Open Access

Complete genome sequence of Corynebacterium pseudotuberculosis biovar ovis strain P54B96 isolated from antelope in South Africa obtained by rapid next generation sequencing technology

  • Syed Shah Hassan
  • , Luis Carlos Guimarães
  • , Ulisses de Pádua Pereira
  • , Arshad Islam
  • , Amjad Ali
  • , Syeda Marriam Bakhtiar
  • , Dayana Ribeiro
  • , Anderson Rodrigues dos Santos
  • , Siomar de Castro Soares
  • , Fernanda Dorella
  • , Anne Cybelle Pinto
  • , Maria Paula Cruz Schneider
  • , Maria Silvanira Barbosa
  • , Síntia Almeida
  • , Vinícius Abreu
  • , Flávia Aburjaile
  • , Adriana Ribeiro Carneiro
  • , Louise Teixeira Cerdeira
  • , Karina Fiaux
  • , Eudes Barbosa
  • , Carlos Diniz
  • , Flavia S. Rocha
  • , Rommel Thiago Jucá Ramos
  • , Neha Jain
  • , Sandeep Tiwari
  • , Debmalya Barh
  • , Anderson Miyoshi
  • , Borna Müller
  • , Artur Silva
  • and Vasco Azevedo
Corresponding author

DOI: 10.4056/sigs.3066455

Received: 15 December 2012

Published: 19 December 2012

Abstract

The Actinobacteria, Corynebacterium pseudotuberculosis strain P54B96, a nonmotile, non-sporulating and a mesophile bacterium, was isolated from liver, lung and mediastinal lymph node lesions in an antelope from South Africa. This strain is interesting in the sense that it has been found together with non-tuberculous mycobacteria (NTMs) which could nevertheless play a role in the lesion formation. In this work, we describe a set of features of C. pseudotuberculosis P54B96, together with the details of the complete genome sequence and annotation. The genome comprises of 2.34 Mbp long, single circular genome with 2,084 protein-coding genes, 12 rRNA, 49 tRNA and 62 pseudogenes and a G+C content of 52.19%. The analysis of the genome sequence provides means to better understanding the molecular and genetic basis of virulence of this bacterium, enabling a detailed investigation of its pathogenesis.

Keyword

s: biovar ovisGram-positive pathogencaseous lymphadenitis/cheesy gland diseaseliver lesionAntelopegenome sequencingIon Torrent

Introduction

Caseous lymphadenitis (CLA) or cheesy gland [1] is highly prevalent in many regions of the world, resulting in huge and significant economic losses in agribusiness since it is responsible for a decrease in wool production and carcass quality [2]. Mainly small ruminant populations like sheep and goats, and other mammals, such as bovines, pigs, deer, ovines, equines, and even, though rarely, in camels and humans, are the victims of Corynebacterium pseudotuberculosis [3-6]. The disease is characterized by the presence of caseous necrosis in external and/or internal lymph nodes [1,7]. Ulcerative lymphangitis, which is confined to the lymph vessels of extremities particularly the hind legs, is a disease caused by this bacterium in the horse [8,9]. The bacterium in some cases of human lymphadenitis, clinical strains are occasionally recovered [10]. The prevalence of CLA in the animals scattered throughout the globe needs effective measures to control the onset of the disease in herds along with the treatment of infected animals. Numerous reports have been published worldwide where mainly small ruminants are the carriers of the C. pseudotuberculosis. They include South Africa, Brazil, United States of America, Canada, Australia, New Zealand, United Kingdom and Egypt [11-18]. Histopathological examination of antelope carcasses from a South African game reserve, a part of their routine meat inspection, showed tuberculosis-like lesions. These lesions were characterized by the presence of encapsulated necrogranulomatous inflammation similar to CLA within the pulmonary tissues, in bronchial lymph nodes, liver, kidney and some other organs of the antelopes [11]. Diseases caused by the bacterium C. pseudotuberculosis are presented in various clinical forms as sheep and goats, affected with CLA [19]. Among the affected animal population, the increased prevalence and rapid transmission of the disease necessitates certain measures to control disease dissemination and prevent the nearby wildlife. The analysis of the genome sequence will help us better understand the molecular and genetic basis of virulence of this bacterium.

