Open Access

Complete genome sequence of Coriobacterium glomerans type strain (PW2T) from the midgut of Pyrrhocoris apterus L. (red soldier bug)

  • Erko Stackebrandt
  • , Ahmet Zeytun,
  • , Alla Lapidus
  • , Matt Nolan
  • , Susan Lucas
  • , Nancy Hammon
  • , Shweta Deshpande
  • , Jan-Fang Cheng
  • , Roxanne Tapia,
  • , Lynne A. Goodwin,
  • , Sam Pitluck
  • , Konstantinos Liolios
  • , Ioanna Pagani
  • , Natalia Ivanova
  • , Konstantinos Mavromatis
  • , Natalia Mikhailova
  • , Marcel Huntemann
  • , Amrita Pati
  • , Amy Chen
  • , Krishna Palaniappan
  • , Yun-juan Chang,
  • , Miriam Land,
  • , Loren Hauser,
  • , Manfred Rohde
  • , Rüdiger Pukall
  • , Markus Göker
  • , John C. Detter
  • , Tanja Woyke
  • , James Bristow
  • , Jonathan A. Eisen,
  • , Victor Markowitz
  • , Philip Hugenholtz,
  • , Nikos C. Kyrpides
  • and Hans-Peter Klenk
Corresponding author

DOI: 10.4056/sigs.3507020

Received: 15 April 2013

Published: 15 April 2013


Coriobacterium glomerans Haas and König 1988, is the only species of the genus Coriobacterium, family Coriobacteriaceae, order Coriobacteriales, phylum Actinobacteria. The bacterium thrives as an endosymbiont of pyrrhocorid bugs, i.e. the red fire bug Pyrrhocoris apterus L. The rationale for sequencing the genome of strain PW2T is its endosymbiotic life style which is rare among members of Actinobacteria. Here we describe the features of this symbiont, together with the complete genome sequence and its annotation. This is the first complete genome sequence of a member of the genus Coriobacterium and the sixth member of the order Coriobacteriales for which complete genome sequences are now available. The 2,115,681 bp long single replicon genome with its 1,804 protein-coding and 54 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.


Gram-positivenon-motilenon-sporulatingobligatory anaerobicchemoorganotrophmesophileendosymbiontinsect intestinal tractCoriobacteriaceaeActinobacteriaGEBA


Strain PW2T (= DSM 20642 = ATCC 49209 = JCM 10262) is the type strain of Coriobacterium glomerans [1]. The absence of sequence data in the original description excluded determination of the phylogenetic position of the genus Coriobacterium, but taxonomic evidence also excluded an affiliation with Bifidobacterium, Eubacterium or Lachnospira. The 16S rRNA gene sequence [2] revealed that Coriobacterium and Atopobium [3] are phylogenetic neighbors. Based upon phylogenetic position within the class Actinobacteria and a unique set of 16S rRNA gene signature nucleotides both genera were placed in the family Coriobacteriaceae, order Coriobacterales, subclass Coriobacteridae [4]. The family was expanded by the description of several new genera which at the time of writing encompasses 13 genera and 29 species [5]. In the 2nd edition of Bergey’s Manual the class Actinobacteria was elevated to phylum rank [6] and subsequently the subclass Coriobacteridae was elevated to class rank [7]. The suborder rank ‘Coriobacterineae has been introduced by Garrity and collaborators [8]. Coriobacterium is a phylogenetic neighbor of Collinsella [9] and both genera form one of four sister clades of Coriobacteriaceae. Besides the type strain a few other closely related strains (e.g. accession numbers FJ554833, FJ554832, FJ554836, FJ554835) were isolated from Pyrrhocoris apterus L. and a related pyrrhocorid host. Their localization in the midgut, the rectum and feces of the red firebug and the vertical transmission route via application of the symbiont to the surface of the eggs was determined via PCR amplification and FISH hybridization. Horizontal transmission also occurred via symbiont-containing material [10]. BLAST re-analysis of 16S rRNA gene sequences of other strains and clones (e.g. accession numbers AJ131149, AJ131150, AJ245921) reported to be members of Coriobacterium [11] revealed that they are actually members of Collinsella.

Here we present a summary classification and a set of features for C. glomerans PW2T together with the description of the complete genomic sequencing and annotation.

Features of the organism

16S rRNA gene sequence analysis

A representative genomic 16S rRNA gene sequence of C. glomerans PW2T was compared using NCBI BLAST [12,13] under default settings (e.g., considering only the high-scoring segment pairs (HSPs) from the best 250 hits) with the most recent release of the Greengenes database [14] and the relative frequencies of taxa and keywords (reduced to their stem [15]) were determined, weighted by BLAST scores. The most frequently occurring genera were Collinsella (61.9%) and Coriobacterium (38.1%) (29 hits in total). Regarding the five hits to sequences from members of the species, the average identity within HSPs was 97.8%, whereas the average coverage by HSPs was 93.4%. Among all other species, the one yielding the highest score was Collinsella tanakaei (AB490807), which corresponded to an identity of 93.4% and an HSP coverage of 99.4%. (Note that the Greengenes database uses the INSDC (= EMBL/NCBI/DDBJ) annotation, which is not an authoritative source for nomenclature or classification.) The highest-scoring environmental sequence was EF399657 (Greengenes short name 'human fecal clone SJTU E 01 75'), which showed an identity of 93.5% and an HSP coverage of 98.4%. The most frequently occurring keywords within the labels of all environmental samples which yielded hits were 'human' (20.6%), 'fecal' (19.8%), 'fece' (10.6%), 'lion' (4.7%) and 'intestin' (1.9%) (221 hits in total). Environmental samples which yielded hits of a higher score than the highest scoring species were not found, indicating that C. glomerans is rarely found in environmental samples.

