This study sought to provide a better understanding of the dynamics of microbial-metal sulphide interfacial processes relevant to heap bioleaching. Attachment and subsequent biofilm formation by Metallosphaera hakonensis (M. hakonensis) on the surface of massive chalcopyrite and pyrite samples, as well as a low-grade chalcopyritic whole ore were investigated. The method made use of a biofilm reactor in which thin sections of mineral ore were mounted. Operating conditions in the reactor simulated those of a bioheap in terms of fluid-flow and mineralogy, where the low-grade chalcopyrite ore sections were used. Pure cultures of M. hakonensis were used to inoculate the reactors and the attachment and subsequent biofilm development visualised in situ after 2, 4 and 8 days using a combination of atomic force and epifluorescent microscopy (AFM–EFM) as well as confocal scanning laser microscopy (CSLM). This revealed insights into biofilm structure and architecture. The effect of varying temperature on the extent of attachment and biofilm development was also assessed after 4 days using three temperature regimes: room temperature (20 ± 1 °C), 45 °C and 65 °C. The density of the attached micro–colonies increased with an increase in time, indicative of an actively growing biofilm. The extent of surface coverage and proliferation of the biofilm was dependent on the temperature, with surface coverage being more extensive at 65 °C, near the optimal temperature for growth. Preferential attachment and biofilm formation to sulphide minerals was observed, with increased surface coverage of pyrite mineral surfaces relative to chalcopyrite and low-grade ore. The AFM–EFM technique enhanced the level of detail at which site specific associations of microorganisms with mineral surfaces could be assessed. Spatial orientation and density of attached micro-colonies were noted.