- Email: [email protected]
‘Colidextribacter massiliensis’ gen. nov., sp. nov., isolated from human right colon
Accepted Manuscript “Colidextribacter massiliensis” gen. nov., sp. nov. isolated from human right colon D. Ricaboni, M. Mailhe, F. Cadoret, V. Vitton, P.-E. Fournier, D. Raoult PII:
S2052-2975(16)30135-4
DOI:
10.1016/j.nmni.2016.11.023
Reference:
NMNI 266
To appear in:
New Microbes and New Infections
Received Date: 25 October 2016 Revised Date:
20 November 2016
Accepted Date: 23 November 2016
Please cite this article as: Ricaboni D, Mailhe M, Cadoret F, Vitton V, Fournier P-E, Raoult D, “Colidextribacter massiliensis” gen. nov., sp. nov. isolated from human right colon, New Microbes and New Infections (2016), doi: 10.1016/j.nmni.2016.11.023. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT “Colidextribacter massiliensis” gen. nov., sp. nov. isolated from human right colon.
Running title : Colidextribacter massiliensis.
1
RI PT
D. Ricaboni1-2,M. Mailhe1,F. Cadoret1, V. Vitton3, P.-E. Fournier1, D. Raoult1* Aix-Marseille Université, URMITE, UM63, CNRS7278, IRD198, Inserm 1095, Institut
Hospitalo-Universitaire Méditerranée-Infection, Faculté de médecine, 27 Boulevard Jean
2
SC
Moulin, 13385, Marseille cedex 05, France
Department of Biomedical and Clinical Sciences, 3rd Division of Clinical Infectious
3
Service de Gastroenterologie, Hopital Nord, Assistance Publique-Hopitaux de Marseille,
13915 Marseille, France.
Corresponding author. E-mail address: [email protected]
TE D
*
M AN U
Disease, University of Milan, Luigi Sacco Hospital, Italy.
AC C
EP
Keywords: Culturomics, taxono-genomics, Colidextribacter massiliensis, human gut.
ACCEPTED MANUSCRIPT
Abstract:
2
We report here the main characteristics of “Colidextribacter massiliensis” strain Marseille-P3083T
3
(CSURP3083) that was isolated from a human right-colon lavage sample.
AC C
EP
TE D
M AN U
SC
RI PT
1
ACCEPTED MANUSCRIPT
In 2016, as a part of culturomics study of the human microbiome [1], we isolated from the right
5
colon of a 27year-old obese patient a bacterial strain that could not be identified by our systematic
6
matrix-assisted laser desorption-ionization time-of-flight mass spectrometry screening on a
7
Microflex spectrometer (Bruker Daltonics, Brenen, Germany) [2]. The patient gave a signed
8
informed consent and the study was validated by the ethics committee of the Institut Fédératif de
9
Recherche IFR48 under number 2016-010.
RI PT
4
10
Strain Marseille-P3083T
11
(bioMérieux, Marcy l’Etoile, France) in anaerobic atmosphere (anaeroGEN, Oxoid Ltd, Dardilly,
12
France) after a 30-day enrichment of the fresh right-colon sample in an anaerobic haemoculture
13
bottle (Becton Dickinson, Pont de Claix, France) added with 5mL of sheep blood (bioMérieux) and
14
5 mL of 0.2µm filtered (Thermo Fisher Scientific, Villebon-sur-Yvette, France) rumen at 37°C.
15
After 96 hours of anaerobic incubation on 5% sheep blood-enriched agar (bioMérieux) colonies
16
were punctiform, approximatively round, translucid, with a mean diameter of 0.1-0.3 mm. Bacterial
17
cells are small Gram-negative cocci, ranging in length from 0.4 to 0.6 µm. Strain Marseille-P3083T
18
tested catalase and oxidase negative. Sporulation test (20 min at 80°C) resulted negative, no growth
19
was achived under aerobic or microaerophilic (campyGen, Oxoid) conditions. The 16S rRNA gene
20
was sequenced using fD1-rP2 primers as previously described [3], using a 3130-XL sequencer
21
(Applied Biosciences, Saint Aubin, France). Strain Marseille-P3083T exhibited a 93.57 % sequence
22
identity with Pseudoflavonifractor capillosus strain ATCC 29799T (Genbank accession number
23
AY136666), the phylogenetically closest species with standing nomenclature (Fig. 1). This value
24
putatively classifies strain Marseille-P3083T as a member of a new genus within the Clostridiales
25
cluster IV in the phylum Firmicutes. Clostridiales cluster IV was created in 1994 and collect
26
microrganisms that exhibit Clostridium and non Clostridium-like characteristics [4], including
27
Gram-negative microrganisms [5]. Intestinimonas butyriciproducens DSM 26588T (16S rRNA
28
similarity of 93.42%) is another phylogenetically very close species (Fig. 1) that exhibits the
29
peculiar characteristic of appearing as a Gram-negative rod while elecron microscopy demonstrated
AC C
EP
TE D
M AN U
SC
growth was obtained on 5% sheep blood-Columbia agar medium
ACCEPTED MANUSCRIPT
30
Gram-positive cell wall characteristics [6].
