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Research questionCan a strategy for scoring oocyte quality, based on cumulus cell (CC) gene expression, prioritize oocytes with the highest implantation potential, while limiting the number of embryos to be processed in culture and the number of supernumerary embryos to be vitrified?DesignAn interventional, blinded, prospective cohort study was retrospectively analyzed. In the original study, patients underwent a fresh Day3 single embryo transfer with embryos ranked based on morphology and CC gene expression (Aurora Test). The additional ranking of the embryos with the Aurora Test resulted in significant higher clinical pregnancy and live birth rates. Now it is investigated if the Aurora Test ranking could be applied to select oocytes. The effect of an Aurora Test based restriction to 2 and 3 2PN or MII oocytes on clinical pregnancy and other outcomes, was analyzed in two subsets of patients with all 2PN (n = 83) or all MII oocytes (n = 45) ranked.ResultsConsidering only the top three ranked 2PN oocytes, 95% of the patients would have received a fresh SET on Day3 resulting in 65% clinical pregnancies. This was not different from the pregnancy rate obtained in a strategy using all oocytes but significantly reduced the need for vitrification of supernumerary embryos by 3-fold. Considering only top-ranked MII oocytes gave similar results.ConclusionsIn countries with legal restrictions on freezing of embryos, gene expression of CC can be used for the selective processing of oocytes and would thus decrease the twin pregnancy rate and workload, especially for embryo morphology scoring and transfers as the handling and processing of lower competence oocytes is prevented, while improving the ART outcome.

An estimated 5.0 million children aged under 5 years died in 2020, with 82% of these deaths occurring in sub-Saharan Africa and southern Asia. Over one-third of Mumbai's population has limited access to healthcare, and child health outcomes are particularly grave among the urban poor. We describe the implementation of a digital technology-based child health programme in Mumbai and evaluate its holistic impact. Using an artificial intelligence (AI)-powered mobile health platform, we developed a programme for community-based management of child health. Leveraging an existing workforce, community health workers (CHW), the programme was designed to strengthen triage and referral, improve access to healthcare in the community, and reduce dependence on hospitals. A Social Return on Investment (SROI) framework is used to evaluate holistic impact. The programme increased the proportion of illness episodes treated in the community from 4% to 76%, subsequently reducing hospitalisations and out-of-pocket expenditure on private healthcare providers. For the total investment of Indian Rupee (INR) 2,632,271, the social return was INR 34,435,827, delivering an SROI ratio of 13. The annual cost of the programme per child was INR 625. Upskilling an existing workforce such as CHWs, with the help of AI-driven decision- support tools, has the potential to extend capacity for critical health services into community settings. This study provides a blueprint for evaluating the holistic impact of health technologies using evidence-based tools like SROI. These findings have applicability across income settings, offering clear rationale for the promotion of technology-supported interventions that strengthen healthcare delivery.

ObjectiveTimely identification of patients who carry multidrug-resistant microorganisms (MDRO) is needed to prevent nosocomial spread to other patients and to the hospital environment. We aimed to compare the yield of a universal screening strategy upon admission to the currently installed universal risk assessment combined with risk-based screening upon admission.MethodsThis observational study was conducted within a prospective cohort study. From January 1, 2018, until September 1, 2019, patients admitted to our hospital were asked to participate. Nasal and perianal samples were taken upon admission and checked for the presence of MDRO. The results of the universal risk assessment and risk-based screening were collected retrospectively from electronic health records.ResultsIn total, 1017 patients with 1069 separate hospital admissions participated in the study. Universal screening identified 38 (3.6%) unknown MDRO carriers upon admission (37 individual patients), all carrying extended-spectrum beta-lactamase-producing Enterobacterales. For 946 of 1069 (88.5%) patients, both the universal risk assessment and universal screening were performed. For 19 (2.0%) admissions, ≥1 risk factor was identified. The universal risk assessment identified one (0.1%) unknown carrier, compared to 37 out of 946 carriers for the universal screening (PConclusionsOur results show that in our low endemic setting, a universal screening strategy identified significantly more MDRO carriers than the currently implemented universal risk-assessment. When implementing a universal risk-assessment, risk factors should be carefully selected to be able to identify ESBL-E carriers. While the universal screening identified more MDRO carriers, further research is needed to determine the cost-effectiveness of this strategy.

Studies assessing patient-centred outcomes of novel rifampicin resistant tuberculosis (RR-TB) diagnostics are rare and mostly apply conventional methods which may not adequately address biases. Even though the Xpert MTB/RIF molecular assay was endorsed a decade ago for simultaneous diagnosis of tuberculosis and RR-TB, the impact of the assay on mortality among people with RR-TB has not yet been assessed. We analysed data of an observational prospective cohort study (EXIT-RIF) performed in South Africa. We applied a causal inference approach using inverse odds of sampling weights to rectify survivor bias and selection bias caused by differing screening guidelines. We also adjusted for confounding using a marginal structural model with inverse probability of treatment weights. We estimated the total effect of an RR-TB diagnosis made by the Xpert assay versus the pre-Xpert diagnostic algorithm (entailing a targeted Line Probe Assay (LPA) among TB-confirmed patients) on two-year mortality and we assessed mediation by RR-treatment initiation. Of the 749 patients diagnosed with RR-TB [247 (33%) by the pre-Xpert diagnostic algorithm and 502 (67%) by the Xpert assay], 42.7% died. Of these, 364 (48.6%) patients died in the pre-Xpert group and 200 (39.8%) in the Xpert group. People diagnosed with RR-TB by the Xpert assay had a higher odds of RR-TB treatment initiation compared to those diagnosed by the targeted LPA-based diagnostic process (OR 2.79; 95%CI 2.19–3.56). Receiving an RR-TB diagnosis by Xpert resulted in a 28% reduction in the odds of mortality within 2 years after presentation to the clinic (ORCI 0.72; 95%CI 0.53–0.99). Causal mediation analysis suggests that the higher rate of RR-TB treatment initiation in people diagnosed by the Xpert assay explains the effect of Xpert on 2-year mortality [natural indirect effect odds ratio 0.90 (95%CI 0.85–0.96). By using causal inference methods in combination with high quality observational data, we could demonstrate that the introduction of the Xpert assay caused a 28% reduction in 2-year odds of mortality of RR-TB. This finding highlights the need for advocacy for a worldwide roll-out of rapid molecular tests. Because the effect is mainly caused by increased RR-TB treatment initiation, health care systems should also ensure timely initiation of effective treatment upon an RR-TB diagnosis.

BackgroundAccess to drug resistant testing for tuberculosis (TB) remains a challenge in high burden countries. Recently, the World Health Organization approved the use of several moderate complexity automated nucleic acid amplification tests (MC-NAAT) that have performance profiles suitable for placement in a range of TB laboratory tiers to improve drug susceptibility tests (DST) coverage.MethodsWe conducted cost analysis of two MC-NAATs with different testing throughput: Lower Throughput (LT, < 24 tests per run) and Higher Throughput (HT, upto 90+ tests per run) for placement in a hypothetical laboratory in a resource limited setting. We used per-test cost as the main indicator to assess 1) drivers of cost by resource types and 2) optimized levels of annual testing volumes for the respective MC-NAATs.ResultsThe base-case per test cost of $18.52 (range: $13.79 - $40.70) for LT test and $15.37 (range: $9.61 - $37.40) for HT test. Per test cost estimates were most sensitive to the number of testing days per week, followed by equipment costs and TB-specific workloads. In general, HT NAATs were cheaper at all testing volume levels, but at lower testing volumes (less than 2,000 per year) LT tests can be cheaper if the durability of the testing system is markedly better and/or procured equipment costs are lower than that of HT NAAT.ConclusionAssuming equivalent performance and infrastructural needs, placement strategies for MC-NAATs need to be prioritized by laboratory system's operational factors, testing demands, and costs.

Background Access to drug resistant testing for tuberculosis (TB) remains a challenge in high burden countries. Recently, the World Health Organization approved the use of several moderate complexity automated nucleic acid amplification tests (MC-NAAT) that have performance profiles suitable for placement in a range of TB laboratory tiers to improve drug susceptibility tests (DST) coverage. Methods We conducted cost analysis of two MC-NAATs with different testing throughput: Lower Throughput (LT, < 24 tests per run) and Higher Throughput (HT, upto 90+ tests per run) for placement in a hypothetical laboratory in a resource limited setting. We used per-test cost as the main indicator to assess 1) drivers of cost by resource types and 2) optimized levels of annual testing volumes for the respective MC-NAATs. Results The base-case per test cost of $18.52 (range: $13.79 - $40.70) for LT test and $15.37 (range: $9.61 - $37.40) for HT test. Per test cost estimates were most sensitive to the number of testing days per week, followed by equipment costs and TB-specific workloads. In general, HT NAATs were cheaper at all testing volume levels, but at lower testing volumes (less than 2,000 per year) LT tests can be cheaper if the durability of the testing system is markedly better and/or procured equipment costs are lower than that of HT NAAT. Conclusion Assuming equivalent performance and infrastructural needs, placement strategies for MC-NAATs need to be prioritized by laboratory system’s operational factors, testing demands, and costs.

