Fig. 1

Study design. Primigravid (Prim.) cows were weighed twice at 240 d of gestation and the mean value was used as their prepartum body weight (ppBW). Multiparous (Mult.) cows were weighted at -2, -1, and 0 days relative to dry-off from the lactation preceding enrolment. The 3 measurements were used to generate a mean ppBW. On 0, 1, and 2 days relative to calving primiparous and multiparous cows were weighted. The 3 measurements were used to generate a mean calving BW (cBW). To only account for the body weight change (BWC) related to tissue accretion or mobilization, the weight of the gravid uterus prepartum and the weight of the empty uterus right after parturition were subtracted from the ppBW and cBW, respectively. The weight of the gravid uterus and empty uterus were calculated using NASEM (2021) equations. After excluding the weight of the gravid uterus from the ppBW and the weight of the empty uterus from the cBW, the BWC prepartum was calculated as the difference between cBW and ppBW divided by the number of days between measurements. To account for any differences in BWC associated with frame, BWC then was calculated as a percentage of cBW. Uterine discharge was evaluated using a Metricheck device (Metricheck, Simcro, New Zealand) at 3 ± 1, 7 ± 1, 10 ± 1 and 13 ± 1 days after calving. Cows diagnosed with metritis up to day 10 were considered cases for this study. There were 52 metritis cases, and 10, 23, and 19 cows were diagnosed with metritis on day 3, 7, and 10, respectively. Two cows were excluded because of several missing values for Bayesian networks on the day of metritis diagnosis. Control cows (n = 50) that did not develop metritis were matched with 50 cows that developed metritis, hence, 100 cows were used for bioinformatic and statistical analyses. All cows had blood collected prepartum (14 ± 6 d before calving), at calving (first 24 h after calving), and at the day of metritis diagnosis (7 ± 2 d after calving). All cows had uterine fluid collected at calving and at diagnosis of metritis. Blood samples were used for flow cytometry, multiplex, and untargeted gas chromatography mass spectrometry (GC-TOF-MS). Flow cytometry and multiplex were performed to assess the systemic immune profile and activation status. Gas chromatography mass spectrometry was performed to assess the systemic metabolomic profile. Uterine fluid samples were used for untargeted GC-TOF-MS and 16 S rRNA gene amplification by PCR. Gas chromatography mass spectrometry was performed to assess the uterine metabolomic profile and the 16 S rRNA gene was amplified by PCR to assess the uterine microbiome