Interfacial interactions between acidophilic bacterial cells and sulphide mineral surfaces play an important role in the metal recovery of bioleaching process. The objective of this study is to develop a reliable bacterial probe technique to quantitatively measure the interaction forces on the nanoscale between various bioleaching microorganisms and mineral surfaces in aqueous solutions.
Atomic force microscopy (AFM) has emerged as a powerful tool to probe the interactions between cell and solid surfaces in liquid environments. The interactions between silica microsphere and silica wafer in salt solutions were measured and modelled with the DLVO theory as a control system. In the past decade, several protocols have been developed to immobilize Gram-positive and Gram-negative bacterial cells to the AFM cantilevers for force measurements. However, for acidophilic bacteria used in bioleaching process, few immobilization approaches were reported for measuring force in acidic and high ionic strength solutions. In this study, we compared different commonly used strategies for constructing reliable bacterial probes for AFM studies on mineral surfaces. The results indicate that compared to other approaches the bacterial colloidal probe technique is a reliable and versatile platform for quantifying true interaction forces between bioleaching bacteria and mineral surfaces, particularly in acidic solutions.
Functionalized colloidal probes covered by bioleaching bacterial cells (Acidithiobacillus thiooxidans and Leptospirillum ferrooxidans) were developed and used to sense specific adhesion forces to a silica surface and a pyrite surface in various solutions. Experimentally, recorded retraction curves of A. thiooxidans revealed sawtooth features that were in good agreement with the wormlike chain model, while that of L. ferrooxidans exhibited stair-step separation. The magnitudes of adhesion forces and snap-off distances were strongly influenced by the ionic strength and pH. Macroscopic surface properties including hydrophobicity and surface potential for bacterial cells and substrates were measured by a sessile drop method and microelectrophoresis. The ATR-FTIR spectra indicated the presence of different types of biopolymers on two strains of bacteria.
Bacteria capable of oxidizing sulphur and iron, known as A. thiooxidans and A. ferrooxidans bacteria, respectively, are important in bioleaching of sulphide minerals. Here, using AFM with bacterial probe technique we report significant differences in the adhesion behaviour of these bacteria when interacting with chalcopyrite surfaces. The bacterial force probes were prepared by attaching bacterial cells onto a silica microsphere (~10 µm radius) glued to the end of an AFM cantilever. Probes were brought into, and separated from, the contact with chalcopyrite surfaces in half-strength 9K medium solutions of various pH conditions. The adhesion forces of nano-Newton resolution versus snap-off distances of nanometre resolutions between the bacterial probes and the mineral surface in aqueous media were obtained during retracting of the probes from the mineral surface. The results show generic different adhesion force curves measured for the two bacteria strains: A. thiooxidans cells exhibit a saw-tooth shaped adhesion force curve, while A. ferrooxidans cells display a stair-step adhesion force curve. Generally, A. ferrooxidans cells show stronger averaged adhesion force with chalcopyrite than A. thiooxidans. Larger adhesion forces of both bacteria were observed at low pH, close to the isoelectric points (~ pH 2) of cells and chalcopyrite. The AFM and FTIR results indicate that the polymers on the bacterial cell surface are critical to the bacterial adhesion onto mineral surfaces. These bacterial probe analyses provide novel insights into the nano-scale mechanism by which bioleaching bacteria interact with mineral surfaces.
The surface appendages and extracellular polymeric substances of cells play an important role in the interaction process. The adhesion and nanomechanical properties of A. ferrooxidans cultured with ferrous sulphate and elemental sulphur were quantitatively investigated under various salt concentrations using the bacterial probe technique with atomic force microscope. Our results show that A. ferrooxidans cells grown with ferrous ion and elemental sulphur exhibit stair-step and sawtooth shape retraction profile, respectively. During the approach of bacterial probes to the substrate surfaces, surface appendages and biopolymers of cells are sequentially compressed. The conformations of surface appendages and biopolymers are significantly affected by the salt concentration of solutions. The bacterial probe technique could be applied to many other investigations combining bacterial species and mineralogical properties in various leaching conditions. It could provide a unique perspective into bacteria-mineral interactions during bioleaching processes.