Whole-genome sequence of Mucor velutinosus NIH1002, a strain associated with disseminated disease.

The draft genome of Mucor velutinosus NIH1002, a 2011 isolate from a case of disseminated disease, was sequenced using PacBio long-read and HiSeq short-read technologies. The genome has 43 contigs, an N50 of 2.65 Mb, and 13,295 protein-coding genes. It is the most complete M. velutinosus genome to date.

The Extent and Impact of Variation in ADME Genes in Sub-Saharan African Populations.

Introduction: Investigating variation in genes involved in the absorption, distribution, metabolism, and excretion (ADME) of drugs are key to characterizing pharmacogenomic (PGx) relationships. ADME gene variation is relatively well characterized in European and Asian populations, but data from African populations are under-studied-which has implications for drug safety and effective use in Africa. Results: We identified significant ADME gene variation in African populations using data from 458 high-coverage whole genome sequences, 412 of which are novel, and from previously available African sequences from the 1,000 Genomes Project. ADME variation was not uniform across African populations, particularly within high impact coding variation. Copy number variation was detected in 116 ADME genes, with equal ratios of duplications/deletions. We identified 930 potential high impact coding variants, of which most are discrete to a single African population cluster. Large frequency differences (i.e., >10%) were seen in common high impact variants between clusters. Several novel variants are predicted to have a significant impact on protein structure, but additional functional work is needed to confirm the outcome of these for PGx use. Most variants of known clinical outcome are rare in Africa compared to European populations, potentially reflecting a clinical PGx research bias to European populations. Discussion: The genetic diversity of ADME genes across sub-Saharan African populations is large. The Southern African population cluster is most distinct from that of far West Africa. PGx strategies based on European variants will be of limited use in African populations. Although established variants are important, PGx must take into account the full range of African variation. This work urges further characterization of variants in African populations including in vitro and in silico studies, and to consider the unique African ADME landscape when developing precision medicine guidelines and tools for African populations.

Non-ambient X-ray and neutron diffraction of novel relaxor ferroelectric xBi2(Zn2/3,Nb1/3)O3-(1 - x)BaTiO3.

The first determination of the phase diagram of the novel ferroelectric relaxor xBi(Zn2/3Nb1/3)O3-(1 - x)BaTiO3 (BZN-BT) has been achieved with a combination of high-resolution X-ray and neutron diffraction up to the miscibility limit near x(BZN) = 20.0% over a temperature range 20 < T < 400 K. The combined X-ray and neutron data show that the instability within the xBZN-(1-x)BT system reaches a maximum at x = 3.9% and is driven by B-site displacement and distortion of the oxygen octahedra in the polar phases. Composition-dependent effects include a narrow Amm2-dominated region focused at x = 3.9%, significant convergence of the lattice parameters in both P4mm and Amm2 phases, and sharp maxima in piezoelectric coefficient d 33 and maximum polarization P max. Lattice parameter dilation at x ≥ 4.0% was observed for both P4mm and Amm2 unit cells, alongside the first appearance of Pm 3 m at 295 K and the onset of significant dielectric relaxation. Low-temperature neutron diffraction indicated a weak or non-existent temperature dependence on the transition from ferroelectric at x = 3.9% to ferroelectric relaxor at x = 4.0%. Temperature-dependent phase transitions were eliminated near x = 3.0%, with the ferroelectric limit observed at x = 5.0% and a transition to a low-loss relaxor dielectric near x = 8.0%.

Skin Metagenomic Sequence Analysis of Early Candida auris Outbreaks in U.S. Nursing Homes.

Candida auris is a human fungal pathogen classified as an urgent threat to the delivery of health care due to its extensive antimicrobial resistance and the high mortality rates associated with invasive infections. Global outbreaks have occurred in health care facilities, particularly, long-term care hospitals and nursing homes. Skin is the primary site of colonization for C. auris. To accelerate research studies, we developed microbiome sequencing protocols, including amplicon and metagenomic sequencing, directly from patient samples at health care facilities with ongoing C. auris outbreaks. We characterized the skin mycobiome with a database optimized to classify Candida species and C. auris to the clade level. While Malassezia species were the predominant skin-associated fungi, nursing home residents also harbored Candida species, including C. albicans, and C. parapsilosis. Amplicon sequencing was concordant with culturing studies to identify C. auris-colonized patients and provided further resolution that distinct clades of C. auris are colonizing facilities in New York and Illinois. Shotgun metagenomic sequencing from a clinical sample with a high fungal bioburden generated a skin-associated profile of the C. auris genome. Future larger scale clinical studies are warranted to more systematically investigate the effects of commensal microbes and patient risk factors on the colonization and transmission of C. auris. IMPORTANCE Candida auris is a human pathogen of high concern due to its extensive antifungal drug resistance and high mortality rates associated with invasive infections. Candida auris skin colonization and persistence on environmental surfaces make this pathogen difficult to control once it enters a health care facility. Residents in long-term care hospitals and nursing homes are especially vulnerable. In this study, we developed microbiome sequencing protocols directly from surveillance samples, including amplicon and metagenomic sequencing, demonstrating concordance between sequencing results and culturing.

