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Ignisphaera Aggregans Type Strain (AQ1.S1T)

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Ignisphaera Aggregans Type Strain (AQ1.S1T) Standards in Genomic Sciences (2010) 3:66-75 DOI:10.4056/sigs.1072907 Complete genome sequence of Ignisphaera aggregans type strain (AQ1.S1T) Markus Göker1, Brittany Held2,4, Alla Lapidus2, Matt Nolan2, Stefan Spring1, Montri Yasawong3, Susan Lucas2, Tijana Glavina Del Rio2, Hope Tice2, Jan-Fang Cheng2, Lynne Goodwin2,4, Roxanne Tapia2,4, Sam Pitluck2, Konstantinos Liolios2, Natalia Ivanova2, Konstantinos Mavromatis2, Natalia Mikhailova2, Amrita Pati2, Amy Chen5, Krishna Palaniap- pan5, Evelyne Brambilla1, Miriam Land2,6, Loren Hauser2,6, Yun-Juan Chang2,6, Cynthia D. Jeffries2,6, Thomas Brettin2, John C. Detter2, Cliff Han2, Manfred Rohde3, Johannes Sikorski1, Tanja Woyke2, James Bristow2, Jonathan A. Eisen2,7, Victor Markowitz5, Philip Hugenholtz2, Nikos C. Kyrpides2, and Hans-Peter Klenk1* 1 DSMZ - German Collection of Microorganisms and Cell Cultures GmbH, Braunschweig, Germany 2 DOE Joint Genome Institute, Walnut Creek, California, USA 3 HZI – Helmholtz Centre for Infection Research, Braunschweig, Germany 4 Los Alamos National Laboratory, Bioscience Division, Los Alamos, New Mexico, USA 5 Biological Data Management and Technology Center, Lawrence Berkeley National Laboratory, Berkeley, California, USA 6 Oak Ridge National Laboratory, Oak Ridge, Tennessee, USA 7 University of California Davis Genome Center, Davis, California, USA *Corresponding author: Hans-Peter Klenk Keywords: hyperthermophile, obligately anaerobic, moderately acidophilic, fermentative, cocci-shaped, hot spring, Crenarchaeota, Desulfurococcaceae, GEBA Ignisphaera aggregans Niederberger et al. 2006 is the type and sole species of genus Ignis- phaera. This archaeal species is characterized by a coccoid-shape and is strictly anaerobic, moderately acidophilic, heterotrophic hyperthermophilic and fermentative. The type strain AQ1.S1T was isolated from a near neutral, boiling spring in Kuirau Park, Rotorua, New Zeal- and. This is the first completed genome sequence of the genus Ignisphaera and the fifth ge- nome (fourth type strain) sequence in the family Desulfurococcaceae. The 1,875,953 bp long genome with its 2,009 protein-coding and 52 RNA genes is a part of the Genomic Encyclo- pedia of Bacteria and Archaea project. Introduction Strain AQ1.S1T (= DSM 17230 = JCM 13409) is the number of polysaccharides and complex proteina- type strain of the species Ignisphaera aggregans, ceous substrates [1]. Here we present a summary which is the type species of the genus Ignisphaera classification and a set of features for I. aggregans [1], one out of nine genera in the family Desulfuro- AQ1.S1T, together with the description of the coccaceae [2-5]. The generic name derives from complete genomic sequencing and annotation. the Latin word ‘ignis’ meaning ‘fire’, and ‘sphaera’ meaning ‘ball’, referring to coccoid cells found in Classification and features the high-temperature environment such as hot Strain AQ1.S1T was isolated from a near neutral, springs [1]. The species epithet is derived from boiling spring situated in Kuirau Park, Rotorua, the Latin word ‘aggregans’ meaning ‘aggregate New Zealand [1]. Interestingly, strains of I. aggre- forming or aggregating clumping’, referring to the gans could not be cultivated from pools with simi- appearance of the cells when grown on mono-, di- lar characteristics in Yellowstone National Park or polysaccharides [1]. Strain AQ1.S1T is of partic- [1]. Only three cultivated strains are reported for ular interest because it is able to ferment quite a the species I. aggregans in addition to AQ1.S1T, The Genomic Standards Consortium Ignisphaera aggregans type strain (AQ1.S1T) these are strains Tok37.S1, Tok10A.S1 and Tok1 thermophilic and grows optimally between 92°C [1]. The 16S rRNA sequence of AQ1.S1T is 99% and 95°C, the temperature range for growth is 85- identical to Tok37.S1, 98% to Tok10A.S1 and 98% 98°C. The pH range for growth is 5.4-7.0, with an to Tok1. Sequence similarities between strain optimum at pH 6.4. The strain grows in the pres- AQ1.S1T and members of the family Pyrodictiaceae ence of up to 0.5% NaCl, however, it grows opti- range from 93.0% for Pyrodictium occultum to mally without NaCl. The doubling time is 7.5 h un- 93.4% for P. abyssi [6] but from 89.7% for Ignicoc- der optimal conditions [1]. I. aggregans strain cus islandicus to 93.5% for Staphylothermus helle- AQ1.S1T is strictly anaerobic and grows hetero- nicus [6] with members of the family Desulfuro- trophically on starch, trypticase peptone, lactose, coccaceae in which I. aggregans is currently classi- glucose, konjac glucomannan, mannose, galactose, fied (Table 1). Genbank [16] currently contains -cyclodextrin. Growth on only three 16S rRNA gene sequences with signifi- beef extract and glucose is weak and not observed cantly high identity values to strain AQ1.S1T: clone onmaltose, yeast glycogen,extract, cellobiose,and β methanol, ethanol, YNP_BP_A32 (96%, DQ243730) from hot springs trehalose, pyruvate, acetate, malate, casamino ac- of