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During spermatogenesis most histones are replaced with protamines. The distribution of the products of Trithorax and Polycomb histone modifications are now examined in both mouse and human sperm, showing conservation of histone methylation distribution across species. The authors propose a role for the Polycomb complex in transgenerational inheritance. In higher eukaryotes, histone methylation is involved in maintaining cellular identity during somatic development. As most nucleosomes are replaced by protamines during spermatogenesis, it is unclear whether histone modifications function in paternal transmission of epigenetic information. Here we show that two modifications important for Trithorax- and Polycomb-mediated gene regulation have methylation-specific distributions at regulatory regions in human spermatozoa. Histone H3 Lys4 dimethylation (H3K4me2) marks genes that are relevant in spermatogenesis and cellular homeostasis. In contrast, histone H3 Lys27 trimethylation (H3K27me3) marks developmental regulators in sperm, as in somatic cells. However, nucleosomes are only moderately retained at regulatory regions in human sperm. Nonetheless, genes with extensive H3K27me3 coverage around transcriptional start sites in particular tend not to be expressed during male and female gametogenesis or in preimplantation embryos. Promoters of orthologous genes are similarly modified in mouse spermatozoa. These data are compatible with a role for Polycomb in repressing somatic determinants across generations, potentially in a variegating manner.

Cellular differentiation entails reprogramming of the transcriptome from a pluripotent to a unipotent fate. This process was suggested to coincide with a global increase of repressive heterochromatin, which results in a reduction of transcriptional plasticity and potential. Here we report the dynamics of the transcriptome and an abundant heterochromatic histone modification, dimethylation of histone H3 at lysine 9 (H3K9me2), during neuronal differentiation of embryonic stem cells. In contrast to the prevailing model, we find H3K9me2 to occupy over 50% of chromosomal regions already in stem cells. Marked are most genomic regions that are devoid of transcription and a subgroup of histone modifications. Importantly, no global increase occurs during differentiation, but discrete local changes of H3K9me2 particularly at genic regions can be detected. Mirroring the cell fate change, many genes show altered expression upon differentiation. Quantitative sequencing of transcripts demonstrates however that the total number of active genes is equal between stem cells and several tested differentiated cell types. Together, these findings reveal high prevalence of a heterochromatic mark in stem cells and challenge the model of low abundance of epigenetic repression and resulting global basal level transcription in stem cells. This suggests that cellular differentiation entails local rather than global changes in epigenetic repression and transcriptional activity.

The Polycomb repressive complex 1 (PRC1) is found to have important gene-dosage-dependent and sex-specific roles in primordial germ cell (PGC) development, including the maintenance of high levels of Oct4 and Nanog and ensuring the proper timing of meiosis through the suppression of retinoic acid signalling in female PGCs. PRC1 role in germ cell development Polycomb group proteins are involved in the transcriptional repression of developmental regulators in embryonic stem cells, where they maintain pluripotency and cell identity during subsequent development. Antoine Peters and colleagues have studied the function of the Polycomb repressive complex 1 (PRC1) in the development of mouse primordial germ cells (PGCs). They observed multiple sex-specific roles of PRC1 in development. PRC1 is required for the maintenance of high levels of expression of the transcription factors Oct4 and Nanog. And by suppressing retinoic acid signalling provided by the somatic compartment of the female genital ridge, PRC1 also ensures proper timing of meiotic induction. In mammals, sex differentiation of primordial germ cells (PGCs) is determined by extrinsic cues from the environment 1 . In mouse female PGCs, expression of stimulated by retinoic acid gene 8 ( Stra8 ) and meiosis are induced in response to retinoic acid provided from the mesonephroi 2 , 3 , 4 , 5 . Given the widespread role of retinoic acid signalling during development 6 , 7 , the molecular mechanisms that enable PGCs to express Stra8 and enter meiosis in a timely manner are unknown 2 , 8 . Here we identify gene-dosage-dependent roles in PGC development for Ring1 and Rnf2 , two central components of the Polycomb repressive complex 1 (PRC1) 9 , 10 . Both paralogues are essential for PGC development between days 10.5 and 11.5 of gestation. Rnf2 is subsequently required in female PGCs to maintain high levels of Oct4 (also known as Pou5f1 ) and Nanog expression 11 , and to prevent premature induction of meiotic gene expression and entry into meiotic prophase. Chemical inhibition of retinoic acid signalling partially suppresses precocious Oct4 downregulation and Stra8 activation in Rnf2 -deficient female PGCs. Chromatin immunoprecipitation analyses show that Stra8 is a direct target of PRC1 and PRC2 in PGCs. These data demonstrate the importance of PRC1 gene dosage in PGC development and in coordinating the timing of sex differentiation of female PGCs by antagonizing extrinsic retinoic acid signalling.