Accurate and user-friendly rapid point-of-care diagnostic tests (POCT) are needed to optimize treatment of Neisseria gonorrhoeae, especially in low-resource settings where syndromic management is the standard of care for sexually transmitted infections. This study aimed to assess the acceptability and usability of a novel lateral flow assay and portable reader for the point-of-care detection of N. gonorrhoeae infection (NG-LFA). This mixed-methods study was conducted as part of a diagnostic performance and usability evaluation of a prototype NG-LFA for detection of N. gonorrhoeae in symptomatic men and women at primary healthcare facilities in the Buffalo City Metro, South Africa. The Standardized System Usability Scale (SUS) was administered, and in-depth interviews were conducted among healthcare professionals (HCPs) and fieldworkers (FWs) at pre-implementation, initial use and 3- and 6-month study implementation to assess user expectations, practical experience, and future implementation considerations for the NG-LFA. Data collection and analysis was guided by the Health Technology Adoption Framework, including new health technology attributes, learnability, satisfaction, and suitability. The framework was adapted to include perceived durability. A total of 21 HCPs and FWs were trained on the NG-LFA use. SUS scores showed good to excellent acceptability ranging from 78.8-90.6 mean scores between HCPs and FWs across study time points. All transcripts were coded using Dedoose and qualitative findings were organized by learnability, satisfaction, suitability, and durability domains. Usability themes are described for each time point. Initial insecurity dissipated and specimen processing dexterity with novel POCT technology was perfected over time especially amongst FWs through practical learning and easy-to-use instructions (learnability). Participants experienced both positive and negative test results, yielding perceived accuracy and minimal testing challenges overall (satisfaction). By 3- and 6-month use, both HCPs and FWs found the NG-LFA convenient to use in primary health care facilities often faced with space constraints and outlined perceived benefits for patients (suitability and durability). Findings show that the NG-LFA device is acceptable and usable even amongst paraprofessionals. High SUS scores and qualitative findings demonstrate high learnability, ease-of-use and suitability that provide valuable information for first-step scale-up requirements at primary healthcare level. Minor prototype adjustments would enhance robustness and durability aspects.

Serological assays have been used in seroprevalence studies to inform the dynamics of COVID-19. Lateral flow immunoassay (LFIA) tests are a very practical technology to use for this objective; however, one of their challenges may be variable diagnostic performance. Given the numerous available LFIA tests, evaluation of their accuracy is critical before real-world implementation. We performed a retrospective diagnostic evaluation study to independently determine the diagnostic accuracy of 4 different antibody-detection LFIA tests: Now Check (Bionote), CareStart (Access bio), Covid-19 BSS (Biosynex) and OnSite (CTK Biotech). The sample panel was comprised of specimens collected and stored in biobanks; specifically, specimens that were RT-PCR positive for SARS-CoV-2 collected at various times throughout the COVID-19 disease course and those that were collected before the pandemic, during 2018 or earlier, from individuals with upper respiratory symptoms but were negative for tuberculosis. Clinical performance (sensitivity and specificity) was analyzed overall, and subset across individual antibody isotypes, and days from symptoms onset. A very high specificity (98% - 100%) was found for all four tests. Overall sensitivity was variable, ranging from 29% [95% CI: 21%-39%] to 64% [95% CI: 54%-73%]. When considering detection of IgM only, the highest sensitivity was 42% [95% CI: 32%-52%], compared to 57% [95% CI: 47%-66%] for IgG only. When the analysis was restricted to at least 15 days since symptom onset, across any isotype, the sensitivity reached 90% for all four brands. All four LFIA tests proved effective for identifying COVID-19 antibodies when two conditions were met: 1) at least 15 days have elapsed since symptom onset and 2) a sample is considered positive when either IgM or IgG is present. With these considerations, the use of this assays could help in seroprevalence studies or further exploration of its potential uses.

IntroductionFollicle stimulating hormone (FSH) is identified to play a role in postmenopausal disease and hypothesized to affect abdominal aortic aneurysm (AAA) onset/progression in postmenopausal women. We aimed to detect FSHR gene expression in AAA tissue and cell types involved in AAA formation.MethodsFSH stimulation of human umbilical cord endothelial cells (HUVECs), smooth muscle cells (HUCs) and PMA-differentiated macrophages to assess gene expression of FSHR and various markers. Human macrophages activated with various stimuli were assessed for FSHR gene expression. AAA dataset, AAA tissue samples and AAA-derived smooth muscle cells (SMC) obtained from elderly female donors were assessed for FSHR gene expression. AAA-SMCs were stimulated with FSH to assess its effect on gene expression. Lastly, oxidized low-density-lipoprotein (ox-LDL) uptake and abundance of cell surface protein markers were assessed by flow cytometry after FSH stimulation of human monocytes.ResultsFSH stimulation showed similar levels of gene expression in HUVECs and HUCs. Only ACTA2 was downregulated in HUCs. In PMA-differentiated macrophages, gene expression of inflammation markers was unchanged after FSH stimulation. FSHR gene expression was found to be low in the AAA datasets. Female AAA-SMCs show occasional FSHR gene expression at a very low level, yet stimulation with FSH did not affect gene expression of SMC- or inflammation markers. FSH stimulation did not impact ox-LDL uptake or alter cell surface protein expression in monocytes. While FSHR gene expression was detected in human testis tissue, it was below quantification level in all other investigated cell types, even upon activation of macrophages with various stimuli.ConclusionDespite previous reports, we did not detect FSHR gene expression in various extragonadal cell types, except in occasional female AAA-SMCs. No clear effect on cell activation was observed upon FSH stimulation in any cell type. Our data suggest that a direct effect of FSH in AAA-related extragonadal cells is unlikely to influence AAA.

Introduction Follicle stimulating hormone (FSH) is identified to play a role in postmenopausal disease and hypothesized to affect abdominal aortic aneurysm (AAA) onset/progression in postmenopausal women. We aimed to detect FSHR gene expression in AAA tissue and cell types involved in AAA formation. Methods FSH stimulation of human umbilical cord endothelial cells (HUVECs), smooth muscle cells (HUCs) and PMA-differentiated macrophages to assess gene expression of FSHR and various markers. Human macrophages activated with various stimuli were assessed for FSHR gene expression. AAA dataset, AAA tissue samples and AAA-derived smooth muscle cells (SMC) obtained from elderly female donors were assessed for FSHR gene expression. AAA-SMCs were stimulated with FSH to assess its effect on gene expression. Lastly, oxidized low-density-lipoprotein (ox-LDL) uptake and abundance of cell surface protein markers were assessed by flow cytometry after FSH stimulation of human monocytes. Results FSH stimulation showed similar levels of gene expression in HUVECs and HUCs. Only ACTA2 was downregulated in HUCs. In PMA-differentiated macrophages, gene expression of inflammation markers was unchanged after FSH stimulation. FSHR gene expression was found to be low in the AAA datasets. Female AAA-SMCs show occasional FSHR gene expression at a very low level, yet stimulation with FSH did not affect gene expression of SMC- or inflammation markers. FSH stimulation did not impact ox-LDL uptake or alter cell surface protein expression in monocytes. While FSHR gene expression was detected in human testis tissue, it was below quantification level in all other investigated cell types, even upon activation of macrophages with various stimuli. Conclusion Despite previous reports, we did not detect FSHR gene expression in various extragonadal cell types, except in occasional female AAA-SMCs. No clear effect on cell activation was observed upon FSH stimulation in any cell type. Our data suggest that a direct effect of FSH in AAA-related extragonadal cells is unlikely to influence AAA.

BackgroundIn healthcare environments, sinks are being increasingly recognized as reservoirs for multidrug-resistant Gram-negative bacteria. In our hospital, carbapenemase-producing, Verona Integron-encoded Metallo-beta-lactamase (VIM)-positive Pseudomonas aeruginosa (VIM-PA) was detected at low endemicity in patients, and environmental culturing revealed that sink drains were primary reservoirs. Therefore, an intervention was initiated in several wards to install sink drain plugs as physical barriers against splashing to prevent transmission of VIM-PA from drain reservoirs to the surrounding sink environment.AimTo assess the efficacy of the intervention on limiting spread of VIM-PA.MethodsSwabs were taken from inner sink environments (i.e. drains), and outer sink environments (i.e. wash basins, faucet aerators, and countertops) twice before and three times after the intervention. Siphon water and drain wells were also sampled before and at the moment of the intervention, respectively. All samples were screened for VIM-PA, and isolates were typed with multiple-locus variable-number tandem repeat analysis (MLVA).ResultsThere was a significant reduction in VIM-PA positivity in both inner (P-value ConclusionsDrain plugs are a simple and effective infection prevention and control measure to contain spread of VIM-PA from drain reservoirs.

There is limited understanding of the dynamic interplay between adolescent girl's and young women's (AGYW) disclosure and social support for using oral pre-exposure prophylaxis (PrEP) and adherence. Towards this, we conducted interviews with 42 AGYW enrolled in The Community PrEP Study who exhibited either high or low blood concentrations of tenofovir-diphosphate (TFV-DP) in dried blood spots. Guided by Theories of Practice, interviews and analysis focused on AGYW perspectives and experiences with PrEP disclosure, support and adherence. AGYW with high TFV-DP blood concentrations described larger social support networks and disclosure events. In contrast, those with low TFV-DP blood concentrations described disclosing to fewer people, resulting in limited social support. Participants discussed partner support, however, this support was not described as consequential to adherence, irrespective of TFV-DP levels. Those with high levels of TFV-DP in their blood described the ability to navigate social scrutiny and changes in social support, while those with low levels of TFV-DP in their blood were more likely to question their own continued use of PrEP. To facilitate AGYW's prevention-effective use of PrEP, expanded skill-building for disclosure and resiliency against changes to social support should be examined as part of PrEP services.

