Background <p>Previous study has found that gastrointestinal tract (GIT) bacteria are involved in the regulation of medium-chain fatty acid triglycerides (MCTs) on the improvement of growth performance in finishing bulls. At present, the effects of MCT on GIT fungi of finishing bulls has not been reported, whether fungi are also involved in the regulation of the growth performance is still unknown. The objective of this study was to investigate GIT fungal composition and diversity of finishing bulls fed different MCT.</p> Results <p>The study employed four ruminally fistulated Yanbian cattle (18-month-old bulls) in a 4 × 4 Latin square design, and were divided into four treatment groups: a control group (CON) receiving a basal total mixed ration (TMR), and three experimental groups (C8, C10, and C12) supplemented with 60 g/bull/day of glycerol monocaprylin (GMC), glycerol monodecanoate (GMD), and glycerol monolaurate (GML), respectively. At the end of each experimental period, rumen fluid, rumen digesta, and rectum feces were collected for ITS sequencing. In the rumen digesta, the relative abundance of o_<i>Eurotiales</i> and f_<i>Aspergillaceae</i> in the C12 group were significantly increased compared with the CON, C8, and C10 groups (<i>P</i> = 0.038 and 0.028, respectively), while in the rumen fluid, the relative abundance of g_<i>Cyllamyces</i> in the C10 group was significantly increased compared with the CON, C8, and C12 groups (<i>P</i> = 0.036).</p> Conclusions <p>There were some differences in the composition and diversity of fungi among different samples and different MCT treatment groups in finishing bulls. Moreover, based on correlation analysis, it is found that GIT fungi may also be involved in the regulation of GMC/GMD/GML on the improvement of feed intake, antioxidant status or nutrient digestibility in finishing bulls.</p>