Genomic insights into Candidatus Amarolinea aalborgensis gen. nov., sp. nov., associated with settleability problems in wastewater treatment plants

Settleability of particles in activated sludge systems can be impaired by an overgrowth of filamentous bacteria, a problem known as bulking. These filaments are often members of the phylum Chloroflexi, sometimes reaching abundances in excess of 30% of the biovolume. The uncultured Chloroflexi phylotype, Candidatus Amarolinea, has been observed in high abundances in Danish full-scale activated sludge systems by 16S rRNA gene amplicon surveys, where it has been associated with bulking. In this study, fluorescence in situ hybridization was applied to confirm their high abundance, filamentous morphology, and contribution to the interfloc bridging that characterizes filamentous bulking. Furthermore, genome-centric metagenomics using both Illumina and Oxford Nanopore sequencing was used to obtain a near complete population genome (5.7 Mbp) of the Ca. Amarolinea phylotype, which belongs to the proposed novel family Amarolineaceae within the order Caldilineales of Chloroflexi. Annotation of the genome indicated that the phylotype is capable of aerobic respiration, fermentation, and dissimilatory nitrate reduction to ammonia. The genome sequence also gives a better insight into the phylogenetic and evolutionary relationships of the organism. The name Candidatus Amarolinea aalborgensis is proposed for the species.