Chromatin compartmentalization regulates the response to DNA damage.

The DNA damage response is essential to safeguard genome integrity. Although the contribution of chromatin in DNA repair has been investigated1,2, the contribution of chromosome folding to these processes remains unclear3. Here we report that, after the production of double-stranded breaks (DSBs) in mammalian cells, ATM drives the formation of a new chromatin compartment (D compartment) through the clustering of damaged topologically associating domains, decorated with γH2AX and 53BP1. This compartment forms by a mechanism that is consistent with polymer-polymer phase separation rather than liquid-liquid phase separation. The D compartment arises mostly in G1 phase, is independent of cohesin and is enhanced after pharmacological inhibition of DNA-dependent protein kinase (DNA-PK) or R-loop accumulation. Importantly, R-loop-enriched DNA-damage-responsive genes physically localize to the D compartment, and this contributes to their optimal activation, providing a function for DSB clustering in the DNA damage response. However, DSB-induced chromosome reorganization comes at the expense of an increased rate of translocations, also observed in cancer genomes. Overall, we characterize how DSB-induced compartmentalization orchestrates the DNA damage response and highlight the critical impact of chromosome architecture in genomic instability.

Loop extrusion as a mechanism for formation of DNA damage repair foci.

The repair of DNA double-strand breaks (DSBs) is essential for safeguarding genome integrity. When a DSB forms, the PI3K-related ATM kinase rapidly triggers the establishment of megabase-sized, chromatin domains decorated with phosphorylated histone H2AX (γH2AX), which act as seeds for the formation of DNA-damage response foci1. It is unclear how these foci are rapidly assembled to establish a 'repair-prone' environment within the nucleus. Topologically associating domains are a key feature of 3D genome organization that compartmentalize transcription and replication, but little is known about their contribution to DNA repair processes2,3. Here we show that topologically associating domains are functional units of the DNA damage response, and are instrumental for the correct establishment of γH2AX-53BP1 chromatin domains in a manner that involves one-sided cohesin-mediated loop extrusion on both sides of the DSB. We propose a model in which H2AX-containing nucleosomes are rapidly phosphorylated as they actively pass by DSB-anchored cohesin. Our work highlights the importance of chromosome conformation in the maintenance of genome integrity and demonstrates the establishment of a chromatin modification by loop extrusion.

A cohesin/HUSH- and LINC-dependent pathway controls ribosomal DNA double-strand break repair.

The ribosomal DNA (rDNA) represents a particularly unstable locus undergoing frequent breakage. DNA double-strand breaks (DSBs) within rDNA induce both rDNA transcriptional repression and nucleolar segregation, but the link between the two events remains unclear. Here we found that DSBs induced on rDNA trigger transcriptional repression in a cohesin- and HUSH (human silencing hub) complex-dependent manner throughout the cell cycle. In S/G2 cells, transcriptional repression is further followed by extended resection within the interior of the nucleolus, DSB mobilization at the nucleolar periphery within nucleolar caps, and repair by homologous recombination. We showed that nuclear envelope invaginations frequently connect the nucleolus and that rDNA DSB mobilization, but not transcriptional repression, involves the nuclear envelope-associated LINC complex and the actin pathway. Altogether, our data indicate that rDNA break localization at the nucleolar periphery is not a direct consequence of transcriptional repression but rather is an active process that shares features with the mobilization of persistent DSB in active genes and heterochromatin.

Comprehensive Mapping of Histone Modifications at DNA Double-Strand Breaks Deciphers Repair Pathway Chromatin Signatures.

Double-strand breaks (DSBs) are extremely detrimental DNA lesions that can lead to cancer-driving mutations and translocations. Non-homologous end joining (NHEJ) and homologous recombination (HR) represent the two main repair pathways operating in the context of chromatin to ensure genome stability. Despite extensive efforts, our knowledge of DSB-induced chromatin still remains fragmented. Here, we describe the distribution of 20 chromatin features at multiple DSBs spread throughout the human genome using ChIP-seq. We provide the most comprehensive picture of the chromatin landscape set up at DSBs and identify NHEJ- and HR-specific chromatin events. This study revealed the existence of a DSB-induced monoubiquitination-to-acetylation switch on histone H2B lysine 120, likely mediated by the SAGA complex, as well as higher-order signaling at HR-repaired DSBs whereby histone H1 is evicted while ubiquitin and 53BP1 accumulate over the entire γH2AX domains.

