Clinical and Laboratory Development of Echinocandin Resistance in Candida glabrata: Molecular Characterization

The pathogenic yeast Candida glabrata has become a public health issue due to the increasing number of echinocandin resistant clinical strains reported. In this study, acquisition and development of resistance to this antifungal class were studied in serial C. glabrata isolates from five patients admitted in two Spanish hospitals with a resistant profile against echinocandins associated with different mutations in hot-spot 1 of FKS2 gene. For two of these patients susceptible FKS wild-type isolates obtained prior to resistant ones were also investigated. Isolates were genotyped using multilocus sequence typing and microsatellite length polymorphism techniques, which yielded comparable results. Susceptible and resistant isolates from the same patient had the same genotype, being sequence type (ST) 3 the most prevalent among them. Isolates with different FKS mutations but the same ST were present in the same patient. MSH2 gene alterations were also studied to investigate their correlation with antifungal resistance acquisition but no association was found with antifungal resistance nor with specific genotypes. In vitro exposure to increasing concentrations of micafungin to susceptible isolates developed colonies carrying FKS mutations in agar plates containing a minimum concentration of 0.06 mg/L of micafungin after less than 48 h of exposure. We investigated the correlation between development of resistance and genotype in a set of susceptible strains after being in vitro exposed to micafungin and anidulafungin but no correlation was found. Mutant prevention concentration values and spontaneous growth frequencies after selection with both echinocandins were statistically similar, although FKS mutant colonies were more abundant after micafungin exposure (p < 0.001). Mutation S663P and F659 deletion were the most common ones found after selection with both echinocandins.This work was supported by the Fondo de Investigación Sanitaria (Grant FI14CIII/00025 to OR-M and research projects PI13/02145 and PI16CIII/00035 to AA-I), and also supported by the Plan Nacional de I+D+i 2013–2016 and Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spanish Network for Research in Infectious Diseases (REIPI RD16CIII/0004/0003) – co-financed by the European Development Regional Fund “A way to achieve Europe,” Operative Program Intelligent Growth 2014–2020.S

TREM1 regulates antifungal immune responses in invasive pulmonary aspergillosis.

Pattern recognition receptors (PRRs) are responsible for Aspergillus fumigatus recognition by innate immunity and its subsequent immune signaling. The triggering receptor expressed on myeloid cells 1 (TREM1) is a recently characterized pro-inflammatory receptor constitutively expressed on the surface of neutrophils and macrophages. A soluble form (sTREM1) of this protein that can be detected in human body fluids has been identified. Here we investigated the role of TREM1 during invasive pulmonary aspergillosis (IPA). IPA patients displayed significantly higher levels of sTREM1 in bronchoalveolar lavages when compared to control patients. Functional analysis in TREM1 showed that the levels of sTREM1 and TREM1 pathway-related cytokines were influenced by single nucleotide polymorphisms in TREM1. In addition, we confirmed a role of TREM1 on antifungal host defense against A. fumigatus in a murine model of IPA. TREM1 deficiency increased susceptibility to infection in the immunosuppressed murine host. Deletion of TREM1 showed delayed innate and adaptive immune responses and impaired pro-inflammatory cytokine responses. The absence of TREM1 in primary macrophages attenuated the TLR signaling by altering the expression of both receptor and effector proteins that are critical to the response against A. fumigatus. In this study, and for the first time, we demonstrate the key role for the TREM1 receptor pathway during IPA.This work was supported by the Fundação para a Ciência e a Tecnologia [PTDC/SAU-SER/29635/2017]; Fundação para a Ciência e a Tecnologia [UIDB/50026/2020 and UIDP/50026/2020]; Fundação para a Ciência e a Tecnologia [PTDC/MED-GEN/28778/2017]; H2020 Excellent Science [NORTE-01-0145-FEDER-000013 and NORTE-01-0145-FEDER-000023)]; Instituto de Salud Carlos III [RD16/CIII/0004/0003]; Instituto de Salud Carlos III [PI18CIII/00045]; Instituto de Salud Carlos III [MPY 1277/15]; Ministerio de Ciencia, Innovación y Universidades [RTI2018-099114-B-I00]; Associação Viver a Ciência (PT) [SFRH/BD/136814/2018]; “la Caixa” Foundation [ID 100010434].S

Clonal chromosomal mosaicism and loss of chromosome Y in elderly men increase vulnerability for SARS-CoV-2

