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Background Barth syndrome is a rare, life-threatening X-linked recessive mitochondrial disorder of lipid metabolism primarily affecting males. Previous research suggests that bezafibrate may ameliorate cellular lipid abnormalities and reduce cardiac dysfunction in an animal model. Objectives Estimate the effect of bezafibrate on clinical, biochemical, and quality-of-life outcomes. Investigate whether within-participant clinical changes parallel in vitro changes in cardiolipin ratio/profile and mitochondrial morphology when each participant’s cells are cultured with bezafibrate. Investigate as for objective 2, culturing each participant’s cells with resveratrol. Describe the most feasible methods and standardised outcome measures to optimise the conduct of future trials and evaluations in Barth syndrome. Describe features of the research infrastructure which optimised recruitment, retention and communication with families and people with Barth syndrome. Describe the perceptions of participants and their families about the research and any important potential barriers to participation. Design Randomised, placebo-controlled, crossover trial of bezafibrate versus placebo. Setting NHS hospital providing UK-wide Barth Syndrome Service. Participants Males aged ≥ 6 years with a confirmed diagnosis of Barth syndrome with stable cardiac status, able to swallow tablets of bezafibrate/placebo. Exclusions were: hypersensitivity or allergy to bezafibrate or any component of bezafibrate; hepatic, liver or renal dysfunction; gallbladder disease; or recent deterioration in general health. Interventions Fifteen weeks of bezafibrate in one period and placebo in a second period, or vice versa (randomly allocated), with at least a 1-month washout between periods. Main outcome measures The primary outcome was peak VO2; secondary outcomes were cardiac function rest and exercise echocardiography and magnetic resonance imaging, cardiolipin ratio, quality of life, dynamic skeletal muscle P-magnetic resonance spectroscopy, mitochondrial studies and neutrophil counts, and adverse events. Outcomes were measured at baseline and the end of each period. Results Eleven males were studied; all attended all three assessments. There was no difference in peak VO2 between periods (0.66 ml/kg/min lower with bezafibrate than placebo, 95% confidence interval 2.34 to 1.03; p = 0.43). There was a trend towards a higher left ventricular ejection fraction with bezafibrate when measured by echocardiography but not magnetic resonance imaging, and better echocardiography-derived rest longitudinal and circumferential strain with bezafibrate. There was no difference in quality of life or cardiolipin ratio between periods. Skeletal muscle 31P magnetic resonance spectroscopy was performed cross-sectionally and showed a trend to higher Tau and lower Qmax indices in the bazafibrate group. Two participants had serious, expected adverse reactions when taking bezafibrate; otherwise, bezafibrate was well tolerated. Limitations The sample size was very small; the bezafibrate dose may have been too low or 15 weeks too short to observe an effect; measurements of mitochondrial content and membrane potential were highly variable; P-magnetic resonance spectroscopy was available only at the final assessment. Conclusions This study did not show significant improvement in the primary and secondary outcomes with bezafibrate treatment. Future work Elamipretide, studied in a small crossover trial in the USA, is another potential intervention which may be worth evaluating in an international study. Trial registration This trial is registered as ISRCTN58006579. Funding This award was funded by the National Institute for Health and Care Research (NIHR) Efficacy and Mechanism Evaluation (EME) programme (NIHR award ref: 12/205/56) and is published in full in Efficacy and Mechanism Evaluation; Vol. 11, No. 13. See the NIHR Funding and Awards website for further award information. Plain language summary Barth syndrome is a rare genetic syndrome with mutations in the tafazzin gene causing long-term effects in several organ systems, leading to significant lifelong comorbidities and a reduced quality of life. Barth syndrome shows an X-linked inheritance, hence it is almost exclusively present in males. Tafazzin gene mutations impair overall cell energy metabolism, and affected individuals show compromise of the immune system with a high risk for life-threatening infections, as well as cardiac dysfunction and cardiomyopathy, muscle and general fatigue, and generally reduced mobility also leading to psychological challenges and dysfunction. Currently, specific medical therapies are limited to symptomatic treatment, and drug therapies targeting the causative mitochondrial energy disturbance are urgently needed. This randomised controlled trial explored the use of oral bezafibrate, a licensed drug that can improve cell energy metabolism. Eleven male patients received either bezafibrate or a placebo for 4 months. Exercise capacity by cardiopulmonary exercise testing on a cycle ergometer, as the primary outcome measurement, as well as cardiac function by echocardiography and magnetic resonance imaging, and cellular anatomy and function by in vitro assays and microscopy, were measured before and after this 4-month period. Bezafibrate did not significantly improve exercise capacity, cardiac function or the cellular metabolism, although several cardiac function parameters showed slight improvements with bezafibrate treatment. No significant adverse effects were recorded. The limitations of the trial were its low participant number and the relatively short duration of the drug treatment phase. The trial did prove that complex randomised controlled drug trials are feasible in people with rare diseases. Adherence was good, with parents reporting few difficulties with pill swallowing. The qualitative interviews also found that the study was acceptable to participants and the burden of participating not too onerous. We hope the research can provide a template for future studies. Scientific summary Introduction Barth syndrome is a very rare, life-threatening, X-linked recessive genetic disease almost exclusively affecting young males, caused by abnormal lipids in mitochondria. The tafazzin gene (TAZ) encodes tafazzin, a phospholipid acyltransferase. Tafazzin transfers unsaturated fatty acids with acyl chains from phospholipids to monolysocardiolipin (MLCL) and regulates remodelling and maturation of cardiolipin, a phospholipid located exclusively in the inner mitochondrial membrane. Mature tetralinoleoyl-cardiolipin (L4-CL) is essential in the mitochondrial membrane for maintaining mitochondrial membrane potential, structural integrity of electron transport chain complexes and mitochondrial cristae architecture. Cardiolipin is a major constituent of inner mitochondrial membranes, and the major effects of the aberrant gene are on muscular tissues. Cardiac and skeletal manifestations of Barth syndrome are thought to result from impaired formation of respiratory chain super-complexes. TAZ mutations decrease L4-CL and increase intermediate species of MLCL, resulting in a marked increase in the MLCL : L4-CL ratio, which provides a highly sensitive and specific biochemical test for Barth syndrome. Barth syndrome is characterised by many gene mutations and profound phenotypic variability between affected patients within families and those in different families. The disease carries many risks and problems for its sufferers, as well as major healthcare costs. Almost one-third (30%) of living UK patients have undergone cardiac transplantation. Several patients who have died due to cardiac rejection or post-transplant lymphoproliferative disease had previously undergone cardiac transplantation. Other patients who have died deteriorated too fast to undergo transplantation or died of complications while awaiting transplantation. Neutropenia is another life-threatening issue. Two-thirds of UK patients are treated with granulocyte colony-stimulating factor by subcutaneous injection. Managing neutropenia in this disease is challenging, since patients have highly variable neutrophil counts, which prevents administration of a consistent daily dose and requires repeated blood counts and physician oversight. Exercise intolerance, lethargy and fatigue are universal, interfering with daily life, schoolwork and play and often necessitating use of a wheelchair. Patients cannot perform strenuous or demanding jobs. Many have major feeding problems from infancy through to adult life; for example, requiring feeding via gastrostomies. There can be rapid deterioration during periods of stable health despite expert medical care. Ventricular arrhythmia (tachycardia or fibrillation) affects 10% of adolescents and can cause sudden cardiac death at any time. There are no specific treatments for Barth syndrome. There is no evidence that standard medications for cardiomyopathy ameliorate long-term poor outcomes. Treatment is multidisciplinary and currently limited to treating intercurrent bacterial infections, bone marrow support, physiotherapy, nutritional support, management of heart failure and arrhythmia, and consideration for heart transplantation. Experiments using lymphoblasts from patients with Barth syndrome show that either bezafibrate or resveratrol can partially normalise the deranged cardiolipin ratio. This may be important, since some patients with Barth syndrome who lack neutropenia and tend to have good exercise tolerance have cardiolipin ratios intermediate between those of typical patients and normal individuals. Bezafibrate also improved left ventricular function and showed other benefits at supraphysiological doses in a TAZ knockdown mouse model. This trial was motivated by the need for disease-specific therapy to prevent morbidity, mortality, psychological distress, and disruption of quality of life (QoL) in affected patients and their families. An effective therapy also has the potential to generate savings for the NHS. Bezafibrate is a lipid-lowering drug used in adults and children, with a good safety record in long-term use. These characteristics made it a candidate for investigation. Bezafibrate has been reported to significantly ameliorate a mitochondrial myopathy in adults and shown promising results in several animal/human cellular models of mitochondrial disease. Several of the laboratory assays and clinical investigations proposed in this study were also considered potentially important beyond the scope of this specific study, to assess associations between genotype and phenotype and for future evaluations of therapies in Barth syndrome. Objectives Estimate the effect on clinical, biochemical and QoL outcome measures of bezafibrate compared to placebo. Investigate whether clinical improvements parallel in vitro changes in cardiolipin ratio and mitochondrial morphology in each participant’s cells when cultured with bezafibrate. Investigate whether clinical improvements and culture findings with bezafibrate parallel in vitro changes in each participant’s cells when exposed to resveratrol in laboratory culture. Describe the most feasible methods and standardised outcome measures for future trials. Describe features of the research infrastructure which optimised recruitment, retention and communication with participants and families. Describe participants’ and their families’ perceptions of research and important potential barriers to participation. Methods Study design and participants CARDIOlipin MANipulation trial (CARDIOMAN) was a double-blind, placebo-controlled crossover randomised trial. Treatment was given in two 15-week periods with a minimum of 1 month’s washout period between these when no treatment was given. Twenty-six males and one female with the disease were alive in the UK when the trial was designed. Males aged ≥ 6 years in the UK diagnosed with Barth syndrome were eligible. Exclusion criteria were: hypersensitivity to bezafibrate or component of the investigational medicinal product (IMP), photoallergic or phototoxic reaction to fibrate, unstable cardiac condition, cardiac chamber shortening fraction < 25% or significant decrease in shortening fraction in the preceding year, atrial or ventricular arrhythmia not stabilised by treatment, inability to swallow the IMP, hepatic dysfunction, renal impairment (creatinine clearance < 90 ml/min), gallbladder disease, recent unspecified significant deterioration in general health, prisoners and adults lacking capacity to provide informed consent. Randomisation and masking Participants were randomised to receive either bezafibrate then placebo, or the reverse order. Allocations were generated in advance by the Bristol Trials Centre and provided to the trial pharmacy which dispensed the IMP. All investigators, members of the research team (apart from pharmacy) and participants remained blind to treatment allocation. Intervention and procedures The intervention IMP was bezafibrate taken orally as 100-mg tablets. Children aged 6–9 years started on 100 mg once daily for the first 4 weeks and, if well tolerated, increased to 100 mg twice daily for the remaining 11 weeks. Children aged 10–17 years started on 200 mg once daily for the first 4 weeks and, if well tolerated, increased to 200 mg twice daily for the remaining 11 weeks. Adults (≥ 18 years) took 200 mg twice daily throughout. The comparator IMP was placebo taken orally as visually identical tablets and manufactured to be as similar as possible in taste and smell to the intervention. The IMP was prescribed at the start of the first period and again at the end of the first period for the second period. Outcomes All in vivo outcomes except magnetic resonance spectroscopy (MRS) were assessed three times: at baseline and in the final week of each treatment phase. The primary outcome measure was peak oxygen consumption on bicycle ergometry [peak volume of oxygen (VO2)]. Secondary outcomes were: MLCL/L4-CL ratio and cardiolipin profile in blood cells; phosphocreatine (PCr)/adenosine 5’ triphosphate (ATP) ratio in cardiac muscle on 31P MRS; skeletal muscle oxidative function/on 31P MRS; QoL [age-appropriate Pediatric Quality of Life Inventory (PedsQLTM) questionnaires]; absolute neutrophil count; amino acid expression (serum arginine and cysteine); cardiac function [left and right ventricular function: left ventricular ejection fraction (LVEF%), right ventricular ejection fraction (RVEF%); and 2-D longitudinal and circumferential strain at rest, peak exercise and after recovery]; mitochondrial size and number in lymphocytes; mitochondrial area in lymphocytes and as a proportion of cytoplasm area; mitochondrial function and cristae organisation in lymphocytes/neutrophils; arrhythmia profile from 12-lead electrocardiogram at rest and during exercise. Qualitative research methods explored participants’ and families’ experiences of the different interventions. Adverse events Serious adverse events (SAEs) and non-serious adverse events (AEs) were collected from participants from the time of consent until 1 month after the final period. Participants’ general practitioners were asked to inform the research team of suspected AEs. Participants were asked questions about SAEs/AEs at each research clinic visit and in monthly phone calls, including during the washout period. SAEs were reviewed by the Chief Investigator, who decided whether the SAE was related to bezafibrate. Statistical analysis A total of 20 males aged ≥ 6 years were potentially eligible. The sample size was dictated by the number of eligible boys willing to take part. The primary analysis estimated the difference in mean peak VO2 between placebo and bezafibrate periods (two-tailed 5% significance level). A sample size of 12 participants allowed the trial to detect a difference of 0.90 (within subject) standard deviations with 80% power. The statistical analysis plan (SAP) prespecified that all analyses should be conducted according to the intention-to-treat (ITT) principle. The ITT population comprised all participants and periods, according to the randomised allocations. The safety population comprised all randomised participants, according to the treatment received in each period, who received at least one dose of IMP. All statistical tests were two-sided and used a 5% significance level, except for interactions that used a 10% significance level. Treatment effects were estimated with 95% confidence intervals (CIs). No formal adjustment was made for multiple testing. The treatment effect for the primary outcome was analysed using mixed linear regression models, adjusting for period (fixed effect; exploring interactions where necessary) and participants (random effects). Model assumptions were tested using standard methods, and the carry-over effect was estimated by including treatment order. Treatment effects for continuously scaled secondary outcomes (prespecified in the SAP) were estimated and tested using the same methods. Results There was no statistically significant difference between bezafibrate and placebo in peak VO2, which was 0.66 ml/kg/min lower (95% CI −2.34 to 1.03; p = 0.43) with bezafibrate. There was a borderline statistically significant increase in LVEF% using echocardiography (3.72%, 95% CI −0.26% to 7.69%; p = 0.065) but no evidence of a difference when using magnetic resonance imaging (MRI) (0.15%, 95% CI −3.21% to 3.51%; p = 0.926). There was also no difference for RVEF% between bezafibrate and placebo (0.32, 95% CI −2.32 to 2.96; p = 0.801). Longitudinal and circumferential strain at rest were better (lower) with bezafibrate than with placebo; differences were –1.67 (95% CI −3.11 to −0.22; p = 0.026) and −2.72 (95% CI −5.03 to −0.40; p = 0.024) respectively. There was no difference in diastolic ratio at rest between bezafibrate and placebo, and no differences in strain or diastolic ratio at peak exercise or after 2 minutes’ recovery. All participants had sinus rhythm at rest and during exercise for all three time points. Magnetic resonance spectroscopy was only performed after the second period for 10 participants. PCr/ATP ratio in cardiac muscle was not collected. Cross-sectionally (five vs. five participants), MRS appeared to increase Tau and reduce Qmax parameters with bezafibrate compared to placebo. Monolysocardiolipin/L4-CL ratio did not differ between bezafibrate and placebo but deteriorated in the second period (20% higher, 95% CI 3% to 40%; p = 0.024). Neutrophil count also showed no difference between bezafibrate and placebo but increased in the second period (2.26, 95% CI 0.08 to 4.45; p = 0.043). Plasma cysteine increased with bezafibrate (32% higher, 95% CI 8% to 62%; p = 0.001) but plasma arginine did not (25% higher, 95% CI −10% to 72%; p = 0.180). Mitochondrial outcomes on blood samples did not show marked differences and were not formally compared. Quality-of-life scores were transformed to a 0–100 scale (higher being better). Bezafibrate had no effect on core (−3.41, 95% CI −8.67 to 1.86; p = 0.192) or fatigue (1.71, 95% CI −6.28 to 9.71; p = 0.659) QoL scores. Emerging themes identified in qualitative interviews were classified as overarching, related to lived experience with Barth syndrome, and trial participation and conduct. Participants and families were pleased to take part in the trial, viewing participation as ‘giving back’ to the medical community. No participant withdrew, and the trial was not perceived to be too onerous for families. Participating in the research gave both individuals and families a sense of responsibility for their condition, partly driven by appreciation for the Barth Syndrome Service and staff expertise. Monolysocardiolipin/L4-CL ratio in vitro was estimated from samples of cells incubated with control, bezafibrate or resveratrol. Each drug reduced the MLCL/L4-CL ratio, largely by increasing intermediate cardiolipin species, and more so with resveratrol than with bezafibrate. Ratios were nevertheless profoundly abnormal in comparison to patients with intermediate Barth phenotypes. Discussion Main findings: study results There was no clinically important or statistically significant improvement in peak VO2 with bezafibrate compared to placebo. There were inconsistent findings for LVEF% by method of measurement; echocardiography showed a borderline improvement that cardiac MRI did not demonstrate. Longitudinal and circumferential strain measurements were better at rest with bezafibrate, but not at peak exercise. There were no clinically important or statistically significant improvements in core or fatigue QoL domains. Plasma cysteine was significantly improved with bezafibrate treatment. Plasma arginine also increased but not significantly. Neutrophil count was unaffected and there was no significant change in cardiolipin ratio in participants’ blood with bezafibrate. There were no significant period effects except for MLCL/L4-CL ratio and neutrophil count, both increased in the second period. In vitro incubation of Epstein–Barr virus-transformed patient lymphoblasts with either bezafibrate or resveratrol resulted in improved MLCL/L4-CL ratios, an effect that was more profound with resveratrol. Treatment with this drug also resulted in a more obvious increase in intermediate MLCL species. Strengths and limitations Strengths of the trial are: randomisation and placebo blinding; crossover design; excellent retention; completeness of the data, good adherence and IMP tolerance. There were several limitations. The sample size (outside our control) was very small, and interpretation of effects for many secondary outcomes was challenging. Longer treatment duration or a higher bezafibrate dose might have produced better outcomes. Bezafibrate may have induced adverse reactions in some participants, inadvertently causing unblinding. The group of participants randomised to receive bezafibrate first was older than the group randomised to the opposite order, which may explain period effects. The electron microscopic and mitochondrial assessments proved problematic and were not completed in vitro. Measurements of mitochondrial content and membrane potential were highly variable. Amino acid levels were not assayed after fasting. Echocardiography images during exercise testing were often suboptimal. Lessons for the future Magnetic resonance spectroscopy in children is still exploratory, and protocols need to be better validated, but it is considered very promising as a potential objective functional outcome. Cardiolipin profile is not a good outcome for research, although it is appropriate for diagnosis because the Barth cardiolipin profile is so extremely different from a normal profile. Electron microscopy measurements of mitochondria should not be considered as outcomes in future trials. Peak VO2 should be the preferred current primary outcome because it is resistant to bias compared to other functional outcomes. Conclusion The trial did not show a significant treatment effect for the primary outcome, peak VO2. Some secondary outcomes such as systolic function measured by echocardiography and amino acid levels provided some evidence of benefit. The trial provided insights about trial methods and conduct in the context of a very rare paediatric disease that should inform future studies. Trial registration This trial is registered as ISRCTN58006579. Funding This award was funded by the National Institute for Health and Care Research (NIHR) Efficacy and Mechanism Evaluation (EME) programme (NIHR award ref: 12/205/56) and is published in full in Efficacy and Mechanism Evaluation; Vol. 11, No. 13. See the NIHR Funding and Awards website for further award information.

