title: activated sludge under copper
design: Activated sludge was collected from an aerobic tank of a sewage treatment plant and was cultivated in an (Sequencing Batch Reactor) SBR with a total effective volume of 5 L. The SBR was operated sequentially in a 6-h cycle: influent filling (15 min), aeration (240 min), settling (95 min), and effluent withdrawal (10 min). The hydraulic retention time was 12 h. After an stable status was reached,  the experiment with intermittent Cu gradient loadings was started. The SBR was operated in five continuous periods, each of which included 8 running cycles with Cu loadings and 24 (or 46) subsequent running cycles without Cu loadings (i.e., the recovery cycles). During the five loading periods, 10, 20, 30, 40, and 50 mg/L were added into a storage tank of influent, respectively. However, measured soluble concentrations of Cu in the influent were only 5.5, 9.2, 10.9, 13.5, and 28.2 mg/L, respectively, and they were used as real dosages of Cu into the SBR. An aliquot of 10.0 mL of activated sludge solution was withdrawn during each settling process at cycles 0 (as the control), 8, 32, 40, 64, 72, 96, 104, 128, 136, and 183. S4 was the activated sludge withdrawn at cycle 64 and was prepared for DNA extraction. The hypervariable V4 region (~240 bp) of the bacterial 16S rRNA gene was amplified for Illumina sequencing.