There is limited understanding of the dynamic interplay between adolescent girl’s and young women’s (AGYW) disclosure and social support for using oral pre-exposure prophylaxis (PrEP) and adherence. Towards this, we conducted interviews with 42 AGYW enrolled in The Community PrEP Study who exhibited either high or low blood concentrations of tenofovir-diphosphate (TFV-DP) in dried blood spots. Guided by Theories of Practice, interviews and analysis focused on AGYW perspectives and experiences with PrEP disclosure, support and adherence. AGYW with high TFV-DP blood concentrations described larger social support networks and disclosure events. In contrast, those with low TFV-DP blood concentrations described disclosing to fewer people, resulting in limited social support. Participants discussed partner support, however, this support was not described as consequential to adherence, irrespective of TFV-DP levels. Those with high levels of TFV-DP in their blood described the ability to navigate social scrutiny and changes in social support, while those with low levels of TFV-DP in their blood were more likely to question their own continued use of PrEP. To facilitate AGYW’s prevention-effective use of PrEP, expanded skill-building for disclosure and resiliency against changes to social support should be examined as part of PrEP services.

Objectives To assess the real-world diagnostic performance of nasal and nasopharyngeal swabs for SD Biosensor STANDARD Q COVID-19 Antigen Rapid Diagnostic Test (Ag-RDT). Methods Individuals ≥5 years with COVID-19 compatible symptoms or history of exposure to SARS-CoV-2 presenting at hospitals in Lesotho received two nasopharyngeal and one nasal swab. Ag-RDT from nasal and nasopharyngeal swabs were performed as point-of-care on site, the second nasopharyngeal swab used for polymerase chain reaction (PCR) as the reference standard. Results Out of 2198 participants enrolled, 2131 had a valid PCR result (61% female, median age 41 years, 8% children), 84.5% were symptomatic. Overall PCR positivity rate was 5.8%. The sensitivity for nasopharyngeal, nasal, and combined nasal and nasopharyngeal Ag-RDT result was 70.2% (95%CI: 61.3–78.0), 67.3% (57.3–76.3) and 74.4% (65.5–82.0), respectively. The respective specificity was 97.9% (97.1–98.4), 97.9% (97.2–98.5) and 97.5% (96.7–98.2). For both sampling modalities, sensitivity was higher in participants with symptom duration ≤ 3days versus ≤ 7days. Agreement between nasal and nasopharyngeal Ag-RDT was 99.4%. Conclusions The STANDARD Q Ag-RDT showed high specificity. Sensitivity was, however, below the WHO recommended minimum requirement of ≥ 80%. The high agreement between nasal and nasopharyngeal sampling suggests that for Ag-RDT nasal sampling is a good alternative to nasopharyngeal sampling.

ObjectivesIn order to generate independent performance data regarding accuracy of COVID-19 antigen-based rapid diagnostic tests (Ag-RDTs), prospective diagnostic evaluation studies across multiple sites are required to evaluate their performance in different clinical settings. This report describes the clinical evaluation the GENEDIA W COVID-19 Ag Device (Green Cross Medical Science Corp., Chungbuk, Korea) and the ActiveXpress+ COVID-19 Complete Testing Kit (Edinburgh Genetics Ltd, UK), in two testing sites Peru and the United Kingdom.MethodsNasopharyngeal swabs collected from 456 symptomatic patients at primary points of care in Lima, Peru and 610 symptomatic participants at a COVID-19 Drive-Through testing site in Liverpool, England were analyzed by Ag-RDT and compared to RT-PCR. Analytical evaluation of both Ag-RDTs was assessed using serial dilutions of direct culture supernatant of a clinical SARS-CoV-2 isolate from the B.1.1.7 lineage.ResultsFor GENEDIA brand, the values of overall sensitivity and specificity were 60.4% [95% CI 52.4-67.9%], and 99.2% [95% CI 97.6-99.7%] respectively; and for Active Xpress+ the overall values of sensitivity and specificity were 66.2% [95% CI 54.0-76.5%], and 99.6% [95% CI 97.9-99.9%] respectively. The analytical limit of detection was determined at 5.0 x 102 pfu/ml what equals to approximately 1.0 x 104 gcn/ml for both Ag-RDTs. The UK cohort had lower median Ct values compared to that of Peru during both evaluations. When split by Ct, both Ag-RDTs had optimum sensitivities at CtConclusionsWhilst the overall clinical sensitivity of the Genedia did not meet WHO minimum performance requirements for rapid immunoassays in either cohort, the ActiveXpress+ did so for the small UK cohort. This study illustrates comparative performance of Ag-RDTs across two global settings and considers the different approaches in evaluation methods.

ObjectivesTo assess the real-world diagnostic performance of nasal and nasopharyngeal swabs for SD Biosensor STANDARD Q COVID-19 Antigen Rapid Diagnostic Test (Ag-RDT).MethodsIndividuals ≥5 years with COVID-19 compatible symptoms or history of exposure to SARS-CoV-2 presenting at hospitals in Lesotho received two nasopharyngeal and one nasal swab. Ag-RDT from nasal and nasopharyngeal swabs were performed as point-of-care on site, the second nasopharyngeal swab used for polymerase chain reaction (PCR) as the reference standard.ResultsOut of 2198 participants enrolled, 2131 had a valid PCR result (61% female, median age 41 years, 8% children), 84.5% were symptomatic. Overall PCR positivity rate was 5.8%. The sensitivity for nasopharyngeal, nasal, and combined nasal and nasopharyngeal Ag-RDT result was 70.2% (95%CI: 61.3-78.0), 67.3% (57.3-76.3) and 74.4% (65.5-82.0), respectively. The respective specificity was 97.9% (97.1-98.4), 97.9% (97.2-98.5) and 97.5% (96.7-98.2). For both sampling modalities, sensitivity was higher in participants with symptom duration ≤ 3days versus ≤ 7days. Agreement between nasal and nasopharyngeal Ag-RDT was 99.4%.ConclusionsThe STANDARD Q Ag-RDT showed high specificity. Sensitivity was, however, below the WHO recommended minimum requirement of ≥ 80%. The high agreement between nasal and nasopharyngeal sampling suggests that for Ag-RDT nasal sampling is a good alternative to nasopharyngeal sampling.

Extracellular histones are cytotoxic molecules involved in experimental acute kidney injury. In patients receiving a renal transplant from donors after circulatory death, who suffer from additional warm ischemia, worse graft outcome is associated with higher machine perfusate extracellular histone H3 concentrations. We now investigated temperature-dependent extracellular histone release in an ex vivo porcine renal perfusion model, and subsequently studied histone release in the absence and presence of non-anticoagulant heparin. Seven pairs of ischemically damaged porcine kidneys were machine perfused at 4°C (cold ischemia) or 28°C (warm ischemia). Perfusate histone H3 concentration was higher after warm as compared to cold ischemia (median (IQR) = 0.48 (0.20-0.83) μg/mL vs. 0.02 (0.00-0.06) μg/mL; p = .045, respectively). Employing immune-electron microscopy (EM), histone containing cytoplasmic protrusions of tubular and endothelial cells were found after warm ischemic injury. Furthermore, abundant histone localization was detected in debris surrounding severely damaged glomerular cells, in a "buck shot" pattern. In vitro, histones were cytotoxic to endothelial and kidney epithelial cells in a temperature-dependent manner. In a separate ex vivo experiment, addition of heparin did not change the total histone H3 levels observed in the perfusate but revealed a continuous increase in the level of a lower molecular weight histone H3 variant. Our findings show that ischemically damaged kidneys release more extracellular histones in warm ischemia, which by EM was due to histone release by renal cells. Blocking of histone-mediated damage during transplantation may be beneficial in prevention of renal injury.

Dog-mediated rabies is responsible for tens of thousands of human deaths annually, and in resource-constrained settings, vaccinating dogs to control the disease at source remains challenging. Currently, rabies elimination efforts rely on mass dog vaccination by the parenteral route. To increase the herd immunity, free-roaming and stray dogs need to be specifically addressed in the vaccination campaigns, with oral rabies vaccination (ORV) of dogs being a possible solution. Using a third-generation vaccine and a standardized egg-flavoured bait, bait uptake and vaccination was assessed under field conditions in Namibia. During this trial, both veterinary staff as well as dog owners expressed their appreciation to this approach of vaccination. Of 1,115 dogs offered a bait, 90% (n = 1,006, 95%CI:91-94) consumed the bait and 72.9% (n = 813, 95%CI:70.2-75.4) of dogs were assessed as being vaccinated by direct observation, while for 11.7% (n = 130, 95%CI:9.9-17.7) the status was recorded as "unkown" and 15.4% (n = 172, 95%CI: 13.4-17.7) were considered as being not vaccinated. Smaller dogs and dogs offered a bait with multiple other dogs had significantly higher vaccination rates, while other factors, e.g. sex, confinement status and time had no influence. The favorable results of this first large-scale field trial further support the strategic integration of ORV into dog rabies control programmes. Given the acceptance of the egg-flavored bait under various settings worldwide, ORV of dogs could become a game-changer in countries, where control strategies using parenteral vaccination alone failed to reach sufficient vaccination coverage in the dog population.

