Several confirmed and probable zoonotic cases of toxigenic Corynebacterium ulcerans, Queensland, Australia.

Background: Toxigenic Corynebacterium ulcerans is an emerging zoonosis globally, causing both cutaneous and respiratory diphtheria-like illness. In Queensland, human infection with toxigenic C. ulcerans is rare, with only three cases reported before October 2015. This case series describes five subsequent cases of toxigenic C. ulcerans in Queensland with links to companion animals. Methods: All data were collected as part of routine public health response, and strains were whole genome sequenced for further characterisation. Household contacts were screened, treated with appropriate antibiotics, and received a diphtheria toxoid-containing vaccine if more than five years had elapsed since their last dose. Findings: No epidemiological or genomic links could be established between any of the five patients, including between the two cases notified from the same locality within eight days of each other. The C. ulcerans strains from Cases Two, Four and Five were closely related to the strains isolated from their respective pets by whole genome sequencing. Domestic dogs were identified as the most likely mode of transmission for Cases One and Three; however, this was unable to be laboratory confirmed, since Case One's dog was treated with antibiotics before it could be tested, and Case Three's dog was euthanised and cremated prior to case notification. Interpretation: These are the first reported Australian cases of this emerging zoonosis with links to companion animals. These cases demonstrate the likely transmission route between companion animals and humans, with no evidence of human-to-human transmission. The existing requirement in the Queensland Health Public Health Management Guidelines, of restrictions on cases and some contacts while awaiting swab results, is currently under review.

Genome-Wide CRISPR Screens Identify Multiple Synthetic Lethal Targets That Enhance KRASG12C Inhibitor Efficacy.

Non-small lung cancers (NSCLC) frequently (∼30%) harbor KRAS driver mutations, half of which are KRASG12C. KRAS-mutant NSCLC with comutated STK11 and/or KEAP1 is particularly refractory to conventional, targeted, and immune therapy. Development of KRASG12C inhibitors (G12Ci) provided a major therapeutic advance, but resistance still limits their efficacy. To identify genes whose deletion augments efficacy of the G12Cis adagrasib (MRTX-849) or adagrasib plus TNO155 (SHP2i), we performed genome-wide CRISPR/Cas9 screens on KRAS/STK11-mutant NSCLC lines. Recurrent, potentially targetable, synthetic lethal (SL) genes were identified, including serine-threonine kinases, tRNA-modifying and proteoglycan synthesis enzymes, and YAP/TAZ/TEAD pathway components. Several SL genes were confirmed by siRNA/shRNA experiments, and the YAP/TAZ/TEAD pathway was extensively validated in vitro and in mice. Mechanistic studies showed that G12Ci treatment induced gene expression of RHO paralogs and activators, increased RHOA activation, and evoked ROCK-dependent nuclear translocation of YAP. Mice and patients with acquired G12Ci- or G12Ci/SHP2i-resistant tumors showed strong overlap with SL pathways, arguing for the relevance of the screen results. These findings provide a landscape of potential targets for future combination strategies, some of which can be tested rapidly in the clinic. SIGNIFICANCE: Identification of synthetic lethal genes with KRASG12C using genome-wide CRISPR/Cas9 screening and credentialing of the ability of TEAD inhibition to enhance KRASG12C efficacy provides a roadmap for combination strategies. See related commentary by Johnson and Haigis, p. 4005.

Integrating remote sensing with ecology and evolution to advance biodiversity conservation.

Remote sensing has transformed the monitoring of life on Earth by revealing spatial and temporal dimensions of biological diversity through structural, compositional and functional measurements of ecosystems. Yet, many aspects of Earth's biodiversity are not directly quantified by reflected or emitted photons. Inclusive integration of remote sensing with field-based ecology and evolution is needed to fully understand and preserve Earth's biodiversity. In this Perspective, we argue that multiple data types are necessary for almost all draft targets set by the Convention on Biological Diversity. We examine five key topics in biodiversity science that can be advanced by integrating remote sensing with in situ data collection from field sampling, experiments and laboratory studies to benefit conservation. Lowering the barriers for bringing these approaches together will require global-scale collaboration.

Patient Perspectives on Detox: Practical and Personal Considerations through a Lens of Patient-Centered Care.

