Abstract: Two green manure experiments in Hunan and Jiangxi provinces were conducted in this study. The soil types at the two experimental sites were purple alluvial soil in Hunan and yellow clayey soil in Jiangxi. Soil properties, nitrification activity, potential nitrification rate, and the abundance of AOA (ammonia-oxidizing archaea) and AOB (ammonia-oxidizing bacteria) amoA gene transcripts were determined to evaluate the effects of application of milk vetch for 8 years on soil fertility, nitrification characteristics, and ammonia oxidizers. The two experiments had the same treatments: 1) no fertilizer and milk vetch (CK); 2) milk vetch but no chemical fertilizer (GM); 3) chemical fertilizer but no milk vetch (CF); 4) chemical fertilizer and milk vetch (GM+CF). The results showed that the application of milk vetch improved the pH of both the paddy soils; i.e., the pH of acidic soil in Jiangxi was increased and that of alkaline soil in Hunan was decreased. Application of milk vetch increased the soil organic matter, total nitrogen, ammonium nitrogen, and available phosphorus. The two paddy soils had different nitrification abilities, both the nitrification activity and potential nitrification rate were much higher in purple alluvial soil than in yellow clayey soil. In purple alluvial soil in Hunan, the nitrification activity ranged from 0.269 to 0.325 μg/(g·h) in all the treatments, and there was no significant difference among the treatments. The potential nitrification rate in purple alluvial soil reached 10.25% at the fifth week of incubation, and the GM+CF treatment inhibited nitrification ability to some extent. In yellow clayey soil in Jiangxi, the nitrification activity ranged from 0.010 to 0.021 μg/(g·h) in all the treatments. The potential nitrification rate in yellow clayey soil increased after the third week of incubation and reached 5.41% by the fifth week. Compared with the control, winter green manuring improved the soil nitrification activity and potential nitrification rate, and had almost the same effects as the CF treatment. The GM+CF treatment had the strongest effect to promote the nitrification ability of yellow clayey soil. Transcripts of AOA were predominant in both purple alluvial soil and yellow clayey soil, and the abundance of AOA-amoA transcripts was significantly higher in purple alluvial soil than in yellow clayey soil. The application of milk vetch did not significantly affect the abundance of AOA-amoA and AOB-amoA transcripts in purple alluvial soil, but increased their abundance in yellow clayey soil, similar to the effects of milk vetch to promote nitrification activity and the potential nitrification rate.