The role that balancing selection plays in the maintenance of genetic diversity remains unresolved. Here, we introduce a new test, based on the McDonald–Kreitman test, in which the number of polymorphisms that are shared between populations is contrasted to those that are private at selected and neutral sites. We show that this simple test is robust to a variety of demographic changes, and that it can also give a direct estimate of the number of shared polymorphisms that are directly maintained by balancing selection. We apply our method to population genomic data from humans and provide some evidence that hundreds of nonsynonymous polymorphisms are subject to balancing selection. What maintains genetic variation remains an unresolved mystery. This study describes the development of a new test and its application to human population genomic data, suggesting that natural selection may have a much more important role than previously thought, with hundreds of non-synonymous polymorphisms subject to balancing selection.

Brain imaging research enjoys increasing adoption of supervised machine learning for single-participant disease classification. Yet, the success of these algorithms likely depends on population diversity, including demographic differences and other factors that may be outside of primary scientific interest. Here, we capitalize on propensity scores as a composite confound index to quantify diversity due to major sources of population variation. We delineate the impact of population heterogeneity on the predictive accuracy and pattern stability in 2 separate clinical cohorts: the Autism Brain Imaging Data Exchange (ABIDE, n = 297) and the Healthy Brain Network (HBN, n = 551). Across various analysis scenarios, our results uncover the extent to which cross-validated prediction performances are interlocked with diversity. The instability of extracted brain patterns attributable to diversity is located preferentially in regions part of the default mode network. Collectively, our findings highlight the limitations of prevailing deconfounding practices in mitigating the full consequences of population diversity.

Background The treatment coverage for major depressive disorder (MDD) is low in many parts of the world despite MDD being a major contributor to disability globally. Most existing reviews of MDD treatment coverage do not account for potential sources of study-level heterogeneity that contribute to variation in reported treatment rates. This study aims to provide a comprehensive review of the evidence and analytically quantify sources of heterogeneity to report updated estimates of MDD treatment coverage and gaps by location and treatment type between 2000 and 2019. Methods and findings A systematic review of the literature was conducted to identify relevant studies that provided data on treatment rates for MDD between January 1, 2000, and November 26, 2021, from 2 online scholarly databases PubMed and Embase. Cohort and cross-sectional studies were included if treatment rates pertaining to the last 12 months or less were reported directly or if sufficient information was available to calculate this along with 95% uncertainty intervals (UIs). Studies were included if they made use of population-based surveys that were representative of communities, countries, or regions under study. Studies were included if they used established diagnostic criteria to diagnose cases of MDD. Sample and methodological characteristics were extracted from selected studies. Treatment rates were modeled using a Bayesian meta-regression approach and adjusted for select covariates that quantified heterogeneity in the data. These covariates included age, sex, treatment type, location, and choice of MDD assessment tool. A total of 149 studies were included for quantitative analysis. Treatment coverage for health service use ranged from 51% [95% UI 20%, 82%] in high-income locations to 20% [95% UI 1%, 53%] in low- and lower middle-income locations. Treatment coverage for mental health service use ranged from 33% [95% UI 8%, 66%] in high-income locations to 8% [95% UI Conclusions In this study, we observed that the treatment coverage for MDD continues to be low in many parts of the world and in particular in low- and lower middle-income countries. There is a continued need for routine data collection that will help obtain more accurate estimates of treatment coverage globally. In a systematic review and Bayesian meta-regression analysis, Modhurima Moitra and colleagues estimate major depressive disorder treatment coverage in 84 countries. Author summary Why was this study done? ➢ Major depressive disorder (MDD) is one of the major contributors to disability worldwide, but treatment rates for this condition are remarkably low. ➢ To the best of our knowledge, previous systematic reviews on this topic provide more descriptive summaries of treatment rates without accounting for differences in study attributes that may contribute to variation in reported treatment rates. ➢ An updated systematic review that is more reflective of the recent literature on treatment rates as well as an improved analytical approach may provide more accurate estimates of treatment rates by resource setting and geography. What did the researchers do and find? ➢ We conducted a systematic review and meta-regression analysis using data on treatment rates for MDD from 149 studies and 84 countries between 2000 and 2021. ➢ We estimated pooled treatment rates adjusted for parameters of interest including age, sex, treatment type, study methods, and location. ➢ Mental health service use ranged from 33% (95% uncertainty interval (UI) 8, 66) in high-income countries to 8% (95% UI What do these findings mean? ➢ This systematic review provides updated evidence on treatment rates for MDD, and results suggest that there are wide disparities in treatment rates by resource setting. ➢ More high-quality data on depression treatment coverage and adequacy are needed from low- and lower middle-income countries. ➢ These findings may help prioritize efforts to scale up depression treatment in locations with clearly identified treatment gaps.

BackgroundThe treatment coverage for major depressive disorder (MDD) is low in many parts of the world despite MDD being a major contributor to disability globally. Most existing reviews of MDD treatment coverage do not account for potential sources of study-level heterogeneity that contribute to variation in reported treatment rates. This study aims to provide a comprehensive review of the evidence and analytically quantify sources of heterogeneity to report updated estimates of MDD treatment coverage and gaps by location and treatment type between 2000 and 2019.Methods and findingsA systematic review of the literature was conducted to identify relevant studies that provided data on treatment rates for MDD between January 1, 2000, and November 26, 2021, from 2 online scholarly databases PubMed and Embase. Cohort and cross-sectional studies were included if treatment rates pertaining to the last 12 months or less were reported directly or if sufficient information was available to calculate this along with 95% uncertainty intervals (UIs). Studies were included if they made use of population-based surveys that were representative of communities, countries, or regions under study. Studies were included if they used established diagnostic criteria to diagnose cases of MDD. Sample and methodological characteristics were extracted from selected studies. Treatment rates were modeled using a Bayesian meta-regression approach and adjusted for select covariates that quantified heterogeneity in the data. These covariates included age, sex, treatment type, location, and choice of MDD assessment tool. A total of 149 studies were included for quantitative analysis. Treatment coverage for health service use ranged from 51% [95% UI 20%, 82%] in high-income locations to 20% [95% UI 1%, 53%] in low- and lower middle-income locations. Treatment coverage for mental health service use ranged from 33% [95% UI 8%, 66%] in high-income locations to 8% [95% UI ConclusionsIn this study, we observed that the treatment coverage for MDD continues to be low in many parts of the world and in particular in low- and lower middle-income countries. There is a continued need for routine data collection that will help obtain more accurate estimates of treatment coverage globally.

Grounded in an ecodevelopment perspective, in the current study we examined unique and moderating effects of daily COVID-19 prevalence (social contexts) on effects of COVID-19 related risk and protective factors such as emotional distress (individual contexts) and employment (working from home and unemployment status; family contexts) on family functioning among 160 recent immigrant families in Israel. In general, results indicate several unique effects of COVID-19 related factors (such as COVID-19 emotional distress, unemployment, and remote work arrangements) on both parents' and adolescents' reports of family functioning. However, results indicated that there were more significant associations between COVID-19 factors (e.g., emotional distress and COVID-19 prevalence) and family functioning indicators with adolescents, than with parents. The effects of COVID-19 factors (e.g., emotional distress and remote work arrangements) were moderated by daily COVID-19 prevalence (new cases and deaths). We discuss ways in which interventionists can contribute to pandemic-related research to promote optimal family functioning among immigrant families.

Genome-wide association studies (GWAS) have identified many single nucleotide polymorphisms (SNPs) that play important roles in the genetic heritability of traits and diseases. With most of these SNPs located on the non-coding part of the genome, it is currently assumed that these SNPs influence the expression of nearby genes on the genome. However, identifying which genes are targeted by these disease-associated SNPs remains challenging. In the past, protein knowledge graphs have often been used to identify genes that are associated with disease, also referred to as "disease genes". Here, we explore whether protein knowledge graphs can be used to identify genes that are targeted by disease-associated non-coding SNPs by testing and comparing the performance of six existing methods for a protein knowledge graph, four of which were developed for disease gene identification. We compare our performance against two baselines: (1) an existing state-of-the-art method that is based on guilt-by-association, and (2) the leading assumption that SNPs target the nearest gene on the genome. We test these methods with four reference sets, three of which were obtained by different means. Furthermore, we combine methods to investigate whether their combination improves performance. We find that protein knowledge graphs that include predicate information perform comparable to the current state of the art, achieving an area under the receiver operating characteristic curve (AUC) of 79.6% on average across all four reference sets. Protein knowledge graphs that lack predicate information perform comparable to our other baseline (genetic distance) which achieved an AUC of 75.7% across all four reference sets. Combining multiple methods improved performance to 84.9% AUC. We conclude that methods for a protein knowledge graph can be used to identify which genes are targeted by disease-associated non-coding SNPs.

