Systematic Comparison of Bacterial Colonization of Endometrial Tissue and Fluid Samples in Recurrent Miscarriage Patients: Implications for Future Endometrial Microbiome Studies

Abstract BACKGROUND A recent study has reported that the microbiota in endometrial fluid of patients receiving in vitro fertilization and embryo transfer (IVF-ET) may predict implantation and pregnancy rates. However, studies are lacking that simultaneously compare the microbiota between endometrial fluid and tissue samples. Whether the microbiota composition in endometrial fluid reflects that in the endometrial tissue remains unclear. METHODS We systematically profiled the microbiota in endometrial fluid and tissue samples of IVF-ET patients using massively parallel sequencing. The bacterial 16S ribosomal RNA gene (V4 region) was PCR-amplified. Sequencing reads with >98% nucleotide identity were clustered as a bacterial taxon. To account for the different number of reads per sample, we normalized the read counts of each taxon before comparing its relative abundances across samples. RESULTS Thirteen taxa, including Verrucomicrobiaceae, Brevundimonas, Achromobacter, Exiguobacterium, and Flavobacterium, were consistently detected only in endometrial tissue samples but not fluid samples. Eight taxa were detected in fluid but not tissue. Twenty-two taxa were differentially abundant between fluid and tissue samples (adjusted P values, 4.1 × 10−25 to 0.025). The numbers of taxa identified per 1000 sequencing reads, diversity, and evenness in fluid samples were smaller than those in tissue samples. CONCLUSIONS Our data suggest that the microbiota composition in endometrial fluid does not fully reflect that in endometrial tissue. Sampling from both endometrial fluid and biopsy allows a more comprehensive view of microbial colonization. Further efforts are needed to identify the preanalytical effects, including sampling sites, methods, and sequencing depth, on profiling endometrial microbiota.