The growth of Mycobacterium PYR-1 on polycyclic aromatic hydrocarbons (PAHs) was examined in a two-phase partitioning bioreactor (TPPB). TPPBs are characterized by a cell-containing aqueous phase, and an immiscible and biocompatible organic phase that partitions toxic/insoluble substrates to the cells based on their metabolic demand and the thermodynamic equilibrium of the system. Particular emphasis was placed on assessing the nature of the organism's interaction with the aqueous-organic interface. Results showed that agitation rate affected cell growth and PAH degradation rates, while substrate concentration did not, two characteristics of systems exhibiting an interfacial uptake mechanism. Mycobacterium PYR-1 associated exclusively with the aqueous-organic interface and contact angle measurements confirmed the organism's high hydrophobicity. This preferential association with the organic phase was also shown to reduce or eliminate the uptake of a water-soluble substrate (glucose) by the organism from the aqueous phase. Moreover, detailed examination using fluorescence microscopy revealed that, in addition to associating with the aqueous-organic interface, this bacterium exists exclusively on the organic side of the interface. This is the first report of an organism actively growing in the organic phase of a TPPB and may have implications on the future study of hydrophobic organisms in TPPBs and in the natural environment. (c)(c) 2004 Elsevier Ltd. All rights reserved.