Open Access

Complete genome sequence of Segniliparus rotundus type strain (CDC 1076T)

  • Johannes Sikorski
  • , Alla Lapidus
  • , Alex Copeland
  • , Monica Misra,
  • , Tijana Glavina Del Rio
  • , Matt Nolan
  • , Susan Lucas
  • , Feng Chen
  • , Hope Tice
  • , Jan-Fang Cheng
  • , Marlen Jando
  • , Susanne Schneider
  • , David Bruce,
  • , Lynne Goodwin,
  • , Sam Pitluck
  • , Konstantinos Liolios
  • , Natalia Mikhailova
  • , Amrita Pati
  • , Natalia Ivanova
  • , Konstantinos Mavromatis
  • , Amy Chen
  • , Krishna Palaniappan
  • , Olga Chertkov,
  • , Miriam Land,
  • , Loren Hauser,
  • , Yun-Juan Chang,
  • , Cynthia D. Jeffries,
  • , Thomas Brettin,
  • , John C. Detter,
  • , Cliff Han,
  • , Manfred Rohde
  • , Markus Göker
  • , Jim Bristow
  • , Jonathan A. Eisen,
  • , Victor Markowitz
  • , Philip Hugenholtz
  • , Nikos C. Kyrpides
  • and Hans-Peter Klenk
Corresponding author

DOI: 10.4056/sigs.791633

Received: 30 March 2010

Published: 30 April 2010


Segniliparus rotundus Butler 2005 is the type species of the genus Segniliparus, which is currently the only genus in the corynebacterial family Segniliparaceae. This family is of large interest because of a novel late-emerging genus-specific mycolate pattern. The type strain has been isolated from human sputum and is probably an opportunistic pathogen. Here we describe the features of this organism, together with the complete genome sequence and annotation. This is the first completed genome sequence of the family Segniliparaceae. The 3,157,527 bp long genome with its 3,081 protein-coding and 52 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.


aerobicnon-sporeformingnovel mycolic acidopportunistic pathogenCorynebacterineaeGEBA


Strain CDC 1076T (= DSM 44985 = ATCC BAA-972 = JCM 13578) is the type strain of the species Segniliparus rotundus [1], which is the type species of the genus Segniliparus. Besides S. rotundus, the genus Segniliparus contains currently only one additional species: S. rugosus at present[1]. Segniliparus is currently the only genus in the family Segniliparaceae. The generic name of the genus derives from the Latin word ‘segnis’, meaning ‘slow’, and the Greek word ‘liparos’, fat/fatty, meaning ‘one with slow fats’, to indicate the possession of slow reacting fatty acids, i.e., late eluting mycolic acids detected with HPLC [1]. The species name is derived from the Latin word ‘rotundus’, rounded, referring to the smooth, round-domed colony forms [1]. Strain CDC 1076T was isolated from human sputum in Tennessee, USA [1]. Currently, only one additional strain of the species, CDC 413 (with identical 16S rRNA gene sequence), is known, which has been isolated from the human nasal region in Missouri, USA [1]. The 16S rRNA gene sequence of the type strain for the second species in the genus, S. rugosus [1], differs by only 1.1% from that of strain CDC 1076T. S. rugosus strains have been isolated from patients with cystic fibrosis in Australia and most probably USA [2,3], suggesting that S. rotundus could also be an opportunistic pathogen. The next closest relatives of S. rotundus outside the genus are the members of the genus Rhodococcus, which share 93.3 to 94.8% 16S rRNA genes sequence similarity with strain CDC 1076T [4]. Environmental screens and metagenomic surveys did not detected a single phylotype with more than 90-92% 16S rRNA gene sequence similarity, indicating a rather limited ecological distribution of the members of the genus Segniliparus (status February 2010). Here we present a summary classification and a set of features for S. rotundus CDC 1076T, together with the description of the complete genomic sequencing and annotation.

