Exploration de la diversité microbienne dans les procédés d'épuration des eaux à boues activées

CHATENAY M.-PARIS 11-BU Pharma. (920192101) / SudocSudocFranceF

Molecular analysis of methanogen populations and their interactions within anaerobic sludge digesters

International audienceKnowledge of archaeal population structure, function and interactions is of great interest for a deeper understanding of the anaerobic digestion step in wastewater treatment process, that represents a bottle neck in the optimization of digesters performance. Although culture-independent techniques have enabled the exploration of archaeal population in such systems, their population dynamics and interactions still require further investigation. In the present study, 2646 almost full archaeal 16S rRNA gene sequences retrieved from 22 anaerobic digesters located worldwide were analyzed and classified into 83 Operational Taxonomic Units (OTUs) for Euryarchaeotes and 2 OTUs for Crenarchaeotes. Among the Euryarchaeotes, Methanosarcinales represent the predominant archaeal population (47.5% of total sequences), followed by the ARC I (WSA2) lineage (25.3%), Methanomicrobiales (19.9%) and Methanobacteriales (1.9%). Theses lineages are predominant in nine, five, two and one digesters respectively. However, the remaining 5 digesters show no predominance of any methanogenic group. According to the predominance of theses lineages, 5 digester profiles were distinguished. This study revealed a clear interaction between the 4 methanogenic lineages. A core of 12 OTUs represented by five, four, two and one OTU for Methanosarcinales, Methanomicrobiales, ARC I and Methanobacteriales respectively were quantitatively abundant in at least 50% of the analyzed digesters. 16S rRNA targeted hybridization oligonucleotide probes targeting the predominant OTUs are being developed to follow their population dynamics under various parameters

Characterization of functional communities and metabolic pathways during methanogenesis in wastewater sludge by isotopic and molecular analysis

International audienceDescription of the subject. Investigation of the relationships between methanogenic community and metabolic pathway dynamics during methanogenesis. Objectives. The objective was to use an isotopic approach coupled with molecular analysis to identify metabolisms and to understand the dynamics of Archaea under different temperature conditions. Method. Sludge was incubated anaerobically under mesophilic and thermophilic conditions. Biogas production was monitored, together with stable isotopic signatures of produced CH4 and CO2. Results. Isotopic signature values for CH4 indicated a change in methanogenic metabolism with time and temperature. CH4 was predominantly produced from H2/CO2 and acetoclastic metabolism at the beginning of the mesophilic incubations and after acetate injection. A progressive shift towards an acetoclastic metabolism was observed at the end of mesophilic incubations. This period was associated with stability within archaeal communities as monitored by automated ribosomal intergenic spacer analysis (ARISA) and FISH with oligonucleotidic probes targeting specifically the Archaea 16S rRNA gene. At the beginning of the thermophilic incubations and after acetate injection, methane was generated mostly from H2/CO2. At the end of the thermophilic incubations, an acetoclastic metabolism was observed and intergenic spacer analysis (ARISA) and FISH showed important shifts in archaeal communities. Conclusions. Isotopic methods coupled with molecular analyses enabled us to better understand methanogenesis and the involvement of archaeal populations, which varied according to temperature. We also observed a preadaptation of archaeal communities in thermophilic conditions, which could be due to the fact that mesophilic microorganisms were progressively eliminated during the thermophilic incubation period.Description du sujet. Cet article traite des relations entre les communautés des archées méthanogènes et les voies métaboliques lors de l’étape de la méthanogenèse. Objectifs. L’objectif est d’utiliser une approche isotopique couplée à des analyses moléculaires afin d'identifier les métabolismes et de comprendre la dynamique des populations d’Archaea sous différentes conditions de températures. Méthodes. Des boues ont été incubées en anaérobiose dans des conditions mésophiles et thermophiles. La production de biogaz et les mesures isotopiques stables de CH4 et CO2 produits ont été analysées. Résultats. Les valeurs isotopiques du CH4 ont exhibé un changement des voies métaboliques de la méthanogenèse en fonction du temps et des températures d’incubation. Au début des incubations mésophiles et après injection de l'acétate, le CH4 a été principalement produit à partir de H2/CO2 et par le métabolisme acétoclastique. À la fin des incubations mésophiles, un changement progressif vers un métabolisme acétoclastique strict a été observé. Une stabilité remarquable de la communauté des archées a été observée dans les conditions mésophiles par les analyses ARISA et FISH. Au début des incubations thermophiles et après injection de l'acétate, le CH4 a été produit essentiellement à partir de H2/CO2. À la fin des incubations thermophiles, un métabolisme acétoclastique s’est installé. De plus, les analyses ARISA et FISH ont montré des changements au niveau des communautés des archées actives. Conclusions. Une méthode isotopique couplée à des méthodes moléculaires nous ont permis de mieux comprendre comment se déroule la méthanogenèse et les populations d’archées impliquées en fonction de la température. Nous avons également observé une pré-adaptation des communautés d’archées dans les conditions thermophiles qui pourrait être due au fait que les micro-organismes mésophiles sont progressivement éliminés

