Accepted Manuscript “Anaerotruncus massiliensis”, A new species identified from human stool from an obese patient after bariatric surgery Amadou H. Togo, René valero, Jérémy Delerce, Didier Raoult, Matthieu Million PII: S2052-2975(16)30086-5 DOI: 10.1016/j.nmni.2016.07.015 Reference: NMNI 217 To appear in: New Microbes and New Infections Received Date: 21 July 2016 Accepted Date: 27 July 2016 Please cite this article as: Togo AH, valero R, Delerce J, Raoult D, Million M, “Anaerotruncus massiliensis”, A new species identified from human stool from an obese patient after bariatric surgery, New Microbes and New Infections (2016), doi: 10.1016/j.nmni.2016.07.015. 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 “Anaerotruncus massiliensis ”, A New Species identified from Human Stool from an Obese Patient after Bariatric Surgery Running head: “Anaerotruncus massiliensis ” Amadou H Togo 1, René valero 2,3 , Jérémy Delerce 1, Didier Raoult 1, 4 and Matthieu Million 1* 1. Aix-Marseille Université, URMITE, UM63, CNRS7278, IRD198, Inserm 1095, Institut Hospitalo-Universitaire Méditerranée-Infection, Faculté de médecine, 27 Boulevard Jean Moulin, 13385, Marseille cedex 05, France 2. Aix Marseille Université, NORT “Nutrition, Obesity and Risk of Thrombosis”, INSERM1062, INRA1260 3. APHM, CHU Hôpital de la Conception, Service Nutrition, Maladies Métaboliques et Endocrinologie, Marseille, France 4. Special Infectious Agents Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia * Corresponding author. E-mail address: [email protected] MANUSCRIPT ACCEPTED ACCEPTED MANUSCRIPT 1 “Anaerotruncus massiliensis”, A New Species identified from Human Stool from an 2 Obese Patient after Bariatric Surgery 3 Running head: “Anaerotruncus massiliensis” 4 Keywords: Culturomics, taxonomy, bariatric surgery, obesity, human gut microbiota MANUSCRIPT ACCEPTED ACCEPTED MANUSCRIPT 5 Abstract: 6 We report the main characteristics of a new bacterial species strain AT3 T (CSUR = P2007, 7 DSM = 100567) that was isolated from the stool sample collected from a 47-year-old obese 8 French man after he underwent bariatric surgery. MANUSCRIPT ACCEPTED ACCEPTED MANUSCRIPT 9 We performed an exploratory study of gut microbiota from obese patients before and after 10 bariatric surgery by microbial culturomics approach as previously described (1). The patient 11 gave a signed informed consent and the study was validated by the ethics committee of the 12 Institut Federatif de Recherche IFR48 under number 09-022. The first growth was obtained 13 after 30 days of culture in a blood culture bottle (BACTEC™ Lytic/10 Anaerobic/F Culture 14 Vials, Le Pont de Claix, Isère, France) enriched with 4 ml of sheep blood and 4 ml of rumen 15 under anaerobic atmosphere at 37°C medium. Agar-grown (Columbia agar + 5% sheep blood, 16 bioMérieux, Marcy l’Etoile, France) colonies were opalescent with a mean diameter of 0.5- 17 1.5 mm. The strain AT3 T is a bacterial strain, whose spectrum could not be identified by our 18 systematic matrix-assisted laser desorption-ionization time-of-flight-mass spectrometer) 19 (MALDI-TOF-MS) screening on Microflex spectrometer (Bruker Daltonics, Germany) (2). 