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Knowledge of temperature variation within and across beach-nesting bird habitat, and how such variation may affect the nesting success and survival of these species, is currently lacking. This type of data is furthermore needed to refine predictions of population changes due to climate change, identify important breeding habitat, and guide habitat restoration efforts. Thermal imagery collected with unmanned aerial vehicles (UAVs) provides a potential approach to fill current knowledge gaps and accomplish these goals. Our research outlines a novel methodology for collecting and implementing active thermal ground control points (GCPs) and assess the accuracy of the resulting imagery using an off-the-shelf commercial fixed-wing UAV that allows for the reconstruction of thermal landscapes at high spatial, temporal, and radiometric resolutions. Additionally, we observed and documented the behavioral responses of beach-nesting birds to UAV flights and modifications made to flight plans or the physical appearance of the UAV to minimize disturbance. We found strong evidence that flying on cloudless days and using sky-blue camouflage greatly reduced disturbance to nesting birds. The incorporation of the novel active thermal GCPs into the processing workflow increased image spatial accuracy an average of 12 m horizontally (mean root mean square error of checkpoints in imagery with and without GCPs was 0.59 m and 23.75 m, respectively). The final thermal indices generated had a ground sampling distance of 25.10 cm and a thermal accuracy of less than 1 °C. This practical approach to collecting highly accurate thermal data for beach-nesting bird habitat while avoiding disturbance is a crucial step towards the continued monitoring and modeling of beach-nesting birds and their habitat.

There is a growing demand for the collection of ultra-high spatial resolution imagery using unmanned aerial systems (UASs). UASs are a cost-effective solution for data collection on small scales and can fly at much lower altitudes, thus yielding spatial resolutions not previously achievable with manned aircraft or satellites. The use of commercially available software for image processing has also become commonplace due to the relative ease at which imagery can be processed and the minimal knowledge of traditional photogrammetric processes required by users. Commercially available software such as AgiSoft Photoscan and Pix4Dmapper Pro are capable of generating the high-quality data that are in demand for environmental remote sensing applications. We quantitatively assess the implications of processing parameter decision-making on UAS product accuracy and quality for orthomosaic and digital surface models for RGB and multispectral imagery. We iterated 40 processing workflows by incrementally varying two key processing parameters in Pix4Dmapper Pro, and conclude that maximizing for the highest intermediate parameters may not always translate into effective final products. We also show that multispectral imagery can effectively be leveraged to derive three-dimensional models of higher quality despite the lower resolution of sensors when compared to RGB imagery, reducing time in the field and the need for multiple flights over the same area when collecting multispectral data is a priority. We conclude that when users plan to use the highest processing parameter values, to ensure quality end-products it is important to increase initial flight coverage in advance.

Abstract Background With the approval of disease-modifying treatments (DMTs) for early Alzheimer’s disease (AD), there is an increased need for efficient and non-invasive detection methods for cerebral amyloid-β (Aβ) pathology. Current methods, including positron emission tomography (PET) and cerebrospinal fluid (CSF) analysis, are costly and invasive methods that may limit access to new treatments. Plasma tau phosphorylated at threonine-217 (P-tau217) presents a promising alternative, yet optimal cutoffs for treatment eligibility with DMTs like aducanumab require further investigation. This study evaluates the efficacy of one- and two-cutoff strategies for determining DMT eligibility at the Butler Hospital Memory & Aging Program (MAP). Methods In this retrospective, cross-sectional diagnostic cohort study, we first developed P-tau217 cutoffs using site-specific and BioFINDER-2 training data, which were then tested in potential DMT candidates from Butler MAP (total n = 150). ROC analysis was used to calculate the area under the curve (AUC) and accuracy of P-tau217 interpretation strategies, using Aβ-PET/CSF testing as the standard of truth. Results Potential DMT candidates at Butler MAP (n = 50), primarily diagnosed with mild cognitive impairment (n = 29 [58%]) or mild dementia (21 [42%]), were predominantly Aβ-positive (38 [76%]), and half (25 [50%]) were subsequently treated with aducanumab. Elevated P-tau217 predicted cerebral Aβ positivity in potential DMT candidates (AUC = 0.97 [0.92–1]), with diagnostic accuracy ranging from 0.88 (0.76–0.95, p = 0.028) to 0.96 (0.86–1, p