Classification and Features

C. pseudotuberculosis is a facultative intracellular pathogen showing pleomorphic forms like coccoids and filamentous rods, with sizes ranging between 0.5-0.6 µm and 1.0-3.0 µm [2]. Cells are described as Gram-positive, non-encapsulated, non-motile, non-sporulating and possessing fimbriae [12,20]. The bacterium was first isolated in 1888 from bovine farcy by Nocard and was first completely described by Preisz, showing its resemblance to diphtheria bacillus. The organism has been previously named Bacillus pseudotuberculosis ovis; Bacillus pseudotuberculosi and, Corynebacterium ovis [8,21]. It is a facultative anaerobe. The best growth temperature and pH are 37o C and 7.0-7.2, respectively [17,22]. After initially growing sparsely, strain P54B96 forms organized clumps on the agar surface, demonstrating dry opaque and concentrically ringed colonies. In liquid media it develops a granular deposit with a surface pellicle [8,22,23].

There exist two biotypes of C. pseudotuberculosis according to their capability of nitrate reduction. Bacteria capable of performing the reduction of nitrate are classified into biovar equi (nitrate reduction positive; mainly isolated from horses and cattle) while the bacteria which can not perform the reduction of nitrate, pertain to biovar ovis (nitrate reduction negative; frequently isolated from sheep and goats) [2,24]. Corynebacteria possess an unusual structural organization in their cell envelope, similar to the Gram-negative bacteria [25] and belong to a very heterogeneous CMNR (Corynebacterium, Mycobacterium, Nocardia and Rhodococcus) group that shares characteristics including an outer lipid layer, mycolic acids in the cell wall along with with its derivatives including phospholipids and lipomannans [4]. Marchand et al. (2012) and others reported the presumed mycomembrane, an atypical outer membrane, pore-forming proteins like PorA and PorB, mycoloyltransferases, the so-called fibronectin-binding proteins like cMytA-D and cMytF, several lipoproteins and some unknown putative C-terminal hydrophobic anchored proteins [26]. Analysis of amino acids and amino sugars of cell wall peptidoglycan reveals the presence of meso-diaminopimelic acid (meso-DAP). Major cell wall sugars are arabinose and galactose [17,27]. In addition, high and low molecular mass glucan, arabinomannan and lipoglycan also make part of the cell wall. Trehalose dimycolate (TDM) and trehalose monomycolate (TMM) are soluble cell envelope lipids [28]. Biochemically, all strains produce acid from glucose, maltose, fructose, sucrose and mannose [21,22]. This bacterium is catalase positive and phospholipase D, beta-hemolysis and oxidase negative [23,29].

Figure 1 shows the phylogenetic neighborhood of C. pseudotuberculosis strain P54B96 in an rpoB gene (β subunit of RNA polymerase) based tree. It has recently been shown that phylogenetic analysis for the identification of Corynebacterium as well as other CMNR species based on rpoB gene sequences are more accurate than analyses based on 16S rRNA [42,43]. The rpoB gene sequences of reference strains from the CMNR group were used to construct the phylogenetic tree.

Figure 1

Phylogenetic tree of C. pseudotuberculosis strain P54B96 representing its position relative to type strains in Corynebacteriaceae along with some other type strains of CMNR group. The tree was inferred from 3,537 aligned characters of the rpoB gene sequence using maximum likelihood method and then checked for its agreement with the current classification Table 1. The branch lengths represent the expected number of substitutions per site. Numbers adjacent to the branches are support values from 1,000 bootstrap replicates, indicated when Larger than 60%. Calculations to determine the phylogenetic distances were done by the software MEGA v5 [30].

Table 1

Classification and general features of C. pseudotuberculosis strain P54B96 according to the MIGS recommendations [31].

