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Large quantities of plastic enter the oceans each year providing extensive attachment surfaces for marine microbes yet understanding their interactions and colonization of plastic debris remains limited. We investigated microbial colonization of various plastic types (polyethylene, polystyrene, polyethylene-terephthalate, and nylon) in ex-situ incubation experiments. Plastic films, both UV-pretreated and untreated, were exposed to seawater from a coastal and an offshore location in the North Sea. 16S rRNA amplicon sequencing was employed to assess microbial community structures after 5, 10, 30, and 45 days of incubation. Our findings show the significant influence of time, seawater origin and plastic type on microbial community succession. We also identified several genera associated with hydrocarbon or plastic degradation potential as well as genera selecting for specific plastics such as Ketobacter and Microbacterium. Our results highlight potential role of microorganisms in plastic biodegradation and support the idea that microbial colonizers on marine plastics debris seemingly select distinct substrate types., 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 Ltd.. All rights reserved.)

Plastic pollution in the marine realm is a severe environmental problem. Nevertheless, plastic may also serve as a potential carbon and energy source for microbes, yet the contribution of marine microbes, especially marine fungi to plastic degradation is not well constrained. We isolated the fungus Parengyodontium album from floating plastic debris in the North Pacific Subtropical Gyre and measured fungal-mediated mineralization rates (conversion to CO 2 ) of polyethylene (PE) by applying stable isotope probing assays with 13 C-PE over 9 days of incubation. When the PE was pretreated with UV light, the biodegradation rate of the initially added PE was 0.044 %/day. Furthermore, we traced the incorporation of PE-derived 13 C carbon into P. album biomass using nanoSIMS and fatty acid analysis. Despite the high mineralization rate of the UV-treated 13 C-PE, incorporation of PE-derived 13 C into fungal cells was minor, and 13 C incorporation was not detectable for the non-treated PE. Together, our results reveal the potential of P. album to degrade PE in the marine environment and to mineralize it to CO 2 . However, the initial photodegradation of PE is crucial for P. album to metabolize the PE-derived carbon., Competing Interests: Declaration of competing interest ME is employed by The Ocean Cleanup, a non-profit organization aimed at advancing scientific understanding and developing solutions to rid the oceans of plastic. The other 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. Published by Elsevier B.V.)

Plastic pollution of the ocean is a top environmental concern. Biodegradable plastics present a potential "solution" in combating the accumulation of plastic pollution, and their production is currently increasing. While these polymers will contribute to the future plastic marine debris budget, very little is known still about the behavior of biodegradable plastics in different natural environments. In this study, we molecularly profiled entire microbial communities on laboratory confirmed biodegradable polybutylene sebacate-co-terephthalate (PBSeT) and polyhydroxybutyrate (PHB) films, and non-biodegradable conventional low-density polyethylene (LDPE) films that were incubated in situ in three different coastal environments in the Mediterranean Sea. Samples from a pelagic, benthic, and eulittoral habitat were taken at five timepoints during an incubation period of 22 months. We assessed the presence of potential biodegrading bacterial and fungal taxa and contrasted them against previously published in situ disintegration data of these polymers. Scanning electron microscopy imaging complemented our molecular data. Putative plastic degraders occurred in all environments, but there was no obvious "core" of shared plastic-specific microbes. While communities varied between polymers, the habitat predominantly selected for the underlying communities. Observed disintegration patterns did not necessarily match community patterns of putative plastic degraders., 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.)

Pigs share anatomical and physiological traits with humans and can serve as a large-animal model for translational medicine. Bona fide porcine pluripotent stem cells (PSCs) could facilitate testing cell and drug therapies. Agriculture and biotechnology may benefit from the ability to produce immune cells for studying animal infectious diseases and to readily edit the porcine genome in stem cells. Isolating porcine PSCs from preimplantation embryos has been intensively attempted over the past decades. We previously reported the derivation of expanded potential stem cells (EPSCs) from preimplantation embryos and by reprogramming somatic cells of multiple mammalian species, including pigs. Porcine EPSCs (pEPSCs) self-renew indefinitely, differentiate into embryonic and extra-embryonic lineages, and permit precision genome editing. Here we present a highly reproducible experimental procedure and data of an optimized and robust porcine EPSC culture system and its use in deriving new pEPSC lines from preimplantation embryos and reprogrammed somatic cells. No particular expertise is required for the protocols, which take ~4-6 weeks to complete. Importantly, we successfully established pEPSC lines from both in vitro fertilized and somatic cell nuclear transfer-derived embryos. These new pEPSC lines proliferated robustly over long-term passaging and were amenable to both simple indels and precision genome editing, with up to 100% targeting efficiency. The pEPSCs differentiated into embryonic cell lineages in vitro and teratomas in vivo, and into porcine trophoblast stem cells in human trophoblast stem cell medium. We show here that pEPSCs have unique epigenetic features, particularly H3K27me3 levels substantially lower than fibroblasts., (© 2024. Springer Nature Limited.)

Background: This article represents the conclusion of the updated German status report on climate change and health, which was jointly written by authors from over 30 national institutions and organisations. The objectives are (a) to synthesise the options for action formulated in the report, (b) to combine them into clusters and guiding principles, (c) to address the success factors for implementation, and (d) to combine the options for action into target parameters., Methods: The options for action from the individual contributions of the status report were systematically recorded and categorised (n=236). Topical clusters were then formed with reference to Essential Public Health Functions, and options for action were assigned to them., Results: Eight topical clusters of options for action and ten guiding principles were identified. These can be summarised in four overarching meta-levels of action: (a) cross-sectorally coordinated structural and behavioural prevention, (b) monitoring, surveillance, and digitalisation (including early warning systems), (c) development of an ecologically sustainable and resilient public health system, and (d) information, communication, and participation. The main success factors for implementation are the design of governance, positive storytelling and risk communication, proactive management of conflicting goals, and a cross-sectoral co-benefit approach., Conclusions: Based on the status report, systematically compiled target parameters and concrete options for action are available for public health., Competing Interests: Conflicts of interest The authors declared no conflicts of interest., (© Robert Koch Institute. All rights reserved unless explicitly granted.)

Ocean plastic pollution is a severe environmental problem but most of the plastic that has been released to the ocean since the 1950s is unaccounted for. Although fungal degradation of marine plastics has been suggested as a potential sink mechanism, unambiguous proof of plastic degradation by marine fungi, or other microbes, is scarce. Here we applied stable isotope tracing assays with 13 C-labeled polyethylene to measure biodegradation rates and to trace the incorporation of plastic-derived carbon into individual cells of the yeast Rhodotorula mucilaginosa, which we isolated from the marine environment. 13 C accumulation in the CO 2 pool during 5-day incubation experiments with R. mucilaginosa and UV-irradiated 13 C-labeled polyethylene as a sole energy and carbon source translated to degradation rates of 3.8% yr -1 of the initially added substrate. Furthermore, nanoSIMS measurements revealed substantial incorporation of polyethylene-derived carbon into fungal biomass. Our results demonstrate the potential of R. mucilaginosa to mineralize and assimilate carbon from plastics and suggest that fungal plastic degradation may be an important sink for polyethylene litter in the marine environment., (© 2023. The Author(s).)

