Here, we report the draft genome sequence of Novosphingobium sp. strain B 225, an iron-depositing bacterium isolated from a phenazone-amended naturally grown biofilm. This biofilm was grown in the Unteres Odertal National Park, Germany. Illumina NextSeq sequencing was used to determine the genome of the strain.DFG, 325093850, Open Access Publizieren 2017 - 2018 / Technische Universität Berli

Braun, Burga

Künzel, Sven

Szewzyka, Ulrich

Genome Announcements

Here, we report the draft genome sequence of Novosphingobium sp. strain B 225, an iron-depositing bacterium isolated from a phenazone-amended naturally grown biofilm. This biofilm was grown in the Unteres Odertal National Park, Germany. Illumina NextSeq sequencing was used to determine the genome of the strain. © 2018 Braun et al

Braun, B.

Künzel, S.

Szewzyk, U.

MPG.PuRe

Draft Genome Sequence of Strain B 225, an Iron-DepositingIsolate of the Genus NovosphingobiumBurga Braun,a Sven Künzel,b Ulrich SzewzykaaTechnische Universität Berlin, Berlin, GermanybMax Planck Institute for Evolutionary Biology, Plön, GermanyABSTRACT Here, we report the draft genome sequence of Novosphingobium sp.strain B 225, an iron-depositing bacterium isolated from a phenazone-amended nat-urally grown biofilm. This biofilm was grown in the Unteres Odertal National Park,Germany. Illumina NextSeq sequencing was used to determine the genome of thestrain.Wastewater effluent poses a potential risk to aquatic ecosystems due to persis-tent organic contaminants which are not eliminated during water treatment.Microbial activity plays a major role in the degradation of environmental pollutants,and members of the genus Novosphingobium have been previously described ascontaminant-degrading organisms (1–3).Based on 16S rRNA gene sequence similarity, strain B 225 belongs to the genusNovosphingobium and shows the highest level of similarity to two Novosphingobiumstrains, namely, Novosphingobium hassiacum strain W-51 (98%) and Novosphingobiumaromaticivorans DSM 12444 (98%), based on BLASTn searches (4), and 97.6% similarityto Novosphingobium ginsenosidimutans by EzBioCloud (5) searches.The Gram-negative rod-shaped Novosphingobium strain B 225 was isolated from a4-month-old iron- and manganese-depositing biofilm grown in a continuous-flowbiofilm reactor exposed in the Bogengraben River of the Unteres Odertal National Park,Germany. The reactor was continuously fed with river water and phenazone (0.5 gliter1). The biofilm was spread on ATA medium [2 g liter1 MnCO3 hydrate, 0.15 gml1 trisodium citrate, 0.2 g liter1 Fe(NH4)2(SO4)2, 0.1 g liter1 cycloheximide, 2 mlvitamin solution (6), 2 ml trace element solution SL 9 (7), 0.94 g phenazone, 20 g agar,and 1 liter filtered water from Lake Daminke] based on Mn agar (8) and incubated for2 weeks at room temperature. Small dark-brown colonies were selected, and their ironand manganese deposition abilities were confirmed according to Schmidt et al. (9).Genomic DNA was extracted using the GeneMATRIX soil DNA purification