Inpatient detoxification is often required before a client can move on to additional substance abuse treatment services. Although often short-term, time spent in inpatient detoxification tends to have long-lasting effects on the recovery process. This qualitative study focuses on one treatment facility in Tampa, Florida that offers a range of recovery services, including inpatient detox and outpatient treatment. Focus groups (N = 70 participants) captured client perceptions of direct clinical care operations, access to resources, and relationships with direct care staff within the inpatient detox program. Perceptions were then assessed using a thematic analysis approach with attention to the literature on person-centered care best practices, behavior change, and patient engagement theories to better understand how facility practices affect treatment engagement and retention. Findings elucidated several practical facilitators and barriers to recovery such as facility resources, services offered, transition to aftercare, and sustainability of treatment. Findings also illuminated several personal facilitators and barriers including patient-staff interactions, personal motivation, and family and community support. The resulting recommendations for practice and research are discussed.

Catalase-Loaded Silica Nanoparticles Formulated via Direct Surface Modification as Potential Oxygen Generators for Hypoxia Relief.

Enzymes are biological catalysts that have many potential industrial and biomedical applications. However, the widespread use of enzymes in the industry has been limited by their instability and poor recovery. In biomedical applications, systemic administration of enzymes has faced two main challenges: limited bioactivity mostly due to rapid degradation by proteases and immunogenic activity, since most enzymes are from nonhuman sources. Herein, we propose a robust enzyme-encapsulation strategy to mitigate these limitations. Catalase (CAT) was encapsulated in nanoporous silica nanoparticles (CAT-SiNPs) by first chemically modifying the enzyme surface with a silica precursor, followed by silica growth and finally poly(ethylene glycol) (PEG) conjugation. The formulation was carried out in mild aqueous conditions and yielded nanoparticles (NPs) with a mean diameter of 230 ± 10 nm and a concentration of 1.3 ± 0.8 × 1012 NPs/mL. CAT-SiNPs demonstrated high enzyme activity, optimal protection from proteolysis by proteinase K and trypsin, and excellent stability over time. In addition, a new electrochemical assay was developed to measure CAT activity in a rapid, simple, and accurate manner without interference from chromophore usually present in biological samples. Concentrations of 2.5 × 1010 to 80 × 1010 CAT-SiNPs/mL not only proved to be nontoxic in cell cultures using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay but also conferred cell protection when cells were exposed to 1 mM hydrogen peroxide (H2O2). Finally, the ability of CAT-SiNPs to release oxygen (O2) when exposed to H2O2 was demonstrated in vivo using a rat model. Following the direct injection of CAT-SiNPs in the left kidney, partial pressure of oxygen (pO2) increased by more than 30 mmHg compared to the contralateral control kidney during the systemic infusion of safe levels of H2O2. This pilot study highlights the potential of CAT-SiNPs to generate O2 to relieve hypoxia in tissues and potentially sensitize tumors against radiation therapy.

Thrombin-Activatable Microbubbles as Potential Ultrasound Contrast Agents for the Detection of Acute Thrombosis.

Acute deep vein thrombosis (DVT) is the formation of a blood clot in the deep veins of the body that can lead to fatal pulmonary embolism. Acute DVT is difficult to distinguish from chronic DVT by ultrasound (US), the imaging modality of choice, and is therefore treated aggressively with anticoagulants, which can lead to internal bleeding. Here we demonstrate that conjugating perfluorobutane-filled (PFB-filled) microbubbles (MBs) with thrombin-sensitive activatable cell-penetrating peptides (ACPPs) could lead to the development of contrast agents that detect acute thrombosis with US imaging. Successful conjugation of ACPP to PFB-filled MBs was confirmed by fluorescence microscopy and flow cytometry. Fluorescein-labeled ACPP was used to evaluate the efficiency of thrombin-triggered cleavage by measuring the mean fluorescence intensity of ACPP-labeled MBs (ACPP-MBs) before and after incubation at 37 °C with thrombin. Lastly, control MBs and ACPP-MBs were infused through a tube containing a clot, and US contrast enhancement was measured with or without the presence of a thrombin inhibitor after washing the clot with saline. With thrombin activity, 91.7 ± 14.2% of the signal was retained after ACPP-MB infusion and washing, whereas only 16.7 ± 4% of the signal was retained when infusing ACPP-MBs in the presence of hirudin, a potent thrombin inhibitor.

