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Planctomycetes of the family Pirellulaceae are commonly addressed as budding aquatic bacteria with a complex lifestyle. Although this family is well represented by cultured and taxonomically characterized isolates, nearly all of them were obtained from brackish or marine habitats. The examples of described freshwater Pirellulaceae planctomycetes arelimited to two species only, Pirellula staley and ‘ Anatilimnocola aggregata’. In this study, we characterized a novel freshwater planctomycete of the genus ‘Anatilimnocola’, strain PX40T, which was isolated from a boreal eutrophic lake. Strain PX40T was represented by budding, unpigmented, ellipsoidal to pear-shaped cells, which often occurred in characteristic flower-like rosettes. Cells were covered by bundles of fimbriae; crateriform-like structures were localized on a reproductive cell pole only. These planctomycetes were obligately aerobic, heterotrophic bacteria that utilized various sugars and some polysaccharides, and were highly sensitive to NaCl. Growth occurred in the pH range 5.0–7.5 (with an optimum at pH 6.5–7.0), and at temperatures between 15 and 30 °C (with an optimum at22–25 °C). The major fatty acids of strain PX40T were C18:1 ω9c, C16:0, and 16:1ω7c; cells also contained a wide variety of hydroxy- and dihydroxy-fatty acids and a C31:9 alkene. The major intact polar lipids were diacylglyceryl-(N,N,N)-trimethylhomoserines. The 16S rRNA gene sequence of strain PX40T displayed 96.6%similarity to that of ‘Anatilimnocola aggregata’ ETA_A8 T. The genome of strain PX40T was 8.93 Mb in size and contained one copy of rRNA operon, 76 tRNA genes and 7092 potential protein-coding genes. The DNA G+C content was 57.8%. The ANI value betweenstrain PX40T and ‘Anatilimnocola aggregata’ ETA_A8 T was 78.3%, suggesting that these planctomycetes representdistinct species. We, therefore, propose a novel species of the genus ‘ Anatilimnocola’, ‘A. floriformis’ sp. nov., with strain PX40T (= KCTC 92369T = VKM B-3621T = UQM 41463T) as the type strain.

Small-magnitude earthquakes induced by gas production in the Groningen field in the Netherlands have prompted the development of seismic risk models that serve both to estimate the impact of these events and to explore the efficacy of different risk mitigation strategies. A core element of the risk modelling is ground-motion prediction models (GMPM) derived from an extensive database of recordings obtained from a dense network of accelerographs installed in the field. For the verification of damage claims, an empirical GMPM for peak ground velocity (PGV) has been developed, which predicts horizontal PGV as a function of local magnitude,ML; hypocentral distance,Rhyp; and the time-averaged shear-wave velocity over the upper 30 m,VS30. For modelling the risk due to potential induced and triggered earthquakes of larger magnitude, a GMPM for response spectral accelerations has been developed from regressions on the outputs from finite-rupture simulations of motions at a deeply buried rock horizon. The GMPM for rock motions is coupled with a zonation map defining frequency-dependent non-linear amplification factors to obtain estimates of surface motions in the region of thick deposits of soft soils. The GMPM for spectral accelerations is formulated within a logic-tree framework to capture the epistemic uncertainty associated with extrapolation from recordings of events ofML ≤ 3.6 to much larger magnitudes.

SUMMARY Rapid onset of subduction tectonics across the western Pacific convergent margins in the early Eocene was followed by a slower phase of margin growth of the proto Tonga-Kermadec subduction system north of Zealandia during a middle Eocene phase to tectonic adjustment. We present new age constraints from International Ocean Discovery Program Expedition 371 borehole data on deformation events in northern Zealandian sediments that document the formation of the convergent margin boundary northwest of New Zealand. The deformation shows a shortening event that lasted up to 20 myr and acted over distances of ∼1000 km inboard of the evolving plate margin, just northwest of New Zealand. Multichannel seismic profiles tied to our new borehole sites show shortening occurred predominantly between 45 Ma and 35 Ma with some deformation related to slope failure continuing into the Oligocene. The termination of shortening is linked to opening of the backarc basins of the southwest Pacific and the migration of the Tonga-Kermadec Trench to the east which may have removed the structural evidence of the Eocene plate margin. Paleogene deformation observed inboard of the evolving proto Tonga-Kermadec subduction system indicates that the lithosphere of northern Zealandia, a region of thin continental crust, was strong enough to act as a stress guide. Compressive stresses that caused intra plate folding and faulting developed behind the initiating subduction system with the finite period of deformation indicating the time frame over which an active convergent margin lay along the northern margin of Zealandia.

