The effect of solid loading (mineral pulp density) on thermophilic bioleaching of pyrite by Sulfolobus metallicus (BC) was investigated in a batch reactor. Different mineral pulp densities in the range 3±18% (w/v) were tested. With mineral pulp densities ranging from 3 to 9% the bioleaching proceeded in a single stage with a relatively constant rate. The bioleaching rates calculated for pulp densities of 3, 6 and 9% were 0.10, 0.11 and 0.09 kg iron m À3 h À1 respectively. By contrast the bioleaching of pyrite at pulp densities of 12 and 15% proceeded in two distinct stages. During the exponential phase of microbial growth a sharp and linear increase in concentration of released iron was achieved. This increasing trend levelled off in the presence of non-growing cells and the second stage of bioleaching continued with a slower rate. For the pulp density of 12% the bioleaching rates of the ®rst and the second stages were 0.09 and 0.02 kg iron m À3 h À1 respectively, whereas the calculated rates in the presence of 15% mineral were 0.07 and 0.017 kg iron m À3 h À1 for the ®rst and the second stages. Application of 18% mineral adversely in¯uenced the activity of the cells and the extent of bioleaching in this case was insigni®cant.

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Semi-passive remediation systems have the potential to treat low-volume, sulphaterich, mining impacted waters in a cost-effective and sustainable way. This paper describes the "proof of concept" evaluation of a hybrid linear flow channel reactor, capable of sustaining efficient biological sulphate reduction and partial oxidation of the sulphide product to elemental sulphur. Key elements include the presence of a sulphate-reducing microbial community, immobilised onto carbon fibres and the rapid development of a floating biofilm at the air-liquid interface. The biofilm consists of heterotrophic species and autotrophic sulphide oxidisers. It impedes oxygen mass transfer into the bulk volume and creates a suitable pH-redox microenvironment for partial sulphide oxidation. Demonstration of the concept was successful, with near 20 complete reduction of the sulphate in the feed (1 g/l), effective management of the sulphide generated (95-100% removal) and recovery of a portion of the sulphur by harvesting the elemental-sulphur-rich biofilm. The biofilm re-formed within 24 hours of harvesting, with no decrease in volumetric sulphate reduction rate during this period. Colonisation of the carbon microfibers by sulphate reducing bacteria ensured biomass retention, suggesting the reactor could remain effective at high volumetric flow rates. 2

Semi-passive bioremediation is a promising strategy to mitigate persistent low volume mine-impacted wastewater containing high sulphate concentrations. Building on the proof of concept demonstration of the hybrid linear flow channel reactor (LFCR), capable of simultaneous biological sulphate reduction and partial sulphide oxidation with elemental sulphur recovery, the impact of key operating parameters, such as temperature, on process performance is critical to real-world application. Temperature fluctuates seasonally and across the diurnal cycle, impacting biological sulphate reduction (BSR) and partial sulphide oxidation. The process is reliant on the metabolic activity and synergistic interactions between sulphate-reducing (SRB) and sulphide-oxidising (SOB) microbial communities that develop within discrete oxic and anoxic microenvironments within the hybrid LFCR. In this study, the impact of operating temperature on process performance was evaluated by decreasing temperature with time from 30 to 10°C in each of three laboratory-scaled hybrid LFCR units operating in pseudo-steady state at 1 g/L sulphate. Using lactate as a carbon source, two reactor sizes (2 and 8 L) were considered, while the impact of lactate vs. acetate as carbon source was evaluated in the 2 L reactors. On incremental decrease in temperature from 30 to 10°C, a decrease in volumetric sulphate reduction rate was observed: from 0.144 to 0.059 mmol/L.h in the 2 L lactate-fed reactor; from 0.128 to 0.042 mmol/L.h in the 8 L lactate-fed reactor; and from 0.127 to 0.010 mmol/L.h in the 2 L acetate-fed reactor. Similarly, sulphate conversion efficiency decreased (2 L lactate-fed: 66% to 27%; 8 L lactate-fed: 61% to 20%; 2 L acetate-fed: 61% to 5%). A decrease in temperature below the critical value (15°C) led to considerable loss in metabolic activity and overall BSR performance. Sessile and planktonic microbial communities were represented by bacterial phyla including Proteobacteria, Synergistetes, Bacteroidetes, and Firmicutes. A diverse group of putative SRB (Deltaproteobacteria) and SOB, including Alpha, Beta, Gamma, and Epsilonproteobacteria phylotypes, were prevalent and shifted in relative abundance and community composition in response to decreasing temperature. Specifically, the decrease in the relative abundance of Deltaproteobacteria with decreasing temperature below 15°C corresponded with a loss of BSR performance across all three reactors. This study demonstrated the impact of low temperature on the physiological selection and ecological differentiation of SRB and SOB communities within the hybrid LFCR and its implications for real-world process performance.