1887

Abstract

Six strictly anaerobic Gram-negative bacteria representing three novel species were isolated from the female reproductive tract. The proposed type strains for each species were designated UPII 199-6, KA00182 and BV3C16-1. Phylogenetic analyses based on 16S rRNA gene sequencing indicated that the bacterial isolates were members of the genus . UPII 199-6 and KA00182 had 16S rRNA gene sequence identities of 99.9 % with 16S rRNA clone sequences previously amplified from the human vagina designated as type 1 and type 2, members of the human vaginal microbiota associated with bacterial vaginosis, preterm birth and HIV acquisition. UPII 199-6 exhibited sequence identities ranging from 92.9 to 93.6 % with validly named isolates and KA00182 had 16S rRNA gene sequence identities ranging from 92.6–94.2 %. BV3C16-1 was most closely related to with a 16S rRNA gene sequence identity of 95.4 %. Cells were coccoid or diplococcoid, non-motile and did not form spores. Genital tract isolates metabolized organic acids but were asaccharolytic. The isolates also metabolized amino acids. The DNA G+C content for the genome sequences of UPII 199-6, KA00182 and BV3C16-1 were 46.4, 38.9 and 49.8 mol%, respectively. Digital DNA–DNA hybridization and average nucleotide identity between the genital tract isolates and other validly named species suggest that each isolate type represents a new species. The major fatty acid methyl esters include the following: C, C, C dimethyl acetal (DMA) and summed feature 5 (C DMA and/or C 3-OH) in UPII 199-6; C and C 9 in KA00182; C; C 3-OH; and summed feature 5 in BV3C16-1. The isolates produced butyrate, isobutyrate, and isovalerate but there were specific differences including production of formate and propionate. Together, these data indicate that UPII 199-6, KA00182 and BV3C16-1 represent novel species within the genus . We propose the following names: sp. nov. for UPII 199-6 representing the type strain of this species (=DSM 111201=ATCC TSD-205), sp. nov. for KA00182 representing the type strain of this species (=DSM 111202=ATCC TSD-206) and sp. nov. for BV3C16-1 representing the type strain of this species (=DSM 111203=ATCC TSD-207).

Funding
This study was supported by the:
  • SharonL. Hillier , National Institutes of Health , (Award U19AI120249)
  • DavidN. Fredricks , National Institutes of Health , (Award HG005816)
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2021-02-22
2021-03-08
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References

  1. Rogosa M. Transfer of Peptostreptococcus elsdenii to a new genus, Megasphaera [M. elsdenii Gutierrez, et al. comb. nov.].. Int J Syst Bact 1971; 21: 187 189 [CrossRef]
    [Google Scholar]
  2. 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 [CrossRef]
    [Google Scholar]
  3. Jeon BS, Kim S, Sang BI. Megasphaera hexanoica sp. nov., a medium-chain carboxylic acid-producing bacterium isolated from a cow rumen. Int J Syst Evol Microbiol 2017; 67: 2114 2120 [CrossRef] [PubMed]
    [Google Scholar]
  4. 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 Evol Microbiol 2006; 56: 695 702 [CrossRef] [PubMed]
    [Google Scholar]
  5. Lanjekar VB, Marathe NP, Ramana VV, Shouche YS, Ranade DR. Megasphaera indica sp. nov., an obligate anaerobic bacteria isolated from human faeces. Int J Syst Evol Microbiol 2014; 64: 2250 2256 [CrossRef] [PubMed]
    [Google Scholar]
  6. Maki JJ, Looft T. Megasphaera stantonii sp. nov., a butyrate-producing bacterium isolated from the cecum of a healthy chicken. Int J Syst Evol Microbiol 2018; 68: 3409 3415 [CrossRef] [PubMed]
    [Google Scholar]
  7. Marchandin H, Jumas-Bilak E, Gay B, Teyssier C, Jean-Pierre H et al. Phylogenetic analysis of some Sporomusa sub-branch members isolated from human clinical specimens: description of Megasphaera micronuciformis sp. nov. Int J Syst Evol Microbiol 2003; 53: 547 553 [CrossRef] [PubMed]
