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Abstract Low-abundance members of microbial communities are difficult to study in their native habitats, including Escherichia coli, a minor but common inhabitant of the gastrointestinal tract, and key opportunistic pathogen of the urinary tract. While multi-omic analyses have detailed interactions between uropathogenic Escherichia coli (UPEC) and the bladder mediating urinary tract infection (UTI), little is known about UPEC in its pre-infection reservoir, the gastrointestinal tract, partly due to its low relative abundance (

Abstract In this study, the concept of rheo-optics was applied to explore the flow birefringence induced by the stress components along the camera’s optical axis since it is often overlooked in the traditional theories of photoelastic flow measurement. A novel aspect of this research is that it involved conducting polarization measurements on simple shear flows, specifically from a perspective in which a shear-velocity gradient exists along the camera’s optical axis. A parallel-plate-type rheometer and a polarization camera are employed for these systematic measurements. The experimental findings for dilute aqueous cellulose nanocrystal suspensions demonstrate that the flow birefringence can be expressed as a power law based on the power of the second invariant of the deformation-rate tensor. This suggests that flow birefringence can be universally characterized by the coordinate-independent invariants and a pre-factor determined by the direction of polarization measurement. By adjusting the nonlinear term in the stress-optic law, its applicability could be expanded to include three-dimensional fluid stress fields in which the stress is distributed along the camera’s optical axis.

Human-associated microbial communities comprise not only complex mixtures of bacterial species, but also mixtures of conspecific strains, the implications of which are mostly unknown since strain level dynamics are underexplored due to the difficulties of studying them. We introduce the Strain Genome Explorer (StrainGE) toolkit, which deconvolves strain mixtures and characterizes component strains at the nucleotide level from short-read metagenomic sequencing with higher sensitivity and resolution than other tools. StrainGE is able to identify strains at 0.1x coverage and detect variants for multiple conspecific strains within a sample from coverages as low as 0.5x.

BackgroundCarbapenem-resistant Enterobacterales (CRE) are an urgent global health threat. Inferring the dynamics of local CRE dissemination is currently limited by our inability to confidently trace the spread of resistance determinants to unrelated bacterial hosts. Whole-genome sequence comparison is useful for identifying CRE clonal transmission and outbreaks, but high-frequency horizontal gene transfer (HGT) of carbapenem resistance genes and subsequent genome rearrangement complicate tracing the local persistence and mobilization of these genes across organisms.MethodsTo overcome this limitation, we developed a new approach to identify recent HGT of large, near-identical plasmid segments across species boundaries, which also allowed us to overcome technical challenges with genome assembly. We applied this to complete and near-complete genome assemblies to examine the local spread of CRE in a systematic, prospective collection of all CRE, as well as time- and species-matched carbapenem-susceptible Enterobacterales, isolated from patients from four US hospitals over nearly 5 years.ResultsOur CRE collection comprised a diverse range of species, lineages, and carbapenem resistance mechanisms, many of which were encoded on a variety of promiscuous plasmid types. We found and quantified rearrangement, persistence, and repeated transfer of plasmid segments, including those harboring carbapenemases, between organisms over multiple years. Some plasmid segments were found to be strongly associated with specific locales, thus representing geographic signatures that make it possible to trace recent and localized HGT events. Functional analysis of these signatures revealed genes commonly found in plasmids of nosocomial pathogens, such as functions required for plasmid retention and spread, as well survival against a variety of antibiotic and antiseptics common to the hospital environment.ConclusionsCollectively, the framework we developed provides a clearer, high-resolution picture of the epidemiology of antibiotic resistance importation, spread, and persistence in patients and healthcare networks.

Healthy development of the gut microbiome provides long-term health benefits. Children raised in countries with high infectious disease burdens are frequently exposed to diarrhoeal pathogens and antibiotics, which perturb gut microbiome assembly. A recent cluster-randomized trial leveraging >4,000 child observations in Dhaka, Bangladesh, found that automated water chlorination of shared taps effectively reduced child diarrhoea and antibiotic use. In this substudy, we leveraged stool samples collected from 130 children 1 year after chlorine doser installation to examine differences between treatment and control children's gut microbiota. Water chlorination was associated with increased abundance of several bacterial genera previously linked to improved gut health; however, we observed no effects on the overall richness or diversity of taxa. Several clinically relevant antibiotic resistance genes were relatively more abundant in the gut microbiome of treatment children, possibly due to increases in Enterobacteriaceae. While further studies on the long-term health impacts of drinking chlorinated water would be valuable, we conclude that access to chlorinated water did not substantially impact child gut microbiome development in this setting, supporting the use of chlorination to increase global access to safe drinking water.

