- Open Access
Non-contiguous finished genome sequence and description of Megasphaera massiliensis sp. nov.
Standards in Genomic Sciences volume 8, pages 525–538 (2013)
Megasphaera massiliensis strain NP3T sp. nov. is the type strain of Megasphaera massiliensis sp. nov., a new species within the genus Megasphaera. This strain, whose genome is described here, was isolated from the fecal flora of an HIV-infected patient. M. massiliensis is a Gram-negative, obligate anaerobic coccobacillus. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 2,661,757 bp long genome (1 chromosome but no plasmid) contains 2,577 protein-coding and 61 RNA genes, including 5 rRNA genes.
Megasphaera massiliensis sp. nov. strain NP3T (= CSUR P245 = DSM 26228) is the type strain of M. massiliensis sp. nov. This bacterium is a Gram-negative, non-sporulating, anaerobic and non-motile coccobacillus that was isolated from the stool of an HIV-infected patient as part of a culturomics study designed to cultivate individually all bacterial species within human feces [1,2].
The current classification of prokaryotes is based on a combination of phenotypic and genotypic characteristics [3,4] including 16S rRNA gene phylogeny, G + C content and DNA-DNA hybridization (DDH). Despite being considered as a “gold standard”, these tools exhibit several drawbacks [5,6]. To date, almost 4,000 bacterial genomes have been sequenced  and the cost of genomic sequencing is constantly decreasing. Therefore, we recently proposed the addition of genomic information to phenotypic criteria, including the protein profile, for the description of new bacterial species [8–29].
The genus Megasphaera (Rogosa 1971), created in 1971 , currently contains 5 species including M. cerevisiae (Engelmann and Weiss 1986) , M. elsdenii (Gutierrez et al. 1959) , M. micronuciformis (Marchandin et al. 2003) , M. paucivorans (Juvonen and Suihko 2006)  and M. sueciensis (Juvonen and Suihko 2006) . The type species, M. elsdenii (Gutierrez et al. 1959) , originally classified in the Peptostreptococcus genus (Gutierrez et al. 1959), was later reclassified within a new genus, Megasphaera (Rogosa 1971), in the family Veillonellaceae (Rogosa 1971) . It is an obligately anaerobic, lactate-fermenting, gastrointestinal microbe of ruminant and non-ruminant mammals, including humans. It was also isolated in a case of human endocarditis . The genome from M. elsdenii strain DSM 20460, isolated from the rumen of sheep, was recently sequenced . M. cerevisiae , M. micronuciformis , M. paucivorans and M. sueciensis  are brewery-associated species. Here we present a summary classification and a set of features for M. massiliensis sp. nov. strain NP3T (= CSUR P245 = DSM 26228) together with the description of the complete genome sequencing and annotation. These characteristics support the circumscription of the species M. massiliensis.
Classification and features
A stool sample was collected from a 32-year-old HIV-infected patient living in Marseille, France. The patient gave written informed consent for the study. The study was approved by the Ethics Committee of the Institut Fédératif de Recherche IFR48, Faculty of Medicine, Marseille, France, under agreement number 09-022.
The fecal specimen was preserved at −80°C after collection. Strain NP3T (Table 1) was isolated in January 2012 by cultivation on 5% sheep blood agar in anaerobic condition at 37°C, following a 7-day preincubation of the stool specimen in an anaerobic blood culture bottle enriched with sterile 5% sheep rumen fluid and 5% sheep blood. The strain exhibited a nucleotide sequence similarity with other members of the genus Megasphaera ranging from 91.5% with M. cerevisiae strain PAT1T to 95.8% with M. elsdenii strain ATCC 25940T, its closest validated phylogenetic neighbor (Figure 1). These values were lower than the 98.7% 16S rRNA gene sequence threshold recommended by Stackebrandt and Ebers to delineate a new species without carrying out DNA-DNA hybridization .
