5-Amino-2,4,6-triiodoisophthalic acid (ATIA), both a precursor and a degradative intermediate of triiodinated contrast media, was anaerobically converted by sludge from a wastewater treatment plant. ATIA conversion took place only when an electron donor such as ethanol was added. A stable mixed culture was established by transfer to a defined synthetic mineral medium with ATIA and ethanol. It could be maintained for 1 year when the sulfate concentration was kept below 30 microM. Transient appearance of 5-amino-2,4-diiodoisophthalic acid, iodide release (2.7 mol iodide/mol ATIA) and accumulation of 5-aminoisophthalic acid indicated that ATIA was reductively dehalogenated. The enriched mixed culture also dehalogenated ATIA derivatives but deiodination remained incomplete. ATIA was the sole terminal electron acceptor used by the mixed culture during deiodination. The ratio of electrons transferred to ATIA, 0.83, was consistent with a respiratory metabolism. Formate, acetate, lactate, butyrate and hydrogen were also used as electron donors. Deiodination was inhibited by a headspace of air or by addition of nitrate, sulfite or thiosulfate. The reaction was 2.6 times slower with sulfate than without.