Targeted sequencing of bacteria using CRISPR-Cas9 enrichment and Oxford Nanopore Technologies.

Unlabelled: Sequencing DNA directly from patient samples enables faster pathogen characterization compared to traditional culture-based approaches, but often yields insufficient sequence data for effective downstream analysis. CRISPR-Cas9 enrichment is designed to improve the yield of low abundance sequences but has not been thoroughly explored with Oxford Nanopore Technologies (ONT) for use in clinical bacterial epidemiology. We designed CRISPR-Cas9 guide RNAs to enrich the human pathogen , by targeting multi-locus sequence type (MLST) and transfer RNA (tRNA) genes, as well as common antimicrobial resistance (AMR) genes and the resistance-associated integron gene .

Quantifying NO Emission Sources in Houston, Texas Using Remote Sensing Aircraft Measurements and Source Apportionment Regression Models.

Air quality managers in areas exceeding air pollution standards are motivated to understand where there are further opportunities to reduce NO emissions to improve ozone and PM air quality. In this project, we use a combination of aircraft remote sensing (i.e.

Retinoic acid and TGF-β orchestrate organ-specific programs of tissue residency.

Tissue-resident memory T (T) cells are integral to tissue immunity, persisting in diverse anatomical sites where they adhere to a common transcriptional framework. How these cells integrate distinct local cues to adopt the common T cell fate remains poorly understood. Here, we show that whereas skin T cells strictly require transforming growth factor β (TGF-β) for tissue residency, those in other locations utilize the metabolite retinoic acid (RA) to drive an alternative differentiation pathway, directing a TGF-β-independent tissue residency program in the liver and synergizing with TGF-β to drive T cells in the small intestine.

Neighborhood-Level Nitrogen Dioxide Inequalities Contribute to Surface Ozone Variability in Houston, Texas.

In Houston, Texas, nitrogen dioxide (NO) air pollution disproportionately affects Black, Latinx, and Asian communities, and high ozone (O) days are frequent. There is limited knowledge of how NO inequalities vary in urban air quality contexts, in part from the lack of time-varying neighborhood-level NO measurements. First, we demonstrate that daily TROPOspheric Monitoring Instrument (TROPOMI) NO tropospheric vertical column densities (TVCDs) resolve a major portion of census tract-scale NO inequalities in Houston, comparing NO inequalities based on TROPOMI TVCDs and spatiotemporally coincident airborne remote sensing (250 m × 560 m) from the NASA TRacking Aerosol Convection ExpeRiment-Air Quality (TRACER-AQ).

Chlamydiae as symbionts of photosynthetic dinoflagellates.

Chlamydiae are ubiquitous intracellular bacteria and infect a wide diversity of eukaryotes, including mammals. However, chlamydiae have never been reported to infect photosynthetic organisms. Here, we describe a novel chlamydial genus and species, Candidatus Algichlamydia australiensis, capable of infecting the photosynthetic dinoflagellate Cladocopium sp.

How low can you go? Short-read polishing of Oxford Nanopore bacterial genome assemblies.

It is now possible to assemble near-perfect bacterial genomes using Oxford Nanopore Technologies (ONT) long reads, but short-read polishing is usually required for perfection. However, the effect of short-read depth on polishing performance is not well understood. Here, we introduce Pypolca (with default and careful parameters) and Polypolish v0.

Hybracter: enabling scalable, automated, complete and accurate bacterial genome assemblies.

Improvements in the accuracy and availability of long-read sequencing mean that complete bacterial genomes are now routinely reconstructed using hybrid (i.e. short- and long-reads) assembly approaches.

Molecular characterization and descriptive analysis of carbapenemase-producing Gram-negative rod infections in Bogota, Colombia.

In this study, the genetic differences and clinical impact of the carbapenemase-encoding genes among the community and healthcare-acquired infections were assessed. This retrospective, multicenter cohort study was conducted in Colombia and included patients infected with carbapenem-resistant Gram-negative rods between 2017 and 2021. Carbapenem resistance was identified by Vitek, and carbapenemase-encoding genes were identified by whole-genome sequencing (WGS) to classify the alleles and sequence types (STs).

Tissue-associated and vertically transmitted bacterial symbiont in the coral Pocillopora acuta.

Coral microhabitats are colonized by a myriad of microorganisms, including diverse bacteria which are essential for host functioning and survival. However, the location, transmission, and functions of individual bacterial species living inside the coral tissues remain poorly studied. Here, we show that a previously undescribed bacterial symbiont of the coral Pocillopora acuta forms cell-associated microbial aggregates (CAMAs) within the mesenterial filaments.

Mosquitoes provide a transmission route between possums and humans for Buruli ulcer in southeastern Australia.

Buruli ulcer, a chronic subcutaneous infection caused by Mycobacterium ulcerans, is increasing in prevalence in southeastern Australia. Possums are a local wildlife reservoir for M. ulcerans and, although mosquitoes have been implicated in transmission, it remains unclear how humans acquire infection.

Hybracter: Enabling Scalable, Automated, Complete and Accurate Bacterial Genome Assemblies.

Improvements in the accuracy and availability of long-read sequencing mean that complete bacterial genomes are now routinely reconstructed using hybrid (i.e. short- and long-reads) assembly approaches.

High-level nitrofurantoin resistance in a clinical isolate of a comparative genomics and metabolomics analysis.

