The invention discloses a regionally cultured aquatic miniature organism nourishing agent and a preparation method thereof. The nourishing agent is composed of culture pond muddy water, an indigenous microorganism flora enriched culture medium, and a microorganism culture material; wherein the weight ratio of the microorganism culture material to the culture pond muddy water is 1: (0.6-1); and the indigenous microorganism flora enriched culture medium accounts for 10 to 20% of the total weight of the microorganism culture material and the culture pond muddy water. The nourishing agent can optimize the diversity of miniature organisms in a culture pond. After using the nourishing agent, the pool nutrient type alga structure is converted into medium nutrient type alga structure composed of beneficial algae communities such as green alga. Moreover, the nourishing agent can reduce the death rate of fish, and the using amount of medicine is reduced. Moreover, the fish quality is improved, the yield of each mu is increased, the feed coefficient is reduced, the fish color becomes healthier, the taste becomes better, and the fish meat does not have any fishy smell and is tight.