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PURPOSE: Movement behaviors (i.e. physical activity [PA], sedentary behavior [SB], and sleep) are intrinsically codependent, an issue resolved using compositional data analysis (CoDA). High-sensitivity C-reactive protein (hs-CRP) is a nonspecific inflammatory marker positively associated with cardiovascular diseases and affected by movement behaviors. Examine the relation between movement behaviors using CoDA and how time reallocation between two movement behaviors was associated with hs-CRP concentration. METHODS: The Jackson Heart Study was designed to investigate cardiovascular disease risk factors among African American participants in the Jackson, MS area. PA and sleep were self-reported with SB calculated as the remaining time in the day. RESULTS: The median untransformed hs-CRP concentration was 0.28 mg·dL (−1) (interquartile range; 0.11, 0.61). Reallocating 15 minutes of PA with SB, the hypothetical change in log hs-CRP concentration was 0.08 mg·dL (−1) (95% CIs; 0.04, 0.11) greater than the average log hs-CRP concentration. Substituting 15 minutes of SB or sleep with PA was associated with a hypothetical change in log hs-CRP concentration difference of −0.05 mg·dL (−1) (−0.08, −0.03) and −0.06 mg·dL (−1) (−0.08, −0.03), respectively. Reallocations between SB and sleep were not associated with the hypothetical difference in log hs-CRP concentration. CONCLUSIONS: Modeling estimates suggest replacing 15 minutes of SB with PA is associated with lower inflammation.

Thirty to seventy percent of all venous thromboembolism (VTE) events are associated with hospitalization. The absolute and relative risks during and after hospitalization are poorly characterized.Quantify the absolute rate and relative risk of VTE during and up to 3 months after medical and surgical hospitalizations.We conducted an observational cohort study between 2010 and 2016 of patients cared for by the University of Vermont (UVM) Health Network's primary care population. Cox proportional hazard models with hospitalization modeled as a time-varying covariate were used to estimate VTE risk.Over 4.3 years of follow-up, 55 220 hospitalizations (156 per 1000 person-years) and 713 first venous thromboembolism events (2.0 per 1000 person-years) occurred. Among individuals not recently hospitalized, the rate of venous thromboembolism was 1.4 per 1000 person-years and 71.8 per 1000 person-years during hospitalization. During the first, second, and third months after discharge, the rates of venous thromboembolism were 35.1, 11.3, and 5.2 per 1000 person-years, respectively. Relative to those not recently hospitalized, the age- and sex-adjusted HRs of venous thromboembolism were 38.0 (95% CI 28.0, 51.5) during hospitalization, and 18.4 (95% CI 15.0, 22.6), 6.3 (95% CI 4.3, 9.0), and 3.0 (95% CI 1.7, 5.4) during the first, second, and third months after discharge, respectively. Stratified by medical versus surgical services the rates were similar.Hospitalization and up to 3 months after discharge were strongly associated with increased venous thromboembolism risk. These data quantify this risk for use in future studies.

ObjectivesTo investigate the COVID-19 infections among staff at our institution and determine the interventions required to prevent subsequent staff infections.DesignRetrospective cohort studyParticipants and settingStaff working at a single tertiary referral hospital who returned a positive test result for SARS-CoV-2 between 25 January 2020 and 25 November 2020.Main outcome measuresSource of COVID-19 infection.ResultsOf 45 staff who returned a positive test result for SARS-CoV-2, 19 were determined to be acquired at Austin Health. Fifteen (15/19; 79% [95% CI: 54–94%]) of these were identified through contact tracing and testing following exposures to other infected staff and were presumed to be staff-staff transmission, including 10 healthcare workers (HCWs) linked to a single ward that cared for COVID-19 patients. Investigation of the outbreak identified the staff tearoom as the likely location for transmission, with subsequent reduction in HCW infections and resolution of the outbreak following implementation of enhanced control measures in tearoom facilities. No HCW contacts (0/204; 0% [95% CI: 0–2%]) developed COVID-19 infection following exposure to unrecognised patients with COVID-19.ConclusionsUnrecognised infections among staff may be a significant driver of HCW infections in healthcare settings. Control measures should be implemented to prevent acquisition from other staff as well as patient-staff transmission.