Summary: Background: Acute myeloid leukaemia (AML) is a bone marrow malignancy with poor prognosis. One of several treatments for AML is midostaurin combined with intensive chemotherapy (MIC), currently approved for FLT3 mutation-positive (FLT3-MP) AML. However, many patients carrying FLT3 mutations are refractory or experience an early relapse following MIC treatment, and might benefit more from receiving a different treatment. Development of a stratification method that outperforms FLT3 mutational status in predicting MIC response would thus benefit a large number of patients. Methods: We employed mass spectrometry phosphoproteomics to analyse 71 diagnosis samples of 47 patients with FLT3-MP AML who subsequently received MIC. We then used machine learning to identify biomarkers of response to MIC, and validated the resulting predictive model in two independent validation cohorts (n = 20). Findings: We identified three distinct phosphoproteomic AML subtypes amongst long-term survivors. The subtypes showed similar duration of MIC response, but different modulation of AML-implicated pathways, and exhibited distinct, highly-predictive biomarkers of MIC response. Using these biomarkers, we built a phosphoproteomics-based predictive model of MIC response, which we called MPhos. When applied to two retrospective real-world patient test cohorts (n = 20), MPhos predicted MIC response with 83% sensitivity and 100% specificity (log-rank p

Objective This study aims to develop a measurement model for health technology acceptability using a theoretical framework and a range of validated instruments to measure user experience, acceptance, usability, health and digital health literacy. Methods A cross-sectional evaluation study using a mixed-methods approach was conducted. An online survey was administered to patients who used a pulse oximeter in a virtual hospital setting during COVID-19. The model development was conducted in three steps: (1) exploratory factor analysis for conceptual model development, (2) measurement model confirmation through confirmatory factor analysis followed by structural equation modelling and (3) test of model external validity on four outcome measures. Finally, the different constructs of the developed model were used to compare two types of pulse oximeters by measuring the standardised scores. Results Two hundred and two participants were included in the analysis, 37.6% were female and the average age was 53 years (SD:15.38). A four-construct model comprising Task Load, Affective Attitude, Self-Efficacy and Value of Use (0.636–0.857 factor loadings) with 12 items resulted from the exploratory factor analysis and yielded a good fit (RMSEA = .026). Health and digital health literacy did not affect the overall reliability of the model. Frustration, performance, trust and satisfaction were identified as outcomes of the model. No significant differences were observed in the acceptability constructs when comparing the two pulse oximeter devices. Conclusions This article proposes a model for the measurement of the acceptability of health technologies used by patients in a remote care setting based on the use of a pulse oximeter in COVID-19 remote monitoring.

There is an urgent need for an effective TB vaccine capable of controlling both acute and chronic Mycobacterium tuberculosis infection in populations with diverse genetic backgrounds. In this study, we characterised the immunogenicity and protective efficacy of a novel protein-in-adjuvant subunit vaccine. The protein component is a fusion protein of three different M. tuberculosis antigens, which we termed CysVac5: CysD, a major component of the M. tuberculosis sulfate activation pathway that is highly expressed during the chronic stage of M. tuberculosis infection, is fused with two major secreted mycobacterial antigens, Ag85B and MPT83. Vaccination of C57BL/6 mice with CysVac5, formulated in a monophosphoryl lipid A (MPLA) and dimethyldioctadecylammonium (DDA) adjuvant combination, resulted in the potent generation of polyfunctional CD4+ T cells secreting multiple cytokines, including IFN-γ, IL-2, TNF and IL-17, against each of the three components of the fusion protein. Furthermore, vaccination with CysVac5-MPLA/DDA conferred significant protection against infection in mouse lungs, which was greater than that afforded by BCG at extended time points post-challenge. The generation of antigen-specific and protective immunity was also observed in CysVac5 vaccinated BALB/c mice, indicating the vaccine could display efficacy across multiple genetic backgrounds. These results indicate that the CysVac5 vaccine has broad immunogenicity, is effective in controlling both acute and chronic phases of M. tuberculosis infection in mice, and warrants further investigation to assess its potential to control pulmonary TB.

Summary: Background: Pneumonia is the leading cause of death in young children globally and is prevalent in the Papua New Guinea highlands. We investigated clinical predictors of hypoxic pneumonia to inform local treatment guidelines in this resource-limited setting. Methods: Between 2013 and 2020, two consecutive prospective observational studies were undertaken enrolling children 0–4 years presenting with pneumonia to health-care facilities in Goroka Town, Eastern Highlands Province. Logistic regression models were developed to identify clinical predictors of hypoxic pneumonia (oxygen saturation

Abstract INTRODUCTION We evaluated the accuracy of remote and in‐person digital tests to distinguish between older adults with and without AD pathological change and used the Montreal Cognitive Assessment (MoCA) as a comparison test. METHODS Participants were 69 cognitively normal older adults with known beta‐amyloid (Aβ) PET status. Participants completed smartphone‐based assessments 3×/day for 8 days, followed by TabCAT tasks, DCTclock™, and MoCA at an in‐person study visit. We calculated the area under the curve (AUC) to compare task accuracies to distinguish Aβ status. RESULTS Average performance on the episodic memory (Prices) smartphone task showed the highest accuracy (AUC = 0.77) to distinguish Aβ status. On in‐person measures, accuracy to distinguish Aβ status was greatest for the TabCAT Favorites task (AUC = 0.76), relative to the DCTclockTM (AUC = 0.73) and MoCA (AUC = 0.74). DISCUSSION Although further validation is needed, our results suggest that several digital assessments may be suitable for more widespread cognitive screening application.