This research reveals the adsorption of cadmium (Cd2+) and zinc (Zn+2) from water using sewage sludge-derived biochar pyrolysed at 700 °C (SSB). The morphology and particle characteristics of SSB were characterised through scanning electron microscopy (SEM), particle size distribution (PSD), fourier transform infrared (FTIR), X-ray diffraction (XRD), and X-ray fluorescence (XRF). The adsorption study showed that the optimum contact times for removing Zn2+ and Cd2+ were 80 and 140 minutes, respectively. 95.51% Zn2+ and 97.54% Cd2+ could be removed from spiked solutions featuring 50 mg/L of Zn2+ and 50 mg/L Cd2+, each treated with 25 g/L biochar. The optimum pH of the solutions was 8–9 at a temperature of 40°C, indicating some precipitation of the metal ions at an alkaline pH. The highest adsorption capacity of SSB for Cd2+ and Zn2+ was found to be 3.02 and 2.51 mg/g, respectively, which compares favourably with other adsorbents. The isotherm studies confirmed experimental data to closely follow the Langmuir isotherm model at an R2 value of 0.9846 and 0.9816 for Cd2+ and Zn2+, respectively. The kinetic study confirmed the physical interaction between the adsorbents and the adsorbate. The spontaneous and exothermic nature of the process was confirmed by negative values of change in Gibbs free energy (ΔG) and enthalpy (ΔH). SSB could be regenerated for 6 cycles. Overall, this study explores sustainability, recycling, and waste management by offering SSB as a potentially cost-effective and environment-friendly solution to remove Cd2+ and Zn2+ from water.