Background: There is no gold standard approach for delirium diagnosis, making the assessment of its epidemiology difficult. Delirium can only be inferred though observation of behavioural disturbance and described with relevant nouns or adjectives. Objective: We aimed to use natural language processing (NLP) and its identification of words descriptive of behavioural disturbance to study the epidemiology of delirium in critically ill patients. Study design: Retrospective study using data collected from the electronic health records of a university-affiliated intensive care unit (ICU) in Melbourne, Australia. Participants: 12 375 patients Intervention: Analysis of electronic progress notes. Identification using NLP of at least one of a list of words describing behavioural disturbance within such notes. Results: We analysed 199 648 progress notes in 12 375 patients. Of these, 5108 patients (41.3%) had NLP-diagnosed behavioural disturbance (NLP-Dx-BD). Compared with those who did not have NLP-Dx-DB, these patients were older, more severely ill, and likely to have medical or unplanned admissions, neurological diagnosis, chronic kidney or liver disease and to receive mechanical ventilation and renal replacement therapy (P < 0.001). The unadjusted hospital mortality for NLP-Dx-BD patients was 14.1% versus 9.6% for patients without NLP-Dx-BD. After adjustment for baseline characteristics and illness severity, NLP-Dx-BD was not associated with increased risk of death (odds ratio [OR], 0.94; 95% CI, 0.80–1.10); a finding robust to multiple sensitivity, subgroups and time of observation subcohort analyses. In mechanically ventilated patients, NLP-Dx-BD was associated with decreased hospital mortality (OR, 0.80; 95% CI, 0.65–0.99) after adjustment for baseline severity of illness and year of admission. Conclusions: NLP enabled rapid assessment of large amounts of data identifying a population of ICU patients with typical high risk characteristics for delirium. Moreover, this technique enabled identification of previously poorly understood associations. Further investigations of this technique appear justified.

Background Due to regional shortages some health services have proposed using surgical masks manufactured from sterilisation wrap. However, there has been little assessment of the safety of this practice. Therefore, we developed our own prototypes and evaluated whether they met regulatory standards. Methods Surgical mask prototypes were manufactured from two thickness grades of commercial sterilisation wrap. Safety was assessed in the context of regulatory standards. As it was not previously reported, we developed and performed differential pressure and synthetic blood penetration resistance experiments in accordance with official methodology. Results Bacterial filtration efficiency was comparable between sterilisation wrap and commercial surgical masks. Both prototypes met regulatory standards for synthetic blood resistance, whilst only our thinner mask fulfilled acceptable differential pressure (‘breathability’) thresholds. Conclusion Acceptable barrier and breathability properties can be achieved with surgical masks produced from sterilisation wrap. Therefore, this may be a reasonable method to supplement stock if required. Unless there are shortages mandating alternatives, health-care workers should always use approved personal protective equipment., Highlights • During the COVID-19 pandemic there has been regional shortages in surgical masks. • Some services have proposed using surgical masks manufactured from sterilisation wrap. • There has been limited assessment of the safety of this practice. • We developed prototypes and evaluated whether they met regulatory standards. • Acceptable barrier and breathability properties were achieved.

Previous studies have shown that anaerobic digestion of food waste can be enhanced by addition of conductive materials that stimulate direct interspecies electron transfer (DIET) between bacteria and methanogens. However, at extremely high organic loading rates (OLRs), volatile fatty acids (VFAs) still tend to accumulate even in the presence of conductive materials because of an imbalance between the formation of fermentation products and the rate of methanogenesis. In this study, granular activated carbon (GAC) immobilized with riboflavin (GAC-riboflavin) was added to an anaerobic digester treating food waste. The GAC-riboflavin reactor operated stably at OLRs as high as 11.5 kgCOD/ (m

All fifteen patients with HLA-A*32:01 restricted vancomycin-induced DRESS, showed negative ex vivo responses to dalbavancin however two showed cross-reactivity to teicoplanin and telavancin. Adjunctive diagnostic testing should be considered to detect potential cross-reactivity amongst glycopeptides.

Essentials Uncontrolled clot breakdown with active bleeding can be seen in advanced cirrhosis. A literature review found little information on optimal management. We report a case of successful treatment with tranexamic acid for persistent subdural hematoma in this setting. A 50‐year‐old woman with advanced cirrhosis presented with spontaneous subdural hematoma. She had a worsening clinical course following craniotomy despite administration of multiple blood products. With elevation in D‐dimer, persistently low fibrinogen and poor response to factor/fibrinogen replacement therapies, we had a suspicion for uncontrolled fibrinolysis. A literature review was conducted on treatment of hyperfibrinolysis in cirrhosis, finding 4 reports in which antifibrinolytics were used to control bleeding with different outcomes. The dose of tranexamic acid used in our patient was employed from previous experience in trauma patients. We transitioned from intravenous to oral administration based on expected pharmacokinetics. Our patient had a successful outcome with resolution of bleeding.

An ancient feature resembling a shod human footprint was recently discovered adjacent to a buried prehistoric housepit dating to ~1840 cal yr BP at the Swan Point site in central Alaska. Recovery of footprints in this context is rare, prompting us to question if the impression was indeed the result of human activity. We tested if the feature’s morphometrics fell within the mean biometrics of a shod human footprint using photogrammetric modeling techniques and classic landmark analyses. The feature was extensively photographed throughout the excavation, and the photos were used to produce three-dimensional photogrammetric digital models from which to obtain landmark biometrics. The results suggest that the feature falls within the expected mean parameters of a prehistoric human footprint. We characterized the footprint as belonging to a healthy, non-obese 9-12-year-old child. The presence of a child and well-constructed house in associated context suggests a healthy prehistoric family. The footprint represents the first of its kind from the subarctic, and the first described from Alaska.

