The anaerobic ammonium oxidizing bacteria (anammox) are the hidden culprit behind the excessive nitrogen loss under a favorable environment, since their detection and abundance get disturbed by several unknown factors. This study intends to find the gap between actual anammox working capacity under different water conditions and fertilizers in the laboratory. The 15N-isotopic tracer technique was used to measure anammox and denitrification rate, and anammox community structure was analyzed through high-throughput sequencing with cytochrome cd-1 nitrite reductase functional gene (an_nirS gene, initially found in Candidatus Scalindua). The experiment consisted of four treatments, i.e., (I) CK_ Control, (II) UR_Urea, (III) PM_Pig Manure, and (IV) SRF_ Slow release fertilizer, under two water conditions, i.e., (a) Continuous flooding_ CF, (b) Alternate wetting and drying_ AWD. Results showed that anammox under CF decreased over time by -40.24%, and denitrification increased up to 39.25%. However, anammox activity under AWD increased up to 10.62% with the availability of NO2-, and surprisingly accompanied by the reduction in denitrification loss (-31.97%), being the most critical factor. We found that soil nifH and AOB genes were strongly favorable for anammox activity, while we observed the presence of anammox and AOB genes co-existing at the same time in paddy soil. The high-throughput sequencing with an_nirS functional gene showed a much higher diversity of anammox genera ever reported, mostly uncultured and unidentified. We concluded that water management is more prominent than fertilizer for anammox, and the most critical factor is the duration of AWD cycle, because of short term air supply could boost anammox activity and gene abundance, and could reduce denitrifier activity as well as nirK gene abundance.