The study of extracellular phosphorylation was initiated in late 19th century when the secreted milk protein, casein, and egg-yolk protein, phosvitin, were shown to be phosphorylated. However, it took more than a century to identify Fam20C, which phosphorylates both casein and phosvitin under physiological conditions. This kinase, along with its family members Fam20A and Fam20B, defined a new family with altered amino acid sequences highly atypical from the canonical 540 kinases comprising the kinome. Fam20B is a glycan kinase that phosphorylates xylose residues and triggers peptidoglycan biosynthesis, a role conserved from sponges to human. The protein kinase, Fam20C, conserved from nematodes to humans, phosphorylates well over 100 substrates in the secretory pathway with overall functions postulated to encompass endoplasmic reticulum homeostasis, nutrition, cardiac function, coagulation, and biomineralization. The preferred phosphorylation motif of Fam20C is SxE/pS, and structural studies revealed that related member Fam20A allosterically activates Fam20C by forming a heterodimeric/tetrameric complex. Fam20A, a pseudokinase, is observed only in vertebrates. Loss-of-function genetic alterations in the Fam20 family lead to human diseases such as amelogenesis imperfecta, nephrocalcinosis, lethal and nonlethal forms of Raine syndrome with major skeletal defects, and altered phosphate homeostasis. Together, these three members of the Fam20 family modulate a diverse network of secretory pathway components playing crucial roles in health and disease. The overarching theme of this review is to highlight the progress that has been made in the emerging field of extracellular phosphorylation and the key roles secretory pathway kinases play in an ever-expanding number of cellular processes.

Lafora disease is a rare recessive form of progressive myoclonic epilepsy, usually diagnosed during adolescence. Patients present with myoclonus, neurological deterioration, and generalized tonic-clonic, myoclonic, or absence seizures. Symptoms worsen until death, usually within the first ten years of clinical onset. The primary histopathological hallmark is the formation of aberrant polyglucosan aggregates called Lafora bodies in the brain and other tissues. Lafora disease is caused by mutations in either the EPM2A gene, encoding laforin, or the EPM2B gene, coding for malin. The most frequent EPM2A mutation is R241X, which is also the most prevalent in Spain. The Epm2a‐/− and Epm2b−/− mouse models of Lafora disease show neuropathological and behavioral abnormalities similar to those seen in patients, although with a milder phenotype. To obtain a more accurate animal model, we generated the Epm2aR240X knock-in mouse line with the R240X mutation in the Epm2a gene, using genetic engineering based on CRISPR-Cas9 technology. Epm2aR240X mice exhibit most of the alterations reported in patients, including the presence of LBs, neurodegeneration, neuroinflammation, interictal spikes, neuronal hyperexcitability, and cognitive decline, despite the absence of motor impairments. The Epm2aR240X knock-in mouse displays some symptoms that are more severe that those observed in the Epm2a−/− knock-out, including earlier and more pronounced memory loss, increased levels of neuroinflammation, more interictal spikes and increased neuronal hyperexcitability, symptoms that more precisely resemble those observed in patients. This new mouse model can therefore be specifically used to evaluate how new therapies affects these features with greater precision.

Fam20C is a secreted protein kinase mutated in Raine syndrome, a human skeletal disorder. In vertebrates, bone and enamel proteins are major Fam20C substrates. However, Fam20 kinases are conserved in invertebrates lacking bone and enamel, suggesting other ancestral functions. We show that FAMK-1, the Caenorhabditis elegans Fam20C orthologue, contributes to fertility, embryogenesis, and development. These functions are not fulfilled when FAMK-1 is retained in the early secretory pathway. During embryogenesis, FAMK-1 maintains intercellular partitions and prevents multinucleation; notably, temperature elevation or lowering cortical stiffness reduces requirement for FAMK-1 in these contexts. FAMK-1 is expressed in multiple adult tissues that undergo repeated mechanical strain, and selective expression in the spermatheca restores fertility. Informatic, biochemical, and functional analysis implicate lectins as FAMK-1 substrates. These findings suggest that FAMK-1 phosphorylation of substrates, including lectins, in the late secretory pathway is important in embryonic and tissue contexts where cells are subjected to mechanical strain.