MIGS ID

    Property

     Term

      Evidence code

    Classification

     Domain Bacteria

      TAS [32]

     Phylum Actinobacteria

      TAS [33]

     Class Actinobacteria

      TAS [34]

     Order Actinomycetales     Suborder Corynebacterineae

      TAS [34-37]

     Family Corynebacteriaceae

      TAS [34,35,37,38]

     Genus Corynebacterium

      TAS [35,38,39]

     Species Corynebacterium pseudotuberculosis

      TAS [35,40]

     Strain P54B96

      TAS [11]

    Gram stain

     Positive

      TAS [21]

    Cell shape

     pleomorphic forms

      TAS [21]

    Motility

     non-motile

      TAS [8]

    Sporulation

     non-sporulating

      TAS [22]

    Temperature range

     mesophilic

      TAS [8,22]

    Optimum temperature

     37°C

      TAS [8,22]

    Salinity

     not reported

      NAS

MIGS-22

    Oxygen requirement

     aerobic and facultatively anaerobic

      TAS [8,22]

    Carbon source

     glucose, fructose, maltose, mannose,     and sucrose

      TAS [8]

    Energy source

  Chemoorganotroph

      TAS [8]

MIGS-6

    Habitat

     Host

      TAS [22]

MIGS-15

    Biotic relationship

     intracellular facultative pathogen

      TAS [22]

MIGS-14

    Pathogenicity

     sheep, goats, horses and cattle,     rarely humans

      TAS [5,6]

    Biosafety level

     2

      TAS [22]

    Isolation

     liver, lung, mediastinal lymph node lesions of antelope

      TAS [11]

MIGS-4

    Geographic location

     Mpumalanga province, South Africa

      TAS [11]

MIGS-5

    Sample collection time

     2009

      TAS [11]

MIGS-4.1MIGS-4.2

    Latitude    Longitude

     not reported     not reported

MIGS-4.3

    Depth

     not reported

MIGS-4.4

    Altitude

     not reported

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 the Gene Ontology project [41]. If the evidence code is IDA, then the property was directly observed for a living isolate by one of the authors or an expert mentioned in the acknowledgements.

Genome sequencing and annotation

Genome project history

This organism was selected for sequencing on the basis of its phylogenetic position. The genome project is deposited in the Genomes OnLine Database [44] and the complete genome sequence is available in GenBank (CP003385.1). Sequencing, finishing and annotation were performed by the Rede Paraense de Genômica e Proteômica (RPGP), Pará, Brazil. A summary of the project information is shown in Table 2.

Table 2

Genome sequencing project information

MIGS ID

     Property

     Term

MIGS-31

     Finishing quality

     Finished

MIGS-28

     Libraries used

     Fragments (mean size 112 bp)

MIGS-29

     Sequencing platforms

     Semiconductor Ion Torrent PGM

MIGS-31.2

     Sequencing coverage

     35-fold

MIGS-30

     Assemblers

     CLC Genome Workbench 4.7.2, Velvet

MIGS-32

     Gene calling method

     Glimmer v3.02

     INSDC ID

     CP003385 (chromosome)

     GenBank Date of Release

     April 05, 2012

     GOLD ID

     Gc02176

     NCBI project ID

     77871

     Database: IMG-GEBA

     2512564058

MIGS-13

     Source material identifier

     BHI broth, P54B96

     Project relevance

     Animal Pathogen, Medical

Growth conditions and DNA isolation

C. pseudotuberculosis P54B96 was grown in brain-heart-infusion broth (BHI-HiMedia Laboratories Pvt. Ltda, India) in shake culture at140 rpm and at 37oC. Extraction of chromosomal DNA was performed by using 50 mL of 48–72 h culture of C. pseudotuberculosis, centrifuged at 4oC and 2000× g for 20 min. Re-suspension of cell pellets was done in 1 mL Tris/EDTA/NaCl [10 mM Tris/HCl (pH7.0), 10 mM EDTA (pH8.0), and 300 mM NaCl] for re-centrifugation under the same conditions. The pellets were re-suspended in 1 mL TE/lysozyme [25 mM Tris/HCl (pH8.0), 10 mM EDTA (pH8.0), 10 mM NaCl, and 10 mg lysozyme/mL]. The sample was then incubated at 37oC for 30 min and then 30 µL of 30% (w/v) sodium N- lauroyl-sarcosine (Sarcosyl) was added to it, incubated for 20 min at 65oC, followed by incubation for 5 min at 4oC. Purification of DNA with phenol/chloroform/isoamylalcohol (25:24:1) was followed by precipitation with ethanol. DNA concentration was determined by spectrophotometer, and the DNA was visualized in ethidium bromide-stained 0.7% agarose gel.

