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River sediments are regarded as hot spots of bacterial density and activity. Moreover, high bacterial densities and biofilm formation are known to promote horizontal gene transfer, the latter playing a vital role in the spread of antimicrobial resistance. It can thus be hypothesized that sediments act as a reservoir of antibiotic resistant bacteria (ARB) and resistance genes (ARGs), particularly in rivers receiving microbes and drug residues from treated sewage. We analyzed the phenotypic susceptibility of 782 Escherichia coli isolates against 24 antimicrobials and we measured the relative abundances of five ARGs in water and sediment extracts of a small stream. We did not find evidence for a general increase in the proportion of resistant E. coli isolated from sediments as compared to those found in stream water. For most antimicrobials, the likelihood of detecting a resistant isolate was similar in water and sediment or it was even lower in the latter compartment. The mean relative abundance of ARGs was moderately increased in sediment-borne samples. Generally, absolute abundances of resistant cells and resistance genes in the sediment exceeded the pelagic level owing to higher bacterial densities. The river bottom thus represents a reservoir of ARB and ARGs that can be mobilized by resuspension. The river bottom represents a reservoir of antibiotic resistance but a dramatic contamination of stream water due to sediment resuspension is unlikely. [ABSTRACT FROM AUTHOR]

Chaoborus is of great interest to many freshwater ecologists. The adults can become pests in certain areas in North America and the larvae are an important food source for fish. In this preliminary study, we identified variable microsatellite loci in three species: Chaoborus astictopus ( HE = 0.52–0.76) , Chaoborus americanus ( HE = 0.46–0.80) and Chaoborus punctipennis ( HE = 0.66–0.81). Using a biotin/streptavidin capture technique of repetitive sequences in a 96-well format, we obtained microsatellite-enriched genomic libraries for all three species and identified six polymorphic microsatellite markers for each species. None of the primers did yield a polymerase chain reaction fragment in a cross-species test. [ABSTRACT FROM AUTHOR]

Twelve new microsatellite loci were isolated from the Green Lizard ( Lacerta viridis viridis). Primers for 28 loci were designed and 18 of these loci amplified well for 10 individuals of four populations. Twelve of these loci were further characterized for a population in Hungary. The results document the suitability of these identified loci for the characterization of the genetic diversity of the endangered species L. viridis viridis. [ABSTRACT FROM AUTHOR]

With the global concerns on antibiotic resistance (AR) as a public health issue, it is pivotal to have data exchange platforms for studies on antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) in the environment. For this purpose, the NORMAN Association is hosting the NORMAN ARB&ARG database, which was developed within the European project ANSWER. The present article provides an overview on the database functionalities, the extraction and the contribution of data to the database. In this study, AR data from three studies from China and Nepal were extracted and imported into the NORMAN ARB&ARG in addition to the existing AR data from 11 studies (mainly European studies) on the database. This feasibility study demonstrates how the scientific community can share their data on AR to generate an international evidence base to inform AR mitigation strategies. The open and FAIR data are of high potential relevance for regulatory applications, including the development of emission limit values / environmental quality standards in relation to AR. The growth in sharing of data and analytical methods will foster collaboration on risk management of AR worldwide, and facilitate the harmonization in the effort for identification and surveillance of critical hotspots of AR. The NORMAN ARB&ARG database is publicly available at: https://www.norman-network.com/nds/bacteria/., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Decentralised wastewater treatment is becoming a suitable strategy to reduce cost and environmental impact. In this research, the performance of two technologies treating black water (BW) and grey water (GW) fractions of urban sewage is carried out in a decentralised treatment of the wastewater produced in three office buildings. An Anaerobic Membrane Bioreactor (AnMBR) treating BW and a Hybrid preanoxic Membrane Bioreactor (H-MBR) containing small plastic carrier elements, treating GW were operated at pilot scale. Their potential on reducing the release of contaminants of emerging concern (CECs) such as Organic Micropollutants (OMPs), Antibiotic Resistance Genes (ARGs) and pathogens was studied. After 226 d of operation, a stable operation was achieved in both systems: the AnMBR removed 92.4 ± 2.5 % of influent COD, and H-MBR removed 89.7 ± 3.5 %. Regarding OMPs, the profile of compounds differed between BW and GW, being BW the matrix with more compounds detected at higher concentrations (up to μg L -1 ). For example, in the case of ibuprofen the concentrations in BW were 23.63 ± 3.97 μg L -1 , 3 orders of magnitude higher than those detected in GW. The most abundant ARGs were sulfonamide resistant genes (sul1) and integron class 1 (intl1) in both BW and GW. Pathogenic bacteria counts were reduced between 1 and 3 log units in the AnMBR. Bacterial loads in GW were much lower than in BW, being no bacterial re-growth observed for the GW effluents after treatment in the H-MBR. None of the selected enteric viruses was detected in GW treatment line., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Escherichia coli ST58 has recently emerged as a globally disseminated uropathogen that often progresses to sepsis. Unlike most pandemic extra-intestinal pathogenic E. coli (ExPEC), which belong to pathogenic phylogroup B2, ST58 belongs to the environmental/commensal phylogroup B1. Here, we present a pan-genomic analysis of a global collection of 752 ST58 isolates from diverse sources. We identify a large ST58 sub-lineage characterized by near ubiquitous carriage of ColV plasmids, which carry genes encoding virulence factors, and by a distinct accessory genome including genes typical of the Yersiniabactin High Pathogenicity Island. This sub-lineage includes three-quarters of all ExPEC sequences in our study and has a broad host range, although poultry and porcine sources predominate. By contrast, strains isolated from cattle often lack ColV plasmids. Our data indicate that ColV plasmid acquisition contributed to the divergence of the major ST58 sub-lineage, and different sub-lineages inhabit poultry, swine and cattle., (© 2022. The Author(s).)