ObjectiveAfter stratifying for age, sex and multimorbidity at baseline, our aim is to analyse time trends in incident multimorbidity and polypharmacy in the 15-year clinical trajectories of individual patients in a family medicine setting.MethodsThis study was carried out using data from the Registration Network Family Medicine in the South of the Netherlands. The clinical trajectories of 10037 subjects during the 15-year period (2000-2014) were analyzed in a repeated measurement of using a generalized estimating equations model as well as a multilevel random intercept model with repeated measurements to determine patterns of incident multimorbidity and polypharmacy. Hierarchical age-period-cohort models were used to generate age and cohort trajectories for comparison with prevalence trends in multimorbidity literature.ResultsMultimorbidity was more common in females than in males throughout the duration of the 15-year trajectory (females: 39.6%; males: 33.5%). With respective ratios of 11.7 and 5.9 between the end and the beginning of the 15-year period, the youngest female and male groups showed a substantial increase in multimorbidity prevalence. Ratios in the oldest female and male groups were 2.2 and 1.9 respectively. Females had higher levels of multimorbidity than males in the 0-24-year and 25-44-year age groups, but the levels converged to a prevalence of 92.2% in the oldest male and 90.7% in the oldest female group. Similar, albeit, moderate differences were found in polypharmacy patterns.ConclusionsWe sought to specify the progression of multimorbidity from an early age. As a result, our study adds to the multimorbidity literature by specifying changes in chronic disease accumulation with relation to polypharmacy, and by tracking differences in patient trajectories according to age and sex. Multimorbidity and polypharmacy are common and their prevalence is accelerating, with a relatively rapid increase in younger groups. From the point of view of family medicine, this underlines the need for a longitudinal approach and a life course perspective in patient care.

Misleading graphs are a source of misinformation that worry many experts. Especially people with a low graph literacy are thought to be persuaded by graphs that misrepresent the underlying data. But we know little about how people interpret misleading graphs and how these graphs influence their opinions. In this study we focus on the effect of truncating the y-axis for a line chart which exaggerates an upgoing trend. In a randomized controlled trial, we showed participants either a normal or a misleading chart, and we did so in two different contexts. After they had seen the graphs, we asked participants their opinion on the trend and to give an estimation of the increase. Finally we measured their graph literacy. Our results show that context is the only significant factor in opinion-forming; the misleading graph and graph literacy had no effect. None of these factors had a significant impact on estimations for the increase. These results show that people might be less susceptible to misleading graphs than we thought and that context has more impact than a misleading y-axis.

Genome-wide association studies (GWAS) have identified many single nucleotide polymorphisms (SNPs) that play important roles in the genetic heritability of traits and diseases. With most of these SNPs located on the non-coding part of the genome, it is currently assumed that these SNPs influence the expression of nearby genes on the genome. However, identifying which genes are targeted by these disease-associated SNPs remains challenging. In the past, protein knowledge graphs have often been used to identify genes that are associated with disease, also referred to as “disease genes”. Here, we explore whether protein knowledge graphs can be used to identify genes that are targeted by disease-associated non-coding SNPs by testing and comparing the performance of six existing methods for a protein knowledge graph, four of which were developed for disease gene identification. We compare our performance against two baselines: (1) an existing state-of-the-art method that is based on guilt-by-association, and (2) the leading assumption that SNPs target the nearest gene on the genome. We test these methods with four reference sets, three of which were obtained by different means. Furthermore, we combine methods to investigate whether their combination improves performance. We find that protein knowledge graphs that include predicate information perform comparable to the current state of the art, achieving an area under the receiver operating characteristic curve (AUC) of 79.6% on average across all four reference sets. Protein knowledge graphs that lack predicate information perform comparable to our other baseline (genetic distance) which achieved an AUC of 75.7% across all four reference sets. Combining multiple methods improved performance to 84.9% AUC. We conclude that methods for a protein knowledge graph can be used to identify which genes are targeted by disease-associated non-coding SNPs.

Background and aimsSubjects with congenital chloride diarrhea (CLD; a defect in solute carrier family 26 member 3 (SLC26A3)) are prone to inflammatory bowel disease (IBD). We investigated fecal microbiota in CLD and CLD-associated IBD. We also tested whether microbiota is modulated by supplementation with the short-chain fatty acid butyrate.Subjects and methodsWe recruited 30 patients with CLD for an observational 3-week follow-up study. Thereafter, 16 consented to oral butyrate substitution for a 3-week observational period. Fecal samples, collected once a week, were assayed for calprotectin and potential markers of inflammation, and studied by 16S ribosomal ribonucleic acid (rRNA) gene amplicon sequencing and compared to that of 19 healthy controls and 43 controls with Crohn's disease. Data on intestinal symptoms, diet and quality of life were collected.ResultsPatients with CLD had increased abundances of Proteobacteria, Veillonella, and Prevotella, and lower abundances of normally dominant taxa Ruminococcaceae and Lachnospiraceae when compared with healthy controls and Crohn´s disease. No major differences in fecal microbiota were found between CLD and CLD-associated IBD (including two with yet untreated IBD). Butyrate was poorly tolerated and showed no major effects on fecal microbiota or biomarkers in CLD.ConclusionsFecal microbiota in CLD is different from that of healthy subjects or Crohn´s disease. Unexpectedly, no changes in the microbiota or fecal markers characterized CLD-associated IBD, an entity with high frequency among patients with CLD.

This work addresses the need for new chemical matter in product development for control of pest insects and vector-borne diseases. We present a barcoding strategy that enables phenotypic screens of blood-feeding insects against small molecules in microtiter plate-based arrays and apply this to discovery of novel systemic insecticides and compounds that block malaria parasite development in the mosquito vector. Encoding of the blood meals was achieved through recombinant DNA-tagged Asaia bacteria that successfully colonised Aedes and Anopheles mosquitoes. An arrayed screen of a collection of pesticides showed that chemical classes of avermectins, phenylpyrazoles, and neonicotinoids were enriched for compounds with systemic adulticide activity against Anopheles. Using a luminescent Plasmodium falciparum reporter strain, barcoded screens identified 48 drug-like transmission-blocking compounds from a 400-compound antimicrobial library. The approach significantly increases the throughput in phenotypic screening campaigns using adult insects and identifies novel candidate small molecules for disease control.

Cytomegalovirus (CMV) infection has a major impact on the T-cell pool, which is thought to be associated with ageing of the immune system. The effect on the T-cell pool has been interpreted as an effect of CMV on non-CMV specific T-cells. However, it remains unclear whether the effect of CMV could simply be explained by the presence of large, immunodominant, CMV-specific memory CD8+ T-cell populations. These have been suggested to establish through gradual accumulation of long-lived cells. However, little is known about their maintenance. We investigated the effect of CMV infection on T-cell dynamics in healthy older adults, and aimed to unravel the mechanisms of maintenance of large numbers of CMV-specific CD8+ T-cells. We studied the expression of senescence, proliferation, and apoptosis markers and quantified the in vivo dynamics of CMV-specific and other memory T-cell populations using in vivo deuterium labelling. Increased expression of late-stage differentiation markers by CD8+ T-cells of CMV+ versus CMV- individuals was not solely explained by the presence of large, immunodominant CMV-specific CD8+ T-cell populations. The lifespans of circulating CMV-specific CD8+ T-cells did not differ significantly from those of bulk memory CD8+ T-cells, and the lifespans of bulk memory CD8+ T-cells did not differ significantly between CMV- and CMV+ individuals. Memory CD4+ T-cells of CMV+ individuals showed increased expression of late-stage differentiation markers and decreased Ki-67 expression. Overall, the expression of senescence markers on T-cell populations correlated positively with their expected in vivo lifespan. Together, this work suggests that i) large, immunodominant CMV-specific CD8+ T-cell populations do not explain the phenotypical differences between CMV+ and CMV- individuals, ii) CMV infection hardly affects the dynamics of the T-cell pool, and iii) large numbers of CMV-specific CD8+ T-cells are not due to longer lifespans of these cells.

BackgroundThe existence of locations with low but stable onchocerciasis prevalence is not well understood. An often suggested yet poorly investigated explanation is that the infection spills over from neighbouring locations with higher infection densities.MethodologyWe adapted the stochastic individual based model ONCHOSIM to enable the simulation of multiple villages, with separate blackfly (intermediate host) and human populations, which are connected through the regular movement of the villagers and/or the flies. With this model we explore the impact of the type, direction and degree of connectedness, and of the impact of localized or full-area mass drug administration (MDA) over a range of connected village settings.Principal findingsIn settings with annual fly biting rates (ABR) below the threshold needed for stable local transmission, persistence of onchocerciasis prevalence can well be explained by regular human traffic and/or fly movement from locations with higher ABR. Elimination of onchocerciasis will then theoretically be reached by only implementing MDA in the higher prevalence area, although lingering infection in the low prevalence location can trigger resurgence of transmission in the total region when MDA is stopped too soon. Expanding MDA implementation to the lower ABR location can therefore shorten the duration of MDA needed. For example, when prevalence spill-over is due to human traffic, and both locations have about equal populations, then the MDA duration can be shortened by up to three years. If the lower ABR location has twice as many inhabitants, the reduction can even be up to six years, but if spill-over is due to fly movement, the expected reduction is less than a year.Conclusions/significanceAlthough MDA implementation might not always be necessary in locations with stable low onchocerciasis prevalence, in many circumstances it is recommended to accelerate achieving elimination in the wider area.

