Universal digital high-resolution melting for the detection of pulmonary mold infections.

Invasive mold infections (IMIs) are associated with high morbidity, particularly in immunocompromised patients, with mortality rates between 40% and 80%. Early initiation of appropriate antifungal therapy can substantially improve outcomes, yet early diagnosis remains difficult to establish and often requires multidisciplinary teams evaluating clinical and radiological findings plus supportive mycological findings. Universal digital high-resolution melting (U-dHRM) analysis may enable rapid and robust diagnoses of IMI. A universal fungal assay was developed for U-dHRM and used to generate a database of melt curve signatures for 19 clinically relevant fungal pathogens. A machine learning algorithm (ML) was trained to automatically classify these pathogen curves and detect novel melt curves. Performance was assessed on 73 clinical bronchoalveolar lavage samples from patients suspected of IMI. Novel curves were identified by micropipetting U-dHRM reactions and Sanger sequencing amplicons. U-dHRM achieved 97% overall fungal organism identification accuracy and a turnaround time of ~4 hrs. U-dHRM detected pathogenic molds (Aspergillus, Mucorales, Lomentospora, and Fusarium) in 73% of 30 samples classified as IMI, including mixed infections. Specificity was optimized by requiring the number of pathogenic mold curves detected in a sample to be >8 and a sample volume to be 1 mL, which resulted in 100% specificity in 21 at-risk patients without IMI. U-dHRM showed promise as a separate or combination diagnostic approach to standard mycological tests. U-dHRM's speed, ability to simultaneously identify and quantify clinically relevant mold pathogens in polymicrobial samples, and detect emerging opportunistic pathogens may aid treatment decisions, improving patient outcomes. IMPORTANCE: Improvements in diagnostics for invasive mold infections are urgently needed. This work presents a new molecular detection approach that addresses technical and workflow challenges to provide fast pathogen detection, identification, and quantification that could inform treatment to improve patient outcomes.

Age difference of patients with and without invasive aspergillosis: a systematic review and meta-analysis.

BACKGROUND: Invasive Aspergillosis (IA) is a life-threatening fungal disease with significant mortality rates. Timely diagnosis and treatment greatly enhance patient outcomes. This study aimed to explore the association between patient age and the development of IA, as well as the potential implications for risk stratification strategies. METHODS: We searched National Center for Biotechnology Information (NCBI) databases for publications until October 2023 containing age characteristics of patients with and without IA. A random-effects model with the application of inverse-variance weighting was used to pool reported estimates from each study, and meta-regression and subgroup analyses were utilized to assess sources of heterogeneity. RESULTS: A systematic review was conducted, resulting in the inclusion of 55 retrospective observational studies with a total of 13,983 patients. Meta-analysis revealed that, on average, patients with IA were approximately two and a half years older (95% Confidence Interval [CI] 1.84-3.31 years; I2 = 26.1%) than those without the disease (p < 0.0001). No significant moderators could explain the observed heterogeneity in age difference. However, subgroup analysis revealed that age differences were more pronounced within particular patient groups compared to others. For example, patients with and without IA who had primary severe lung infections exhibited a greater difference in mean age than other patient cohorts. CONCLUSIONS: Further research, such as individual patient data meta-analysis, is necessary to better understand the potential relationship between increasing age and the likelihood of IA. Improved risk stratification strategies based on patient age could potentially enhance the early detection and treatment of IA, ultimately improving patient outcomes.

Knowledge, Attitudes, Practices, and Prevention Barriers Related to Childhood Lead Poisoning Among Nepali-Speaking Bhutanese Parents in Northeast Ohio, United States.

The study objectives were: (i) to develop and administer a survey to assess childhood lead poisoning (CLP) knowledge, attitudes, practices and prevention barriers (KAP-B) among the Nepali-Speaking Bhutanese (NSB) community in Northeast Ohio; and (ii) to examine the association between socio-demographic characteristics of NSB parents and their understanding of CLP as measured by the constructs of knowledge and attitudes. A Nepali language KAP-B questionnaire was developed and 200 NSB parents with at least one child ≤ 7 years of age from the Akron Metropolitan Area, Ohio were interviewed. NSB parents demonstrated a low level of knowledge about CLP prevention measures. While 82% lived in pre-1978 houses, only 27.5% perceived their house/neighborhood to be potentially lead contaminated. Only 33% of the parents reported understanding lead-related information provided by their child's healthcare provider. Low-level CLP awareness among NSB community emphasizes a need for culturally tailored and linguistically appropriate community-level CLP educational intervention programs in this vulnerable community.