Genome sequencing and assembly

The complete genome sequence of C. pseudotuberculosis P54B96 was obtained using the Ion Torrent PGM (Life Technologies) Sequencing Platform. A total, of 562,812 reads were generated, each with a mean size of 112 nts usable sequence (35-fold coverage). Furthermore, a hybrid de novo assembly approach was applied using 376,642 Ion filtered reads (19-fold coverage). This was carried out after quality filtering process during which reads representing an average Phred quality of less than 20, were removed. This strategy allowed closing gaps without bench work time cost [45].

For homopolymer correction, an inherent problem of the Ion Torrent [46], CLCBio Genome Workbench 4.7.2 was used. Having detected a high number of frameshifts, manual curation was required prior to analysis to prevent false-positive identification of pseudogenes. The genome of P54B96 strain consists of 2,337,657 bp circular chromosome and the average G+C content of the chromosome is 52.2%. The genome was predicted to contain 2,084 coding sequences (CDS), four rRNA operons, 49 tRNA and 62 pseudogenes.

Genome annotation

For automatic annotation, different programs were used. These include; Glimmer: gene predictor [47], RNAmmer: rRNA predictor [48]; tRNA-scan-SE: tRNA predictor [49]; and Tandem Repeat Finder: repetitive DNA predictor [50]. Functional annotation was performed by similarity analyses, using public databases of National Center for Biotechnology Information (NCBI) non-redundant database, Pfam and InterProScan software [51], which integrates multiple domain and protein family databases. Manual annotation was performed using Artemis [52].

Metabolic network analysis

The metabolic Pathway/Genome Database (PGDB) was computationally generated using Pathway Tools software version 15.0 [53] and MetaCyc version 15.0 [54], based on annotated EC numbers and a customized enzyme name mapping file. There has been no manual curation in the database and it may contain errors, similar to a Tier 3 BioCyc PGDB [55].

Genome properties

The genome is 2,337,657 bp long and comprises one main circular chromosome with a 52.19% GC content. A total of 2,207 genes were predicted, among which 2,146 were protein coding genes, and 61 RNAs; 62 pseudogenes were also identified. Of the whole genome, 69.01% comprise genes that were assigned with putative functions, while the remaining genes were annotated as hypothetical proteins. The properties and statistics of the C. pseudotuberculosis genome are listed in Table 3. The distributions of genes into COGs functional categories is presented in Figure 2 and Table 4, followed by a cellular overview diagram in Figure 3 and a summary of metabolic network statistics shown in Table 5.

Table 3

Genome Statistics

Attribute

    Value

      % of Total

Genome size (bp)

    2,337,657

      100.00%

DNA coding region (bp)

    2,005,391

      85.79%

DNA G+C content (bp)