Plasmodium species, the causative agent of malaria, have a complex life cycle involving two hosts. The sporozoite life stage is characterized by an extended phase in the mosquito salivary glands followed by free movement and rapid invasion of hepatocytes in the human host. This transmission stage has been the subject of many transcriptomics and proteomics studies and is also targeted by the most advanced malaria vaccine. We applied Bayesian data integration to determine which proteins are not only present in sporozoites but are also specific to that stage. Transcriptomic and proteomic Plasmodium data sets from 26 studies were weighted for how representative they are for sporozoites, based on a carefully assembled gold standard for Plasmodium falciparum (Pf) proteins known to be present or absent during the sporozoite life stage. Of 5418 Pf genes for which expression data were available at the RNA level or at the protein level, 975 were identified as enriched in sporozoites and 90 specific to them. We show that Pf sporozoites are enriched for proteins involved in type II fatty acid synthesis in the apicoplast and GPI anchor synthesis, but otherwise appear metabolically relatively inactive in the salivary glands of mosquitos. Newly annotated hypothetical sporozoite-specific and sporozoite-enriched proteins highlight sporozoite-specific functions. They include PF3D7_0104100 that we identified to be homologous to the prominin family, which in human has been related to a quiescent state of cancer cells. We document high levels of genetic variability for sporozoite proteins, specifically for sporozoite-specific proteins that elicit antibodies in the human host. Nevertheless, we can identify nine relatively well-conserved sporozoite proteins that elicit antibodies and that together can serve as markers for previous exposure. Our understanding of sporozoite biology benefits from identifying key pathways that are enriched during this life stage. This work can guide studies of molecular mechanisms underlying sporozoite biology and potential well-conserved targets for marker and drug development.

DNA methyltransferase (Dnmt)3b mediates de novo DNA methylation and modulation of Dnmt3b in respiratory epithelial cells has been shown to affect the expression of multiple genes. Respiratory epithelial cells provide a first line of defense against pulmonary pathogens and play a crucial role in the immune response during pneumonia caused by Pseudomonas (P.) aeruginosa, a gram-negative bacterium that expresses flagellin as an important virulence factor. We here sought to determine the role of Dntm3b in respiratory epithelial cells in immune responses elicited by P. aeruginosa. DNMT3B expression was reduced in human bronchial epithelial (BEAS-2B) cells as well as in primary human and mouse bronchial epithelial cells grown in air liquid interface upon exposure to P. aeruginosa (PAK). Dnmt3b deficient human bronchial epithelial (BEAS-2B) cells produced more CXCL1, CXCL8 and CCL20 than control cells when stimulated with PAK, flagellin-deficient PAK (PAKflic) or flagellin. Dnmt3b deficiency reduced DNA methylation at exon 1 of CXCL1 and enhanced NF-ĸB p65 binding to the CXCL1 promoter. Mice with bronchial epithelial Dntm3b deficiency showed increased Cxcl1 mRNA expression in bronchial epithelium and CXCL1 protein release in the airways during pneumonia caused by PAK, which was associated with enhanced neutrophil recruitment and accelerated bacterial clearance; bronchial epithelial Dnmt3b deficiency did not modify responses during pneumonia caused by PAKflic or Klebsiella pneumoniae (an un-flagellated gram-negative bacterium). Dnmt3b deficiency in type II alveolar epithelial cells did not affect mouse pulmonary defense against PAK infection. These results suggest that bronchial epithelial Dnmt3b impairs host defense during Pseudomonas induced pneumonia, at least in part, by dampening mucosal responses to flagellin.

Burkholderia multivorans is a member of the Burkholderia cepacia complex (Bcc), notorious for its pathogenicity in persons with cystic fibrosis. Epidemiological surveillance suggests that patients predominantly acquire B. multivorans from environmental sources, with rare cases of patient-to-patient transmission. Here we report on the genomic analysis of thirteen isolates from an endemic B. multivorans strain infecting four cystic fibrosis patients treated in different pediatric cystic fibrosis centers in Belgium, with no evidence of cross-infection. All isolates share an identical sequence type (ST-742) but whole genome analysis shows that they exhibit peculiar patterns of genomic diversity between patients. By combining short and long reads sequencing technologies, we highlight key differences in terms of small nucleotide polymorphisms indicative of low rates of adaptive evolution within patient, and well-defined, hundred kbps-long segments of high enrichment in mutations between patients. In addition, we observed large structural genomic variations amongst the isolates which revealed different plasmid contents, active roles for transposase IS3 and IS5 in the deactivation of genes, and mobile prophage elements. Our study shows limited within-patient B. multivorans evolution and high between-patient strain diversity, indicating that an environmental microdiverse reservoir must be present for this endemic strain, in which active diversification is taking place. Furthermore, our analysis also reveals a set of 30 parallel adaptations across multiple patients, indicating that the specific genomic background of a given strain may dictate the route of adaptation within the cystic fibrosis lung.

ObjectivesDiagnostics are essential for controlling the pandemic. Identifying a reliable and fast diagnostic device is needed for effective testing. We assessed performance and ease-of-use of the Abbott PanBio antigen-detecting rapid diagnostic test (Ag-RDT).MethodsThis prospective, multi-centre diagnostic accuracy study enrolled at two sites in Germany. Following routine testing with reverse-transcriptase polymerase chain reaction (RT-PCR), a second study-exclusive swab was performed for Ag-RDT testing. Routine swabs were nasopharyngeal (NP) or combined NP/oropharyngeal (OP) whereas the study-exclusive swabs were NP. To evaluate performance, sensitivity and specificity were assessed overall and in predefined sub-analyses accordingly to cycle-threshold values, days after symptom onset, disease severity and study site. Additionally, an ease-of-use assessment (EoU) and System Usability Scale (SUS) were performed.Results1108 participants were enrolled between Sept 28 and Oct 30, 2020. Of these, 106 (9.6%) were PCR-positive. The Abbott PanBio detected 92/106 PCR-positive participants with a sensitivity of 86.8% (95% CI: 79.0% - 92.0%) and a specificity of 99.9% (95% CI: 99.4%-100%). The sub-analyses indicated that sensitivity was 95.8% in Ct-values ConclusionThe Abbott PanBio Ag-RDT performs well for SARS-CoV-2 testing in this large manufacturer independent study, confirming its WHO recommendation for Emergency Use in settings with limited resources.

IntroductionTargeted therapy against tumor angiogenesis is widely used in clinical practice for patients with colorectal liver metastases (CRLM). Possible predictive biomarkers for tumor angiogenesis, such as, microvessel density (MVD), hypoxia and cell proliferation, can be determined using immunohistochemical staining. However, patients ineligible for surgical treatment need to undergo invasive diagnostic interventions in order to determine these biomarkers. CT perfusion (CTP) is an emerging functional imaging technique, which can non-invasively determine vascular properties of solid tumors. The purpose of this study was to evaluate CTP with histological biomarkers in CRLM.Material and methodsPatients with CRLM underwent CTP one day before liver surgery. CTP analysis was performed on the entire volume of the largest metastases in each patient. Dual-input maximum slope analysis was used and data concerning arterial flow (AF), portal flow (PF) and perfusion index (PI) were recorded. Immunohistochemical staining with CD34, M75/CA-IX and MIB-1 was performed on the rim in the midsection of the tumor to determine respectively MVD, hypoxia and cell proliferation.ResultsTwenty CRLM in 20 patients were studied. Mean size of the largest CRLM was 37 mm (95% CI 21-54 mm). Mean AF and PF were respectively 64 ml/min/100ml (95% CI 48-79) and 30 ml/min/100ml (95% CI 22-38). Mean PI was 68% (95% CI 62-73). No significant correlation was found between tumor growth patterns and CTP (p = 0.95). MVD did not significantly correlate to AF (r = 0.05; p = 0.84), PF (r = 0.17; p = 0.47) and PI (r = -0.12; p = 0.63). Cell proliferation also did not significantly correlate to AF (r = 0.07; p = 0.78), PF (r = -0.01; p = 0.95) and PI (r = 0.15; p = 0.52). Hypoxia did not significantly correlate to AF (r = -0.05; p = 0.83), however, significantly to PF (r = 0.51; p = 0.02) and a trend to negative correlation with PF (r = -0.43; p = 0.06). However, after controlling the false discovery rate, no significant correlation between CTP and used immunohistochemical biomarkers was found.ConclusionIn conclusion, this feasibility study found a trend to negative correlation between PI and hypoxia, CTP might therefore possibly evaluate this prognostic marker in CRLM non-invasively. However, CTP is not an appropriate technique for the assessment of microvessels or cell proliferation in CRLM.