Classification and features

Figure 1 shows the phylogenetic neighborhood of for S. rotundus CDC 1076T in a 16S rRNA based tree. The sequence of the sole 16S rRNA gene in the genome is identical with the previously published 16S rRNA sequence generated from DSM 44985 (AY608918).

Figure 1

Phylogenetic tree highlighting the position of S. rotundus CDC 1076T relative to the other type strains within the suborder Corynebacterineae. The tree was inferred from 1,436 aligned characters [5,6] of the 16S rRNA gene sequence under the maximum likelihood criterion [7] and rooted with the type strains of the order Actinomycetales. The branches are scaled in terms of the expected number of substitutions per site. Numbers above branches are support values from 350 bootstrap replicates [8] if larger than 60%. Lineages with type strain genome sequencing projects registered in GOLD [9] are shown in blue, published genomes in bold [10,11].

CDC 1076T cells are short rods with 0.4µm width by 1.0-1.3 µm length (Table 1 and Figure 2), forming round, smooth, dense and domed colonies [1]. Occasionally, v-forms are produced, but no true branching, mycelium, or spores have been reported. The colonies are non-pigmented, non-photochromogenic and do not produce a diagnostic odor [1]. It is negative for arylsulfatase after three days but positive after 14 days. Strain CDC 1076T does not grow on MacConkey agar, is weakly positive for iron uptake, Tween opacity and Tween hydrolysis, but negative for nitrate and tellurite reduction and for growth in lysozyme (21 days) [1]. Strain CDC 1076T does not produce niacin and develops bubbles in the semi-quantitative catalase test [1]. Using the API CORYNE test kit, strain CDC 1076T is positive for β-glucosidase and pyrazinamidase activities and negative for alkaline phosphatase, β-galactosidase, β-glucuronidase, α-glucosidase, N-acetyl-β-glucosaminidase and pyrrolidonyl arylamidase activity at 33°C [1]. Strain CDC 1076T is susceptible to amikacin, cefoxitan, clarithromycin, ciprofloxacin, doxycycline, imipenem and sulfamethoxazole at or below the respective MIC breakpoints but intermediate to tobramycin [1]. Glucose, maltose, D-fructose and trehalose are used as carbon source for growth with acid production, but not adonitol, L-arabinose, cellobiose, dulcitol, i-erythritol, galactose, i-myo-inositol, lactose, mannose, melibiose, raffinose, L-rhamnose, salicin, D-mannitol, D-sorbitol and sodium citrate [1]. Strain CDC 1076T hydrolyzes urea but not acetamide, adenine, casein, citrate, aesculin, hypoxanthine, tyrosine and xanthine [1].

Table 1

Classification and general features of S. rotundus CDC 1076T according to the MIGS recommendations [12]




      Evidence code

     Current classification

     Domain Bacteria

      TAS [13]

     Phylum Actinobacteria

      TAS [14]

     Class Actinobacteria

      TAS [15]

     Subclass Actinobacteridae

      TAS [15]

     Order Actinomycetales

      TAS [15]

     Suborder Corynebacterineae

      TAS [15]

     Family Segniliparaceae

      TAS [1]

     Genus Segniliparus

      TAS [1]

     Species Segniliparus rotundus

      TAS [1]

     Type strain CDC 1076

      TAS [1]

     Gram stain



     Cell shape

     short rods

      TAS [1]



      TAS [1]



      TAS [1]

     Temperature range

     mesophile, 28°C - 37°C

      TAS [1]

     Optimum temperature


      TAS [1]


     not determined


     Oxygen requirement


      TAS [1]

     Carbon source

     glucose, maltose, D-fructose, trehalose

      TAS [1]

     Energy source


      TAS [1]



     unknown, but probably host associated

      TAS [1]


     Biotic relationship




     most probably opportunistic pathogen

      TAS [1-3]

     Biosafety level


      TAS [16]


     human sputum

      TAS [1]