This study focuses on the impact of infiltration rates on colloidal transport and reactive processes associated with Enterococcus faecalis JH2-2 using water-saturated sediment columns. The infiltration rates influence the physical transport of bacteria by controlling the mean flow velocity. This, in turn, impacts biological processes in pore water owing to the higher or lower residence time of the bacteria in the column. In the present study, continuous injection of E. faecalis (suspended in saline water with varying conditions of dissolved oxygen and nutrient concentrations) into a lab-scale sediment column was performed at flow velocities of 0.02 cm min -1 and 0.078 cm min -1 , i.e., at residence times of 1-5 hours. The impact of residence times on reactive processes is significant for field scale setups. A process-based model with a first-order rate coefficient for each biological process was fitted for each obtained condition-specific dataset from the experimental observations (breakthrough curves). The coefficients were converted to a dimensionless form to facilitate the comparison of biological processes. These results indicate that the processes of attachment and growth were flow-dependent. The growth process in the absence of dissolved oxygen was the most dominant process, with a Damkoehler number of approximately 48.

Ciliates are unicellular eukaryotes with both a germline genome and a somatic genome in the same cytoplasm. The somatic macronucleus (MAC), responsible for gene expression, is not sexually transmitted but develops from a copy of the germline micronucleus (MIC) at each sexual generation. In the MIC genome of Paramecium tetraurelia, genes are interrupted by tens of thousands of unique intervening sequences called internal eliminated sequences (IESs), which have to be precisely excised during the development of the new MAC to restore functional genes. To understand the evolutionary origin of this peculiar genomic architecture, we sequenced the MIC genomes of 9 Paramecium species (from approximately 100 Mb in Paramecium aurelia species to >1.5 Gb in Paramecium caudatum). We detected several waves of IES gains, both in ancestral and in more recent lineages. While the vast majority of IESs are single copy in present-day genomes, we identified several families of mobile IESs, including nonautonomous elements acquired via horizontal transfer, which generated tens to thousands of new copies. These observations provide the first direct evidence that transposable elements can account for the massive proliferation of IESs in Paramecium. The comparison of IESs of different evolutionary ages indicates that, over time, IESs shorten and diverge rapidly in sequence while they acquire features that allow them to be more efficiently excised. We nevertheless identified rare cases of IESs that are under strong purifying selection across the aurelia clade. The cases examined contain or overlap cellular genes that are inactivated by excision during development, suggesting conserved regulatory mechanisms. Similar to the evolution of introns in eukaryotes, the evolution of Paramecium IESs highlights the major role played by selfish genetic elements in shaping the complexity of genome architecture and gene expression., Competing Interests: The authors have declared that no competing interests exist.