Introduction: The COVID-19 pandemic and restrictions placed on movement to prevent its transmission has led to a surge in demand for remote medical care. We investigated whether COVID-Care, a patient-reported telehealth symptom monitoring system, was successful at delivering safe monitoring and care for these patients leading to decreased hospital presentations. Methods: We performed a single centre, prospective, interventional cohort study with symptomatic outpatients who presented for COVID-19 screening at Austin Health, Australia. Participants were invited to take part in the COVID-Care program, entering common COVID-19 symptoms on a purpose-built, online survey monitored by infectious diseases physicians, and matched with clinical data including date of symptom onset, hospital admission, and screening clinic presentations. Results: 42,158 COVID-19 swabs were performed in 31,626 patients from March to October 2020, with 414 positive cases. 20,768 people used the COVID-Care survey at least once. COVID-Care users were significantly younger than non-users. Of the 414 positive cases, 254 used COVID-Care, with 160 non-users. Excluding presentations on the same day or prior to the COVID-19 swab, there were 56 hospital presentations. 4.3% of COVID-Care users and 28.1% non-users were admitted to hospital or the Emergency Department (p

Chaperone proteins are required to maintain the overall fold and function of proteins in the cell. As part of the Hsp70 family, Ssa1 acts to maintain cellular proteostasis through a variety of diverse pathways aimed to preserve the native conformation of target proteins, thereby preventing aggregation and future states of cellular toxicity. Studying the structural dynamics of Ssa1 in vitro is essential to determining their precise mechanisms and requires the development of purification methods that result in highly pure chaperones. Current methods of expressing and purifying Ssa1 utilize affinity tagged constructs expressed in Escherichia coli, however, expression in an exogenous source produces proteins that lack post-translational modifications leading to undesired structural and functional effects. Current protocols to purify Ssa1 from Saccharomyces cerevisiae require large amounts of starting material, multiple steps of chromatography, and result in low yield. Our objective was to establish a small-scale purification of Ssa1 expressed from its endogenous source, Saccharomyces cerevisiae, with significant yield and purity. We utilized a protein A affinity tag that was previously used to purify large protein complexes from yeast, combined with magnetic Dynabeads that are conjugated with rabbit immunoglobulin G (IgG). Our results show that we can produce native, highly pure, active Ssa1 via this one-step purification with minimal amounts of starting material, and this Ssa1-protein A fusion does not alter cellular phenotypes. This methodology is a significant improvement in Ssa1 purification and will facilitate future experiments that will elucidate the biochemical and biophysical properties of Hsp70 chaperones.

Anaerobic digestion of municipal solid waste (MSW) incineration leachate is limited by high organic loading rates, high concentrations of complex organic matter, and the presence of compounds like ammonia, heavy metals, and salts that can inhibit microbial activity. The purpose of this study was to investigate whether the addition of magnetite could help reactors treating MSW incineration leachate overcome these limitations and promote efficient conversion of complex organic matter to methane. These studies showed that magnetite amendments improved chemical oxygen demand (COD) removal efficiencies (78.8% vs 89.0%) and methane production rates (3.7 m3STP/(m3·d) vs 4.8 m3STP/(m3·d)) at an organic loading rate (OLR) of 18.2 kg COD/(m3·d). Significant differences in microbial community structures were also observed between non-amended and magnetite-amended reactors. The majority of sequences from the magnetite-amended reactor clustered with acetogenic and mixed acid fermentative bacteria. Sequences from bacteria capable of extracellular electron transfer and methanogens from the genera Methanosarcina and Methanosaeta were also more abundant in magnetite-amended reactors. It is possible that these species were participating in direct interspecies electron transfer (DIET) facilitated by magnetite amendments. In addition, ammonia nitrogen, calcium, and heavy metals were lower in effluent collected from magnetite-amended reactors than non-amended controls, suggesting that magnetite adsorbed these inhibitory compounds and created an environment more amenable to microbial growth. These results are significant and should be used to develop strategies designed to optimize bio-methanogenic treatment of complex waste with elevated contaminant concentrations.

The metabotropic glutamatergic receptor subtype 5 (mGluR5) may represent a promising therapeutic target for stress-related psychiatric disorders. Here, we describe mGluR5 findings in stress disorders, particularly major depressive disorder (MDD), highlighting insights from positron emission tomography studies. Positron emission tomography studies report either no differences or lower mGluR5 in MDD, potentially reflecting MDD heterogeneity. Unlike the rapidly acting glutamatergic agent ketamine, mGluR5-specific modulation has not yet shown antidepressant efficacy in MDD and bipolar disorder. Although we recently showed that ketamine may work, in part, through significant mGluR5 modulation, the specific role of mGluR5 downregulation in ketamine's antidepressant response is unclear. In contrast to MDD, there has been much less investigation of mGluR5 in bipolar disorder, yet initial studies indicate that mGluR5-specific treatments may aid in both depressed and manic mood states. The direction of modulation needed may be state dependent, however, limiting clinical feasibility. There has been relatively little study of posttraumatic stress disorder or obsessive-compulsive disorder to date, although there is evidence for the upregulation of mGluR5 in these disorders. However, while antagonism of mGluR5 may reduce fear conditioning, it may also reduce fear extinction. Therefore, studies are needed to determine the role mGluR5 modulation might play in the treatment of these conditions. Further challenges in modulating this prevalent neurotransmitter system include potential induction of significant side effects. As such, more research is needed to identify level and type (positive/negative allosteric modulation or full antagonism) of mGluR5 modulation required to translate existing knowledge into improved therapies.

