The search for halotolerant acidophilic microorganisms to increase the efficiency of bioleaching processes in regions of fresh water scarcity has been ongoing for the past two decades. In this study, three pure cultures (V6, V8 and M8) and four enrichment cultures (14C, L2.21, L4-9 and L6.11) from low pH, high saline environments were characterized for their ability to oxidize soluble iron and inorganic sulfur in the presence of increasing concentrations of chloride ion. The mixed cultures 14C and L2-21 contained predominantly Acidihalobacter and Acidithiobacillus spp., respectively, while L4-9 and L6-11 predominantly contained Perroplasma spp. Cultures V6, V8, 14C and L2-21 were assessed for their ability to oxidize 1% pyrite at 9, 15 and 30 g/L chloride ion. Results showed that pure cultures V6 and V8 and mixed culture 14C were able to oxidize pyrite at chloride ion concentrations of 30 g/L, which is higher than the chloride concentration found in seawater (19 g/L). L2-21 was unable to oxidise pyrite, possibly due to the predominant presence of sulfur oxidizing microorganisms in the mixed culture. This illustrates the potential applicability of the cultures for saline water bioleaching and biooxidation of sulfide ores.