A global metagenomic map of urban microbiomes and antimicrobial resistance.

We present a global atlas of 4,728 metagenomic samples from mass-transit systems in 60 cities over 3 years, representing the first systematic, worldwide catalog of the urban microbial ecosystem. This atlas provides an annotated, geospatial profile of microbial strains, functional characteristics, antimicrobial resistance (AMR) markers, and genetic elements, including 10,928 viruses, 1,302 bacteria, 2 archaea, and 838,532 CRISPR arrays not found in reference databases. We identified 4,246 known species of urban microorganisms and a consistent set of 31 species found in 97% of samples that were distinct from human commensal organisms. Profiles of AMR genes varied widely in type and density across cities. Cities showed distinct microbial taxonomic signatures that were driven by climate and geographic differences. These results constitute a high-resolution global metagenomic atlas that enables discovery of organisms and genes, highlights potential public health and forensic applications, and provides a culture-independent view of AMR burden in cities.

Haplotype diversity and sequence heterogeneity of human telomeres.

Telomeres are regions of repetitive nucleotide sequences capping the ends of eukaryotic chromosomes that protect against deterioration, and whose lengths can be correlated with age and adverse health risk factors. Yet, given their length and repetitive nature, telomeric regions are not easily reconstructed from short-read sequencing, thus making telomere sequencing, mapping, and variant resolution challenging problems. Recently, long-read sequencing, with read lengths measuring in hundreds of kilobase pairs, has made it possible to routinely read into telomeric regions and inspect their sequence structure. Here, we describe a framework for extracting telomeric reads from whole-genome single-molecule sequencing experiments, including de novo identification of telomere repeat motifs and repeat types, and also describe their sequence variation. We find that long, complex telomeric stretches and repeats can be accurately captured with long-read sequencing, observe extensive sequence heterogeneity of human telomeres, discover and localize noncanonical telomere sequence motifs (both previously reported, as well as novel), and validate them in short-read sequence data. These data reveal extensive intra- and inter-population diversity of repeats in telomeric haplotypes, reveal higher paternal inheritance of telomeric variants, and represent the first motif composition maps of multi-kilobase-pair human telomeric haplotypes across three distinct ancestries (Ashkenazi, Chinese, and Utah), which can aid in future studies of genetic variation, aging, and genome biology.

Chromatin-wide profiling of DYRK1A reveals a role as a gene-specific RNA polymerase II CTD kinase.

DYRK1A is a dosage-sensitive protein kinase that fulfills key roles during development and in tissue homeostasis, and its dysregulation results in human pathologies. DYRK1A is present in both the nucleus and cytoplasm of mammalian cells, although its nuclear function remains unclear. Genome-wide analysis of DYRK1A-associated loci reveals that the kinase is recruited preferentially to promoters of genes actively transcribed by RNA polymerase II (RNAPII), which are functionally associated with translation, RNA processing, and cell cycle. DYRK1A-bound promoter sequences are highly enriched in a conserved palindromic motif, which is necessary to drive DYRK1A-dependent transcriptional activation. DYRK1A phosphorylates the C-terminal domain (CTD) of RNAPII at Ser2 and Ser5. Depletion of DYRK1A results in reduced association of RNAPII at the target promoters as well as hypophosphorylation of the RNAPII CTD along the target gene bodies. These results are consistent with DYRK1A being a transcriptional regulator by acting as a CTD kinase.

Minerva: an alignment- and reference-free approach to deconvolve Linked-Reads for metagenomics.

Emerging Linked-Read technologies (aka read cloud or barcoded short-reads) have revived interest in short-read technology as a viable approach to understand large-scale structures in genomes and metagenomes. Linked-Read technologies, such as the 10x Chromium system, use a microfluidic system and a specialized set of 3' barcodes (aka UIDs) to tag short DNA reads sourced from the same long fragment of DNA; subsequently, the tagged reads are sequenced on standard short-read platforms. This approach results in interesting compromises. Each long fragment of DNA is only sparsely covered by reads, no information about the ordering of reads from the same fragment is preserved, and 3' barcodes match reads from roughly 2-20 long fragments of DNA. However, compared to long-read technologies, the cost per base to sequence is far lower, far less input DNA is required, and the per base error rate is that of Illumina short-reads. In this paper, we formally describe a particular algorithmic issue common to Linked-Read technology: the deconvolution of reads with a single 3' barcode into clusters that represent single long fragments of DNA. We introduce Minerva, a graph-based algorithm that approximately solves the barcode deconvolution problem for metagenomic data (where reference genomes may be incomplete or unavailable). Additionally, we develop two demonstrations where the deconvolution of barcoded reads improves downstream results, improving the specificity of taxonomic assignments and of k-mer-based clustering. To the best of our knowledge, we are the first to address the problem of barcode deconvolution in metagenomics.