BackgroundAs a response to the COVID-19 pandemic, the Sydney Local Health District in New South Wales, Australia, launched the rpavirtual program, the first full-scale virtual hospital in Australia, to remotely monitor and follow up stable patients with COVID-19. As part of the intervention, a pulse oximeter wearable device was delivered to patients to monitor their oxygen saturation levels, a critical indicator of COVID-19 patient deterioration. Understanding users’ perceptions toward the device is fundamental to assessing its usability and acceptability and contributing to the effectiveness of the intervention, but no research to date has explored the user experience of the pulse oximeter for remote monitoring in this setting. ObjectiveThis study aimed to explore the use, performance, and acceptability of the pulse oximeter by clinicians and patients in rpavirtual during COVID-19. MethodsSemistructured interviews and usability testing were conducted. Stable adult patients with COVID-19 (aged ≥18 years) who used the pulse oximeter and were monitored by rpavirtual, and rpavirtual clinicians monitoring these patients were interviewed. Clinicians could be nurses, doctors, or staff who were part of the team that assisted patients with the use of the pulse oximeter. Usability testing was conducted with patients who had the pulse oximeter when they were contacted. Interviews were coded using the Theoretical Framework of Acceptability. Usability testing was conducted using a think-aloud protocol. Data were collected until saturation was reached. ResultsTwenty-one patients (average age 51, SD 13 years) and 15 clinicians (average age 41, SD 11 years) completed the interview. Eight patients (average age 51, SD 13 years) completed the usability testing. All participants liked the device and thought it was easy to use. They also had a good understanding of how to use the device and the device’s purpose. Patients’ age and device use–related characteristics (eg, the warmth of hands and hand steadiness) were identified by users as factors negatively impacting the accurate use of the pulse oximeter. ConclusionsPatients and clinicians had very positive perceptions of the pulse oximeter for COVID-19 remote monitoring, indicating high acceptability and usability of the device. However, factors that may impact the accuracy of the device should be considered when delivering interventions using the pulse oximeter for remote monitoring. Targeted instructions about the use of the device may be necessary for specific populations (eg, older people and patients unfamiliar with technology). Further research should focus on the integration of the pulse oximeter data into electronic medical records for real-time and secure patient monitoring.

Current vaccines against SARS-CoV-2 reduce mortality but are less effective in preventing infection. Here the authors show that intranasal vaccination with a subunit vaccine including an TLR2-stimulating adjuvant induces strong neutralising antibody and T-cell responses against SARS-CoV-2 in the lungs that protect against infection.

Abstract Background The gut microbiome plays an important role in autoimmunity including multiple sclerosis and its mouse model called experimental autoimmune encephalomyelitis (EAE). Prior studies have demonstrated that the multiple sclerosis gut microbiota can contribute to disease, hence making it a potential therapeutic target. In addition, antibiotic treatment has been shown to ameliorate disease in the EAE mouse model of multiple sclerosis. Yet, to this date, the mechanisms mediating these antibiotic effects are not understood. Furthermore, there is no consensus on the gut-derived bacterial strains that drive neuroinflammation in multiple sclerosis. Results Here, we characterized the gut microbiome of untreated and vancomycin-treated EAE mice over time to identify bacteria with neuroimmunomodulatory potential. We observed alterations in the gut microbiota composition following EAE induction. We found that vancomycin treatment ameliorates EAE, and that this protective effect is mediated via the microbiota. Notably, we observed increased abundance of bacteria known to be strong inducers of regulatory T cells, including members of Clostridium clusters XIVa and XVIII in vancomycin-treated mice during the presymptomatic phase of EAE, as well as at disease peak. We identified 50 bacterial taxa that correlate with EAE severity. Interestingly, several of these taxa exist in the human gut, and some of them have been implicated in multiple sclerosis including Anaerotruncus colihominis, a butyrate producer, which had a positive correlation with disease severity. We found that Anaerotruncus colihominis ameliorates EAE, and this is associated with induction of RORγt+ regulatory T cells in the mesenteric lymph nodes. Conclusions We identified vancomycin as a potent modulator of the gut-brain axis by promoting the proliferation of bacterial species that induce regulatory T cells. In addition, our findings reveal 50 gut commensals as regulator of the gut-brain axis that can be used to further characterize pathogenic and beneficial host-microbiota interactions in multiple sclerosis patients. Our findings suggest that elevated Anaerotruncus colihominis in multiple sclerosis patients may represent a protective mechanism associated with recovery from the disease. Video Abstract