The mechanisms driving dynamics of many epidemiologically important mosquito-borne pathogens are complex, involving combinations of vector and host factors (e.g., species composition and life-history traits), and factors associated with transmission and reporting. Understanding which intrinsic mechanisms contribute most to observed disease dynamics is important, yet often poorly understood. Ross River virus (RRV) is Australia's most important mosquito-borne disease, with variable transmission dynamics across geographic regions. We used deterministic ordinary differential equation models to test mechanisms driving RRV dynamics across major epidemic centers in Brisbane, Darwin, Mandurah, Mildura, Gippsland, Renmark, Murray Bridge, and Coorong. We considered models with up to two vector species (Aedes vigilax, Culex annulirostris, Aedes camptorhynchus, Culex globocoxitus), two reservoir hosts (macropods, possums), seasonal transmission effects, and transmission parameters. We fit models against long-term RRV surveillance data (1991-2017) and used Akaike Information Criterion to select important mechanisms. The combination of two vector species, two reservoir hosts, and seasonal transmission effects explained RRV dynamics best across sites. Estimated vector-human transmission rate (average β = 8.04x10-4per vector per day) was similar despite different dynamics. Models estimate 43% underreporting of RRV infections. Findings enhance understanding of RRV transmission mechanisms, provide disease parameter estimates which can be used to guide future research into public health improvements and offer a basis to evaluate mitigation practices.

Background: The work on the ESGB guidelines for diagnosis and treatment of biofilm infections began in 2012 and the result was published in 2014. The guidelines have been and still are frequently cited in the literature proving its usefulness for people working with biofilm infections. At the ESGB Biofilm conference in Mallorca 2022 (Eurobiofilms2022) the board of the ESGB decided to evaluate the 2014-guidelines and relevant publications since 2014 based on a lecture given at the Eurobiofilms2022. Guideline methods: The Delphi method for working on production of guidelines and the current ESCMID rules for guidelines are presented. The criteria for evaluation of relevant literature are very strict and especially for treatment, most clinicians and regulatory authorities require convincing results from Level I (randomized controlled trials) publications to justify changes of treatments. The relevant new biofilm literature and the relevant biofilm presentations from the Eurobiofilms meetings and ECCMID conferences was used for evaluating the contemporary relevance of the ESGB 2014 guidelines. Diagnosis of biofilm infections: Several infectious diseases have been recognized as biofilm infections since 2014, but the diagnostic methods and therapeutic strategies are still the same as recommended in the 2014 ESGB guidelines which are summarized in this opinion paper. Treatment of biofilm infections: Some promising new in vitro and in vivo (animal experiments) observations and reports for therapy of biofilm infections are mentioned, but they still await clinical trials. Conclusion: The interim opinion at the present time (2022) is therefore, that the guidelines do not need revision now, but there is a need for survey articles discussing new methods of diagnosis and treatment of biofilm infections in order - hopefully – to give inspiration to conduct clinical trials which may lead to progress in diagnosis and treatment of patients with biofilm infections.

Diagnosing biofilm infections has remained a constant challenge for the last 50 years. Existing diagnostic methods struggle to identify the biofilm phenotype. Moreover, most methods of biofilm analysis destroy the biofilm making the resultant data interpretation difficult. In this study we introduce Fourier Transform Infra-Red (FTIR) spectroscopy as a label-free, non-destructive approach to monitoring biofilm progression. We have utilised FTIR in a novel application to evaluate the chemical composition of bacterial biofilms without disrupting the biofilm architecture. S. epidermidis (RP62A) was grown onto calcium fluoride slides for periods of 30 min–96 h, before semi-drying samples for analysis. We report the discovery of a chemical marker to distinguish between planktonic and biofilm samples. The appearance of new proteins in biofilm samples of varying maturity is exemplified in the spectroscopic data, highlighting the potential of FTIR for identifying the presence and developmental stage of a single biofilm.

The global clinical and socioeconomic impact of chronic wounds is substantial. The main difficulty that clinicians face during the treatment of chronic wounds is the risk of infection at the wound site. Infected wounds arise from an accumulation of microbial aggregates in the wound bed, leading to the formation of polymicrobial biofilms that can be largely resistant to antibiotic therapy. Therefore, it is essential for studies to identify novel therapeutics to alleviate biofilm infections. One innovative technique is the use of cold atmospheric plasma (CAP) which has been shown to possess promising antimicrobial and immunomodulatory properties. Here, different clinically relevant biofilm models will be treated with cold atmospheric plasma to assess its efficacy and killing effects. Biofilm viability was assessed using live dead qPCR, and morphological changes associated with CAP evaluated using scanning electron microscopy (SEM). Results indicated that CAP was effective against Candida albicans and Pseudomonas aeruginosa, both as mono-species biofilms and when grown in a triadic model system. CAP also significantly reduced viability in the nosocomial pathogen, Candida auris. Staphylococcus aureus Newman exhibited a level of tolerance to CAP therapy, both when grown alone or in the triadic model when grown alongside C. albicans and P. aeruginosa. However, this degree of tolerance exhibited by S. aureus was strain dependent. At a microscopic level, biofilm treatment led to subtle changes in morphology in the susceptible biofilms, with evidence of cellular deflation and shrinkage. Taken together, these results indicate a promising application of direct CAP therapy in combatting wound and skin-related biofilm infections, although biofilm composition may affect the treatment efficacy.

Candida albicans is the most prevalent and notorious of the Candida species involved in bloodstream infections, which is characterised by its capacity to form robust biofilms. Biofilm formation is an important clinical entity shown to be highly variable among clinical isolates. There are various environmental and physiological factors, including nutrient availability which influence the phenotype of Candida species. However, mechanisms underpinning adaptive biofilm heterogeneity have not yet been fully explored.Within this study we have profiled previously characterised and phenotypically distinct C. albicans bloodstream isolates. We assessed the dynamic susceptibility of these differing populations to antifungal treatments using population analysis profiling in addition to assessing biofilm formation and morphological changes. High throughput methodologies of RNA-Seq and LC-MS were employed to map and integrate the transcriptional and metabolic reprogramming undertaken by heterogenous C. albicans isolates in response to biofilm and hyphal inducing serum.We found a significant relationship between biofilm heterogeneity and azole resistance (P

Abstract Repeated maximal breath‐holds have been demonstrated to induce bradycardia, increase haematocrit and haemoglobin and prolong subsequent breath‐hold duration by 20%. Freedivers use non‐maximal breath‐hold techniques (BHTs) to improve breath‐hold duration. The aim of this study was to investigate the cardiorespiratory and haematological responses to various BHTs. Ten healthy men (34.5 ± 1.9 years) attended five randomized experimental trials and performed a 40 min period of quiet rest or one of three BHTs followed by a maximal breath‐hold challenge during facial immersion in water at 30 or 10°C. Cardiovascular and respiratory parameters were measured continuously using finger plethysmography and breath‐by‐breath gas analysis, respectively, and venous blood samples were collected throughout. Facial immersion in cold water caused marked bradycardia (74.1 vs. 50.2 beats/min after 40 s) but did not increase breath‐hold duration compared with warm water control conditions. Facial immersion breath‐hold duration was 30.8–43.3% greater than the control duration when preceded by BHTs that involved repeated breath‐holds of constant duration (P = 0.021), increasing duration (P 0.05). In conclusion, the duration of apnoea can be extended by manipulating blood gases through repeated prior breath‐holds, but changes in cardiac output and red blood cell mass do not appear essential.