Annotating unknown species of urban microorganisms on a global scale unveils novel functional diversity and local environment association.

In urban ecosystems, microbes play a key role in maintaining major ecological functions that directly support human health and city life. However, the knowledge about the species composition and functions involved in urban environments is still limited, which is largely due to the lack of reference genomes in metagenomic studies comprises more than half of unclassified reads. Here we uncovered 732 novel bacterial species from 4728 samples collected from various common surface with the matching materials in the mass transit system across 60 cities by the MetaSUB Consortium. The number of novel species is significantly and positively correlated with the city population, and more novel species can be identified in the skin-associated samples. The in-depth analysis of the new gene catalog showed that the functional terms have a significant geographical distinguishability. Moreover, we revealed that more biosynthetic gene clusters (BGCs) can be found in novel species. The co-occurrence relationship between BGCs and genera and the geographical specificity of BGCs can also provide us more information for the synthesis pathways of natural products. Expanded the known urban microbiome diversity and suggested additional mechanisms for taxonomic and functional characterization of the urban microbiome. Considering the great impact of urban microbiomes on human life, our study can also facilitate the microbial interaction analysis between human and urban environment.

Description of Citrobacter cronae sp. nov., isolated from human rectal swabs and stool samples.

Nine independent Gram-negative bacterial strains were isolated from rectal swabs or stool samples of immunocompromised patients from two different wards of a university hospital. All isolates were phylogenetically analysed based on their 16S rRNA gene sequence, housekeeping gene recN, multilocus sequence analysis of concatenated partial fusA, leuS, pyrG and rpoB sequences, and by whole genome sequencing data. The analysed strains of the new species cluster together and form a separate branch with Citrobacter werkmanii NBRC105721T as the most closely related species. An average nucleotide identity value of 95.9-96% and computation of digital DNA-DNA hybridization values separate the new species from all other type strains of the genus Citrobacter. Biochemical characteristics further delimit the isolates from closely related Citrobacter type strains. As a result of the described data, a new Citrobacter species is introduced, for which the name Citrobacter cronae sp. nov. is proposed. The type strain is Tue2-1T with a G+C DNA content of 52.2 mol%.

eDiVA-Classification and prioritization of pathogenic variants for clinical diagnostics.

Mendelian diseases have shown to be an and efficient model for connecting genotypes to phenotypes and for elucidating the function of genes. Whole-exome sequencing (WES) accelerated the study of rare Mendelian diseases in families, allowing for directly pinpointing rare causal mutations in genic regions without the need for linkage analysis. However, the low diagnostic rates of 20-30% reported for multiple WES disease studies point to the need for improved variant pathogenicity classification and causal variant prioritization methods. Here, we present the exome Disease Variant Analysis (eDiVA; http://ediva.crg.eu), an automated computational framework for identification of causal genetic variants (coding/splicing single-nucleotide variants and small insertions and deletions) for rare diseases using WES of families or parent-child trios. eDiVA combines next-generation sequencing data analysis, comprehensive functional annotation, and causal variant prioritization optimized for familial genetic disease studies. eDiVA features a machine learning-based variant pathogenicity predictor combining various genomic and evolutionary signatures. Clinical information, such as disease phenotype or mode of inheritance, is incorporated to improve the precision of the prioritization algorithm. Benchmarking against state-of-the-art competitors demonstrates that eDiVA consistently performed as a good or better than existing approach in terms of detection rate and precision. Moreover, we applied eDiVA to several familial disease cases to demonstrate its clinical applicability.

Chromosome-level assembly of Arabidopsis thaliana Ler reveals the extent of translocation and inversion polymorphisms.

Resequencing or reference-based assemblies reveal large parts of the small-scale sequence variation. However, they typically fail to separate such local variation into colinear and rearranged variation, because they usually do not recover the complement of large-scale rearrangements, including transpositions and inversions. Besides the availability of hundreds of genomes of diverse Arabidopsis thaliana accessions, there is so far only one full-length assembled genome: the reference sequence. We have assembled 117 Mb of the A. thaliana Landsberg erecta (Ler) genome into five chromosome-equivalent sequences using a combination of short Illumina reads, long PacBio reads, and linkage information. Whole-genome comparison against the reference sequence revealed 564 transpositions and 47 inversions comprising ∼3.6 Mb, in addition to 4.1 Mb of nonreference sequence, mostly originating from duplications. Although rearranged regions are not different in local divergence from colinear regions, they are drastically depleted for meiotic recombination in heterozygotes. Using a 1.2-Mb inversion as an example, we show that such rearrangement-mediated reduction of meiotic recombination can lead to genetically isolated haplotypes in the worldwide population of A. thaliana Moreover, we found 105 single-copy genes, which were only present in the reference sequence or the Ler assembly, and 334 single-copy orthologs, which showed an additional copy in only one of the genomes. To our knowledge, this work gives first insights into the degree and type of variation, which will be revealed once complete assemblies will replace resequencing or other reference-dependent methods.

