In the ecological environment, cadmium, a heavy metal produced from human activities and industry toxic material, has polluted the water and affects the reproductive health of aquatic biota. Many fish farmers use water from the river for freshwater fish cultivation. This study examined the effects of supplementation feed (probiotics and vitamin C) on sperm quality and total bacteria in fish intestine, Oreochromis niloticus, after Cadmium (Cd) exposure. We found that probiotics did not seem to colonise fish intestine or change the overall amount of the intestinal microbiota. However, probiotic supplementation actually changed the total amount of bacteria in the fish intestine. In particular, the administration of probiotics in fish exposed to Cd has increased more intestinal bacteria compared to the administration of vitamin C. The addition of vitamin C as a supplement did not affect the total amount of bacteria in the fish intestine. These results indicated that probiotics have a significant capacity to recover sperm motility in freshwater fish exposed to Cd pollution.