The coupled nitrification-denitrification process plays a pivotal role in cycling and removal of nitrogen in aquatic ecosystems. In the present study, the communities of ammonia oxidizers and denitrifiers in the sediments of 2 basins (Guozhenghu Basin and Tuanhu Basin) of a large urban eutrophic lake (Lake Donghu) were determined using the ammonia monooxygenase subunit A (amoA) gene and the nitrite reductase gene. At all sites of this study, the archaeal amoA gene predominated over the bacterial amoA gene, whereas the functional gene for denitrification nirK gene far outnumbered the nirS gene. Spatially, compared with the Tuanhu Basin, the Guozhenghu Basin showed a significantly greater abundance of the archaeal amoA gene but less abundance of the nirK and nirS genes, while there was no significant difference of bacterial amoA gene copy numbers between the 2 basins. Unlike the archaeal amoA gene, the nirK gene showed a significant difference in community structure between the 2 basins. Archaeal amoA diversity was limited to the water-sediment cluster of Crenarchaeota, in sharp contrast with nirK for which 22 distinct operational taxonomic units were found. Accumulation of organic substances were found to be positively related to nirK and nirS gene copy numbers but negatively related to archaeal amoA gene copy numbers, whereas the abundance of the bacterial amoA gene was related to ammonia concentration.