Enhanced oxidation of Mn(II) and As(III) by aerobic granular sludge via ferrous citrate: Key roles of colloidal iron and extracellular superoxide radical

Microbial manganese (Mn) oxidation plays a crucial role in shaping the fate of various elements, including arsenic (As). However, this process faces challenges in wastewater environments due to its inherent inefficiency and instability. In our initial research, a serendipitous discovery occurred: the addition of citrate to Fe(II)-containing wastewater stimulated the oxidation of Mn(II) by aerobic granular sludge (AGS). Subsequent experiments in four sequencing batch reactors (SBRs) over a 67-day period confirmed this stimulatory effect. The presence of Fe(II)-citrate led to a remarkable twofold increase in the oxidation of Mn(II) and As(III). The removal efficiency improved from 21±4 % to 87±7 % for Mn(II) and from 77.1 ± 1.8 % to 93.6 ± 0.2 % for As(III). The verification experiments demonstrated that the simultaneous addition of manganese-oxidizing bacteria (MnOB) and Fe(II)-citrate is an effective strategy for enhancing the oxidation and removal of Mn(II) and As(III) by AGS. Through a combination of genomic analysis, cell-free filtrate incubation, and bacterial batch cultivations (including monitoring the time-course changes of 17 substances and 2 free radicals), we elucidated a novel Mn(II) oxidation pathway in Pseudomonas, along with its stimulation method and mechanism. First, bacteria rapidly degrade citrate possibly via the citrate-Mg