Nielsen, Per Halkjær

Abstract ‘Candidatus Microthrix parvicella’ is a lipid-accumulating, filamentous bacterium so far found only in activated sludge wastewater treatment plants, where it is a common causative agent of sludge separation problems. Despite attracting considerable interest, its detailed physiology is still unclear. In this study, the genome of the RN1 strain was sequenced and annotated, which facilitated the construction of a theoretical metabolic model based on available in situ and axenic experimental data. This model proposes that under anaerobic conditions, this organism accumulates preferentially long-chain fatty acids as triacylglycerols. Utilisation of trehalose and/or polyphosphate stores or partial oxidation of long-chain fatty acids may supply the energy required for anaerobic lipid uptake and storage. Comparing the genome sequence of this isolate with metagenomes from two full-scale wastewater treatment plants with enhanced biological phosphorus removal reveals high similarity, with few metabolic differences between the axenic and the dominant community ‘Ca. M. parvicella’ strains. Hence, the metabolic model presented in this paper could be considered generally applicable to strains in full-scale treatment systems. The genomic information obtained here will provide the basis for future research into in situ gene expression and regulation. Such information will give substantial insight into the ecophysiology of this unusual and biotechnologically important filamentous bacterium.

Summary Twelve strains of filamentous bacteria morphologically identified as ‘ Microthrix parvicella ’ were isolated from industrial activated sludge wastewater treatment plants. 16S rRNA gene sequences analysis showed that these strains were all closely related to ‘ Candidatus Microthrix parvicella’. Six of them, however, had a 16S rRNA gene similarity of only 95.7% and 96.7% to ‘ Candidatus Microthrix parvicella’ suggesting the presence of a new species. The name ‘ Candidatus Microthrix calida’ is proposed for this new microorganism. The physiological properties of these six isolates supported the description of a new taxon. The ‘ Candidatus Microthrix calida’ strains produced thin filaments (0.3–0.7 µm diameter), they did not grow on the media supporting the growth of ‘ Candidatus Microthrix parvicella’ and could be cultivated at higher temperature (up to 36.5°C). Preliminary data on substrate uptake were obtained by microautoradiography on pure culture. Two new fluorescence in situ hybridization (FISH) probes, Mpa‐T1‐1260 specific for ‘ Candidatus Microthrix calida’ and Mpa‐all‐1410 targeting both Microthrix species, were designed. The presence of Microthrix spp. was investigated in 114 activated sludge plants. ‘ Microthrix parvicella ’ morphotype was detected in 23% of the analysed samples and FISH analysis revealed that ‘ Candidatus Microthrix calida’ was present in 5% of them. The remaining ‘ M. parvicella ’ filaments were positive with probe Mpa‐all‐1410 but could not all be identified as ‘ Candidatus Microthrix parvicella’ suggesting the presence of more hitherto undescribed biodiversity within this morphotype.