BackgroundAntigen-detecting rapid diagnostic tests (Ag-RDTs) for the detection of SARS-CoV-2 offer new opportunities for testing in the context of the COVID-19 pandemic. Nasopharyngeal swabs (NPS) are the reference sample type, but oropharyngeal swabs (OPS) may be a more acceptable sample type in some patients.MethodsWe conducted a prospective study in a single screening center to assess the diagnostic performance of the Panbio™ COVID-19 Ag Rapid Test (Abbott) on OPS compared with reverse-transcription quantitative PCR (RT-qPCR) using NPS during the second pandemic wave in Switzerland.Results402 outpatients were enrolled in a COVID-19 screening center, of whom 168 (41.8%) had a positive RT-qPCR test. The oropharyngeal Ag-RDT clinical sensitivity compared to nasopharyngeal RT-qPCR was 81% (95%CI: 74.2-86.6). Two false positives were noted out of the 234 RT-qPCR negative individuals, which resulted in a clinical specificity of 99.1% (95%CI: 96.9-99.9) for the Ag-RDT. For cycle threshold values ≤ 26.7 (≥ 1E6 SARS-CoV-2 genomes copies/mL, a presumed cut-off for infectious virus), 96.3% sensitivity (95%CI: 90.7-99.0%) was obtained with the Ag-RDT using OPS.InterpretationBased on our findings, the diagnostic performance of the Panbio™ Covid-19 RDT with OPS samples, if taken by a trained person and high requirements regarding quality of the specimen, meet the criteria required by the WHO for Ag-RDTs (sensitivity ≥80% and specificity ≥97%) in a high incidence setting in symptomatic individuals.

BackgroundStudies from high income countries report that HIV-positive people have an impaired systolic and diastolic cardiac function compared to HIV-negative people. It is unclear if results can be translated directly to the Sub-Saharan Africa context. This study assesses electro- and echocardiographic characteristics in an urban African population, comparing HIV-positive people (treated and not yet treated) with HIV-negative controls.MethodsWe conducted a cross-sectional study in Johannesburg, South Africa. We enrolled HIV-positive participants from three randomized controlled trials that had recruited participants from routine HIV testing programs. HIV-negative controls were recruited from the community. Data were collected on demographics, cardiovascular risk factors, medical history and electrocardiographic and echocardiographic characteristics.ResultsIn total, 394 HIV-positive participants and 153 controls were enrolled. The mean age of HIV-positive participants was 40±9 years (controls: 35±10 years), and 34% were male (controls: 50%). Of HIV-positive participants 36% were overweight or obese (controls: 44%), 23% had hypertension (controls: 28%) and 12% were current smoker (controls: 37%). Median time since HIV diagnosis was 6.0 years (IQR 2.3-10.0) and median treatment duration was 4.0 years (IQR 0.0-8.0), 50% had undetectable viral load. The frequency of anatomical cardiac abnormalities was low and did not differ between people with and without HIV. We observed no relation between HIV or anti-retroviral therapy (ART) and systolic or diastolic heart function. There was an association between ART use and corrected QT interval: +11.8 ms compared to HIV-negative controls (pConclusionThe low number of major cardiac abnormalities in this relatively young, well managed urban African HIV-positive population is reassuring. The increase in corrected QT interval and left ventricular mass may contribute to higher cardiac mortality and morbidity in people living with HIV in the long term.

There is a maldistribution of human resources for health globally, with many Lower- and Middle-Income Countries experiencing significant shortages. We examined healthcare workers' job preferences in South Africa to identify factors which potentially influence employment decisions. A discrete choice experiment was conducted among 855 South African healthcare workers critical to its national HIV testing and treatment programs. Job characteristics included workload, workplace culture, availability of equipment, training opportunities, sector and facility type, location, salary and benefits. Main effects analysis was conducted using fixed effects logistic regression. Interaction effects identified divergence in preferences. Heavy workload (OR = 0.78; 95% C.I. 0.74-0.83), poor workplace culture (odds ratio 0.66; 95% C.I. 0.62-0.69), insufficient availability of equipment (OR = 0.67; 95% C.I. 0.63-0.70) and infrequent training opportunities (OR = 0.75; 95% C.I. 0.71-0.80) had large, significant effects on worker preferences. An increase in salary of 20% (OR = 1.29; 95% C.I. 1.16-1.44) had a positive effect on preferences, while a salary decrease of 20% (OR = 0.55; 95% C.I. 0.49-0.60) had a strong negative effect. Benefits packages had large positive effects on preferences: respondents were twice as likely to choose a job that included medical aid, pension and housing contributions worth 40% of salary (OR = 2.06; 95% C.I. 1.87-2.26), holding all else constant. Although salary was important across all cadres, benefits packages had larger effects on job preferences than equivalent salary increases. Improving working conditions is critical to attracting and retaining appropriate health cadres responsible for the country's HIV services, especially in the public sector and underserved, often rural, communities. Crucially, our evidence suggests that factors amenable to improvement such as workplace conditions and remuneration packages have a greater influence on healthcare workers employment decisions than employment sector or location.

ObjectiveWe investigated the potential of [18F]fluorodeoxyglucose ([18F]FDG) and [18F]Fluoromethylcholine ([18F]FCho) PET, compared to contrast-enhanced MRI, for the early detection of treatment response in F98 glioblastoma (GB) rats.MethodsWhen GB was confirmed on T2- and contrast-enhanced T1-weighted MRI, animals were randomized into a treatment group (n = 5) receiving MRI-guided 3D conformal arc micro-irradiation (20 Gy) with concomitant temozolomide, and a sham group (n = 5). Effect of treatment was evaluated by MRI and [18F]FDG PET on day 2, 5, 9 and 12 post-treatment and [18F]FCho PET on day 1, 6, 8 and 13 post-treatment. The metabolic tumor volume (MTV) was calculated using a semi-automatic thresholding method and the average tracer uptake within the MTV was converted to a standard uptake value (SUV).ResultsTo detect treatment response, we found that for [18F]FDG PET (SUVmean x MTV) is superior to MTV only. Using (SUVmean x MTV), [18F]FDG PET detects treatment effect starting as soon as day 5 post-therapy, comparable to contrast-enhanced MRI. Importantly, [18F]FDG PET at delayed time intervals (240 min p.i.) was able to detect the treatment effect earlier, starting at day 2 post-irradiation. No significant differences were found at any time point for both the MTV and (SUVmean x MTV) of [18F]FCho PET.ConclusionsBoth MRI and particularly delayed [18F]FDG PET were able to detect early treatment responses in GB rats, whereas, in this study this was not possible using [18F]FCho PET. Further comparative studies should corroborate these results and should also include (different) amino acid PET tracers.

The interpretation of archaeological features often requires a combined methodological approach in order to make the most of the material record, particularly from sites where this may be limited. In practice, this requires the consultation of different sources of information in order to cross validate findings and combat issues of ambiguity and equifinality. However, the application of a multiproxy approach often generates incompatible data, and might therefore still provide ambiguous results. This paper explores the potential of a simple digital framework to increase the explanatory power of multiproxy data by enabling the incorporation of incompatible, ambiguous datasets in a single model. In order to achieve this, Bayesian confirmation was used in combination with decision trees. The results of phytolith and geochemical analyses carried out on soil samples from ephemeral sites in Jordan are used here as a case study. The combination of the two datasets as part of a single model enabled us to refine the initial interpretation of the use of space at the archaeological sites by providing an alternative identification for certain activity areas. The potential applications of this model are much broader, as it can also help researchers in other domains reach an integrated interpretation of analysis results by combining different datasets.

Background Bacterial vaginosis (BV) is the most common gynecological condition in women of reproductive age and associated with adverse pregnancy outcomes. In the Democratic Republic of the Congo (DRC), neonatal mortality rate is as high as 2.8 percent with preterm birth (PTB) and low birth weight (LBW) as leading causes. Because no studies have addressed BV in DRC, we aimed to investigate the prevalence of BV, the risk factors and the association between BV and adverse pregnancy outcomes in a population of pregnant women from Bukavu, DRC. Methods A total of 533 pregnant women in the second trimester of pregnancy were recruited in the Provincial Reference Hospital of Bukavu, DRC, between January and October 2017, and followed until delivery. Clinical and sociodemographic data of mother and newborn, and data on (vaginal) hygiene practices, sexual behavior and reproductive history were collected. BV was diagnosed by Nugent scoring of Gram-stained vaginal smears. Two multivariate regression models were built to identify risk factors for BV and to investigate BV as a risk factor for adverse pregnancy outcomes. Results The prevalence of BV was 26.3% and approximately half of the women with BV were asymptomatic. Independent risk factors for BV were the use of alternatives to water for intravaginal washing, concurrent partners, unemployed status, the presence of vaginal Candida and clay consumption. BV was independently associated with both LBW and PTB of an infant with LBW. Conclusion The prevalence of BV in Bukavu is high but in line with the global average. BV was associated with adverse pregnancy outcomes in our study population. Hence, research on modifiable risk factor-based interventions to reduce the prevalence of BV, and on screening/treatment of BV during antenatal care should be explored to reduce neonatal mortality and morbidity.