Background: Pig-derived tissues could overcome the shortage of human donor organs in transplantation. However, the glycans with terminal α-Gal and Neu5Gc, which are synthesized by enzymes, encoded by the genes GGTA1 and CMAH, are known to play a major role in immunogenicity of porcine tissue, ultimately leading to xenograft rejection., Methods: The N-glycome and glycosphingolipidome of native and decellularized porcine pericardia from wildtype (WT), GGTA1-KO and GGTA1/CMAH-KO pigs were analyzed by multiplexed capillary gel electrophoresis coupled to laser-induced fluorescence detection., Results: We identified biantennary and core-fucosylated N-glycans terminating with immunogenic α-Gal- and α-Gal-/Neu5Gc-epitopes on pericardium of WT pigs that were absent in GGTA1 and GGTA1/CMAH-KO pigs, respectively. Levels of N-glycans terminating with galactose bound in β(1-4)-linkage to N-acetylglucosamine and their derivatives elongated by Neu5Ac were increased in both KO groups. N-glycans capped with Neu5Gc were increased in GGTA1-KO pigs compared to WT, but were not detected in GGTA1/CMAH-KO pigs. Similarly, the ganglioside Neu5Gc-GM3 was found in WT and GGTA1-KO but not in GGTA1/CMAH-KO pigs. The applied detergent based decellularization efficiently removed GSL glycans., Conclusion: Genetic deletion of GGTA1 or GGTA1/CMAH removes specific epitopes providing a more human-like glycosylation pattern, but at the same time changes distribution and levels of other porcine glycans that are potentially immunogenic., (© 2023 The Authors. Xenotransplantation published by John Wiley & Sons Ltd.)

Global warming of 1.5°C and even 2°C is likely to be exceeded during the 21 st century. Climate change poses a worldwide threat and has direct and indirect effects on infectious diseases, on non-communicable diseases and on mental health. Not all people are equally able to protect themselves against the impacts of climate change; particularly populations that are vulnerable due to individual factors (children, older persons, those immunocompromised or with pre-existing conditions), social factors (the socially disadvantaged), or living and working conditions (e. g. people who work outdoors) are subject to an increased risk. Concepts such as One Health or Planetary Health provide a framework to frame both climate change itself and adaptation strategies or sets of actions for environmental human and animal health. Knowledge of climate change impacts has grown in recent years, and mitigation and adaptation strategies have been developed., Competing Interests: Conflicts of interest The authors declared no conflicts of interest., (© Robert Koch Institute. All rights reserved unless explicitly granted.)

In the 21st century, an accumulation of complex crises such as climate change, biodiversity loss, environmental pollution, wars, and pandemics is leading to economic, social, and health problems for people of current and future generations. These problems are essentially due to the disregard of natural regenerative capacities of ecosystems. Health approaches such as One Health and Planetary Health have gained popularity since the early 2000s and are increasingly used in politics, science, and the health professions. Environmental medicine is also influenced by the growing number of crises and environmental problems and the increasing interest in the concepts of One Health and Planetary Health.In this discussion paper, the development of the two concepts is first presented. Then, their significance for environmental medicine will be discussed. Health professionals working on environmental medicine issues must now consider both local and global causes and health effects when assessing environmental impacts, which can complicate decisions and lead to conflicting objectives., (© 2023. The Author(s).)

We investigated microbial methane oxidation in the water column of two connected but hydrodynamically contrasting basins of Lake Lugano, Switzerland. Both basins accumulate large amounts of methane in the water column below their chemoclines, but methane oxidation efficiently prevents methane from reaching surface waters. Here we show that in the meromictic North Basin water column, a substantial fraction of methane was eliminated through anaerobic methane oxidation (AOM) coupled to nitrite reduction by Candidatus Methylomirabilis. Incubations with 14 CH 4 and concentrated biomass from this basin showed enhanced AOM rates with nitrate (+62%) and nitrite (+43%). In the more dynamic South Basin, however, aerobic methanotrophs prevailed, Ca. Methylomirabilis was absent in the anoxic water column, and no evidence was found for nitrite-dependent AOM. Here, the duration of seasonal stratification and anoxia seems to be too short, relative to the slow growth rate of Ca. Methylomirabilis, to allow for the establishment of anaerobic methanotrophs, in spite of favorable hydrochemical conditions. Using 16 S rRNA gene sequence data covering nearly ten years of community dynamics, we show that Ca. Methylomirabilis was a permanent element of the pelagic methane filter in the North Basin, which proliferated during periods of stable water column conditions and became the dominant methanotroph in the system. Conversely, more dynamic water column conditions led to a decline of Ca. Methylomirabilis and induced blooms of the faster-growing aerobic methanotrophs Methylobacter and Crenothrix. Our data highlight that physical (mixing) processes and ecosystem stability are key drivers controlling the community composition of aerobic and anaerobic methanotrophs., (© 2023. The Author(s).)

Ocean plastic pollution is a problem of increasing magnitude; yet, the amount of plastic at the sea surface is much lower than expected. Solar ultraviolet (UV) radiation can induce photodegradation, but its importance in determining the longevity of floating plastic remains unconstrained. Here, we measured photodegradation rates of different plastic types slightly larger than microplastics (virgin polymers and floating plastic debris) under simulated marine conditions. UV irradiation caused all plastic types to leach dissolved organic carbon, and to a lesser degree carbon dioxide, carbon monoxide, methane, and other hydrocarbon gases. The release of photodegradation products translates to degradation rates of 1.7-2.3 % yr -1 of the tested plastic particles normalized to conditions as found in the subtropical surface ocean. Modelling the accumulation of floating plastic debris, our results show that solar UV radiation could already have degraded 7 to 22 % of all floating plastic that has ever been released to the sea., 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 © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Methods that unambiguously prove microbial plastic degradation and allow for quantification of degradation rates are necessary to constrain the influence of microbial degradation on the marine plastic budget. We developed an assay based on stable isotope tracer techniques to determine microbial plastic mineralization rates in liquid medium on a lab scale. For the experiments, 13 C-labeled polyethylene ( 13 C-PE) particles (irradiated with UV-light to mimic exposure of floating plastic to sunlight) were incubated in liquid medium with Rhodococcus ruber as a model organism for proof of principle. The transfer of 13 C from 13 C-PE into the gaseous and dissolved CO 2 pools translated to microbially mediated mineralization rates of up to 1.2 % yr -1 of the added PE. After incubation, we also found highly 13 C-enriched membrane fatty acids of R. ruber including compounds involved in cellular stress responses. We demonstrated that isotope tracer techniques are a valuable tool to detect and quantify microbial plastic degradation., 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 © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Plastic pollution in the marine environment has been identified as a global problem; different polymer types and fragment sizes have been detected across all marine regions, from sea ice to the equator and the surface to the deep sea. However, quantification of marine plastics debris in the size range of nanoplastics (<1 μm) and ultrafine microplastics (<10 μm) is not constrained, because such minuscule particles are challenging to measure. In this work, we applied a novel analytical assay using Thermal Desorption - Proton Transfer Reaction - Mass Spectrometry (TD-PTR-MS), which is suitable to detect and identify plastics in the nanogram range. From two stations in the Wadden Sea (the Netherlands), we measured nanoplastics directly from seawater aliquots, and from filters with different mesh sizes. Our results show the presence of Polystyrene (PS) and Polyethylene terephthalate (PET) nanopalstics as well as ultrafine microplastics in the Wadden Sea water column. The mass concentration of PS nanoplastics was 4.2 μg/L on average, indicating a substantial contribution of nanoplastics to the Wadden Sea's total plastic budget., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The long-term fate of plastics in the ocean and their interactions with marine microorganisms remain poorly understood. In particular, the role of sinking plastic particles as a transport vector for surface microbes towards the deep sea has not been investigated. Here, we present the first data on the composition of microbial communities on floating and suspended plastic particles recovered from the surface to the bathypelagic water column (0-2000 m water depth) of the North Pacific Subtropical Gyre. Microbial community composition of suspended plastic particles differed from that of plastic particles afloat at the sea surface. However, in both compartments, a diversity of hydrocarbon-degrading bacteria was identified. These findings indicate that microbial community members initially present on floating plastics are quickly replaced by microorganisms acquired from deeper water layers, thus suggesting a limited efficiency of sinking plastic particles to vertically transport microorganisms in the North Pacific Subtropical Gyre., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