kit(Roboklon, Berlin, Germany). The paired-end library was prepared by using the TruSeqDNA high-throughput (HT) sample prep kit (Illumina Netherlands, Eindhoven, TheNetherlands), and mate-pair libraries were established with the Nextera mate-pairsample preparation kit (Illumina Netherlands). The paired-end library was preparedfollowing the Illumina Nextera XT DNA library prep kit protocol. Genome sequencingwas done on an Illumina NextSeq 500 sequencer using a NextSeq mid-output kitversion 2 with 300-cycle chemistry by generating 10,292,075 raw reads. Demultiplexingwas done with bcl2fastq version 2.18.0.12, and quality filtering of raw reads wasperformed using Trimmomatic version 0.36 (10). The reads were then checked forambiguous base calls and low complexity, employing the DUST algorithm (11), andfiltered accordingly with an R script in Microsoft R Open version 3.3.2 (http://www.r-project.org/), followed by preassembly with SPAdes version 3.10.0 (12) using defaultk-mer lengths up to 99 bp. Scaffolds 500 bp of this preassembly were subject toextension and second-round scaffolding with the SSPACE standard version 3.0 (13).Received 15 March 2018 Accepted 20 March2018 Published 12 April 2018Citation Braun B, Künzel S, Szewzyk U. 2018.Draft genome sequence of strain B 225, aniron-depositing isolate of the genusNovosphingobium. Genome Announc 6:e00325-18. https://doi.org/10.1128/genomeA.00325-18.Copyright © 2018 Braun et al. This is an open-access article distributed under the terms ofthe Creative Commons Attribution 4.0International license.Address correspondence to Burga Braun,Burga.braun@tu-berlin.de.PROKARYOTEScrossmVolume 6 Issue 15 e00325-18 genomea.asm.org 1 on June 22, 2018 by MAX PLANCK INST F EVOLUTIONSBIOLOGIEhttp://genomea.asm.org/Downloaded from Scaffolds 2,500 bp were assigned to genome bins by MetaBAT version 0.32.4 (14), andfunctional annotation of the draft genomes was performed with Prokka version 1.12b(15).The draft genome included 73 contigs, with an N50 assembly quality of 107,389 bpand an L50 value of 11 bp. The shortest sequence was 2,912 bp, and the longestsequence was 400,854 bp. The total size of the draft genome was 3,712,107 bp, with aGC content of 64%. Annotation included 3,500 coding regions for 3,561 genes, 463signal peptides, 3 rRNAs (16S and 23S), 49 tRNAs, 1 transfer-messenger RNA (tmRNA),and 8 miscellaneous RNAs (miscRNAs).Accession number(s). This whole-genome shotgun project has been deposited atDDBJ/ENA/GenBank under the accession number MWLM00000000. The version de-scribed in this paper is version MWLM01000000.ACKNOWLEDGMENTSBioinformatic consulting was provided by omics2view.consulting GbR (Kiel, Germany).This work was supported by a grant of the Bundesministerium für Bildung undForschung (BMBF), 02WU0715.REFERENCES1. Pieper C, Risse D, Schmidt B, Braun B, Szewzyk U, Rotard W. 2010.Investigation of the microbial degradation of phenazone-type drugs andtheir metabolites by natural biofilms derived from river water usingliquid chromatography/tandem