Different growth temperatures (25, 30, 37, 45°C) were tested. Growth occurred between 30 and 45°C, and optimal growth was observed at 37°C. Colonies were transparent and smooth with a diameter of 0.5 to 1 mm on blood-enriched Columbia agar (BioMérieux). Growth of the strain was tested in 5% sheep blood agar (BioMérieux) under anaerobic and microaerophilic conditions using GENbag anaer and GENbag microaer systems, respectively (BioMérieux), and in the presence of air, with or without 5% CO2. Growth only occurred in anaerobic atmosphere. No growth was observed under aerobic conditions and microaerophilic conditions. A motility test was negative. Cells grown on agar are Gram-negative coccobacilli (Figure 2), with a mean diameter of 0.87 µm and the presence of phages in electron microscopy (Figure 3).
Strain NP3T exhibited oxidase, but no catalase activity. Using RAPID 32A identification strips (BioMérieux), positive reactions were observed for α-glucosidase and β-glucosidase. Negative reactions were observed for urease, arginine dihydrolase, α and β-galactosidase, β-galactosidase-6-phosphate, α-arabinosidase, β-glucuronidase, N-acetyl-β-glucosanimidase, mannose and raffinose fermentation, α-fucosidase, alkanine phosphatase, arginine arylamidase, proline arylamidase, leucyl glycine arylamidase, phenylalanine arylamidase, leucine arylamidase, pyroglutamic acid arylamidase, tyrosine arylamidase, alanine arylamidase, glycine arylamidase, histidine arylamidase, glutamyl glutamic acid arylamidase and serine arylamidase. Carbohydrate metabolism was examined using an API 50CH strip (BioMerieux). Positive reactions were observed for potassium gluconate, potassium 5-cetogluconate, aesculin, salicine, N-acetylglucosamine, and arbutine production, and L-arabinose, D-ribose, D-xylose, D-galactose, D-glucose, D-fructose, D-mannose, L-rhamnose, D-mannitol, D-sorbitol, D-celiobiose, D-maltose, D-lactose, D-trehalose, gentiobiose, L-fucose and D-arabitol fermentation. Weak reactions were observed for amygdaline and potassium 2-cetogluconate production, and glycerol and D-arabinose fermentation. Table 2 summarizes the differential phenotypic characteristics of M. massiliensis, M. elsdenii and M. micronuciformis. M. massiliensis strain NP3T was susceptible to amoxicillin, amoxicillin-clavulanic acid, ceftriaxone, imipenem and doxycycline but resistant to vancomycin, erythromycin, rifampicin, trimethoprim-sulfamethoxazole, metronidazole and ciprofloxacin.
Matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) MS protein analysis was carried out as previously described  using a Microflex spectrometer (Brüker Daltonics, Germany). Briefly, a pipette tip was used to pick one isolated bacterial colony from a culture agar plate and spread it as a thin film on a MTP 384 MALDI-TOF target plate (Bruker Daltonics). Twelve distinct deposits were done for strain NP3T from 12 isolated colonies. Each smear was overlaid with 2 µL of matrix solution (a saturated solution of alpha-cyano-4-hydroxycinnamic acid) in 50% acetonitrile, 2.5% tri-fluoracetic acid and allowed to dry for five minutes. Spectra were recorded in the positive linear mode for the mass range from 2,000 to 20,000 Da (parameter settings: ion source 1 (ISI), 20kV; IS2, 18.5 kV; lens, 7 kV). A spectrum was obtained after 675 shots with variable laser power. The time of acquisition was between 30 seconds and 1 minute per spot. The 12 NP3T spectra were imported into the MALDI Bio Typer software (version 2.0, Bruker) and analyzed by standard pattern matching (with default parameter settings) against the main spectra of 3,769 bacteria, including the spectra from M. micronuciformis, Veillonella atypica, V. caviae, V. criceti, V. denticariosi, V. dispar, V. montpellierensis, V. parvula, V. ratti and V. rogosae, that were used as reference data (Figures 4 and 5). The method of identification included the m/z from 3,000 to 15,000 Da. For every spectrum, 100 peaks at most were taken into account and compared with the spectra in the database. The MALDI-TOF score enabled the predictive identification and discrimination of the tested species from those in a database: a score > 2 with a validated species enabled identification at the species level, and a score < 1.7 did not enable any identification. No significant score was obtained for strain NP3T against the Brüker database, suggesting that our isolate was not a member of a known species. We added the spectrum from strain NP3T to our database for future reference (Figure 4). Figure 5 shows the MALDI-TOF MS spectrum differences between M. massiliensis and other Megasphaera and Veillonella species (Figure 5).