20 The 16S rRNA gene was sequenced using fD1-rP2 primers as previously described (3), using 21 a 3130-XL sequencer (Applied Biosciences, Saint Aubin, France). Strain AT3 T exhibited a 22 96.6% sequence similarity with Anaerotruncus colihominisMANUSCRIPT strain WAL 14565 T = CCUG 23 45055 T = CIP 107754 T, the phylogenetically closest species with standing nomenclature (Fig. 24 1), which putatively classifies it as a member of the genus Anaerotruncus established in 2004 25 by Lawson (4) within Clostridiaceae family in Firmicutes phylum. Bacterial cells were 26 Gram-negative stain; rod-shaped with 0.3 x 2.9µm. The strain AT3T was catalase-positive and 27 oxidase-negative. The closest species, Anaerotruncus colihominis is a Gram-positive bacillus 28 whereas the strain AT3 T remained Gram-negative after three assays. Strain AT3 Texhibits a 29 16S rRNA gene ACCEPTEDsequence divergence higher than 1.35% (3.4%) with its phylogenetically 30 closest species with standing in nomenclature (5); we propose the creation of a new species 31 that we named Anaerotruncus massiliensis (ma.si.li.en ′.sis. L. fem. adj. massiliensis , of 32 Massilia, the Latin name of Marseille where A. massiliensis was first isolated). The strain 33 AT3T is the type strain of the new species A. massiliensis . ACCEPTED MANUSCRIPT 34 MALDITOF-MS spectrum accession number 35 The MALDITOF-MS spectrum of this strain is available on http://www.mediterranee- 36 infection.com/article.php?laref=256&titre=urms-database 37 Nucleotide sequence accession number 38 The 16S rRNA gene sequence was deposited in GenBank beneath accession number 39 LN_866995. 40 Deposit in a culture collection 41 Strain AT3T was deposited in the Collection de Souches de l’Unité des Rickettsies (CSUR,) 42 and Deutsche Sammlung von Mikroorganismen (DSM) under number CSUR= P2007, 43 DSM=100567. MANUSCRIPT ACCEPTED ACCEPTED MANUSCRIPT 44 Conflict of interest 45 No conflict of interest to declare. 46 Acknowledgements 47 The authors thank Magdalen Lardière for English reviewing. This work was funded by 48 Mediterrannée-Infection Foundation. MANUSCRIPT ACCEPTED ACCEPTED MANUSCRIPT 49 50 Figure 1: Phylogenetic tree showing the position of strain AT3T relative to other 51 phylogenetically closest species. Sequences were aligned using CLUSTALW, and 52 phylogenetic inferences obtained using the maximum-MANUSCRIPTlikelihood method within the MEGA 53 software. Numbers at the nodes depict bootstrap values obtained by repeating the analysis 500 54 times to yield a consensus tree. Just the bootstraps values with at least 90% were retained. The 55 scale bar indicates a 1% nucleotide sequence divergence. ACCEPTED ACCEPTED MANUSCRIPT 56 References 57 1. Lagier J-C, Armougom F, Million M, Hugon P, Pagnier I, Robert C, et al . 58 Microbial culturomics: paradigm shift in the human gut microbiome study. Clin 59 Microbiol Infect. 2012 Dec;18(12):1185-93. 60 2. Seng P, Drancourt M, Gouriet F, La Scola B, Fournier P-E, Rolain JM, et al . Ongoing 61 revolution in bacteriology: routine identification of bacteria by matrix-assisted laser 62 desorption ionization time-of-flight mass spectrometry. Clin Infect Dis Off Publ Infect 63 Dis Soc Am. 2009 Aug; 49 (4):543–51. 64 3. Drancourt M, Bollet C, Carlioz A, Martelin R, Gayral JP, Raoult D. 16S ribosomal 65 DNA sequence analysis of a large collection of environmental and clinical 66 unidentifiable bacterial isolates. J Clin Microbiol. 2000 Oct;38(10):3623–30. 67 4. Lawson PA, Song Y, Liu C, Molitoris DR, Vaisanen M-L, Collins MD, et al . 68 Anaerotruncus colihominis gen. nov., sp. nov., from human faeces. Int J Syst Evol 69 Microbiol. 2004 Mar; 54(Pt 2):413–7. MANUSCRIPT 70 5. Huson DH, Richter DC, Rausch C, Dezulian T, Franz M, Rupp R. Dendroscope: An 71 interactive viewer for large phylogenetic trees. BMC Bioinformatics. 2007; 8:460. ACCEPTED.