Background The human brain is inherently organized into distinct networks, as reported widely by resting-state functional magnetic resonance imaging (rs-fMRI), which are based on blood-oxygen-level-dependent (BOLD) signal fluctuations. 11C-UCB-J PET maps synaptic density via synaptic vesicle protein 2A, which is a more direct structural measure underlying brain networks than BOLD rs-fMRI. Methods The aim of this study was to identify maximally independent brain source networks, i.e., “spatial patterns with common covariance across subjects”, in 11C-UCB-J data using independent component analysis (ICA), a data-driven analysis method. Using a population of 80 healthy controls, we applied ICA to two 40-sample subsets and compared source network replication across samples. We examined the identified source networks at multiple model orders, as the ideal number of maximally independent components (IC) is unknown. In addition, we investigated the relationship between the strength of the loading weights for each source network and age and sex. Results Thirteen source networks replicated across both samples. We determined that a model order of 18 components provided stable, replicable components, whereas estimations above 18 were not stable. Effects of sex were found in two ICs. Nine ICs showed age-related change, with 4 remaining significant after correction for multiple comparison. Conclusion This study provides the first evidence that human brain synaptic density can be characterized into organized covariance patterns. Furthermore, we demonstrated that multiple synaptic density source networks are associated with age, which supports the potential utility of ICA to identify biologically relevant synaptic density source networks.

We contribute to the understanding of megafauna extinction and human dispersal in subarctic eastern Beringia by focusing on changes in the trophic dynamics of the large mammal community as well as the ecological role of humans as a predator and competitor. We reconstruct habitat use by megafauna and humans throughout the Pleistocene-Holocene boundary based on zooarchaeological data and stable isotope ratios of collagen. Our results are consistent with habitat heterogeneity and availability being important factors in the changing abundance of large herbivores. We argue that an increase in herbivore diversity and biomass at the beginning of the Bolling-Allerod interstadial and a relative lack of competitors favored the initial human colonization of subarctic eastern Beringia. As herbivore resources dwindled later in the Late Glacial, people increasingly relied on bison and wapiti. By efficiently extracting some of the highest-ranked resources in the landscape, people are likely to have contributed to the trophic displacement or regional extirpation of other large predators. The ecological patterns that we observe in subarctic eastern Beringia are consistent with a mixture of both top-down and bottom-up controls over biotic turnover.

Growth of bacterial and archaeal species capable of interspecies electron exchange was stimulated by addition of conductive materials (carbon cloth or granular activated carbon (GAC)) to anaerobic digesters treating dog food (a substitute for the dry-organic fraction of municipal solid waste (OFMSW)). Methane production (772-1428mmol vs

Recent archaeological and paleoecological work along both interior and coastal routes for early colonization of the New World has suggested that the interior route was impossible, leaving the coastal route as the only colonization route taken by Clovis ancestors. We review the geological, paleoecological, and archaeological record for Eastern Beringia and adjacent areas. Spatio-temporal patterning of known sites and evaluation of early interior and coastal route radiocarbon, luminescence, and cosmogenic dating, along with new analyses of obsidian distribution and adaptive strategies of early Beringians, indicate this assessment is premature and the interior route remains a viable hypothesis, available by at least 13,560 and possibly 14,900 years ago.