Genome sequencing information
Genome project history
The organism was selected for sequencing on the basis of its phenotypic differences, phylogenetic position and 16S rRNA similarity to other members of the genus Megasphaera, and is part of a study of the human digestive flora aiming at isolating all bacterial species within human feces [1,2]. It was the third genome of a Megasphaera species and the first sequenced genome of M. massiliensis sp. nov. The GenBank ID is CAVO00000000 and consists of 106 large contigs. Table 3 shows the project information and its association with MIGS version 2.0 compliance .
Growth conditions and DNA isolation
Megasphaera massiliensis strain NP3T sp. nov. (= CSUR P245 = DSM 26228) was grown anaerobically on 5% sheep blood-enriched agar (BioMérieux) at 37°C. Ten petri dishes were spread and resuspended in 3 × 100µl of G2 buffer (EZ1 DNA Tissue kit, Qiagen). A first mechanical lysis was performed using glass powder on the Fastprep-24 device (MP Biomedicals, Ilkirch, France) during 2 × 20 seconds. DNA was then treated with 2.5 µg/µL lysozyme treatment (30 minutes at 37°C) and extracted using a BioRobot EZ 1 Advanced XL (Qiagen). The DNA was then concentrated and purified using a Qiamp kit (Qiagen). The yield and the concentration were measured using the Quant-it Picogreen kit (Invitrogen) on the Genios_Tecan fluorometer at 82.2 ng/µl.
Genome sequencing and assembly
A paired-end sequencing strategy was used (Roche). The library was pyrosequenced on a GS FLX Titanium sequencere (Roche). This project was loaded on a 1/4 region on PTP Picotiterplate (Roche). Five µg of DNA were mechanically fragmented on the Covaris device (KBioScience-LGC Genomics, Teddington, UK) using miniTUBE-Red 5Kb. The DNA fragmentation was visualized through the Agilent 2100 BioAnalyzer on a DNA labchip 7500 with an optimal size of 3.3 kb. After PCR amplification through 17 cycles followed by double size selection, the single stranded paired-end library was then loaded on a DNA labchip RNA pico 6000 on the BioAnalyzer. The pattern showed an optimum at 613 bp and the concentration was quantified on a Genios Tecan fluorometer at 3.48 pg/µL. The library concentration equivalence was calculated at 5.21E+09 molecules/µL. The library was stored at −20°C until further use, and the library was clonally amplified with 0.5 cpb in 3 emPCR reactions with the GS Titanium SV emPCR Kit (Lib-L) v2 (Roche). The yield of the emPCR was 9.99%, in the range of 5 to 20% from the Roche procedure. Approximately 790,000 beads were loaded on the GS Titanium PicoTiterPlates PTP Kit 70x75 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 186,153 passed filter wells generated 61.97 Mb with a length average of 332 bp. The filter-passed sequences were assembled using Newbler with 90% identity and 40 bp as overlap. The final assembly identified 114 large contigs (>1,500 bp) arranged into 28 scaffolds and generated a genome size of 2.66 Mb, which corresponds to a coverage of 23.3× genome equivalent.