Figure 1 shows the phylogenetic neighborhood of C. glomerans PW2T in a 16S rRNA based tree. The sequences of the two identical 16S rRNA gene copies in the genome differ by six nucleotides from the previously published 16S rRNA sequence (X79048), which contains three ambiguous base calls.

Figure 1

Phylogenetic tree highlighting the position of C. glomerans relative to the type strains of the other species within the family Coriobacteriaceae. The tree was inferred from 1,401 aligned characters [16,17] of the 16S rRNA gene sequence under the maximum likelihood (ML) criterion [18]. Rooting was done initially using the midpoint method [19] and then checked for its agreement with the current classification (Table 1). The branches are scaled in terms of the expected number of substitutions per site. Numbers adjacent to the branches are support values from 300 ML bootstrap replicates [20] (left) and from 1,000 maximum-parsimony bootstrap replicates [21] (right) if larger than 60%. Lineages with type strain genome sequencing projects registered in GOLD [22] are labeled with one asterisk, those also listed as 'Complete and Published' with two asterisks ([23-27], see FP929047 for Gordonibacter pamelaeae).

Table 1

Classification and general features of C. glomerans PW2T according to the MIGS recommendations [28].




    Evidence code

   Current classification

    Domain Bacteria

    TAS [29]

    Phylum Actinobacteria

    TAS [30]

    Class Actinobacteria

    TAS [4]

    Subclass Coriobacteridae

    TAS [4,31]

    Order Coriobacteriales

    TAS [4,31]

    Suborder Coriobacterineae

    TAS [32]

    Family Coriobacteriaceae

    TAS [4,31]

    Genus Coriobacterium

    TAS [1]

    Species Coriobacterium glomerans

    TAS [1]


   Subspecific genetic lineage (strain)


    TAS [1]


   Reference for biomaterial

    Haas and König 1988

    TAS [1]

   Gram stain


    TAS [1]

   Cell shape


    TAS [1]



    TAS [1]



    TAS [1]

   Temperature range


    TAS [1]

   Optimum temperature


    TAS [1]


    not reported


   Relationship to oxygen

    obligate anaerobe

    TAS [1]

   Carbon source

    not reported

   Energy metabolism


    TAS [1]



    host, intestinal tract

    TAS [1]



    not reported


   Biotic relationship



   Known pathogenicity


    TAS [1]


   Specific host

    Pyrrhocoris apterus L.

    TAS [1]


   Health status of Host


   Biosafety level


    TAS [33]


   Trophic level




    intestinal tract of the red soldier bug

    TAS [1]


   Geographic location

    Bavaria, Germany

    TAS [1]


   Time of sample collection

    December 1981




    not reported



    not reported



    not reported



    not reported

Evidence codes - 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). Evidence codes are from the Gene Ontology project [34].

Morphology and physiology

Cells of C. glomerans PW2T stain Gram-positive which is consistent with an electron-dense cell wall, 40 nm wide [35]. The cells are pear-shaped to irregularly shaped rods with length that varies from 0.44 to 1.80 µm (Figure 2). Spherical involution forms are common. When attached to the epithelia of the intestines, the bacteria form chains which may reach more than 150 µm in length. Flagella are absent. Colonies, grown anaerobically in an atmosphere of N2-CO2 (80:20) on blood agar (Columbia agar base, BBL), supplemented Schaedler agar (BBL), and TPY agar at 25 and 30°C are about 0.6 mm in diameter and consist of long filamentous chains, bent into hairpins, curls, and loops. A flocculent, wooly sediment with a clear supernatant is formed in fluid media. The bacteria are strictly anaerobic. In TPY medium glucose is fermented to acetic acid, L-lactic acid, and ethanol in a molar ratio of 1.16:1.00:0.95. CO2 and H2 also are produced [1] but D-lactic acid, formic acid, volatile short-chain alcohols, or other volatile fatty acids are not formed. Under more stringent anaerobic conditions with N2-CO2 (80:20) as the gas phase, but lacking H2, the formation of ethanol occurs only at a lower concentration. C. glomerans PW2T ferments glucose, L-arabinose, D-xylose, D-ribose, mannose, sucrose, maltose, cellobiose, mannitol, and salicin but not lactose, melibiose, raffinose, inulin, starch, and inositol [1].