T-cell responses to SARS-CoV-2-derived peptide pools have been documented, however it remains largely unclear whether prominent SARS-CoV-2-specific T cell populations originate from naive or pre-existing memory sets. As HLA-B*07:02-restricted N105-113 epitope (B7/N105) is the most dominant SARS-CoV-2 CD8+ T-cell specificity to date, we dissected CD8+ T-cell responses directed at this epitope by direct ex vivo analyses in peripheral blood mononuclear cells (PBMCs) from acute and convalescent COVID-19 patients, and pre-pandemic PBMCs, tonsils, lungs and spleens. Using peptide-HLA tetramers, immunodominant B7/N105+CD8+ T-cells were detected at a high frequency (∼2.18x10-4) in COVID-19 patients, comparable to the well-established influenza-specific A2/M158+CD8+ T-cell population. Remarkably, frequencies of B7/N105+CD8+ T-cells were also readily detectable in pre-pandemic PBMCs and tonsils (at 6.55x10-5 and 2.76x10-4, respectively), although they mainly displayed a naive phenotype, indicating a lack of previous cross-reactive exposures. Ex vivo TCRαβ analyses revealed that a diverse TCRαβ repertoire and promiscuity in αβ TCR pairing underlie such high naive precursor frequencies of B7/N105+CD8+ T-cells. Overall, our study demonstrates that high precursor frequency and plasticity of TCRα-TCRβ pairing underpin immunodominance of SARS-CoV-2-specific B7/N105+CD8+ T-cell responses and advocates for vaccine strategies which include the nucleocapsid protein to elicit immunodominant CD8+ T-cell responses in HLA-B*07:02 individuals. Funding: This work was supported by theClifford Craig Foundation to KLF and KK, NHMRC Leadership Investigator Grant to KK (1173871), NHMRC Program Grant to DLD (#1132975), Research Grants Council of theHong Kong Special Administrative Region, China (#T11-712/19-N) to KK, the Victorian Government (SJK, AKW), MRFF award (#2002073) to SJK and AKW, MRFFAward (#1202445) to KK, NHMRC program grant 1149990 (SJK) and NHMRC project grant 1162760 (AKW). AKW is supported by Emerging Leadership 1 Investigator Grant (#1173433), JAJ by an NHMRC Early Career Fellowship (ECF) (#1123673), KK by NHMRC SeniorResearch Fellowship (1102792), DLD by a NHMRC Principal Research Fellowship(#1137285) and SJK by NHMRC Senior Principal Research Fellowship (#1136322). CES has received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement (#792532). JR is supported by an ARC Laureate fellowship. JRH and WZ are supported by the Melbourne Research Scholarship from The University of Melbourne. LH is supported by the Melbourne International Research Scholarship (MIRS) and the Melbourne International Fee Remission Scholarship (MIFRS) from The University of Melbourne. Ethical Approval: Experiments conformed to the Declaration of Helsinki Principles and theAustralian National Health and Medical Research Council Code of Practice. Written informed consents were obtained from all blood donors prior to the study. Lung and spleen tissues were obtained from deceased organ donors after written informed consents from the next of kin.Written informed consents were obtained from participants’ parental or guardians for underage tonsil tissue donors. The study was approved by the Alfred Hospital (#280/14), Austin Health (HREC/63201/Austin-2020); the University of Melbourne (#2057366.1, #2056901.1,#2056689, #2056761, #1442952, #1955465, and #1443389), the Australian Red CrossLifeblood (ID 2015#8), the Tasmanian Health and Medical (ID H0017479) and the James Cook University (H7886) Human Research Ethics Committees.

SARS-CoV-2 causes a spectrum of illness, ranging from asymptomatic to severe COVID-19. As the immunological basis for severity remains ill-defined, we analysed 78 SARS-CoV-2-infected individuals at acute and/or convalescent timepoints, up to 102 days post-symptom onset, quantifying 184 innate and adaptive immunological parameters. Acute COVID-19 in hospitalised patients was associated with high levels of IL-6, IL-18 and IL-10, elevated neutrophil-to-lymphocyte and neutrophil-to-T cell ratios, and high proportions of activated CD38+ neutrophils, CD38+ eosinophils, CD38+ HLA-DRlo monocytes, CD38+CD56dim NK cells, CD38+ γδ T cells, antibody-secreting cells, CD38+ICOS+ circulating T follicular helper (cTFH) cells, CD38+HLA-DR+CD4+ T cells, effector CD27-CD45RA+ and CD38+ CD8+ T cells. During convalescence, elevated seroconversion and neutralising antibodies were prominent and correlated with acute CXCR3+ cTFH cell activation. Strikingly, intensive care unit (ICU) patients with severe COVID-19 displayed higher levels of soluble IL-6R, IL-18, and hyperactivation of innate, adaptive and myeloid compartments than ward patients with moderate disease. Our analyses provide a comprehensive map of longitudinal immunological responses in COVID-19 patients during acute and convalescent phases of SARS-CoV-2 infection, and integrate key cellular pathways of complex perturbed immune networks that underpin severe COVID-19, providing important insights into potential biomarkers and immunotherapies. Funding: This work was supported by the Australian National Health and Medical Research Council (NHMRC) Leadership Investigator Grant to KK (#1173871), NHMRC Program Grant to DLD (#1132975), Research Grants Council of the Hong Kong Special Administrative Region, China (#T11-712/19-N) to KK, the Jack Ma Foundation to KK, KS, DIG and AWC, the a2 milk foundation to KS, the Victorian Government MRFF award (#2002073) to SJK, DIG and AKW, MRFF Award (#1202445) to KK, NHMRC program grant 1149990 (SJK), NHMRC project grant 1162760 (AKW), NHMRC Program Grant (#1071916) to KK and DCJ, NHMRC Program Grant (1113293) to DIG, the Merridew Foundation to NJCK. KK is supported by NHMRC Senior Research Fellowship (1102792), DLD by a NHMRC Principal Research Fellowship (#1137285), KS by an NHMRC Investigator grant (#1177174), SYCT by a NHMRC Career Development Fellowship (#1145033), DIG by an NHMRC Senior Principal Research Fellowship (#1117766) and SJK by NHMRC Senior Principal Research Fellowship (#1136322). SRL is supported by an NHMRC program grant (#1149990) and practitioner fellowship (#1135851). LH and WZ are supported by the Melbourne International Research Scholarship (MIRS) and the Melbourne International Fee Remission Scholarship (MIFRS) from The University of Melbourne. XJ is supported by China Scholarship Council-University of Melbourne joint Scholarship. CES has received funding from the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement (#792532). JRH and WZ are supported by the Melbourne Research Scholarship from The University of Melbourne. TMA and FM-W are International Society for Advancement of Cytometry Marylou Ingram Scholars. GHP is supported by the Australian Government Research Training Program (RTP) Scholarship. JAJ is supported by an NHMRC Early Career Fellowship (ECF) (#1123673). EBC. is supported by a NHMRC Peter Doherty Fellowship (#1091516). KK and AC were supported by the University of Melbourne Dame Kate Campbell Fellowship. The Melbourne WHO Collaborating Centre for Reference and Research on Influenza is supported by the Australian Government Department of Health. Conflict of Interest: The authors declare no conflict of interest. Ethical Approval: The study was approved by the Alfred Hospital (#280/14), Melbourne Health (HREC/17/MH/53), Monash Health (HREC/15/MonH/64/2016.196), The University of Melbourne (#2056689, #2056761, #1442952, #1955465 and #1443389) and James Cook University (H7886) Human Research Ethics Committees.

