The invention discloses a nested PCR (polymerase chain reaction) kit for detecting duck-manure pollution in a water body and a detection method thereof, and belongs to the field of the detection of pollution in water bodies. The nested PCR kit for detecting the duck-manure pollution in the water body comprises two pairs of nested PCR primers, dNTP (deoxyribonucleoside triphosphate), a PCR buffer, a Taq polymerase, a Mg<2+> solution and ddH2O (double distilled water). Two rounds of PCR amplification are used by a nested PCR method. The detection method comprises the following steps of extracting a DNA (deoxyribonucleic acid) in a to-be-detected water sample, and carrying out a first round of PCR amplification by using the DNA in the to-be-detected water sample as a template, wherein the primers are an SDF (stroma derived factor) and SDR (short-chain dehydrogenase / reductase); after a first reaction is terminated, carrying out a second round of PCR amplification by using a product of the first round of PCR amplification as a template, wherein the primers are an NDF (neutral detergent fiber)and NDR, and after a second reaction is terminated, detecting the existence of a 158bp strip, which shows that the water body is subjected to the duck-manure pollution, through agarose gel electrophoresis. According to the method, the detection sensitivity is greatly increased through nested PCR amplification; a little duck-manure pollution which exists in water can be detected; moreover, the method has a favorable specificity, and can be directly applied to the detection and the preventive treatment work of fecal pollution in the water body.