Abstract Background Carbapenem-resistant Enterobacterales (CRE) are an urgent global health threat. Inferring the dynamics of local CRE dissemination is currently limited by our inability to confidently trace the spread of resistance determinants to unrelated bacterial hosts. Whole-genome sequence comparison is useful for identifying CRE clonal transmission and outbreaks, but high-frequency horizontal gene transfer (HGT) of carbapenem resistance genes and subsequent genome rearrangement complicate tracing the local persistence and mobilization of these genes across organisms. Methods To overcome this limitation, we developed a new approach to identify recent HGT of large, near-identical plasmid segments across species boundaries, which also allowed us to overcome technical challenges with genome assembly. We applied this to complete and near-complete genome assemblies to examine the local spread of CRE in a systematic, prospective collection of all CRE, as well as time- and species-matched carbapenem-susceptible Enterobacterales, isolated from patients from four US hospitals over nearly 5 years. Results Our CRE collection comprised a diverse range of species, lineages, and carbapenem resistance mechanisms, many of which were encoded on a variety of promiscuous plasmid types. We found and quantified rearrangement, persistence, and repeated transfer of plasmid segments, including those harboring carbapenemases, between organisms over multiple years. Some plasmid segments were found to be strongly associated with specific locales, thus representing geographic signatures that make it possible to trace recent and localized HGT events. Functional analysis of these signatures revealed genes commonly found in plasmids of nosocomial pathogens, such as functions required for plasmid retention and spread, as well survival against a variety of antibiotic and antiseptics common to the hospital environment. Conclusions Collectively, the framework we developed provides a clearer, high-resolution picture of the epidemiology of antibiotic resistance importation, spread, and persistence in patients and healthcare networks.

Abstract Human-associated microbial communities comprise not only complex mixtures of bacterial species, but also mixtures of conspecific strains, the implications of which are mostly unknown since strain level dynamics are underexplored due to the difficulties of studying them. We introduce the Strain Genome Explorer (StrainGE) toolkit, which deconvolves strain mixtures and characterizes component strains at the nucleotide level from short-read metagenomic sequencing with higher sensitivity and resolution than other tools. StrainGE is able to identify strains at 0.1x coverage and detect variants for multiple conspecific strains within a sample from coverages as low as 0.5x.

As many malaria-endemic countries move towards elimination of Plasmodium falciparum, the most virulent human malaria parasite, effective tools for monitoring malaria epidemiology are urgent priorities. P. falciparum population genetic approaches offer promising tools for understanding transmission and spread of the disease, but a high prevalence of multi-clone or polygenomic infections can render estimation of even the most basic parameters, such as allele frequencies, challenging. A previous method, COIL, was developed to estimate complexity of infection (COI) from single nucleotide polymorphism (SNP) data, but relies on monogenomic infections to estimate allele frequencies or requires external allele frequency data which may not available. Estimates limited to monogenomic infections may not be representative, however, and when the average COI is high, they can be difficult or impossible to obtain. Therefore, we developed THE REAL McCOIL, Turning HEterozygous SNP data into Robust Estimates of ALelle frequency, via Markov chain Monte Carlo, and Complexity Of Infection using Likelihood, to incorporate polygenomic samples and simultaneously estimate allele frequency and COI. This approach was tested via simulations then applied to SNP data from cross-sectional surveys performed in three Ugandan sites with varying malaria transmission. We show that THE REAL McCOIL consistently outperforms COIL on simulated data, particularly when most infections are polygenomic. Using field data we show that, unlike with COIL, we can distinguish epidemiologically relevant differences in COI between and within these sites. Surprisingly, for example, we estimated high average COI in a peri-urban subregion with lower transmission intensity, suggesting that many of these cases were imported from surrounding regions with higher transmission intensity. THE REAL McCOIL therefore provides a robust tool for understanding the molecular epidemiology of malaria across transmission settings.

