Research article Special Issues

Use of high throughput amplicon sequencing and ethidium monoazide dye to track microbiota changes in an equol-producing menopausal woman receiving a long-term isoflavones treatment

  • Received: 17 October 2018 Accepted: 26 February 2019 Published: 22 March 2019
  • This work describes the impact of long term consumption of an isoflavone-rich dietary daily supplement on the composition and diversity of the faecal microbiota of a menopausal, equol-producing woman. Sequencing of 16S rDNA amplicons was performed on faecal samples taken at 0, 1, 3 and 6 months of treatment. Additionally, and for comparative purposes, ethidium monoazide (EMA) was used to avoid detection of DNA from dead bacteria. Members of two genera of the family Coriobacteriaceae (Eggerthella and Collinsella) were found in greater proportions at all sampling points during isoflavone supplementation. Different genera of the family Ruminococcaceae (e.g., Ruminococcus and Faecalibacterium), as well as members of the family Lachnospiraceae (Coprococcus) also underwent significant increases. For this last genus a positive correlation with the levels of equol excretion in urine was found. Currently bacterial strains known to be involved in isoflavone metabolism and equol production have been assigned to these taxa. The use of EMA dye allowed us to unravel those bacterial gut linages (e.g., Lachnospiraceae) that were more susceptible to damage. Our study contributes to the identification of microorganisms possibly involved in the production of isoflavone-desirable metabolites (such as equol), which could ultimately be isolated and further used as probiotics by people who cannot naturally benefit from isoflavones.

    Citation: Lucía Guadamuro, M. Andrea Azcárate-Peril, Rafael Tojo, Baltasar Mayo, Susana Delgado. Use of high throughput amplicon sequencing and ethidium monoazide dye to track microbiota changes in an equol-producing menopausal woman receiving a long-term isoflavones treatment[J]. AIMS Microbiology, 2019, 5(1): 102-116. doi: 10.3934/microbiol.2019.1.102

    Related Papers:

  • This work describes the impact of long term consumption of an isoflavone-rich dietary daily supplement on the composition and diversity of the faecal microbiota of a menopausal, equol-producing woman. Sequencing of 16S rDNA amplicons was performed on faecal samples taken at 0, 1, 3 and 6 months of treatment. Additionally, and for comparative purposes, ethidium monoazide (EMA) was used to avoid detection of DNA from dead bacteria. Members of two genera of the family Coriobacteriaceae (Eggerthella and Collinsella) were found in greater proportions at all sampling points during isoflavone supplementation. Different genera of the family Ruminococcaceae (e.g., Ruminococcus and Faecalibacterium), as well as members of the family Lachnospiraceae (Coprococcus) also underwent significant increases. For this last genus a positive correlation with the levels of equol excretion in urine was found. Currently bacterial strains known to be involved in isoflavone metabolism and equol production have been assigned to these taxa. The use of EMA dye allowed us to unravel those bacterial gut linages (e.g., Lachnospiraceae) that were more susceptible to damage. Our study contributes to the identification of microorganisms possibly involved in the production of isoflavone-desirable metabolites (such as equol), which could ultimately be isolated and further used as probiotics by people who cannot naturally benefit from isoflavones.


    加载中

    Acknowledgments



    The study was partially supported by projects from the Spanish Ministry of Economy and Competitiveness (MINECO) (AGL-2014-57820-R) and Asturias Principality (GRUPIN14-137). The Microbiome Core Facility is supported in part by the NIH/National Institute of Diabetes and Digestive and Kidney Diseases grant P30 DK34987. LG was supported by a research contract of the FPI Program from MINECO (BES-2012-062502). SD was supported by a research contract from MINECO associated to the project BIO2014-55019-JIN.

    Conflict of Interest



    The authors declare that there is no conflict of interests regarding the publication of this article.

