJ620501) and Inanidrilus exumae (GenBank accession no. FM202060), for which no symbiont genomes are available.
ANNOUNCEMENT
Gutless phallodriline annelids are a species-rich, monophyletic group of marine oligochaetes that lack a digestive system (1). Instead, they rely on phylogenetically diverse bacterial symbionts that provide them with nutrition via chemosynthesis (2–5). Symbionts of different host species show host specificity but often form closely related bacterial clades. To date, none of the symbionts has been cultivated, but comparative genomic analyses have been performed using high-throughput metagenomic sequencing (3, 6). Here, we present two draft genomes of a deltaproteobacterial symbiont from the host Olavius algarvensis, termed OalgDelta3. OalgDelta3 is closely related to uncultured Delta3 endosymbionts from two other gutless phallodrilinids (6, 7), namely, Olavius ilvae and Inanidrilus exumae, sharing 98% sequence identity with their 16S rRNA genes (GenBank accession no. AJ620501 and FM202060, respectively), and to deltaproteobacteria (SEEP-SRB1) that form consortia with anaerobic methane oxidizers in anoxic marine sediments (8, 9).
Two O. algarvensis specimens were collected from Elba, Italy (specimen B2SA from Sant’Andrea [42°48'26.00"N, 10°8'28.00"E] in April 2013 and specimen A1CA from Cavoli [42°44'4.15"N, 10°11'10.47"E] in June 2014), as described previously (4), and stored at 4°C in RNAlater (Ambion Life Technologies). DNA was extracted from individual specimens using the Qiagen blood and tissue kit. Two separate DNA libraries were prepared from mechanically sheared (Covaris LE220), size-selected (using Beckman Coulter solid-phase reversible immobilization beads targeting 300 bp) total DNA using the KAPA library creation kit for Illumina platforms (Kapa Biosystems) and underwent paired-end sequencing on the Illumina HiSeq 2000 platform (2 × 150 bp) at the DOE Joint Genome Institute. A total of 53,614,071 and 36,269,211 raw read pairs were obtained from B2SA and A1CA, respectively.
Reads were quality controlled with BBDuk (minimum kmer, 11; minimum length, 36 bp; minimum Phred quality score, 2) of the BBMap toolkit v36.86 (https://sourceforge.net/projects/bbmap) and BayesHammer implemented in SPAdes v3.9.1 (10, 11), and 53,474,255 (B2SA) and 36,195,816 (A1CA) clean read pairs were separately assembled with MEGAHIT v1.0.6 (12). Metagenome-assembled genomes (MAGs) were obtained using MetaBAT v0.26.3 (13) and refined using Bandage v0.08.1 (14). Genome completeness and contamination were estimated using CheckM v1.0.5 (15). Average nucleotide identity (ANI) was calculated with enveomics tools (16). Genome annotation and analysis were performed with RASTtk (17–19) and Pathway Tools v21.0 (20). Default parameters were used for all software unless otherwise noted.
MAGs were assigned as OalgDelta3 by identifying the previously reported 16S rRNA gene sequence (GenBank accession no. AM493254) with >99.5% identity. The OalgDelta3 MAG from B2SA has 5.58 Mbp (401 contigs, with an N50 value of 76,958 bp and the largest contig of 424,721 bp; GC content, 54.2%; genome coverage, 50×) and contains 5,795 protein-coding sequences (CDSs) and 43 tRNA-coding regions. The OalgDelta3 MAG from A1CA has 5.74 Mbp (350 contigs, with an N50 value of 89,163 bp and the largest contig of 348,870 bp; GC content, 54.1%; genome coverage, 350×) and includes 5,876 CDSs and 45 tRNA-coding regions. The two MAGs share 99.86% identical 16S rRNA gene sequences, with a genome-wide ANI of 99.87%. Both MAGs have 94.4% estimated completeness, with no (B2SA) or very low (A1CA; 0.03%) contamination estimates, and conform to the MIMAG standards for high-quality draft genomes (21). Similar to other deltaproteobacterial symbionts of O. algarvensis, OalgDelta3 harbors pathways for sulfate reduction, carbon monoxide and H2 oxidation, utilization of sugars, short-chain fatty acids and oligopeptides, and a multitude of associated ATP-binding cassette (ABC) transporters, tripartite ATP-independent periplasmic (TRAP)-type transporters, and other transporters.
