First detection of Leptospira spp. in cumulus-oocyte complexes from naturally infected cows

Although a few studies have reported Leptospira spp. in cumulus-oocyte complexes (COCs) from experimentally infected cows, it remains unclear whether this bacterium can penetrate COCs in naturally infected animals. This study aimed to detect the presence of Leptospira spp. in COCs from naturally infected cows. Ovaries and uterine body fragments were collected from 40 cows after slaughter. Follicular fluid (FF) and uterine fragments (UF) were screened using conventional PCR (cPCR), and negative results were confirmed by quantitative PCR (qPCR). COCs were immunolabeled using a monoclonal anti-LipL41 antibody, followed by an Alexa Fluor 488-conjugated secondary antibody, and counterstained with propidium iodide. Detection of Leptospira spp. was performed by epifluorescence microscopy. Cows were then classified as positive (POS-FF, POS-UF, or both) or negative (NEG). In POS-FF animals (n = 8), 9.1 COCs/female were recovered, with 51.1 ± 12.7 % showing Leptospira presence (not significant; n.s.). In POS-UF cows (n = 12), 6.6 COCs/female were obtained, and 81.0 ± 5.5 % showed bacterial presence (n.s.). In animals positive for both FF and UF (n = 10), 5.4 COCs/female were collected, with 63.7 ± 12.9 % testing positive (n.s.). In NEG animals (n = 10), which were both cPCR- and qPCR-negative, 69.3 ± 9.7 % of COCs exhibited bacterial labeling, suggesting a low bacterial load (n.s.). These cows yielded 9.7 ± 3.7 COCs/female (n.s.). Although no difference was observed between the groups, this is the first report demonstrating the presence of Leptospira spp. in COCs from naturally infected cows, highlighting a mechanism contributing to reproductive failure in cattle.