Background In microbial diagnostics, whole-genome sequencing (WGS) is used to address key questions such as species identification, presence of antimicrobial resistance genes (ARGs), virulence genes, and outbreak detection. The choice of sequencing technology is crucial to ensure high-quality data, cost-effectiveness, and efficient reporting times. We aimed to compare Illumina (short-read) and ONT (long-read) sequencing methods for WGS on different bacterial species for base accuracy and reliable taxonomic and ARG identification. Materials and methods We used clinical isolates of ESKAPE pathogens ( n  = 12) and ATCC strains ( n  = 8) of varying %G + C. Illumina sequencing was performed on MiSeq (PE150) and ONT sequencing using GridION with R9.4.1 and R10.4.1 flowcells. Base-calling was performed using Guppy, Dorado, and Rerio software. We performed de novo assembly with Unicycler for Illumina and Flye for ONT, and two types of hybrid assemblies, Unicycler and Polypolish. We annotated genomes with Bakta and assessed the quality (QUAST, GTDB-Tk). We identified ARGs (AMRFinderPlus) and plasmids (MOB-suite). We mapped reads and called SNPs using Minimap2, Pilon, vcftools, and Snippy (Illumina). Core genome MLST analysis was conducted with Ridom Seqsphere+. Results We observed that Illumina sequencing provided consistently high-quality reads (median Q-score 35), whereas for ONT R10.4.1, SUP model showed higher median quality (median Q-score 15.3) compared to R9.4.1 (median Q-score 13.9, SUP model). We observed that Illumina-based assemblies generated fewer genes annotated as disrupted; for ONT assemblies, the base-caller affects assembly annotation accuracy, with High accuracy (HAC) and Super accuracy (SUP) base-calling models perform better than FAST model. ONT assemblies resolved rRNA operons better than Illumina assemblies. Sequencing errors were determined by SNP calling, and varied widely by species, with ONT often generating more sequencing errors compared to Illumina. Hybrid assemblies combine accuracy and completeness effectively. Taxonomic identification and ARG detection were reliable across all methods. Conclusion Combining Illumina and ONT technologies yielded optimal bacterial genome sequencing results, leveraging the high accuracy of short reads and improved contiguity of ONT long reads. The HAC and SUP ONT models with Dorado notably enhance genome assembly annotation and resolution of complex regions, although species-specific issues, likely due to repeat regions and base modifications, remain challenging even in SUP model with Dorado. Hybrid approaches currently offer the most comprehensive and accurate genome assemblies for clinical microbiology. For reliable cgMLST even using the most recent ONT methods, resolution must be assessed on a species-by-species basis.

Despite the discovery of familial cases with mutations in Cu/Zn-superoxide dismutase (SOD1), Guanine nucleotide exchange C9orf72, TAR DNA-binding protein 43 (TARDBP) and RNA-binding protein FUS as well as a number of other genes linked to Amyotrophic Lateral Sclerosis (ALS), the etiology and molecular pathogenesis of this devastating disease is still not understood. As proteins do not act alone, conducting an analysis of ALS at the system level may provide new insights into the molecular biology of ALS and put it into relationship to other neurological diseases. A set of ALS-associated genes/proteins were collected from publicly available databases and text mining of scientific literature. We used these as seed proteins to build protein-protein interaction (PPI) networks serving as a scaffold for further analyses. From the collection of networks, a set of core modules enriched in seed proteins were identified. The molecular biology of the core modules was investigated, as were their associations to other diseases. To assess the core modules' ability to describe unknown or less well-studied ALS biology, they were queried for proteins more recently associated to ALS and not involved in the primary analysis. We describe a set of 26 ALS core modules enriched in ALS-associated proteins. We show that these ALS core modules not only capture most of the current knowledge about ALS, but they also allow us to suggest biological interdependencies. In addition, new associations of ALS networks with other neurodegenerative diseases, e.g. Alzheimer's, Huntington's and Parkinson's disease were found. A follow-up analysis of 140 ALS-associated proteins identified since 2014 reveals a significant overrepresentation of new ALS proteins in these 26 disease modules. Using protein-protein interaction networks offers a relevant approach for broadening the understanding of the biological context of known ALS-associated genes. Using a bottom-up approach for the analysis of protein-protein interaction networks is a useful method to avoid bias caused by over-connected proteins. Our ALS-enriched modules cover most known biological functions associated with ALS. The presence of recently identified ALS-associated proteins in the core modules highlights the potential for using these as a scaffold for identification of novel ALS disease mechanisms.