Incidence of and risk factors for local complications of peripheral venous catheters in patients older than 70 years: Empirical research quantitative.

AIMS: To assess the incidence density of local complications of peripheral venous catheters in patients aged 70 years and older, to identify risk factors for local complications of peripheral venous catheters, to describe microbiological epidemiology and to assess the impact of complications on patient outcomes. DESIGN: Prospective, observational, single-centre study. METHODS: Patients 70 years and older admitted to the geriatric department of a teaching hospital in France between December 2019 and May 2020 were considered for inclusion if they had a peripheral venous catheter during their stay. Nurses checked the catheter insertion site three times a day for local complications; physicians ensured the follow-up of complications. The STROBE checklist was used in this prospective observational study. RESULTS: A total of 322 patients were included, with 849 peripheral venous catheters; the median age was 88 years and 182 (56.5%) were women. The incidence density of local complications was 50.5/1000 peripheral venous catheter-days. Risk factors for local complications on multivariate analysis were dressing replacement (OR 1.18), furosemide (OR 1.11) and vancomycin (OR 1.60) infusion, urinary continence (OR 1.09) and hematoma at the catheter insertion site (OR 1.15). Thirteen cellulitis and three abscesses were diagnosed. Occurrence of a local complication was associated with a 3-day increased duration of hospital stay (17 vs. 14 days). CONCLUSION: Risk factors for peripheral venous catheter local complications include urinary continence, furosemide or vancomycin infusion, hematoma at the peripheral venous catheter insertion site or dressing replacement. IMPLICATION FOR THE PATIENT CARE: Closer clinical monitoring may help reduce the occurrence of local peripheral venous catheters complication in patients 70 years and older. RELEVANCE TO CLINICAL PRACTICE: Patients at greater risk of peripheral venous catheter local complications deserve closer clinical monitoring or improved preventive measures, which may be beneficial to reduce the length of hospital stay. NO PATIENT OR PUBLIC CONTRIBUTION: The study was designed to describe risk factors for local complications of peripheral venous catheters in order to reinforce surveillance in this specific population by nurses and medical staffs. Patients had their peripheral venous catheter insertion site checked thrice a day by the nurse in charge as part of usual care. They, as service users, caregivers or members of the public, were not solicited for data collection, analysis, interpretation or preparation of the manuscript.

DeepG4: A deep learning approach to predict cell-type specific active G-quadruplex regions.

DNA is a complex molecule carrying the instructions an organism needs to develop, live and reproduce. In 1953, Watson and Crick discovered that DNA is composed of two chains forming a double-helix. Later on, other structures of DNA were discovered and shown to play important roles in the cell, in particular G-quadruplex (G4). Following genome sequencing, several bioinformatic algorithms were developed to map G4s in vitro based on a canonical sequence motif, G-richness and G-skewness or alternatively sequence features including k-mers, and more recently machine/deep learning. Recently, new sequencing techniques were developed to map G4s in vitro (G4-seq) and G4s in vivo (G4 ChIP-seq) at few hundred base resolution. Here, we propose a novel convolutional neural network (DeepG4) to map cell-type specific active G4 regions (e.g. regions within which G4s form both in vitro and in vivo). DeepG4 is very accurate to predict active G4 regions in different cell types. Moreover, DeepG4 identifies key DNA motifs that are predictive of G4 region activity. We found that such motifs do not follow a very flexible sequence pattern as current algorithms seek for. Instead, active G4 regions are determined by numerous specific motifs. Moreover, among those motifs, we identified known transcription factors (TFs) which could play important roles in G4 activity by contributing either directly to G4 structures themselves or indirectly by participating in G4 formation in the vicinity. In addition, we used DeepG4 to predict active G4 regions in a large number of tissues and cancers, thereby providing a comprehensive resource for researchers. Availability: https://github.com/morphos30/DeepG4.