31
On the basis of the morphologic differences (morphology and Gram stain) and the 16S rRNA
32
sequence divergence of strain Marseille-P3083T with the phylogenetically closest species with
33
standing in nomenclature [7-8] we propose here the creation of the new genus Colidextribacter
34
(Co.li.dex.tri.bac.ter,
35
bacter a rod ; N.L. masc. n. Colidextribacter, a rod from the right colon); and a new species
36
Colidextribacter massiliensis for which Marseille-P3083T (=CSURP3083) is the type strain.
37
Strain Marseille-P3083T is the type strain of Colidextribacter massiliensis gen. nov., sp. nov.,
38
(mas.si.li.en’sis, L., masc. adj., massiliensis for Massilia, the Roman name of Marseille).
SC
RI PT
L. neut. n. colon or colum, the colon; L. adj. dexter right; N.L. masc. n.
M AN U
39
The MALDITOF spectrum of “Colidextribacter massiliensis” strain Marseille-P23083 T is
41
available at: (http://www.mediterranee-infection.com/article.php?laref=256&titre=urms-database)
42
Nucleotide sequence accession number. The 16S r RNA gene sequence was deposited in Genbank
43
under accession number LT598546.
44
Deposit in a culture collection. Strain Marseille-P3083T was deposited in the Collection de
45
Souches de l’Unité des Rickettsies (CSUR, WDCM 875) under number P3083, ( deposit in progress
46
in DSMZ collection).
47
Conflict of interest:
48
None to declare.
49
Funding sources:
50
This work was funded by Mediterrannée-Infection Foundation.
EP
AC C
51
TE D
40
52 53 54
References
55
[1] Lagier JC, Hugon P, Khelaifia S, Fournier PE, La Scola B, Raoult D. The rebirth of culture in
ACCEPTED MANUSCRIPT
microbiology through the example of culturomics to study human gut microbiota. Clin Microbiol
57
Rev 2015;28:237–64.
58
[2] Seng P, Abat C, Rolain JM, Colson P, Lagier JC, Gouriet F, Fournier PE, Drancourt M, La Scola
59
B and Raoult D. Identification of rare pathogenic bacteria in a clinical microbiology laboratory:
60
impact of matrix-assisted laser desorption ionization-time of flight mass spectrometry. J Clin
61
Microbiol 2013;51:2182-94
62
[3] Drancourt M, Bollet C, Carlioz A,Martelin R, Gayral J-P, Raoult D. 16S ribosomal DNA
63
sequence analysis of a large collection of environmental and clinical unidentifiable bacterial
64
isolates. J Clin Microbiol 2000; 38:3623-3630.
65
[4] Collins MD, Lawson PA, Willems A, Cordoba JJ, Fernandez-Garayzabal J, Garcia P, et al. The
66
phylogeny of the genus Clostridium: proposal of five new genera and eleven new species
67
combinations. Int J Syst Bacteriol. 1994;44:812–26.
68
[5] Carlier J-P, Bedora-Faure M, K’ouas G, Alauzet C, Mory F. Proposal to unify Clostridium
69
orbiscindens Winter et al. 1991 and Eubacterium plautii (Séguin 1928) Hofstad and Aasjord 1982,
70
with description of Flavonifractor plautii gen. nov., comb. nov., and reassignment of Bacteroides
71
capillosus to Pseudoflavonifractor capillosus gen. nov., comb. nov. Int J Syst Evol Microbiol.
72
2010;60:585–90.
73
[6] Kläring K, Hanske L, Bui N, Charrier C, Blaut M, Haller D, et al. Intestinimonas
74
butyriciproducens gen. nov., sp. nov., a butyrate-producing bacterium from the mouse intestine.
75
International Journal of Systematic and Evolutionary Microbiology. 2013;63:4606–12.