Multi-drug resistant Enterobacter bugandensis species isolated from the International Space Station and comparative genomic analyses with human pathogenic strains.

BACKGROUND: The antimicrobial resistance (AMR) phenotypic properties, multiple drug resistance (MDR) gene profiles, and genes related to potential virulence and pathogenic properties of five Enterobacter bugandensis strains isolated from the International Space Station (ISS) were carried out and compared with genomes of three clinical strains. Whole genome sequences of ISS strains were characterized using the hybrid de novo assembly of Nanopore and Illumina reads. In addition to traditional microbial taxonomic approaches, multilocus sequence typing (MLST) analysis was performed to classify the phylogenetic lineage. Agar diffusion discs assay was performed to test antibiotics susceptibility. The draft genomes after assembly and scaffolding were annotated with the Rapid Annotations using Subsystems Technology and RNAmmer servers for downstream analysis. RESULTS: Molecular phylogeny and whole genome analysis of the ISS strains with all publicly available Enterobacter genomes revealed that ISS strains were E. bugandensis and similar to the type strain EB-247T and two clinical isolates (153_ECLO and MBRL 1077). Comparative genomic analyses of all eight E. bungandensis strains showed, a total of 4733 genes were associated with carbohydrate metabolism (635 genes), amino acid and derivatives (496 genes), protein metabolism (291 genes), cofactors, vitamins, prosthetic groups, pigments (275 genes), membrane transport (247 genes), and RNA metabolism (239 genes). In addition, 112 genes identified in the ISS strains were involved in virulence, disease, and defense. Genes associated with resistance to antibiotics and toxic compounds, including the MDR tripartite system were also identified in the ISS strains. A multiple antibiotic resistance (MAR) locus or MAR operon encoding MarA, MarB, MarC, and MarR, which regulate more than 60 genes, including upregulation of drug efflux systems that have been reported in Escherichia coli K12, was also observed in the ISS strains. CONCLUSION: Given the MDR results for these ISS Enterobacter genomes and increased chance of pathogenicity (PathogenFinder algorithm with > 79% probability), these species pose important health considerations for future missions. Thorough genomic characterization of the strains isolated from ISS can help to understand the pathogenic potential, and inform future missions, but analyzing them in in-vivo systems is required to discern the influence of microgravity on their pathogenicity.

Genomic characterisation of clinical and environmental Pseudomonas putida group strains and determination of their role in the transfer of antimicrobial resistance genes to Pseudomonas aeruginosa.

BACKGROUND: Pseudomonas putida is a Gram-negative, non-fermenting bacterium frequently encountered in various environmental niches. P. putida rarely causes disease in humans, though serious infections and outbreaks have been reported from time to time. Some have suggested that P. putida functions as an exchange platform for antibiotic resistance genes (ARG), and thus represents a serious concern in the spread of ARGs to more pathogenic organisms within a hospital. Though poorly understood, the frequency of ARG exchange between P. putida and the more virulent Pseudomonas aeruginosa and its clinical relevance are particularly important for designing efficient infection control strategies, such as deciding whether high-risk patients colonized with a multidrug resistant but typically low pathogenic P. putida strain should be contact isolated or not. RESULTS: In this study, 21,373 screening samples (stool, rectal and throat swab) were examined to determine the presence of P. putida in a high-risk group of haemato-oncology patients during a 28-month period. A total of 89 P. putida group strains were isolated from 85 patients, with 41 of 89 (46.1%) strains harbouring the metallo-beta-lactamase gene bla VIM. These 41 clinical isolates, plus 18 bla VIM positive environmental P. putida isolates, and 17 bla VIM positive P. aeruginosa isolates, were characterized by whole genome sequencing (WGS). We constructed a maximum-likelihood tree to separate the 59 bla VIM positive P. putida group strains into eight distinct phylogenetic clusters. Bla VIM-1 was present in 6 clusters while bla VIM-2 was detected in 4 clusters. Five P. putida group strains contained both, bla VIM-1 and bla VIM-2 genes. In contrast, all P. aeruginosa strains belonged to a single genetic cluster and contained the same ARGs. Apart from bla VIM-2 and sul genes, no other ARGs were shared between P. aeruginosa and P. putida. Furthermore, the bla VIM-2 gene in P. aeruginosa was predicted to be only chromosomally located. CONCLUSION: These data provide evidence that no exchange of comprehensive ARG harbouring mobile genetic elements had occurred between P. aeruginosa and P. putida group strains during the study period, thus eliminating the need to implement enhanced infection control measures for high-risk patients colonized with a bla VIM positiv P. putida group strains in our clinical setting.

