Non contiguous-finished genome sequence and description of Peptoniphilus obesi sp. nov.
© The Author(s) 2013
Published: 25 February 2013
Peptoniphilus obesi strain ph1T sp. nov., is the type strain of P. obesi sp. nov., a new species within the genus Peptoniphilus. This strain, whose genome is described here, was isolated from the fecal flora of a 26-year-old woman suffering from morbid obesity. P. obesi strain ph1T is a Gram-positive, obligate anaerobic coccus. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 1,774,150 bp long genome (1 chromosome but no plasmid) contains 1,689 protein-coding and 29 RNA genes, including 5 rRNA genes.
KeywordsPeptoniphilus obesi genome
Peptoniphilus obesi strain ph1T (=CSUR=P187, =DSM =25489) is the type strain of P. obesi sp. nov. This bacterium is a Gram-positive, anaerobic, indole-negative coccus that was isolated from the stool of a 26-year-old woman suffering from morbid obesity and is part of a study aiming at cultivating all species within human feces, individually .
Widespread use of gene sequencing, notably 16SrRNA, for the identification of bacteria recovered from clinical specimens, has enabled the description of a great number of bacterial species and genera of clinical importance [2,3]. The recent development of high throughput genome sequencing and mass spectrometric analyses has provided unprecedented access to a wealth of genetic and proteomic information .
The current classification of prokaryotes, known as polyphasic taxonomy, relies on a combination of phenotypic and genotypic characteristics . However, as more than 3,000 bacterial genomes have been sequenced  and the cost of genomic sequencing is decreasing, we recently proposed to integrate genomic information in addition to their main phenotypic characteristics (habitat, Gram-stain reaction, culture and metabolic characteristics, and when applicable, pathogenicity) in the description of new bacterial species [7–18].
The commensal microbiota of humans and animals consists, in part, of many Gram-positive anaerobic cocci. These bacteria are also commonly associated with a variety of human infections . Extensive taxonomic changes have occurred among this group of bacteria, especially in clinically-important genera such as Finegoldia, Parvimonas, and Peptostreptococcus . Members of genus Peptostreptococcus were divided into three new genera, Peptoniphilus, Anaerococcus and Gallicola by Ezaki . The genus Peptoniphilus currently contains eight species that produce butyrate, are non-saccharolytic and use peptone and amino acids as major energy sources: P. asaccharolyticus, P. harei, P. indolicus, P. ivorii, P. lacrimalis , P. gorbachii, P. olsenii, and P. methioninivorax [21,22].
Members of the genus Peptoniphilus have been isolated mainly from various human clinical specimens such as vaginal discharges, ovarian, peritoneal, sacral and lachrymal gland abscesses . In addition, P. indolicus causes summer mastitis in cattle .
Here we present a summary classification and a set of features for P. obesi sp. nov. strain ph1T (CSUR=P187, DSM=25489) together with the description of the complete genomic sequence and its annotation. These characteristics support the circumscription of the species P. obesi.
Classification and features
Classification and general features of Peptoniphilus obesi strain ph1T according to the MIGS recommendations 
Family Clostridiales family XI Incertae sedis
Species Peptoniphilus obesi
Type strain ph1T
Sample collection time
0 m above sea level
Strain ph1T exhibited neither catalase nor oxidase activities. Using the API rapid ID 32A system (BioMérieux), positive reactions were observed for arginine arylamidase and leucine arylamidase. Negative reactions were found for urease, nitrate reduction, arginine dihydrolase, indole production, α-arabinosidase, α-glucosidase, α-fucosidase, β-galactosidase, glutamic acid decarboxylase, 6-phospho-β-galactosidase β-glucosidase, β-glucuronidase, N-acetyl-β-glucosaminidase, D-mannose, D-raffinose, alkaline phosphatase, alanine arylamidase, glutamyl glutamic acid arylamidase, glycine arylamidase, histidine arylamidase, leucyl glycine arylamidase, phenylalanine arylamidase, proline arylamidase, pyroglutamic acid arylamidase, serine arylamidase and tyrosine arylamidase. P. obesi is susceptible to penicillin G, amoxicillin, amoxicillin + clavulanic acid, imipenem, nitrofurantoin, erythromycin, doxycyclin, rifampicine, vancomycin, gentamicin 500, metronidazole and resistant to ceftriaxon, ciprofloxacin, gentamicin 10 and trimetoprim + sulfamethoxazole.
Differential characteristics of P. obesi sp. nov strain ph1T, Peptoniphilus grossensis strain ph5 T, Peptoniphilus timonensis strain JC401T and Peptoniphilus gorbachii WAL 10418T.