Abstract The Prostate, Lung, Colorectal and Ovarian (PLCO) Cancer Screening Trial is a prospective cohort study of nearly 155,000 U.S. volunteers aged 55–74 at enrollment in 1993–2001. We developed the PLCO Atlas Project, a large resource for multi-trait genome-wide association studies (GWAS), by genotyping participants with available DNA and genomic consent. Genotyping on high-density arrays and imputation was performed, and GWAS were conducted using a custom semi-automated pipeline. Association summary statistics were generated from a total of 110,562 participants of European, African and Asian ancestry. Application programming interfaces (APIs) and open-source software development kits (SKDs) enable exploring, visualizing and open data access through the PLCO Atlas GWAS Explorer website, promoting Findable, Accessible, Interoperable, and Re-usable (FAIR) principles. Currently the GWAS Explorer hosts association data for 90 traits and >78,000,000 genomic markers, focusing on cancer and cancer-related phenotypes. New traits will be posted as association data becomes available. The PLCO Atlas is a FAIR resource of high-quality genetic and phenotypic data with many potential reuse opportunities for cancer research and genetic epidemiology.

Introduction: Accurate and rapid diagnostics paired with effective tracking and tracing systems are key to halting the spread of infectious diseases, limiting the emergence of new variants and to monitor vaccine efficacy. The current gold standard test (RT-qPCR) for COVID-19 is highly accurate and sensitive, but is time-consuming, and requires expensive specialised, lab-based equipment.Methods: Herein, we report on the development of a SARS-CoV-2 (COVID-19) rapid and inexpensive diagnostic platform that relies on a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay and a portable smart diagnostic device. Automated image acquisition and an Artificial Intelligence (AI) deep learning model embedded in the Virus Hunter 6 (VH6) device allow to remove any subjectivity in the interpretation of results. The VH6 device is also linked to a smartphone companion application that registers patients for swab collection and manages the entire process, thus ensuring tests are traced and data securely stored.Results: Our designed AI-implemented diagnostic platform recognises the nucleocapsid protein gene of SARS-CoV-2 with high analytical sensitivity and specificity. A total of 752 NHS patient samples, 367 confirmed positives for coronavirus disease (COVID-19) and 385 negatives, were used for the development and validation of the test and the AI-assisted platform. The smart diagnostic platform was then used to test 150 positive clinical samples covering a dynamic range of clinically meaningful viral loads and 250 negative samples. When compared to RT-qPCR, our AI-assisted diagnostics platform was shown to be reliable, highly specific (100%) and sensitive (98–100% depending on viral load) with a limit of detection of 1.4 copies of RNA per µL in 30 min. Using this data, our CE-IVD and MHRA approved test and associated diagnostic platform has been approved for medical use in the United Kingdom under the UK Health Security Agency’s Medical Devices (Coronavirus Test Device Approvals, CTDA) Regulations 2022. Laboratory and in-silico data presented here also indicates that the VIDIIA diagnostic platform is able to detect the main variants of concern in the United Kingdom (September 2023).Discussion: This system could provide an efficient, time and cost-effective platform to diagnose SARS-CoV-2 and other infectious diseases in resource-limited settings.

Abstract Wounds can commonly become infected with polymicrobial biofilms containing bacterial and fungal microorganisms. Microbial colonization of the wound can interfere with sufficient healing and repair, leading to high rates of chronicity in certain individuals, which can have a huge socioeconomic burden worldwide. One route for alleviating biofilm formation in chronic wounds is sufficient treatment of the infected area with topical wound washes and ointments. Thus, the primary aim here was to create a complex in vitro biofilm model containing a range of microorganisms commonly isolated from the infected wound milieu. These polymicrobial biofilms were treated with three conventional anti-biofilm wound washes, chlorhexidine (CHX), povidone-iodine (PVP-I), and hydrogen peroxide (H2O2), and efficacy against the microorganisms assessed using live/dead qPCR. All treatments reduced the viability of the biofilms, although H2O2 was found to be the most effective treatment modality. These biofilms were then co-cultured with 3D skin epidermis to assess the inflammatory profile within the tissue. A detailed transcriptional and proteomic profile of the epidermis was gathered following biofilm stimulation. At the transcriptional level, all treatments reduced the expression of inflammatory markers back to baseline (untreated tissue controls). Olink technology revealed a unique proteomic response in the tissue following stimulation with untreated and CHX-treated biofilms. This highlights treatment choice for clinicians could be dictated by how the tissue responds to such biofilm treatment, and not merely how effective the treatment is in killing the biofilm.

The present work involves the study of the removal of Cu2+, Fe3+, Pb2+and Zn2+from synthetic metal solutions using natural zeolite. Laboratory experiments were used to investigate the efficiency of adsorbents in the uptake of heavy metals from industrial wastewater. The kinetic study was used to identify the effect of parameters that affect the rate of adsorption and evaluated their impact on the efficiency of the zeolite in the removal of heavy metals from industrial wastewater. Natural zeolite (clinoptilolite) as adsorbent contacted with multi-component synthetic solutions containing Cu2+, Fe3+, Pb2+and Zn2+ions without any pre-modifications and every hour 15 ml of the samples were filtered and taken for metal ion concentration analysis using the ICP-OES. The pH values were monitored and adjusted regularly. The results showed that the capacity of the adsorbents for the removal of heavy metals increased with a greater mass of absorbent, increased initial solution pH, increased agitation speed and higher solution concentration.

Background Helicobacter pylori (H. pylori), a bacterium implicated in the development of peptic ulcer and gastric cancer, is estimated to infect around half the world’s population. Its prevalence in Australia is unclear. This scoping review aimed to evaluate all Australian literature providing estimates of the prevalence of H. pylori. Methods Australian studies examining H. pylori prevalence from 1982 onwards were eligible for inclusion. Medline, Embase and Scopus databases, and grey literature sources, were searched. Two independent reviewers undertook a two-stage screening process. Data were extracted by two independent reviewers using a pre-specified template. Results Of 444 identified studies, 75 were included in the review. H. pylori prevalence in Australian population-based studies (n = 8) ranged from 38.0% in 1991 to 15.1% in 2002; however, estimated prevalence across all non-clinical population studies in diverse sub-groups (n = 29) has varied dramatically. Decreased prevalence has been more marked in populations with gastrointestinal symptoms and conditions compared to non-clinical populations. Data on H. pyloriprevalence in vulnerable populations are lacking. Conclusions This is the first scoping review of Australian studies reporting H. pylori prevalence. A wide range of study designs, population groups, geographic regions, and diagnostic methods was included, involving data collected over a 50-year period (1969 to 2018). The summary of H. pylori prevalence estimates over time in this review points to a decrease in prevalence in Australia, particularly among populations with gastrointestinal symptoms and illnesses; however, it is unknown whether there is inequity in prevalence trends across vulnerable sub-groups of the Australian population. Future research and interventions supporting the health and wellbeing of vulnerable populations is required to ensure equitable health gains are made for all.

The synthesis of zeolite materials by hydrothermal transformation of kaolin using a conventional hydrothermal method was investigated. Different analytical techniques were used to characterize the starting kaolin and produced zeolite A samples, including scanning electron microscopy (SEM), energy-dispersive spectroscopy (EDS), x-ray diffraction (XRD), x-ray fluorescence (XRF), thermogravimetric analysis (TGA), and Fourier transform infrared (FT-IR) spectroscopy. The synthetic zeolite type A was obtained after activation of kaolin and metakaolin followed by different thermal and chemical treatments. The metakaolinization phase was achieved by calcining the kaolin in air at 600°C for 3 hours, a much lower temperature than previously reported in the literature. Metakaolin was treated with 3 M sodium hydroxide solution at a ratio of 1:5 and, using stainless steel autoclaves with teflon liners, heated the mixture to 200°C in a microwave for 24 hours. The results from this synthesis route showed that zeolite A with a cubic crystal habit has been successfully synthesized.

Global concern regarding mosquito-borne disease emergence and re-emergence has driven the development of citizen science mosquito surveillance initiatives. Although these initiatives have shown great potential to assist local health authorities, ensuring outcomes are translatable to improved public health policy and practice remains challenging. Here we present a summary of citizen science mosquito surveillance programs worldwide, their focus, strategies, and outcomes, with a view to how best to apply this approach in their local areas. A scoping review of studies and reports was conducted through systematic search on electronic databases (Scopus, PubMed, Web of Science, and Google Scholar), grey literature, and other documents listed in the references of selected articles. A total of 33 citizen science studies included in this review described 29 citizen science mosquito surveillance projects operating in 16 countries, besides three programs with wide geographic coverage. The selected programs focused on different strategies and methods according to their local and national contextual needs. The majority of the programs reported being free or low in cost, and amenable to participants. Also, citizen scientists valued the opportunity to actively contribute to a scientific activity in which they saw value. Local and national programs have been successful in involving the broader public and yielding data on mosquito populations. However, to ensure the best public health outcomes, sustainability, and scalability, there is a need to continue engaging with stakeholders, including community members, researchers, public health agents, industry, and policymakers, and to bridge existing collaborations across different sectors.

