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Filamentous Chloroflexi (green non-sulfur bacteria) are abundant in wastewater treatment processes with biological nutrient removal^c

Advanced Wastewater Management Centre, Department of Microbiology and Parasitology, The University of Queensland, Brisbane 4072, Australia¹
ComBinE group, Advanced Computational Modelling Centre, The University of Queensland, Brisbane 4072, Australia²

Author for correspondence: Linda L. Blackall. Tel: +61 7 3365 4645. Fax: +61 7 3365 4620. e-mail: blackall{at}biosci.uq.edu.au

Most filamentous bacteria in biological nutrient removal (BNR) processes have not been identified beyond their morphotype and simple staining reactions. Furthermore, the majority of sludge filaments observed under the microscope do not hybridize to commonly used phylogenetic probes for well characterized bacterial phyla such as the Proteobacteria, Actinobacteria, Firmicutes and Bacteroidetes. Specific 16S rRNA-targeted oligonucleotide probes were designed for the phylum Chloroflexi (green non-sulfur bacteria) and optimized for use in fluorescence in situ hybridization. Chloroflexi have been implicated in BNR systems by phylogenetic identification of filamentous bacteria isolated by micromanipulation from sludge and culture-independent molecular phylogenetic surveys. The predominant morphotype responding to the probes was filamentous and these filaments were generally abundant in 10 Australian full-scale and two laboratory-scale BNR samples examined. Filamentous bacteria responding to a subdivision 1 Chloroflexi probe were rare in the samples, whereas subdivision 3 Chloroflexi filaments were very common in some sludges. This is in direct contrast to results obtained from molecular phylogenetic surveys of BNR systems where most sludge 16S rDNA clones belong to subdivision 1 and only a few to subdivision 3. It is suggested that filamentous bacteria belonging to the Chloroflexi phylum account for a large fraction of phylogenetically uncharacterized filaments in BNR systems and are likely to be abundant in such systems on a global scale.

Keywords: activated sludge, filamentous bacteria, fluorescence in situ hybridization (FISH), phylogeny, microbial ecology

Abbreviations: BNR, biological nutrient removal; FISH, fluorescent in situ hybridization

^c The EMBL accession numbers for the sequences reported in this paper are X84472 (strain SBR1029 16S rDNA), X84474 (strain SBR1031 16S rDNA), X84498 (strain SBR1064 16S rDNA), X84565 (strain SBR2022 16S rDNA), X84576 (strain SBR2037 16S rDNA) and X84607 (strain SBR2076 16S rDNA).

^a Present address: Department of Biotechnology, Centre of Chemistry and Chemical Engineering, Lund University, PO Box 124, SE-22100 Lund, Sweden.

^b Present address: Department of Environmental Science, Policy and Management, Hilgard Hall, University of California Berkeley, Berkeley CA 94720, USA.

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