Cell diameter (µm)
Glutamyl glutamic acid arylamidase
Genome sequencing and annotation
Genome project history
454 GS paired-end 3-kb library
454 GS FLX Titanium
Newbler version 2.5.3
Gene calling method
Genbank Date of Release
May 30, 2012
NCBI project ID
Study of the human gut microbiome
Growth conditions and DNA isolation
P. obesi sp. nov. strain ph1T(CSUR=P187, =DSM=25489), was grown anaerobically on 5% sheep blood-enriched BHI agar at 37°C. Four petri dishes were spread and resuspended in 3x500 µl of TE buffer and stored at 80°C. Then, 500 µl of this suspension were thawed, centrifuged for 3 minutes at 10,000 rpm and resuspended in 3×100µL of G2 buffer (EZ1 DNA Tissue kit, Qiagen). A first mechanical lysis was performed by glass powder on the Fastprep-24 device (Sample Preparation system, MP Biomedicals, USA) using 2×20 seconds cycles. DNA was then treated with 2.5 µg/µL lysozyme (30 minutes at 37°C) and extracted using the BioRobot EZ1 Advanced XL (Qiagen). The DNA was then concentrated and purified using the Qiamp kit (Qiagen). The yield and the concentration was measured by the Quant-it Picogreen kit (Invitrogen) on the Genios Tecan fluorometer at 37.2 ng/µl.
Genome sequencing and assembly
DNA (5 µg) was mechanically fragmented on a Hydroshear device (Digilab, Holliston, MA, USA) with an enrichment size of 3–4kb. DNA fragmentation was visualized through an Agilent 2100 BioAnalyzer on a DNA labchip 7500 with an optimal size of 3.287kb. The library was constructed according to the 454 GS FLX Titanium paired-end protocol. Circularization and nebulization were performed and generated a pattern with an optimum at 665 bp. After PCR amplification through 15 cycles followed by double size selection, the single stranded paired end library was then quantified on the Quant-it Ribogreen kit (Invitrogen) on the Genios Tecan fluorometer at 72 pg/µL. The library concentration equivalence was calculated as 1.99E+08 molecules/µL. The library was stored at −20°C until further use.
The shotgun library was clonally amplified with 0.5 cpb and 1 cpb in 2 SV-emPCR reactions per condition, with the GS Titanium SV emPCR Kit (Lib-L) v2 (Roche). The yield of the emPCR was 9.2% for 0.5 cpb and 12% for 1 cpb in the range of 5 to 20% from the Roche procedure. Approximately 790,000 beads were loaded on 1/4 region of a GS Titanium PicoTiterPlate PTP Kit 70×75 and sequenced with the GS FLX Titanium Sequencing Kit XLR70 (Roche). The run was performed overnight and then analyzed on the cluster through the gsRunBrowser and Newbler assembler (Roche). A total of 228,882 passed filter wells were obtained and generated 76.8Mb of DNA sequence with a average length of 336 bp. The global passed filter sequences were assembled using Newbler with 90% identity and 40 bp as overlap. The final assembly identified 5 scaffolds and 32 large contigs (>1,500 bp) generating a genome size of 1.7 Mb.
Open Reading Frames (ORFs) were predicted using Prodigal  with default parameters but the predicted ORFs were excluded if they spanned a sequencing gap region. The predicted bacterial protein sequences were searched against the GenBank database  and the Clusters of Orthologous Groups (COG) databases using BLASTP. The tRNAScanSE tool  was used to find tRNA genes, whereas ribosomal RNAs were found by using RNAmmer  and BLASTN against the GenBank database. Signal peptides and numbers of transmembrane helices were predicted using SignalP  and TMHMM , respectively. ORFans were identified if their BLASTP E-value was lower than 1e-03 for alignment length greater than 80 amino acids. If alignment lengths were smaller than 80 amino acids, we used an E-value of 1e-05. To estimate the mean level of nucleotide sequence similarity at the genome level between Peptoniphilus obesi and other members of the Peptoniphilus genera, we compared genomes two by two and determined the mean percentage of nucleotide sequence identity among orthologous ORFs using BLASTn Orthologous genes were detected using the Proteinortho software .