mass spectrometry (LC-MS/MS). WaterRes 44:4559 – 4569. https://doi.org/10.1016/j.watres.2010.05.028.2. Sohn JH, Kwon KK, Kang JH, Jung HB, Kim SJ. 2004. Novosphingobiumpentaromativorans sp. nov., a high-molecular-mass polycyclic aro-matic hydrocarbon-degrading bacterium isolated from estuarine sed-iment. Int J Syst Evol Microbiol 54:1483–1487. https://doi.org/10.1099/ijs.0.02945-0.3. Yuan J, Lai Q, Zheng T, Shao Z. 2009. Novosphingobium indicum sp. nov.,a polycyclic aromatic hydrocarbon-degrading bacterium isolated from adeep-sea environment. Int J Syst Evol Microbiol 59:2084 –2088. https://doi.org/10.1099/ijs.0.002873-0.4. Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, LipmanDJ. 1997. Gapped BLAST and PSI-BLAST: a new generation of proteindatabase search programs. Nucleic Acids Res 25:3389 –3402. https://doi.org/10.1093/nar/25.17.3389.5. Yoon S, Ha S, Kwon S, Lim J, Kim Y, Seo H, Chun J. 2017. IntroducingEzBioCloud: a taxonomically united database of 16S rRNA gene se-quences and whole-genome assemblies. Int J Syst Evol Microbiol 67:1613–1617. https://doi.org/10.1099/ijsem.0.001755.6. Schlegel HG, Zaborosch C. 1992. Allgemeine Mikrobiologie Thieme Ver-lag, Stuttgart, Germany.7. Tschech A, Pfennig N. 1984. Growth yield increase linked to caffeatereduction in Acetobacterium woodii. Arch Microbiol 137:163–167. https://doi.org/10.1007/BF00414460.8. Mulder EG, Van Veen W. 1963. Investigations on the Sphaerotilus-Leptothrix group. Antonie Van Leeuwenhoek 29:121–153. https://doi.org/10.1007/BF02046045.9. Schmidt B, Sánchez LA, Fretschner T, Kreps G, Ferrero MA, Siñeriz F,Szewzyk U. 2014. Isolation of Sphaerotilus-Leptothrix strains from ironbacteria communities in Tierra del Fuego wetlands. FEMS Microbiol Ecol90:454 – 466. https://doi.org/10.1111/1574-6941.12406.10. Bolger AM, Lohse M, Usadel B. 2014. Trimmomatic: a flexible trimmer forIllumina sequence data. Bioinformatics 30:2114 –2120. https://doi.org/10.1093/bioinformatics/btu170.11. Morgulis A, Gertz EM, Schäffer AA, Agarwala R. 2006. A fast and sym-metric DUST implementation to mask low-complexity DNA sequences. JComput Biol 13:1028 –1040. https://doi.org/10.1089/cmb.2006.13.1028.12. 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 newgenome assembly algorithm and its applications to single-cell sequenc-ing. J Comput Biol 19:455– 477. https://doi.org/10.1089/cmb.2012.0021.13. Boetzer M, Henkel CV, Jansen HJ, Butler D, Pirovano W. 2011. Scaf-folding pre-assembled contigs using SSPACE. Bioinformatics 27:578 –579. https://doi.org/10.1093/bioinformatics/btq683.14. Kang DD, Froula J, Egan R. 2014. A robust statistical framework forreconstructing genomes from metagenomic data. bioRxiv https://doi.org/10.1101/011460.15. Seemann T. 2014. Prokka: rapid prokaryotic genome annotation. Bioin-formatics 30:2068 –2069. https://doi.org/10.1093/bioinformatics/btu153.Braun et al.Volume 6 Issue 15 e00325-18 genomea.asm.org 2 on June 22, 2018 by MAX PLANCK INST F EVOLUTIONSBIOLOGIEhttp://genomea.asm.org/Downloaded from 