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, COVID-19) had an estimated overall case fatality ratio of 1.38% (pre-vaccination), being 53% higher in males and increasing exponentially with age. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, we found 133 cases (1.42%) with detectable clonal mosaicism for chromosome alterations (mCA) and 226 males (5.08%) with acquired loss of chromosome Y (LOY). Individuals with clonal mosaic events (mCA and/or LOY) showed a 54% increase in the risk of COVID-19 lethality. LOY is associated with transcriptomic biomarkers of immune dysfunction, pro-coagulation activity and cardiovascular risk. Interferon-induced genes involved in the initial immune response to SARS-CoV-2 are also down-regulated in LOY. Thus, mCA and LOY underlie at least part of the sex-biased severity and mortality of COVID-19 in aging patients. Given its potential therapeutic and prognostic relevance, evaluation of clonal mosaicism should be implemented as biomarker of COVID-19 severity in elderly people. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, individuals with clonal mosaic events (clonal mosaicism for chromosome alterations and/or loss of chromosome Y) showed an increased risk of COVID-19 lethality

The Helicobacter pylori Genome Project : insights into H. pylori population structure from analysis of a worldwide collection of complete genomes

Helicobacter pylori, a dominant member of the gastric microbiota, shares co-evolutionary history with humans. This has led to the development of genetically distinct H. pylori subpopulations associated with the geographic origin of the host and with differential gastric disease risk. Here, we provide insights into H. pylori population structure as a part of the Helicobacter pylori Genome Project (HpGP), a multi-disciplinary initiative aimed at elucidating H. pylori pathogenesis and identifying new therapeutic targets. We collected 1011 well-characterized clinical strains from 50 countries and generated high-quality genome sequences. We analysed core genome diversity and population structure of the HpGP dataset and 255 worldwide reference genomes to outline the ancestral contribution to Eurasian, African, and American populations. We found evidence of substantial contribution of population hpNorthAsia and subpopulation hspUral in Northern European H. pylori. The genomes of H. pylori isolated from northern and southern Indigenous Americans differed in that bacteria isolated in northern Indigenous communities were more similar to North Asian H. pylori while the southern had higher relatedness to hpEastAsia. Notably, we also found a highly clonal yet geographically dispersed North American subpopulation, which is negative for the cag pathogenicity island, and present in 7% of sequenced US genomes. We expect the HpGP dataset and the corresponding strains to become a major asset for H. pylori genomics

Healthcare Access and Quality Index based on mortality from causes amenable to personal health care in 195 countries and territories, 1990-2015 : a novel analysis from the Global Burden of Disease Study 2015

Background National levels of personal health-care access and quality can be approximated by measuring mortality rates from causes that should not be fatal in the presence of effective medical care (ie, amenable mortality). Previous analyses of mortality amenable to health care only focused on high-income countries and faced several methodological challenges. In the present analysis, we use the highly standardised cause of death and risk factor estimates generated through the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) to improve and expand the quantification of personal health-care access and quality for 195 countries and territories from 1990 to 2015. Methods We mapped the most widely used list of causes amenable to personal health care developed by Nolte and McKee to 32 GBD causes. We accounted for variations in cause of death certification and misclassifications through the extensive data standardisation processes and redistribution algorithms developed for GBD. To isolate the effects of personal health-care access and quality, we risk-standardised cause-specific mortality rates for each geography-year by removing the joint effects of local environmental and behavioural risks, and adding back the global levels of risk exposure as estimated for GBD 2015. We employed principal component analysis to create a single, interpretable summary measure-the Healthcare Quality and Access (HAQ) Index-on a scale of 0 to 100. The HAQ Index showed strong convergence validity as compared with other health-system indicators, including health expenditure per capita (r= 0.88), an index of 11 universal health coverage interventions (r= 0.83), and human resources for health per 1000 (r= 0.77). We used free disposal hull analysis with bootstrapping to produce a frontier based on the relationship between the HAQ Index and the Socio-demographic Index (SDI), a measure of overall development consisting of income per capita, average years of education, and total fertility rates. This frontier allowed us to better quantify the maximum levels of personal health-care access and quality achieved across the development spectrum, and pinpoint geographies where gaps between observed and potential levels have narrowed or widened over time. Findings Between 1990 and 2015, nearly all countries and territories saw their HAQ Index values improve; nonetheless, the difference between the highest and lowest observed HAQ Index was larger in 2015 than in 1990, ranging from 28.6 to 94.6. Of 195 geographies, 167 had statistically significant increases in HAQ Index levels since 1990, with South Korea, Turkey, Peru, China, and the Maldives recording among the largest gains by 2015. Performance on the HAQ Index and individual causes showed distinct patterns by region and level of development, yet substantial heterogeneities emerged for several causes, including cancers in highest-SDI countries; chronic kidney disease, diabetes, diarrhoeal diseases, and lower respiratory infections among middle-SDI countries; and measles and tetanus among lowest-SDI countries. While the global HAQ Index average rose from 40.7 (95% uncertainty interval, 39.0-42.8) in 1990 to 53.7 (52.2-55.4) in 2015, far less progress occurred in narrowing the gap between observed HAQ Index values and maximum levels achieved; at the global level, the difference between the observed and frontier HAQ Index only decreased from 21.2 in 1990 to 20.1 in 2015. If every country and territory had achieved the highest observed HAQ Index by their corresponding level of SDI, the global average would have been 73.8 in 2015. Several countries, particularly in eastern and western sub-Saharan Africa, reached HAQ Index values similar to or beyond their development levels, whereas others, namely in southern sub-Saharan Africa, the Middle East, and south Asia, lagged behind what geographies of similar development attained between 1990 and 2015. Interpretation This novel extension of the GBD Study shows the untapped potential for personal health-care access and quality improvement across the development spectrum. Amid substantive advances in personal health care at the national level, heterogeneous patterns for individual causes in given countries or territories suggest that few places have consistently achieved optimal health-care access and quality across health-system functions and therapeutic areas. This is especially evident in middle-SDI countries, many of which have recently undergone or are currently experiencing epidemiological transitions. The HAQ Index, if paired with other measures of health-systemcharacteristics such as intervention coverage, could provide a robust avenue for tracking progress on universal health coverage and identifying local priorities for strengthening personal health-care quality and access throughout the world. Copyright (C) The Author(s). Published by Elsevier Ltd.Peer reviewe