Nucleotide content and gene count levels of the genome
% of totala
Genome size (bp)
DNA coding region (bp)
G+C content (bp)
Number of replicons
Genes with function prediction
Genes assigned to COGs
Genes with peptide signals
Genes with transmembrane helices
Number of genes associated with the 25 general COG functional categories
% age a
RNA processing and modification
Replication, recombination and repair
Chromatin structure and dynamics
Cell cycle control, mitosis and meiosis
Signal transduction mechanisms
Cell wall/membrane biogenesis
Intracellular trafficking and secretion
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
Not in COGs
Comparison with the genomes from other Peptoniphilus species
Here, we compared the genome sequence of P. obesi strain ph1T with those of P. harei strain ACS-146-V-Sch2b, P. lacrimalis strain 315-B, Peptoniphilus senegalensis JC140T, Peptoniphilus timonensis JC401T, Peptoniphilus grossensis ph5 T and Peptoniphilus indolicus strain ATCC BAA-1640.
Number of orthologous genes (upper right) and average nucleotide identity levels (lower left) between pairs of genomes determined using the Proteinortho software .
On the basis of phenotypic (Table 2), phylogenetic and genomic analyses (Table 6), we formally propose the creation of Peptoniphilus obesi sp. nov. that contains the strain ph1T. This strain has been found in Marseille, France.
Description of Peptoniphilus obesi sp. nov.
Peptoniphilus obesi (o.be.si. L. masc. gen. adj. obesi of an obese, the disease presented by the patient from whom the type strain ph1T was isolated).
Colonies are 0.4 mm in diameter on blood-enriched Columbia agar and stain gray, transparent, opaque, colonies are not bright. Cells are coccoid, diameter range from 0.77µm to 0.93 µm with a mean diameter of 0.87 µm. Optimal growth is achieved anaerobically. No growth is observed in aerobic conditions. Growth occurs between 30–45°C, with optimal growth observed at 37°C, on blood-enriched Columbia agar. Cells stain Gram-positive, are non endospore-forming, and non-motile. Arginine arylamidase and leucine arylamidase activities are present. Cells are negative for the following activities: catalase, oxidase, urease, nitrate reduction, arginine dihydrolase, indole production, α-arabinosidase, α-glucosidase, α-fucosidase, β-galactosidase, glutamic acid decarboxylase, 6-phospho-β-galactosidase β-glucosidase, β-glucuronidase, N-acetyl-β-glucosaminidase, D-mannose, D-raffinose, alkaline phosphatase, alanine arylamidase, glutamyl glutamic acid arylamidase, glycine arylamidase, histidine arylamidase, leucyl glycine arylamidase, phenylalanine arylamidase, proline arylamidase, pyroglutamic acid arylamidase, serine arylamidase and tyrosine arylamidase. Cells are susceptible to penicillin G, amoxicillin, amoxicillin + clavulanic acid, imipenem, nitrofurantoin, erythromycin, doxycycline, rifampicine, vancomycin, gentamicin 500, metronidazole and resistant to ceftriaxone, gentamicin 10, ciprofloxacin and trimethoprim + sulfamethoxazole.
The G+C content of the genome is 30.1%. The 16S rRNA and genome sequences are deposited in GenBank under accession numbers CAHB00000000 and JN837495, respectively. The type strain ph1T (= CSUR P187 = DSM 25489) was isolated from the fecal flora of an obese French patient.
- Lagier JC, Armougom F, Million M, Hugon P, Pagnier I, Robert C, Bittar F, Fournous G, Gimenez G, Maraninchi M, et al. Microbial culturomics: paradigm shift in the human gut microbiome study. Clin Microbiol Infect 2012; 18:1185–1193. PubMedView ArticlePubMedGoogle Scholar
- Clarridge JE. Impact of 16S rRNA gene sequence analysis for identification of bacteria on clinical microbiology and infectious diseases. Clin Microbiol Rev 2004; 17:840–862. PubMed 0.1128/CMR.17.4.840-862.2004PubMed CentralView ArticlePubMedGoogle Scholar
- Stackebrandt E, Ebers J. Taxonomic parameters revisited: tarnished gold standards. Microbiol Today 2006; 33:152–155.Google Scholar
- Welker M, Moore ER. Applications of whole-cell matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry in systematic microbiology. Syst Appl Microbiol 2011; 34:2–11. PubMed http://dx.doi.org/10.1016/j.syapm.2010.11.013View ArticlePubMedGoogle Scholar