The cis-regulatory code of Hox function in Drosophila.

Precise gene expression is a fundamental aspect of organismal function and depends on the combinatorial interplay of transcription factors (TFs) with cis-regulatory DNA elements. While much is known about TF function in general, our understanding of their cell type-specific activities is still poor. To address how widely expressed transcriptional regulators modulate downstream gene activity with high cellular specificity, we have identified binding regions for the Hox TF Deformed (Dfd) in the Drosophila genome. Our analysis of architectural features within Hox cis-regulatory response elements (HREs) shows that HRE structure is essential for cell type-specific gene expression. We also find that Dfd and Ultrabithorax (Ubx), another Hox TF specifying different morphological traits, interact with non-overlapping regions in vivo, despite their similar DNA binding preferences. While Dfd and Ubx HREs exhibit comparable design principles, their motif compositions and motif-pair associations are distinct, explaining the highly selective interaction of these Hox proteins with the regulatory environment. Thus, our results uncover the regulatory code imprinted in Hox enhancers and elucidate the mechanisms underlying functional specificity of TFs in vivo.

Analysis of a long-term outbreak of XDR Pseudomonas aeruginosa: a molecular epidemiological study.

OBJECTIVES: Here we report on a long-term outbreak from 2009 to 2012 with an XDR Pseudomonas aeruginosa on two wards at a university hospital in southern Germany. METHODS: Whole-genome sequencing was performed on the outbreak isolates and a core genome was constructed for molecular epidemiological analysis. We applied a time-place-sequence algorithm to improve estimation of transmission probabilities. RESULTS: By using conventional infection control methods we identified 49 P. aeruginosa strains, including eight environmental isolates that belonged to ST308 (by MLST) and carried the metallo-ß-lactamase IMP-8. Phylogenetic analysis on the basis of a non-recombinant core genome that contained 22 outbreak-specific SNPs revealed a pattern of four dominant clades with a strong phylogeographic structure and allowed us to determine the potential temporal origin of the outbreak to July 2008, 1 year before the index case was diagnosed. Superspreaders at the root of clades exhibited a high number of probable and predicted transmissions, indicating their exceptional position in the outbreak. CONCLUSIONS: Our results suggest that the initial expansion of dominant sublineages was driven by a few superspreaders, while environmental contamination seemed to sustain the outbreak for a long period despite regular environmental control measures.

Antagonistic regulation of apoptosis and differentiation by the Cut transcription factor represents a tumor-suppressing mechanism in Drosophila.

Apoptosis is essential to prevent oncogenic transformation by triggering self-destruction of harmful cells, including those unable to differentiate. However, the mechanisms linking impaired cell differentiation and apoptosis during development and disease are not well understood. Here we report that the Drosophila transcription factor Cut coordinately controls differentiation and repression of apoptosis via direct regulation of the pro-apoptotic gene reaper. We also demonstrate that this regulatory circuit acts in diverse cell lineages to remove uncommitted precursor cells in status nascendi and thereby interferes with their potential to develop into cancer cells. Consistent with the role of Cut homologues in controlling cell death in vertebrates, we find repression of apoptosis regulators by Cux1 in human cancer cells. Finally, we present evidence that suggests that other lineage-restricted specification factors employ a similar mechanism to put the brakes on the oncogenic process.

Astronaut omics and the impact of space on the human body at scale.

Future multi-year crewed planetary missions will motivate advances in aerospace nutrition and telehealth. On Earth, the Human Cell Atlas project aims to spatially map all cell types in the human body. Here, we propose that a parallel Human Cell Space Atlas could serve as an openly available, global resource for space life science research. As humanity becomes increasingly spacefaring, high-resolution omics on orbit could permit an advent of precision spaceflight healthcare. Alongside the scientific potential, we consider the complex ethical, cultural, and legal challenges intrinsic to the human space omics discipline, and how philosophical frameworks may benefit from international perspectives.