Studying information recurrence, gatekeeping, and the role of communities during internet outages in Venezuela.

Many authoritarian regimes have taken to censoring internet access in order to stop the spread of misinformation, restrict citizens from discussing certain topics, and prevent mobilization, among other reasons. There are several theories about the effectiveness of censorship. Some suggest that censorship will effectively limit the flow of information, whereas others predict that a backlash will form, resulting in ultimately more discussion about the topic. In this work, we analyze the role of communities and gatekeepers during multiple internet outages in Venezuela in January 2019. First, we measure how critical information (e.g., entities and hashtags) spreads during outages focusing on information recurrence and burstiness within and across language and location communities. We discover that information bursts tend to cross both language and location community boundaries rather than being limited to a single community during several outages. Then we identify users who play central roles and propose a novel method to detect gatekeepers-users who prevent critical information from spreading across communities during outages. We show that bilingual and English-speaking users play more central roles compared to Spanish-speaking users, but users inside and outside Venezuela have similar distribution of centrality. Finally, we measure the differences in social network structure before and after each outage event and discuss its effect on how information spreads. We find that with each outage event social connections tend to get less connected with higher mean shortest path, indicating that the effect of censorship makes it harder for information to spread.

Structure-based optimisation of orally active & reversible MetAP-2 inhibitors maintaining a tight 'molecular budget'.

Structure-based led optimisation of orally active reversible Methionine Aminopeptidase-2 (MetAP-2) inhibitors utilising a 'molecular budget' medicinal chemistry strategy is described. The key physicochemical parameters of target molecules (cLogP, molecular size and H-bond donor count) were monitored through straightforward and intuitive use of atom count and distribution. The balance between structure-based design and an awareness of the physicochemical properties of the compounds synthesised enabled the rapid identification of a potent molecule with good oral pharmacokinetic (PK) characteristics by making fewer, higher quality compounds. The resulting candidate quality molecule was validated in a mechanistic cellular assay and a rodent secondary immunisation model.

Discovery of a Bromodomain and Extraterminal Inhibitor with a Low Predicted Human Dose through Synergistic Use of Encoded Library Technology and Fragment Screening.

The bromodomain and extraterminal (BET) family of bromodomain-containing proteins are important regulators of the epigenome through their ability to recognize N-acetyl lysine (KAc) post-translational modifications on histone tails. These interactions have been implicated in various disease states and, consequently, disruption of BET-KAc binding has emerged as an attractive therapeutic strategy with a number of small molecule inhibitors now under investigation in the clinic. However, until the utility of these advanced candidates is fully assessed by these trials, there remains scope for the discovery of inhibitors from new chemotypes with alternative physicochemical, pharmacokinetic, and pharmacodynamic profiles. Herein, we describe the discovery of a candidate-quality dimethylpyridone benzimidazole compound which originated from the hybridization of a dimethylphenol benzimidazole series, identified using encoded library technology, with an N-methyl pyridone series identified through fragment screening. Optimization via structure- and property-based design led to I-BET469, which possesses favorable oral pharmacokinetic properties, displays activity in vivo, and is projected to have a low human efficacious dose.

Plasmid Dissemination and Selection of a Multidrug-Resistant Klebsiella pneumoniae Strain during Transplant-Associated Antibiotic Therapy.