- Genome Online Database. http://www.genomesonline.org/cgi->bin/GOLD/index.cgi
- Tindall BJ, Rosselló-Móra R, Busse HJ, Ludwig W, Kämpfer P. Notes on the characterization of prokaryote strains for taxonomic purposes. Int J Syst Evol Microbiol 2010; 60:249–266. PubMed http://dx.doi.org/10.1099/ijs.0.016949-0View ArticlePubMedGoogle Scholar
- Kokcha S, Mishra AK, Lagier JC, Million M, Leroy Q, Raoult D, Fournier PE. Non contiguous-finished genome sequence and description of Bacillus timonensis sp. nov. Stand Genomic Sci 2012; 6:346–355.http://dx.doi.org/10.4056/sigs.2776064PubMed CentralView ArticlePubMedGoogle Scholar
- Lagier JC, El Karkouri K, Nguyen TT, Armougom F, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Anaerococcus senegalensis sp. nov. Stand Genomic Sci 2012; 6:116–125. PubMed http://dx.doi.org/10.4056/sigs.2415480PubMed CentralView ArticlePubMedGoogle Scholar
- Mishra AK, Gimenez G, Lagier JC, Robert C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Alistipes senegalensis sp. nov. Stand Genomic Sci 2012; 6:304–314. http://dx.doi.org/10.4056/sigs.2625821View ArticleGoogle Scholar
- Lagier JC, Armougom F, Mishra AK, Ngyuen TT, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Alistipes timonensis sp. nov. Stand Genomic Sci 2012; 6:315–324.http://dx.doi.org/10.4056/sigs.2685917PubMed CentralView ArticlePubMedGoogle Scholar
- Mishra AK, Lagier JC, Robert C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Clostridium senegalense sp. nov. Stand Genomic Sci 2012; 6:386–395.PubMed CentralPubMedGoogle Scholar
- Mishra AK, Lagier JC, Robert C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Peptoniphilus timonensis sp. nov. Stand Genomic Sci 2012; 7:1–11. http://dx.doi.org/10.4056/sigs.2956294PubMed CentralView ArticlePubMedGoogle Scholar
- Mishra AK, Lagier JC, Rivet R, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Paenibacillus senegalensis sp. nov. Stand Genomic Sci 2012; 7:70–81. http://dx.doi.org/10.4056/sigs.3054650PubMed CentralView ArticlePubMedGoogle Scholar
- Lagier JC, Gimenez G, Robert C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Herbaspirillum massiliense sp. nov. Stand Genomic Sci 2012; 7:200–209.http://dx.doi.org/10.4056/sigs.3086474PubMed CentralPubMedGoogle Scholar
- Roux V, El Karkouri K, Lagier JC, Robert C, Raoult D. Non-contiguous finished genome sequence and description of Kurthia massiliensis sp. nov. Stand Genomic Sci 2012; 7:221–232. http://dx.doi.org/10.4056/sigs.3206554PubMed CentralView ArticlePubMedGoogle Scholar
- Kokcha S, Ramasamy D, Lagier JC, Robert C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Brevibacterium senegalense sp. nov. Stand Genomic Sci 2012; 7:233–245. http://dx.doi.org/10.4056/sigs.3256677PubMed CentralView ArticlePubMedGoogle Scholar
- Ramasamy D, Kokcha S, Lagier JC, N’Guyen TT, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Aeromicrobium massilense sp. nov. Stand Genomic Sci 2012; 7:246–257. http://dx.doi.org/10.4056/sigs.3306717PubMed CentralView ArticlePubMedGoogle Scholar
- Lagier JC, Ramasamy D, Rivet R, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Cellulomonas massiliensis sp. nov. Stand Genomic Sci 2012;7:258–270. http://dx.doi.org/10.4056/sigs.3316719PubMed CentralView ArticlePubMedGoogle Scholar
- Jousimies-Somer HP, Summanen DM, Citron EJ, Baron HM. Wexler, Finegold SM. Wadsworth-KTL anaerobic bacteriology manual, 6th ed. Star Publishing, Belmont, 2002.Google Scholar
- Ezaki T, Kawamura Y, Li N, Li ZY, Zhao L, Shu S. Proposal of the genera Anaerococcus gen. nov., Peptoniphilus gen. nov. and Gallicola gen. nov. for members of the genus Peptostreptococcus. Int J Syst Evol Microbiol 2001; 51:1521–1528. PubMed http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=11491354&dopt=AbstractView ArticlePubMedGoogle Scholar
- Song Y, Liu C, Finegold SM. Peptoniphilus gorbachii sp. nov., Peptoniphilus olsenii sp. nov. and Anaerococcus murdochii sp. nov. isolated from clinical specimens of human origin. J Clin Microbiol 2007; 45:1746–1752. PubMed http://dx.doi.org/10.1128/JCM.00213-07PubMed CentralView ArticlePubMedGoogle Scholar