    1,219,912

      52.19%

Number of replicons

    1

Extrachromosomal elements

    0

Total genes

    2,145

      100.00%

RNA genes

    61

      2.76%

rRNA operons

    4

Protein-coding genes

    2,084

      97.16%

Pseudo genes

    62

      2.81%

Genes with function prediction

    1,511

      68.46%

Genes in paralog clusters

    425

      19.26%

Genes assigned to COGs

    1,552

      70.32%

Genes assigned Pfam domains

    1,596

      72.32%

Genes with signal peptides

    651

      29.50%

Genes with transmembrane helices

    584

      26.46%

CRISPR repeats

    0

Figure 2

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

Code

     Value

    %age

      Description

J

     140

    6.72

      Translation, ribosomal structure and biogenesis

A

     1

    0.1

      RNA processing and modification

K

     121

    5.8

      Transcription

L

     88

    4.2

      Replication, recombination and repair

B

     0

    0.0

      Chromatin structure and dynamics

D

     21

    1.0

      Cell cycle control, cell division, chromosome partitioning

Y

     0

    0.0

      Nuclear structure

V

     25

    1.2

      Defense mechanisms

T

     54

    2.6

      Signal transduction mechanisms

M

     87

    4.2

      Cell wall/membrane biogenesis

N

     1

    0.1

      Cell motility

Z

     0

    0.0

      Cytoskeleton

W

     0

    0.0

      Extracellular structures

U

     27

    1.3

      Intracellular trafficking and secretion

O

     77

    3.7

      Posttranslational modification, protein turnover, chaperones

C

     90

    4.3

      Energy production and conversion

G

     113

    5.4

      Carbohydrate transport and metabolism

E

     177

    8.5

      Amino acid transport and metabolism

F

     73

    3.5

      Nucleotide transport and metabolism

H

     102

    4.9

      Coenzyme transport and metabolism

I

     57

    2.7

      Lipid transport and metabolism

P

     122

    5.9

      Inorganic ion transport and metabolism

Q

     26

    1.3

      Secondary metabolites biosynthesis, transport and catabolism

R

     169

    8.1

      General function prediction only

S

     136

    6.5

      Function unknown

-

     655

    31.4

      Not in COGs

Figure 3

Schematic cellular overview of all pathways of the C. pseudotuberculosis P54B96 metabolism. Nodes represent metabolites, with shape indicating class of metabolite. Lines represent reactions.

Table 5

Metabolic Network Statistics

Attribute

      Value

Total genes

      2,145

Enzymes

      500

Enzymatic reactions

      764

Metabolic pathways

      152

Metabolites

      622

Declarations

Acknowledgement

We would like to gratefully acknowledge the help of all the team members & the financing agencies. Hassan S.S acknowledges the receipt of a Scholarship from the CNPq under the “TWAS-CNPq Postgraduate Fellowship Programme” for doctoral studies. This work was partially executed by Rede Paraense de Genômica e Proteômica supported by FAPESPA (Fundação de Amparo à Pesquisa do Estado do Pará), CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico, Brasil), CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, Brasil) and FAPEMIG (Fundação de Amparo à Pesquisa do Estado de Minas Gerais, Brasil).