Candida albicans is an opportunistic pathogen found throughout multiple body sites and is frequently co-isolated from infections of the respiratory tract and oral cavity with Staphylococcus aureus. Herein we present the first report of the effects that S. aureus elicits on the C. albicans transcriptome. Dual-species biofilms containing S. aureus and C. albicans mutants defective in ALS3 or ECE1 were optimised and characterised, followed by transcriptional profiling of C. albicans by RNA-sequencing (RNA-seq). Altered phenotypes in C. albicans mutants revealed specific interaction profiles between fungus and bacteria. The major adhesion and virulence proteins Als3 and Ece1, respectively, were found to have substantial effects on the Candida transcriptome in early and mature biofilms. Despite this, deletion of ECE1 did not adversely affect biofilm formation or the ability of S. aureus to interact with C. albicans hyphae. Upregulated genes in dual-species biofilms corresponded to multiple gene ontology terms, including those attributed to virulence, biofilm formation and protein binding such as ACE2 and multiple heat-shock protein genes. This shows that S. aureus pushes C. albicans towards a more virulent genotype, helping us to understand the driving forces behind the increased severity of C. albicans-S. aureus infections.

ABSTRACT Despite the strikingly high worldwide prevalence of vulvovaginal candidiasis (VVC), treatment options for recurrent VVC (RVVC) remain limited, with many women experiencing failed clinical treatment with frontline azoles. Further, the cause of onset and recurrence of disease is largely unknown, with few studies identifying potential mechanisms of treatment failure. This study aimed to assess a panel of clinical samples from healthy women and those with RVVC to investigate the influence of Candida, the vaginal microbiome, and how their interaction influences disease pathology. 16S rRNA sequencing characterized disease by a reduction in specific health-associated Lactobacillus species, such as Lactobacillus crispatus, coupled with an increase in Lactobacillus iners. In vitro analysis showed that Candida albicans clinical isolates are capable of heterogeneous biofilm formation, and we found the presence of hyphae and C. albicans aggregates in vaginal lavage fluid. Additionally, the ability of Lactobacillus to inhibit C. albicans biofilm formation and biofilm-related gene expression was demonstrated. Using RNA sequencing technology, we were able to identify a possible mechanism by which L. crispatus may contribute to re-establishing a healthy vaginal environment through amino acid acquisition from C. albicans. This study highlights the potential formation and impact of Candida biofilms in RVVC. Additionally, it suggests that RVVC is not entirely due to an arbitrary switch in C. albicans from commensal to pathogen and that understanding interactions between this yeast and vaginal Lactobacillus species may be crucial to elucidating the cause of RVVC and developing appropriate therapies. IMPORTANCE RVVC is a significant burden, both economically and for women's health, but its prevalence is poorly documented globally due to the levels of self-treatment. Identifying triggers for development and recurrence of VVC and the pathogenesis of the microbes involved could considerably improve prevention and treatment options for women with recurrent, azole-resistant cases. This study therefore aimed to examine the interkingdom dynamics from healthy women and those with RVVC using next-generation sequencing techniques and to further investigate the molecular interactions between C. albicans and L. crispatus in a relevant biofilm coculture system.

Background: Dengue is a significant climate-sensitive disease. Public health professionals play an important role in prevention and control of the disease. This study aimed to explore dengue control and prevention in the context of climate change in China. Methods: A cross-sectional survey was conducted among 630 public health professionals in 2015. Descriptive analysis and logistic regression were performed. Results: More than 80% of participants from southwest and central China believed climate change would affect dengue. However, participants from northeast China were less likely to believe so (65%). Sixty-nine percent of participants in Yunnan perceived that dengue had emerged/re-emerged in recent years, compared with 40.6% in Henan and 23.8% in Liaoning. Less than 60% of participants thought current prevention and control programs had been effective. Participants believed mosquitoes in high abundance, imported cases and climate change were main risk factors for dengue in China. Conclusion: There were varying views of dengue in China. Professionals in areas susceptible to dengue were more likely to be concerned about climate change and dengue. Current prevention and control strategies need to be improved. Providing more information for staff in lower levels of Centers for Disease Control and Prevention may help in containing a possible increase of dengue. Keywords: Climate change, Dengue control, Infectious disease, Public health professional, China

Introduction Cardiopulmonary exercise testing (CPET) provides a series of biomarkers, such as peak oxygen uptake, which could assess the development of disease status in interstitial lung disease (ILD). However, despite use in research and clinical settings, the feasibility of CPET in this patient group has yet to be established.Methods Twenty-six patients with ILD (19 male) were recruited to this study. Following screening for contraindications to maximal exercise, participants underwent an incremental CPET to volitional exhaustion. Feasibility of CPET was assessed by the implementation, practicality, acceptability and demand, thus providing clinical-driven and patient-driven information on this testing procedure.Results Of the 26 recruited participants, 24 successfully completed at least one CPET, with 67/78 prospective tests being completed. Contraindications included hypertension, low resting oxygen saturation and recent pulmonary embolism. Of the CPETs undertaken, 63% successfully reached volitional exhaustion, with 31% being terminated early by clinicians due to excessive desaturation. Quantitative and qualitative feedback from participants revealed a positive experience of CPET and desire for it to be included as a future monitoring tool.Conclusion CPET is feasible in patients with ILD. Identification of common clinical contraindications, and understanding of patient perspectives will allow for effective design of future studies utilising CPET as a monitoring procedure.

ABSTRACT Candida auris is an enigmatic yeast that provides substantial global risk in health care facilities and intensive care units. A unique phenotype exhibited by certain isolates of C. auris is their ability to form small clusters of cells known as aggregates, which have been to a limited extent described in the context of pathogenic traits. In this study, we screened several nonaggregative and aggregative C. auris isolates for biofilm formation, where we observed a level of heterogeneity among the different phenotypes. Next, we utilized an RNA sequencing approach to investigate the transcriptional responses during biofilm formation of a nonaggregative and aggregative isolate of the initial pool. Observations from these analyses indicate unique transcriptional profiles in the two isolates, with several genes identified relating to proteins involved in adhesion and invasion of the host in other fungal species. From these findings, we investigated for the first time the fungal recognition and inflammatory responses of a three-dimensional skin epithelial model to these isolates. In these models, a wound was induced to mimic a portal of entry for C. auris. We show that both phenotypes elicited minimal response in the model minus induction of the wound, yet in the wounded tissue, both phenotypes induced a greater response, with the aggregative isolate more proinflammatory. This capacity of aggregative C. auris biofilms to generate such responses in the wounded skin highlights how this opportunistic yeast is a high risk within the intensive care environment where susceptible patients have multiple indwelling lines. IMPORTANCE Candida auris has recently emerged as an important cause of concern within health care environments due to its ability to persist and tolerate commonly used antiseptics and disinfectants, particularly when attached to a surface (biofilms). This yeast is able to colonize and subsequently infect patients, particularly those that are critically ill or immunosuppressed, which may result in death. We have undertaken analysis on two different phenotypic types of this yeast, using molecular and immunological tools to determine whether either of these has a greater ability to cause serious infections. We describe that both isolates exhibit largely different transcriptional profiles during biofilm development. Finally, we show that the inability to form small aggregates (or clusters) of cells has an adverse effect on the organism’s immunostimulatory properties, suggesting that the nonaggregative phenotype may exhibit a certain level of immune evasion.

Abstract Background Though there was the significant decrease in the incidence of malaria in central and southwest China during the 1980s and 1990s, there has been a re-emergence of malaria since 2000. Methods A cross-sectional survey was conducted amongst the staff of eleven Centers for Disease Control and Prevention (CDC) in China to gauge their perceptions regarding the impacts of climate change on malaria transmission and its control and prevention. Descriptive analysis was performed to study CDC staff’s knowledge, attitudes, perceptions and suggestions for malaria control in the face of climate change. Results A majority (79.8%) of CDC staff were concerned about climate change and 79.7% believed the weather was becoming warmer. Most participants (90.3%) indicated climate change had a negative effect on population health, 92.6 and 86.8% considered that increasing temperatures and precipitation would influence the transmission of vector-borne diseases including malaria. About half (50.9%) of the surveyed staff indicated malaria had re-emerged in recent years, and some outbreaks were occurring in new geographic areas. The main reasons for such re-emergence were perceived to be: mosquitoes in high-density, numerous imported cases, climate change, poor environmental conditions, internal migrant populations, and lack of health awareness. Conclusions This study found most CDC staff endorsed the statement that climate change had a negative impact on infectious disease transmission. Malaria had re-emerged in some areas of China, and most of the staff believed that this can be managed. However, high densities of mosquitoes and the continuous increase in imported cases of malaria in local areas, together with environmental changes are bringing about critical challenges to malaria control in China. This study contributes to an understanding of climate change related perceptions of malaria control and prevention amongst CDC staff. It may help to formulate in-house training guidelines, community health promotion programmes and policies to improve the capacity of malaria control and prevention in the face of climate change in China.