Biogas produced from anaerobic digestion usually contains 30%-50% CO2, much of which must be removed, before utilization. Bioelectrochemical biogas upgrading approaches show promise, however, they have not yet been optimized for practical applications. In this study, a bioelectrochemical system with low energy input (applied cathode potential of -0.5 V vs. standard hydrogen electrode, SHE) was used for in-situ biogas upgrading. High efficiency CO2 conversion (318.5 mol/d/m2) was achieved when the system was operated with an organic load of 1.7 kgCOD/(m3 d). Methane content in the upgraded biogas was 97.0% and CO2 concentrations stayed below 3%, which is comparable to biogas upgraded with more expensive and less sustainable physiochemical approaches. The high efficiency of this approach could likely be attributed to a significant enrichment of Methanothrix (92.7%) species on the cathode surface that were expressing genes involved in both acetogenic methanogenesis and direct electron transfer (DET). Electromethanogenesis by these organisms also increased proton consumption and created a higher pH that increased the solubility of CO2 in the bioreactor. In addition, CO2 removal from the biogas was likely further enhanced by an enrichment of Actinobacillus species known to be capable of CO2 fixation. Artificial neural network (ANN) models were also used to estimate CH4 production under different loading conditions. The ANN architecture with 10 neurons at hidden layers fit best with a mean square error of 6.06 × 10−3 and R2 of 0.99.

Background The use of in vivo and ex vivo diagnostic tools for delayed immune-mediated adverse drug reactions is currently ill defined. Objective To determine whether the combination of skin testing and/or IFN-γ enzyme-linked immunoSpot assay (ELISpot) can aid diagnosis of these allergy phenotypes. Methods Patients with antibiotic-associated severe delayed immune-mediated adverse drug reaction hypersensitivity, including Stevens-Johnson syndrome and toxic epidermal necrolysis, drug reaction with eosinophilia and systemic symptoms (DRESS), acute generalized exanthematous pustulosis, generalized bullous fixed drug eruption, and severe maculopapular exanthema, were prospectively recruited. In vivo testing was completed to the implicated drug(s), and ex vivo testing was performed with the patient's PBMCs stimulated with the relevant antibiotic concentrations for IFN-γ release ELISpot measurement. Results Eighty-one patients met the inclusion criteria, with DRESS (42; 51.9%) accounting for most cases. Among the 63 (78%) who had an ELISpot assay performed, 34 (54%) were positive to at least 1 implicated antibiotic (median spot-forming units/million cells, 99.5; interquartile range, 68-187), with glycopeptide being a strong predictor of positivity (adjusted odds ratio, 6.11; 95% CI, 1.74-21.42). In combination (in vivo and ex vivo), 51 (63%) of those tested were positive to an implicated antibiotic. For DRESS and severe maculopapular exanthema associated with penicillins and cephalosporins, this combination confirmed the culprit agent in 11 of the 12 cases and in 6 of 7 for DRESS associated with glycopeptides. Conclusions This study demonstrates that using in vivo in combination with ex vivo testing can enhance the diagnostic approach in these severe phenotypes by assisting with the identification of possible culprit antibiotics.