At present, the distribution of plastic debris in the ocean water column remains largely unknown. Such information, however, is required to assess the exposure of marine organisms to plastic pollution as well as to calculate the ocean plastic mass balance. Here, we provide water column profiles (0-300 m water depth) of plastic (0.05-5 cm in size) concentration and key planktonic species from the eastern North Atlantic Ocean. The amount of plastic decreases rapidly in the upper few meters, from ~ 1 item/m 3 (~ 1000 µg/m 3 ) at the sea surface to values of ~ 0.001-0.01 items/m 3 (~ 0.1-10 µg/m 3 ) at 300 m depth. Ratios of plastic to plankton varied between ~ 10 -5 and 1 plastic particles per individual with highest ratios typically found in the surface waters. We further observed that pelagic ratios were generally higher in the water column below the subtropical gyre compared to those in more coastal ecosystems. Lastly, we show plastic to (non-gelatinous) plankton ratios could be as high as ~ 10 2 -10 7 plastic particles per individual when considering reported concentrations of small microplastics < 100 μm. Plastic pollution in our oceans may therefore soon exceed estimated safe concentrations for many pelagic species., (© 2022. The Author(s).)

Background: 2018-2020 were unusually warm years in Germany, and the summer of 2018 was the second warmest summer since record-keeping began in 1881. Higher temperatures regularly lead to increased mortality, particularly among the elderly., Methods: We used weekly data on all-cause mortality and mean temperature from the period 1992-2021 and estimated the number of heat-related deaths in all of Germany, and in the northern, central, and southern regions of Germany, employing a generalized additive model (GAM). To characterize long-term trends, we compared the effect of heat on mortality over the decades., Results: Our estimate reveals that the unusually high summer temperatures in Germany between 2018 and 2020 led to a statistically significant number of deaths in all three years. There were approximately 8700 heat-related deaths in 2018, 6900 in 2019, and 3700 in 2020. There was no statistically significant heat-related increase in deaths in 2021. A comparison of the past three decades reveals a slight overall decline in the effect of high temperatures on mortality., Conclusion: Although evidence suggests that there has been some adaptation to heat over the years, the data from 2018-2020 in particular show that heat events remain a significant threat to human health in Germany.

Freshwater lakes represent an important source of the potent greenhouse gas methane (CH 4 ) to the atmosphere. Methane emissions are regulated to large parts by aerobic (MOx) and anaerobic (AOM) oxidation of methane, which are important CH 4 sinks in lakes. In contrast to marine benthic environments, our knowledge about the modes of AOM and the related methanotrophic microorganisms in anoxic lake sediments is still rudimentary. Here, we demonstrate the occurrence of AOM in the anoxic sediments of Lake Sempach (Switzerland), with maximum in situ AOM rates observed within the surface sediment layers in presence of multiple groups of methanotrophic bacteria and various oxidants known to support AOM. However, substrate-amended incubations (with NO 2 - , NO 3 - , SO 4 2- , Fe-, and Mn-oxides) revealed that none of the electron acceptors previously reported to support AOM enhanced methane turnover in Lake Sempach sediments under anoxic conditions. In contrast, the addition of oxygen to the anoxic sediments resulted in an approximately 10-fold increase in methane oxidation relative to the anoxic incubations. Phylogenetic and isotopic evidence indicate that both Type I and Type II aerobic methanotrophs were growing on methane under both oxic and anoxic conditions, although methane assimilation rates were an order of magnitude higher under oxic conditions. While the anaerobic electron acceptor responsible for AOM could not be identified, these findings expand our understanding of the metabolic versatility of canonically aerobic methanotrophs under anoxic conditions, with important implications for future investigations to identify methane oxidation processes. Bacterial AOM by facultative aerobic methane oxidizers might be of much larger environmental significance in reducing methane emissions than previously thought., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Su, Zopfi, Niemann and Lehmann.)

A growing number of sperm methylome analyses have identified genomic loci that are susceptible to paternal age effects in a variety of mammalian species, including human, bovine, and mouse. However, there is little overlap between different data sets. Here, we studied whether or not paternal age effects on the sperm epigenome have been conserved in mammalian evolution and compared methylation patterns of orthologous regulatory regions (mainly gene promoters) containing both conserved and non-conserved CpG sites in 94 human, 36 bovine, and 94 mouse sperm samples, using bisulfite pyrosequencing. We discovered three ( NFKB2 , RASGEF1C , and RPL6 ) age-related differentially methylated regions (ageDMRs) in humans, four ( CHD7 , HDAC11 , PAK1 , and PTK2B ) in bovines, and three ( Def6 , Nrxn2 , and Tbx19 ) in mice. Remarkably, the identified sperm ageDMRs were all species-specific. Most ageDMRs were in genomic regions with medium methylation levels and large methylation variation. Orthologous regions in species not showing this age effect were either hypermethylated (>80%) or hypomethylated (<20%). In humans and mice, ageDMRs lost methylation, whereas bovine ageDMRs gained methylation with age. Our results are in line with the hypothesis that sperm ageDMRs are in regions under epigenomic evolution and may be part of an epigenetic mechanism(s) for lineage-specific environmental adaptations and provide a solid basis for studies on downstream effects in the genes analyzed here.

Background: Infectious diseases of farmed and wild animals pose a recurrent threat to food security and human health. The macrophage, a key component of the innate immune system, is the first line of defence against many infectious agents and plays a major role in shaping the adaptive immune response. However, this phagocyte is a target and host for many pathogens. Understanding the molecular basis of interactions between macrophages and pathogens is therefore crucial for the development of effective strategies to combat important infectious diseases., Results: We explored how porcine pluripotent stem cells (PSCs) can provide a limitless in vitro supply of genetically and experimentally tractable macrophages. Porcine PSC-derived macrophages (PSCdMs) exhibited molecular and functional characteristics of ex vivo primary macrophages and were productively infected by pig pathogens, including porcine reproductive and respiratory syndrome virus (PRRSV) and African swine fever virus (ASFV), two of the most economically important and devastating viruses in pig farming. Moreover, porcine PSCdMs were readily amenable to genetic modification by CRISPR/Cas9 gene editing applied either in parental stem cells or directly in the macrophages by lentiviral vector transduction., Conclusions: We show that porcine PSCdMs exhibit key macrophage characteristics, including infection by a range of commercially relevant pig pathogens. In addition, genetic engineering of PSCs and PSCdMs affords new opportunities for functional analysis of macrophage biology in an important livestock species. PSCs and differentiated derivatives should therefore represent a useful and ethical experimental platform to investigate the genetic and molecular basis of host-pathogen interactions in pigs, and also have wider applications in livestock., (© 2021. The Author(s).)