To evaluate and compare the risk of emerging vector-borne diseases (VBDs), a Model for INTegrated RISK assessment, MINTRISK, was developed to assess the introduction risk of VBDs for new regions in an objective, transparent and repeatable manner. MINTRISK is a web-based calculation tool, that provides semi-quantitative risk scores that can be used for prioritization purposes. Input into MINTRISK is entered by answering questions regarding entry, transmission, establishment, spread, persistence and impact of a selected VBD. Answers can be chosen from qualitative answer categories with accompanying quantitative explanation to ensure consistent answering. The quantitative information is subsequently used as input for the model calculations to estimate the risk for each individual step in the model and for the summarizing output values (rate of introduction; epidemic size; overall risk). The risk assessor can indicate his uncertainty on each answer, and this is accounted for by Monte Carlo simulation. MINTRISK was used to assess the risk of four VBDs (African horse sickness, epizootic haemorrhagic disease, Rift Valley fever, and West Nile fever) for the Netherlands with the aim to prioritise these diseases for preparedness. Results indicated that the overall risk estimate was very high for all evaluated diseases but epizootic haemorrhagic disease. Uncertainty intervals were, however, wide limiting the options for ranking of the diseases. Risk profiles of the VBDs differed. Whereas all diseases were estimated to have a very high economic impact once introduced, the estimated introduction rates differed from low for Rift Valley fever and epizootic haemorrhagic disease to moderate for African horse sickness and very high for West Nile fever. Entry of infected mosquitoes on board of aircraft was deemed the most likely route of introduction for West Nile fever into the Netherlands, followed by entry of infected migratory birds.

Background The Danish Physiotherapy Research Database for chronic patients receiving Free of Charge Physiotherapy (PhysDB-FCP) was piloted over a 1-year period. The purpose of the PhysDB-FCP is to provide a user friendly digital online structured tool that standardizes initial and follow up clinical assessments generating data that can be used for clinical decision making and support future research in physiotherapy for patients with chronic disease. Although initial assessments were completed, the attrition rate was 73% and 90% at 3- and 6- months, respectively, which suggests problems with the current tool. Objective To evaluate the perspectives of the physiotherapists that used the PhysDB-FCP and propose changes to the tool based on this feedback. Materials and methods Fifty of the 103 physiotherapists introduced to the PhysDB-FCP completed an anonymous online survey. Physiotherapists were asked Likert/categorical and yes/no questions on experiences with the PhysDB-FCP within their practice, perceptions of patient experiences, suitability of the resources and support provided by the PhysDB-FCP working group and the ideal administration frequency of the assessments within the PhysDB-FCP. Open ended feedback on possible improvements to the PhysDB-FCP was also collected. Results Physiotherapists agreed that the PhysDB-FCP was useful for taking a physiotherapy assessment (74%) and the patient survey was useful for goal setting (72%). Although physiotherapists felt the PhysDB-FCP was well-defined (82%), only 36% would like to use a similar tool again. Generally, the PhysDB-FCP was too time-consuming, administered too frequently and included irrelevant items. For example, 72% of physiotherapists took >45 min to administer the assessment in the first consultation which was performed over multiple sessions. Conclusions The perspectives of physiotherapists using The PhysDB-FCP suggest specific changes that will ensure better use of the tool in future practice. Changes will likely involve administering the assessment less frequently (every 6-months to 1-year), shortening the assessment, and using diagnosis-specific assessment items.

BackgroundThe Danish Physiotherapy Research Database for chronic patients receiving Free of Charge Physiotherapy (PhysDB-FCP) was piloted over a 1-year period. The purpose of the PhysDB-FCP is to provide a user friendly digital online structured tool that standardizes initial and follow up clinical assessments generating data that can be used for clinical decision making and support future research in physiotherapy for patients with chronic disease. Although initial assessments were completed, the attrition rate was 73% and 90% at 3- and 6- months, respectively, which suggests problems with the current tool.ObjectiveTo evaluate the perspectives of the physiotherapists that used the PhysDB-FCP and propose changes to the tool based on this feedback.Materials and methodsFifty of the 103 physiotherapists introduced to the PhysDB-FCP completed an anonymous online survey. Physiotherapists were asked Likert/categorical and yes/no questions on experiences with the PhysDB-FCP within their practice, perceptions of patient experiences, suitability of the resources and support provided by the PhysDB-FCP working group and the ideal administration frequency of the assessments within the PhysDB-FCP. Open ended feedback on possible improvements to the PhysDB-FCP was also collected.ResultsPhysiotherapists agreed that the PhysDB-FCP was useful for taking a physiotherapy assessment (74%) and the patient survey was useful for goal setting (72%). Although physiotherapists felt the PhysDB-FCP was well-defined (82%), only 36% would like to use a similar tool again. Generally, the PhysDB-FCP was too time-consuming, administered too frequently and included irrelevant items. For example, 72% of physiotherapists took >45 min to administer the assessment in the first consultation which was performed over multiple sessions.ConclusionsThe perspectives of physiotherapists using The PhysDB-FCP suggest specific changes that will ensure better use of the tool in future practice. Changes will likely involve administering the assessment less frequently (every 6-months to 1-year), shortening the assessment, and using diagnosis-specific assessment items.

To evaluate and compare the risk of emerging vector-borne diseases (VBDs), a Model for INTegrated RISK assessment, MINTRISK, was developed to assess the introduction risk of VBDs for new regions in an objective, transparent and repeatable manner. MINTRISK is a web-based calculation tool, that provides semi-quantitative risk scores that can be used for prioritization purposes. Input into MINTRISK is entered by answering questions regarding entry, transmission, establishment, spread, persistence and impact of a selected VBD. Answers can be chosen from qualitative answer categories with accompanying quantitative explanation to ensure consistent answering. The quantitative information is subsequently used as input for the model calculations to estimate the risk for each individual step in the model and for the summarizing output values (rate of introduction; epidemic size; overall risk). The risk assessor can indicate his uncertainty on each answer, and this is accounted for by Monte Carlo simulation. MINTRISK was used to assess the risk of four VBDs (African horse sickness, epizootic haemorrhagic disease, Rift Valley fever, and West Nile fever) for the Netherlands with the aim to prioritise these diseases for preparedness. Results indicated that the overall risk estimate was very high for all evaluated diseases but epizootic haemorrhagic disease. Uncertainty intervals were, however, wide limiting the options for ranking of the diseases. Risk profiles of the VBDs differed. Whereas all diseases were estimated to have a very high economic impact once introduced, the estimated introduction rates differed from low for Rift Valley fever and epizootic haemorrhagic disease to moderate for African horse sickness and very high for West Nile fever. Entry of infected mosquitoes on board of aircraft was deemed the most likely route of introduction for West Nile fever into the Netherlands, followed by entry of infected migratory birds.

BackgroundBacterial vaginosis (BV) is the most common gynecological condition in women of reproductive age and associated with adverse pregnancy outcomes. In the Democratic Republic of the Congo (DRC), neonatal mortality rate is as high as 2.8 percent with preterm birth (PTB) and low birth weight (LBW) as leading causes. Because no studies have addressed BV in DRC, we aimed to investigate the prevalence of BV, the risk factors and the association between BV and adverse pregnancy outcomes in a population of pregnant women from Bukavu, DRC.MethodsA total of 533 pregnant women in the second trimester of pregnancy were recruited in the Provincial Reference Hospital of Bukavu, DRC, between January and October 2017, and followed until delivery. Clinical and sociodemographic data of mother and newborn, and data on (vaginal) hygiene practices, sexual behavior and reproductive history were collected. BV was diagnosed by Nugent scoring of Gram-stained vaginal smears. Two multivariate regression models were built to identify risk factors for BV and to investigate BV as a risk factor for adverse pregnancy outcomes.ResultsThe prevalence of BV was 26.3% and approximately half of the women with BV were asymptomatic. Independent risk factors for BV were the use of alternatives to water for intravaginal washing, concurrent partners, unemployed status, the presence of vaginal Candida and clay consumption. BV was independently associated with both LBW and PTB of an infant with LBW.ConclusionThe prevalence of BV in Bukavu is high but in line with the global average. BV was associated with adverse pregnancy outcomes in our study population. Hence, research on modifiable risk factor-based interventions to reduce the prevalence of BV, and on screening/treatment of BV during antenatal care should be explored to reduce neonatal mortality and morbidity.

ObjectivesDetermine the diagnostic accuracy of two antigen-detecting rapid diagnostic tests (Ag-RDT) for SARS-CoV-2 at the point of care and define individuals' characteristics providing best performance.MethodsWe performed a prospective, single-center, point of care validation of two Ag-RDT in comparison to RT-PCR on nasopharyngeal swabs.ResultsBetween October 9th and 23rd, 2020, 1064 participants were enrolled. The PanbioTM Covid-19 Ag Rapid Test device (Abbott) was validated in 535 participants, with 106 positive Ag-RDT results out of 124 positive RT-PCR individuals, yielding a sensitivity of 85.5% (95% CI: 78.0-91.2). Specificity was 100.0% (95% CI: 99.1-100) in 411 RT-PCR negative individuals. The Standard Q Ag-RDT (SD Biosensor, Roche) was validated in 529 participants, with 170 positive Ag-RDT results out of 191 positive RT-PCR individuals, yielding a sensitivity of 89.0% (95%CI: 83.7-93.1). One false positive result was obtained in 338 RT-PCR negative individuals, yielding a specificity of 99.7% (95%CI: 98.4-100). For individuals presenting with fever 1-5 days post symptom onset, combined Ag-RDT sensitivity was above 95%. Lower sensitivity of 88.2% was seen on the same day of symptom development (day 0).ConclusionsWe provide an independent validation of two widely available commercial Ag-RDTs, both meeting WHO criteria of ≥80% sensitivity and ≥97% specificity. Although less sensitive than RT-PCR, these assays could be beneficial due to their rapid results, ease of use, and independence from existing laboratory structures. Testing criteria focusing on patients with typical symptoms in their early symptomatic period onset could further increase diagnostic value.