Abstract
Recent progress in bovine intestinal organoid research has expanded opportunities for creating improved in vitro models to study intestinal physiology and pathology. However, the establishment of a culture condition capable of generating organoids from all segments of the cattle intestine has remained elusive. Although previous research has described the development of bovine jejunal, ileal, and colonic organoids, this study marks the first report of successful bovine duodenal and rectal organoid development. Maintenance of these organoids through serial passages and cryopreservation was achieved, with higher success rates observed in large intestinal organoids compared to their small intestinal counterparts. A novel approach involving the use of biopsy forceps during initial tissue sampling streamlined the subsequent tissue processing, simplifying the procedure compared to previously established protocols in cattle. Additionally, our study introduced a more cost-effective culture medium based on Advanced DMEM/F12, diverging from frequently used commercially available organoid culture media. This enhancement improves accessibility to organoid technology by reducing culture costs. Crucially, the derived organoids from jejunum, ileum, colon and rectum faithfully preserved the structural, cellular, and genetic characteristics of in vivo intestinal tissue. This research underscores the significant potential of adult bovine intestinal organoids as a physiologically and morphologically relevant in vitro model. Such organoids provide a renewable and sustainable resource for a broad spectrum of studies, encompassing investigations into normal intestinal physiology in cattle and the intricate host-pathogen interactions of clinically and economically significant enteric pathogens.