Prodigal  with default parameters was used to predict the Open Reading Frames (ORFs). The predicted ORFs were excluded if they spanned a sequencing gap region. Protein functional assessment was obtained by comparison with sequences in the GenBank  and Clusters of Orthologs Groups (COG) databases using BLASTP. The rRNA and tRNA were identified using RNAmmer  and tRNAscan-SE 1.21  respectively. SignalP  and TMHMM  were used to predict signal peptides and transmembrane helices, 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. Such parameter thresholds have already been used in previous works to define ORFans. Artemis  was used for data management and DNA Plotter  was used for visualization of genomic features. PHAST was used to identify, annotate and graphically display prophage sequences within bacterial genomes or plasmids . To estimate the mean level of nucleotide sequence similarity at the genome level between M. massiliensis and another 5 members of the family Veillonellaceae, orthologous proteins were detected using the Proteinortho software with the following parameters: e-value 1e-5, 30% percentage of identity, 50% coverage and algebraic connectivity of 50% , and genomes compared two by two. For each pair of genomes, we determined the mean percentage of nucleotide sequence identity among orthologous ORFs using BLASTn.
The genome of M. massiliensis strain NP3T is 2,661,757 bp long (in 28 scaffolds, 1 chromosome, and no plasmid) with a 50.2% GC content (Table 3 and Figure 6). Of the 2,577 predicted genes, 2,516 were protein-coding genes and there were 61 RNA genes. A total of 1,697 genes (65.8%) were assigned a putative function. A total of 248 genes (9.6%) were annotated as hypothetical proteins. The properties and the statistics of the genome are summarized in Tables 4 and 5. The distribution of genes into COGs functional categories is presented in Table 5.
Comparison with the genomes from M. elsdenii, Megasphaera species, Veillonella dispar, V. parvula and Anaeroglobus geminatus
The draft genome of M. massiliensis strain NP3T (2.66 Mb) has a larger size than that of M. elsdenii (2.47 Mb), V. parvula (2.13 Mb), V. dispar (2.12 Mb), A. geminatus (1.79 Mb) and M. micronuciformis (1.77 Mb) respectively. M. massiliensis has a lower G + C content (50.2%) than M. elsdenii (52.8%) but higher than V. parvula, V. dispar, M. micronuciformis and A. geminatus (38.6, 38.8, 46.8 and 48.7%, respectively). M. massiliensis (2,516) has more predicted protein-coding genes than M. elsdenii, A. geminatus, V. dispar, V. parvula, and M. micronuciformis (2,219, 2,148, 1,954, 1,844 and 1,774, respectively) (Table 6). In addition, M. massiliensis shared a mean genomic sequence similarity of 81.84, 69.44, 63.68, 62.92 and 70.27% with M. elsdenii, M. micronuciformis, V. dispar, V. parvula and A. geminatus respectively (Table 6).
M. massiliensis harbors two intact bacteriophages. Based on PHAST results, phage 1 of M. massiliensis was most closely related to Clostridium phage phi CD119 whereas phage 2 was most similar to Bacillus phage BCJA1c.
On the basis of phenotypic, phylogenetic and genomic analyses, we formally propose the creation of Megasphaera massiliensis sp. nov. that contains the strain NP3T. This bacterial strain has been found in Marseille, France.
Description of Megasphaera massiliensis sp. nov.
Megasphaera massiliensis (mas.il.ien’sis. L. gen. fem. n. massiliensis, of Massilia, the Latin name of Marseille where was cultivated strain NP3T). It has been isolated from the feces of a 32-year-old HIV-infected French patient.
