This study investigated the performance of an ecological filter (EF) and vermifiltration (VF) system, the effects of substrate and earthworms on the diversity and abundance of denitrifying genes coding for nitrite (nirS and nirK) reductases and on denitrifying rate, and the factors influencing denitrification. The majority of organic matter, ammonia nitrogen and total nitrogen from sewage was removed by the soil layer in both reactors, and their total removal efficiencies increased in VF compared with those in EF. Additionally, substrate in the reactors significantly influenced the Shannon diversity index and abundance of nirS and nirK, as well as the denitrifying rate. However, the earthworms only significantly influenced nirS diversity. Furthermore, evaluation of the factors controlling denitrification implied that increasing NH3-N availability, diversity and abundance of nirS and nirK or decreasing available NO3-N might be responsible for the enhanced denitrification activity obtained using VF for rural domestic wastewater treatment. (C) 2016 Elsevier Ltd. All rights reserved.