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

Abstract

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 (5.7Mbp) 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.

The Phylogeny, Biodiversity, and Ecology of the Chloroflexi in Activated Sludge

“…The phylum ‘ Chloroflexi , embraces an ecologically and physiologically diverse group of bacteria, which have been detected in an increasingly wide range of anaerobic habitats including sediments, hot springs, methanogenic anaerobic sludge digesters where they are highly abundant and play an important fermentative role as well as contributing to sludge granulation (Hug et al, 2013; McIlroy et al, 2016; Sun et al, 2016; Xia et al, 2016; Petriglieri et al, 2018; Bovio et al, 2019), the human oral cavity (Campbell et al, 2014), Anammox reactors (Kindaichi et al, 2012; Cao et al, 2016; Wang et al, 2016) and activated sludge communities (Björnsson et al, 2002; Kragelund et al, 2007a, 2011; Speirs et al, 2009, 2011, 2017; Yamada and Sekiguchi, 2009; Hanada, 2014; McIlroy et al, 2018; Andersen et al, 2019; Nierychlo et al, 2019). This is not the place to explore in detail their still evolving systematics, which have been dealt with elsewhere (Hanada, 2014).…”

Targeted Cell Sorting Combined With Single Cell Genomics Captures Low Abundant Microbial Dark Matter With Higher Sensitivity Than Metagenomics

“…Bioinformatic assembly and binning have been the method of choice to obtain (draft) genomes of not-yet-culturable microorganisms like Chloroflexi ( Tyson et al, 2004 ; McIlroy et al, 2017 ; Andersen et al, 2018 ) from metagenomes (so-called MAGs, metagenome-assembled genomes). There has been a massive influx of MAGs from novel uncultured microorganisms in the recent past ( Hug et al, 2016 ; Parks et al, 2017 ; Tully et al, 2018a ), and even the recovery of genomes from lower abundant species is now possible due to improvements in sequencing depths and bioinformatics algorithms ( Albertsen et al, 2013 ; Vollmers et al, 2017a , b ).…”

Advantages outweigh concerns about using genome sequence as type material for prokaryotic taxonomy

“…Third, the standards to use have been outlined already previously by us (Konstantinidis et al ., ) and others (Bowers et al ., ), and are of similar stringency to those used for isolate genomes; the reader is referred to these publications for further details. Furthermore, long‐read sequencing for routine taxonomic descriptions, even on environmental samples, is coming up soon (e.g., Andersen et al ., ), and is strongly expected to circumvent several of the low‐quality issues reported for MAGs and SAGs in the literature, e.g. provide complete genome of similar quality to the isolate genomes, and/or help to identify and fix genome sequences that may be chimeric.…”