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Abstract Background The goal of this study was to assess the microbial ecology and diversity present in the uterus of post-partum dairy cows with and without metritis from 24 commercial California dairy farms using shotgun metagenomics. A set subset of 95 intrauterine swab samples, taken from a larger selection of 307 individual cow samples previously collected, were examined for α and β diversity and differential abundance associated with metritis. Cows within 21 days post-partum were categorized into one of three clinical groups during sample collection: control (CT, n = 32), defined as cows with either no vaginal discharge or a clear, non-purulent mucus vaginal discharge; metritis (MET, n = 33), defined as a cow with watery, red or brown colored, and fetid vaginal discharge; and purulent discharge cows (PUS, n = 31), defined as a non-fetid purulent or mucopurulent vaginal discharge. Results All three clinical groups (CT, MET, and PUS) were highly diverse, with the top 12 most abundant genera accounting for 10.3%, 8.8%, and 10.1% of mean relative abundance, respectively. The α diversity indices revealed a lower diversity from samples collected from MET and PUS when compared to CT cows. PERMANOVA statistical testing revealed a significant difference (P adjusted |