    [Google Scholar]
  8. Antunes LC, Poppleton D, Klingl A, Criscuolo A, Dupuy B et al. Phylogenomic analysis supports the ancestral presence of LPS-outer membranes in the Firmicutes. eLife 2016; 5: e14589 [CrossRef] [PubMed]
    [Google Scholar]
  9. Marchandin H, Juvonen R, Haikara A et al. Megasphaera. In Whitman WB, DeVos P, Dedysh S, Hedlund B, Kampfer P. (editors) Bergey’s Manual of Systematics of Archaea and Bacteria John Wiley & Sons, Inc., in association with Bergey’s Manual Trust; 2015
    [Google Scholar]
  10. Padmanabhan R, Lagier JC, Dangui NP, Michelle C, Couderc C et al. Non-contiguous finished genome sequence and description of Megasphaera massiliensis sp. nov. Stand Genomic Sci 2013; 8: 525 538 [CrossRef] [PubMed]
    [Google Scholar]
  11. Zhou X, Bent SJ, Schneider MG, Davis CC, Islam MR et al. Characterization of vaginal microbial communities in adult healthy women using cultivation-independent methods. Microbiology 2004; 150: 2565 2573 [CrossRef] [PubMed]
    [Google Scholar]
  12. Fredricks DN, Fiedler TL, Marrazzo JM. Molecular identification of bacteria associated with bacterial vaginosis. N Engl J Med 2005; 353: 1899 1911 [CrossRef] [PubMed]
    [Google Scholar]
  13. Fredricks DN, Fiedler TL, Thomas KK, Mitchell CM, Marrazzo JM. Changes in vaginal bacterial concentrations with intravaginal metronidazole therapy for bacterial vaginosis as assessed by quantitative PCR. J Clin Microbiol 2009; 47: 721 726 [CrossRef] [PubMed]
    [Google Scholar]
  14. Fredricks DN, Fiedler TL, Thomas KK, Oakley BB, Marrazzo JM. Targeted PCR for detection of vaginal bacteria associated with bacterial vaginosis. J Clin Microbiol 2007; 45: 3270 3276 [CrossRef] [PubMed]
    [Google Scholar]
  15. Cruciani F, Biagi E, Severgnini M, Consolandi C, Calanni F et al. Development of a microarray-based tool to characterize vaginal bacterial fluctuations and application to a novel antibiotic treatment for bacterial vaginosis. Antimicrob Agents Chemother 2015; 59: 2825 2834 [CrossRef] [PubMed]
    [Google Scholar]
  16. Fethers K, Twin J, Fairley CK, Fowkes FJI, Garland SM et al. Bacterial vaginosis (bv) candidate bacteria: associations with bv and behavioural practices in sexually-experienced and inexperienced women. PLoS One 2012; 7: e30633 [CrossRef] [PubMed]
    [Google Scholar]
  17. Fredricks DN, Plantinga A, Srinivasan S, Oot A, Wiser A et al. Vaginal and extra-vaginal bacterial colonization and risk for incident bacterial vaginosis in a population of women who have sex with men. J Infect Dis 2020; 28: [CrossRef]
    [Google Scholar]
  18. Hilbert DW, Smith WL, Chadwick SG, Toner G, Mordechai E et al. Development and validation of a highly accurate quantitative real-time PCR assay for diagnosis of bacterial vaginosis. J Clin Microbiol 2016; 54: 1017 1024 [CrossRef] [PubMed]
    [Google Scholar]
  19. Marrazzo JM, Fiedler TL, Srinivasan S, Thomas KK, Liu C et al. Extravaginal reservoirs of vaginal bacteria as risk factors for incident bacterial vaginosis. J Infect Dis 2012; 205: 1580 1588 [CrossRef] [PubMed]
    [Google Scholar]
  20. Marrazzo JM, Thomas KK, Fiedler TL, Ringwood K, Fredricks DN. Relationship of specific vaginal bacteria and bacterial vaginosis treatment failure in women who have sex with women. Ann Intern Med 2008; 149: 20 28 [CrossRef] [PubMed]
    [Google Scholar]
  21. Muzny CA, Blanchard E, Taylor CM, Aaron KJ, Talluri R et al. Identification of key bacteria involved in the induction of incident bacterial vaginosis: a prospective study. J Infect Dis 2018; 218: 966 978 [CrossRef] [PubMed]
    [Google Scholar]
  22. Shipitsyna E, Roos A, Datcu R, Hallen A, Fredlund H et al. Composition of the vaginal microbiota in women of reproductive age--sensitive and specific molecular diagnosis of bacterial vaginosis is possible?. PLoS One 2013; 8: e60670 [CrossRef] [PubMed]