The Andes are the world's most biodiverse mountain chain, encompassing a complex array of ecosystems from tropical rainforests to alpine habitats. We provide a synthesis of Andean vascular plant diversity by estimating a list of all species with publicly available records, which we integrate with a phylogenetic dataset of 14 501 Neotropical plant species in 194 clades. We find that (i) the Andean flora comprises at least 28 691 georeferenced species documented to date, (ii) Northern Andean mid-elevation cloud forests are the most species-rich Andean ecosystems, (iii) the Andes are a key source and sink of Neotropical plant diversity, and (iv) the Andes, Amazonia, and other Neotropical biomes have had a considerable amount of biotic interchange through time., Trends in Plant Science, 27 (4), ISSN:1360-1385, ISSN:1878-4372

Robust paleomagnetic results through geological time are one of the keys to understand the drift history of the eastern Qiangtang terrane (EQT). Here, we presented comprehensive petrographic observations and rock magnetic and paleomagnetic analyses of the early Carboniferous Upper Zaduo (ZD) limestone Formation (C1z2) from the Sulucun (SLC) section in the Zaduo area, EQT, to investigate its magnetic originality and geological significance. A total of 12 sites (131 samples) were collected. Photomicrograph observations indicate that the limestone samples were characterized by widespread carbonate veinlets. Electron microprobe and energy dispersive spectrometry analyses confirm that authigenic magnetite formed after pyrite. Rock magnetic analyses reveal the dominant magnetic minerals of pyrite and magnetite, with ‘wasp-waisted’ hysteresis loops and close to the “remagnetization trend” hysteresis parameters. Based on both thermal and alternating field demagnetizations, the characteristic remanent magnetization directions for most samples were isolated: Dg = 6.3°, Ig = 50.1°, kg = 54.9, α95 = 6.2° in-situ, and Ds = 330.2°, Is = 58.9°, ks = 5.9, and α95 = 20.5° after 2-step tilt correction. The κ (α95) value decreases (increases) after tilt-correction, and the ChRM directions failed both the McFadden (1990), Watson and Enkin (1993) fold tests, indicating post-folding magnetizations. The 11 site-mean directions yield a mean in-situ paleopole of 84.4°N, 200.3°E, and A95 = 6.8°, which is coincident with the post ∼53 Myr (especially around 40 Ma) paleopoles of the region. We therefore interpreted that these early Carboniferous limestone samples contain remagnetized magnetizations and that they were obtained after 53 Ma, most likely around 40 Ma, due to the far-field effect of the India–Eurasia collision.

Membrane-spanning lipids are present in a wide variety of archaea but they are rarely in bacteria. Nevertheless, the (hyper)thermophilic members of the order Thermotogales harbor tetraester, tetraether, and mixed ether/ester membrane-spanning lipids mostly composed of core lipids derived from diabolic acids, C30, C32 and C34 dicarboxylic acids with two adjacent mid-chain methyl substituents. Lipid analysis of Thermotoga maritima across growth phases revealed a decrease of the relative abundance of fatty acids together with an increase of diabolic acids with independence of growth temperature. We also identified isomers of C30 and C32 diabolic acids, i.e. dicarboxylic acids with only one methyl group at C-15. Their distribution suggests they are products of the condensation reaction but preferably produced when the length of the acyl chains is not optimal. In comparison with growth at the optimal temperature of 80°C, an increase of glycerol ether-derived lipids was observed at 55°C. Besides, our analysis only detected diabolic acid-containing intact polar lipids with phosphoglycerol (PG) headgroups. Considering these findings, we hypothesize a biosynthetic pathway for the synthesis of membrane-spanning lipids based on PG polar lipid formation, suggesting that the protein catalyzing this process could be a membrane protein. We also identified, by genomic and protein domain analyses, a gene coding for a putative plasmalogen synthase homologue in T. maritima, which is also present in other bacteria producing sn1-alkyl ether lipids but not plasmalogens, suggesting it could be involved in the conversion of the ester to ether bond in the diabolic acids bound in membrane-spanning lipids. Importance Membrane-spanning lipids are unique compounds found in most archaeal membranes, but they are also present in specific bacterial groups like the Thermotogales. The synthesis and physiological role of membrane-spanning lipids in bacteria represent an evolutionary and biochemical open question that points to the differentiation of the membrane lipids composition. Understanding the formation of membrane-spanning lipids is crucial to solving this question and identifying the enzymatic and biochemical mechanism performing this procedure. In the present work, we found changes at the core lipid level, and we propose that the growth phase drives the biosynthesis of these lipids rather than temperature. Our results identified physiological conditions influencing the membrane-spanning lipids biosynthetic process which can further clarify the pathway leading to the biosynthesis of these compounds.