The intestinal epithelium plays a variety of roles including providing an effective physical barrier and innate immune protection against infection. Two-dimensional models of the intestinal epithelium, 2D enteroids, are a valuable resource to investigate intestinal cell biology and innate immune functions and are suitable for high throughput studies of paracellular transport and epithelial integrity. We have developed a chicken 2D enteroid model that recapitulates all major differentiated cell lineages, including enterocytes, Paneth cells, Goblet cells, enteroendocrine cells and leukocytes, and self-organises into an epithelial and mesenchymal sub-layer. Functional studies demonstrated the 2D enteroids formed a tight cell layer with minimal paracellular flux and a robust epithelial integrity, which was maintained or rescued following damage. The 2D enteroids were also able to demonstrate appropriate innate immune responses following exposure to bacterial endotoxins, from Salmonella enterica serotype Typhimurium and Bacillus subtilis. Frozen 2D enteroids cells when thawed were comparable to freshly isolated cells. The chicken 2D enteroids provide a useful ex vivo model to study intestinal cell biology and innate immune function, and have potential uses in screening of nutritional supplements, pharmaceuticals, and bioactive compounds., (© 2021. The Author(s).)

The developmental competence of male and female gametes is frequently reduced under in vitro conditions, mainly due to oxidative stress during handling. The amino-acid derived hormone melatonin has emerged as a potent non-enzymatic antioxidant in many biological systems. The goal of the present study was to evaluate the effects of melatonin on post-thaw sperm quality, fertilizing ability, and embryo development and competence in vitro after in vitro fertilization. Frozen-thawed bovine spermatozoa were incubated either in the presence of 10-11 M melatonin (MT), or its solvent (ethanol; Sham-Control), or plain Tyrode's Albumin Lactate Pyruvate medium (TALP, Control). Computer-Assisted Sperm Analysis (CASA) and flow cytometry data after 30 min, 120 min, and 180 min incubation did not reveal any significant effects of melatonin on average motility parameters, sperm subpopulation structure as determined by hierarchical cluster, or on the percentage of viable, acrosome intact sperm, or viable sperm with active mitochondria. Nevertheless, in vitro matured cumulus-oocyte-complexes fertilized with spermatozoa which had been preincubated with 10-11 M melatonin (MT-Sperm) showed higher (P < 0.01) rates of monospermic fertilization, reduced (P < 0.05) polyspermy and enhanced (P < 0.05) embryo development compared to the Control group. Moreover, the relative abundance of MAPK13 in the in vitro-derived blastocysts was greater (P < 0.05) than observed in the Control group. In conclusion, adding melatonin to the sperm-preparation protocol for bovine IVF improved proper fertilization and enhanced embryonic development and competence in vitro., Competing Interests: The authors declare that they have no conflicts of interest.

Decellularization of xenogeneic heart valves might lead to excellent regenerative implants, from which many patients could benefit. However, this material carries various xenogeneic epitopes and thus bears a considerable inherent immunological risk. Here, we investigated the regenerative and immunogenic potential of xenogeneic decellularized heart valve implants using pigs deficient for the galactosyltransferase gene (GGTA1-KO) as novel large animal model. Decellularized aortic and pulmonary heart valves obtained from sheep, wild-type pigs or GGTA1-KO pigs were implanted into GGTA1-KO pigs for 3, or 6 months, respectively. Explants were analyzed histologically, immunhistologically (CD3, CD21 and CD172a) and anti-αGal antibody serum titers were determined by ELISA. Xenogeneic sheep derived implants exhibited a strong immune reaction upon implantation into GGTA1-KO pigs, characterized by massive inflammatory cells infiltrates, presence of foreign body giant cells, a dramatic increase of anti-αGal antibody titers and ultimately destruction of the graft, whereas wild-type porcine grafts induced only a mild reaction in GGTA1-KO pigs. Allogeneic implants, wild-type/wild-type and GGTA1-KO/GGTA1-KO valves did not induce a measurable immune reaction. Thus, GGTA1-KO pigs developed a 'human-like' immune response toward decellularized xenogeneic implants showing that immunogenicity of xenogeneic implants is not sufficiently reduced by decellularization, which detracts from their regenerative potential., (© Crown copyright 2021.)

Plastic particles in the ocean are typically covered with microbial biofilms, but it remains unclear whether distinct microbial communities colonize different polymer types. In this study, we analyzed microbial communities forming biofilms on floating microplastics in a bay of the island of Elba in the Mediterranean Sea. Raman spectroscopy revealed that the plastic particles mainly comprised polyethylene (PE), polypropylene (PP), and polystyrene (PS) of which polyethylene and polypropylene particles were typically brittle and featured cracks. Fluorescence in situ hybridization and imaging by high-resolution microscopy revealed dense microbial biofilms on the polymer surfaces. Amplicon sequencing of the 16S rRNA gene showed that the bacterial communities on all plastic types consisted mainly of the orders Flavobacteriales, Rhodobacterales, Cytophagales, Rickettsiales, Alteromonadales, Chitinophagales, and Oceanospirillales. We found significant differences in the biofilm community composition on PE compared with PP and PS (on OTU and order level), which shows that different microbial communities colonize specific polymer types. Furthermore, the sequencing data also revealed a higher relative abundance of archaeal sequences on PS in comparison with PE or PP. We furthermore found a high occurrence, up to 17% of all sequences, of different hydrocarbon-degrading bacteria on all investigated plastic types. However, their functioning in the plastic-associated biofilm and potential role in plastic degradation needs further assessment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Vaksmaa, Knittel, Abdala Asbun, Goudriaan, Ellrott, Witte, Vollmer, Meirer, Lott, Weber, Engelmann and Niemann.)

