Evidence that the endometrial microbiota has an effect on implantation success or failure

Published:October 05, 2016DOI:


      Bacterial cells in the human body account for 1–3% of total body weight and are at least equal in number to human cells. Recent research has focused on understanding how the different bacterial communities in the body (eg, gut, respiratory, skin, and vaginal microbiomes) predispose to health and disease. The microbiota of the reproductive tract has been inferred from the vaginal bacterial communities, and the uterus has been classically considered a sterile cavity. However, while the vaginal microbiota has been investigated in depth, there is a paucity of consistent data regarding the existence of an endometrial microbiota and its possible impact in reproductive function.


      This study sought to test the existence of an endometrial microbiota that differs from that in the vagina, assess its hormonal regulation, and analyze the impact of the endometrial microbial community on reproductive outcome in infertile patients undergoing in vitro fertilization.

      Study Design

      To identify the existence of an endometrial microbiota, paired samples of endometrial fluid and vaginal aspirates were obtained simultaneously from 13 fertile women in prereceptive and receptive phases within the same menstrual cycle (total samples analyzed n = 52). To investigate the hormonal regulation of the endometrial microbiota during the acquisition of endometrial receptivity, endometrial fluid was collected at prereceptive and receptive phases within the same cycle from 22 fertile women (n = 44). Finally, the reproductive impact of an altered endometrial microbiota in endometrial fluid was assessed by implantation, ongoing pregnancy, and live birth rates in 35 infertile patients undergoing in vitro fertilization (total samples n = 41) with a receptive endometrium diagnosed using the endometrial receptivity array. Genomic DNA was obtained either from endometrial fluid or vaginal aspirate and sequenced by 454 pyrosequencing of the V3–V5 region of the 16S ribosomal RNA (rRNA) gene; the resulting sequences were taxonomically assigned using QIIME. Data analysis was performed using R packages. The χ2 test, Student t test, and analysis of variance were used for statistical analyses.


      When bacterial communities from paired endometrial fluid and vaginal aspirate samples within the same subjects were interrogated, different bacterial communities were detected between the uterine cavity and the vagina of some subjects. Based on its composition, the microbiota in the endometrial fluid, comprising up to 191 operational taxonomic units, was defined as a Lactobacillus-dominated microbiota (>90% Lactobacillus spp.) or a non-Lactobacillus-dominated microbiota (<90% Lactobacillus spp. with >10% of other bacteria). Although the endometrial microbiota was not hormonally regulated during the acquisition of endometrial receptivity, the presence of a non-Lactobacillus-dominated microbiota in a receptive endometrium was associated with significant decreases in implantation [60.7% vs 23.1% (P = .02)], pregnancy [70.6% vs 33.3% (P = .03)], ongoing pregnancy [58.8% vs 13.3% (P = .02)], and live birth [58.8% vs 6.7% (P = .002)] rates.


      Our results demonstrate the existence of an endometrial microbiota that is highly stable during the acquisition of endometrial receptivity. However, pathological modification of its profile is associated with poor reproductive outcomes for in vitro fertilization patients. This finding adds a novel microbiological dimension to the reproductive process.

