Drinking water production from anaerobic groundwater is usually achieved by so called conventional techniques such as aeration and sand filtration. The notion conventional implies a long history and general acceptation of the application, but doesn’t necessarily mean a thorough understanding of the processes involved. This is certainly the case for groundwater filtration, with groundwater being the major source for drinking water production in the world. During infiltration and soil passage groundwater may become deeply anaerobic and be loaded with methane, iron, ammonium and manganese that have to be removed during drinking water production. The removal processes for these compounds may be physical-chemical or biological. Since the important PhD-researchers Lerk and Graveland used a chemical approach in the 1960th, the general perception is that only methane and ammonium removal is biological under environmental conditions. Biological iron and manganese removal would be more exceptional as a result of the specific conditions required. The origin of this PhD lies in a persistent nitrification problem in the filters of Oasen Drinking Water Company. When no strong chemical oxidizers are used, like in the Netherlands, nitrification is the only applicable process to fully remove ammonium. It comprises the two-step biological conversion of ammonia via nitrite to nitrate. The first step, the microbial oxidation of ammonia to nitrite, becomes incomplete during the aging of the filter. The relapse typically becomes visible three to six months after the startup of a filter with new filter material. Oasen has only one technique to counteract these nitrification problems, namely subsurface aeration. In that technique, a limited amount of aerated water is periodically injected into the groundwater aquifer, resulting in in situ iron oxidation. The iron colloids that are also formed in the aquifer stimulate the nitrification in the filters, but the working mechanism is unknown. As this “Wonder van Nieuw Lekkerland” is not understood and restricted by licenses, Oasen looks for alternative techniques to maintain a sound nitrification. The general hypothesis for this problem was that the nitrification problem resulted from the interaction with the other removal processes. The problem was studied in full-scale filters and lab-scale setups. Molecular techniques, such as DGGE and clone libraries, were used to identify the major groups of microorganisms present in the groundwater and filters. Ammonia oxidation is performed by Nitrosomonas and archaea, nitrite oxidation by Nitrospira, while Nitrobacter bacteria are not found in drinking water filters. Notable was the presence of the iron-oxidizing Gallionella bacteria in the subsurface aerated groundwater. Another molecular technique, quantitative PCR, was used to quantify Gallionella and ammonia-oxidizing bacteria and archaea in all incoming and outgoing water flows and attached to the filter material. From these numbers balances were made for the filters. The activity of the ammonia-oxidizers was assessed in standardized batch experiments. One of the major findings was that Gallionella grew extensively in a groundwater filter with nitrification problems. In a filter fed with subsurface aerated water, however, Gallionella pumped up with the subsurface aerated water did not continue to grow in the filter. In fact, clone libraries showed, that Gallionella growing in situ deviated from the ones growing in the filter. The growth of Gallionella in well-ventilated trickling filters is remarkable, because these organisms are supposed to be micro-aerophilic. Trickling filters are used for their efficient gas transfer, resulting in effective aeration and stripping of methane and carbon dioxide. The effluent water has a pH of 7.5 to 8 and is almost saturated with oxygen. To verify the growth of Gallionella under these conditions Gallionella bacteria were cultured in continuously operated oxidation and filtration columns (Figure 1). These experiments confirmed the growth of Gallionella under oxygen saturated conditions and at a pH up to 7.7 (Figure 2). The balance approach of ammonia-oxidizing bacteria (AOB) showed that the nitrification problem was not caused by the excessive washout of these microorganisms. In fact, the number of AOB was higher in a filter with nitrification problems, but their activity was much lower. This low cell-specific activity was caused by limitation of the essential nutrient phosphate that could be corrected by addition of phosphate (see Figure 3). So what is the relation between the growth of Gallionella and nitrification problems? While phosphate in groundwater is readily removed for the greater part by co-precipitation with chemically formed iron oxyhydroxides, the biogenic iron precipitates of Gallionella have a higher adsorption capacity for phosphate and further lower the phosphate concentration to limiting levels for the growth of AOB. The outcome of this PhD research provides solutions for the groundwater nitrification problem (such as phosphate dosage and suppression of Gallionella growth) and perspective to further optimize trickling filtration, the most efficient process to remove methane and iron from groundwater.