The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fmicb.2022.816605/full#supplementary-material, In acid drainage environments, biosulfidogenesis by sulfate-reducing bacteria (SRB) attenuates the extreme conditions by enabling the precipitation of metals as their sulfides, and the neutralization of acidity through proton consumption. So far, only a handful of moderately acidophilic SRB species have been described, most of which are merely acidotolerant. Here, a novel species within a novel genus of moderately acidophilic SRB is described, Acididesulfobacillus acetoxydans gen. nov. sp. nov. strain INE, able to grow at pH 3.8. Bioreactor studies with strain INE at optimum (5.0) and low (3.9) pH for growth showed that strain INE alkalinized its environment, and that this was more pronounced at lower pH. These studies also showed the capacity of strain INE to completely oxidize organic acids to CO2, which is uncommon among acidophilic SRB. Since organic acids are mainly in their protonated form at low pH, which increases their toxicity, their complete oxidation may be an acid stress resistance mechanism. Comparative proteogenomic and membrane lipid analysis further indicated that the presence of saturated ether-bound lipids in the membrane, and their relative increase at lower pH, was a protection mechanism against acid stress. Interestingly, other canonical acid stress resistance mechanisms, such as a Donnan potential and increased active charge transport, did not appear to be active., This work was financed by ERC grants to AS (project 323009) and JS (project 694569), the research program TTW under project number 14797, which is financed by the Dutch Research Council (NWO) to IS-A, and a Gravitation grant (SIAM 024.002.002) of the Netherlands Ministry of Education, Culture and Science to AS and JS., info:eu-repo/semantics/publishedVersion

Abstract Bacterial membranes are composed of fatty acids (FAs) ester-linked to glycerol-3-phosphate, while archaea possess membranes made of isoprenoid chains ether-linked to glycerol-1-phosphate. Many archaeal species organize their membrane as a monolayer of membrane-spanning lipids (MSLs). Exceptions to this ‘lipid divide’ are the production by some bacterial species of (ether-bound) MSLs, formed by tail-tail condensation of fatty acids resulting in the formation of (iso) diabolic acids (DAs), which are the likely precursors of paleoclimatological relevant branched glycerol dialkyl glycerol tetraether molecules. However, the enzymes responsible for their production are unknown. Here, we report the discovery of bacterial enzymes responsible for the condensation reaction of fatty acids and for ether bond formation, and confirm that the building blocks of iso-DA are branched iso-FAs. Phylogenomic analyses of the key biosynthetic genes reveal a much wider diversity of potential MSL (ether)-producing bacteria than previously thought, with significant implications for our understanding of the evolution of lipid membranes. # Description of contents of this directory ## 1_Phylogenies.tar.gz Contains all files generated for the phylogenies of the membrane spanning lipid (MSL) synthase and the glycerol ester reductase (Ger, aka ether bond synthase here). Each foldercontains: DIAMOND output (2 files): Confirmed MSL synthase of T. ethanolicus and glycerol ester reductase (Ger) of T. maritima MSB8) were queried with DIAMOND v2.0.6 against the NCBI non-redundant protein sequence database (nr)downloaded on 7 January 2021. Hits with an e-value =< 1e-50 and query coverage of >= 30% were selected cd-hitoutput(2 files): Protein sequences from the previous step were clustered with cd-hit v4.8.1 using a sequence identity threshold of 0.7 Clustal_Omega output(1 file): Alignment of the representative sequences obtained from the previous step trimAloutput (1 file): Alignment of the previous step with the gaps removed withtrimAl v1.4.rev15 in -gappyout mode IQ-TREE output(9 files):Phylogenetic trees were constructed with IQ-TREE v2.1.2 with 1,000 ultrafast bootstraps. Model selectionwas based on nuclear models and the best-fit model was chosen according to BIC (LG+R10 for both genes). Maximum-likelihood trees were visualized in iTOL and are available as PDF in the Supplementary Information of the paper aligned sequences aligned and trimmed sequences output from IQ-TREE PDFs for each individual tree ## 2_Supplementary Files.rar Contains the SVG files for the Data S1 and Data S2, and the generated pdf files for Data S1 and Data S2. ## 3_GC GC MS.rar Contains the Files generated with the lipid analyses, including: • GC Files for the Supplementary Table1 and Figure 1 • GC-irmMS Files for Supplementary Table 2 and Figure 1 • GC Files for the Supplementary Table3 , Figure 1 and Figure S2 • GC-MS Files for samples mentioned along the manuscript and for Figure 2, 3, S7, S8, S10 and S11. A pdf file with a descriptive list of the corresponding GC, GC-MS, GC-irmMS files within the manuscript &nbsp