Quality of Care Is Improved by Rapid Short Incubation MALDI-ToF Identification from Blood Cultures as Measured by Reduced Length of Stay and Patient Outcomes as Part of a Multi-Disciplinary Approach to Bacteremia in Pediatric Patients.

Sepsis has seen an incremental increase in cases of about 13% annually in the USA and accounts for approximately 4400 deaths among pediatric patients. Early identification of the specific pathogen allows the clinician to ensure that the antibiotic coverage is optimal, an intervention that has been shown to improve patient outcomes in sepsis. Our study's objective was to assess the impact of a rapid Bruker MALDI-Tof identification protocol on pediatric sepsis cases by assessing various indicators. We assessed the quality of care by measuring the following indicators; time to identification of the pathogen, initiation of the most appropriate antibiotic, length of stay (LOS) in hospital and patient outcomes, using a retrospective review over three consecutive years. In total 92 pediatric patients, similar in age and gender distributions were assessed; 37 in 2012, 33 in 2013 and 22 in 2014. The introduction of MALDI-TOF identification in 2013 led to a significant decrease in time to identify a pathogen by 21.03 hours (p = 1.95E-05). A short incubation MALDI-TOF identification protocol in 2014 further reduced time to identification by 17.75 hours (p = 2.48E-3). Overall in 2014 this led to a trend to earlier optimization of antibiotics by 20.2 hours (p = 0.14) and a reduction in length of stay after the implementation of MALDI-ToF identification in 2013 of 3.07 days and a further reduction of 8.92 days after the introduction of the rapid short incubation identification protocol using MALDI-Tof in 2014 (P = 0.12). By evaluating the subgroup of patients where antibiotics were changed, our study confirmed that patients received appropriate therapy 48.8% (20.2 hours) earlier compared to conventional methods leading to a decrease in length of stay of 23.65 days after the implementation of MALDI-ToF identification and a further reduction of 9.82 days in 2014 compared to 2012 (p = 0.02). In 2014 outcomes between the patients needing a change in their antibiotic compared to the patients where the empirical therapy was considered to be optimal were similar with respect to length of stay; 13.04 and 10.93 days (p = 0.34). In the 2012 group there was a significant increase in the length of stay in the group needing change in excess of 30 days (p = 0.02) compared to the group where empirical therapy was considered to be optimal, this clearly showed an improvement in the quality of care received after the rapid identification was instituted in 2014. The 2012 group had a four times overall increased sepsis associated mortality risk compared to the 2014 group and when empirical antibiotics needed to be optimized this risk was 7 times compared to the 2014 group. We conclude that rapid identification of bacterial pathogens in pediatric blood cultures with a rapid incubation MALDI-TOF identification protocol plays an important role in improving quality of care as part of a multidisciplinary approach to pediatric bacteremia and sepsis.

Sparse Spectro-Temporal Receptive Fields Based on Multi-Unit and High-Gamma Responses in Human Auditory Cortex.

Spectro-Temporal Receptive Fields (STRFs) were estimated from both multi-unit sorted clusters and high-gamma power responses in human auditory cortex. Intracranial electrophysiological recordings were used to measure responses to a random chord sequence of Gammatone stimuli. Traditional methods for estimating STRFs from single-unit recordings, such as spike-triggered-averages, tend to be noisy and are less robust to other response signals such as local field potentials. We present an extension to recently advanced methods for estimating STRFs from generalized linear models (GLM). A new variant of regression using regularization that penalizes non-zero coefficients is described, which results in a sparse solution. The frequency-time structure of the STRF tends toward grouping in different areas of frequency-time and we demonstrate that group sparsity-inducing penalties applied to GLM estimates of STRFs reduces the background noise while preserving the complex internal structure. The contribution of local spiking activity to the high-gamma power signal was factored out of the STRF using the GLM method, and this contribution was significant in 85 percent of the cases. Although the GLM methods have been used to estimate STRFs in animals, this study examines the detailed structure directly from auditory cortex in the awake human brain. We used this approach to identify an abrupt change in the best frequency of estimated STRFs along posteromedial-to-anterolateral recording locations along the long axis of Heschl's gyrus. This change correlates well with a proposed transition from core to non-core auditory fields previously identified using the temporal response properties of Heschl's gyrus recordings elicited by click-train stimuli.