The reuse of treated wastewater (e.g. for irrigation) is a common practice to combat water scarcity problems world-wide. However, the potential spread of opportunistic pathogens and fecal contaminants like Enterococci within the subsoil could pose serious health hazards. Additional sources (e.g., leaky sewer systems, livestock farming) aggravate this situation. This study contributes to an understanding of pathogen spread in the environment, using a combined modelling and experimental approach. The impact of quartz sediment and certain wastewater characteristics on the dissemination of Enterococcus faecalis JH2-2 is investigated. The transport processes of advection-dispersion and straining were studied by injecting conservative saline tracer and fluorescent microspheres through sediment packed columns, and evaluating resulting breakthrough curves using models. Similarly, simultaneously occurring reactive processes of microbial attachment, decay, respiration and growth were studied by injecting Enterococcus faecalis JH2-2 suspended in water with or without dissolved oxygen (DO) and nutrients through sediment, and evaluating resulting inlet and outlet concentration curves. The processes of straining, microbial decay and growth, were important when DO was absent. Irreversible attachment was important when DO was present. Sensitivity analysis of each parameter was conducted, and field scale behavior of the processes was predicted, to facilitate future work., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

The occurrence of chemical and biological contaminants of emerging concern (CECs) was investigated in treated wastewater intended for reuse in agriculture. An agarose hydrogel diffusion-based passive sampler was exposed to the outlet of a wastewater treatment plant (WWTP) located in Cyprus, which is equipped with membrane bioreactor (MBR). Passive samplers in triplicate were exposed according to a time-series exposure plan with maximum exposure duration of 28 days. Composite flow-proportional wastewater samples were collected in parallel with the passive sampling exposure plan and were processed by solid phase extraction using HORIZON SPE-DEX 4790 and the same sorbent material (Oasis HLB) as in the passive sampler. The analysis of passive samplers and wastewater samples enabled (i) the field-scale calibration of the passive sampler prototype by the calculation of in situ sampling rates of target substances, and (ii) the investigation of in silico predicted transformation products of the four most ecotoxicologically hazardous antibiotics (azithromycin, clarithromycin, erythromycin, ofloxacin). Additionally, the wastewater samples were subjected to the analysis of seven preselected antibiotic resistant genes (ARGs) and one mobile resistant element (int1). All extracts were analyzed for chemicals in a single batch using a highly sensitive method for pharmaceuticals, antibiotics and illicit drugs by liquid chromatography tandem MS/MS (LC-QQQ) and for various other target compounds (2316 compounds in total) by liquid chromatography high-resolution mass spectrometry (LC-HRMS). 279 CECs and all investigated ARGs (except for bla CTX-M-32 ) were detected, highlighting potential chemical and biological hazards related to wastewater reuse practices. 16 CECs were prioritized following ecotoxicological risk assessment, whereas sul1 and the mobile resistant element (int1) showed the highest abundance. Comprehensive monitoring efforts using novel sampling methods such as passive sampling, wide-scope target screening and molecular analysis are required to assure safe application of wastewater reuse and avoid spread and crop uptake of potentially hazardous chemicals., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier Ltd.)

Next-Generation Sequencing (NGS) technologies are expected to play a crucial role in the surveillance of infectious diseases, with their unprecedented capabilities for the characterisation of genetic information underlying the virulence and antimicrobial resistance (AMR) properties of microorganisms.  In the implementation of any novel technology for regulatory purposes, important considerations such as harmonisation, validation and quality assurance need to be addressed.  NGS technologies pose unique challenges in these regards, in part due to their reliance on bioinformatics for the processing and proper interpretation of the data produced.  Well-designed benchmark resources are thus needed to evaluate, validate and ensure continued quality control over the bioinformatics component of the process.  This concept was explored as part of a workshop on "Next-generation sequencing technologies and antimicrobial resistance" held October 4-5 2017.   Challenges involved in the development of such a benchmark resource, with a specific focus on identifying the molecular determinants of AMR, were identified. For each of the challenges, sets of unsolved questions that will need to be tackled for them to be properly addressed were compiled. These take into consideration the requirement for monitoring of AMR bacteria in humans, animals, food and the environment, which is aligned with the principles of a "One Health" approach., Competing Interests: Competing interests: JOG receives some research funding from Oxford Nanopore Technologies. ER is consultant for Pathoquest.

