Integrated analysis reveals strong reproducible signals within and across studies of the built environment

Background Built environment microbiome studies have identified numerous factors that shape indoor microbiomes, yet the reproducibility of these findings across buildings, timepoints, and research groups remains unclear. Differences in sequencing protocols, sampling design, and environments pose major challenges for cross-study comparisons, particularly in low-biomass environments where technical variation can obscure biological signal. To address this gap, we constructed a simple ontology which groups samples into one of three categories: hand, hand-associated surfaces, and floor then applied it to four publicly available 16S rRNA gene datasets: a hospital, university dormitory, Air Force dormitory, and private residential houses. Results We identified strong and reproducible separation between floors and surfaces with frequent human contact. We found that floors consistently harbored soil-associated taxa, including KD4-96, 67-14, Skermanella, and Sphingobacterium , whereas hands and hand-associated surfaces were enriched with skin-associated genera, such as Lawsonella and Cutibacterium . Within studies, these results were generally consistent across timepoints. Across studies, mixed-model PERMANOVA analysis revealed significant clustering by sample type, with modest effects of study, suggesting that biological signal outweighed differences in laboratory or sequencing methods. Leave-one-study-out random forest models achieved high AUCs for hand vs. floor comparisons (0.865 to 0.921), moderate AUCs for hand-associated vs. floor comparisons, and weaker performance for hand vs. hand-associated comparisons. Application of the batch-correction method DEBIAS-M did not improve effect sizes or classification performance, indicating that reproducible structure was already discernible without batch adjustment. Conclusions Despite substantial temporal and environmental heterogeneity among studies, we found that the built environment microbiome has a reproducible bacterial signal. There was consistent enrichment of soil-derived taxa on floors and human-associated taxa on hands and hand-associated surfaces suggesting a stable microbiome despite differences in building type, occupancy, and methodology. These findings establish an important foundation for future studies, suggesting cross-study comparability, the accuracy of ecological inference, and the ability to support the development of predictive applications in indoor microbiome research.