Background: Cities are home to over half the global population; that proportion is expected to rise to 70% by mid-century. The urban environment differs greatly from that in which humans evolved, with potentially important consequences for health. Rates for allergic, inflammatory and auto-immune diseases appear to rise with urbanization and be higher in the more urbanized nations of the world which has led some to suggest that cities promote the occurrence of these diseases. However, there are no syntheses outlining what urban-associated diseases are and what characteristics of cities promote their occurrence. Objectives: To synthesize the current understanding of “urban-associated diseases”, and discover the common, potentially modifiable features of cities that may be driving these associations. Methods: We focus on any diseases that have been associated with cities or are particularly prominent in today’s urban societies. We draw on expertise across diverse health fields to examine the evidence for urban connections and drivers. Discussion: We found evidence for urban associations across allergic, auto-immune, inflammatory, lifestyle and infectious disease categories. Some conditions (e.g. obesity and diabetes) have complex relationships with cities that have been insufficiently explored. Other conditions (e.g. allergies and asthma) have more evidence demonstrating their relationship with cities and the mechanisms driving that association. Unsurprisingly, air pollution was the characteristic of cities most frequently associated with disease. Other identified urban risk factors are not as widely known: altered microbial exposure and a disconnect from environmental microbiomes, vitamin D deficiency, noise and light pollution, and a transient, over-crowded, impoverished population. However, many complexities and caveats to these relationships beg clarification; we highlight the current knowledge gaps and outline ways to fill those gaps. Identifying urban-associated diseases and their drivers will allow us to prepare for the urban-disease burden of the future and create healthy cities that mitigate that disease burden. Keywords: Urban, Health, Environmental microbiome, Environmental risk factors, Disease, Ecology

The synthesis of zeolite materials by hydrothermal transformation of kaolin using an alkaline fusion prior to hydrothermal synthesis method was investigated. The kaolin clay used in the present investigation was supplied from Iraq. The physical and chemical characterization of the starting kaolin and produced zeolite Na-LSX samples were carried out using different analytical techniques such as Scanning Electron Microscopy (SEM), Energy Dispersive Spectroscopy (EDS), X – Ray Diffraction (XRD), X – Ray Fluorescence (XRF), Thermogravimetric Analysis (TGA), Fourier Transform Infrared (FT-IR) Spectroscopy and Inductively Coupled Plasma-Optical Emission Spectrometer (ICP-OES). An alkaline fusion method was introduced prior to the hydrothermal treatment while kaolin clay powder was mixed manually with NaOH powder (ratio = 1/1.2 in weight). The metakaolinisation phase was achieved by calcining the mixture in air at 600 oC for 1hr. The result from this route shows that zeolite Na-LSX with cubic rounded edge crystal habit have been successfully synthesised. Finally, The kinetic study indicated the suitability of the zeolite Na-LSX for the removal of Cu2+, Fe3+, Pb2+ and Zn2+ ions from synthetic wastewater.

This study reports for the first time the geologic occurrence of natural zeolite A and associated minerals in mudstones from the Cretaceous Paja Formation in the urban area of the municipality of Vélez (Santander), Colombia. These rocks are mainly composed of quartz, muscovite, pyrophyllite, kaolinite and chlorite group minerals, framboidal and cubic pyrite, as well as marcasite, with minor feldspar, sulphates, and phosphates. Total organic carbon (TOC), total sulfur (TS), and millimeter fragments of algae are high, whereas few centimeters and not biodiverse small ammonite fossils, and other allochemical components are subordinated. Na–A zeolite and associated mineral phases as sodalite occur just beside the interparticle micropores (honeycomb from framboidal, cube molds, and amorphous cavities). It is facilitated by petrophysical properties alterations, due to processes of high diagenesis, temperatures up to 80–100 °C, with weathering contributions, which increase the porosity and permeability, as well as the transmissivity (fluid flow), allowing the geochemistry remobilization and/or recrystallization of pre-existing silica, muscovite, kaolinite minerals group, salts, carbonates, oxides and peroxides. X-ray diffraction analyses reveal the mineral composition of the mudstones and scanning electron micrographs show the typical cubic morphology of Na–A zeolite of approximately 0.45 mμ in particle size. Our data show that the sequence of the transformation of phases is: Poorly crystalline aluminosilicate → sodalite → Na–A zeolite. A literature review shows that this is an unusual example of the occurrence of natural zeolites in sedimentary marine rocks recognized around the world.

Background: Existing standardized biofilm assays focus on simple mono-species or bacterial-only models. Incorporating Candida albicans into complex biofilm models can offer a more appropriate and relevant polymicrobial biofilm for the development of oral health products. Aims: This study aimed to assess the importance of interkingdom interactions in polymicrobial oral biofilm systems with or without C. albicans, and test how these models respond to oral therapeutic challenges in vitro. Materials and Methods: Polymicrobial biofilms (two models containing 5 and 10 bacterial species, respectively) were created in parallel in the presence and absence of C. albicans and challenged using clinically relevant antimicrobials. The metabolic profiles and biomasses of these complex biofilms were estimated using resazurin dye and crystal violet stain, respectively. Quantitative PCR was utilized to assess compositional changes in microbial load. Additional assays, for measurements of pH and lactate, were included to monitor fluctuations in virulence “biomarkers.” Results: An increased level of metabolic activity and biomass in the presence of C. albicans was observed. Bacterial load was increased by more than a factor of 10 in the presence of C. albicans. Assays showed inclusion of C. albicans impacted the biofilm virulence profiles. C. albicans did not affect the biofilms’ responses to the short-term incubations with different treatments. Conclusions: The interkingdom biofilms described herein are structurally robust and exhibit all the hallmarks of a reproducible model. To our knowledge, these data are the first to test the hypothesis that yeasts may act as potential “keystone” components of oral biofilms.

Until vaccines and effective therapeutics become available, the practical solution to transit safely out of the current coronavirus disease 19 (CoVID-19) lockdown may include the implementation of an effective testing, tracing and tracking system. However, this requires a reliable and clinically validated diagnostic platform for the sensitive and specific identification of SARS-CoV-2. Here, we report on the development of a de novo, high-resolution and comparative genomics guided reverse-transcribed loop-mediated isothermal amplification (LAMP) assay. To further enhance the assay performance and to remove any subjectivity associated with operator interpretation of results, we engineered a novel hand-held smart diagnostic device. The robust diagnostic device was further furnished with automated image acquisition and processing algorithms and the collated data was processed through artificial intelligence (AI) pipelines to further reduce the assay run time and the subjectivity of the colorimetric LAMP detection. This advanced AI algorithm-implemented LAMP (ai-LAMP) assay, targeting the RNA-dependent RNA polymerase gene, showed high analytical sensitivity and specificity for SARS-CoV-2. A total of ~200 coronavirus disease (CoVID-19)-suspected NHS patient samples were tested using the platform and it was shown to be reliable, highly specific and significantly more sensitive than the current gold standard qRT-PCR. Therefore, this system could provide an efficient and cost-effective platform to detect SARS-CoV-2 in resource-limited laboratories.

ABSTRACT Candida auris has emerged as a significant global nosocomial pathogen. This is primarily due to its antifungal resistance profile but also its capacity to form adherent biofilm communities on a range of clinically important substrates. While we have a comprehensive understanding of how other Candida species resist and respond to antifungal challenge within the sessile phenotype, our current understanding of C. auris biofilm-mediated resistance is lacking. In this study, we are the first to perform transcriptomic analysis of temporally developing C. auris biofilms, which were shown to exhibit phase- and antifungal class-dependent resistance profiles. A de novo transcriptome assembly was performed, where sequenced sample reads were assembled into an ~11.5-Mb transcriptome consisting of 5,848 genes. Differential expression (DE) analysis demonstrated that 791 and 464 genes were upregulated in biofilm formation and planktonic cells, respectively, with a minimum 2-fold change. Adhesin-related glycosylphosphatidylinositol (GPI)-anchored cell wall genes were upregulated at all time points of biofilm formation. As the biofilm developed into intermediate and mature stages, a number of genes encoding efflux pumps were upregulated, including ATP-binding cassette (ABC) and major facilitator superfamily (MFS) transporters. When we assessed efflux pump activity biochemically, biofilm efflux was greater than that of planktonic cells at 12 and 24 h. When these were inhibited, fluconazole sensitivity was enhanced 4- to 16-fold. This study demonstrates the importance of efflux-mediated resistance within complex C. auris communities and may explain the resistance of C. auris to a range of antimicrobial agents within the hospital environment. IMPORTANCE Fungal infections represent an important cause of human morbidity and mortality, particularly if the fungi adhere to and grow on both biological and inanimate surfaces as communities of cells (biofilms). Recently, a previously unrecognized yeast, Candida auris, has emerged globally that has led to widespread concern due to the difficulty in treating it with existing antifungal agents. Alarmingly, it is also able to grow as a biofilm that is highly resistant to antifungal agents, yet we are unclear about how it does this. Here, we used a molecular approach to investigate the genes that are important in causing the cells to be resistant within the biofilm. The work provides significant insights into the importance of efflux pumps, which actively pump out toxic antifungal drugs and therefore enhance fungal survival within a variety of harsh environments.