Colonies were smooth and transparent with 0.5 to 1 mm in diameter on blood-enriched Columbia agar. Optimal growth is only achieved anaerobically and grows between 30 and 45°C, with optimal growth observed at 37°C. The strain is a Gram-negative, non-endospore forming, non motile coccobacillus. Positive for α-glucosidase, β-glucosidase, potassium gluconate, potassium 5-cetogluconate, aesculin, salicine, N-acetylglucosamine, and arbutine production. Positive for L-arabinose, D-ribose, D-xylose, D-galactose, D-glucose, D-fructose, D-mannose, L-rhamnose, D-mannitol, D-sorbitol, D-celiobiose, D-maltose, D-lactose, D-trehalose, gentiobiose, L-fucose and D-arabitol fermentation. Negative for urease, arginine dihydrolase, α and β-galactosidase, β-galactosidase-6-phosphate, α-arabinosidase, β-glucuronidase, N-acetyl-β-glucosanimidase, mannose and raffinose fermentation, α-fucosidase, alkanine phosphatase, arginine arylamidase, proline arylamidase, leucyl glycine arylamidase, phenylalanine arylamidase, leucine arylamidase, pyroglutamic acid arylamidase, tyrosine arylamidase, alanine arylamidase, glycine arylamidase, histidine arylamidase, glutamyl glutamic acid arylamidase and serine arylamidase. Weak reactions observed for amygdaline and potassium 2-cetogluconate production, and glycerol and D-arabinose fermentation. Cells are susceptible to amoxicillin, amoxicillin-clavulanic acid, ceftriaxone, imipenem and doxycycline, but resistant to vancomycin, erythromycin, rifampicin, trimethoprim/sulfamethoxazole, metronidazole, and ciprofloxacin. The G+C content of the genome is 50.2%. The 16S rRNA and genome sequences are deposited in Genbank under accession numbers JX424772 and CAVO00000000, respectively. The type strain NP3T (= CSUR P245 = DSM 26228) was isolated from the fecal flora of an HIV-infected patient in Marseille, France.
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. PubMed
Dubourg G, Lagier JC, Armougom F, Robert C, Hamad I, Brouqui P. The gut microbiota of a patient with resistant tuberculosis is more comprehensively studied by culturomics than by metagenomics. Eur J Clin Microbiol Infect Dis 2013; 32:637–645. PubMed http://dx.doi.org/10.1007/s10096-012-1787-3
Tindall BJ, Rossello-Mora R, Busse HJ, Ludwig W, Kampfer 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.0.016949-0
Stackebrandt E, Ebers J. Taxonomic parameters revisited: tarnished gold standards. Microbiol Today 2006; 33:152–155.
Wayne LG, Brenner DJ, Colwell RR, Grimont PAD, Kandler O, Krichevsky MI, Moore LH, Moore WEC, Murray RGE, Stackebrandt E, et al. Report of the ad hoc committee on reconciliation of approaches to bacterial systematic. Int J Syst Bacteriol 1987; 37:463–464. http://dx.doi.org/10.1099/00207713-37-4-463
Rossello-Mora R. DNA-DNA Reassociation Methods Applied to Microbial Taxonomy and Their Critical Evaluation. In: Stackebrandt E (ed), Molecular Identification, Systematics, and population Structure of Prokaryotes. Springer, Berlin, 2006; p. 23–50.
Genome Online Database. http://genomesonline.org/cgi-bin/GOLD/index.cgi.