Molecular Evidence for Novel Planctomycete Diversity in a Municipal Wastewater Treatment Plant

We examined anoxic and aerobic basins and an anaerobic digestor of a municipal wastewater treatment plant for the presence of novel planctomycete-like diversity. Three 16S rRNA gene libraries were constructed by using a 16S rRNA-targeted universal reverse primer and a forward PCR primer specific for Planctomycetes. Phylogenetic analysis of 234 16S rRNA gene sequences defined 110 operational taxonomic units. The majority of these sequences clustered with the four known genera, Pirellula (32%), Planctomyces (18.4%), Gemmata (3.8%), and Isosphaera (0.4%). More interestingly, 42.3% of the sequences appeared to define two distantly separated monophyletic groups. The first group, represented by 35.5% of the sequences, was related to the Planctomyces group and branched as a monophyletic cluster. It exhibited between 11.9 and 20.3% 16S rRNA gene sequence dissimilarity in comparisons with cultivated planctomycetes. The second group, represented by 6.8% of the sequences, was deeply rooted within the Planctomycetales tree. It was distantly related to the anammox sequences (level of dissimilarity, 20.3 to 24.4%) and was a monophyletic cluster. The retrieved sequences extended the intralineage phylogenetic depth of the Plantomycetales from 23 to 30.6%. The lineages described here may have a broad diversity of undiscovered biochemical and metabolic novelty. We developed a new 16S rRNA-targeted oligonucleotide probe and localized members of one of the phylogenetic groups using the fluorescent in situ hybridization technique. Our results indicate that activated sludge contains very diverse representatives of this group, which grow under aerobic and anoxic conditions and even under anaerobic conditions. The majority of species in this group remain poorly characterized

Microbial Analysis and Efficiency of Biofiltration Packing Systems for Hydrogen Sulfide Removal from Wastewater Off Gas

International audienceHydrogen sulfide (H2S) is commonly detected among waste gases from wastewater treatment plants causing atmospheric pollution, which affects human and environmental health. The aim of this study is to characterize the microbial diversity present within two different H2S biofiltration systems packed with either pozzolan or marble and to assess their efficiency for H2S removal. Few examples were reported about the relationship between the characteristics of the packing material and biofilter performance. Physicochemical parameters such as sulfate concentrations and pH are measured along this study. Sulfate concentrations produced by H2S oxidation were higher using marble biofilter and a higher efficiency of H2S removal was observed. Microbial diversity was characterized using a culture-independent 16S rRNA gene approach. Our results show that the biofilter packed with marble is very acidic (pH <3) and exhibits a higher bacterial diversity with three dominant bacterial families: Xanthomonadaceae, Hydrogenophilaceae, and Spirillaceae. However the biofilter packed with pozzolan is less acidic (pH 5.7–6.8) and shows completely different phylotypes where the plastid 16S rRNA gene of the red algae Cyanidium caldarium and Acidithiobacillus bacterium were retrieved. Such results show that the selection of an appropriate packing material is essential to the odor removal performance of a biofilter system and should allow for the design of better reactor with optimal operating conditions

Eukaryotic molecular diversity at different steps of the wastewater treatment plant process reveals more phylogenetic novel lineages