Most human embryos are aneuploid. Aneuploidy frequently arises during the early mitotic divisions of the embryo, but its origin remains elusive. Human zygotes that cluster their nucleoli at the pronuclear interface are thought to be more likely to develop into healthy euploid embryos. Here, we show that the parental genomes cluster with nucleoli in each pronucleus within human and bovine zygotes, and clustering is required for the reliable unification of the parental genomes after fertilization. During migration of intact pronuclei, the parental genomes polarize toward each other in a process driven by centrosomes, dynein, microtubules, and nuclear pore complexes. The maternal and paternal chromosomes eventually cluster at the pronuclear interface, in direct proximity to each other, yet separated. Parental genome clustering ensures the rapid unification of the parental genomes on nuclear envelope breakdown. However, clustering often fails, leading to chromosome segregation errors and micronuclei, incompatible with healthy embryo development., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Methane is the final product of the anaerobic decomposition of organic matter. The conversion of organic matter to methane (methanogenesis) as a mechanism for energy conservation is exclusively attributed to the archaeal domain. Methane is oxidized by methanotrophic microorganisms using oxygen or alternative terminal electron acceptors. Aerobic methanotrophic bacteria belong to the phyla Proteobacteria and Verrucomicrobia, while anaerobic methane oxidation is also mediated by more recently discovered anaerobic methanotrophs with representatives in both the bacteria and the archaea domains. The anaerobic oxidation of methane is coupled to the reduction of nitrate, nitrite, iron, manganese, sulfate, and organic electron acceptors (e.g., humic substances) as terminal electron acceptors. This review highlights the relevance of methanotrophy in natural and anthropogenically influenced ecosystems, emphasizing the environmental conditions, distribution, function, co-existence, interactions, and the availability of electron acceptors that likely play a key role in regulating their function. A systematic overview of key aspects of ecology, physiology, metabolism, and genomics is crucial to understand the contribution of methanotrophs in the mitigation of methane efflux to the atmosphere. We give significance to the processes under microaerophilic and anaerobic conditions for both aerobic and anaerobic methane oxidizers. In the context of anthropogenically influenced ecosystems, we emphasize the current and potential future applications of methanotrophs from two different angles, namely methane mitigation in wastewater treatment through the application of anaerobic methanotrophs, and the biotechnological applications of aerobic methanotrophs in resource recovery from methane waste streams. Finally, we identify knowledge gaps that may lead to opportunities to harness further the biotechnological benefits of methanotrophs in methane mitigation and for the production of valuable bioproducts enabling a bio-based and circular economy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Guerrero-Cruz, Vaksmaa, Horn, Niemann, Pijuan and Ho.)

The pig is an important model organism for biomedical research, mainly due to its extensive genetic, physiological and anatomical similarities with humans. Until date, direct conversion of somatic cells into hepatocyte-like cells (iHeps) has only been achieved in rodents and human cells. Here, we employed lentiviral vectors to screen a panel of 12 hepatic transcription factors (TF) for their potential to convert porcine fibroblasts into hepatocyte-like cells. We demonstrate for the first time, hepatic conversion of porcine somatic cells by over-expression of CEBPα, FOXA1 and HNF4α2 (3TF-piHeps). Reprogrammed 3TF-piHeps display a hepatocyte-like morphology and show functional characteristics of hepatic cells, including albumin secretion, Dil-AcLDL uptake, storage of lipids and glycogen and activity of cytochrome P450 enzymes CYP1A2 and CYP2C33 (CYP2C9 in humans). Moreover, we show that markers of mature hepatocytes are highly expressed in 3TF-piHeps, while fibroblastic markers are reduced. We envision piHeps as useful cell sources for future studies on drug metabolism and toxicity as well as in vitro models for investigation of pig-to-human infectious diseases.

The presence of plastics in the marine environment poses a threat to ocean life and has received much scientific and public attention in recent years. Plastics were introduced to the market in the 1950s and since then, global production figures and ocean plastic littering have increased exponentially. Of the 359 million tonnes (Mt) produced in 2018, an estimated 14.5 Mt has entered the ocean. In particular smaller plastic particles can be ingested by marine biota causing hazardous effects. Plastic marine debris (PMD) is exposed to physical, chemical and biological stressors. These cause macro and microplastic to break down into smaller fragments, including sub micrometre sized nanoplastic particles, which may account for an important but so far unevaluated fraction of the ocean plastic budget. Physicochemical and biological deterioration of PMD also leads to the release of more volatile compounds and the terminal oxidation of PMD, which most likely accounts for an important but also unevaluated fraction in the ocean plastic budget. This minireview provides an overview on (1) the quantity of plastic production and waste, pathways for plastics to enter the marine realm, the inventory of PMD and the negative effects of PMD to ocean life. (2) We discuss plastic degradation mechanisms in the ocean, expanding on the processes of photodegradation and biodegradation. (3) This review also highlights the emerging topic of nanoplastics in the sea and provides an overview on their specific physical and chemical properties, potential harm to ocean life, and nanoplastic detection techniques.

Bovine oocytes from prepubertal donors have been used for in vitro embryo production to decrease the generation interval. However, reduced cumulus-oocyte competence, mainly attributed to increased apoptosis, has been observed in oocytes/embryos collected from prepubertal donors. Here, we investigated the effects of the potent antioxidative molecule melatonin on cumulus-oocyte competence and embryo development in prepubertal and adult dairy cattle in vitro. A total of fifteen Holstein Friesian calves, six to ten months old (7.6 ± 1.34 months of age). And fifteen adult cows with one to four calvings (2.3 ± 0.96 calvings) were enrolled as ovum pick up (OPU) donors in this study. Cumulus-oocyte complexes (COCs) were cultured either in the presence or absence of melatonin (0.01 nM). The proportion of cleavage stages, blastocysts, and advanced blastocysts was determined. Embryo quality was assessed via differential staining to determine the total embryonic cells and allocation to the inner cell mass (ICM) and trophectoderm (TE) cells. Melatonin treatment yielded a greater percentage of blastocysts compared to the control group, i.e. oocytes from both adult cows (P = 0.0485; 24.8 ± 3.5% vs. 16.0 ± 3.4%, respectively), and from prepubertal donors (P = 0.0007; Melatonin 23.1 ± 5.1% vs. Control: 11.1 ± 3.5%). Adult cows had significantly (P = 0.0370) greater advanced blastocyst rates than those found in the prepubertal group (13.9%± vs. 7.0±%, respectively). Additionally, the number of ICM, total cells, and the ratios ICM: Total, ICM: TE, respectively, were greater (P < 0.05) after melatonin treatment compared with the control group (39.1 ± 2.8, 98.6 ± 5.7, 0.4 ± 0.01, and 0.7 ± 0.04 vs. 27.3 ± 2.9, 81.2 ± 5.8, 0.34 ± 0.01, and 0.52 ± 0.04, respectively). Blastocysts derived from adult cows had a greater number of TE (P = 0.01) and total embryonic cells (P = 0.0095) compared to the prepubertal donor group (63.5 ± 3.2 and 101.05 ± 4.8 vs. 48.9 ± 4.3 and 78.8 ± 6.5, respectively). Nevertheless, embryonic cell counting in embryos derived from prepubertal COCs equated to that observed from adult donors after melatonin exposure. In conclusion, these results indicate that the presence of melatonin during in vitro maturation improves cumulus-oocyte competence, embryo development, and quality by increasing the allocation of embryonic cells to the ICM compartment and the total number of embryonic cells in both adult and prepubertal bovine donors., Competing Interests: Declaration of competing interest No conflicts of interest occurred to carry out this experiment., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The sex-determining region on the Y chromosome (SRY) is thought to be the central genetic element of male sex development in mammals. Pathogenic modifications within the SRY gene are associated with a male-to-female sex reversal syndrome in humans and other mammalian species, including rabbits and mice. However, the underlying mechanisms are largely unknown. To understand the biological function of the SRY gene, a site-directed mutational analysis is required to investigate associated phenotypic changes at the molecular, cellular, and morphological level. Here, we successfully generated a knockout of the porcine SRY gene by microinjection of two CRISPR-Cas ribonucleoproteins, targeting the centrally located "high mobility group" (HMG), followed by a frameshift mutation of the downstream SRY sequence. This resulted in the development of genetically male (XY) pigs with complete external and internal female genitalia, which, however, were significantly smaller than in 9-mo-old age-matched control females. Quantitative digital PCR analysis revealed a duplication of the SRY locus in Landrace pigs similar to the known palindromic duplication in Duroc breeds. Our study demonstrates the central role of the HMG domain in the SRY gene in male porcine sex determination. This proof-of-principle study could assist in solving the problem of sex preference in agriculture to improve animal welfare. Moreover, it establishes a large animal model that is more comparable to humans with regard to genetics, physiology, and anatomy, which is pivotal for longitudinal studies to unravel mammalian sex determination and relevant for the development of new interventions for human sex development disorders., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