Antibiotics, which are used both to prevent and to treat infections, are a mainstay therapy for lifesaving procedures such as transplantation. For this reason, and many others, increased antibiotic resistance among human-associated pathogens, such as the carbapenem-resistant Enterobacteriaceae species, is of grave concern. In this study, we report on a hematopoietic stem cell transplant recipient in whom cultures detected the emergence of carbapenem resistance and spread across five strains of bacteria that persisted for over a year. Carbapenem resistance in Citrobacter freundii, Enterobacter cloacae, Klebsiella aerogenes, and Klebsiella pneumoniae was linked to a pair of plasmids, each carrying the Klebsiella pneumoniae carbapenemase gene (blaKPC). Surveillance cultures identified a carbapenem-susceptible strain of Citrobacter freundii that may have become resistant through horizontal gene transfer of these plasmids. Selection of a multidrug-resistant Klebsiella pneumoniae strain was also detected following combination antibiotic therapy. Here we report a plasmid carrying the blaKPC gene with broad host range that poses the additional threat of spreading to endogenous members of the human gut microbiome.IMPORTANCE Antibiotic-resistant bacteria are a serious threat to medically fragile patient populations. The spread of antibiotic resistance through plasmid-mediated mechanisms is of grave concern as it can lead to the conversion of endogenous patient-associated strains to difficult-to-treat pathogens.

Design and development of a series of borocycles as selective, covalent kallikrein 5 inhibitors.

The connection between Netherton syndrome and overactivation of epidermal/dermal proteases, particularly Kallikrein 5 (KLK5) has been well established and it is expected that a KLK5 inhibitor would improve the dermal barrier and also reduce the pain and itch that afflict Netherton syndrome patients. One of the challenges of covalent protease inhibitors has been achieving selectivity over closely related targets. In this paper we describe the use of structural insight to design and develop a selective and highly potent reversibly covalent KLK5 inhibitor from an initial weakly binding fragment.

A novel unembalmed human cadaveric limb model for assessing conformational changes in self-expanding nitinol stents in the popliteal artery.

OBJECTIVE: To develop an unembalmed human cadaveric lower limb model as a more realistic environment for testing self-expanding nitinol stents. We studied conformational changes and strain induced by knee flexion in nitinol stents deployed in the popliteal artery (PA). METHODS: One Lifestent® each was deployed into one limb of four cadavers (control group), while the contralateral leg received a different stent (Absolute®, Protégé Everflex®, Supera®, and Gore Viabahn®). The limbs were mounted on a quasi-static knee rig (QKR) and X-ray imaging was performed at pre-defined knee flexion angles. A least-squares solution to the equation of a circle was used to assess radius of curvature at flexion points (FP), and nominal strain was calculated for each stented artery. RESULTS: There were differences but also some similarities in conformational changes seen in the various stents. Knee flexion produced at least two FP in all stents. The mean radius of curvature decreased with increasing degrees of flexion but more so in distal (main) than proximal (accessory) FP (22 mm vs. 11 mm) in all stents. Supera® stent had the greatest relative radii of curvatures, and Absolute® stent had the highest strain value in comparison to the contralateral PA as the control group. CONCLUSION: This study describes a novel human cadaveric limb model for testing self-expanding nitinol stents implanted in the PA. Several parameters of conformational change in the stented PA such as FP formation, FP translocation and strain induced by axial compression were described. These may be useful for developing new stents for the PA location.

Predicting onset of complications from diabetes: a graph based approach.

Diabetes is a significant health concern with more than 30 million Americans living with diabetes. Onset of diabetes increases the risk for various complications, including kidney disease, myocardial infractions, heart failure, stroke, retinopathy, and liver disease. In this paper, we study and predict the onset of these complications using a network-based approach by identifying fast and slow progressors. That is, given a patient's diagnosis of diabetes, we predict the likelihood of developing one or more of the possible complications, and which patients will develop complications quickly. This combination of "if a complication will be developed" with "how fast it will be developed" can aid the physician in developing better diabetes management program for a given patient.

Investigation of a Cluster of Sphingomonas koreensis Infections.

BACKGROUND: Plumbing systems are an infrequent but known reservoir for opportunistic microbial pathogens that can infect hospitalized patients. In 2016, a cluster of clinical sphingomonas infections prompted an investigation. METHODS: We performed whole-genome DNA sequencing on clinical isolates of multidrug-resistant Sphingomonas koreensis identified from 2006 through 2016 at the National Institutes of Health (NIH) Clinical Center. We cultured S. koreensis from the sinks in patient rooms and performed both whole-genome and shotgun metagenomic sequencing to identify a reservoir within the infrastructure of the hospital. These isolates were compared with clinical and environmental S. koreensis isolates obtained from other institutions. RESULTS: The investigation showed that two isolates of S. koreensis obtained from the six patients identified in the 2016 cluster were unrelated, but four isolates shared more than 99.92% genetic similarity and were resistant to multiple antibiotic agents. Retrospective analysis of banked clinical isolates of sphingomonas from the NIH Clinical Center revealed the intermittent recovery of a clonal strain over the past decade. Unique single-nucleotide variants identified in strains of S. koreensis elucidated the existence of a reservoir in the hospital plumbing. Clinical S. koreensis isolates from other facilities were genetically distinct from the NIH isolates. Hospital remediation strategies were guided by results of microbiologic culturing and fine-scale genomic analyses. CONCLUSIONS: This genomic and epidemiologic investigation suggests that S. koreensis is an opportunistic human pathogen that both persisted in the NIH Clinical Center infrastructure across time and space and caused health care-associated infections. (Funded by the NIH Intramural Research Programs.).