Recruiting large numbers of naïve lymphocytes to lymph nodes is critical for mounting an effective adaptive immune response. While most naïve lymphocytes utilize homing molecule L-selectin to enter lymph nodes, some circulating cells can traffic to the lung-draining mediastinal lymph node (mLN) through lymphatics via the intermediate organ, lung. However, whether this alternative trafficking mechanism operates in infection and contributes to T cell priming are unknown. We report that in pulmonary Mycobacterium tuberculosis-infected mice, homing of circulating lymphocytes to the mLN is significantly less efficient than to non-draining lymph node. CD62L blockade only partially reduced the homing of naïve T lymphocytes, consistent with L-selectin-independent routing of naïve lymphocytes to the site. We further demonstrated that lymphatic vessels in infected mLN expanded significantly and inhibiting lymphangiogenesis with a vascular endothelial growth factor receptor 3 kinase inhibitor reduced the recruitment of intravenously injected naïve lymphocytes to the mLN. Finally, mycobacterium-specific T cells entering via the L-selectin-independent route were readily activated in the mLN. Our study suggests that both L-selectin-dependent and -independent pathways contribute to naïve lymphocyte entry into mLN during M. tuberculosis infection and the latter pathway may represent an important mechanism for orchestrating host defence in the lungs.

Introduction Progress towards leprosy elimination is threatened by increasing incidence in ‘hot-spot’ areas where more effective control strategies are urgently required. In these areas, active case finding and leprosy prevention limited to known contacts is insufficient for control. Population-wide active case-finding together with universal prevention through mass drug administration (MDA) has been shown to be effective in ‘hot-spot’ areas, but is logistically challenging and expensive. Combining leprosy screening and MDA with other population-wide screening activities such as for tuberculosis may increase programme efficiency. There has been limited evaluation of the feasibility and effectiveness of combined screening and MDA interventions. The COMBINE study aims to bridge this knowledge gap.Methods and analysis This implementation study will assess the feasibility and effectiveness of active leprosy case-finding and treatment, combined with MDA using either single-dose rifampicin or rifamycin-containing tuberculosis preventive or curative treatment, for reducing leprosy incidence in Kiribati. The leprosy programme will run over 2022–2025 in concert with population-wide tuberculosis screening-and-treatment in South Tarawa. The primary research question is to what extent the intervention reduces the annual leprosy new case detection rate (NCDR) in adults and children compared with routine screening and postexposure prophylaxis (PEP) among close contacts (baseline leprosy control activities). Comparisons will be made with (1) the preintervention NCDR separably among adults and children in South Tarawa (before–after study) and (2) the corresponding NCDRs in the rest of the country. Additionally, the postintervention prevalence of leprosy obtained from a survey of a ‘hot-spot’ sub-population will be compared with prevalence documented during the intervention. The intervention will be implemented in collaboration with the Kiribati National Leprosy Programme.Ethics and dissemination Approval has been obtained from the Kiribati Ministry of Health and Medical Services (MHMS), the University of Otago (H22/111) and the University of Sydney (2021/127) Human Research Ethics Committees. Findings will be shared with the MHMS, local communities and internationally through publication.

Tuberculosis (TB), caused by Mycobacterium tuberculosis, results in approximately 1.6 million deaths annually. BCG is the only TB vaccine currently in use and offers only variable protection; however, the development of more effective vaccines is hindered by a lack of defined correlates of protection (CoP) against M. tuberculosis. Pulmonary vaccine delivery is a promising strategy since it may promote lung-resident immune memory that can respond rapidly to respiratory infection. In this study, CysVac2, a subunit protein previously shown to be protective against M. tuberculosis in mouse models, was combined with either Advax® adjuvant or a mixture of alum plus MPLA and administered intratracheally into mice. Peripheral immune responses were tracked longitudinally, and lung-local immune responses were measured after challenge. Both readouts were then correlated with protection after M. tuberculosis infection. Although considered essential for the control of mycobacteria, induction of IFN-γ-expressing CD4+ T cells in the blood or lungs did not correlate with protection. Instead, CD4+ T cells in the lungs expressing IL-17A correlated with reduced bacterial burden. This study identified pulmonary IL-17A-expressing CD4+ T cells as a CoP against M. tuberculosis and suggests that mucosal immune profiles should be explored for novel CoP.