The first case of SARS-CoV-2 in Basel, Switzerland was detected on February 26 th 2020. We present a phylogenetic study to explore viral introduction and evolution during the exponential early phase of the local COVID-19 outbreak from February 26 th until March 23 rd . We sequenced SARS-CoV-2 naso-oropharyngeal swabs from 746 positive tests that were performed at the University Hospital Basel during the study period. We successfully generated 468 high quality genomes from unique patients and called variants with our COVID-19 Pipeline (COVGAP), and analysed viral genetic diversity using PANGOLIN taxonomic lineages. To identify introduction and dissemination events we incorporated global SARS-CoV-2 genomes and inferred a time-calibrated phylogeny. Epidemiological data from patient questionnaires was used to facilitate the interpretation of phylogenetic observations. The early outbreak in Basel was dominated by lineage B.1 (83·6%), detected first on March 2 nd , although the first sample identified belonged to B.1.1. Within B.1, 68·2% of our samples fall within a clade defined by the SNP C15324T (‘Basel cluster’), including 157 identical sequences at the root of the ‘Basel cluster’, some of which we can specifically trace to regional spreading events. We infer the origin of B.1-C15324T to mid-February in our tri-national region. The other genomes map broadly over the global phylogenetic tree, showing several introduction events from and/or dissemination to other regions of the world via travellers. Family transmissions can also be traced in our data. A single lineage variant dominated the outbreak in the Basel area while other lineages, such as the first (B.1.1), did not propagate. A mass gathering event was the predominant initial source of cases, with travel returners and family transmissions to a lesser extent. We highlight the importance of adding specific questions to epidemiological questionnaires, to obtain data on attendance of large gatherings and their locations, as well as travel history, to effectively identify routes of transmissions in up-coming outbreaks. This phylogenetic analysis in concert with epidemiological and contact tracing data, allows connection and interpretation of events, and can inform public health interventions. Trial Registration: ClinicalTrials.gov NCT04351503 .

Blood cultures (BC) are critical for the diagnosis of bloodstream infections, pathogen identification, and resistance testing. Guidelines recommend a blood volume of 8-10 mL per bottle as lower volumes result in decreased sensitivity. We aimed to evaluate factors for non-adherence to recommended volumes and assess the effects on diagnostic performance. From February to April 2020, we measured collected blood volumes by weighing all BC containers from inpatient samples at the University Hospital Basel. Information on BC volumes was merged with clinical and microbiological data, as well as nursing staff schedules. We analyzed factors associated with (i) BC sampling volume, (ii) reaching recommended volumes (≥8 mL), (iii) BC positivity, and (iv) time to positivity using linear and generalized linear mixed effect models. We evaluated a total of 4'118 BC bottles collected from 686 patients. A total of 1'495 (36.3%) of all bottles contained the recommended filling volume of ≥8 mL. Using a central venous and arterial catheter for drawing blood resulted in an increase of filling volume by 0.26 mL (95% CI 0.10, 0.41) and 0.50 mL (95% CI 0.31, 0.69) compared to peripheral venipuncture, respectively. Each additional nursing staff working at the time of blood drawing was associated with 6% higher odds of achieving the recommended filling volume. We found no significant correlation between the filling volume and the positivity rate. Our results indicate critical pre-analytical quality markers linked to BC collection procedures to reach recommended collection volumes. No significant impact on the positivity rate was found.