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Draft genome sequence of strain B 225, an iron-depositing isolate of the genus Novosphingobium

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Draft Genome Sequence of Strain B 225, an Iron-DepositingIsolate of the Genus NovosphingobiumBurga Braun,a Sven Künzel,b Ulrich SzewzykaaTechnische Universität Berlin, Berlin, GermanybMax Planck Institute for Evolutionary Biology, Plön, GermanyABSTRACT Here, we report the draft genome sequence of Novosphingobium sp.strain B 225, an iron-depositing bacterium isolated from a phenazone-amended nat-urally grown bioﬁlm. This bioﬁlm was grown in the Unteres Odertal National Park,Germany. Illumina NextSeq sequencing was used to determine the genome of thestrain.Wastewater efﬂuent poses a potential risk to aquatic ecosystems due to persis-tent organic contaminants which are not eliminated during water treatment.Microbial activity plays a major role in the degradation of environmental pollutants,and members of the genus Novosphingobium have been previously described ascontaminant-degrading organisms (1–3).Based on 16S rRNA gene sequence similarity, strain B 225 belongs to the genusNovosphingobium and shows the highest level of similarity to two Novosphingobiumstrains, namely, Novosphingobium hassiacum strain W-51 (98%) and Novosphingobiumaromaticivorans DSM 12444 (98%), based on BLASTn searches (4), and 97.6% similarityto Novosphingobium ginsenosidimutans by EzBioCloud (5) searches.The Gram-negative rod-shaped Novosphingobium strain B 225 was isolated from a4-month-old iron- and manganese-depositing bioﬁlm grown in a continuous-ﬂowbioﬁlm reactor exposed in the Bogengraben River of the Unteres Odertal National Park,Germany. The reactor was continuously fed with river water and phenazone (0.5 gliter1). The bioﬁlm was spread on ATA medium [2 g liter1 MnCO3 hydrate, 0.15 gml1 trisodium citrate, 0.2 g liter1 Fe(NH4)2(SO4)2, 0.1 g liter1 cycloheximide, 2 mlvitamin solution (6), 2 ml trace element solution SL 9 (7), 0.94 g phenazone, 20 g agar,and 1 liter ﬁltered water from Lake Daminke] based on Mn agar (8) and incubated for2 weeks at room temperature. Small dark-brown colonies were selected, and their ironand manganese deposition abilities were conﬁrmed according to Schmidt et al. (9).Genomic DNA was extracted using the GeneMATRIX soil DNA puriﬁcation kit(Roboklon, Berlin, Germany). The paired-end library was prepared by using the TruSeqDNA high-throughput (HT) sample prep kit (Illumina Netherlands, Eindhoven, TheNetherlands), and mate-pair libraries were established with the Nextera mate-pairsample preparation kit (Illumina Netherlands). The paired-end library was preparedfollowing the Illumina Nextera XT DNA library prep kit protocol. Genome sequencingwas done on an Illumina NextSeq 500 sequencer using a NextSeq mid-output kitversion 2 with 300-cycle chemistry by generating 10,292,075 raw reads. Demultiplexingwas done with bcl2fastq version 2.18.0.12, and quality ﬁltering of raw reads wasperformed using Trimmomatic version 0.36 (10). The reads were then checked forambiguous base calls and low complexity, employing the DUST algorithm (11), andﬁltered accordingly with an R script in Microsoft R Open version 3.3.2 (http://www.r-project.org/), followed by preassembly with SPAdes version 3.10.0 (12) using defaultk-mer lengths up to 99 bp. Scaffolds 500 bp of this preassembly were subject toextension and second-round scaffolding with the SSPACE standard version 3.0 (13).Received 15 March 2018 Accepted 20 March2018 Published 12 April 2018Citation Braun B, Künzel S, Szewzyk U. 2018.Draft genome sequence of strain B 225, aniron-depositing isolate of the genusNovosphingobium. Genome Announc 6:e00325-18. https://doi.org/10.1128/genomeA.00325-18.Copyright © 2018 Braun et al. This is an open-access article distributed under the terms ofthe Creative Commons Attribution 4.0International license.Address correspondence to Burga Braun,Burga.braun@tu-berlin.de.PROKARYOTEScrossmVolume 6 Issue 15 e00325-18 genomea.asm.org 1 on April 27, 2018 by guesthttp://genomea.asm.org/Downloaded from Scaffolds2,500 bp were assigned to genome bins by MetaBAT version 0.32.4 (14), andfunctional annotation of the draft genomes was performed with Prokka