Mycobacterium tuberculosis is a major human pathogen, and new vaccines are needed to prevent transmission. Mucosal vaccination may confer protection against M. tuberculosis by stimulating tissue-resident memory (TRM) CD4+ T cells in the lungs. The chemokine receptor CXCR3 promotes lung recruitment of T cells, but its role in TRM development is unknown. This study demonstrates the recombinant influenza A virus vaccine PR8.p25, expressing the immunodominant M. tuberculosis T cell epitope p25, induces CXCR3 expression on p25-specific CD4+ T cells in the lungs so that the majority of vaccine-induced CD4+ TRM expresses CXCR3 at 6 weeks. However, CXCR3−/− mice developed equivalent antigen-specific CD4+ T cell responses to wild-type (WT) mice following PR8.p25, and surprisingly retained more p25-specific CD4+ TRM in the lungs than WT mice at 6 weeks. The adoptive transfer of CXCR3−/− and WT P25 T cells into WT mice revealed that the initial recruitment of vaccine-induced CD4+ T cells into the lungs was independent of CXCR3, but by 6 weeks, CXCR3-deficient P25 T cells, and especially CXCR3−/− TRM, were significantly reduced compared to CXCR3-sufficient P25 T cells. Therefore, although CXCR3 was not essential for CD4+ TRM recruitment or retention, it provided a competitive advantage for the induction of M. tuberculosis-specific CD4+ TRM in the lungs following pulmonary immunization.

Abstract Objectives SARS‐CoV‐2 infection causes a spectrum of clinical disease presentation, ranging from asymptomatic to fatal. While neutralising antibody (NAb) responses correlate with protection against symptomatic and severe infection, the contribution of the T‐cell response to disease resolution or progression is still unclear. As newly emerging variants of concern have the capacity to partially escape NAb responses, defining the contribution of individual T‐cell subsets to disease outcome is imperative to inform the development of next‐generation COVID‐19 vaccines. Methods Immunophenotyping of T‐cell responses in unvaccinated individuals was performed, representing the full spectrum of COVID‐19 clinical presentation. Computational and manual analyses were used to identify T‐cell populations associated with distinct disease states. Results Critical SARS‐CoV‐2 infection was characterised by an increase in activated and cytotoxic CD4+ lymphocytes (CTL). These CD4+ CTLs were largely absent in asymptomatic to severe disease states. In contrast, non‐critical COVID‐19 was associated with high frequencies of naïve T cells and lack of activation marker expression. Conclusion Highly activated and cytotoxic CD4+ T‐cell responses may contribute to cell‐mediated host tissue damage and progression of COVID‐19. Induction of these potentially detrimental T‐cell responses should be considered when developing and implementing effective COVID‐19 control strategies.

An effective adaptive immune response depends on the organized architecture of secondary lymphoid organs, including the lymph nodes (LNs). While the cellular composition and microanatomy of LNs under steady state are well defined, the impact of chronic tissue inflammation on the structure and function of draining LNs is incompletely understood. Here we showed that Mycobacterium tuberculosis infection remodeled LN architecture by increasing the number and paracortical translocation of B cells. The formation of paracortical B lymphocyte and CD35+ follicular dendritic cell clusters dispersed CCL21-producing fibroblastic reticular cells and segregated pathogen-containing myeloid cells from antigen-specific CD4+ T cells. Depletion of B cells restored the chemokine and lymphoid structure and reduced bacterial burdens in LNs of the chronically infected mice. Importantly, this remodeling process impaired activation of naive CD4+ T cells in response to mycobacterial and unrelated antigens during chronic tuberculosis infection. Our studies reveal a mechanism in the regulation of LN microanatomy during inflammation and identify B cells as a critical element limiting the T cell response to persistent intracellular infection in LNs.

Background and objective Data on the prevalence of anti-tuberculous drug resistance and its association with genetic mutations in Mycobacterium tuberculosis are limited. Our study explores the genomics of tuberculosis in Ca Mau, Vietnam. Methods Patients ≥15 years in Ca Mau Province, Vietnam, were screened annually for tuberculosis between 2014 and 2017. Isolates underwent drug susceptibility testing (DST) using the breakpoint method. DNA was extracted and whole genome sequencing (WGS) was performed. Results We identified 365 positive sputum cultures for M. tuberculosis and processed 237 for DST and 265 for WGS. Resistance to isoniazid was present in 19.8% (95% CI 14.7 to 24.9%), rifampicin in 3.5% (1.1 to 5.7%) and ethambutol in 2.5% (0.9 to 5.4%) of isolates. Relevant mutations in rpoB gene were detected in 3.8% (1.8 to 6.8%). katG, inhA or fabG1 mutations were found in 19.6% (15.0 to 24.9%) with KatG being most common at 12.8% (9.1–17.5%). We found 38.4% of isolates were of Beijing lineage, 49.4% East-African-Indian lineage and 8.4% European-American lineage. There were no associations between resistance profiles and clinical features. Conclusion The high burden of isoniazid resistance and the katG mutation highlights the challenges facing Vietnam in its efforts to achieve its EndTB goals.