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References

  1. Williamson LH. Caseous lymphadenitis in small ruminants. [vii.]. Vet Clin North Am Food Anim Pract. 2001; 17:359-371PubMed
  2. Dorella FA, Pacheco LG, Oliveira SC, Miyoshi A and Azevedo V. Corynebacterium pseudotuberculosis: microbiology, biochemical properties, pathogenesis and molecular studies of virulence. Vet Res. 2006; 37:201-218 View ArticlePubMed
  3. Ayers JL. Caseous lymphadenitis in goats and sheep: a review of diagnosis, pathogenesis, and immunity. J Am Vet Med Assoc. 1977; 171:1251-1254PubMed
  4. Marchand CH, Salmeron C, Raad RB, Meniche X, Chami M, Masi M, Blanot D, Daffe M, Tropis M and Huc E. Biochemical disclosure of the mycolate outer membrane of Corynebacterium glutamicum. J Bacteriol. 2012; 194:587-597 View ArticlePubMed
  5. Brown CC, Olander HJ and Alves SF. Synergistic hemolysis-inhibition titers associated with caseous lymphadenitis in a slaughterhouse survey of goats and sheep in Northeastern Brazil. Can J Vet Res. 1987; 51:46-49PubMed
  6. Lipsky BA, Goldberger AC, Tompkins LS and Plorde JJ. Infections caused by nondiphtheria corynebacteria. Rev Infect Dis. 1982; 4:1220-1235 View ArticlePubMed
  7. Aleman M, Spier SJ, Wilson WD and Doherr M. Corynebacterium pseudotuberculosis infection in horses: 538 cases (1982-1993). J Am Vet Med Assoc. 1996; 209:804-809PubMed
  8. Merchant IA, Packer RA. Veterinary bacteriology and virology. Ames: Iowa State University Press; 1967, p. 752.
  9. Piontkowski MD and Shivvers DW. Evaluation of a commercially available vaccine against Corynebacterium pseudotuberculosis for use in sheep. J Am Vet Med Assoc. 1998; 212:1765-1768PubMed
  10. Trost E, Ott L, Schneider J, Schroder J, Jaenicke S, Goesmann A, Husemann P, Stoye J, Dorella FA and Rocha FS. The complete genome sequence of Corynebacterium pseudotuberculosis FRC41 isolated from a 12-year-old girl with necrotizing lymphadenitis reveals insights into gene-regulatory networks contributing to virulence. BMC Genomics. 2010; 11:728 View ArticlePubMed
  11. Müller B, de Klerk-Lorist LM, Henton MM, Lane E, Parsons S, Gey van Pittius NC, Kotze A, van Helden PD and Tanner M. Mixed infections of Corynebacterium pseudotuberculosis and non-tuberculous mycobacteria in South African antelopes presenting with tuberculosis-like lesions. Vet Microbiol. 2011; 147:340-345 View ArticlePubMed
  12. Connor KM, Quirie MM, Baird G and Donachie W. Characterization of United Kingdom isolates of Corynebacterium pseudotuberculosis using pulsed-field gel electrophoresis. J Clin Microbiol. 2000; 38:2633-2637PubMed
  13. Ben Saïd MS, Ben Maitigue H, Benzarti M, Messadi L, Rejeb A and Amara A. Epidemiological and clinical studies of ovine caseous lymphadenitis. Arch Inst Pasteur Tunis. 2002; 79:51-57PubMed
  14. Binns SH, Bailey M and Green LE. Postal survey of ovine caseous lymphadenitis in the United Kingdom between 1990 and 1999. Vet Rec. 2002; 150:263-268 View ArticlePubMed
  15. Arsenault J, Girard C, Dubreuil P, Daignault D, Galarneau JR, Boisclair J, Simard C and Belanger D. Prevalence of and carcass condemnation from maedi-visna, paratuberculosis and caseous lymphadenitis in culled sheep from Quebec, Canada. Prev Vet Med. 2003; 59:67-81 View ArticlePubMed
  16. Paton MW, Walker SB, Rose IR and Watt GF. Prevalence of caseous lymphadenitis and usage of caseous lymphadenitis vaccines in sheep flocks. Aust Vet J. 2003; 81:91-95 View ArticlePubMed
  17. Selim SA. Oedematous skin disease of buffalo in Egypt. J Vet Med B Infect Dis Vet Public Health. 2001; 48:241-258 View ArticlePubMed
  18. Pinheiro RRGA, Alves FSF and Haddad JP. Aspectos epidemiológicos da caprinocultura cearense. Arquivo Brasileiro Med Veterinaria Zootecnia. 2000; 52:10 View Article
