A validated low-abundance uterine microbial signature associated with chronic endometritis

ABSTRACT Background: Chronic endometritis (CE) is a persistent inflammatory endometrial disorder associated with infertility, recurrent implantation failure, and pregnancy loss. While previous microbiome studies have focused on Lactobacillus depletion and common genital tract bacteria, low-abundance microbial lineages remain poorly characterised. Objective: To identify reproducible low-abundance microbial signatures associated with CE and evaluate their robustness through external validation and complementary analyses. Methods: Uterine lavage specimens from infertile women with CE ( n = 29) and non-CE controls ( n = 16) were analysed by 16S rRNA sequencing in a discovery cohort. An independent cohort of 58 participants was used for external validation. CE was diagnosed by hysteroscopy and CD138 immunohistochemistry (≥5 stromal plasma cells/10 high-power fields). Multiple negative controls were included, and contaminants were assessed using decontam. Prespecified analyses included cervicovaginal source tracking, shotgun metagenomics in a subset, cytokine profiling, and post-treatment follow-up. Results: CE samples exhibited increased richness and modestly higher diversity. Presence/absence-based beta diversity demonstrated clearer separation between CE and controls. CE was associated with enrichment of Patescibacteria, Planctomycetota, Bdellovibrio nota, and selected archaeal-lineage signals. After contaminant filtering, major rare-lineage features remained enriched. Patescibacteria and Bdellovibrio -related signals correlated with CD138-positive plasma-cell counts and inflammatory cytokines. A nested discovery model achieved an area under the receiver operating characteristic curve (AUC) of 0.84, and the locked marker panel achieved an AUC of 0.81 in external validation. Conclusions: CE is associated with a reproducible low-abundance microbial signature in uterine lavage samples. These findings highlight candidate microbial lineages for further investigation but do not establish causality or uterine residency.