Workers in the mining and construction industries are at increased risk of respiratory and other diseases as a result of being exposed to harmful levels of airborne particulate matter (PM) for extended periods of time. While clear links have been established between PM exposure and the development of occupational lung disease, the mechanisms are still poorly understood. A greater understanding of how exposures to different levels and types of PM encountered in mining and construction workplaces affect pathophysiological processes in the airways and lungs and result in different forms of occupational lung disease is urgently required. Such information is needed to inform safe exposure limits and monitoring guidelines for different types of PM and development of biomarkers for earlier disease diagnosis. Suspended particles with a 50% cut-off aerodynamic diameter of 10 µm and 2.5 µm are considered biologically active owing to their ability to bypass the upper respiratory tract's defences and penetrate deep into the lung parenchyma, where they induce potentially irreversible damage, impair lung function and reduce the quality of life. Here we review the current understanding of occupational respiratory diseases, including coal worker pneumoconiosis and silicosis, and how PM exposure may affect pathophysiological responses in the airways and lungs. We also highlight the use of experimental models for better understanding these mechanisms of pathogenesis. We outline the urgency for revised dust control strategies, and the need for evidence-based identification of safe level exposures using clinical and experimental studies to better protect workers’ health.

Abstract Assessments of fish communities tend to rely on capture‐based methods that, due to sampling biases, can underestimate actual species richness. Alternatively, environmental DNA (eDNA) based metabarcoding is a noncapture approach that infers species richness and distribution by collecting and sequencing DNA present in the ecosystem. Here, eDNA metabarcoding was applied to the lower River Severn, a highly modified and impounded river, to identify the species present in the fish assemblage. Using a universal primer for fish (12S mtDNA region), comparisons were made between the species identified as present by eDNA metabarcoding versus long‐term data available from fisheries monitoring data based on capture methods. Depending on the stringency of detection thresholds applied, the two methods detected between 15 and 25 fish species present in the river, with the eDNA metabarcoding detecting most species previously reported in the capture surveys, although with differences in the relative abundance of species between the methods. Notably, eDNA metabarcoding detected species of high conservation importance that were never sampled by capture techniques, including native European shads (Alosa spp.). Differences in the similarity indices of species detection were greater between the sampling methods than between sampling sites on each river. These results highlight the high potential of eDNA metabarcoding to provide an effective monitoring tool for biodiversity and conservation in rivers, but also indicate the need for complementary multi‐method sampling for robust estimates of fish species richness.

Introduction Population-wide interventions offer a pathway to tuberculosis (TB) and leprosy elimination, but ‘real-world’ implementation in a high-burden setting using a combined approach has not been demonstrated. This implementation study aims to demonstrate the feasibility and evaluate the effect of population-wide screening, treatment and prevention on TB and leprosy incidence rates, as well as TB transmission.Methods and analysis A non-randomised ‘screen-and-treat’ intervention conducted in the Pacific atoll of South Tarawa, Kiribati. Households are enumerated and all residents ≥3 years, as well as children

Background: The Nav 1.8 sodium channel has a key role in generating repetitive action potentials in nociceptive human dorsal root ganglion neurons. Nav 1.8 is differentiated from other voltage-gated sodium channels by its unusually slow inactivation kinetics and depolarised voltage-dependence of activation. These features are particularly pronounced in the human Nav 1.8 channel and allow the channel to remain active during repolarisation. Gain-of-function mutations in Nav 1.8 have been linked to neuropathic pain and selective blockers of Nav 1.8 have been developed as potential new analgesics. However, it is not well understood how modulating the Nav 1.8 conductance alters neuronal excitability and how this depends on the balance of other ion channels expressed by nociceptive neurons. Methods: To investigate this, we developed a novel computational model of the human dorsal root ganglion neuron and used it to construct a population of models that mimicked inter-neuronal heterogeneity in ionic conductances and action potential morphology Results: By simulating changes to the Nav 1.8 conductance in the population of models, we found that moderately increasing the Nav 1.8 conductance led to increased firing rate, as expected, but increasing Nav 1.8 conductance beyond an inflection point caused firing rate to decrease. We found that the delayed rectifier and M-type potassium conductances were also critical for determining neuronal excitability. In particular, altering the delayed rectifier potassium conductance shifted the position of the Nav 1.8 inflection point and therefore the relationship between Nav 1.8 conductance and firing rate. Conclusions: Our results suggest that the effects of modulating Nav 1.8 in a nociceptive neuron can depend significantly on other conductances, particularly potassium conductances.