Wastewater microbiota represents important actors of organic depollution. Nowadays, some species used as bioindicators of the effluent quality are still identified by microscopy. In the present study, we investigated eukaryotic diversity at the different steps of the treatment process of a wastewater treatment plant (aerobic, anaerobic, clarifier basins and anaerobic digester) using the 18S rRNA gene sequencing approach. Of the 1519 analysed sequences, we identified 160 operational taxonomic units. Interestingly, 56.9% of the phylotypes were assigned to novel phylogenetic molecular species since they show \textless97% sequence identity with their nearest affiliated representative within public databases. Peritrichia ciliates were the most predominant group, with Epistylis as the most common genus. Although anaerobic, the digester appears to harbor many unclassified phylotypes of protozoa species. Novel lineages such as LKM11 and LKM118 were widely represented in the digester. Diversity values given by Shannon indexes show that the clarifier is the most diversified. This work will help designing molecular tools that are fast, reliable, and reproducible for monitoring wastewater depollution and studying phylogenetic relationships among the wonderful world of protists within this anthropogenic ecosystem

Microbial community structure associated with the high loading anaerobic codigestion of olive mill and abattoir wastewaters

International audienceThe effect of increasing the organic loading rates (OLRs) on the performance of the anaerobic codigestion of olive mill (OMW) and abattoir wastewaters (AW) was investigated under mesophilic and thermophilic conditions. The structure of the microbial community was also monitored. Increasing OLR to 9 g of chemical oxygen demand (COD) L−1 d−1 affected significantly the biogas yield and microbial diversity at 35 °C. However, at 55 °C digester remained stable until OLR of 12 g of COD L−1 d−1 with higher COD removal (80%) and biogas yield (0.52 L g−1 COD removed). Significant differences in the bacterial communities were detected between mesophilic and thermophilic conditions. The dominant phyla detected in the digester at both phases were the Firmicutes, Actinobacteria, Bacteroidetes, Synergistetes and Spirochaete. However, Verrucomicrobia, Proteobacteria and the candidate division BRC1 were only detected at thermophilic conditions. The Methanobacteriales and the Thermoplasmales were found as a high predominant archaeal member in the anaerobic sludge

Molecular analysis of methanogen populations and their interactions within anaerobic sludge digesters

International audienceKnowledge of archaeal population structure, function and interactions is of great interest for a deeper understanding of the anaerobic digestion step in wastewater treatment process, that represents a bottle neck in the optimization of digesters performance. Although culture-independent techniques have enabled the exploration of archaeal population in such systems, their population dynamics and interactions still require further investigation. In the present study, 2646 almost full archaeal 16S rRNA gene sequences retrieved from 22 anaerobic digesters located worldwide were analyzed and classified into 83 Operational Taxonomic Units (OTUs) for Euryarchaeotes and 2 OTUs for Crenarchaeotes. Among the Euryarchaeotes, Methanosarcinales represent the predominant archaeal population (47.5% of total sequences), followed by the ARC I (WSA2) lineage (25.3%), Methanomicrobiales (19.9%) and Methanobacteriales (1.9%). Theses lineages are predominant in nine, five, two and one digesters respectively. However, the remaining 5 digesters show no predominance of any methanogenic group. According to the predominance of theses lineages, 5 digester profiles were distinguished. This study revealed a clear interaction between the 4 methanogenic lineages. A core of 12 OTUs represented by five, four, two and one OTU for Methanosarcinales, Methanomicrobiales, ARC I and Methanobacteriales respectively were quantitatively abundant in at least 50% of the analyzed digesters. 16S rRNA targeted hybridization oligonucleotide probes targeting the predominant OTUs are being developed to follow their population dynamics under various parameters

“Candidatus Cloacamonas Acidaminovorans”: Genome Sequence Reconstruction Provides a First Glimpse of a New Bacterial Division▿ †

Many microorganisms live in anaerobic environments. Most of these microorganisms have not yet been cultivated. Here, we present, from a metagenomic analysis of an anaerobic digester of a municipal wastewater treatment plant, a reconstruction of the complete genome of a bacterium belonging to the WWE1 candidate division. In silico proteome analysis indicated that this bacterium might derive most of its carbon and energy from the fermentation of amino acids, and hence, it was provisionally classified as “Candidatus Cloacamonas acidaminovorans.” “Candidatus Cloacamonas acidaminovorans” is probably a syntrophic bacterium that is present in many anaerobic digesters. This report highlights how environmental sequence data might provide genomic and functional information about a new bacterial clade whose members are involved in anaerobic digestion