Fate of SARS-CoV-2 coronavirus in wastewater treatment sludge during storage and thermophilic anaerobic digestion.

Since the COVID-19 outbreak has started in late 2019, SARS-CoV-2 has been widely detected in human stools and in urban wastewater. No infectious SARS-CoV-2 particles have been detected in raw wastewater until now, but it has been reported occasionally in human stools. This has raised questions on the fate of SARS-CoV-2 during wastewater treatment and notably in its end-product, wastewater treatment sludge, which is classically valorized by land spreading for agricultural amendment. In the present work, we focused on SARS-CoV-2 stability in wastewater treatment sludge, either during storage (4 °C, room temperature) or thermophilic anaerobic digestion (50 °C). Anaerobic digestion is one of the possible processes for sludge valorization. Experiments were conducted in laboratory pilots; SARS-CoV-2 detection was based on RT-quantitative PCR or RT-digital droplet PCR. In addition to SARS-CoV-2, Bovine Coronavirus (BCoV) particles were used as surrogate virus. The RNA from SARS-CoV-2 particles, inactivated or not, was close to the detection limit but stable in wastewater treatment sludge, over the whole duration of the assays at 4 °C (55 days) and at ambient temperature (∼20 °C, 25 days). By contrast, the RNA levels of BCoV and inactivated SARS-CoV-2 particles decreased rapidly during the thermophilic anaerobic digestion of wastewater treatment sludge lasting for 5 days, with final levels that were close to the detection limit. Although the particles' infectivity was not assessed, these results suggest that thermophilic anaerobic digestion is a suitable process for sludge sanitation, consistent with previous knowledge on other coronaviruses.

A dual role of dLsd1 in oogenesis: regulating developmental genes and repressing transposons.

The histone demethylase LSD1 is a key chromatin regulator that is often deregulated in cancer. Its ortholog, dLsd1 plays a crucial role in Drosophila oogenesis; however, our knowledge of dLsd1 function is insufficient to explain its role in the ovary. Here, we have performed genome-wide analysis of dLsd1 binding in the ovary, and we document that dLsd1 is preferentially associated to the transcription start site of developmental genes. We uncovered an unanticipated interplay between dLsd1 and the GATA transcription factor Serpent and we report an unexpected role for Serpent in oogenesis. Besides, our transcriptomic data show that reducing dLsd1 levels results in ectopic transposable elements (TE) expression correlated with changes in H3K4me2 and H3K9me2 at TE loci. In addition, our results suggest that dLsd1 is required for Piwi dependent TE silencing. Hence, we propose that dLsd1 plays crucial roles in establishing specific gene expression programs and in repressing transposons during oogenesis.

First evidence of SARS-CoV-2 genome detection in zebra mussel (Dreissena polymorpha).

The uses of bivalve molluscs in environmental biomonitoring have recently gained momentum due to their ability to indicate and concentrate human pathogenic microorganisms. In the context of the health crisis caused by the COVID-19 epidemic, the objective of this study was to determine if the SARS-CoV-2 ribonucleic acid genome can be detected in zebra mussels (Dreissena polymorpha) exposed to raw and treated urban wastewaters from two separate plants to support its interest as bioindicator of the SARS-CoV-2 genome contamination in water. The zebra mussels were exposed to treated wastewater through caging at the outlet of two plants located in France, as well as to raw wastewater in controlled conditions. Within their digestive tissues, our results showed that SARS-CoV-2 genome was detected in zebra mussels, whether in raw and treated wastewaters. Moreover, the detection of the SARS-CoV-2 genome in such bivalve molluscans appeared even with low concentrations in raw wastewaters. This is the first detection of the SARS-CoV-2 genome in the tissues of a sentinel species exposed to raw and treated urban wastewaters. Despite the need for development for quantitative approaches, these results support the importance of such invertebrate organisms, especially zebra mussel, for the active surveillance of pathogenic microorganisms and their indicators in environmental waters.

Modelling the benefits of urine source separation scenarios on wastewater treatment plants within an urban water basin.