Introduction: The development of more effective tuberculosis (TB) vaccines is essential for the global control of TB. Recently, there have been major advances in the field, but an important hindrance remains the lack of correlates of protection against TB. This requires each vaccine candidate to undergo clinical efficacy trials based on data from animal protection studies, but the results from animal models do not necessarily predict efficacy in humans. Area covered: In this review we summarize our current knowledge of immune mechanisms that may contribute to protective immunity against TB following vaccination and relate these to protective efficacy in animal models and recent clinical trials. Although some initial trials did not reproduce protection against TB in humans, recent trials have demonstrated promising efficacy for three vaccine approaches. Expert opinion: Although CD4+ T lymphocytes are essential for protection against TB, there is no clear correlation between conventional CD4+ or CD8+ T cell responses and protective efficacy of TB vaccines. Recent attention has focused on other immune responses, including donor unrestricted T cells, B lymphocytes, and antibodies. Prospective studies on samples from vaccinated individuals protected in recent trials will allow evaluation of these alternative immune mechanisms as potential correlates of protection.

Pathogenic mycobacteria actively dysregulate protective host immune signalling pathways during infection to drive the formation of permissive granuloma microenvironments. Dynamic regulation of host microRNA (miRNA) expression is a conserved feature of mycobacterial infections across host-pathogen pairings. Here we examine the role of miR-206 in the zebrafish model of Mycobacterium marinum infection, which allows investigation of the early stages of granuloma formation. We find miR-206 is upregulated following infection by pathogenic M. marinum and that antagomir-mediated knockdown of miR-206 is protective against infection. We observed striking upregulation of cxcl12a and cxcr4b in infected miR-206 knockdown zebrafish embryos and live imaging revealed enhanced recruitment of neutrophils to sites of infection. We used CRISPR/Cas9-mediated knockdown of cxcl12a and cxcr4b expression and AMD3100 inhibition of Cxcr4 to show that the enhanced neutrophil response and reduced bacterial burden caused by miR-206 knockdown was dependent on the Cxcl12/Cxcr4 signalling axis. Together, our data illustrate a pathway through which pathogenic mycobacteria induce host miR-206 expression to suppress Cxcl12/Cxcr4 signalling and prevent protective neutrophil recruitment to granulomas.

Abstract Objectives Non‐sputum‐based tests to accurately identify active tuberculosis (TB) disease and monitor response to therapy are urgently needed. This study examined the biomarker capacity of a panel of plasma proteins alone, and in conjunction with a previously identified miRNA signature, to identify active TB disease. Methods The expression of nine proteins (IP‐10, MCP‐1, sTNFR1, RANTES, VEGF, IL‐6, IL‐10, TNF and Eotaxin) was measured in the plasma of 100 control subjects and 100 TB patients, at diagnosis (treatment naïve) and over the course of treatment (1‐, 2‐ and 6‐month intervals). The diagnostic performance of the nine proteins alone, and with the miRNA, was assessed. Results Six proteins were significantly up‐regulated in the plasma of TB patients at diagnosis compared to controls. Receiver operator characteristic curve analysis demonstrated that IP‐10 with an AUC = 0.874, sensitivity of 75% and specificity of 87% was the best single biomarker candidate to distinguish TB patients from controls. IP‐10 and IL‐6 levels fell significantly within one month of commencing treatment and may have potential as indicators of a positive response to therapy. The combined protein and miRNA panel gave an AUC of 1.00. A smaller panel of only five analytes (IP‐10, miR‐29a, miR‐146a, miR‐99b and miR‐221) showed an AUC = 0.995, sensitivity of 96% and specificity of 97%. Conclusions A novel combination of miRNA and proteins significantly improves the sensitivity and specificity as a biosignature over single biomarker candidates and may be useful for the development of a non‐sputum test to aid the diagnosis of active TB disease.

Short-chain fatty acids (SCFAs) are produced by microbial fermentation of dietary fiber in the gut. Butyrate is a particularly important SCFA with anti-inflammatory properties and is generally present at lower levels in inflammatory diseases associated with gut microbiota dysbiosis in mammals. We aimed to determine if SCFAs are produced by the zebrafish microbiome and if SCFAs exert conserved effects on zebrafish immunity as an example of the non-mammalian vertebrate immune system. We demonstrate that bacterial communities from adult zebrafish intestines synthesize all three main SCFA in vitro, although SCFA were below our detectable limits in zebrafish intestines in vivo. Immersion in butyrate, but not acetate or propionate, reduced the recruitment of neutrophils and M1-type pro-inflammatory macrophages to wounds. We found conservation of butyrate sensing by neutrophils via orthologs of the hydroxycarboxylic acid receptor 1 (hcar1) gene. Neutrophils from Hcar1-depleted embryos were no longer responsive to the anti-inflammatory effects of butyrate, while macrophage sensitivity to butyrate was independent of Hcar1. Our data demonstrate conservation of anti-inflammatory butyrate effects and identify the presence of a conserved molecular receptor in fish.