Data availability.
The MAGs of the OalgDelta3 endosymbiont have been deposited in the European Nucleotide Archive under accession no. PRJEB28157 (assemblies GCA_903231395 and GCA_903231505), using the data brokerage service of the German Federation for Biological Data (GFBio) (22), in compliance with the Minimal Information about any (x) Sequence (MIxS) standard (23). The raw sequences are available at the Sequence Read Archive (SRA) under accession no. SRX2712534 and SRX2554373.
ACKNOWLEDGMENTS
This work was supported by the Max Planck Society, a Moore Foundation Marine Microbial Initiative Investigator Award to N.D. (grant GBMF3811), the North Carolina State Chancellor’s Faculty Excellence Program Cluster on Microbiomes and Complex Microbial Communities (M.K.), the USDA National Institute of Food and Agriculture Hatch Project 1014212 (M.K.), and the European Union’s Horizon 2020 research and innovation program under Marie Skłodowska-Curie grant agreement 660280 (C.W.). Sequencing was conducted by the U.S. DOE Joint Genome Institute, a DOE Office of Science User Facility, and was supported under contract DE-AC02-05CH11231.
REFERENCES
- 1.Dubilier N, Blazejak A, Ruehland C (2005) Symbioses between bacteria and gutless marine oligochaetes, p 251–275. In Overmann J. (ed), Molecular basis of symbiosis. progress in molecular and subcellular biology, vol 41 Springer, Berlin, Germany. [DOI] [PubMed] [Google Scholar]
- 2.Dubilier N, Mülders C, Ferdelman T, de Beer D, Pernthaler A, Klein M, Wagner M, Erséus C, Thiermann F, Krieger J, Giere O, Amann R. 2001. Endosymbiotic sulphate-reducing and sulphide-oxidizing bacteria in an oligochaete worm. Nature 411:298–302. doi: 10.1038/35077067. [DOI] [PubMed] [Google Scholar]
- 3.Woyke T, Teeling H, Ivanova NN, Huntemann M, Richter M, Gloeckner FO, Boffelli D, Anderson IJ, Barry KW, Shapiro HJ, Szeto E, Kyrpides NC, Mussmann M, Amann R, Bergin C, Ruehland C, Rubin EM, Dubilier N. 2006. Symbiosis insights through metagenomic analysis of a microbial consortium. Nature 443:950–955. doi: 10.1038/nature05192. [DOI] [PubMed] [Google Scholar]
- 4.Kleiner M, Wentrup C, Lott C, Teeling H, Wetzel S, Young J, Chang YJ, Shah M, VerBerkmoes NC, Zarzycki J, Fuchs G, Markert S, Hempel K, Voigt B, Becher D, Liebeke M, Lalk M, Albrecht D, Hecker M, Schweder T, Dubilier N. 2012. Metaproteomics of a gutless marine worm and its symbiotic microbial community reveal unusual pathways for carbon and energy use. Proc Natl Acad Sci U S A 109:E1173–E1182. doi: 10.1073/pnas.1121198109. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Wippler J, Kleiner M, Lott C, Gruhl A, Abraham PE, Giannone RJ, Young JC, Hettich RL, Dubilier N. 2016. Transcriptomic and proteomic insights into innate immunity and adaptations to a symbiotic lifestyle in the gutless marine worm Olavius algarvensis. BMC Genomics 17:942. doi: 10.1186/s12864-016-3293-y. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Kleiner M, Woyke T, Ruehland C, Dubilier N. 2011. The Olavius algarvensis metagenome revisited: lessons learned from the analysis of the low-diversity microbial consortium of a gutless marine worm, p 319–333. In de Brujin FJ. (ed), Handbook of molecular microbial ecology II: metagenomics in different habitats. John Wiley & Sons, Hoboken, NJ. [Google Scholar]
- 7.Bergin C. 2009. Phylogenetic diversity and metabolic versatility of the bacterial endosymbionts in marine gutless oligochaete worms. Ph.D. thesis University of Bremen, Bremen, Germany. [Google Scholar]