Protective alleles and precision healthcare in crewed spaceflight.

Common and rare alleles are now being annotated across millions of human genomes, and omics technologies are increasingly being used to develop health and treatment recommendations. However, these alleles have not yet been systematically characterized relative to aerospace medicine. Here, we review published alleles naturally found in human cohorts that have a likely protective effect, which is linked to decreased cancer risk and improved bone, muscular, and cardiovascular health. Although some technical and ethical challenges remain, research into these protective mechanisms could translate into improved nutrition, exercise, and health recommendations for crew members during deep space missions.

Enhancing European capabilities for application of multi-omics studies in biology and biomedicine space research.

Following on from the NASA twins' study, there has been a tremendous interest in the use of omics techniques in spaceflight. Individual space agencies, NASA's GeneLab, JAXA's ibSLS, and the ESA-funded Space Omics Topical Team and the International Standards for Space Omics Processing (ISSOP) groups have established several initiatives to support this growth. Here, we present recommendations from the Space Omics Topical Team to promote standard application of space omics in Europe. We focus on four main themes: i) continued participation in and coordination with international omics endeavors, ii) strengthening of the European space omics infrastructure including workforce and facilities, iii) capitalizing on the emerging opportunities in the commercial space sector, and iv) capitalizing on the emerging opportunities in human subjects research.

Multifactorial regulation of a hox target gene.

Hox proteins play fundamental roles in controlling morphogenetic diversity along the anterior-posterior body axis of animals by regulating distinct sets of target genes. Within their rather broad expression domains, individual Hox proteins control cell diversification and pattern formation and consequently target gene expression in a highly localized manner, sometimes even only in a single cell. To achieve this high-regulatory specificity, it has been postulated that Hox proteins co-operate with other transcription factors to activate or repress their target genes in a highly context-specific manner in vivo. However, only a few of these factors have been identified. Here, we analyze the regulation of the cell death gene reaper (rpr) by the Hox protein Deformed (Dfd) and suggest that local activation of rpr expression in the anterior part of the maxillary segment is achieved through a combinatorial interaction of Dfd with at least eight functionally diverse transcriptional regulators on a minimal enhancer. It follows that context-dependent combinations of Hox proteins and other transcription factors on small, modular Hox response elements (HREs) could be responsible for the proper spatio-temporal expression of Hox targets. Thus, a large number of transcription factors are likely to be directly involved in Hox target gene regulation in vivo.

Building the Space Omics Topical Team to boost European space researchers' role in the international consortia redefining spaceflight-generated datasets.

In a broadening and more competitive space exploration landscape, playing at scale is necessary to obtain results. European researchers share their lessons learned on growing a research program where omics techniques can feed new knowledge, both fundamental and practical, for space exploration. Sending people to new space destinations will require interdisciplinary research centered around omics and personalized medicine, with added constraints of low-gravity and high-radiation environments.

Ozone Disinfection for Elimination of Bacteria and Degradation of SARS-CoV2 RNA for Medical Environments.

Pathogenic bacteria and viruses in medical environments can lead to treatment complications and hospital-acquired infections. Current disinfection protocols do not address hard-to-access areas or may be beyond line-of-sight treatment, such as with ultraviolet radiation. The COVID-19 pandemic further underscores the demand for reliable and effective disinfection methods to sterilize a wide array of surfaces and to keep up with the supply of personal protective equipment (PPE). We tested the efficacy of Sani Sport ozone devices to treat hospital equipment and surfaces for killing Escherichia coli, Enterococcus faecalis, Bacillus subtilis, and Deinococcus radiodurans by assessing Colony Forming Units (CFUs) after 30 min, 1 h, and 2 h of ozone treatment. Further gene expression analysis was conducted on live E. coli K12 immediately post treatment to understand the oxidative damage stress response transcriptome profile. Ozone treatment was also used to degrade synthetic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA as assessed by qPCR CT values. We observed significant and rapid killing of medically relevant and environmental bacteria across four surfaces (blankets, catheter, remotes, and syringes) within 30 min, and up to a 99% reduction in viable bacteria at the end of 2 h treatment cycles. RNA-seq analysis of E. coli K12 revealed 447 differentially expressed genes in response to ozone treatment and an enrichment for oxidative stress response and related pathways. RNA degradation of synthetic SARS-CoV-2 RNA was seen an hour into ozone treatment as compared to non-treated controls, and a non-replicative form of the virus was shown to have significant RNA degradation at 30 min. These results show the strong promise of ozone treatment of surfaces for reducing the risk of hospital-acquired infections and as a method for degradation of SARS-CoV-2 RNA.