Background: The present study reports the development of a sensitive dot blot protocol for determining the level of preformed antibodies against porcine heart valve tissue derived from wild-type (WT) and α-Gal-KO (GGTA1-KO) pigs in human sera., Methods: The assay uses decellularized and solubilized heart valve tissue; antibody binding found in this dot blot assay could be correlated with antibody titers of preformed anti-α-Gal and anti-Neu5Gc antibodies detected by a sensitive ELISA., Results: The ultimate protocol had an inter-assay variance of 9.5% and an intra-assay variance of 9.2%, showing that the test is reliable and highly reproducible. With the aid of this dot blot assay, we found significant variation with regard to antibody contents among twelve human sera. Binding of preformed antibodies to WT tissue was significantly higher than to GGTA1-KO tissue., Conclusions: The dot blot assay described herein could be a valuable tool to measure preformed antibody levels in human sera against unknown epitopes on decellularized tissue prior to implantation. Ultimately, this prescreening may allow a matching of the porcine xenograft with the respective human recipients in demand and thus may become an important tool for graft long-term survival similar to current allotransplantation settings., (© 2020 The Authors. Xenotransplantation published by John Wiley & Sons Ltd.)

Cold seeps are characterized by high biomass, which is supported by the microbial oxidation of the available methane by capable microorganisms. The carbon is subsequently transferred to higher trophic levels. South of Svalbard, five geological mounds shaped by the formation of methane gas hydrates, have been recently located. Methane gas seeping activity has been observed on four of them, and flares were primarily concentrated at their summits. At three of these mounds, and along a distance gradient from their summit to their outskirt, we investigated the eukaryotic and prokaryotic biodiversity linked to 16S and 18S rDNA. Here we show that local methane seepage and other environmental conditions did affect the microbial community structure and composition. We could not demonstrate a community gradient from the summit to the edge of the mounds. Instead, a similar community structure in any methane-rich sediments could be retrieved at any location on these mounds. The oxidation of methane was largely driven by anaerobic methanotrophic Archaea-1 (ANME-1) and the communities also hosted high relative abundances of sulfate reducing bacterial groups although none demonstrated a clear co-occurrence with the predominance of ANME-1. Additional common taxa were observed and their abundances were likely benefiting from the end products of methane oxidation. Among these were sulfide-oxidizing Campilobacterota, organic matter degraders, such as Bathyarchaeota, Woesearchaeota, or thermoplasmatales marine benthic group D, and heterotrophic ciliates and Cercozoa., (Copyright © 2020 Carrier, Svenning, Gründger, Niemann, Dessandier, Panieri and Kalenitchenko.)

Background: As a consequence of global warming, heat waves are expected to become more frequent, more intense, and longer. The elderly and persons with chronic diseases are especially vulnerable to health problems due to heat. This article is devoted to the question of the extent to which the effects of heat waves in Germany are changing over time, and whether preventive health measures are working., Methods: We use a statistical model to quantify the effect of high mean temperatures on mortality. Within this model, different exposure-response curves for the three temporal intervals 1992-2000, 2001-2010, and 2011-2017 are estimated. Attention is also paid to the delayed effect on mortality of high mean temperatures in the preceding week., Results: Our analysis reveals a clear, systematic association of the mean temperature in the current week, as well as the mean temperature in the preceding week, with weekly mortality. This association is more pronounced for higher age groups and decreases over the years under analysis, with the exception of a relatively weak effect of heat in southern Germany in 1992-2000. The strongest effects were related to the heat waves in 1994 and 2003, with approximately 10 200 and 9600 fatalities, respectively. Approximately 7800 fatalities were estimated for the summer of 2006, and 4700 and 5200 for 2010 and 2015, respectively., Conclusion: In Germany, as elsewhere, climate change has been causing more frequent, more intense, and longer periods of heat in the summer. The harmful effect of heat on health is reduced by adaptive processes, presumably including successful preventive measures. Such measures should be extended in the future, and perhaps complemented by other measures in order to further diminish the effect of heat on mortality .

In modern livestock farming horned cattle pose an increased risk of injury for each other as well as for the farmers. Dehorning without anesthesia is associated with stress and pain for the calves and raises concerns regarding animal welfare. Naturally occurring structural variants causing polledness are known for most beef cattle but are rare within the dairy cattle population. The most common structural variant in beef cattle consists of a 202 base pair insertion-deletion (Polled Celtic variant). For the generation of polled offspring from a horned Holstein-Friesian bull, we isolated the Polled Celtic variant from the genome of an Angus cow and integrated it into the genome of fibroblasts taken from the horned bull using the CRISPR/Cas12a system (formerly Cpf1). Modified fibroblasts served as donor cells for somatic cell nuclear transfer and reconstructed embryos were transferred into synchronized recipients. One resulting pregnancy was terminated on day 90 of gestation for the examination of the fetus. Macroscopic and histological analyses proved a polled phenotype. The remaining pregnancy was carried to term and delivered one calf with a polled phenotype which died shortly after birth. In conclusion, we successfully demonstrated the practical application of CRISPR/Cas12a in farm animal breeding and husbandry.

Environmental noise is a great burden for the population in Germany, especially in urban areas. People are often exposed not only to one but several noise sources. Long-term exposure to environmental noise can have several and severe adverse effects on human health, such as noise annoyance, sleep disturbances, ischaemic heart disease and depression. Additional burdens arise from multiple exposure to particulate matter or air pollutants. The current article describes the relevant adverse health effects due to long-term environmental noise exposure, discusses the challenges of environmental noise in urban areas different and introduces measures and instruments to abate environmental noise.