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. PubMed http://dx.doi.org/10.4056/sigs.2776064
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.2415480
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.2625821
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. PubMed
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
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. PubMed http://dx.doi.org/10.4056/sigs.2956294
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. PubMed
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. PubMed
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. PubMed http://dx.doi.org/10.4056/sigs.3206554
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. PubMed http://dx.doi.org/10.4056/sigs.3256677
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. PubMed http://dx.doi.org/10.4056/sigs.3306717
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. PubMed http://dx.doi.org/10.4056/sigs.3316719
Lagier JC, El Karkouri K, Rivet R, Couderc C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Senegalemassilia anaerobia sp. nov. Stand Genomic Sci 2013; 7:343–356. http://dx.doi.org/10.4056/sigs.3246665
Mishra AK, Hugon P, Lagier JC, Nguyen TT, Robert C, Couderc C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Peptoniphilus obesi sp. nov. Stand Genomic Sci 2013; 7:357–369. http://dx.doi.org/10.4056/sigs.32766871
Mishra AK, Lagier JC, Nguyen TT, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Peptoniphilus senegalensis sp. nov. Stand Genomic Sci 2013; 7:370–381. http://dx.doi.org/10.4056/sigs.3366764
Lagier JC, El Karkouri K, Mishra AK, Robert C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Enterobacter massiliensis sp. nov. Stand Genomic Sci 2013; 7:399–412. http://dx.doi.org/10.4056/sigs.3396830
Hugon P, Ramasamy D, Lagier JC, Rivet R, Couderc C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Alistipes obesi sp. nov. Stand Genomic Sci 2013; 7:427–439. http://dx.doi.org/10.4056/sigs.3336746
Mishra AK, Hugon P, Robert C, Couderc C, Raoult D, Fournier PE. Non-contiguous finished genome sequence and description of Peptoniphilus grossensis sp. nov. Stand Genomic Sci 2012; 7:320–330. PubMed
Mishra AK, Hugon P, Lagier JC, Nguyen TT, Couderc C, Raoult D, Fournier PE. Non contiguous-finished genome sequence and description of Enorma massiliensis gen. nov., sp. nov., a new member of the Family Coriobacteriaceae. Stand Genomic Sci 2013; 8:290–305. http://dx.doi.org/10.4056/sigs.3426906
Ramasamy D, Lagier JC, Gorlas A, Raoult D, Fournier PE. Non contiguous-finished genome sequence and description of Bacillus massiliosenegalensis sp. nov. Stand Genomic Sci 2013; 8:264–278. http://dx.doi.org/10.4056/sigs.3496989
Ramasamy D, Lagier JC, Nguyen TT, Raoult D, Fournier PE. Non contiguous-finished genome sequence and description of of Dielma fastidiosa gen. nov., sp. nov., a new member of the Family Erysipelotrichaceae. Stand Genomic Sci 2013; 8:336–351. http://dx.doi.org/10.4056/sigs.3567059
Mishra AK, Lagier JC, Robert C, Raoult D, Fournier PE. Genome sequence and description of Timonella senegalensis gen. nov., sp. nov., a new member of the suborder Micrococcinae. Stand Genomic Sci 2013; 8:318–335. http://dx.doi.org/10.4056/sigs.3476977
Rogosa M. Transfer of Peptostreptococcus elsdenii Gutierrez et al. to a New Genus, Megasphaera [M. elsdenii (Gutierrez et al.) comb. nov.]. Int J Syst Bacteriol 1971; 21:187–189. http://dx.doi.org/10.1099/00207713-21-2-187
Engelmann U, Weiss N. Megasphaera cerevisiae sp. nov.: a new Gram-negative obligately anaerobic coccus isolated from spoiled beer. Syst Appl Microbiol 1985; 6:287–290. http://dx.doi.org/10.1016/S0723-2020(85)80033-3
Marchandin H, Jumas-Bilak E, Gay B, Teyssier C, Jean-Pierre H, de Buochberg MS, Carriere C, Carlier JP. Phylogenetic analysis of some Sporomusa sub-branch members isolated from human clinical specimens: description of Megasphaera micronuciformis sp. nov. Int J Syst Bacteriol 2003; 53:547–553. PubMed
Juvonen R, Suihko ML. Megasphaera paucivorans sp. nov., Megasphaera sueciensis sp. nov. and Pectinatus haikarae sp. nov., isolated from brewery samples, and emended description of the genus Pectinatus. Int J Syst Bacteriol 2006; 56:695–702. PubMed
Brancaccio M, Legendre GG. Megasphaera elsdenii endocarditis. J Clin Microbiol 1979; 10:72–74. PubMed
Marx H, Graf AB, Tatto NE, Thallinger GG, Mattanovich D, Sauer M. Genome sequence of the ruminal bacterium Megasphaera elsdenii. J Bacteriol 2011; 193:5578–5579. PubMed http://dx.doi.org/10.1128/JB.05861-11
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. Nat Biotechnol 2008; 26:541–547. PubMed http://dx.doi.org/10.1038/nbt1360
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.87.12.4576
Gibbons NE, Murray RGE. Proposals Concerning the Higher Taxa of Bacteria. Int J Syst Bacteriol 1978; 28:1–6. http://dx.doi.org/10.1099/00207713-28-1-1
Garrity GM, Holt JG. The Road Map to the Manual. In: Garrity GM, Boone DR, Castenholz RW (eds), Bergey’s Manual of Systematic Bacteriolo-gy, Second Edition, Volume 1, Springer, New York, 2001, p. 119–169.