     Geographic location

     Tennessee, USA

      TAS [1]


     Sample collection time

     2005 or before

      TAS [1]


     Latitude     Longitude








Evidence codes - IDA: Inferred from Direct Assay (first time in publication); TAS: Traceable Author Statement (i.e., a direct report exists in the literature); NAS: Non-traceable Author Statement (i.e., not directly observed for the living, isolated sample, but based on a generally accepted property for the species, or anecdotal evidence). These evidence codes are from of the Gene Ontology project [17]. If the evidence code is IDA, then the property was directly observed for a live isolate by one of the authors or an expert mentioned in the acknowledgements.

Figure 2

Scanning electron micrograph of S. rotundus CDC 1076T


The cell wall of strain CDC 1076T contains mycolic acids and meso-diaminopimelic acid [1]. The mycolic acid HPLC pattern is a triple cluster of contiguous eluting peaks starting at approx. 6.0 min and ending with the last peak co-eluting with the internal standard. The TLC mycolic acid pattern reveals α+- and α-mycolates [1]. The fatty acids composition of the strain is dominated by straight-chain saturated acids such as the taxon-specific C10:0 (21.0%), C16:0 (18.5%), C14:0 (15.3%), 10-methyl-C18:0 (7.4%, tuberculostearic acid), C20:0 (4.9%), C12:0 (2.4%), C18:0 (1.9%), with some by straight-chain desaturated acids, C18:1 cis (15.1%) and C16:1ω9t (9.7%); (personal communication with R.M. Kroppenstedt). Quinones are mainly MK 8(H4) and MK 8(H2) with some MK 8(H6) and traces of MK 9(H2) (R.M. Kroppenstedt, personal communication).

Genome sequencing and annotation

Genome project history

This organism was selected for sequencing on the basis of its phylogenetic position, and is part of the Genomic Encyclopedia of Bacteria and Archaea project [18]. The genome project is deposited in the Genome OnLine Database [9] and the complete genome sequence is deposited in GenBank. Sequencing, finishing and annotation were performed by the DOE Joint Genome Institute (JGI). A summary of the project information is shown in Table 2.

Table 2

Genome sequencing project information





     Finishing quality



     Libraries used

    Two genomic 454 libraries: one standard    and one 4kb PE; one Illumina shotgun library


     Sequencing platforms

    454 GS FLX Titanium, Illumina GAii


     Sequencing coverage

    58.1× 454 pyrosequence, 73.3× Illumina



    Newbler version 12.0.1 PreRelease     3/30/2009.1.02.15, Velvet, phrap


     Gene calling method




     GenBank Date of Release

    not yet

     GOLD ID


     NCBI project ID


     Database: IMG-GEBA



     Source material identifier

    DSM 44985

     Project relevance

    Tree of Life, GEBA

Growth conditions and DNA isolation

S. rotundus CDC 1076T, DSM 44985, was grown in DSMZ medium 645 (Middlebrook Medium) [19] at 28°C. DNA was isolated from 1-1.5 g of cell paste using Qiagen Genomic 500 DNA Kit (Qiagen, Hilden, Germany) with lysis modification LALMP according to Wu et al. [18].

