Microbial mats were developed in the Bioremediation Laboratory at CAU and have been used successfully to remediate heavy metals from surface water. The cellular and molecular mechanisms involved in the bioremediation processes are not well understood. The objective of this study was to provide basic understanding of the cellular and molecular basis of heavy metal remediation by mats. The central hypotheses are: (1) that there are metal resistant and/or reducing bacteria in the mats; and (2) that mats produce specific biomolecules that bind to and sequester heavy metals from solution. The specific aims of this investigation were: (1) to isolate, identify, and characterize metal resistant and/or reducing bacteria from the mats, and (2) to isolate, purify, and characterize specific metal-binding biomolecules (bioflocculants) secreted by microbial mats. In this study a facultative, photoorganotrophic, purple, non-sulfur bacterium was isolated from the mixed-species microbial mats that were dominated by cyanobacteria and contained heterotrophic and purple autorophic bacteria. The isolated bacterium was a motile, gram negative rod. Electron micrographs of thin sections of the bacterium showed a lamellar intracytoplasmic membrane (1CM) system. Based on its morphology, nutrient requirements, absorption spectra, GC content, RAPD-PCR fingerprints, and partial sequences of 16 S rDNA, this isolate has been identified as a new strain of Rhodopseudomonas (Rhodopseudomonas mehrabi UME-1). This bacterium was resistant to high concentrations of several heavy metals (50-1000 mg/1) including As, Cd, Cr(III), Co, Cs, Cu(I), Cu(II), Fe, Hg(I), Mn, Pb, Sr, and Zn. The isolate reduced high concentrations (100 mg/1) of Cr(VI) and selenite. An extracellular, acidic polymer with metal binding and flocculating properties was also isolated from the microbial mats. Production of this acidic polysaccharide was highest at the end of the exponential growth phase, and decreased with increased formation of biofilm. The exopolymer has a high molecular weight (500,000 Dalton) and an isoelectric point of 4.2. Its carbohydrate composition was determined by GC/MS and fluorophore-assisted electrophoresis (FACE), and contained Arabinose, rhamnose, fticose, xylose, mannose, glucose, galactose, glucuronic acid, galacturonic acid, N-aetylglucosamine, and N-aetylgalactosamine. Rheological studies showed that bioflocculants produced a highly viscous solution, 1000 fold greater than the viscosity of glycerol at equivalent concentrations. Interaction of bioflocculant with mono- and divalent metals were examined using equilibrium and continuous flow dialysis. Bioflocculant was bound to 200-480 mg of metal per mg bioflocculant. Its strong metal binding and flocculating properties make this exopolysaccharide a good flocculating agent and may find useful commercial applications.