    [1] Xiao CW (2008) Health effects of soy protein and isoflavones in humans. J Nutr 138: 1244S–1249S. doi: 10.1093/jn/138.6.1244S
    [2] Bolanos R, Del Castillo A, Francia J (2010) Soy isoflavones versus placebo in the treatment of climacteric vasomotor symptoms: systematic review and meta-analysis. Menopause 17: 660–666.
    [3] North American Menopause Society (2011) The role of soy isoflavones in menopausal health: report of The North American Menopause Society/Wulf H. Utian Translational Science Symposium in Chicago, IL (October 2010). Menopause 18: 732–753.
    [4] EFSA (European Food Science Authority) (2012) Scientific Opinion on the substantiation of health claims related to soy isoflavones and maintenance of bone mineral density (ID 1655) and reduction of vasomotor symptoms associated with menopause (ID 1654, 1704, 2140, 3093, 3154, 3590) (further assessment) pursuant to Article 13(1) of Regulation (EC) No 1924/2006. EFSA J 10:2847. doi: 10.2903/j.efsa.2012.2847
    [5] Crozier A, Jaganath IB, Clifford MN (2009) Dietary phenolics: chemistry, bioavailability and effects on health. Nat Prod Rep 26: 1001–1043. doi: 10.1039/b802662a
    [6] de Cremoux P, This P, Leclercq G, et al. (2010) Controversies concerning the use of phytoestrogens in menopause management: bioavailability and metabolism. Maturitas 65: 334–339. doi: 10.1016/j.maturitas.2009.12.019
    [7] Sánchez-Calvo JM, Rodríguez-Iglesias MA, Molinillo JMG, et al. (2013) Soy isoflavones and their relationship with microflora: beneficial effects on human health in equol producers. Phytochem Rev 12: 979–1000. doi: 10.1007/s11101-013-9329-x
    [8] Atkinson C, Frankenfeld CL, Lampe JW (2005) Gut bacterial metabolism of the soy isoflavone daidzein: exploring the relevance to human health. Exp Biol Med (Maywood) 230: 155–170. doi: 10.1177/153537020523000302
    [9] Messina M (2016) Soy and Health Update: Evaluation of the Clinical and Epidemiologic Literature. Nutrients 8: E754. doi: 10.3390/nu8120754
    [10] Setchell KD, Clerici C (2010) Equol: history, chemistry, and formation. J Nutr 140: 1355S–1362S. doi: 10.3945/jn.109.119776
    [11] Kemperman RA, Bolca S, Roger LC, et al. (2010) Novel approaches for analysing gut microbes and dietary polyphenols: challenges and opportunities. Microbiology 156: 3224–3231. doi: 10.1099/mic.0.042127-0
    [12] Clavel T, Lepage P, Charrier C (2014) The Family Coriobacteriaceae. In: Rosenberg E, DeLong EF, Lory S, et al. The Prokaryotes, 11 Eds., Berlin: Springer, 201–238.
    [13] Dueñas M, Muñoz-González I, Cueva C, et al. (2015) A survey of modulation of gut microbiota by dietary polyphenols. Biomed Res Int 2015: 850902.
    [14] Nakatsu CH, Armstrong A, Clavijo AP, et al. (2014) Fecal bacterial community changes associated with isoflavone metabolites in postmenopausal women after soy bar consumption. PLoS One 9: e108924. doi: 10.1371/journal.pone.0108924
    [15] Bolca S, Possemiers S, Herregat A, et al. (2007) Microbial and dietary factors are associated with the equol producer phenotype in healthy postmenopausal women. J Nutr 137: 2242–2246. doi: 10.1093/jn/137.10.2242
    [16] Clavel T, Fallani M, Lepage P, et al. (2005) Isoflavones and functional foods alter the dominant intestinal microbiota in postmenopausal women. J Nutr 135: 2786–2792. doi: 10.1093/jn/135.12.2786
    [17] Guadamuro L, Delgado S, Redruello B, et al. (2015) Equol status and changes in fecal microbiota in menopausal women receiving long-term treatment for menopause symptoms with a soy-isoflavone concentrate. Front Microbiol 6: 777
    [18] Nocker A, Cheung CY, Camper AK (2006) Comparison of propidium monoazide with ethidium monoazide for differentiation of live vs. dead bacteria by selective removal of DNA from dead cells. J Microbiol Methods 67: 310–320.
    [19] Redruello B, Guadamuro L, Cuesta I, et al. (2015) A novel UHPLC method for the rapid and simultaneous determination of daidzein, genistein and equol in human urine. J Chromatogr B Analyt Technol Biomed Life Sci 1005: 1–8. doi: 10.1016/j.jchromb.2015.09.029
    [20] Zoetendal EG, Heilig HG, Klaassens ES, et al. (2006) Isolation of DNA from bacterial samples of the human gastrointestinal tract. Nat Protoc 1: 870–873 doi: 10.1038/nprot.2006.142
    [21] Young JP, Downer HL, Eardly BD (1991) Phylogeny of the phototrophic rhizobium strain BTAi1 by polymerase chain reaction-based sequencing of a 16S rRNA gene segment. J Bacteriol 173: 2271–2277. doi: 10.1128/jb.173.7.2271-2277.1991