    [Google Scholar]
  23. Srinivasan S, Hoffman NG, Morgan MT, Matsen FA, Fiedler TL et al. Bacterial communities in women with bacterial vaginosis: high resolution phylogenetic analyses reveal relationships of microbiota to clinical criteria. PLoS One 2012; 7: e37818 [CrossRef] [PubMed]
    [Google Scholar]
  24. Tamrakar R, Yamada T, Furuta I, Cho K, Morikawa M et al. Association between Lactobacillus species and bacterial vaginosis-related bacteria, and bacterial vaginosis scores in pregnant Japanese women. BMC Infect Dis 2007; 7: 128 [CrossRef] [PubMed]
    [Google Scholar]
  25. Zozaya-Hinchliffe M, Lillis R, Martin DH, Ferris MJ. Quantitative PCR assessments of bacterial species in women with and without bacterial vaginosis. J Clin Microbiol 2010; 48: 1812 1819 [CrossRef] [PubMed]
    [Google Scholar]
  26. Zozaya-Hinchliffe M, Martin DH, Ferris MJ. Prevalence and abundance of uncultivated Megasphaera-like bacteria in the human vaginal environment. Appl Environ Microbiol 2008; 74: 1656 1659 [CrossRef] [PubMed]
    [Google Scholar]
  27. Cartwright CP, Lembke BD, Ramachandran K, Body BA, Nye MB et al. Development and validation of a semiquantitative, multitarget PCR assay for diagnosis of bacterial vaginosis. J Clin Microbiol 2012; 50: 2321 2329 [CrossRef] [PubMed]
    [Google Scholar]
  28. Coleman JS, Gaydos CA. Molecular diagnosis of bacterial vaginosis: an update. J Clin Microbiol 2018; 56: e00342 18 [CrossRef] [PubMed]
    [Google Scholar]
  29. Gaydos CA, Beqaj S, Schwebke JR, Lebed J, Smith B et al. Clinical validation of a test for the diagnosis of vaginitis. Obstet Gynecol 2017; 130: 181 189 [CrossRef] [PubMed]
    [Google Scholar]
  30. Kusters JG, Reuland EA, Bouter S, Koenig P, Dorigo-Zetsma JW. A multiplex real-time PCR assay for routine diagnosis of bacterial vaginosis. Eur J Clin Microbiol Infect Dis 2015; 34: 1779 1785 [CrossRef] [PubMed]
    [Google Scholar]
  31. Singh R, Ramsuran V, Mitchev N, Niehaus AJ, Han KSS et al. Assessing a diagnosis tool for bacterial vaginosis. Eur J Clin Microbiol Infect Dis 2020; 39: 1481 1485 [CrossRef] [PubMed]
    [Google Scholar]
  32. Gosmann C, Anahtar MN, Handley SA, Farcasanu M, Abu-Ali G et al. Lactobacillus-deficient cervicovaginal bacterial communities are associated with increased HIV acquisition in young South African women. Immunity 2017; 46: 29 37 [CrossRef] [PubMed]
    [Google Scholar]
  33. McClelland RS, Lingappa JR, Srinivasan S, Kinuthia J, John-Stewart GC et al. Evaluation of the association between the concentrations of key vaginal bacteria and the increased risk of HIV acquisition in African women from five cohorts: a nested case-control study. Lancet Infect Dis 2018; 18: 554 564 [CrossRef] [PubMed]
    [Google Scholar]
  34. Srinivasan S, Richardson BA, Wallis J, Fiedler TL, Dezzutti CS. Vaginal microbiota and HIV acquisition risk among African women. Conference on Retroviruses and Opportunistic Infections USA: Boston; 2018
    [Google Scholar]
  35. Nelson DB, Hanlon A, Nachamkin I, Haggerty C, Mastrogiannis DS et al. Early pregnancy changes in bacterial vaginosis-associated bacteria and preterm delivery. Paediatr Perinat Epidemiol 2014; 28: 88 96 [CrossRef] [PubMed]
    [Google Scholar]
  36. Elovitz MA, Gajer P, Riis V, Brown AG, Humphrys MS et al. Cervicovaginal microbiota and local immune response modulate the risk of spontaneous preterm delivery. Nat Commun 2019; 10: 1305 [CrossRef] [PubMed]
    [Google Scholar]
  37. Haggerty CL, Ness RB, Totten PA, Farooq F, Tang G et al. Presence and concentrations of select bacterial vaginosis-associated bacteria are associated with increased risk of pelvic inflammatory disease. Sex Transm Dis 2020; 47: 344 346 [CrossRef] [PubMed]
    [Google Scholar]
  38. Petrina MAB, Cosentino LA, Rabe LK, Hillier SL. Susceptibility of bacterial vaginosis (BV)-associated bacteria to secnidazole compared to metronidazole, tinidazole and clindamycin. Anaerobe 2017; 47: 115 119 [CrossRef] [PubMed]