Stringent discharge regulations are encouraging researchers to create innovative and sustainable wastewater treatment solutions. Urine source separation (USS) is among the potent approaches that may reduce nutrient peak loads in the influent wastewater and improve nutrient recovery. A phenomenological model was used to simulate dynamic influent properties and predict the advantages gained from implementing USS in an urban water basin. Several scenarios were investigated assuming different levels of deployment: at the entire city, or specifically in office buildings for men's urine only, or for both men and women employees. The results confirmed that all scenarios of urine source separation offered benefits at the treatment plant in terms of reducing nitrogen influent load. The economic benefits in terms of reducing energy consumption for nitrification and decreasing methanol addition for denitrification were quantified, and results confirmed environmental advantages gained from different USS scenarios. Despite larger advantages gained from a global USS rate in an entire city, implementation of a specific USS in office buildings would remain more feasible from a logistical perspective. A significant benefit in terms of reducing greenhouse gas emissions is demonstrated and this was especially due to the high level of N2O emissions avoided in nitrifying biological aerated filter.

Monitoring freshwater fish communities in large rivers using environmental DNA metabarcoding and a long-term electrofishing survey.

Monitoring freshwater fish communities in a large human-impacted river is a challenging task. The structure of fish assemblages has been monitored yearly in the Marne and the Seine Rivers, across the Paris conurbation, France, using traditional electrofishing (EF) surveys since 1990, in accordance with the European Water Framework Directive. In addition, metabarcoding of DNA extracted from environmental samples (eDNA) was concomitantly conducted in nine sampling sites in 2017 and in 2018 to compare the estimates of species richness and relative abundance among three methods: annual, long-term EF monitoring and eDNA. The present study confirms better detection of fish species using eDNA compared to annual EF. eDNA metabarcoding was also more efficient for species detection than a 3-6-year EF survey but was similar or less efficient than a long-term EF survey of 14 years of monitoring. In addition, the numbers of reads per species relative to the total number of reads significantly increased with (a) increasing relative abundance (relative percentage of individuals caught per species) and (b) increasing number of years that a fish species was detected during the 2000-2018 period. These results suggest that eDNA could reflect local population persistence.

An environmentally friendly surrogate method for measuring the soluble chemical oxygen demand in wastewater: use of three-dimensional excitation and emission matrix fluorescence spectroscopy in wastewater treatment monitoring.

Gaining rapid knowledge of dissolved organic matter (DOM) proves to be decisive for wastewater treatment plant operators in efforts to achieve good treatment efficiency in light of current legislation. DOM can be monitored by application of fluorescence spectroscopy both online and in real time in order to derive an assessment of DOM oxidation potential. This work presents an eco-friendly alternative method for measuring the soluble chemical oxygen demand (COD) in raw sewage by means of three-dimensional fluorescence spectroscopy. A peak-picking approach has been developed based on a previous parallel factor analysis (PARAFAC) model dedicated to Paris raw sewage. Fluorescence spectroscopy parameters were used to obtain a good prediction model of soluble COD (r2 = 0.799; p < 0.0001; n = 80) for raw sewage. The approach employed in this study serves as a guideline for purposes of implementing online wastewater monitoring and conducting environmentally friendly soluble COD measurements in the laboratory.

Modelling the nitrification in a full-scale tertiary biological aerated filter unit.

A wastewater biofiltration model is used to assess its capacity to reproduce the treatment behaviour of a plant-sized tertiary nitrifying biofilter unit. It is calibrated on two different types of datasets collected at the Seine-Aval biofiltration plant (Achères, France): grab samples at several heights inside the media bed and a long-term daily plant monitoring over a 1-year period. The model parameters are first calibrated to fit the dynamics observed in the media bed, after which the model is compared to the second dataset. Further parameter changes are then made if necessary and the model is once again compared to both datasets to ensure its ability to predict the treatment behaviour on both size scales. The calibrated model provides correct predictions for most observed nutrient variables for both datasets. An overestimation of the oxygen transfer under a summer, low ammonia load period however leads to a slight underestimation of the nitrifying efficiency of the biofilters. Statistical score computation corroborates the model accuracy as the mean error scores usually remain low. They also point to a certain weakness of the model regarding the suspended solids filtration. Both datasets are overall correctly modelled using a single parameter set. Most of this parameter set is close to or contained in value ranges found in the literature. The parameters related to aeration, however, seem to be slightly higher than what is reported elsewhere.