76
[7] Huson DH, Auch AF, Qi J, Schuster SC. MEGAN analysis of metagenomic data. Genome Res
77
2007;17:377–86.
78
[8] Kim M, Oh H-S, Park S-C, Chun J. Towards a taxonomic coherence between average nucleotide
79
identity and 16S rRNA gene sequence similarity for species demarcation of prokaryotes. Int J Syst
80
Evol Microbiol 2014;64:346–51.
AC C
EP
TE D
M AN U
SC
RI PT
56
ACCEPTED MANUSCRIPT
Figure 1: Phylogenetic tree showing the position of “Colidextribacter massiliensis” strain
82
Marseille-P3083T relative to other phylogenetically-close neighbours. Sequences were aligned
83
using CLUSTALW, and phylogenetic inferences obtained using the maximum-likelihood method
84
within the MEGA software. Numbers at the nodes are percentages of bootstrap values obtained by
85
repeating the analysis 1,000 times to generate a majority consensus tree. Only the bootstraps score
86
at least 90% were retained.
RI PT
81
87
SC
88 89
AC C
EP
TE D
M AN U
90
ACCEPTED MANUSCRIPT
Pseudoflavonifractor capillosus (NR 025670) 99
RI PT
Flavonifractor plautii (NR 029356) Intestinimonas butyriciproducens (NR 118554)
Colidextribacter massiliensis (LT598546) Clostridium viride (NR 026204)
98
Oscillibacter valericigenes (NR 074793) 100
Oscillibacter ruminantium (NR 118156)
SC
Sporobacter termitidis (NR 044972) 91
Papillibacter cinnamivorans (NR 025025)
98
Butyricicoccus pullicaecorum (NR 044490) 100
M AN U
Eubacterium desmolans (NR 044644)
Eubacterium siraeum (NR 102935)
Hydrogenoanaerobacterium saccharovorans (NR 044425)
100
Clostridium methylpentosum (NR 029355) Ruminococcus champanellensis (NR 102884)
92
Ruminococcus albus (NR 115230)
Natranaerobaculum magadiense (NR 135712)
TE D
Saccharofermentans acetigenes (NR 115340) Clostridium stercorarium (NR 102513)
Clostridium alkalicellulosi (NR 115345) Ruminiclostridium thermocellum (NR 074629) 96
AC C
0.02
Clostridium clariflavum (NR 102987)
EP
95
S2052-2975(16)30135-4
DOI:
10.1016/j.nmni.2016.11.023
Reference:
NMNI 266
To appear in:
New Microbes and New Infections
Received Date: 25 October 2016 Revised Date:
20 November 2016
Accepted Date: 23 November 2016
Please cite this article as: Ricaboni D, Mailhe M, Cadoret F, Vitton V, Fournier P-E, Raoult D, “Colidextribacter massiliensis” gen. nov., sp. nov. isolated from human right colon, New Microbes and New Infections (2016), doi: 10.1016/j.nmni.2016.11.023. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
ACCEPTED MANUSCRIPT “Colidextribacter massiliensis” gen. nov., sp. nov. isolated from human right colon.
Running title : Colidextribacter massiliensis.
1
RI PT
D. Ricaboni1-2,M. Mailhe1,F. Cadoret1, V. Vitton3, P.-E. Fournier1, D. Raoult1* Aix-Marseille Université, URMITE, UM63, CNRS7278, IRD198, Inserm 1095, Institut
Hospitalo-Universitaire Méditerranée-Infection, Faculté de médecine, 27 Boulevard Jean
2
SC
Moulin, 13385, Marseille cedex 05, France
Department of Biomedical and Clinical Sciences, 3rd Division of Clinical Infectious
3
Service de Gastroenterologie, Hopital Nord, Assistance Publique-Hopitaux de Marseille,
13915 Marseille, France.
Corresponding author. E-mail address: [email protected]
TE D
*
M AN U
Disease, University of Milan, Luigi Sacco Hospital, Italy.
AC C
EP
Keywords: Culturomics, taxono-genomics, Colidextribacter massiliensis, human gut.
ACCEPTED MANUSCRIPT
Abstract:
2
We report here the main characteristics of “Colidextribacter massiliensis” strain Marseille-P3083T
3
(CSURP3083) that was isolated from a human right-colon lavage sample.