In the Miocene (23-5 Ma), a large wetland known as the Pebas System characterized western Amazonia. During the Middle Miocene Climatic Optimum (c. 17-15 Ma), this system reached its maximum extent and was episodically connected to the Caribbean Sea, while receiving sediment input from the Andes in the west, and the craton (continental core) in the east. Towards the late Miocene (c. 10 Ma) the wetland transitioned into a fluvial-dominated system. In biogeographic models, the Pebas System is often considered in two contexts: one describing the system as a cradle of speciation for aquatic or semi-aquatic taxa such as reptiles, molluscs and ostracods, and the other describing the system as a barrier for dispersal and gene flow for amphibians and terrestrial taxa such as plants, insects and mammals. Here we highlight a third scenario in which the Pebas System is a permeable biogeographical system. This model is inspired by the geological record of the mid-Miocene wetland, which indicates that sediment deposition was cyclic and controlled by orbital forcing and sea-level change, with environmental conditions repeatedly altered. This dynamic landscape favoured biotic exchange at the interface of (1) aquatic and terrestrial, (2) brackish and freshwater and (3) eutrophic to oligotrophic conditions. In addition, the intermittent connections between western Amazonia and the Caribbean Sea, the Andes and eastern Amazonia favoured two-way migrations. Therefore, biotic exchange and adaptation was probably the norm, not the exception, in the Pebas System. The myriad of environmental conditions contributed to the Miocene Amazonian wetland system being one of the most species-rich systems in geological history., Botanical Journal of the Linnean Society, 199 (1), ISSN:0024-4074, ISSN:1095-8339

A pure culture of alkaliphilic haloarchaeon strain AArc-ST capable of anaerobic growth by carbohydrate-dependent sulfur respiration was obtained from hypersaline lakes in southwestern Siberia . According to phylogenetic analysis , AArc-ST formed a new genus level branch most related to the genus Natronoarchaeum in the order Halobacteriales . The strain is facultatively anaerobic with strictly respiratory metabolism growing either by anaerobic respiration with elemental sulfur and thiosulfate as the electron acceptors or by aerobic respiration at microoxic conditions. Thiosulfate is reduced partially to sulfide and sulfite. It is a first sulfur-reducing alkaliphilic haloarchaeon utilizing sugars, starch and glycerol as substrates for anaerobic growth. It is extremely halophilic (optimum at 3.5 M total Na+) and obligately alkaliphilic (optimum at pH 9.5). The dominant polar lipids include PG and PGP-Me with the archaeol (C20-C20) or extended archaeol (C20-C25) cores. The dominant respiratory lipoquinone is MK-8:8. On the basis of unique physiological properties and results of phylogenetic analysis, the soda lake isolate is suggested to beclassified into a novel genus and speciesNatranaeroarchaeum sulfidigenes gen. nov., sp. nov. (=JCM 34033 T = UNIQEM U1000T). Furthermore, on the bases of phylogenomic reconstruction, a new family Natronoarchaeaceae fam. nov. isproposed within the order Halobacteriales incorporatingNatranaeroarchaeum and three related genera: Natronoarchaeum, Salinarchaeum and Halostella.

