Nitrogen pollution in water is a growing concern. Anthropogenic activities have increased the amount of nitrogen released into watercourses, which harms human health and the environment, and causes serious problems, such as eutrophication. Feammox is a recently discovered biological pathway associated with the nitrogen cycle that has gained scientific interest. This process couples anaerobic ammonium oxidation with iron reduction. This work presents a study on the Feammox mechanism from the enrichment of an activated sludge obtained from a sewage treatment plant. The enrichment was carried out at neutral pH to study the N-2 pathway, that is, the Feammox process with the oxidation of ammonium (NH4+) directly to N-2. In addition, different sources of iron were studied: iron chloride (FeCl3); ferrihydrite; and goethite. The characterization of the sludge showed the genes associated with ammonia monooxygenase, nitrate and nitrite reductases processes, along with relevant microbial species. The enrichment, carried out for 42 days and monitored every 14 days, showed that FeCl3 as a source of Fe was more effective for the coupled process of oxidation of NH4+ and the reduction of Fe(III) to Fe(II). At the end of the enrichment period, a removal of 31% and 32.2% of NH4+, and an increase in Fe(II) concentration by 52.4 and 63.9 times regarding the initial value were achieved in aerobic and anaerobic sludge, respectively. This study provides information on the potential of Feammox in the removal of N from wastewater, and the oxidation/reduction yields in the initial enrichment phase.