Five strains of 'Candidatus Nostocoida limicola' were isolated by micromanipulation from two activated sludge plants. Two (Ben17 and Ben18) were from Sunbury, Victoria, Australia, and three (Ben67, Ver1 and Ver2) were from Verona, Italy. The near complete 16S rDNA sequences were determined for five strains and the phylogenetic location of this important bulking filament in the actinomycete subphylum is reported for the first time. Phylogenetically, the Ben strains formed one group with 99.4% 16S rDNA similarity, and the Ver strains formed another with 99.9% 16S rDNA similarity. The mean similarity between the two groups was 97.4%. By 16S rDNA comparison, the closest relative to all strains was Terrabacter sp. strain DPO1361 (95.0-95.5% identical). On R2A medium, all strains generally grew as short filaments or clumps of cocci, whereas on glucose sulfide (GS) medium, all grew as irregular twisting filaments comprising Gram-positive and Gram-negative cells, which is close to their in situ morphology. Polyphosphate was stored either as granules (R2A) or throughout the trichomes (GS). None of the strains could grow without added nitrogen, reduce nitrate to nitrogen gas or grow anaerobically, whereas all could grow at 15-30 degrees C, produce catalase and reduce nitrate to nitrite. All were inactive in the Hugh & Leifson test. This paper describes 'Candidatus Nostocoida limicola'.