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.
Marchandin H, Teyssier C, Campos J, Jean-Pierre H, Roger F, Gay B, Carlier JP, Jumas-Bilak E. Negativicoccus succinicivorans gen. nov., sp. nov., isolated from human clinical samples, emended description of the family Veillonellaceae and description of Negativicutes classis nov., Selenomonadales ord. nov. and Acidaminococcaceae fam. nov. in the bacterial phylum Firmicutes. Int J Syst Evol Microbiol 2010; 60:1271–1279. PubMed http://dx.doi.org/10.1099/ijs.0.013102-0
Rogosa M. Transfer of Veillonella Prévot and Acidaminococcus Rogosa from Neisseriaceae to Veillonellaceae fam. nov. and the inclusion of Megasphaera Rogosa in Veillonellaceae. Int J Syst Bacteriol 1971; 21:231–233. http://dx.doi.org/10.1099/00207713-21-3-231
Skerman VBD, McGowan V, Sneath PHA. Approved Lists of Bacterial Names. Int J Syst Bacteriol 1980; 30:225–420. http://dx.doi.org/10.1099/00207713-30-1-225
Rogosa M. Genus III. Megasphaera Rogosa 1971, 187. In: Buchanan RE, Gibbons NE (eds), Bergey’s Manual of Determinative Bacteriology, Eighth Edition, The Williams and Wilkins Co., Baltimore, 1974, p. 448–449.
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/10.1038/75556
Seng P, Drancourt M, Gouriet F, La Scola B, Fournier PE, 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/10.1086/600885
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. Nat Biotechnol 2008; 26:541–547. PubMed http://dx.doi.org/10.1038/nbt1360
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/gkr1202
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/gkm160
Lowe TM, Eddy SR. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res 1997; 25:955–964. PubMed
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.2004.05.028
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.2000.4315
Rutherford K, Parkhill J, Crook J, Horsnell T, Rice P, Rajandream MA, Barrell B. Artemis: sequence visualization and annotation. Bioinformatics 2000; 16:944–945. PubMed http://dx.doi.org/10.1093/bioinformatics/16.10.944
Carver T, Thomson N, Bleasby A, Berriman M, Parkhill J. DNAPlotter: circular and linear interactive genome visualization. Bioinformatics 2009; 25:119–120. PubMed http://dx.doi.org/10.1093/bioinformatics/btn578
Zhou Y, Liang Y, Lynch KH, Dennis JJ, Wishart DS. PHAST: a fast phage search tool. Nucleic Acids Res 2011; 39:W347–W352. PubMed http://dx.doi.org/10.1093/nar/gkr485
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/10.1186/1471-2105-12-124
The authors thank the Xegen Company (www.xegen.fr) for automating the genomic annotation process. This study was funded by the Mediterranee-Infection Foundation.
About this article
Cite this article
Padmanabhan, R., Lagier, JC., Dangui, N.P.M. et al. Non-contiguous finished genome sequence and description of Megasphaera massiliensis sp. nov.. Stand in Genomic Sci 8, 525–538 (2013). https://doi.org/10.4056/sigs.4077819
- Megasphaera massiliensis