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Differential Diagnosis of Fungal Pneumonias vs. Tuberculosis in AIDS Patients by Using Two New Molecular Methods.

Opportunistic fungal pneumonias (OFP) are the main cause of death in AIDS patients worldwide. Diagnosis of these infections is often late as tuberculosis (TB) is frequently the first suspicion. In addition, diagnostic tools have limitations and are unavailable in disadvantaged regions. To perform the differential diagnosis of the main fungi causing OFP in AIDS patients (Histoplasma capsulatum, Cryptococcus neoformans/C. gattii and Pneumocystis jirovecii) vs. the Mycobacterium tuberculosis complex (MTBC), two new assays were developed: (i) a multiplex real-time PCR (MRT-PCR) and (ii) a simple and cost-effective method based on real-time PCR and the analysis of melting curves after amplification (MC-PCR). Both of the techniques were optimized and standardized "in vitro", showing a suitable reproducibility (CV ranged between 1.84 and 3.81% and 1.41 and 4.83%, respectively), a 100% specificity and detection limits between 20 and 2 fg of genomic DNA per 20 µL of reaction. A validation study was performed by retrospectively using 42 clinical samples from 37 patients with proven fungal infection or TB, and 33 controls. The overall sensitivity for the MRT-PCR assay and the MC-PCR assay was 88% and 90.4%, respectively. Both techniques were fast, sensitive and reproducible, allowing for the detection of these pathogens and the performance of a differential diagnosis.This work was supported by research projects PI14CIII/00045 and PI17CIII/00033 from Spanish Fondo de Investigaciones Sanitarias of the Instituto de Salud Carlos III. L.B-M. has a contract supported by the Ministerio de Ciencia e Innovación, Instituto de Salud Carlos III, cofinanced by the European Development Regional Fund (EDRF) “A Way to Achieve Europe” and the Spanish Network for the Research in Infectious Diseases (REIPI; RD16/CIII/0004/0003). A.C.M-A had a short fellowship from Fundación Carolina (call 2017–2018).S

In Vitro Activities of 35 Double Combinations of Antifungal Agents against Scedosporium apiospermum and Scedosporium prolificans▿

Activities of 35 combinations of antifungal agents against Scedosporium spp. were analyzed by a checkerboard microdilution design and the summation of fractional concentration index. An average indifferent effect was detected apart from combinations of azole agents and echinocandins against Scedosporium apiospermum. Antagonism was absent for all antifungal combinations against both species

Value of Serial Quantification of Fungal DNA by a Real-Time PCR-Based Technique for Early Diagnosis of Invasive Aspergillosis in Patients with Febrile Neutropenia ▿

A study was designed to assess the reliability of the serial detection of Aspergillus sp. DNA to diagnose invasive aspergillosis (IA) in patients with febrile neutropenia. Two blood and two serum samples were taken weekly from 83 patients. A total of 2,244 samples were analyzed by real-time quantitative PCR. Twelve (14.4%) patients were diagnosed with IA. Taking two consecutive positive results as the diagnostic criterion, PCR detected 11 cases, with 4 false positives, giving sensitivity, specificity, positive, and negative predictive values of 91.6%, 94.4%, 73.3%, and 98.5%, respectively. On analyzing in conjunction with high-resolution chest tomography (HRCT) and galactomannan (GM) testing, the combination of serial PCR and GM detected 100% of aspergillosis cases, with a positive predictive value of 75.1%. This diagnostic strategy presented, according to CART analysis, a receiver-operator curve with an area under the curve of 0.97 (95% confidence interval, 0.895 to 1.032; P < 0.01), with a relative risk of IA 6.92 times higher than the control population and with predictive success of 95.2%. As regards early diagnosis, the serial detection of Aspergillus DNA took on average 21 days less than HRCT and 68 days less than GM. The serial detection of Aspergillus DNA using real-time quantitative PCR has great diagnostic applicability, which increases when combined with GM quantification