  19. Barakat AASSA, Atef A, Saber MS and Nafie EK. Two serotypes of Corynebacterium pseudotuberculosis isolated from different animal species. Revue Scientifique et Technique Office International des Epizooties. 1984; 3:151-163
  20. Hard GC. Electron microscopic examination of Corynebacterium ovis. J Bacteriol. 1969; 97:1480-1485PubMed
  21. Jones DCM. Irregular, nonsporing Gram-positive rods. In: Smeath PHA, Mair NS, Sharpe ME, Holt JG (eds), Bergey's Manual of Systematic Bacteriology. Williams and Wilkins, Baltimore; 1986, p. 1261.
  22. Buxton A, Fraser G. Corynebacterium. In: Buxton A, Fraser G (eds) Animal Microbiology. Blackwell Scientific Publications, Edinburgh, 1977, p. 177.
  23. Muckle CA and Gyles CL. Characterization of strains of corynebacterium pseudotuberculosis. Can J Comp Med. 1982; 46:206-208PubMed
  24. Biberstein EL, Knight HD and Jang S. Two biotypes of Corynebacterium pseudotuberculosis. Vet Rec. 1971; 89:691-692 View ArticlePubMed
  25. Bayan N, Houssin C, Chami M and Leblon G. Mycomembrane and S-layer: two important structures of Corynebacterium glutamicum cell envelope with promising biotechnology applications. J Biotechnol. 2003; 104:55-67 View ArticlePubMed
  26. De Sousa-D'Auria C, Kacem R, Puech V, Tropis M, Leblon G, Houssin C and Daffe M. New insights into the biogenesis of the cell envelope of corynebacteria: identification and functional characterization of five new mycoloyltransferase genes in Corynebacterium glutamicum. FEMS Microbiol Lett. 2003; 224:35-44 View ArticlePubMed
  27. Puech V, Chami M, Lemassu A, Laneelle MA, Schiffler B, Gounon P, Bayan N, Benz R and Daffe M. Structure of the cell envelope of corynebacteria: importance of the non-covalently bound lipids in the formation of the cell wall permeability barrier and fracture plane. Microbiology. 2001; 147:1365-1382PubMed
  28. Gebhardt H, Meniche X, Tropis M, Kramer R, Daffe M and Morbach S. The key role of the mycolic acid content in the functionality of the cell wall permeability barrier in Corynebacterineae. Microbiology. 2007; 153:1424-1434 View ArticlePubMed
  29. Songer JG, Beckenbach K, Marshall MM, Olson GB and Kelley L. Biochemical and genetic characterization of Corynebacterium pseudotuberculosis. Am J Vet Res. 1988; 49:223-226PubMed
  30. Tamura K, Peterson D, Peterson N, Stecher G, Nei M and Kumar S. MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol. 2011; 28:2731-2739 View ArticlePubMed
  31. Field D, Garrity G, Gray T, Morrison N, Selengut J, Sterk P, Tatusova T, Thomson N, Allen MJ and Angiuoli SV. The minimum information about a genome sequence (MIGS) specification. Nat Biotechnol. 2008; 26:541-547 View ArticlePubMed
  32. Woese CR, Kandler O and Wheelis ML. Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc Natl Acad Sci USA. 1990; 87:4576-4579 View ArticlePubMed
  33. Garrity GM, Holt JG. The Road Map to the Manual. In: Garrity GM, Boone DR, Castenholz RW (eds), Bergey's Manual of Systematic Bacteriology, Second Edition, Volume 1, Springer, New York, 2001, p. 119-169.
  34. Stackebrandt E, Rainey FA and Ward-Rainey NL. Proposal for a new hierarchic classification system, Actinobacteria classis nov. Int J Syst Bacteriol. 1997; 47:479-491 View Article
  35. Skerman VBD, McGowan V and Sneath PHA. Approved Lists of Bacterial Names. Int J Syst Bacteriol. 1980; 30:225-420 View Article
  36. Buchanan RE. Studies in the nomenclature and classification of bacteria. II. The primary subdivisions of the Schizomycetes. J Bacteriol. 1917; 2:155-164PubMed
  37. Zhi XY, Li WJ and Stackebrandt E. An update of the structure and 16S rRNA gene sequence-based definition of higher ranks of the class Actinobacteria, with the proposal of two new suborders and four new families and emended descriptions of the existing higher taxa. Int J Syst Evol Microbiol. 2009; 59:589-608 View ArticlePubMed