In this study, an Escherichia coli (E. coli) whole-cell biosensor for the specific detection of bioavailable arsenic was developed by placing a green fluorescent protein (GFP) reporter gene under the control of the ArsR1 (GSU2952) regulatory circuit from Geobacter sulfurreducens. E. coli cells only emitted green fluorescence in the presence of arsenite and were more sensitive to arsenite when they were grown in M9 supplemented medium compared to LB medium. Under optimal test conditions, the Geobacter arsR1 promoter had a detection limit of 0.01 μM arsenite and the GFP expression was linear within a range of 0.03–0.1 μM (2.25–7.5 μg/l). These values were well below World Health Organization’s drinking water quality standard, which is 10 μg/l. The feasibility of using this whole-cell biosensor to detect arsenic in water samples, such as arsenic polluted tap water and landfill leachate was verified. The biosensor was determined to be just as sensitive as atomic fluorescence spectrometry. This study examines the potential applications of biosensors constructed with Geobacter ArsR-Pars regulatory circuits and provides a rapid and cost-effective tool that can be used for arsenic detection in water samples.

Background: We performed whole-exome sequencing of pretreatment biopsies and examined whether genome-wide metrics of overall mutational load, clonal heterogeneity or alterations at variant, gene, and pathway levels are associated with treatment response and survival. Patients and methods: Two hundred and three biopsies from the NeoALTTO trial were analyzed. Mutations were called with MuTect, and Strelka, using pooled normal DNA. Associations between DNA alterations and outcome were evaluated by logistic and Cox-proportional hazards regression. Results: There were no recurrent single gene mutations significantly associated with pathologic complete response (pCR), except PIK3CA [odds ratio (OR) = 0.42, P = 0.0185]. Mutations in 33 of 714 pathways were significantly associated with response, but different genes were affected in different individuals. PIK3CA was present in 23 of these pathways defining a ` trastuzumab resistance-network' of 459 genes. Cases with mutations in this network had low pCR rates to trastuzumab (2/50, 4%) compared with cases with no mutations (9/16, 56%), OR = 0.035;P< 0.001. Mutations in the ` Regulation of RhoA activity' pathway were associated with higher pCR rate to lapatinib (OR = 14.8, adjusted P = 0.001), lapatinib_trastuzumab (OR = 3.0, adjusted P = 0.09), and all arms combined (OR v 3.77, adjusted P = 0.02). Patients (n = 124) with mutations in the trastuzumab resistance network but intact RhoA pathway had 2% (1/41) pCR rate with trastuzumab alone (OR = 0.026, P = 0.001) but adding lapatinib increased pCR rate to 45% (17/38, OR = 1.68, P = 0.3). Patients (n = 46) who had no mutations in either gene set had 6% pCR rate (1/15) with lapatinib, but had the highest pCR rate, 52% (8/15) with trastuzumab alone. Conclusions: Mutations in the RhoA pathway are associated with pCR to lapatinib and mutations in a PIK3CA-related network are associated with resistance to trastuzumab. The combined mutation status of these two pathways could define patients with very low response rate to trastuzumab alone that can be augmented by adding lapatinib or substituting trastuzumab with lapatinib.

Better understanding of the biology of heterotopic ossification (HO) formation will lead to treatment and prevention modalities that can be directed specifically at the cellular level. Early identification of HO precursor cells and target genes may provide prognostic value that guides individualized prophylactic treatment. Better understanding of molecular signaling and proteomics variability will allow surgeons to individualize preemptive treatment to suppress inflammation and formation of HO. Careful surgical technique to avoid muscle damage is important. Damaged muscle should be debrided as a prophylactic measure. Hemostasis and avoidance of a postoperative hematoma may decrease the chance of formation of HO.

Bio-methanogenic digestion of incineration leachate is hindered by high OLRs, which can lead to build-up of VFAs, drops in pH and ultimately in reactor souring. It was hypothesized that incorporation of carbon cloth into reactors treating leachate would promote DIET and enhance reactor performance. To examine this possibility, carbon cloth was added to laboratory-scale UASB reactors that were fed incineration leachate. As expected, the carbon-cloth amended reactor could operate stably with a 34.2% higher OLR than the control (49.4 vs 36.8 kg COD/(m 3 d)). Microbial community analysis showed that bacteria capable of extracellular electron transfer and methanogens known to participate in DIET were enriched on the carbon cloth surface, and conductivity of sludge from the carbon cloth amended reactor was almost twofold higher than sludge from the control (9.77 vs 5.47 μS/cm), suggesting that microorganisms in the experimental reactor may have been expressing electrically conductive filaments.