AC C
EP
TE D
M AN U
SC
RI PT
1
ACCEPTED MANUSCRIPT
In 2016, as a part of culturomics study of the human microbiome [1], we isolated from the right
5
colon of a 27year-old obese patient a bacterial strain that could not be identified by our systematic
6
matrix-assisted laser desorption-ionization time-of-flight mass spectrometry screening on a
7
Microflex spectrometer (Bruker Daltonics, Brenen, Germany) [2]. The patient gave a signed
8
informed consent and the study was validated by the ethics committee of the Institut Fédératif de
9
Recherche IFR48 under number 2016-010.
RI PT
4
10
Strain Marseille-P3083T
11
(bioMérieux, Marcy l’Etoile, France) in anaerobic atmosphere (anaeroGEN, Oxoid Ltd, Dardilly,
12
France) after a 30-day enrichment of the fresh right-colon sample in an anaerobic haemoculture
13
bottle (Becton Dickinson, Pont de Claix, France) added with 5mL of sheep blood (bioMérieux) and
14
5 mL of 0.2µm filtered (Thermo Fisher Scientific, Villebon-sur-Yvette, France) rumen at 37°C.
15
After 96 hours of anaerobic incubation on 5% sheep blood-enriched agar (bioMérieux) colonies
16
were punctiform, approximatively round, translucid, with a mean diameter of 0.1-0.3 mm. Bacterial
17
cells are small Gram-negative cocci, ranging in length from 0.4 to 0.6 µm. Strain Marseille-P3083T
18
tested catalase and oxidase negative. Sporulation test (20 min at 80°C) resulted negative, no growth
19
was achived under aerobic or microaerophilic (campyGen, Oxoid) conditions. The 16S rRNA gene
20
was sequenced using fD1-rP2 primers as previously described [3], using a 3130-XL sequencer
21
(Applied Biosciences, Saint Aubin, France). Strain Marseille-P3083T exhibited a 93.57 % sequence
22
identity with Pseudoflavonifractor capillosus strain ATCC 29799T (Genbank accession number
23
AY136666), the phylogenetically closest species with standing nomenclature (Fig. 1). This value
24
putatively classifies strain Marseille-P3083T as a member of a new genus within the Clostridiales
25
cluster IV in the phylum Firmicutes. Clostridiales cluster IV was created in 1994 and collect
26
microrganisms that exhibit Clostridium and non Clostridium-like characteristics [4], including
27
Gram-negative microrganisms [5]. Intestinimonas butyriciproducens DSM 26588T (16S rRNA
28
similarity of 93.42%) is another phylogenetically very close species (Fig. 1) that exhibits the
29
peculiar characteristic of appearing as a Gram-negative rod while elecron microscopy demonstrated
AC C
EP
TE D
M AN U
SC
growth was obtained on 5% sheep blood-Columbia agar medium
ACCEPTED MANUSCRIPT
30
Gram-positive cell wall characteristics [6].
31
On the basis of the morphologic differences (morphology and Gram stain) and the 16S rRNA
32
sequence divergence of strain Marseille-P3083T with the phylogenetically closest species with
33
standing in nomenclature [7-8] we propose here the creation of the new genus Colidextribacter
34
(Co.li.dex.tri.bac.ter,
35
bacter a rod ; N.L. masc. n. Colidextribacter, a rod from the right colon); and a new species
36
Colidextribacter massiliensis for which Marseille-P3083T (=CSURP3083) is the type strain.
37
Strain Marseille-P3083T is the type strain of Colidextribacter massiliensis gen. nov., sp. nov.,
38
(mas.si.li.en’sis, L., masc. adj., massiliensis for Massilia, the Roman name of Marseille).
SC
RI PT
L. neut. n. colon or colum, the colon; L. adj. dexter right; N.L. masc. n.
M AN U
39
The MALDITOF spectrum of “Colidextribacter massiliensis” strain Marseille-P23083 T is
41
available at: (http://www.mediterranee-infection.com/article.php?laref=256&titre=urms-database)
42
Nucleotide sequence accession number. The 16S r RNA gene sequence was deposited in Genbank
43
under accession number LT598546.
44
Deposit in a culture collection. Strain Marseille-P3083T was deposited in the Collection de
45
Souches de l’Unité des Rickettsies (CSUR, WDCM 875) under number P3083, ( deposit in progress
46
in DSMZ collection).
47
Conflict of interest:
48
None to declare.
49
Funding sources:
50
This work was funded by Mediterrannée-Infection Foundation.