The Aptian Romualdo Formation in the Araripe Basin represents the establishment of an epeiric sea in the Brazilian Northeast during the Early Cretaceous. This unit is composed by a succession of limestones, shales, sandstones, and coquinas, which constitute shell beds near the top. This last interval strongly suggests a marine influence in the whole basin based on the fossil assemblage. However, even though the establishment of marine waters during the deposition of the Romualdo Formation is a consensus, paleogeographical and paleoenvironmental aspects still need to be better understood. This study aims to perform a sedimentological analysis of the coquina interval, based on petrographic and taphonomic analysis of field data and in a new fully recovered well-core (2-AB-1-CE) drilled in the Serra do Maozinha at Abaiara county in the Ceara State. The described rocks were subdivided into eight facies constituted by bivalves, gastropods, ostracods, peloids, micrite, siliciclastic grains, punctual foraminifers, and plant debris. Based on facies' vertical stacking at well 2-AB-1-CE, a main regressive pattern was observed. It was also possible to subdivide the coquina interval into high-frequency transgressive–regressive cycles. The regional distribution of the facies permitted the identification of a shallow epeiric sea throughout the basin, with variations in the paleoenvironment energy and siliciclastic input. These variations were mainly controlled by paleotopography, fluvial discharge, and sea-level oscillations. The data allowed the identification of two subsequent paleogeographical configurations controlled by sea-level change. The first one, related to a sea-level high, was characterized by a protected bay on the western border and a gulf in the rest of the basin. Then a sea-level fall led to the formation of closed and open lagoons associated with higher terrigenous supply. As such, this study adds new data for a better understanding of the Araripe Basin and the impacts of sea-level fluctuations in South America during the Aptian.

The interpretation of local redox indicators in marine sediments relies on their preservation. By their nature, such biogeochemical tracers are susceptible to diagenetic alteration, particularly through post-depositional re-oxidation of the sediment. This can result in the mobilization and loss of distinctive redox dependant signatures, leading to the classical problem that the presence of oxygenated conditions cannot be robustly inferred by the absence of evidence for anoxia. Authigenic uranium enrichments, and their isotope signatures (δ238U), are widely used to infer bottom-water and pore-water redox conditions but are susceptible to later oxidative diagenetic disturbance. Here we explore the preservation of authigenic δ238U signatures in sediment samples from Sapropel S1 at Site 64PE406-E1 in the Eastern Mediterranean Sea (~1760 m water depth), which was originally deposited under reducing conditions but severely affected by later oxidative diagenesis and the variable loss of authigenic U(IV) from the upper sapropel. To this end, we compare U isotope signatures from bulk measurements (δ238Ubulk) with detrital corrected authigenic U (δ238Uauth) and carbonate associated U (δ238UCAU). In contrast to open ocean carbonates deposited under fully oxic conditions, sapropelic carbonate leachates yield δ238UCAU similar to calculated δ238Uauth, thus recording (predominantly) redox dependant isotope fractionation. There is no evidence for significant isotope fractionation resulting from post-depositional oxidative diagenesis, but associated authigenic U(IV) removal results in larger relative detrital contributions to δ238Ubulk, and hence large uncertainties on δ238Uauth estimates. By contrast, δ238UCAU successfully avoids detrital phases, and the uncertainty associated with detrital corrections, providing a primary record of changes in local redox conditions. Thus, we propose that δ238UCAU can be used for accurate reconstructions of benthic de-oxygenation in sediments with low U enrichments, including sediments with post-depositional U(IV) loss and less reducing environments., Geochimica et Cosmochimica Acta, 311, ISSN:0016-7037, ISSN:1872-9533

The Permian magmatic rocks from Morocco contain crustal xenoliths that sample the Variscan crust in a context of widespread magmatism. A series of such xenoliths was collected in Permian dykes of the Central Massif, in the region of Mrirt. The metapelitic xenoliths are silica poor to intermediate (44 < SiO2

Ordination is the name given to a group of methods used to analyse multiple variables without preceding hypotheses. Over the last few decades, the use of these methods in Earth science in general, and notably in analyses of sedimentary sources, has dramatically increased. However, with limited resources oriented towards Earth scientists on the topic, the application of ordination analysis is at times suboptimal and misuse by authors can occur. This text was written for researchers with little to no experience with ordination with the aim of exposing them to the utility and the pitfalls of this branch of exploratory statistics. To do so, a detailed review of three ordination methods is offered: principal component analysis, non‐metric multidimensional scaling and detrended correspondence analysis. A survey of 163 publications in Earth science is presented, in which these ordination methods were used together with a summary of how, why and on what type of data ordination was used. With common mistakes outlined and misuses in those publications identified. Notably, issues were found with reproducibility, documentation, data set dimensions and transformations. Based on this survey, a recommended workflow is offered for Earth scientists who wish to apply ordination. Additionally, this article is accompanied by highly annotated R scripts for novice users to use these methods.