Figure 2

Scanning electron micrograph of C. glomerans PW2T


The peptidoglycan of stain PW2T contains lysine as the diagnostic amino acid in position 3 of the peptide subunit with the interpeptide bridge containing aspartic acid (Lys-Asp type; A4α according to [36]; A11.31 according to [37]). Information on major cell wall sugars, fatty acids, menaquinones and polar lipids is not available. The mol% G+C of DNA was reported to be about 61, and is here confirmed by the genome sequence.

Genome sequencing and annotation

Genome project history

This organism was selected for sequencing on the basis of its phylogenetic position [38], and is part of the Genomic Encyclopedia of Bacteria and Archaea project [39]. The genome project is deposited in the Genomes OnLine Database [22] and the complete genome sequence is deposited in GenBank. Sequencing, finishing and annotation were performed by the DOE Joint Genome Institute (JGI) using state of the art sequencing technology [40]. A summary of the project information is shown in Table 2.

Table 2

Genome sequencing project information





   Finishing quality



   Libraries used

   Three genomic libraries: one 454 pyrosequence standard library,   one 454 PE library (6.7 kb insert size), one Illumina library


   Sequencing platforms

   Illumina GAii, 454 GS FLX Titanium


   Sequencing coverage

   35.8 × Illumina; 68.3 × pyrosequence



   Newbler version 2.3, Velvet, Phrap version SPS - 4.24


   Gene calling method

   Prodigal 1.4, GenePRIMP



   GenBank Date of Release

   October 11, 2011



   NCBI project ID


   Database: IMG-GEBA



   Source material identifier

   DSM 20642

   Project relevance

   Tree of Life, GEBA

Growth conditions and DNA isolation

C. glomerans strain PW2T, DSM 20642, was grown anaerobically in an atmosphere of N2-CO2 (80:20) in DSMZ medium 104 (modified PYG medium) at 30°C. DNA was isolated from 1-1.5 g of cell paste using MasterPure Gram-positive DNA purification kit (Epicentre MGP04100) following the standard protocol as recommended by the manufacturer with modification st/LALM for cell lysis as described in Wu et al. 2009 [39]. DNA is available through the DNA Bank Network [41].

Genome sequencing and assembly

The genome was sequenced using a combination of Illumina and 454 sequencing platforms. All general aspects of library construction and sequencing can be found at the JGI website [42]. Pyrosequencing reads were assembled using the Newbler assembler (Roche). The initial Newbler assembly consisting of 14 contigs in one scaffold was converted into a phrap [43] assembly by making fake reads from the consensus, to collect the read pairs in the 454 paired end library. Illumina GAii sequencing data (75.8 Mb) was assembled with Velvet [44] and the consensus sequences were shredded into 1.5 kb overlapped fake reads and assembled together with the 454 data. The 454 draft assembly was based on 128.4 Mb 454 draft data and all of the 454 paired end data. Newbler parameters are -consed -a 50 -l 350 -g -m -ml 20. The Phred/Phrap/Consed software package [43] was used for sequence assembly and quality assessment in the subsequent finishing process. After the shotgun stage, reads were assembled with parallel phrap (High Performance Software, LLC). Possible mis-assemblies were corrected with gapResolution [42], Dupfinisher [45], or sequencing cloned bridging PCR fragments with subcloning. Gaps between contigs were closed by editing in Consed, by PCR and by Bubble PCR primer walks (J.-F. Chang, unpublished). A total of 40 additional reactions were necessary to close gaps and to raise the quality of the finished sequence. Illumina reads were also used to correct potential base errors and increase consensus quality using a software Polisher developed at JGI [46]. The error rate of the completed genome sequence is less than 1 in 100,000. Together, the combination of the Illumina and 454 sequencing platforms provided 104.1 × coverage of the genome. The final assembly contained 456,305 pyrosequence and 2,106,317 Illumina reads.

Genome annotation

Genes were identified using Prodigal [47] as part of the DOE-JGI genome annotation pipeline [48], followed by a round of manual curation using the JGI GenePRIMP pipeline [49]. The predicted CDSs were translated and used to search the National Center for Biotechnology Information (NCBI) nonredundant database, UniProt, TIGR-Fam, Pfam, PRIAM, KEGG, COG, and InterPro databases. Additional gene prediction analysis and functional annotation was performed within the Integrated Microbial Genomes – Expert Review (IMG-ER) platform [50].

Genome properties

The genome statistics are provided in Table 3 and Figure 3. The genome consists of one chromosome with a total length of 2,115,681 bp and a G+C content of 60.4%. Of the 1,858 genes predicted, 1,804 were protein-coding genes, and 54 RNAs; 36 pseudogenes were also identified. The majority of the protein-coding genes (74.2%) were assigned a putative function while the remaining ones 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


   tRNA genes



   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 map of the chromosome. 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 (purple/olive).

Table 4

Number of genes associated with the general COG functional categories



   % age





    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 and 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



We would like to gratefully acknowledge the help of Gabriele Gehrich-Schröter for growing C. glomerans cultures, and Susanne Schneider for DNA extraction and quality control (both at DSMZ). This work was performed under the auspices of the US Department of Energy 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, and Los Alamos National Laboratory under contract No. DE-AC02-06NA25396, UT-Battelle and Oak Ridge National Laboratory under contract DE-AC05-00OR22725.


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