    [22] Goecks J, Nekrutenko A, Taylor J, et al. (2010) Galaxy: a comprehensive approach for supporting accessible, reproducible, and transparent computational research in the life sciences. Genome Biol 11: R86. doi: 10.1186/gb-2010-11-8-r86
    [23] Edgar RC, Haas BJ, Clemente JC, et al. (2011) UCHIME improves sensitivity and speed of chimera detection. Bioinformatics 27: 2194–2200. doi: 10.1093/bioinformatics/btr381
    [24] Wang Q, Garrity GM, Tiedje JM, et al. (2007) Naive Bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy. Appl Environ Microbiol 73: 5261–5267. doi: 10.1128/AEM.00062-07
    [25] Moreno CE, Halffter G (2001) On the measure of sampling effort used in species accumulation curves. J Appl Ecol 38: 487–490. doi: 10.1046/j.1365-2664.2001.00590.x
    [26] Yarza P, Richter M, Peplies J, et al. (2008) The All-Species Living Tree project: a 16S rRNA-based phylogenetic tree of all sequenced type strains. Syst Appl Microbiol 31: 241–250. doi: 10.1016/j.syapm.2008.07.001
    [27] Li W, Godzik A (2006) Cd-hit: a fast program for clustering and comparing large sets of protein or nucleotide sequences. Bioinformatics 22: 1658–1659. doi: 10.1093/bioinformatics/btl158
    [28] Graf D, Di Cagno R, Fak F, et al. (2015) Contribution of diet to the composition of the human gut microbiota. Microb Ecol Health Dis 26: 26164.
    [29] David LA, Maurice CF, Carmody RN, et al. (2014) Diet rapidly and reproducibly alters the human gut microbiome. Nature 505: 559–563. doi: 10.1038/nature12820
    [30] Singh RK, Chang HW, Yan D, et al. (2017) Influence of diet on the gut microbiome and implications for human health. J Transl Med, 15: 73. doi: 10.1186/s12967-017-1175-y
    [31] van Duynhoven J, Vaughan EE, Jacobs DM, et al. (2011) Metabolic fate of polyphenols in the human superorganism. Proc Natl Acad Sci USA 108: 4531–4538. doi: 10.1073/pnas.1000098107
    [32] Rowland IR, Wiseman H, Sanders TA, et al. (2000) Interindividual variation in metabolism of soy isoflavones and lignans: influence of habitual diet on equol production by the gut microflora. Nutr Cancer 36: 27–32. doi: 10.1207/S15327914NC3601_5
    [33] Guadamuro L, Dohrmann AB, Tebbe CC, et al. (2017) Bacterial communities and metabolic activity of faecal cultures from equol producer and non-producer menopausal women under treatment with soy isoflavones. BMC Microbiol 17: 93. doi: 10.1186/s12866-017-1001-y
    [34] Meehan CJ, Beiko RG (2014) A phylogenomic view of ecological specialization in the Lachnospiraceae, a family of digestive tract-associated bacteria. Genome Biol Evol 6: 703–713. doi: 10.1093/gbe/evu050
    [35] Rios-Covian D, Ruas-Madiedo P, Margolles A, et al. (2016) Intestinal short chain fatty acids and their link with diet and human health. Front Microbiol 7: 185.
    [36] Kim MG, Lee HS (2009) Growth-inhibiting activities of phenethyl isothiocyanate and its derivatives against intestinal bacteria. J Food Sci 74: M467–471. doi: 10.1111/j.1750-3841.2009.01333.x
    [37] Sklenickova O, Flesar J, Kokoska L, et al. (2010) Selective growth inhibitory effect of biochanin A against intestinal tract colonizing bacteria. Molecules 15: 1270–1279. doi: 10.3390/molecules15031270
    [38] Vazquez L, Guadamuro L, Giganto F, et al. (2017) Development and use of a real-time quantitative PCR method for detecting and quantifying equol-producing bacteria in human faecal samples and slurry cultures. Front Microbiol 8: 1155. doi: 10.3389/fmicb.2017.01155
    [39] Wagner AO, Malin C, Knapp BA, et al. (2008) Removal of free extracellular DNA from environmental samples by ethidium monoazide and propidium monoazide. Appl Environ Microbiol 74: 2537–2539. doi: 10.1128/AEM.02288-07
    [40] Nocker A, Richter-Heitmann T, Montijn R, et al. (2010) Discrimination between live and dead cellsin bacterial communities from environmental water samples analyzed by 454 pyrosequencing. Int Microbiol 13: 59–65.
    [41] Hein I, Flekna G, Wagner M, et al. (2006) Possible errors in the interpretation of ethidium bromide and PicoGreen DNA staining results from ethidium monoazide-treated DNA. Appl Environ Microbiol 72: 6860–6861. doi: 10.1128/AEM.01243-06
  • Reader Comments
  • © 2019 the Author(s), licensee AIMS Press. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0)
通讯作者: 陈斌, bchen63@163.com
  • 1. 

    沈阳化工大学材料科学与工程学院 沈阳 110142

  1. 本站搜索
  2. 百度学术搜索
  3. 万方数据库搜索
  4. CNKI搜索

Metrics

Article views(4577) PDF downloads(1576) Cited by(10)

Article outline

Figures and Tables

Figures(4)  /  Tables(2)

/

DownLoad:  Full-Size Img  PowerPoint
Return
Return

Catalog