- Rooney AP, Swezey JL, Pukall R, Schumann P, Spring S. Peptoniphilus methioninivorax sp. nov., a Gram-positive anaerobic coccus isolated from retail ground beef. Int J Syst Evol Microbiol 2011; 61:1962–1967. PubMed http://dx.doi.org/10.1099/ijs.0.024232-0View ArticlePubMedGoogle Scholar
- List of Prokaryotic names with Standing in Nomenclature.Google Scholar
- Field D, Garrity G, Gray T, Morrison N, Selengut J, Sterk P, Tatusova T, Thomson N, Allen MJ, Angiuoli SV, et al. The minimum information about a genome sequence (MIGS) specification.Google Scholar
- Woese CR, Kandler O, 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. PubMed http://dx.doi.org/10.1073/pnas.87.12.4576PubMed CentralView ArticlePubMedGoogle Scholar
- Gibbons NE, Murray RGE. Proposals concerning the higher taxa of Bacteria. Int J Syst Bacteriol 1978; 28:1–6.http://dx.doi.org/10.1099/00207713-28-1-1View ArticleGoogle Scholar
- 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.View ArticleGoogle Scholar
- Murray RGE. The Higher Taxa, or, a Place for Everything…? In: Holt JG (ed), Bergey’s Manual of Systematic Bacteriology, First Edition, Volume 1, The Williams and Wilkins Co., Baltimore, 1984, p. 31–34.Google Scholar
- List of new names and new combinations previously effectively, but not validly, published. List no. 132. Int J Syst Evol Microbiol 2010; 60:469–472. http://dx.doi.org/10.1099/ijs.0.022855-0
- Rainey FA. Class II. Clostridia class nov. In: De Vos P, Garrity G, Jones D, Krieg NR, Ludwig W, Rainey FA, Schleifer KH, Whitman WB (eds), Bergey’s Manual of Systematic Bacteriology, Second Edition, Volume 3, Springer-Verlag, New York, 2009, p. 736.Google Scholar
- Skerman VBD, Sneath PHA. Approved list of bacterial names. Int J Syst Bact 1980; 30:225–420. http://dx.doi.org/10.1099/00207713-30-1-225View ArticleGoogle Scholar
- Prévot AR. Dictionnaire des bactéries pathogens. In: Hauduroy P, Ehringer G, Guillot G, Magrou J, Prevot AR, Rosset, Urbain A (eds). Paris, Masson, 1953, p. 1–692.Google Scholar
- Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, Davis AP, Dolinski K, Dwight SS, Eppig JT, et al. Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat Genet 2000; 25:25–29. PubMed http://dx.doi.org/10.1038/75556PubMed CentralView ArticlePubMedGoogle Scholar
- Seng P, Drancourta M, Gouriet F, La Scola B, Fournier PF, Rolain JM, Raoult D. Ongoing revolution in bacteriology: routine identification of bacteria by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Clin Infect Dis 2009; 49:543–551. PubMed http://dx.doi.org/10.1086/600885View ArticlePubMedGoogle Scholar
- Prodigal. http://prodigal.ornl.gov
- Benson DA, Karsch-Mizrachi I, Clark K, Lipman DJ, Ostell J, Sayers EW. GenBank. Nucleic Acids Res 2012; 40:D48–D53. PubMed http://dx.doi.org/10.1093/nar/gkr1202PubMed CentralView ArticlePubMedGoogle Scholar
- Lowe TM, Eddy SR. t-RNAscan-SE: a program for imroved detection of transfer RNA gene in genomic sequence. Nucleic Acids Res 1997; 25:955–964. PubMedPubMed CentralView ArticlePubMedGoogle Scholar
- Lagesen K, Hallin P, Rodland EA, Staerfeldt HH, Rognes T, Ussery DW. RNAmmer: consistent and rapid annotation of ribosomal RNA genes. Nucleic Acids Res 2007; 35:3100–3108. PubMed http://dx.doi.org/10.1093/nar/gkm160PubMed CentralView ArticlePubMedGoogle Scholar
- Bendtsen JD, Nielsen H, von Heijne G, Brunak S. Improved prediction of signal peptides: SignalP 3.0. J Mol Biol 2004; 340:783–795. PubMed http://dx.doi.org/10.1016/j.jmb.2004.05.028View ArticlePubMedGoogle Scholar
- Krogh A, Larsson B, von Heijne G, Sonnhammer EL. Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes. J Mol Biol 2001; 305:567–580. PubMed http://dx.doi.org/10.1006/jmbi.2000.4315View ArticlePubMedGoogle Scholar
- Lechner M, Findeib S, Steiner L, Marz M, Stadler PF, Prohaska SJ. Proteinortho: Detection of (Co-)orthologs in large-scale analysis. BMC Bioinformatics 2011; 12:124. PubMed http://dx.doi.org/10.1186/1471-2105-12-124PubMed CentralView ArticlePubMedGoogle Scholar