Differences in adhesion of A. thiooxidans and A. ferrooxidans on chalcopyrite as revealed by atomic force microscopy with bacterial probes
Introduction
The use of a diverse group of acidophilic microorganisms to perform bioleaching of sulphide minerals has been employed industrially for over half a century. These acidophiles can accelerate the dissolution of sulphide minerals via the oxidation of ferrous ion and/or sulphur compounds. A. thiooxidans and A. ferrooxidans are the most commonly described mesophilic microorganisms in bioleaching systems (Norris, 2007). A. thiooxidans is a sulphur-oxidizer and grows autotrophically with various sulphur compounds, while A. ferrooxidans can utilize both ferrous ions and sulphur compounds. The chalcopyrite (CuFeS2) is the world’s most abundant source of copper. However, the bioleaching of chalcopyrite remains a major challenge due to its refractory nature (Watling, 2006).
Bacterial adhesion is a prerequisite for contact bioleaching and likely to affect the efficiency of bioleaching. Adhesion of bacterial cells onto mineral surfaces is a complex process, which is bacterial strain and mineral selective. Bacterial surface constituents play pivotal roles in establishing firm adhesion between the cell and their energy source, the mineral surfaces (Tuson and Weibel, 2013). Additionally, the pH value of the leaching solution is of great importance in the bioleaching process due to its strong impact on the metabolic activity and surface properties of bacteria and minerals.
Microbial adhesion has been studied by various macroscopic assays. Long-term (Harneit et al., 2006, Mangold et al., 2008, Vilinska and Hanumantha, 2009) and short-term (Tan and Chen, 2012, Wang et al., 2012) adsorption experiments have been carried out to investigate the adsorption behaviour of bioleaching microorganisms. However, these approaches probe large ensembles of bacterial cells and do not provide information on the fundamental forces governing the bacterial adhesion. Although many studies have revealed that the extracellular polymeric substance and surface components of bioleaching bacteria are composed of a mixture of macromolecules, such as proteins, polysaccharides and lipids (Gehrke et al., 1998, Govender and Gericke, 2011), the exact nature of the adhesion is still to be revealed.
Direct quantification of bacterial adhesion force is critical to our understanding of bacteria–mineral interactions. The invention of AFM has allowed the nano-scale exploration of microorganisms. As a powerful imaging tool, AFM has been extensively used to provide three-dimensional topographic views and structural details of microorganisms and biofilms in bioleaching systems (Becker et al., 2011, Florian et al., 2010, Noël et al., 2010). In addition, AFM is being increasingly used to investigate the interaction forces of biological systems. While much progress has been made by using AFM to quantify the interactions from single molecules to whole cells (Beaussart et al., 2013, Francius et al., 2009, Taylor and Lower, 2008, Xu et al., 2013), the nano-scaled interactions between the bioleaching microorganisms and mineral surfaces remain largely unknown.
The motivation of the present work is to unravel the adhesion behaviour of A. thiooxidans and A. ferrooxidans to the chalcopyrite surface under the influence of various pH conditions that ranged from pH 2 to pH 6. A bacterial probe technique was employed to measure the adhesion forces toward the mineral substrate. The magnitude and frequency of adhesion force were used to understand and estimate the bacteria–mineral affinity of interaction. Our results emphasize the differences in the pattern of retraction curves from these bacteria as well as the responses of their adhesion behaviours, i.e. adhesion forces and snap-off distances, to various pH conditions.
Section snippets
Bacterial strains and growth conditions
Bacterial strains used in this study were kindly provided by Professor Guohua Gu (School of Mineral Processing and Bioengineering, Central South University, China). A. ferrooxidans and A. thiooxidans were cultured in 9 K medium at pH 2.0 (Silverman and Lundgren, 1959): (NH4)2SO4 3.0 g/L, KCl 0.1 g/L, K2HPO4⋅3H2O 0.5 g/L, MgSO4⋅7H2O 0.5 g/L, Ca(NO3)2 0.01 g/L. 4.47% (w/v) FeSO4 and 3% (w/v) elemental sulphur were added for A. ferrooxidans and A. thiooxidans as energy source, respectively. All bacteria
Bacterial probes and surface characterization
Adhesion forces between the two strains of bioleaching bacteria and the chalcopyrite surface were investigated using AFM with a bacterial probe. We combine the early work of Razatos et al. (1998) with the use of colloidal probes to construct bacterial probes. Fig. 1(A and C) shows the SEM images of an A. thiooxidans probe and an A. ferrooxidans probe respectively. After completing consecutive force measurements (over 100 curves) in various pH conditions, the contact area was still coated with a
Conclusion
Understanding and controlling bacteria–mineral interactions in the context of bioleaching requires elucidation of the mechanisms by which bacteria adhere to the mineral surfaces. We have shown that bacteria-coated colloidal probe is a reliable approach for quantifying the adhesion forces of bacteria–mineral interactions under various pH conditions, especially in acidic solutions. Collectively, our results demonstrate that A. thiooxidans and A. ferrooxidans cells exhibited two striking
Acknowledgements
Authors gratefully acknowledge the University of Queensland postgraduate scholarship (UQRS). We thank Professor Guohua Gu (School of Mineral Processing and Bioengineering, Central South University, China) for providing the bacterial strains and Dr. Lei Ge (School of Chemical Engineering, University of Queensland) for assistance with FTIR measurements.
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