Background Urinary tract infections (UTIs) are among the most common bacterial infections worldwide, often caused by uropathogenic Escherichia coli . Multiple bacterial virulence factors or patient characteristics have been linked separately to progressive, more invasive infections. In this study, we aim to identify pathogen- and patient-specific factors that drive the progression to urosepsis by jointly analysing bacterial and host characteristics. Methods We analysed 1076 E. coli strains isolated from 825 clinical cases with UTI and/or bacteraemia by whole-genome sequencing (Illumina). Sequence types (STs) were determined via srst2 and capsule loci via fastKaptive. We compared the isolates from urine and blood to confirm clonality. Furthermore, we performed a bacterial genome-wide association study (bGWAS) (pyseer) using bacteraemia as the primary clinical outcome. Clinical data were collected by an electronic patient chart review. We concurrently analysed the association of the most significant bGWAS hit and important patient characteristics with the clinical endpoint bacteraemia using a generalised linear model (GLM). Finally, we designed qPCR primers and probes to detect papGII -positive E. coli strains and prospectively screened E. coli from urine samples ( n  = 1657) at two healthcare centres. Results Our patient cohort had a median age of 75.3 years (range: 18.00–103.1) and was predominantly female (574/825, 69.6%). The bacterial phylogroups B2 (60.6%; 500/825) and D (16.6%; 137/825), which are associated with extraintestinal infections, represent the majority of the strains in our collection, many of which encode a polysaccharide capsule (63.4%; 525/825). The most frequently observed STs were ST131 (12.7%; 105/825), ST69 (11.0%; 91/825), and ST73 (10.2%; 84/825). Of interest, in 12.3% (13/106) of cases, the E. coli pairs in urine and blood were only distantly related. In line with previous bGWAS studies, we identified the gene papGII ( p -value < 0.001), which encodes the adhesin subunit of the E. coli P-pilus, to be associated with ‘bacteraemia’ in our bGWAS. In our GLM, correcting for patient characteristics, papGII remained highly significant (odds ratio = 5.27, 95% confidence interval = [3.48, 7.97], p -value < 0.001). An independent cohort of cases which we screened for papGII -carrying E. coli at two healthcare centres further confirmed the increased relative frequency of papGII -positive strains causing invasive infection, compared to papGII -negative strains ( p -value = 0.033, chi-squared test). Conclusions This study builds on previous work linking papGII with invasive infection by showing that it is a major risk factor for progression from UTI to bacteraemia that has diagnostic potential.

We show that elevated levels of Ret receptor are found in different sub‐types of human breast cancers and that high Ret correlates with decreased metastasis‐free survival. The role of Ret in ER+ breast cancer models was explored combining in vitro and in vivo approaches. Our analyses revealed that ligand‐induced Ret activation: (i) stimulates migration of breast cancer cells; (ii) rescues cells from anti‐proliferative effects of endocrine treatment and (iii) stimulates expression of cytokines in the presence of endocrine agents. Indeed, we uncovered a positive feed‐forward loop between the inflammatory cytokine IL6 and Ret that links them at the expression and the functional level. In vivo inhibition of Ret in a metastatic breast cancer model inhibits tumour outgrowth and metastatic potential. Ret inhibition blocks the feed‐forward loop by down‐regulating Ret levels, as well as decreasing activity of Fak, an integrator of IL6‐Ret signalling. Our results suggest that Ret kinase should be considered as a novel therapeutic target in subsets of breast cancer. Graphical Abstract Receptor tyrosine kinases (RTK) are validated targets for cancer therapy, but only subsets of patients are eligible for this treatment. Ret inhibition blocks a novel Ret‐IL6 feed‐forward loop, tumor growth and metastasis of ER+ breast cancers.

At our tertiary children’s hospital, infections with newly detected methicillin-resistant Staphylococcus aureus (MRSA) among children attending primary (age 6–12 years) and secondary school (age 13–16 years) nearly doubled in 2018 compared to previous years. This observation initiated an epidemiological outbreak investigation including phenotypic (susceptibility testing) and genotypic (whole genome sequencing) characterization of the isolates. In addition, a cross-sectional study was conducted to determine source of the outbreak, colonization frequency and to identify risk factors for transmission using a questionnaire. As a result, 49 individuals were detected with 57 corresponding isolates. Based on the case definition combined with whole genome sequencing, a core cluster was identified that shared common genetic features and a similar antimicrobial susceptibility pattern (efflux-mediated macrolide resistance, tetracycline susceptibility along with presence of Panton-Valentine leukocidin). Epidemiologic evaluation identified a distinct school as a common risk factor. However, the source of the clustered infections within that school could not be further specified. No further cases could be detected after decolonization of infected and colonized children.