Nitrofurantoin is a commonly used chemotherapeutic agent in the treatment of uncomplicated urinary tract infections caused by the problematic multidrug resistant Gram-negative pathogen . The present study aims to elucidate the mechanism of nitrofurantoin action and high-level resistance in using whole-genome sequencing (WGS), qPCR analysis, mutation structural modeling and untargeted metabolomic analysis. WGS profiling of evolved highly resistant mutants (nitrofurantoin minimum inhibitory concentrations > 256 mg/L) revealed modified expression of several genes related to membrane transport (porin and efflux pump regulator ) and nitroreductase activity ( and , involved in nitrofurantoin reduction).

High-performance enrichment-based genome sequencing to support the investigation of hepatitis A virus outbreaks.

This proof-of-concept study introduces a hybrid capture oligo panel for whole-genome sequencing of all six human pathogenic hepatitis A virus (HAV) subgenotypes, exhibiting a higher sensitivity than some conventional genotyping assays. The ability of hybrid capture to enrich multiple targets allows for a single, streamlined workflow, thus facilitating the potential harmonization of molecular surveillance of HAV with other enteric viruses. Even challenging sample matrices can be accommodated, making them suitable for broad implementation in clinical and public health laboratories.

Functional potential and evolutionary response to long-term heat selection of bacterial associates of coral photosymbionts.

Symbiotic microorganisms are crucial for the survival of corals and their resistance to coral bleaching in the face of climate change. However, the impact of microbe-microbe interactions on coral functioning is mostly unknown but could be essential factors for coral adaption to future climates. Here, we investigated interactions between cultured dinoflagellates of the Symbiodiniaceae family, essential photosymbionts of corals, and associated bacteria.

Bactabolize is a tool for high-throughput generation of bacterial strain-specific metabolic models.

Metabolic capacity can vary substantially within a bacterial species, leading to ecological niche separation, as well as differences in virulence and antimicrobial susceptibility. Genome-scale metabolic models are useful tools for studying the metabolic potential of individuals, and with the rapid expansion of genomic sequencing there is a wealth of data that can be leveraged for comparative analysis. However, there exist few tools to construct strain-specific metabolic models at scale.

Photochemical model assessment of single source NO and O plumes using field study data.

Single source contribution to ambient O and PM has been estimated with photochemical grid models to support policy demonstrations for National Ambient Air Quality Standards, regional haze, and permit related programs. Limited field data exists to evaluate model representation of the spatial extent and chemical composition of plumes emitted by specific facilities. New tropospheric column measurements of NO and in-plume chemical measurements downwind of specific facilities allows for photochemical model evaluation of downwind plume extent, grid resolution impacts on plume concentration gradients, and source attribution methods.

Direct inference and control of genetic population structure from RNA sequencing data.

RNAseq data can be used to infer genetic variants, yet its use for estimating genetic population structure remains underexplored. Here, we construct a freely available computational tool (RGStraP) to estimate RNAseq-based genetic principal components (RG-PCs) and assess whether RG-PCs can be used to control for population structure in gene expression analyses. Using whole blood samples from understudied Nepalese populations and the Geuvadis study, we show that RG-PCs had comparable results to paired array-based genotypes, with high genotype concordance and high correlations of genetic principal components, capturing subpopulations within the dataset.

Discovery and Biosynthesis of the Cytotoxic Polyene Terpenomycin in Human Pathogenic .

are opportunistic human pathogens that can cause a range of debilitating and difficult to treat infections of the lungs, brain, skin, and soft tissues. Despite their close relationship to the well-known secondary metabolite-producing genus, , comparatively few natural products are known from the , and even less is known about their involvement in the pathogenesis. Here, we combine chemistry, genomics, and molecular microbiology to reveal the production of terpenomycin, a new cytotoxic and antifungal polyene from a human pathogenic isolate.

Intimately tracking NO pollution over the New York City - Long Island Sound land-water continuum: An integration of shipboard, airborne, satellite observations, and models.

Nitrogen dioxide (NO) pollution remains a serious global problem, particularly near highly populated urbanized coasts that face increasing challenges with climate change. Yet, the combined impact of urban emissions, pollution transport, and complex meteorology on the spatiotemporal dynamics of NO along heterogeneous urban coastlines remains poorly characterized. Here, we integrated measurements from different platforms - boats, ground-based networks, aircraft, and satellites - to characterize total column NO (TCNO) dynamics across the land-water continuum in the New York metropolitan area, the most populous area in the United States that often experiences the highest national NO levels.

Smoking Cessation Interventions and Abstinence Outcomes for People Living in Rural, Regional, and Remote Areas of Three High-Income Countries: A Systematic Review.

Introduction: Tobacco smoking rates in high-income countries are greater in rural, regional, and remote (RRR) areas compared to cities. Yet, there is limited knowledge about interventions targeted to RRR smokers. This review describes the effectiveness of smoking cessation interventions for RRR smokers in supporting smoking abstinence.

Assembling the perfect bacterial genome using Oxford Nanopore and Illumina sequencing.

A perfect bacterial genome assembly is one where the assembled sequence is an exact match for the organism's genome-each replicon sequence is complete and contains no errors. While this has been difficult to achieve in the past, improvements in long-read sequencing, assemblers, and polishers have brought perfect assemblies within reach. Here, we describe our recommended approach for assembling a bacterial genome to perfection using a combination of Oxford Nanopore Technologies long reads and Illumina short reads: Trycycler long-read assembly, Medaka long-read polishing, Polypolish short-read polishing, followed by other short-read polishing tools and manual curation.