Integrons are extensively targeted as a proxy for anthropogenic impact in the environment. We developed a novel high-throughput amplicon sequencing pipeline that enables characterization of thousands of integron gene cassette-associated reads, and applied it to acquire a comprehensive overview of gene cassette composition in effluents from wastewater treatment facilities across Europe. Between 38 100 and 172 995 reads per-sample were generated and functionally characterized by screening against nr, SEED, ARDB and β-lactamase databases. Over 75% of the reads were characterized as hypothetical, but thousands were associated with toxin-antitoxin systems, DNA repair, cell membrane function, detoxification and aminoglycoside and β-lactam resistance. Among the reads characterized as β-lactamases, the carbapenemase bla OXA was dominant in most of the effluents, except for Cyprus and Israel where bla GES was also abundant. Quantitative PCR assessment of bla OXA and bla GES genes in the European effluents revealed similar trends to those displayed in the integron amplicon sequencing pipeline described above, corroborating the robustness of this method and suggesting that these integron-associated genes may be excellent targets for source tracking of effluents in downstream environments. Further application of the above analyses revealed several order-of-magnitude reductions in effluent-associated β-lactamase genes in effluent-saturated soils, suggesting marginal persistence in the soil microbiome.

Nanomaterial (NM) release into wastewater treatment plants (WWTPs) is inevitable due to increased production and application throughout past decades and in the future. Concern arose about environmental risks and impact on activated sludge. Environmental risk assessment (ERA) for NMs according to established guidelines is considered not suitable, because NMs exhibit unique characteristics. For hazard identification on activated sludge, standard test organisms for aquatic toxicity testing are not meaningful. In this study, we developed an acute toxicity test for ciliates (Paramecium tetraurelia) as representatives of the important functional group of microbial predators and filter feeders. We chose silver nanoparticles (nAg) exemplarily for ion releasing nanoparticles and regarded toxicity by ions as well. Our results indicate that ions are more toxic (EC₅₀ 0.73 mg/L) than nanoparticles themselves (EC₅₀ 2.15 mg/L). However, nAg must be considered as a source of ions and requires size, surface coating, and compartment-specific ERA. We strived to develop such ERA based on our results, modeled environmental concentration data from literature, and surface area concentrations. Results indicated a probable risk toward activated sludge. This likely has effects on effluent water quality. We conclude that carefully modeled environmental concentrations are vital for more exact ERA for nAg and other NMs.

We investigated the morphology, phylogeny of the 18S rDNA, and pH response of Oxytricha acidotolerans sp. nov. and Urosomoida sp. (Ciliophora, Hypotricha) isolated from two chemically similar acid mining lakes (pH~2.6) located at Langau, Austria, and in Lusatia, Germany. Oxytricha acidotolerans sp. nov. from Langau has 18 frontal-ventral-transverse cirri but a very indistinct kinety 3 fragmentation so that the assignment to Oxytricha is uncertain. The somewhat smaller species from Lusatia has a highly variable cirral pattern and the dorsal kineties arranged in the Urosomoida pattern and is, therefore, preliminary designated as Urosomoida sp. The pH response was measured as ciliate growth rates in laboratory experiments at pH ranging from 2.5 to 7.0. Our hypothesis was that the shape of the pH reaction norm would not differ between these closely related (3% difference in their SSU rDNA) species. Results revealed a broad pH niche for O. acidotolerans, with growth rates peaking at moderately acidic conditions (pH 5.2). Cyst formation was positively and linearly related to pH. Urosomoida sp. was more sensitive to pH and did not survive at circumneutral pH. Accordingly, we reject our hypothesis that similar habitats would harbour ciliate species with virtually identical pH reaction norm., (Copyright © 2012 Elsevier GmbH. All rights reserved.)

For the first time the complete mitochondrial genome was sequenced for a member of Lacertidae. Lacerta viridis viridis was sequenced in order to compare the phylogenetic relationships of this family to other reptilian lineages. Using the long-polymerase chain reaction (long PCR) we characterized a mitochondrial genome, 17,156 bp long showing a typical vertebrate pattern with 13 protein coding genes, 22 transfer RNAs (tRNA), two ribosomal RNAs (rRNA) and one major noncoding region. The noncoding region of L. v. viridis was characterized by a conspicuous 35 bp tandem repeat at its 5' terminus. A phylogenetic study including all currently available squamate mitochondrial sequences demonstrates the position of Lacertidae within a monophyletic squamate group. We obtained a narrow relationship of Lacertidae to Scincidae, Iguanidae, Varanidae, Anguidae, and Cordylidae. Although, the internal relationships within this group yielded only a weak resolution and low bootstrap support, the revealed relationships were more congruent with morphological studies than with recent molecular analyses.