      Key words

      To read this article in full you will need to make a payment


        • Sirota I.
        • Zarek S.M.
        • Segars J.H.
        Potential influence of the microbiome on infertility and assisted reproductive technology.
        Semin Reprod Med. 2014; 32: 35-42
        • González A.
        • Vázquez-Baeza Y.
        • Knight R.
        SnapShot: the human microbiome.
        Cell. 2014; 158: 690-691
        • Burton J.P.
        • Reid G.
        Evaluation of the bacterial vaginal flora of 20 postmenopausal women by direct (Nugent score) and molecular (polymerase chain reaction and denaturing gradient gel electrophoresis) techniques.
        J Infect Dis. 2002; 186: 1770-1780
        • Pavlova S.I.
        • Kilic A.O.
        • Kilic S.S.
        • et al.
        Genetic diversity of vaginal lactobacilli from women in different countries based on 16S rRNA gene sequences.
        J Appl Microbiol. 2002; 92: 451-459
        • Gajer P.
        • Brotman R.M.
        • Bai G.
        • et al.
        Temporal dynamics of the human vaginal microbiota.
        Sci Transl Med. 2012; 4 (132ra52)
        • Onderdonk A.B.
        • Delaney M.L.
        • Fichorova R.N.
        The human microbiome during bacterial vaginosis.
        Clin Microbiol Rev. 2016; 29: 223-238
        • Romero R.
        • Hassan S.S.
        • Gajer P.
        • et al.
        The composition and stability of the vaginal microbiota of normal pregnant women is different from that of non-pregnant women.
        Microbiome. 2014; 2: 4
        • Ravel J.
        • Gajer P.
        • Abdo Z.
        • et al.
        Vaginal microbiome of reproductive-age women.
        Proc Natl Acad Sci U S A. 2011; 108: 4680-4687
        • Romero R.
        • Hassan S.S.
        • Gajer P.
        • et al.
        The vaginal microbiota of pregnant women who subsequently have spontaneous preterm labor and delivery and those with a normal delivery at term.
        Microbiome. 2014; 2: 18
        • Ralph S.G.
        • Rutherford A.J.
        • Wilson J.D.
        Influence of bacterial vaginosis on conception and miscarriage in the first trimester: cohort study.
        BMJ. 1999; 319: 220-223
        • Hay P.E.
        • Lamont R.F.
        • Taylor-Robinson D.
        • Morgan D.J.
        • Ison C.
        • Pearson J.
        Abnormal bacterial colonization of the genital tract and subsequent preterm delivery and late miscarriage.
        BMJ. 1994; 308: 295-298
        • Romero R.
        • Chaiworapongsa T.
        • Kuivaniemi H.
        • Tromp G.
        Bacterial vaginosis, the inflammatory response and the risk of preterm birth: a role for genetic epidemiology in the prevention of preterm birth.
        Am J Obstet Gynecol. 2004; 190: 1509-1519
        • Fanchin R.
        • Harmas A.
        • Benaoudia F.
        • Lundkvist U.
        • Olivennes F.
        • Frydman R.
        Microbial flora of the cervix assessed at the time of embryo transfer adversely affects in vitro fertilization outcome.
        Fertil Steril. 1998; 70: 866-870
        • Egbase P.E.
        • al-Sharhan M.
        • al-Othman S.
        • al-Mutawa M.
        • Udo E.E.
        • Grudzinskas J.G.
        Incidence of microbial growth from the tip of the embryo transfer catheter after embryo transfer in relation to clinical pregnancy rate following in-vitro fertilization and embryo transfer.
        Hum Reprod. 1996; 11: 1687-1689
        • Moore D.E.
        • Soules M.R.
        • Klein N.A.
        • Fujimoto V.Y.
        • Agnew K.J.
        • Eschenbach D.A.
        Bacteria in the transfer catheter tip influence the live-birth rate after in vitro fertilization.
        Fertil Steril. 2000; 74: 1118-1124
        • Salim R.
        • Ben-Shlomo I.
        • Colodner R.
        • Keness Y.
        • Shalev E.
        Bacterial colonization of the uterine cervix and success rate in assisted reproduction: results of a prospective survey.
        Hum Reprod. 2002; 17: 337-340
        • Selman H.
        • Mariani M.
        • Barnocchi N.
        • et al.
        Examination of bacterial contamination at the time of embryo transfer, and its impact on the IVF/pregnancy outcome.
        J Assist Reprod Genet. 2007; 24: 395-399
        • Romero R.
        • Espinoza J.
        • Mazor M.
        Can endometrial infection/inflammation explain implantation failure, spontaneous abortion, and preterm birth after in vitro fertilization?.
        Fertil Steril. 2004; 82: 799-804
        • Møller B.R.
        • Kristiansen F.V.
        • Thorsen P.
        • Frost L.
        • Mogensen S.C.
        Sterility of the uterine cavity.
        Acta Obstet Gynecol Scand. 1995; 74: 216-219
        • Mitchell C.M.
        • Haick A.
        • Nkwopara E.
        • et al.
        Colonization of the upper genital tract by vaginal bacterial species in nonpregnant women.
        Am J Obstet Gynecol. 2015; 212: 611.e1-611.e9
        • Racicot K.
        • Cardenas I.
        • Wunsche V.
        • et al.
        Viral infection of the pregnant cervix predisposes to ascending bacterial infection.
        J Immunol. 2013; 191: 934-941
        • Cho I.
        • Blaser M.J.
        The human microbiome: at the interface of health and disease.
        Nat Rev Genet. 2012; 13: 260-270
        • Vilella F.
        • Ramirez L.
        • Berlanga O.
        • et al.
        PGE2 and PGF2 concentrations in human endometrial fluid as biomarkers for embryonic implantation.
        J Clin Endocrinol Metab. 2013; 98: 4123-4132
        • Ruiz-Alonso M.
        • Blesa D.
        • Díaz-Gimeno P.
        • et al.
        The endometrial receptivity array for diagnosis and personalized embryo transfer as a treatment for patients with repeated implantation failure.
        Fertil Steril. 2013; 100: 818-824
        • Sim K.
        • Cox M.J.
        • Wopereis H.
        • Martin R.
        • Knol J.
        • Li M.S.
        Improved detection of bifidobacteria with optimized 16S rRNA-gene based pyrosequencing.
        PLoS One. 2012; 7: e32543
        • Caporaso J.G.
        • Kuczynski J.
        • Stombaugh J.
        • et al.
        QIIME allows analysis of high-throughput community sequencing data.
        Nat Methods. 2010; 7: 335-336
        • Edgar R.C.
        Search and clustering orders of magnitude faster than BLAST.
        Bioinformatics. 2010; 26: 2460-2461
        • Cole J.R.
        • Wang Q.
        • Cardenas E.
        • et al.
        The ribosomal database project: improved alignments and new tools for rRNA analysis.
        Nucleic Acids Res. 2009; 37: D141-D145
      1. Shannon CE. The mathematical theory of communication. University of Illinois Press. Illinis book edition, 1963.