Environmental and biotic responses to early Eocene hyperthermal events in the southwest Pacific are critical for global paleoclimate reconstructions during Cenozoic greenhouse intervals, but detailed multidisciplinary studies are generally missing from this time and location. Eocene carbonate sediments were recovered during International Ocean Discovery Program Expedition 371 at Site U1510 on southern Lord Howe Rise in the Tasman Sea. Part of the Early Eocene Climatic Optimum (EECO; 53.26–49.14 Ma) and superimposed hyperthermal events have been identified based on refined calcareous nannofossil biostratigraphic data and carbon stable isotope records on bulk sediment and benthic foraminifera. Four negative carbon isotope excursions (CIEs) associated with negative oxygen isotope excursions are recognized within the EECO. Comparison with a global compilation of sites indicates these CIEs correlate to the K event (Eocene Thermal Maximum 3), and tentatively to the S, T, and U events. Sediments with a high carbonate content throughout the EECO provide an excellent opportunity to examine these CIEs, as carbonate dissolution often impacts correlative records elsewhere. Benthic foraminifera and calcareous nannoplankton taxa indicative of warm waters are most abundant during the K event, the most prominent hyperthermal of the EECO. Eutrophication of surface waters during the K event did not lead to increased trophic conditions at the seafloor, whereas a coupled response is observed during smaller hyperthermals. The biotic turnover sheds new light on the paleoenvironmental consequences of hyperthermal events.

Middle Jurassic sauropod taxa are poorly known, due to a stratigraphic bias of localities yielding body fossils. One such locality is Cerro Cóndor North, Cañadón Asfalto Formation, Patagonia, Argentina, dated to latest Early-Middle Jurassic. From this locality, the holotype of Patagosaurus fariasi Bonaparte 1986 is revised. Th e material consists of the axial skeleton, the pelvic girdle, and the right femur. Patagosaurus is mainly characterised by a combination of features mainly identifi ed on the axial skeleton, including the following: 1) cervical centra with low Elongation Index; 2) high projection of the postzygodiapophyseal lamina; 3) deep anterior pleurocoels that are sometimes compartmentalized in cervicals; 4) high projection of the neural arch and spine in dorsal vertebrae and anterior(most) caudal vertebrae; 5) deep pneumatic foramina in posterior dorsals which connect into an internal pneumatic chamber; and 6) anterior caudal vertebrae with ‘saddle’ shaped neural spines. Diagnostic features on the appendicular skeleton include: 1) a transversely wide and anteroposteriorly short femur; 2) a medial placement of the fourth trochanter on the femur; and 3) an anteroposteriorly elongated ilium with a rounded dorsal rim, with hook-shaped anterior lobe. Th e characters that are diagnostic for Patagosaurus are discussed, and the osteology of Patagosaurus is compared to that of Early and Middle Jurassic (eu)sauropods from both Laurasia and Gondwana.

Carbapenemase-producing Enterobacterales (CPE) pose a threat to public health necessitating restriction of further spread. Tools to efficiently monitor the prevalence of these still relatively rare resistant bacteria and insight in routes of dissemination are pivotal for the development of prevention strategies. By analysis of untreated and treated wastewater from 100 municipal wastewater treatment plants (WWTPs) together serving over 40% of the Dutch population, this study investigated both the distribution of CPE in the Dutch population, and WWTPs as a source of CPE in the aquatic environment. CPE were detected at 89% of the WWTPs, in 87 influents and 53 effluents. Overall, 15 different CPE-types were detected based on species and carbapenemase gene. The most widely distributed were E. coli carrying blaOXA-48-like genes, which were detected at 88 WWTPs including small WWTPs without connected health care institutions (HCI). BlaOXA-48-like-positive-K. pneumoniae, blaNDM- or blaKPC-positive E. coli, and blaNDM- or blaKPC-positive K. pneumoniae were detected at 33, 20, and 14 WWTPs, respectively. Mean influent and effluent CPE concentrations were 7.9×102 cfu/l and 11 cfu/l. The total daily number of CPE discharged by the 100 WWTPs was estimated to be 2.2×1011 cfu. In multivariate analysis, CPE concentrations in untreated wastewater were positively associated with WWTP size and E. coli concentrations, but not with the presence of HCI. Based on the total number of CPE (9.7×1012 cfu) and ESBL-E. coli (2.41015 cfu) in influents, and a prevalence of approximately 5% in the Dutch population for ESBL-E. coli, the prevalence of CPE in the Dutch population was roughly estimated to be 0.02%. Wastewater surveillance is an efficient tool to monitor the distribution of CPE in the population at a national level and may supplement human surveillance data. CPE are emitted to the aquatic environment with treated wastewater and associated public health risks need to be determined.