Spatial and temporal characteristics of microbial communities in the Seine river in the greater Paris area under anthropogenic perturbation.

Microorganisms play an important role in maintaining the proper functioning of river ecosystems and are promising candidates for environmental indicators. They are also highly sensitive to environmental changes. It is necessary to have basic knowledge about them in order to know the ecological status of river ecosystem. To our knowglege, there is very little information on the status of microorganisms in surface water of the Seine River, although the Seine River is one of the rivers that suffers the greatest impact from humain activities in the world due to a weak dilution effect. It is therefore necessary to carry out a microbial analysis to assess the ecological status of the Seine River and to use it as a reference to compare with the future state when, for instance, new disinfection technologies of wastewater are implemented. To this end, the microbial communities of the Seine surface water were analyzed, taking into account the spatial effect, including the tributaries, and from upstream to downstream of the Paris conurbation and the temporal aspect, with a monitoring over 4 seasons. The results showed that the microbiome of the water is highly diverse and involved a variety of functions. The main phyla making up the surface water microbiome were Proteobacteria, Actinobacteriota, Firmicutes, Bacteroidota, while other minor phyla were Deinococcota, Patescibacteria, Gemmatimonadota, Cyanobacteria, Bdellovibrionota, Acidobacteriota, Campilobacterota, Myxococcota, and Desulfobacterota. Overall, the microbial community did not change spatially (with the exception of some minor differences between upstream and downstream), but did vary seasonally. The main factors influencing this microbiome were temperature, nitrate and orthophosphate concentrations. The main predicted functions were related to cell metabolism, in particular carbohydrates, amino acids, lipids, energy, vitamins and cofactors, and cell mobility. The microbial compositions showed a strong balance between microbial groups and were involved in the degradation of recalcitrant compounds.

Assessing the effectiveness of performic acid disinfection on effluents: focusing on bacterial abundance and diversity.

Poorly-treated wastewater harbors harmful microorganisms, posing risks to both the environment and public health. To mitigate this, it is essential to implement robust disinfection techniques in wastewater treatment plants. The use of performic acid (PFA) oxidation has emerged as a promising alternative, due to its powerful disinfection properties and minimal environmental footprint. While PFA has been used to inactivate certain microbial indicators, its potential to tackle the entire microbial community in effluents, particularly resistant bacterial strains, remains largely unexplored. The present study evaluates the efficacy of PFA disinfection on the microbial communities of a WWTP effluent, through microbial resistance mechanisms due to their membrane structure. The effluent microbiome was quantified and identified. The results showed that the number of damaged cells increases with CT, reaching a maximum for CT = 240 mg/L•min and plateauing around 60 mg/L•min, highlighting the optimal conditions for PFA-disinfection against microbial viability. A low PFA level with a 10-min contact time significantly affected the microbial composition. It is worth noting the sensitivity of several bacterial genera such as Flavobacterium, Pedobacter, Massilia, Exiguobacterium, and Sphingorhabdus to PFA, while others, Acinetobacter, Leucobacter, Thiothrix, Paracoccus, and Cloacibacterium, showed resistance. The results detail the resistance and sensitivity of bacterial groups to PFA, correlated with their Gram-positive or Gram-negative membrane structure. These results underline PFA effectiveness in reducing microbial levels and remodeling bacterial composition, even with minimal concentrations and short contact times, demonstrating its suitability for widespread application in WWTPs.

Improving monitoring of dissolved organic matter from the wastewater treatment plant to the receiving environment: A new high-frequency in situ fluorescence sensor capable of analyzing 29 pairs of Ex/Em wavelengths.