Genome sequencing and assembly

The genome was sequenced using a combination of Illumina and 454 technologies [20]. An Illumina GAii shotgun library with reads of 443 Mb, a 454 Titanium draft library with average read length of 304 bases, and a paired-end 454 library with average insert size of 4 Kb were generated for this genome. All general aspects of library construction and sequencing can be found at Web Site. Illumina sequencing data was assembled with VELVET [21] and the consensus sequences were shredded into 1.5 kb overlapped fake reads and assembled together with the 454 data. Draft assemblies were based on 183 Mb 454 data, and 454 paired-end data. Newbler parameters are -consed -a 50 -l 350 -g -m -ml 20. The initial assembly contained 26 contigs in one scaffold. We converted the initial 454 assembly into a phrap assembly by making fake reads from the consensus, collecting the read pairs in the 454 paired-end library. The Phred/Phrap/Consed software package (Web Site) was used for sequence assembly and quality assessment [18] in the following finishing process. After the shotgun stage, reads were assembled with parallel phrap (High Performance Software, LLC). Possible mis-assemblies were corrected with gapResolution (unpublished, Web Site), Dupfinisher [22], or sequencing cloned bridging PCR fragments with subcloning or transposon bombing (Epicentre Biotechnologies, Madison, WI). Gaps between contigs were closed by editing in Consed, by PCR and by Bubble PCR (J-F Cheng, unpublished) primer walks. A total of 108 additional reactions were necessary to close gaps and to raise the quality of the finished sequence. The completed genome sequences had an error rate less than one in 100,000 bp.

Genome annotation

Genes were identified using Prodigal [23] as part of the Oak Ridge National Laboratory genome annotation pipeline, followed by a round of manual curation using the JGI GenePRIMP pipeline [24]. The predicted CDSs were translated and used to search the National Center for Biotechnology In-formation (NCBI) nonredundant database, UniProt, TIGRFam, Pfam, PRIAM, KEGG, COG, and InterPro databases. Additional gene prediction analysis and manual functional annotation was performed within the Integrated Microbial Genomes Expert Review (IMG-ER) platform [25].

Genome properties

The genome consists of a 3,157,527 bp long chromosome (Table 3 and Figure 3). Of the 3,133 genes predicted, 3,081 were protein-coding genes, and 52 RNAs; 75 pseudogenes were also identified. The majority of the protein-coding genes (63.0%) were assigned with a putative function while those remaining were annotated as hypothetical proteins. The distribution of genes into COGs functional categories is presented in Table 4.

Table 3

Genome Statistics



  % of Total

Genome size (bp)



DNA coding region (bp)



DNA G+C content (bp)



Number of replicons


Extrachromosomal elements


Total genes



RNA genes



rRNA operons


Protein-coding genes



Pseudo genes



Genes with function prediction



Genes in paralog clusters



Genes assigned to COGs



Genes assigned Pfam domains



Genes with signal peptides



Genes with transmembrane helices



CRISPR repeats


Figure 3

Graphical circular map of the genome. From outside to the center: Genes on forward strand (color by COG categories), Genes on reverse strand (color by COG categories), RNA genes (tRNAs green, rRNAs red, other RNAs black), GC content, GC skew.

Table 4

Number of genes associated with the general COG functional categories








     Translation, ribosomal structure and biogenesis




     RNA processing and modification








     Replication, recombination and repair




     Chromatin structure and dynamics




     Cell cycle control, cell division, chromosome partitioning




     Nuclear structure




     Defense mechanisms




     Signal transduction mechanisms




     Cell wall/membrane biogenesis




     Cell motility








     Extracellular structures




     Intracellular trafficking, secretion, and vesicular transport




     Posttranslational modification, protein turnover, chaperones




     Energy production and conversion




     Carbohydrate transport and metabolism




     Amino acid transport and metabolism




     Nucleotide transport and metabolism




     Coenzyme transport and metabolism




     Lipid transport and metabolism




     Inorganic ion transport and metabolism




     Secondary metabolites biosynthesis, transport and catabolism




     General function prediction only




     Function unknown




     Not in COGs



This work was performed under the auspices of the US Department of Energy's Office of Science, Biological and Environmental Research Program, and by the University of California, Lawrence Berkeley National Laboratory under contract No. DE-AC02-05CH11231, Lawrence Livermore National Laboratory under Contract No. DE-AC52-07NA27344, Los Alamos National Laboratory under contract No. DE-AC02-06NA25396, and Oak Ridge National Laboratory under contract DE-AC05-00OR22725, as well as German Research Foundation (DFG) INST 599/1-1 and SI 1352/1-2.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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