BackgroundMethicillin-resistant Staphylococcus aureus (MRSA) is one of the most common healthcare-associated pathogens. To examine the role of inter-hospital patient sharing on MRSA transmission, a previous study collected 2,214 samples from 30 hospitals in Orange County, California and showed by spa typing that genetic differentiation decreased significantly with increased patient sharing. In the current study, we focused on the 986 samples with spa type t008 from the same population.MethodsWe used genome sequencing to determine the effect of patient sharing on genetic differentiation between hospitals. Genetic differentiation was measured by between-hospital genetic diversity, F ST , and the proportion of nearly identical isolates between hospitals.ResultsSurprisingly, we found very similar genetic diversity within and between hospitals, and no significant association between patient sharing and genetic differentiation measured by F ST . However, in contrast to F ST , there was a significant association between patient sharing and the proportion of nearly identical isolates between hospitals. We propose that the proportion of nearly identical isolates is more powerful at determining transmission dynamics than traditional estimators of genetic differentiation (F ST ) when gene flow between populations is high, since it is more responsive to recent transmission events. Our hypothesis was supported by the results from coalescent simulations.ConclusionsOur results suggested that there was a high level of gene flow between hospitals facilitated by patient sharing, and that the proportion of nearly identical isolates is more sensitive to population structure than F ST when gene flow is high.

The classic model of non-contact ACL injury includes environmental, anatomical, hormonal and biomechanical risk factors which directly impact either the amount of stress placed on the ligament or the relative capacity of ligament to withstand the forces placed on it. However, cognition also clearly plays a role in successful athletic performance, yet diminished cognitive function is rarely considered a risk factor for injury. # Objective To examine the existing literature to determine the extent to which cognitive function (both cognitive ability and task cognitive load) influences non-contact lower extremity injury risk in male and female athletes with a broad variety of athletic expertise. # Study Design Scoping Review # Methods An electronic search was conducted of CINAHL, SPORTDiscus, Google Scholar, and MEDLINE using the PRISMA method. Search terms included Boolean combinations of “cognition”, “concussion”, “ImPACT”, “cognitive deficit”, “mild traumatic brain injury (mTBI)”, and “neuropsychological function” as cognitive descriptors and the terms “injury risk” and “lower extremity injury” as injury descriptors. Inclusion criteria included papers written in English published between 2000-2021. Exclusion criteria included neurological and cognitively atypical populations, except for concussion (included). Included articles were appraised using the Quality Assessment Tool for Observational Cohort and Cross-Sectional Studies. # Results Fifty-six studies utilizing across the spectrum of levels of evidence met inclusion criteria. Forty-one articles had good, fourteen had fair, and one had poor methodological quality. Studies examined baseline cognitive function in healthy athletes (n=7); performance during dual-task paradigms (n=13); and the impact of concussion on dual-task performance (n=4), LE injury risk (n=22), or post-concussion testing (n=10). Six articles examining cognitive function and all dual-task studies (including concussion studies) found altered biomechanics associated with injury or increased processing demands. Studies related to concussion and injury incidence consistently found an increased risk of LE injury following concussion. Half of the studies that examined concussion and post-concussion cognitive testing demonstrated significant effects. # Discussion Consistent across participant demographics, tasks, and dependent measures, fifty-one of fifty-six assessed articles concluded that decreased cognitive ability or increased cognitive load led to risky LE mechanics or a direct increase in non-contact LE injury risk. # Conclusion The robustness of results across gender, performance level, sport, cognitive ability, task cognitive load suggest that the inclusion of cognitive training in the design of optimal LE injury prevention programs warrants further study. Level of Evidence: Ia

Recommendations regarding the use of face masks as a preventive measure for COVID-19 are inconsistent. Here, the authors show that optimal distribution of surgical-standard face masks in the population, or universal coverage of homemade face coverings, could reduce total infections and deaths.

Undocumented Latino day laborers in the United States are vulnerable to being arrested and expelled at any time. This social fact shapes their everyday lives in terms of actions taken and strategies deployed to mitigate being confronted, profiled, and possibly incarcerated and deported. While perceptions of threat and bouts of discrimination are routine among undocumented Latino day laborers, their specific nature vary according to multiple social factors and structural forces that differ significantly from locale to locale. The experience of discrimination is often tacitly negotiated through perceptions, decisions, and actions toward avoiding or moderating its ill effects. This essay examines urban undocumented Latino day laborers over a variety of sites in the greater San Francisco Bay Area, which, compared to many metropolitan areas in the U.S. is as good as it gets in terms of being socially tolerated and relatively safe from persecution. Nonetheless, tacit negotiations are necessary to withstand or overcome challenges presented by idiosyncratic and ever changing global, national/state, and local dynamics of discrimination. [undocumented Latino laborers, social exclusion, discrimination, tacit negotiation].

Introduction The widespread use of antibiotics, in both healthcare and agriculture, has led to the emergence of antibiotic-resistant bacteria, decreasing our ability to effectively treat common infections. With predictions of [...]