A high-frequency, in situ fluorescence probe, called Fluocopée®, has been developed in order to better monitor variations in both the quality and quantity of dissolved organic matter within various aquatic environments (e.g. wastewater, receiving environments) thanks to a wide choice of 29 measured Excitation/Emission wavelength pairs. This advance pave the way to new measurement possibilities in comparison with existing probes, which are usually only able to measure 1-4 fluorophores. The qualification tests of the Fluocopée® probe indicate a high level of accuracy for the measurements of tyrosine, tryptophan and humic acids solutions. Good repeatability and reproducibility are also observed. For the first time, this tool has been deployed in an urban watershed (Bougival, Seine River, downstream of Paris) and in the settled effluent from a wastewater treatment plant (Seine aval, Achères, France). This new high-frequency in situ probe offers great application potential, including organic matter quality and quantity monitoring at drinking and wastewater treatment plants (treatment optimization) and in continental and marine waters (the fate of organic matter in biogeochemical cycles).

Microplastic accumulation in sewer sediments and its potential entering the environment via combined sewer overflows: a study case in Paris.

During wet weather events, combined sewer overflows (CSOs) transfer large amount of particulate matter and associated pollutants into surrounding water bodies, thereby deteriorating the recipients' ecological health. Resuspension of sewer sediments during these events contributes significantly to pollution level of these discharges. However, how much this in-sewer process contributes to CSOs' quality regarding microplastic (MP) pollution is little known. Therefore, an investigation on sewer deposits inside the Parisian combined sewer network was carried out. The study found high MP concentrations stored in this matrix, ranging from 5 × 103 to 178 × 103 particle/kg dry weight. Polymer composition is similar to what found in raw wastewater, containing a high proportion of polyethylene and polypropylene. Thus, the results indicated the persistence of MPs in sewer network during transport during dry weather periods to treatment facilities. Once resuspension of sewer deposits happens, MPs can be released into water flow and get discharged along with CSOs. This highlights another potential pathway of MPs into freshwater environment.

Municipal wastewater treatment by biofiltration: comparisons of various treatment layouts. Part 2: assessment of the operating costs in optimal conditions.

This work aims to compare the operation costs (energy, reagents, waste management) for the three layouts usually used in wastewater treatment plants incorporating biofilters, using technical and economical data acquired during 10 years of operation of a Parisian plant (Seine Centre, 240,000 m(3) d(-1) -800,000 equivalent inhabitants). The final objective is to establish general economical data and tendencies that can be translated from our study to any biofiltration plant. Our results evidenced the savings achieved through the treatment process combining upstream and downstream denitrification. To use this layout reduced the operating costs by some 10% as compared with conventional processing only comprising downstream denitrification. Operating costs were respectively estimated at 37 and 34 €/1,000 m(3) for downstream denitrification and combining upstream and downstream denitrification layouts.

Municipal wastewater treatment by biofiltration: comparisons of various treatment layouts. Part 1: assessment of carbon and nitrogen removal.

One of the largest wastewater treatment plants in the Paris conurbation (240,000 m(3)/d) has been studied over several years in order to provide technical and economical information about biological treatment by biofiltration. Biofiltration systems are processes in which carbon and nitrogen pollution of wastewater are treated by ascendant flow through immersed fixed cultures. This paper, focused on technical information, aims: (1) to compare performances of the three biological treatment layouts currently used in biofiltration systems: upstream denitrification (UD), downstream denitrification (DD) and combined upstream-downstream denitrification (U-DD) layouts; and (2) to describe in detail each treatment step. Our study has shown that more than 90% of the carbon and ammoniacal pollution is removed during biological treatment, whatever the layout used. Nitrate, produced during nitrification, is then reduced to atmospheric nitrogen. This reduction is more extensive when the denitrification stage occurs downstream from the treatment (DD layout with methanol addition), whereas it is only partial when it is inserted upstream from the treatment (UD layout - use of endogenous carbonaceous substrate). So, the UD layout leads to a nitrate concentration that exceeds the regulatory threshold in the effluent, and the treatment must be supplemented with a post-denitrification step (U-DD layout). Our work has also shown that the optimal ammonium-loading rate is about 1.1-1.2 kg N-NH(4)(+) per m(3) media (polystyrene) and day. For denitrification, the optimal nitrate-loading rate is about 2.5 kg N per m(3) media (expanded clay) and day in the case of DD with methanol, and is about 0.25 kg N-NO(3)(-) per m(3) media and day in the case of UD with exogenous carbonaceous substrate.