Approaches to target tractability assessment - a practical perspective.

The assessment of the suitability of novel targets to intervention by different modalities, e.g. small molecules or antibodies, is increasingly seen as important in helping to select the most progressable targets at the outset of a drug discovery project. This perspective considers differing aspects of tractability and how it can be assessed using in silico and experimental approaches. We also share some of our experiences in using these approaches.

Health Workers' Knowledge and Perceptions on Dementia in Skilled Nursing Homes: A Pilot Implementation of Teepa Snow's Positive Approach to Care Certification Course.

INTRODUCTION: A knowledge gap on caring for people with dementia exists among health workers employed in skilled nursing facilities. This article reports on knowledge gained and perceptions changed among 24 skilled nursing facility health workers who completed Teepa Snow's innovative Positive Approach to Care Certification course as a regional initiative. METHODS: This pilot study used a quasi-experiment with a one-group pretest-posttest design to assess the dementia knowledge and perceptions of health workers who participated in the course. Paired responses (n = 22) for the 54-item dementia knowledge and training/coaching perceptions survey were compared using the McNemar test, Paired t test, and Wilcoxon signed-rank test to analyze the improvement in knowledge and perceptions of dementia. RESULTS: Significant knowledge gain was found among health workers after completing the program (t(21) = -7.46, P < .001). Statistical differences (P < .05) were present in eight knowledge items (ie, working memory, mental health, temporal lobe, "sapphire", "space and awareness", personal space, binocular vision, and hippocampus) and four perception areas (ie, approaching, understanding, calming, and instructing) about dementia. Significant changes were also found in four areas of health workers' perceptions about dementia (P < .05). DISCUSSION: The certification course can improve knowledge and facilitate perceptual change on dementia. Implementing this program as a novel regional initiative has robust potential in ensuring continuing workforce development in health care settings challenged with ever-changing people living with dementia needs and high staff turnover.

Genomic Analysis of Hospital Plumbing Reveals Diverse Reservoir of Bacterial Plasmids Conferring Carbapenem Resistance.

The hospital environment is a potential reservoir of bacteria with plasmids conferring carbapenem resistance. Our Hospital Epidemiology Service routinely performs extensive sampling of high-touch surfaces, sinks, and other locations in the hospital. Over a 2-year period, additional sampling was conducted at a broader range of locations, including housekeeping closets, wastewater from hospital internal pipes, and external manholes. We compared these data with previously collected information from 5 years of patient clinical and surveillance isolates. Whole-genome sequencing and analysis of 108 isolates provided comprehensive characterization of blaKPC/blaNDM-positive isolates, enabling an in-depth genetic comparison. Strikingly, despite a very low prevalence of patient infections with blaKPC-positive organisms, all samples from the intensive care unit pipe wastewater and external manholes contained carbapenemase-producing organisms (CPOs), suggesting a vast, resilient reservoir. We observed a diverse set of species and plasmids, and we noted species and susceptibility profile differences between environmental and patient populations of CPOs. However, there were plasmid backbones common to both populations, highlighting a potential environmental reservoir of mobile elements that may contribute to the spread of resistance genes. Clear associations between patient and environmental isolates were uncommon based on sequence analysis and epidemiology, suggesting reasonable infection control compliance at our institution. Nonetheless, a probable nosocomial transmission of Leclercia sp. from the housekeeping environment to a patient was detected by this extensive surveillance. These data and analyses further our understanding of CPOs in the hospital environment and are broadly relevant to the design of infection control strategies in many infrastructure settings.IMPORTANCE Carbapenemase-producing organisms (CPOs) are a global concern because of the morbidity and mortality associated with these resistant Gram-negative bacteria. Horizontal plasmid transfer spreads the resistance mechanism to new bacteria, and understanding the plasmid ecology of the hospital environment can assist in the design of control strategies to prevent nosocomial infections. A 5-year genomic and epidemiological survey was undertaken to study the CPOs in the patient-accessible environment, as well as in the plumbing system removed from the patient. This comprehensive survey revealed a vast, unappreciated reservoir of CPOs in wastewater, which was in contrast to the low positivity rate in both the patient population and the patient-accessible environment. While there were few patient-environmental isolate associations, there were plasmid backbones common to both populations. These results are relevant to all hospitals for which CPO colonization may not yet be defined through extensive surveillance.