    [Google Scholar]
  39. Petrina MAB, Cosentino LA, Wiesenfeld HC, Darville T, Hillier SL. Susceptibility of endometrial isolates recovered from women with clinical pelvic inflammatory disease or histological endometritis to antimicrobial agents. Anaerobe 2019; 56: 61 65 [CrossRef] [PubMed]
    [Google Scholar]
  40. Srinivasan S, Munch MM, Sizova MV, Fiedler TL, Kohler CM et al. More easily cultivated than identified: classical isolation with molecular identification of vaginal bacteria. J Infect Dis 2016; 214 Suppl 1: S21 S28 [CrossRef] [PubMed]
    [Google Scholar]
  41. Dang AT, Cotton S, Sankaran-Walters S, Li C-S, Lee CY et al. Evidence of an increased pathogenic footprint in the lingual microbiome of untreated HIV infected patients. BMC Microbiol 2012; 12: 153 [CrossRef] [PubMed]
    [Google Scholar]
  42. Sato N, Kakuta M, Hasegawa T, Yamaguchi R, Uchino E et al. Metagenomic analysis of bacterial species in tongue microbiome of current and never smokers. NPJ Biofilms Microbiomes 2020; 6: 11 [CrossRef] [PubMed]
    [Google Scholar]
  43. de la Cuesta-Zuluaga J, Mueller NT, Corrales-Agudelo V, Velásquez-Mejía EP, Carmona JA et al. Metformin Is associated with higher relative abundance of mucin-degrading Akkermansia muciniphila and several short-chain fatty acid-producing microbiota in the gut. Diabetes Care 2017; 40: 54 62 [CrossRef] [PubMed]
    [Google Scholar]
  44. Gaike AH, Paul D, Bhute S, Dhotre DP, Pande P et al. The gut microbial diversity of newly diagnosed diabetics but not of prediabetics is significantly different from that of healthy nondiabetics. mSystems 2020; 5: e00578 19 [CrossRef]
    [Google Scholar]
  45. Patrone V, Vajana E, Minuti A, Callegari ML, Federico A et al. Postoperative changes in fecal bacterial communities and fermentation products in obese patients undergoing bilio-intestinal bypass. Front Microbiol 2016; 7: 200 [CrossRef] [PubMed]
    [Google Scholar]
  46. Frolund M, Falk L, Ahrens P, Jensen JS. Detection of ureaplasmas and bacterial vaginosis associated bacteria and their association with non-gonococcal urethritis in men. PLoS One 2019; 14: e0214425 [CrossRef] [PubMed]
    [Google Scholar]
  47. Nelson DE, Dong Q, Van der Pol B, Toh E, Fan B et al. Bacterial communities of the coronal sulcus and distal urethra of adolescent males. PLoS One 2012; 7: e36298 [CrossRef] [PubMed]
    [Google Scholar]
  48. Srinivasan S, Chambers LC, Tapia KA, Hoffman NG, Munch MM et al. Urethral microbiota in men: association of Haemophilus influenzae and Mycoplasma penetrans with nongonococcal urethritis. Clin Infect Dis 2020 [CrossRef] [PubMed]
    [Google Scholar]
  49. Wiesenfeld HC, Meyn LA, Darville T, Macio IS, Hillier SL. A randomized controlled trial of ceftriaxone and doxycycline, with or without metronidazole, for the treatment of acute pelvic inflammatory disease. Clin Infect Dis 2020 [CrossRef] [PubMed]
    [Google Scholar]
  50. Atlas RM. Handbook of Microbiological Media , Fourth Edition. Washington D. C: ASM Press; 2010
    [Google Scholar]
  51. Koransky JR, Allen SD, Dowell VR. Use of ethanol for selective isolation of sporeforming microorganisms. Appl Environ Microbiol 1978; 35: 762 765 [CrossRef] [PubMed]
    [Google Scholar]
  52. Song Y, Finegold S et al. Peptostreptococcus, Finegoldia, Anaerococcus, Peptoniphilus, Veillonella, and other anaerobic cocci. In Jorgensen J, Pfaller M, Carroll K, Funke G, Landry M. (editors) Manual of Clinical Microbiology , 11th edn. Washington D. C: ASM Press; 2015 pp 909 919
    [Google Scholar]
  53. Summanen P, Baron EJ, Citron DM, Strong CA, Wexler HM. Belmont Wadsworth Anaerobic Bacteriology Manual , 5th ed. Belmont, CA: Star Publishing; 1993
    [Google Scholar]