version 1.12b(15).The draft genome included 73 contigs, with an N50 assembly quality of 107,389 bpand an L50 value of 11 bp. The shortest sequence was 2,912 bp, and the longestsequence was 400,854 bp. The total size of the draft genome was 3,712,107 bp, with aGC content of 64%. Annotation included 3,500 coding regions for 3,561 genes, 463signal peptides, 3 rRNAs (16S and 23S), 49 tRNAs, 1 transfer-messenger RNA (tmRNA),and 8 miscellaneous RNAs (miscRNAs).Accession number(s). This whole-genome shotgun project has been deposited atDDBJ/ENA/GenBank under the accession number MWLM00000000. The version de-scribed in this paper is version MWLM01000000.ACKNOWLEDGMENTSBioinformatic consulting was provided by omics2view.consulting GbR (Kiel, Germany).This work was supported by a grant of the Bundesministerium für Bildung undForschung (BMBF), 02WU0715.REFERENCES1. Pieper C, Risse D, Schmidt B, Braun B, Szewzyk U, Rotard W. 2010.Investigation of the microbial degradation of phenazone-type drugs andtheir metabolites by natural bioﬁlms derived from river water usingliquid chromatography/tandem mass spectrometry (LC-MS/MS). WaterRes 44:4559–4569. https://doi.org/10.1016/j.watres.2010.05.028.2. Sohn JH, Kwon KK, Kang JH, Jung HB, Kim SJ. 2004. Novosphingobiumpentaromativorans sp. nov., a high-molecular-mass polycyclic aro-matic hydrocarbon-degrading bacterium isolated from estuarine sed-iment. Int J Syst Evol Microbiol 54:1483–1487. https://doi.org/10.1099/ijs.0.02945-0.3. Yuan J, Lai Q, Zheng T, Shao Z. 2009. Novosphingobium indicum sp. nov.,a polycyclic aromatic hydrocarbon-degrading bacterium isolated from adeep-sea environment. Int J Syst Evol Microbiol 59:2084–2088. https://doi.org/10.1099/ijs.0.002873-0.4. Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, LipmanDJ. 1997. Gapped BLAST and PSI-BLAST: a new generation of proteindatabase search programs. Nucleic Acids Res 25:3389–3402. https://doi.org/10.1093/nar/25.17.3389.5. Yoon S, Ha S, Kwon S, Lim J, Kim Y, Seo H, Chun J. 2017. IntroducingEzBioCloud: a taxonomically united database of 16S rRNA gene se-quences and whole-genome assemblies. Int J Syst Evol Microbiol 67:1613–1617. https://doi.org/10.1099/ijsem.0.001755.6. Schlegel HG, Zaborosch C. 1992. Allgemeine Mikrobiologie Thieme Ver-lag, Stuttgart, Germany.7. Tschech A, Pfennig N. 1984. Growth yield increase linked to caffeatereduction in Acetobacterium woodii. Arch Microbiol 137:163–167. https://doi.org/10.1007/BF00414460.8. Mulder EG, Van Veen W. 1963. Investigations on the Sphaerotilus-Leptothrix group. Antonie Van Leeuwenhoek 29:121–153. https://doi.org/10.1007/BF02046045.9. Schmidt B, Sánchez LA, Fretschner T, Kreps G, Ferrero MA, Siñeriz F,Szewzyk U. 2014. Isolation of Sphaerotilus-Leptothrix strains from ironbacteria communities in Tierra del Fuego wetlands. FEMS Microbiol Ecol90:454–466. https://doi.org/10.1111/1574-6941.12406.10. Bolger AM, Lohse M, Usadel B. 2014. Trimmomatic: a ﬂexible trimmer forIllumina sequence data. Bioinformatics 30:2114–2120. https://doi.org/10.1093/bioinformatics/btu170.11. Morgulis A, Gertz EM, Schäffer AA, Agarwala R. 2006. A fast and sym-metric DUST implementation to mask low-complexity DNA sequences. JComput Biol 13:1028–1040. https://doi.org/10.1089/cmb.2006.13.1028.12. 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 newgenome assembly algorithm and its applications to single-cell sequenc-ing. J Comput Biol 19:455–477. https://doi.org/10.1089/cmb.2012.0021.13. Boetzer M, Henkel CV, Jansen HJ, Butler D, Pirovano W. 2011. Scaf-folding pre-assembled contigs using SSPACE. Bioinformatics 27:578–579. https://doi.org/10.1093/bioinformatics/btq683.14. Kang DD, Froula J, Egan R. 2014. A robust statistical framework forreconstructing genomes from metagenomic data. bioRxiv https://doi.org/10.1101/011460.15. Seemann T. 2014. Prokka: rapid prokaryotic genome annotation. Bioin-formatics 30:2068–2069. https://doi.org/10.1093/bioinformatics/btu153.Braun et al.Volume 6 Issue 15 e00325-18 genomea.asm.org 2 on April 27, 2018 by guesthttp://genomea.asm.org/Downloaded from 

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Draft genome sequence of strain B 225, an iron-depositing isolate of the genus Novosphingobium

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