The Use and Intended Outcomes of Presence: A Focus Group Study.

PURPOSE: To explore and understand the use and intended outcomes of presence from the perspective of and as experienced by nurses. METHODS: Twenty-seven nurses participated in one of four focus groups. Data were analyzed using Giorgi's phenomenological method. FINDINGS: Four themes emerged: (1) therapeutic communication; (2) nurse well-being; (3) dimensions of presence; and (4) intention to improve client outcomes. CONCLUSIONS: Presence was described as a multidimensional intervention that required therapeutic communication and nurse well-being with the intention of improving client outcomes. Study findings provide evidence of the significance of presence in the face of human interaction that is shifting to virtual, impersonal communication.

Unprecedented phase transition sequence in the perovskite Li0.2Na0.8NbO3.

The perovskite Li0.2Na0.8NbO3 is shown, by powder neutron diffraction, to display a unique sequence of phase transitions at elevated temperature. The ambient temperature polar phase (rhombohedral, space group R3c) transforms via a first-order transition to a polar tetragonal phase (space group P42mc) in the region 150-300°C; these two phases correspond to Glazer tilt systems a-a-a- and a+a+c-, respectively. At 500°C a ferroelectric-paraelectric transition takes place from P42mc to P42/nmc, retaining the a+a+c- tilt. Transformation to a single-tilt system, a0a0c+ (space group P4/mbm), occurs at 750°C, with the final transition to the aristotype cubic phase at 850°C. The P42mc and P42/nmc phases have each been seen only once and twice each, respectively, in perovskite crystallography, in each case in compositions prepared at high pressure.

Plasmid Dynamics in KPC-Positive Klebsiella pneumoniae during Long-Term Patient Colonization.

UNLABELLED: Carbapenem-resistant Klebsiella pneumoniae strains are formidable hospital pathogens that pose a serious threat to patients around the globe due to a rising incidence in health care facilities, high mortality rates associated with infection, and potential to spread antibiotic resistance to other bacterial species, such as Escherichia coli Over 6 months in 2011, 17 patients at the National Institutes of Health (NIH) Clinical Center became colonized with a highly virulent, transmissible carbapenem-resistant strain of K. pneumoniae Our real-time genomic sequencing tracked patient-to-patient routes of transmission and informed epidemiologists' actions to monitor and control this outbreak. Two of these patients remained colonized with carbapenemase-producing organisms for at least 2 to 4 years, providing the opportunity to undertake a focused genomic study of long-term colonization with antibiotic-resistant bacteria. Whole-genome sequencing studies shed light on the underlying complex microbial colonization, including mixed or evolving bacterial populations and gain or loss of plasmids. Isolates from NIH patient 15 showed complex plasmid rearrangements, leaving the chromosome and the blaKPC-carrying plasmid intact but rearranging the two other plasmids of this outbreak strain. NIH patient 16 has shown continuous colonization with blaKPC-positive organisms across multiple time points spanning 2011 to 2015. Genomic studies defined a complex pattern of succession and plasmid transmission across two different K. pneumoniae sequence types and an E. coli isolate. These findings demonstrate the utility of genomic methods for understanding strain succession, genome plasticity, and long-term carriage of antibiotic-resistant organisms. IMPORTANCE: In 2011, the NIH Clinical Center had a nosocomial outbreak involving 19 patients who became colonized or infected with blaKPC-positive Klebsiella pneumoniae Patients who have intestinal colonization with blaKPC-positive K. pneumoniae are at risk for developing infections that are difficult or nearly impossible to treat with existing antibiotic options. Two of those patients remained colonized with blaKPC-positive Klebsiella pneumoniae for over a year, leading to the initiation of a detailed genomic analysis exploring mixed colonization, plasmid recombination, and plasmid diversification. Whole-genome sequence analysis identified a variety of changes, both subtle and large, in the blaKPC-positive organisms. Long-term colonization of patients with blaKPC-positive Klebsiella pneumoniae creates new opportunities for horizontal gene transfer of plasmids encoding antibiotic resistance genes and poses complications for the delivery of health care.