  54. Miller JM, Rhoden DL. Preliminary evaluation of Biolog, a carbon source utilization method for bacterial identification. J Clin Microbiol 1991; 29: 1143 1147 [CrossRef] [PubMed]
    [Google Scholar]
  55. Beaucage CM, Onderdonk AB. Evaluation of a prereduced anaerobically sterilized medium (PRAS II) system for identification anaerobic microorganisms. J Clin Microbiol 1982; 16: 570 572 [CrossRef] [PubMed]
    [Google Scholar]
  56. Nagana Gowda GA, Gowda YN, Raftery D. Expanding the limits of human blood metabolite quantitation using NMR spectroscopy. Anal Chem 2015; 87: 706 715 [CrossRef] [PubMed]
    [Google Scholar]
  57. Ulrich EL, Akutsu H, Doreleijers JF, Harano Y, Ioannidis YE et al. BioMagResBank. Nucleic Acids Res 2008; 36: D402 D408 [CrossRef] [PubMed]
    [Google Scholar]
  58. Wishart DS, Jewison T, Guo AC, Wilson M, Knox C et al. HMDB 3.0-the human metabolome database in 2013. Nucleic Acids Res 2013; 41: D801 D807 [CrossRef] [PubMed]
    [Google Scholar]
  59. Srinivasan S, Morgan MT, Fiedler TL, Djukovic D, Hoffman NG et al. Metabolic signatures of bacterial vaginosis. mBio 2015; 6: e00204-15 [CrossRef] [PubMed]
    [Google Scholar]
  60. Wayne PA. Clinical and Laboratory Standards Institute editor Performance Standards for Antimicrobial Susceptibility Testing , 26th ed. 2016
    [Google Scholar]
  61. Workowski KA. Centers for disease control and prevention sexually transmitted diseases treatment guidelines. Clin Infect Dis 2015; 61 Suppl 8: S759 S762 [CrossRef] [PubMed]
    [Google Scholar]
  62. Hillier SL, Nyirjesy P, Waldbaum AS, Schwebke JR, Morgan FG et al. Secnidazole treatment of bacterial vaginosis: a randomized controlled trial. Obstet Gynecol 2017; 130: 379 386 [CrossRef] [PubMed]
    [Google Scholar]
  63. Pentikis H, Adetoro N, Tipping D, Levy S. An integrated efficacy and safety analysis of single-dose secnidazole 2 g in the treatment of bacterial vaginosis. Reprod Sci 2020; 27: 523 528 [CrossRef] [PubMed]
    [Google Scholar]
  64. Schwebke JR, Morgan FG, Koltun W, Nyirjesy P. A phase-3, double-blind, placebo-controlled study of the effectiveness and safety of single oral doses of secnidazole 2 G for the treatment of women with bacterial vaginosis. Am J Obstet Gynecol 2017; 217: 678.e1 67678 [CrossRef] [PubMed]
    [Google Scholar]
  65. Yoon SH, Ha S-M, Kwon S, Lim J, Kim Y et al. Introducing EzBioCloud: a taxonomically United database of 16S rRNA gene sequences and whole-genome assemblies. Int J Syst Evol Microbiol 2017; 67: 1613 1617 [CrossRef] [PubMed]
    [Google Scholar]
  66. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool. J Mol Biol 1990; 215: 403 410 [CrossRef] [PubMed]
    [Google Scholar]
  67. Tamura K, Nei M. Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Mol Biol Evol 1993; 10: 512 526 [CrossRef] [PubMed]
    [Google Scholar]
  68. Kumar S, Stecher G, Li M, Knyaz C, Tamura K. mega X: molecular evolutionary genetics analysis across computing platforms. Mol Biol Evol 2018; 35: 1547 1549 [CrossRef] [PubMed]
    [Google Scholar]
  69. Wattam AR, Davis JJ, Assaf R, Boisvert S, Brettin T et al. Improvements to PATRIC, the all-bacterial bioinformatics database and analysis resource center. Nucleic Acids Res 2017; 45: D535 D542 [CrossRef] [PubMed]
    [Google Scholar]
  70. Meier-Kolthoff JP, Klenk HP, Goker M. Taxonomic use of DNA G+C content and DNA-DNA hybridization in the genomic age. Int J Syst Evol Microbiol 2014; 64: 352 356 [CrossRef] [PubMed]
    [Google Scholar]
  71. Meier-Kolthoff JP, Auch AF, Klenk HP, Goker M. Genome sequence-based species delimitation with confidence intervals and improved distance functions. BMC Bioinform 2013; 14: 60 [CrossRef] [PubMed]
    [Google Scholar]
  72. Rodriguez-R LM K. The enveomics collection: a toolbox for specialized analyses of microbial genomes and metagenomes. Peer J Prepr 2016; 4: e1900v1
    [Google Scholar]
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