  38. Lehmann KB, Neumann R. Lehmann's Medizin, Handatlanten. X Atlas und Grundriss der Bakteriologie und Lehrbuch der speziellen bakteriologischen Diagnostik., Fourth Edition, Volume 2, J.F. Lehmann, München, 1907, p. 270.
  39. Bernard KA, Wiebe D, Burdz T, Reimer A, Ng B, Singh C, Schindle S and Pacheco AL. Assignment of Brevibacterium stationis (ZoBell and Upham 1944) Breed 1953 to the genus Corynebacterium, as Corynebacterium stationis comb. nov., and emended description of the genus Corynebacterium to include isolates that can alkalinize citrate. Int J Syst Evol Microbiol. 2010; 60:874-879 View ArticlePubMed
  40. Eberson F. A bacteriologic study of the diphtheroid organisms with special reference to Hodgkin's disease. J Infect Dis. 1918; 23:1-42
  41. Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, Davis AP, Dolinski K, Dwight SS and Eppig JT. Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat Genet. 2000; 25:25-29 View ArticlePubMed
  42. Khamis A, Raoult D and La Scola B. rpoB gene sequencing for identification of Corynebacterium species. J Clin Microbiol. 2004; 42:3925-3931 View ArticlePubMed
  43. Khamis A, Raoult D and La Scola B. Comparison between rpoB and 16S rRNA gene sequencing for molecular identification of 168 clinical isolates of Corynebacterium. J Clin Microbiol. 2005; 43:1934-1936 View ArticlePubMed
  44. Liolios K, Chen IM, Mavromatis K, Tavernarakis N, Hugenholtz P, Markowitz VM and Kyrpides NC. The Genomes On Line Database (GOLD) in 2009: status of genomic and metagenomic projects and their associated metadata. Nucleic Acids Res. 2010; 38(Database issue):D346-D354 View ArticlePubMed
  45. Cerdeira LT, Carneiro AR, Ramos RT, de Almeida SS, D'Afonseca V, Schneider MP, Baumbach J, Tauch A, McCulloch JA and Azevedo VA. Rapid hybrid de novo assembly of a microbial genome using only short reads: Corynebacterium pseudotuberculosis I19 as a case study. J Microbiol Methods. 2011; 86:218-223 View ArticlePubMed
  46. Mellmann A, Harmsen D, Cummings CA, Zentz EB, Leopold SR, Rico A, Prior K, Szczepanowski R, Ji Y and Zhang W. Prospective genomic characterization of the German enterohemorrhagic Escherichia coli O104:H4 outbreak by rapid next generation sequencing technology. PLoS ONE. 2011; 6:e22751 View ArticlePubMed
  47. Delcher AL, Harmon D, Kasif S, White O and Salzberg SL. Improved microbial gene identification with GLIMMER. Nucleic Acids Res. 1999; 27:4636-4641 View ArticlePubMed
  48. Lagesen K, Hallin P, Rodland EA, Staerfeldt HH, Rognes T and Ussery DW. RNAmmer: consistent and rapid annotation of ribosomal RNA genes. Nucleic Acids Res. 2007; 35:3100-3108 View ArticlePubMed
  49. Lowe TM and Eddy SR. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res. 1997; 25:955-964PubMed
  50. Benson G. Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Res. 1999; 27:573-580 View ArticlePubMed
  51. Zdobnov EM and Apweiler R. InterProScan--an integration platform for the signature-recognition methods in InterPro. Bioinformatics. 2001; 17:847-848 View ArticlePubMed
  52. Rutherford K, Parkhill J, Crook J, Horsnell T, Rice P, Rajandream MA and Barrell B. Artemis: sequence visualization and annotation. Bioinformatics. 2000; 16:944-945 View ArticlePubMed
  53. Karp PD, Paley S and Romero P. The Pathway Tools software. Bioinformatics. 2002; 18(Suppl 1):S225-S232 View ArticlePubMed
  54. Caspi R, Foerster H, Fulcher CA, Kaipa P, Krummenacker M, Latendresse M, Paley S, Rhee SY, Shearer AG and Tissier C. The MetaCyc Database of metabolic pathways and enzymes and the BioCyc collection of Pathway/Genome Databases. Nucleic Acids Res. 2007; 36(Database issue):D623-D631 View ArticlePubMed
  55. Karp PD, Ouzounis CA, Moore-Kochlacs C, Goldovsky L, Kaipa P, Ahren D, Tsoka S, Darzentas N, Kunin V and Lopez-Bigas N. Expansion of the BioCyc collection of pathway/genome databases to 160 genomes. Nucleic Acids Res. 2005; 33:6083-6089 View ArticlePubMed