ENZYME-LINKED IMMUNOFILTRATION ASSAY TO ESTIMATE ATTACHMENT OF THIOBACILLI TO PYRITE

An enzyme-linked immunofiltration assay (ELIFA) has been developed in order to estimate directly and specifically Thiobacillus ferrooxidans attachment on sulfide minerals. This method derives from the enzyme-li nked immunosorbent assay but is performed on filtration membranes whic h allow the retention of mineral particles for a subsequent immunoenzy matic reaction in microtiter plates. The polyclonal antiserum used in this study was raised against T, ferrooxidans DSM 583 and recognized c ell surface antigens present on bacteria belonging to the genus Thioba cillus. This antiserum and the ELIFA allowed the direct quantification of attached bacteria with high sensitivity (10(4) bacteria were detec ted per well of the microtiter plate). The mean value of bacterial att achment has been estimated to be about 10(5) bacteria mg(-1) of pyrite at a particle size of 56 to 65 mu m. The geometric coverage ratio of pyrite by T, ferrooxidans ranged from 0.25 to 2.25%. This suggests an attachment of T, ferrooxidans on the pyrite surface to well-defined li mited sites with specific electrochemical or surface properties. ELIFA was shown to be compatible with the measurement of variable levels of adhesion. Therefore, this method may be used to establish adhesion is otherms of T, ferrooxidans on various sulfide minerals exhibiting diff erent physicochemical properties in order to understand the mechanisms of bacterial interaction with mineral surfaces.