Haemorrhagic fever with renal syndrome (HFRS) is a rodent-borne disease that poses a serious public health threat in China. HFRS is caused by hantaviruses, mainly Seoul virus in urban areas and Hantaan virus in agricultural areas. Although preventive measures including vaccination programs and rodent control measures have resulted in a decline in cases in recent years, there has been an increase in incidence in some areas and new endemic areas have emerged. This review summarises the recent literature relating to the effects of climatic factors on the incidence of HFRS in China and discusses future research directions. Temperature, precipitation and humidity affect crop yields, rodent breeding patterns and disease transmission, and these can be influenced by a changing climate. Detailed surveillance of infections caused by Hantaan and Seoul viruses and further research on the viral agents will aid in interpretation of spatiotemporal patterns and a better understanding of the environmental and ecological drivers of HFRS amid China's rapidly urbanising landscape and changing climate.

The use of natural compounds as an alternative source of antimicrobials has become a necessity given the growing concern over global antimicrobial resistance. Polyphenols, found in various edible plants, offers one potential solution to this. We aimed to investigate the possibility of using curcumin within the context of oral health as a way of inhibiting and preventing the harmful development of Candida albicans biofilms. We undertook a series of adsorption experiments with varying concentrations of curcumin, showing that 50 μg/ml could prevent adhesion. This effect could be further synergized by the curcumin pre-treatment of yeast cells to obtain significantly greater inhibition (>90%, p < 0.001). Investigation of the biological impact of curcumin showed that it preferentially affected immature morphological forms (yeast and germlings), and actively promoted aggregation of the cells. Transcriptional analyses showed that key adhesins were down-regulated (ALS1 and ALS3), whereas aggregation related genes (ALS5 and AAF1) were up-regulated. Collectively, these data demonstrated that curcumin elicits anti-adhesive effects and that induces transcription of genes integrally involved in the processes related to biofilm formation. Curcumin and associated polyphenols therefore have the capacity to be developed for use in oral healthcare to augment existing preventative strategies for candidal biofilms on the denture surface.

Extreme heat events will become more frequent under anthropogenic climate change, especially in Mediterranean ecosystems. Microhabitats can considerably moderate (buffer) the effects of extreme weather events and hence facilitate the persistence of some components of the biodiversity. We investigate the microclimatic moderation provided by two important microhabitats (cavities formed by the leaves of the grass-tree Xanthorrhoea semiplana F.Muell., Xanthorrhoeaceae; and inside the leaf-litter) during the summer of 2015/16 on the Fleurieu Peninsula of South Australia. We placed microsensors inside and outside these microhabitats, as well as above the ground below the forest canopy. Grass-tree and leaf-litter microhabitats significantly buffered against high temperatures and low relative humidity, compared to ground-below-canopy sensors. There was no significant difference between grass-tree and leaf-litter temperatures: in both microhabitats, daily temperature variation was reduced, day temperatures were 1-5°C cooler, night temperatures were 0.5-3°C warmer, and maximum temperatures were up to 14.4°C lower, compared to ground-below-canopy sensors. Grass-tree and leaf-litter microhabitats moderated heat increase at an average rate of 0.24°C temperature per 1°C increase of ambient temperature in the ground-below-canopy microhabitat. The average daily variation in temperature was determined by the type (grass-tree and leaf-litter versus ground-below-canopy) of microhabitat (explaining 67%), the amount of canopy cover and the area of the vegetation fragment (together explaining almost 10% of the variation). Greater canopy cover increased the amount of microclimatic moderation provided, especially in the leaf-litter. Our study highlights the importance of microhabitats in moderating macroclimatic conditions. However, this moderating effect is currently not considered in species distribution modelling under anthropogenic climate change nor in the management of vegetation. This shortcoming will have to be addressed to obtain realistic forecasts of future species distributions and to achieve effective management of biodiversity.

The synthesis of low-silica zeotypes by hydrothermal transformation of kaolinite-rich clay and the nucleation and growth processes of sodalite and cancrinite in the system Na2O-Al2O3-SiO2-H2O at 100 °C were investigated. The synthesis products were characterized by X-ray powder diffraction (XRPD), scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FT-IR), 29Si and 27Al Magic Angle Spinning Nuclear Magnetic Resonance (MAS-NMR) and thermogravimetric analysis (TGA). Our data show that the sequence of the transformation of phases is: Poorly crystalline aluminosilicate → zeolite LTA → sodalite → sodalite + cancrinite → cancrinite. Synthesized materials appeared stable thermodynamically under the experimental conditions, with zeolite LTA (a metastable phase) occurring as a minor phase, compared with the presence of sodalite and cancrinite.

Despite their clinical significance and substantial human health burden, fungal infections remain relatively under-appreciated. The widespread overuse of antibiotics and the increasing requirement for indwelling medical devices provides an opportunistic potential for the overgrowth and colonization of pathogenic Candida species on both biological and inert substrates. Indeed, it is now widely recognized that biofilms are a highly important part of their virulence repertoire. Candida albicans is regarded as the primary fungal biofilm forming species, yet there is also increasing interest and growing body of evidence for non-Candida albicans species (NCAS) biofilms, and interkingdom biofilm interactions. C. albicans biofilms are heterogeneous structures by definition, existing as three-dimensional populations of yeast, pseudo-hyphae, and hyphae, embedded within a self-produced extracellular matrix. Classical molecular approaches, driven by extensive studies of laboratory strains and mutants, have enhanced our knowledge and understanding of how these complex communities develop, thrive, and cause host-mediated damage. Yet our clinical observations tell a different story, with differential patient responses potentially due to inherent biological heterogeneity from specific clinical isolates associated with their infections. This review explores some of the recent advances made in an attempt to explore the importance of working with clinical isolates, and what this has taught us.

This paper investigates the physicomechanical properties of mortar containing high volume of fly ash (FA) as partial replacement of cement in presence of copolymer latexes. Portland cement (PC) was partially replaced with 0, 10, 20, 30 50, and 60% FA. Copolymer latexes were used based on 2-hydroxyethyl acrylate (2-HEA) and 2-hydroxymethylacrylate (2-HEMA). Testing included workability, setting time, absorption, chemically combined water content, compressive strength, and scanning electron microscopy (SEM). The addition of FA to mortar as replacement of PC affected the physicomechanical properties of mortar. As the content of FA in the concrete increased, the setting times (initial and final) were elongated. The results obtained at 28 days of curing indicate that the maximum properties of mortar occur at around 30% FA. Beyond 30% FA the properties of mortar reduce and at 60% FA the properties of mortar are lower than those of the reference mortar without FA. However, the addition of polymer latexes into mortar containing FA improved most of the physicomechanical properties of mortar at all curing times. Compressive strength, combined water, and workability of mortar containing FA premixed with latexes are higher than those of mortar containing FA without latexes.

The physical and mechanical properties of mortar containing synthetic cosurfactants as air entraining agent are investigated. The cosurfactants consist of a combination of 2% dodecyl benzene sodium sulfonate (DBSS) and either 1.5% polyvinyl alcohol (PVA) or 1.5% polyoxyethylene glycol monomethyl ether (POE). Also these cosurfactants were used to prepare copolymers latex: styrene/butyl methacrylate (St/BuMA), styrene/methyl methacrylate (St/MMA), and styrene/glycidyl methacrylate (St/GMA), in order to study their effects on the properties of mortar. The properties of mortar examined included flow table, W/C ratio, setting time, water absorption, compressive strength, and combined water. The results indicate that the latex causes improvement in mortar properties compared with cosurfactants. Also polymer latex containing DBSS/POE is more effective than that containing DBSS/PVA.

Investigations at Happisburgh, UK, have revealed the oldest known hominin footprint surface outside Africa at between ca. 1 million and 0.78 million years ago. The site has long been recognised for the preservation of sediments containing Early Pleistocene fauna and flora, but since 2005 has also yielded humanly made flint artefacts, extending the record of human occupation of northern Europe by at least 350,000 years. The sediments consist of sands, gravels and laminated silts laid down by a large river within the upper reaches of its estuary. In May 2013 extensive areas of the laminated sediments were exposed on the foreshore. On the surface of one of the laminated silt horizons a series of hollows was revealed in an area of ca. 12 m(2). The surface was recorded using multi-image photogrammetry which showed that the hollows are distinctly elongated and the majority fall within the range of juvenile to adult hominin foot sizes. In many cases the arch and front/back of the foot can be identified and in one case the impression of toes can be seen. Using foot length to stature ratios, the hominins are estimated to have been between ca. 0.93 and 1.73 m in height, suggestive of a group of mixed ages. The orientation of the prints indicates movement in a southerly direction on mud-flats along the river edge. Early Pleistocene human fossils are extremely rare in Europe, with no evidence from the UK. The only known species in western Europe of a similar age is Homo antecessor, whose fossil remains have been found at Atapuerca, Spain. The foot sizes and estimated stature of the hominins from Happisburgh fall within the range derived from the fossil evidence of Homo antecessor.

Taking the adjective mollis and related words (mollitia, mollire, etc.) as my example, I suggest how some of the tools used by linguists working in the area of lexical semantics can supplement the rich resources of lexicography (TLL) by shedding further light on the question of what it means to ask what a word “means” in general, and how this set of words in particular fits into the sex-gender system informing the Latin textual tradition. Out of the wide range of topics in lexical semantics, I here focus on two: sense-relations and polysemy. The paper concludes with an illustration of how use of these tools can add nuance to our reading of the language of mollitia in Catullus 16.