In somatic cells/tissues, methylation of ribosomal DNA (rDNA) increases with age and age-related pathologies, which has a direct impact on the regulation of nucleolar activity and cellular metabolism. Here, we used bisulfite pyrosequencing and show that methylation of the rDNA transcription unit including upstream control element (UCE), core promoter, 18S rDNA, and 28S rDNA in human sperm also significantly increases with donor's age. This positive correlation between sperm rDNA methylation and biological age is evolutionarily conserved among mammals with widely different life spans such as humans, marmoset, bovine, and mouse. Similar to the tandemly repeated rDNA, methylation of human α-satellite and interspersed LINE1 repeats, marmoset α-satellite, bovine alpha- and testis satellite I, mouse minor and major satellite, and LINE1-T repeats increases in the aging male germline, probably related to their sperm histone packaging. Deep bisulfite sequencing of single rDNA molecules in human sperm revealed that methylation does not only depend on donor's age, but also depend on the region and sequence context (A vs. G alleles). Both average rDNA methylation of all analyzed DNA molecules and the number of fully (>50%) methylated alleles, which are thought to be epigenetically silenced, increase with donor's age. All analyzed CpGs in the sperm rDNA transcription unit show comparable age-related methylation changes. Unlike other epigenetic aging markers, the rDNA clock appears to operate in similar ways in germline and soma in different mammalian species. We propose that sperm rDNA methylation, directly or indirectly, influences nucleolar formation and developmental potential in the early embryo., (© 2020 The Authors. Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Chimeric pigs harboring organs derived from human stem cells are promising for patient-specific regenerative therapies. Induced pluripotent stem cells (iPSCs) can contribute to all cell types of the fetus, including germline after injection into embryos. However, ethical concerns prohibit testing human iPSCs in chimera assays. Here, we evaluated porcine embryos as hosts for an interspecies chimera assay using iPSCs from either cynomolgus monkeys (cyiPSCs) or mouse (miPSCs). To establish an in vitro culture system compatible for cyiPSCs and porcine embryos, we determined blastocyst development in eight different stem cell media. The highest developmental rates of blastocysts were achieved in Knockout Dulbecco's modified Eagle's medium with 20% knockout serum replacement. We found that cyiPSCs injected into porcine embryos survived in vitro and were mostly located in the trophectoderm (TE). Instead, when miPSCs were injected into porcine embryos, the cells rapidly proliferated. The behavior of chimeras developed in vitro was recapitulated in vivo ; cyiPSCs were observed in the TE, but not in the porcine epiblast. However, when miPSCs were injected into in vivo derived porcine embryos, mouse cells were found in both, the epiblast and TE. These results demonstrate that porcine embryos could be useful for evaluating the interspecies chimera-forming ability of iPSCs from different species.

Xenotransplantation of pancreatic islets offers a promising alternative to overcome the shortage of allogeneic donors. Despite significant advances, either immune rejection or oxygen supply in immune protected encapsulated islets remains major bottlenecks for clinical application. To decrease xenogeneic immune responses, we generated tissue engineered swine leucocyte antigen (SLA)-silenced islet cell clusters (ICC). Single-cell suspensions from pancreatic islets were generated by enzymatic digestion of porcine ICCs. Cells were silenced for SLA class I and class II by lentiviral vectors encoding for short hairpin RNAs targeting beta2-microglobulin or class II transactivator, respectively. SLA-silenced ICCs-derived cells were then used to form new ICCs in stirred bioreactors in the presence of collagen VI. SLA class I silencing was designed to reach a level of up to 89% and class II by up to 81% on ICCs-derived cells. Xenogeneic T cell immune responses, NK cell and antibody-mediated cellular-dependent immune responses were significantly decreased in SLA-silenced cells. In stirred bioreactors, tissue engineered islets showed the typical 3D structure and insulin production. These data show the feasibility to generate low immunogenic porcine ICCs after single-cell engineering and post-transduction islet reassembling that might serve as an alternative to allogeneic pancreatic islet cell transplantation., (© 2020 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Porcine xenografts lacking swine leukocyte antigen (SLA) class I are thought to be protected from human T cell responses. We have previously shown that SLA class I deficiency can be achieved in pigs by CRISPR/Cas9-mediated deletion of β 2 -microglobulin (B2M). Here, we characterized another line of genetically modified pigs in which targeting of the B2M locus did not result in complete absence of B2M and SLA class I but rather in significantly reduced expression levels of both molecules. Residual SLA class I was functionally inert, because no proper differentiation of the CD8 + T cell subset was observed in B2M low pigs. Cells from B2M low pigs were less capable in triggering proliferation of human peripheral blood mononuclear cells in vitro, which was mainly due to the nonresponsiveness of CD8 + T cells. Nevertheless, cytotoxic effector cells developing from unaffected cell populations (eg, CD4 + T cells, natural killer cells) lysed targets from both SLA class I + wildtype and SLA class I low pigs with similar efficiency. These data indicate that the absence of SLA class I is an effective approach to prevent the activation of human CD8 + T cells during the induction phase of an anti-xenograft response. However, cytotoxic activity of cells during the effector phase cannot be controlled by this approach., (© 2019 The Authors. American Journal of Transplantation published by Wiley Periodicals, Inc. on behalf of The American Society of Transplantation and the American Society of Transplant Surgeons.)

How much of the greenhouse gas methane is transported from the seafloor to the atmosphere is unclear. Here, we present data describing an extensive ebullition event that occurred in Eckernförde Bay, a shallow gas-hosting coastal inlet in the Baltic Sea, in the fall of 2014. A weak storm induced hydrostatic pressure fluctuations that in turn stimulated gas ebullition from the seabed. In a finely tuned sonar survey of the bay, we obtained a hydroacoustic dataset with exceptionally high sensitivity for bubble detection. This allowed us to identify 2849 bubble seeps rising within 28 h from the seafloor across the 90 km² study site. Based on our calculations, the estimated bubble-driven episodic methane flux from the seafloor across the bay is 1,900 μMol m -2 d -1 . Our study demonstrates that storm-associated fluctuations of hydrostatic pressure induce bulk gas-driven ebullitions. Given the extensive occurrence of shallow gas-hosting sediments in coastal seas, similar ebullition events probably take place in many parts of the Western Baltic Sea. However, these are likely to be missed during field investigations, due to the lack of high-quality data acquisition during storms, such that atmospheric inputs of marine-derived methane will be highly underestimated.

Several mud volcanoes are active in the Canadian Beaufort Sea. In this study, we investigated vertical variations in methanotrophic communities in sediments of the mud volcano MV420 (420 m water depth) by analyzing geochemical properties, microbial lipids, and nucleic acid signatures. Three push cores were collected with a remotely operated vehicle from visually discriminative habitats that were devoid of megafauna and/microbial mats (DM) to the naked eye, covered with bacterial mats (BM), or colonized by siboglinid tubeworms (ST). All MV420 sites showed the presence of aerobic methane oxidation (MOx)- and anaerobic methane oxidation (AOM)-related lipid biomarkers (4α-methyl sterols and sn-2-hydroxyarchaeol, respectively), which were distinctly different in comparison with a reference site at which these compounds were not detected. Lipid biomarker results were in close agreement with 16S rRNA analyses, which revealed the presence of MOx-related bacteria (Methylococcales) and AOM-related archaea (ANME-2 and ANME-3) at the MV420 sites. 4α-methyl sterols derived from Methylococcales predominated in the surface layer at the BM site, which showed a moderate methane flux (0.04 mmol cm -2 y -1 ), while their occurrence was limited at the DM (0.06 mmol cm -2 y -1 ) and ST (0.01 mmol cm -2 y -1 ) sites. On the other hand, 13 C-depleted sn-2-hydroxyarchaeol potentially derived from ANME-2 and/or ANME-3 was abundant in down-core sediments at the ST site. Our study indicates that a niche diversification within this mud volcano system has shaped distinct methanotrophic communities due to availability of electron acceptors in association with varying degrees of methane flux and bioirrigation activity.