Reassortment of segmented viruses can be an important source of genetic diversity underlying viral evolution and emergence. Methods for the quantification of reassortment have been described but are often cumbersome and best suited for the analysis of reassortment between highly divergent parental strains. While it is useful to understand the potential of divergent parents to reassort, outcomes of such heterologous reassortment are driven by differential selection acting on the progeny and are typically strain specific. To quantify reassortment robustly, a system free of differential selection is needed. We have generated such a system for influenza A virus and for mammalian orthoreovirus by constructing well-matched parental viruses carrying small genetic tags. The method utilizes high-resolution melt technology for the identification of reassortant viruses. Ease of sample preparation and data analysis enables streamlined genotyping of a large number of virus clones. The method described here thereby allows quantification of the efficiency of reassortment and can be applied to diverse segmented viruses.

The H2-assisted biogas upgrading approach has recently attracted much interest as a low-cost and environmentally friendly alternative to commonly used ex-situ/ physiochemical biogas upgrading techniques. However, most studies conducted to date have been limited to anaerobic solid-waste treatment characterized by flocculant sludge and low organic loading rates (OLR). In an attempt to expand its application to high-rate anaerobic wastewater treatment, an innovative two-stage up-flow anaerobic sludge blanket reactor system was employed using anaerobic granular sludge. We found that the CH4 content of product gas was consistently >90% and that H2 and CO2 concentrations stayed below 5%, even when OLR was increased from 1 to 5 g L−1 d−1 and H2 feeding rates were increased from 0.13 to 0.63 g L−1 d−1. We were also able to show that CO (5–10%) in H2-rich syngas didn’t inhibit methanogenesis or had significant impact on microbial community structure, suggesting that H2-assisted biogas upgrading with H2-rich syngas is feasible.

Summary Background Patients with metastatic colon cancer (mCRC) are at increased risk of venous thromboembolism (VTE). Limited preclinical data suggest that the oncogene (KRAS) mutational status of the tumor represents a plausible clinical link to systemic hypercoagulability in cancer patients. Objectives To determine if a tumor genetic characteristic, KRAS mutational status, is associated with an increased risk of VTE in patients with mCRC. Patients/methods A retrospective cohort study of patients with mCRC and KRAS test results was conducted at multiple practice sites across New England in the United States. The primary outcome was a VTE event, defined as deep venous thrombosis (DVT) and/or pulmonary embolism (PE), either 6 months before or at any time after the diagnosis of mCRC. KRAS status (mutated vs. wild type) and other relevant predictors of thrombosis were collected. Results Of 172 histologically confirmed patients with mCRC, 40 developed a VTE (23.3%). Sixty-five patients (37.8%) had a mutant KRAS status. The incidence of VTE and DVT among patients with mutated KRAS was 32.3 and 23.1%, respectively. The corresponding incidence among patients with wild-type KRAS was 17.8 and 9.4%. Odd ratios for the association were 2.21 (95% CI, 1.08–4.53) for VTE and 2.62 (95% CI, 1.12–6.12) for DVT, and remained significant despite adjustment for Khorana score and bevacizumab use. Conclusion Tumor mutant KRAS status is associated with an increased risk of VTE in patients with mCRC. The tumor genetic profile may represent a novel and important risk factor for thrombosis in patients with cancer.

Highlights • Sediment influence on hydrate dissolution was studied in the field and lab • In the field, CH4(aq) concentrations were at or below theoretical saturation • Dissolution of natural hydrate blanketed by 15 cm thick sediment was measured • This covered hydrate dissolved at a slower rate than nearby exposed hydrates • In the lab, exposed hydrate dissolved faster than covered hydrate Abstract Dissolution rates of naturally occurring gas hydrates vary by orders of magnitude across studies suggesting that environmental factors may influence hydrate dissolution. To determine the role that sediment cover plays in hydrate dissolution, we used a mini-pore fluid array sampler (mPFA) to continuously collect sediment porewater adjacent to a hydrate outcrop and maintain it at in situ pressure for later analysis. This allowed us to measure in situ dissolved hydrocarbon concentrations in the porewater over time without sample loss due to degassing. We deployed the mPFA at a hydrate outcrop at Barkley Canyon on the Cascadia Margin for nine months. This novel approach yielded concentration data that were used to determine the steady-state dissolution rate of the hydrate outcrop and test predictions of the diffusion-control model for dissolution in the field. In the lab, we conducted a series of experiments with artificial hydrate to directly compare dissolution rates between exposed and sediment-covered hydrate. The dissolution rate of the natural hydrate outcrop covered with sediment was 0.06 cm y− 1. The laboratory experiments of sediment-covered hydrate yielded dissolution rates of 0.6 ± 0.5 cm y− 1 (n = 5). In both laboratory and field observations, the dissolution rate of hydrates exposed directly to bulk water (3.9 ± 1.7 cm y− 1 and 3.5 cm y− 1 respectively) was at least an order of magnitude faster than the dissolution rate of sediment covered hydrate. These results are consistent with expectations of diffusion-control and support this model of hydrate dissolution. In nature, sediment may account for the persistence of hydrate in otherwise methane-depleted environments by increasing the diffusive boundary layer and slowing the rate of molecular diffusion via porosity/tortuosity effects. We provide a number of “Lessons Learned” for improving the instrument design and for consideration during future studies.