EP
AC C
51
TE D
40
52 53 54
References
55
[1] Lagier JC, Hugon P, Khelaifia S, Fournier PE, La Scola B, Raoult D. The rebirth of culture in
ACCEPTED MANUSCRIPT
microbiology through the example of culturomics to study human gut microbiota. Clin Microbiol
57
Rev 2015;28:237–64.
58
[2] Seng P, Abat C, Rolain JM, Colson P, Lagier JC, Gouriet F, Fournier PE, Drancourt M, La Scola
59
B and Raoult D. Identification of rare pathogenic bacteria in a clinical microbiology laboratory:
60
impact of matrix-assisted laser desorption ionization-time of flight mass spectrometry. J Clin
61
Microbiol 2013;51:2182-94
62
[3] Drancourt M, Bollet C, Carlioz A,Martelin R, Gayral J-P, Raoult D. 16S ribosomal DNA
63
sequence analysis of a large collection of environmental and clinical unidentifiable bacterial
64
isolates. J Clin Microbiol 2000; 38:3623-3630.
65
[4] Collins MD, Lawson PA, Willems A, Cordoba JJ, Fernandez-Garayzabal J, Garcia P, et al. The
66
phylogeny of the genus Clostridium: proposal of five new genera and eleven new species
67
combinations. Int J Syst Bacteriol. 1994;44:812–26.
68
[5] Carlier J-P, Bedora-Faure M, K’ouas G, Alauzet C, Mory F. Proposal to unify Clostridium
69
orbiscindens Winter et al. 1991 and Eubacterium plautii (Séguin 1928) Hofstad and Aasjord 1982,
70
with description of Flavonifractor plautii gen. nov., comb. nov., and reassignment of Bacteroides
71
capillosus to Pseudoflavonifractor capillosus gen. nov., comb. nov. Int J Syst Evol Microbiol.
72
2010;60:585–90.
73
[6] Kläring K, Hanske L, Bui N, Charrier C, Blaut M, Haller D, et al. Intestinimonas
74
butyriciproducens gen. nov., sp. nov., a butyrate-producing bacterium from the mouse intestine.
75
International Journal of Systematic and Evolutionary Microbiology. 2013;63:4606–12.
76
[7] Huson DH, Auch AF, Qi J, Schuster SC. MEGAN analysis of metagenomic data. Genome Res
77
2007;17:377–86.
78
[8] Kim M, Oh H-S, Park S-C, Chun J. Towards a taxonomic coherence between average nucleotide
79
identity and 16S rRNA gene sequence similarity for species demarcation of prokaryotes. Int J Syst
80
Evol Microbiol 2014;64:346–51.
AC C
EP
TE D
M AN U
SC
RI PT
56
ACCEPTED MANUSCRIPT
Figure 1: Phylogenetic tree showing the position of “Colidextribacter massiliensis” strain
82
Marseille-P3083T relative to other phylogenetically-close neighbours. Sequences were aligned
83
using CLUSTALW, and phylogenetic inferences obtained using the maximum-likelihood method
84
within the MEGA software. Numbers at the nodes are percentages of bootstrap values obtained by
85
repeating the analysis 1,000 times to generate a majority consensus tree. Only the bootstraps score
86
at least 90% were retained.
RI PT
81
87
SC
88 89
AC C
EP
TE D
M AN U
90
ACCEPTED MANUSCRIPT
Pseudoflavonifractor capillosus (NR 025670) 99
RI PT
Flavonifractor plautii (NR 029356) Intestinimonas butyriciproducens (NR 118554)
Colidextribacter massiliensis (LT598546) Clostridium viride (NR 026204)
98
Oscillibacter valericigenes (NR 074793) 100
Oscillibacter ruminantium (NR 118156)
SC
Sporobacter termitidis (NR 044972) 91
Papillibacter cinnamivorans (NR 025025)
98
Butyricicoccus pullicaecorum (NR 044490) 100
M AN U
Eubacterium desmolans (NR 044644)
Eubacterium siraeum (NR 102935)
Hydrogenoanaerobacterium saccharovorans (NR 044425)
100
Clostridium methylpentosum (NR 029355) Ruminococcus champanellensis (NR 102884)
92
Ruminococcus albus (NR 115230)
Natranaerobaculum magadiense (NR 135712)
TE D
Saccharofermentans acetigenes (NR 115340) Clostridium stercorarium (NR 102513)
Clostridium alkalicellulosi (NR 115345) Ruminiclostridium thermocellum (NR 074629) 96
AC C
0.02
Clostridium clariflavum (NR 102987)
EP
95