- 8.Schreiber L, Holler T, Knittel K, Meyerdierks A, Amann R. 2010. Identification of the dominant sulfate-reducing bacterial partner of anaerobic methanotrophs of the ANME-2 clade. Environ Microbiol 12:2327–2340. doi: 10.1111/j.1462-2920.2010.02275.x. [DOI] [PubMed] [Google Scholar]
- 9.Kleindienst S, Ramette A, Amann R, Knittel K. 2012. Distribution and in situ abundance of sulfate-reducing bacteria in diverse marine hydrocarbon seep sediments. Environ Microbiol 14:2689–2710. doi: 10.1111/j.1462-2920.2012.02832.x. [DOI] [PubMed] [Google Scholar]
- 10.Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, Lesin VM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin AV, Vyahhi N, Tesler G, Alekseyev MA, Pevzner PA. 2012. SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol 19:455–477. doi: 10.1089/cmb.2012.0021. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Nikolenko SI, Korobeynikov AI, Alekseyev MA. 2013. BayesHammer: Bayesian clustering for error correction in single-cell sequencing. BMC Genomics 14:S7. doi: 10.1186/1471-2164-14-S1-S7. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Li D, Luo R, Liu CM, Leung CM, Ting HF, Sadakane K, Yamashita H, Lam TW. 2016. MEGAHIT v1.0: a fast and scalable metagenome assembler driven by advanced methodologies and community practices. Methods 102:3–11. doi: 10.1016/j.ymeth.2016.02.020. [DOI] [PubMed] [Google Scholar]
- 13.Kang DD, Froula J, Egan R, Wang Z. 2015. MetaBAT, an efficient tool for accurately reconstructing single genomes from complex microbial communities. PeerJ 3:e1165. doi: 10.7717/peerj.1165. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.Wick RR, Schultz MB, Zobel J, Holt KE. 2015. Bandage: interactive visualization of de novo genome assemblies. Bioinformatics 31:3350–3352. doi: 10.1093/bioinformatics/btv383. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15.Parks DH, Imelfort M, Skennerton CT, Hugenholtz P, Tyson GW. 2015. CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes. Genome Res 25:1043–1055. doi: 10.1101/gr.186072.114. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Rodriguez-R LM, Konstantinidis KT. 2016. The enveomics collection: a toolbox for specialized analyses of microbial genomes and metagenomes. PeerJ Prepr 4:e1900v1. doi: 10.7287/peerj.preprints.1900v1. [DOI] [Google Scholar]
- 17.Aziz RK, Bartels D, Best AA, DeJongh M, Disz T, Edwards RA, Formsma K, Gerdes S, Glass EM, Kubal M, Meyer F, Olsen GJ, Olson R, Osterman AL, Overbeek RA, McNeil LK, Paarmann D, Paczian T, Parrello B, Pusch GD, Reich C, Stevens R, Vassieva O, Vonstein V, Wilke A, Zagnitko O. 2008. The RAST server: Rapid Annotations using Subsystems Technology. BMC Genomics 9:75. doi: 10.1186/1471-2164-9-75. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 18.Overbeek R, Olson R, Pusch GD, Olsen GJ, Davis JJ, Disz T, Edwards RA, Gerdes S, Parrello B, Shukla M, Vonstein V, Wattam AR, Xia F, Stevens R. 2014. The SEED and the Rapid Annotation of microbial genomes using Subsystems Technology (RAST). Nucleic Acids Res 42:D206–D214. doi: 10.1093/nar/gkt1226. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 19.Brettin T, Davis JJ, Disz T, Edwards RA, Gerdes S, Olsen GJ, Olson R, Overbeek R, Parrello B, Pusch GD, Shukla M, Thomason JA, Stevens R, Vonstein V, Wattam AR, Xia F. 2015. RASTtk: a modular and extensible implementation of the RAST algorithm for building custom annotation pipelines and annotating batches of genomes. Sci Rep 5:8365. doi: 10.1038/srep08365. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 20.Karp PD, Latendresse M, Paley SM, Krummenacker M, Ong QD, Billington R, Kothari A, Weaver D, Lee T, Subhraveti P, Spaulding A, Fulcher C, Keseler IM, Caspi R. 2016. Pathway Tools version 19.0 update: software for pathway/genome informatics and systems biology. Brief Bioinform 17:877–890. doi: 10.1093/bib/bbv079. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 21.Bowers RM, Kyrpides NC, Stepanauskas R, Harmon-Smith M, Doud D, Reddy TBK, Schulz F, Jarett J, Rivers AR, Eloe-Fadrosh EA, Tringe SG, Ivanova NN, Copeland A, Clum A, Becraft ED, Malmstrom RR, Birren B, Podar M, Bork P, Weinstock GM, Garrity GM, Dodsworth JA, Yooseph S, Sutton G, Glöckner FO, Gilbert JA, Nelson WC, Hallam SJ, Jungbluth SP, Ettema TJG, Tighe S, Konstantinidis KT, Liu WT, Baker BJ, Rattei T, Eisen JA, Hedlund B, McMahon KD, Fierer N, Knight R, Finn R, Cochrane G, Karsch-Mizrachi I, Tyson GW, Rinke C, Lapidus A, Meyer F, Yilmaz P, Parks DH, Eren AM, Schriml L, Banfield JF, Hugenholtz P, Woyke T, Genome Standards Consortium . 2017. Minimum information about a single amplified genome (MISAG) and a metagenome-assembled genome (MIMAG) of bacteria and archaea. Nat Biotechnol 35:725–731. doi: 10.1038/nbt.3893. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 22.Diepenbroek M, Glöckner F, Grobe P, Güntsch A, Huber R, König-Ries B, Kostadinov I, Nieschulze J, Seeger B, Tolksdorf R, Triebel D. 2014. Towards an integrated biodiversity and ecological research data management and archiving platform: the German Federation for the Curation of Biological Data (GFBio), p 1711–1724. In Plödereder E, Grunske L, Schneider E, Ull D (ed), Informatik 2014: Big Data Komplexität meistern. GI ed.: Lecture Notes in Informatics (LNI) Proceedings 232 Köllen Verlag, Bonn, Germany. [Google Scholar]
- 23.Yilmaz P, Kottmann R, Field D, Knight R, Cole JR, Amaral-Zettler L, Gilbert JA, Karsch-Mizrachi I, Johnston A, Cochrane G, Vaughan R, Hunter C, Park J, Morrison N, Rocca-Serra P, Sterk P, Arumugam M, Bailey M, Baumgartner L, Birren BW, Blaser MJ, Bonazzi V, Booth T, Bork P, Bushman FD, Buttigieg PL, Chain PS, Charlson E, Costello EK, Huot-Creasy H, Dawyndt P, DeSantis T, Fierer N, Fuhrman JA, Gallery RE, Gevers D, Gibbs RA, San Gil I, Gonzalez A, Gordon JI, Guralnick R, Hankeln W, Highlander S, Hugenholtz P, Jansson J, Kau AL, Kelley ST, Kennedy J, Knights D, Koren O, Kuczynski J, Kyrpides N, Larsen R, Lauber CL, Legg T, Ley RE, Lozupone CA, Ludwig W, Lyons D, Maguire E, Methé BA, Meyer F, Muegge B, Nakielny S, Nelson KE, Nemergut D, Neufeld JD, Newbold LK, Oliver AE, Pace NR, Palanisamy G, Peplies J, Petrosino J, Proctor L, Pruesse E, Quast C, Raes J, Ratnasingham S, Ravel J, Relman DA, Assunta-Sansone S, Schloss PD, Schriml L, Sinha R, Smith MI, Sodergren E, Spo A, Stombaugh J, Tiedje JM, Ward DV, Weinstock GM, Wendel D, White O, Whiteley A, Wilke A, Wortman JR, Yatsunenko T, Glöckner FO. 2011. Minimum information about a marker gene sequence (MIMARKS) and minimum information about any (x) sequence (MIxS) specifications. Nat Biotechnol 29:415–420. doi: 10.1038/nbt.1823. [DOI] [PMC free article] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The MAGs of the OalgDelta3 endosymbiont have been deposited in the European Nucleotide Archive under accession no. PRJEB28157 (assemblies GCA_903231395 and GCA_903231505), using the data brokerage service of the German Federation for Biological Data (GFBio) (22), in compliance with the Minimal Information about any (x) Sequence (MIxS) standard (23). The raw sequences are available at the Sequence Read Archive (SRA) under accession no. SRX2712534 and SRX2554373.