Yellowstone National Park, clone SSW_L4_A01 ids (0.1% w/v), carboxymethylcellulose, amylo- (95%, EU635921) from mud hot springs, Nevada, pectin (corn), xanthan gum, locust gum (bean), USA, and clone DDP-A02 (94%, AB462559) from a guar gum, dextran, xylan (oat spelts, larch or Japanese alkaline geothermal pool, which does not birch), xylitol, xylose or amylose (corn and potato) necessarily indicate the presence of I. aggregans [1]. Mono- and disaccharides are accumulated in but probably the presence of yet to be identified AQ1.S1T cultures grown in media containing kon- other species in the genus Ignisphaera. Environ- jac glucomannan, but not in sterile media that had mental samples and metagenomic surveys fea- been exposed to the same temperature as the in- tured in Genbank contain not a single sequence oculated medium or the stock of konjac gluco- with >87% sequence identity (as of June 2010), mannan [1]. As hypothesized by Niederberger et indicating that I. aggregans might play a rather al. [1], this most probably indicates that the konjac limited and regional role in the environment. glucomannan is being hydrolyzed enzymatically by AQ1.S1T into sugars for metabolism. Removal The cells of strain AQ1.S1T are regular to irregular of cystine from the growth medium does not affect cocci which occur singly, in pairs or as aggregates cell density significantly. Hydrogen sulfide is also of many cells [1]. They usually have dimensions detected in AQ1.S1T cultures grown in enrichment between 1- Figure 1). Aggregation of cells media. Strain AQ1.S1T is resistant to novobiocin is common when AQ1.S1T is grown on mono-, di- and streptomycin but sensitive to erythromycin, or polysaccharides1.5 μm ( [1]. Strain AQ1.S1T is hyper- chloramphenicol and rifampicin [1]. Figure 1. Scanning electron micrograph of I. aggregans AQ1.S1T 67 Standards in Genomic Sciences Göker et al. Chemotaxonomy No chemotaxonomic data are currently available Genome sequencing and annotation T for I. aggregans strain AQ1.S1 . Also, chemotax- Genome project history onomic information for the family Desulfurococca- This organism was selected for sequencing on the ceae is scarce. What is known is that the type spe- basis of its phylogenetic position [26], and is part cies of this family, Desulfurococcus mucosus, lacks of the Genomic Encyclopedia of Bacteria and Arc- a murein cell wall and contains phytanol and po- haea project [27]. The genome project is depo- lyisopreonoid dialcohols as major components of sited in the Genome OnLine Database [21] and the the cellular lipids [3]. complete genome sequence is deposited in Gen- Figure 2 shows the phylogenetic neighborhood of Bank. Sequencing, finishing and annotation were I. aggregans AQ1.S1T in a 16S rRNA based tree. performed by the DOE Joint Genome Institute The sequence of the single 16S rRNA gene copy in (JGI). A summary of the project information is the genome of strain AQ1.S1 does not differ from shown in Table 2. the previously published 16S rRNA sequence from DSM 17230 (DQ060321). Figure 2. Phylogenetic tree highlighting the position of I. aggregans AQ1.S1T relative to the type strains of the other genera within the order Desulfurococcales. The tree was inferred from 1,329 aligned characters [17,18] of the 16S rRNA gene sequence under the maximum likelihood criterion [19] and rooted with the type strains of the genera of the neighboring order Acidilobales. The branches are scaled in terms of the expected number of substitutions per site. Numbers above branches are support values from 250 bootstrap replicates [20] ,if greater than 60%. Lineages with type strain genome sequencing projects registered in GOLD [21] are shown in blue, published genomes in bold ([22-25], CP000504 and CP000852). Growth conditions and DNA isolation Genome sequencing and assembly I. aggregans AQ1.S1T, DSM 17230, was grown The genome of strain AQ1.S1T was sequenced using anaerobically in DSMZ medium 1043 (Ignisphaera a combination of Illumina and 454 technologies. An medium) [28] at 92°C. DNA was isolated from 0.5- Illumina GAii shotgun library with reads of 152 Mb, 1 g of cell paste using MasterPure Gram Positive a 454 Titanium draft library with average read DNA Purification Kit (Epicentre MGP04100). One length of 320 bases, and a paired end 454 library µl lysozyme and five µl mutanolysin and lysosta- with average insert size of 15 kb were generated phine, each, were added to the standard lysis solu- for this genome. All general aspects of library con- tion for one hour at 37°C followed by 30 min incu- struction and sequencing can be found at bation on ice after the MPC-step. http://www.jgi.doe.gov/. Illumina sequencing data was assembled with VELVET and the consensus http://standardsingenomics.org 68 Ignisphaera aggregans type strain (AQ1.S1T) sequences were shredded into 1.5 kb overlapped Software, LLC). Possible mis-assemblies were cor- fake reads and assembled together with the 454 rected with gapResolution, Dupfinisher [29], or se- data. Draft assemblies were based on 177 Mb 454 quencing cloned bridging PCR fragments with sub- draft data, and 454 paired end data.
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