Background: Despite major improvements in pig-to-primate xenotransplantation, long-term survival of xenografts is still challenging. The major histocompatibility complex (MHC) class I, which is crucial in cellular immune response, is an important xenoantigen. Abrogating MHC class I expression on xenografts might be beneficial for extending graft survival beyond current limits., Methods: In this study, we employed the CRISPR/Cas9 system to target exon 2 of the porcine beta-2-microglobulin (B2M) gene to abrogate SLA class I expression on porcine cells. B2M-KO cells served as donor cells for somatic cell nuclear transfer, and cloned embryos were transferred to three recipient sows. The offspring were genotyped for mutations at the B2M locus, and blood samples were analyzed via flow cytometry for the absence of SLA class I molecules., Results: Pregnancies were successfully established and led to the birth of seven viable piglets. Genomic sequencing proved that all piglets carried biallelic modifications at the B2M locus leading to a frameshift, a premature stop codon, and ultimately a functional knockout. However, survival times of these animals did not exceed 4 weeks due to unexpected disease processes., Conclusion: Here, we demonstrate the feasibility of generating SLA class I knockout pigs by targeting the porcine beta-2-microglobulin gene using the CRISPR/Cas9 system. Additionally, our findings indicate for the first time that this genetic modification might have a negative impact on the viability of the animals. These issues need to be solved to unveil the real value for xenotransplantation in the future., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The regulations for fitness to drive after a cerebrovascular accident in the German Driving License Regulations (FeV) and the German Evaluation Guidelines for Driving Ability (BGL). are not up to date with the current medical knowledge and not consistent with regulations regarding cardiovascular diseases. This position paper presented by six medical and neuropsychological societies in Germany provides a guideline for the assessment of driving ability after diagnosis of a cerebrovascular disease and addresses three major questions: If there is a functional limitation, how can it be compensated for? What is the risk of sudden loss of control while driving in the future? Are there behavioral or personality changes or cognitive deficiencies interfering with safety while driving? Recommendations for the assessment of driving ability in different cerebrovascular diseases are presented. This article is a translation of the position paper published in Nervenarzt: Marx, P., Hamann, G.F., Busse, O. et al. Nervenarzt 90(4): 388-398., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2019.)

For the first time, the optimized stellarator Wendelstein 7-X has operated with an island divertor. An operation regime in hydrogen was found in which the total plasma radiation approached the absorbed heating power without noticeable loss of stored energy. The divertor thermography recorded simultaneously a strong reduction of the heat load on all divertor targets, indicating almost complete power detachment. This operation regime was stably sustained over several energy confinement times until the preprogrammed end of the discharge. The plasma radiation is mainly due to oxygen and is located at the plasma edge. This plasma scenario is reproducible and robust at various heating powers, plasma densities, and gas fueling locations. These experimental results show that the island divertor concept actually works and displays good power dissipation potential, producing a promising exhaust concept for the stellarator reactor line.

Sedimentary biofilms comprising microbial communities mediating the anaerobic oxidation of methane are rare. Here, we describe two biofilm communities discovered in sediment cores recovered from Arctic cold seep sites (gas hydrate pingos) in the north-western Barents Sea, characterized by steady methane fluxes. We found macroscopically visible biofilms in pockets in the sediment matrix at the depth of the sulphate-methane-transition zone. 16S rRNA gene surveys revealed that the microbial community in one of the two biofilms comprised exclusively of putative anaerobic methanotrophic archaea of which ANME-1 was the sole archaeal taxon. The bacterial community consisted of relatives of sulphate-reducing bacteria (SRB) belonging to uncultured Desulfobacteraceae clustering into SEEP-SRB1 (i.e. the typical SRB associated to ANME-1), and members of the atribacterial JS1 clade. Confocal laser scanning microscopy demonstrates that this biofilm is composed of multicellular strands and patches of ANME-1 that are loosely associated with SRB cells, but not tightly connected in aggregates. Our discovery of methanotrophic biofilms in sediment pockets closely associated with methane seeps constitutes a hitherto overlooked and potentially widespread sink for methane and sulphate in marine sediments.

We recently derived mouse expanded potential stem cells (EPSCs) from individual blastomeres by inhibiting the critical molecular pathways that predispose their differentiation. EPSCs had enriched molecular signatures of blastomeres and possessed developmental potency for all embryonic and extra-embryonic cell lineages. Here, we report the derivation of porcine EPSCs, which express key pluripotency genes, are genetically stable, permit genome editing, differentiate to derivatives of the three germ layers in chimeras and produce primordial germ cell-like cells in vitro. Under similar conditions, human embryonic stem cells and induced pluripotent stem cells can be converted, or somatic cells directly reprogrammed, to EPSCs that display the molecular and functional attributes reminiscent of porcine EPSCs. Importantly, trophoblast stem-cell-like cells can be generated from both human and porcine EPSCs. Our pathway-inhibition paradigm thus opens an avenue for generating mammalian pluripotent stem cells, and EPSCs present a unique cellular platform for translational research in biotechnology and regenerative medicine.

Background: During the summers of 2003 and 2015, heat was found to be the cause of a substantial number of deaths in Germany. Until now, estimates for the total number of heat-related deaths were only available regionally in Germany. For the summer of 2003, an analysis for Baden-Württemberg was extrapolated to the whole of Germany., Objectives: Our analysis tries to prove a stable statistical relationship between heat and mortality and to use this to quantify the number of heat-related deaths in Germany between the years 2001 and 2015., Materials and Methods: By fitting a nonlinear statistical model, we estimated exposure-response curves that describe the influence of heat on the mortality rate. The performance of different indicators for heat stress was compared., Results: The comparison of the different indicators for heat showed that the weekly mean temperature was most useful to explain the course of the weekly mortality during the summer. The relation between mortality rate and weekly mean temperature varied between age groups and regions in Germany (north, middle, south). The age groups (75-84) and (85+) were most affected by heat. The highest number of heat-related deaths was 7600 (95% CI 5500-9900), found for the summer 2003, followed by 6200 (4000; 8000) in the summer 2006 and 6100 (4000; 8300) in the summer 2015., Conclusions: We could show that even in weekly data on mortality, a clear influence of heat could be identified. A national surveillance of mortality that allows real-time monitoring would be desirable.

The regulations for ability to drive with cerebrovascular diseases in the German Driving License Regulations (Fahrerlaubnisverordnung, FeV) and German Guidelines for the Evaluation of Driving Ability of the Federal Highway Research Institute (BASt) are not up to date with the current medical knowledge and are not consistent with comparable regulations regarding cardiovascular diseases. This is particularly true for the assessment of future risks for a sudden loss of control during driving. The present position paper of six medical and neuropsychological societies in Germany presents the current conditions for the assessment of driving ability of patients a cerebrovascular diesease and recommends an estimation of the ability to drive founded on the current state of scientific knowledge. It addresses the following: 1. Physical and mental functional limitations and the possibilities for compensation, which if necessary enable a fitness to drive under conditions or within limits, including the importance of behavioral or personality changes and cognitive deficiencies that interfere with safety. 2. The potential danger due to a sudden loss of control as a result of a transient ischemic attack (TIA) new stroke event, or another cardiovascular event while driving. A summary in the form of a table provides physicians and expert assessors with assistance for the most important cerebrovascular diseases.