Genome-centric metagenomic insights into the role of Chloroflexi in anammox, activated sludge and methanogenic reactors.

BACKGROUND: The phylum Chloroflexi is highly abundant in a wide variety of wastewater treatment bioreactors. It has been suggested that they play relevant roles in these ecosystems, particularly in degrading carbon compounds and on structuring flocs or granules. Nevertheless, their function is not yet well understood as most species have not been isolated in axenic cultures. Here we used a metagenomic approach to investigate Chloroflexi diversity and their metabolic potential in three environmentally different bioreactors: a methanogenic full-scale reactor, a full-scale activated sludge reactor and a lab scale anammox reactor. RESULTS: Differential coverage binning approach was used to assemble the genomes of 17 new Chloroflexi species, two of which are proposed as new Candidatus genus. In addition, we recovered the first representative genome belonging to the genus 'Ca. Villigracilis'. Even though samples analyzed were collected from bioreactors operating under different environmental conditions, the assembled genomes share several metabolic features: anaerobic metabolism, fermentative pathways and several genes coding for hydrolytic enzymes. Interestingly, genome analysis from the anammox reactor indicated a putative role of Chloroflexi in nitrogen conversion. Genes related to adhesiveness and exopolysaccharides production were also detected. Complementing sequencing analysis, filamentous morphology was detected by Fluorescent in situ hybridization. CONCLUSION: Our results suggest that Chloroflexi participate in organic matter degradation, nitrogen removal and biofilm aggregation, playing different roles according to the environmental conditions.

Ubiquity and Diversity of Cold Adapted Denitrifying Bacteria Isolated From Diverse Antarctic Ecosystems.

Nitrogen cycle has been poorly investigated in Antarctic ecosystems. In particular, how extreme conditions of low temperature, dryness, and high radiation select the microorganisms involved in the cycle is not yet understood. Denitrification is an important step in the nitrogen cycle in which nitrate is reduced stepwise to the gases NO, N2O, and N2. Denitrification is carried out by a wide group of microorganisms spread in the phylogenetic tree. The aim of this work was to isolate and characterize denitrifying bacteria present in different cold environments from Antarctica. Bacterial isolates were obtained from lake, meltwater, sea, glacier ice, ornithogenic soil, and penguin feces samples from King George Island, Fildes peninsula in the Antarctic. Samples were taken during the deicing season in five sampling campaigns. From all the samples we were able to isolate denitrifying strains. A total of 199 bacterial isolates with the capacity to grow in anaerobic mineral media reducing nitrate at 4°C were obtained. The characterization of the isolates by 16S rRNA gene sequence analysis showed a high predominance of the genus Pseudomonas, followed by Janthinobacterium, Flavobacterium, Psychrobacter, and Yersinia. Other minor genera detected were Cryobacterium, Iodobacter, Kaistella, and Carnobacterium. The capacity to denitrify was not previously described for most of the bacteria related to our isolates and in many of them denitrifying genes were not present suggesting the presence of new genes in this extreme environment. Our work demonstrates the ubiquity of denitrification in the Maritime Antarctica and gives important information linking denitrification at cold temperature with taxa in an unequivocal way.

Database Mining to Unravel the Ecology of the Phylum Chloroflexi in Methanogenic Full Scale Bioreactors.

Although microbial communities of anaerobic bioreactors have been extensively studied using DNA-based tools, there are still several knowledge gaps regarding the microbiology of the process, in particular integration of all generated data is still limited. One understudied core phylum within anaerobic bioreactors is the phylum Chloroflexi, despite being one of the most abundant groups in anaerobic reactors. In order to address the abundance, diversity and phylogeny of this group in full-scale methanogenic reactors globally distributed, a compilation of 16S ribosomal RNA gene sequence data from 62 full-scale methanogenic reactors studied worldwide, fed either with wastewater treatment anaerobic reactors (WTARs) or solid-waste treatment anaerobic reactors (STARs), was performed. One of the barriers to overcome was comparing data generated using different primer sets and different sequencing platforms. The sequence analysis revealed that the average abundance of Chloroflexi in WTARs was higher than in STARs. Four genera belonging to the Anaerolineae class dominated both WTARs and STARs but the core populations were different. According to the phylogenetic analysis, most of the sequences formed clusters with no cultured representatives. The Anaerolineae class was more abundant in reactors with granular biomass than in reactors with disperse biomass supporting the hypothesis that Anaerolineae play an important role in granule formation and structure due to their filamentous morphology. Cross-study comparisons can be fruitfully used to understand the complexity of the anaerobic digestion process. However, more efforts are needed to standardize protocols and report metadata information.

Towards centralized biogas plants: Co-digestion of sewage sludge and pig manure maintains process performance and active microbiome diversity.

The aim of this study is to assess the performance of anaerobic digestion against co-digestion systems during the start-up stages based on key process parameters and biological indicators. Two parallel experiments treating sewage sludge alone or co-digested with low concentration of pig manure (8% vol., 2-3% in COD basis) were carried out in two lab-scale CSTR at mesophilic conditions. Same inoculant and organic loading rate sequences were applied for two consecutive runs of 79 and 90 days. According to the removal efficiencies achieved, no significant differences were encountered amongst mono-digestion and co-digestion. This observation was reinforced with the analysis of the total/active microbiome, sequencing 16S rRNA genes and transcripts. The addition of a co-substrate at low concentration had a negligible effect on the total/active microbial communities; they evolved following the same pattern. This might be an advantage in order to upgrade existing wastewater treatment plants to become centralized biogas facilities.