Dissimilative Iron Reduction by the Marine Eubacterium Alteromonas putrefaciens Strain 200

The ability of microorganisms to mediate the transformation of iron coupled with the participation of iron as a reactive redox species in natural waters and sediments has prompted recent speculation that such bacteria play an important role in such activities as global iron cycling, biologically induced mineral formation, and localized competition for available organic substrates. Microaerophilic soil bacteria dominate the list of microorganisms capable of iron reduction. However, screening experiments have shown that the marine eubacterium Alteromonas putrefaciens strain 200 is one of the most efficient iron reducers yet isolated. Respiratory inhibitor studies have suggested that a branched respiratory chain and two ferri-reductases (one constitutive and one induced by growth under microaerophilic conditions (2 µM O2)) are involved in iron reduction. Further complementary biochemical and genetic studies have concentrated on characterizing the components of the induced iron reduction system. Ethyl methane sulfonate (EMS) has been used to develop mutagenesis procedures for A.putrefaciens 200; under optimal conditions, the frequency of total auxotrophy for EMS-treated cultures is approximately 0.4%. Tests for specific auxotrophic requirements have indicated the presence of a complex distribution of loci required for biosynthetic pathways. Mutants in both high-rate and low-rate ferri-reductase activity have been identified via a newly developed technique for screening EMS-treated A.putrefaciens 200 cultures. Ferri-reductase-deficient mutants generated by these techniques have been shown to contain reduced cytochrome content. Preliminary results indicate that the broad host range cloning vector pVK100 (Inc P) is mobilizable into A.putrefaciens 200. Current work is aimed at construction of an A.putrefaciens 200 clone bank, and complementation of the ferri-reductase mutants.