Fungal biofilm infections have become increasingly recognised as a significant clinical problem. One of the major reasons behind this is the impact that these have upon treatment, as antifungal therapy often fails and surgical intervention is required. This places a large financial burden on health care providers. This paper aims to illustrate the importance of fungal biofilms, particularly Candida albicans, and discusses some of the key fungal biofilm resistance mechanisms that include, extracellular matrix (ECM), efflux pump activity, persisters, cell density, overexpression of drug targets, stress responses, and the general physiology of the cell. The paper demonstrates the multifaceted nature of fungal biofilm resistance, which encompasses some of the newest data and ideas in the field.

Objective: Youths with disruptive behavior disorders, including conduct disorder and oppositional defiant disorder, show major impairments in reinforcement-based decision making. However, the neural basis of these difficulties remains poorly understood. This partly reflects previous failures to differentiate responses during decision making and feedback processing and to take advantage of computational model-based functional MRI (fMRI). Method: Participants were 38 community youths ages 10-18 (20 had disruptive behavior disorders, and 18 were healthy comparison youths). Model-based fMRI was used to assess the computational processes involved in decision making and feedback processing in the ventromedial prefrontal cortex, insula, and caudate. Results: Youths with disruptive behavior disorders showed reduced use of expected value information within the ventromedial prefrontal cortex when choosing to respond and within the anterior insula when choosing not to respond. In addition, they showed reduced responsiveness to positive prediction errors and increased responsiveness to negative prediction errors within the caudate during feed back. Conclusions: This study is the first to determine impairments in the use of expected value within the ventromedial prefrontal cortex and insula during choice and in prediction error-signaling within the caudate during feedback in youths with disruptive behavior disorders. [ABSTRACT FROM AUTHOR]

Abstract  We have used density-functional theory to investigate the neighboring-group stabilization of iodine, arsenic, and phosphorus-centered oxyanion moieties in species such as deprotonated 2-iodoxybenzoic acid (IBX) and its analogs. The magnitudes of different stabilizing effects and further candidates for analogous stabilization are analyzed. Figure The σ-hole colinear with the in-plane I-O bond in PhIO2 [ABSTRACT FROM AUTHOR]

Abstract  Previous investigations of Aedes aegypti response to human odor components have revealed a number of compounds that attract host-seeking females. However, such studies have utilized only a small number of long-term laboratory Ae. aegypti colonies. Using laboratory y-olfactometers, we studied the attraction of four different Ae. aegypti populations (North Queensland, Australia; Florida, USA; Singapore; and Minas Gerais, Brazil) to a key attractant compound from human skin, lactic acid. Combinations of lactic acid with ammonia and a fatty acid (caproic acid) were also investigated. The aims were to determine the extent of variation in lactic acid dose response among populations and to see whether all four populations responded equally to combinations of human odor components. Although all Ae. aegypti populations were attracted to lactic acid, there were differences in the threshold dose: Florida 0.03 μg/min, Singapore 0.17 μg/min, North Queensland 1.92 μg/min, and Brazil 10.27 μg/min. Attraction to lactic acid alone (maximum <40%) was significantly lower than for human odor (>87% for all populations). Significant increases in attraction were observed when lactic acid was combined with ammonia or caproic acid, although not for all populations. In addition, the highest doses of caproic acid tested decreased attraction when combined with lactic acid. The divergent responses to host kairomones seen here may be evidence of adaptation to locally available hosts in different parts of the geographic range of Ae. aegypti. [ABSTRACT FROM AUTHOR]

The objective of testing is to determine whether an implementation under test conforms to its specification. In distributed test architectures involving multiple remote testers, this objective can be complicated by the fact that testers may encounter coordination problems relating to controllability (synchronization) and observability during the application of tests. Based on a finite state machine (FSM) specification of the externally observable behaviour of a distributed system and a distinguishing sequence, this paper proposes a method for constructing a checking sequence where there is no potential controllability or observability problems, and where the use of external coordination message exchanges among testers is minimized. The proposed method does not assume a reliable reset feature in the implementations of the given FSM to be tested by the resulting checking sequence. [ABSTRACT FROM AUTHOR]

A series of lipophilic and hydrophilic fac tricarbonyl rhenium bisimine complexes have been prepared, their membrane-permeabilities explored in liposomes and their potential for application in fluorescence microscopy cell imaging demonstrated in the first application of MLCT-fluorescent rhenium complexes in cell imaging. [ABSTRACT FROM AUTHOR]

Abstract Face masks slow the spread of SARS-CoV-2, but it has been unknown how masks might reshape social interaction. One important possibility is that masks may influence how individuals communicate emotion through facial expressions. Here, we clarify to what extent—and how—masks influence facial emotion communication, through drift–diffusion modeling (DDM). Over two independent pre-registered studies, conducted three and 6 months into the COVID-19 pandemic, online participants judged expressions of 6 emotions (anger, disgust, fear, happiness, sadness, surprise) with the lower or upper face “masked” or unmasked. Participants in Study 1 (N = 228) correctly identified expressions above chance with lower face masks. However, they were less likely—and slower—to correctly identify these expressions relative to without masks, and they accumulated evidence for emotion more slowly—via decreased drift rate in DDM. This pattern replicated and intensified 3 months later in Study 2 (N = 264). These findings highlight how effectively individuals still communicate with masks, but also explain why they can experience difficulties communicating when masked. By revealing evidence accumulation as the underlying mechanism, this work suggests that time-sensitive situations may risk miscommunication with masks. This research could inform critical interventions to promote continued mask wearing as needed.

: Objectives Aspergillus fumigatus undergoes morphological transition throughout its growth and development. These changes have direct implications for the effectiveness of antifungal treatment. Here we report the in vitro antifungal activity of voriconazole, amphotericin B and caspofungin against three specific phases of multicellular development of A. fumigatus. : Methods A. fumigatus conidia were propagated for 8, 12 and 24 h prior to antifungal challenge. The resultant activity of the three agents tested was determined using an XTT reduction assay to assess both endpoint and time–kill susceptibility profiles. : Results Endpoint susceptibility testing demonstrated a time-dependent decrease in efficacy for all three antifungal agents as the complexity of the A. fumigatus hyphal structure developed. Overall, amphotericin B exhibited the best spectrum of activity at each phase of growth, but was comparable to voriconazole against germinated conidial growth (8 h). Later, both voriconazole and caspofungin were ineffective against complex mycelial structures (12 and 24 h). Time–kill studies demonstrated that amphotericin B was significantly more efficacious at reducing A. fumigatus metabolism than both voriconazole and caspofungin for all three growth phases examined, most notably after 1 h of drug exposure (P < 0.001). : Conclusions Overall, the data presented demonstrate that treatment of actively growing A. fumigatus cells with antifungal agents is more efficacious than treating mature structures in vitro. Amphotericin B was consistently more effective against each phase and displayed rapid effects, and therefore may be a suitable option for managing patient groups at risk from aspergillosis infections. [ABSTRACT FROM AUTHOR]

Background: There are suggestions that patients with attention-deficit/hyperactivity disorder (ADHD) show impairment in executive attention control and emotion regulation. This study investigated emotion regulation as a function of the recruitment of executive attention in patients with ADHD. Methods: Thirty-five healthy children/adolescents (mean age = 13.91) and twenty-six children/adolescents with ADHD (mean age = 14.53) participated in this fMRI study. They completed the affective Stroop paradigm viewing positive, neutral and negative images under varying cognitive loads. A 3-way ANOVA (diagnosis-by-condition-by-emotion) was conducted on the BOLD response data. Following this, 2 3-way ANOVAs (diagnosis-by-condition-by-emotion) were applied to context-dependent psychophysiological interaction (gPPI) analyses generated from a dorsomedial frontal cortex and an amygdala seed (identified from the BOLD response ANOVA main effects of condition and emotion respectively). Results: A diagnosis-by-condition interaction within dorsomedial frontal cortex revealed reduced recruitment of dorsomedial frontal cortex as a function of increased task demands in the children/adolescents with ADHD relative to healthy children/adolescents. The level of reduction in recruitment of dorsomedial frontal cortex was significantly correlated with symptom severity (total and hyperactivity) measured by Conner's Parent Report Scale in the children/adolescents with ADHD. In addition, analysis of gPPI data from a dorsomedial frontal cortex seed revealed significant diagnosis-by-condition interactions within lateral frontal cortex; connectivity between dorsomedial frontal cortex and lateral frontal cortex was reduced in the patients with ADHD relative to comparison youth during congruent and incongruent task trials relative to view trials. There were no interactions of group, or main effect of group, within the amygdala in the BOLD response ANOVA (though children/adolescents with ADHD showed increased responses to positive images within temporal cortical regions during task trials; identified by the diagnosis-by-condition-by-emotion interaction). However, analysis of gPPI data from an amygdala seed revealed decreased connectivity between amygdala and lentiform nucleus in the presence of emotional stimuli in children/adolescents with ADHD (diagnosis-by-emotion interaction). Conclusion: The current study demonstrated disrupted recruitment of regions implicated in executive function and impaired connectivity within those regions in children/adolescents with ADHD. There were also indications of heightened representation of emotional stimuli in patients with ADHD. However, as the findings were specific for positive stimuli, the suggestion of a general failure in emotion regulation in ADHD was not supported.