Cystic fibrosis (CF) is a genetic disorder causing dysfunctional ion transport resulting in accumulation of viscous mucus that fosters chronic bacterial biofilm-associated infection in the airways. Achromobacter xylosoxidans and Stenotrophomonas maltophilia are increasingly prevalent CF pathogens and while Burkholderia cencocepacia is slowly decreasing; all are complicated by multidrug resistance that is enhanced by biofilm formation. This study investigates potential synergy between the antibiotics ciprofloxacin (0.5–128 µg/mL), colistin (0.5–128 µg/mL) and tobramycin (0.5–128 µg/mL) when combined with the neutral pH form of N-Acetylcysteine (NACneutral) (0.5–16.3 mg/mL) against 11 cystic fibrosis strains of Burkholderia, Stenotrophomonas and Achromobacter sp. in planktonic and biofilm cultures. We screened for potential synergism using checkerboard assays from which fraction inhibitory concentration indices (FICI) were calculated. Synergistic (FICI ≤ 0.5) and additive (0.5 > FICI ≥ 1) combinations were tested on irreversibly attached bacteria and 48 h mature biofilms via time-course and colony forming units (CFU/mL) assays. This study suggests that planktonic FICI analysis does not necessarily translate to reduction in bacterial loads in a biofilm model. Future directions include refining synergy testing and determining further mechanisms of action of NAC to understand how it may interact with antibiotics to better predict synergy.

Currently available vaccines fail to provide consistent protection against tuberculosis (TB). New, improved vaccines are urgently needed for controlling the disease. The mycobacterial antigen fusions H4 (Ag85B-TB10.4) and H28 (Ag85B-TB10.4-Rv2660c) have been shown to be very immunogenic and have been considered as potential candidates for TB vaccine development. However, soluble protein vaccines are often poorly immunogenic, but augmented immune responses can be induced when selected antigens are delivered in particulate form. This study investigated whether the mycobacterial antigen fusions H4 and H28 can induce protective immunity when assembled into particulate vaccines (polyester nanoparticle-H4, polyester nanoparticle-H28, H4 nanoparticles and H28 nanoparticles). The particulate mycobacterial vaccines were assembled inside an engineered endotoxin-free production strain of Escherichia coli at high yield. Vaccine nanoparticles were purified and induced long-lasting antigen-specific T cell responses and protective immunity in mice challenged by aerosol with virulent Mycobacterium tuberculosis. A significant reduction of M. tuberculosis CFU, up to 0.7-log10 protection, occurred in the lungs of mice immunized with particulate vaccines in comparison to placebo-vaccinated mice (p < 0.0001). Polyester nanoparticles displaying the mycobacterial antigen fusion H4 induced a similar level of protective immunity in the lung when compared to M. bovis bacillus Calmette-Guérin (BCG), the currently approved TB vaccine. The safe and immunogenic polyester nanoparticle-H4 vaccine is a promising subunit vaccine candidate, as it can be cost-effectively manufactured and efficiently induces protection against TB.

Peripheral immune regulation is critical for the maintenance of self-tolerance. Here we have investigated signaling processes that distinguish T cells with regulatory capability from effector T cells. The murine Tg4 T cell receptor recognizes a peptide derived from the self-antigen myelin basic protein. T cells from Tg4 T cell receptor transgenic mice can be used to generate effector T cells and three types of T cells with regulatory capability, inducible regulatory T cells, T cells tolerized by repeated in vivo antigenic peptide exposure or T cells treated with the tolerogenic drug UCB9608 (a phosphatidylinositol 4 kinase IIIβ inhibitor). We comparatively studied signaling in all of these T cells by activating them with the same antigen presenting cells presenting the same myelin basic protein peptide. Supramolecular signaling structures, as efficiently detected by large-scale live cell imaging, are critical mediators of T cell activation. The formation of a supramolecular signaling complex anchored by the adaptor protein linker for activation of T cells (LAT) was consistently terminated more rapidly in Tg4 T cells with regulatory capability. Such termination could be partially reversed by blocking the inhibitory receptors CTLA-4 and PD-1. Our work suggests that attenuation of proximal signaling may favor regulatory over effector function in T cells.