      2. Simpson EH. Measurement of diversity. Nature 1949;163:688-8.

        • DiGiulio D.B.
        • Callahan B.J.
        • McMurdie P.J.
        • et al.
        Temporal and spatial variation of the human microbiota during pregnancy.
        Proc Natl Acad Sci U S A. 2015; 112: 11060-11065
        • Spurbeck R.R.
        • Arvidson C.G.
        Inhibition of Neisseria gonorrhoeae epithelial cell interactions by vaginal Lactobacillus species.
        Infect Immun. 2008; 76: 3124-3130
        • Hyman R.W.
        • Herndon C.N.
        • Jiang H.
        • et al.
        The dynamics of the vaginal microbiome during infertility therapy with in vitro fertilization-embryo transfer.
        J Assist Reprod Genet. 2012; 29: 105-115
        • Liversedge N.H.
        • Turner A.
        • Horner P.J.
        • Keay S.D.
        • Jenkins J.M.
        • Hull M.G.
        The influence of bacterial vaginosis on in-vitro fertilization and embryo implantation during assisted reproduction treatment.
        Hum Reprod. 1999; 14: 2411-2415
        • van Oostrum N.
        • De Sutter P.
        • Meys J.
        • Verstraelen H.
        Risks associated with bacterial vaginosis in infertility patients: a systematic review and meta-analysis.
        Hum Reprod. 2013; 28: 1809-1815
        • Egbase P.E.
        • Udo E.E.
        • al-Sharhan M.
        • Grudzinskas J.G.
        Prophylactic antibiotics and endocervical microbial inoculation of the endometrium at embryo transfer.
        Lancet. 1999; 354: 651-652
        • Franasiak J.M.
        • Werner M.D.
        • Juneau C.R.
        • et al.
        Endometrial microbiome at the time of embryo transfer: next-generation sequencing of the 16S ribosomal subunit.
        J Assist Reprod Genet. 2016; 33: 129-136
        • Skarin A.
        • Sylwan J.
        Vaginal lactobacilli inhibiting growth of Gardnerella vaginalis, Mobiluncus and other bacterial species cultured from vaginal content of women with bacterial vaginosis.
        Acta Pathol Microbiol Immunol Scand B. 1986; 94: 399-403
        • Yamamoto T.
        • Zhou X.
        • Williams C.J.
        • Hochwalt A.
        • Forney L.J.
        Bacterial populations in the vaginas of healthy adolescent women.
        J Pediatr Adolesc Gynecol. 2009; 22: 11-18
        • Dominguez F.
        • Gadea B.
        • Mercader A.
        • Esteban F.J.
        • Pellicer A.
        • Simón C.
        Embryologic outcome and secretome profile of implanted blastocysts obtained after coculture in human endometrial epithelial cells versus the sequential system.
        Fertil Steril. 2010; 93: 774-782.e1
      3. Cha J, Vilella F, Dey SK, Simón C. Molecular interplay in successful implantation. Science. S. Sanders. Washington; Nov. 12, 2013; Ten critical topics in reproductive medicine: 44-8.

      Supplementary References

        • van der Gaast M.H.
        • Beier-Hellwig K.
        • Fauser B.C.J.M.
        • Beier H.M.
        • Macklon N.S.
        Endometrial secretion aspiration prior to embryo transfer does not reduce implantation rates.
        Reprod Biomed Online. 2003; 7: 105-109
        • Díaz-Gimeno P.
        • Horcajadas J.A.
        • Martínez-Conejero J.A.
        • et al.
        A genomic diagnostic tool for human endometrial receptivity based on the transcriptomic signature.
        Fertil Steril. 2011; 95: 50-60.e1-15
        • Díaz-Gimeno P.
        • Ruiz-Alonso M.
        • Blesa D.
        • et al.
        The accuracy and reproducibility of the endometrial receptivity array is superior to histology as a diagnostic method for endometrial receptivity.
        Fertil Steril. 2013; 99: 508-517
        • Pearson W.R.
        • Wood T.
        • Zhang Z.
        • Miller W.
        Comparison of DNA sequences with protein sequences.
        Genomics. 1997; 46: 24-36
        • Martin M.
        Cutadapt removes adapter sequences from high-throughput sequencing reads.
        EMBnet J. 2011; 17: 10
        • Edgar R.C.
        • Haas B.J.
        • Clemente J.C.
        • Quince C.
        • Knight R.
        UCHIME improves sensitivity and speed of chimera detection.
        Bioinformatics. 2011; 27: 2194-2200
        • Fawcett T.
        An introduction to ROC analysis.
        ROC Analysis in Pattern Recognition. 2006; 27: 861-874