Universal Digital High Resolution Melt for the detection of pulmonary mold infections.

Background: Invasive mold infections (IMIs) such as aspergillosis, mucormycosis, fusariosis, and lomentosporiosis are associated with high morbidity and mortality, particularly in immunocompromised patients, with mortality rates as high as 40% to 80%. Outcomes could be substantially improved with early initiation of appropriate antifungal therapy, yet early diagnosis remains difficult to establish and often requires multidisciplinary teams evaluating clinical and radiological findings plus supportive mycological findings. Universal digital high resolution melting analysis (U-dHRM) may enable rapid and robust diagnosis of IMI. This technology aims to accomplish timely pathogen detection at the single genome level by conducting broad-based amplification of microbial barcoding genes in a digital polymerase chain reaction (dPCR) format, followed by high-resolution melting of the DNA amplicons in each digital reaction to generate organism-specific melt curve signatures that are identified by machine learning. Methods: A universal fungal assay was developed for U-dHRM and used to generate a database of melt curve signatures for 19 clinically relevant fungal pathogens. A machine learning algorithm (ML) was trained to automatically classify these 19 fungal melt curves and detect novel melt curves. Performance was assessed on 73 clinical bronchoalveolar lavage (BAL) samples from patients suspected of IMI. Novel curves were identified by micropipetting U-dHRM reactions and Sanger sequencing amplicons. Results: U-dHRM achieved an average of 97% fungal organism identification accuracy and a turn-around-time of 4hrs. Pathogenic molds (Aspergillus, Mucorales, Lomentospora and Fusarium) were detected by U-dHRM in 73% of BALF samples suspected of IMI. Mixtures of pathogenic molds were detected in 19%. U-dHRM demonstrated good sensitivity for IMI, as defined by current diagnostic criteria, when clinical findings were also considered. Conclusions: U-dHRM showed promising performance as a separate or combination diagnostic approach to standard mycological tests. The speed of U-dHRM and its ability to simultaneously identify and quantify clinically relevant mold pathogens in polymicrobial samples as well as detect emerging opportunistic pathogens may provide information that could aid in treatment decisions and improve patient outcomes.

An Evaluation of the OLM PneumID Real-Time Polymerase Chain Reaction to Aid in the Diagnosis of Pneumocystis Pneumonia.

BACKGROUND: The use of the PCR to aid in the diagnosis of Pneumocystis pneumonia (PcP) has demonstrated excellent clinical performance, as evidenced through various systematic reviews and meta-analyses, yet there are concerns over the interpretation of positive results due to the potential presence of Pneumocystis colonization of the airways. While this can be overcome by applying designated positivity thresholds to PCR testing, the shear number of assays described limits the development of a universal threshold. Commercial assays provide the opportunity to overcome this problem, provided satisfactory performance is determined through large-scale, multi-centre evaluations. METHODS: Retrospective case/control and consecutive cohort performance evaluations of the OLM PneumID real-time PCR assay were performed on DNA eluates from a range of samples sent from patients where "in-house" PCR had been performed as part of routine diagnostic testing. The clinical performance of the PneumID assay was determined before including it in a diagnostic algorithm to provide the probability of PcP (dependent on diagnostic evidence). RESULTS: After being used to test 317 patients (32 with PcP), the overall performance of the PneumID assay was found to be excellent (Sensitivity/Specificity: 96.9%/95.1%). False positivity could be removed by applying a threshold specific to sample type (<33.1 cycles for BAL fluid; <37.0 cycles for throat swabs), whereas considering any positive respiratory samples as significant generated 100% sensitivity, making absolute negativity sufficient to exclude PcP. Incorporating the PneumID assay into diagnostic algorithms alongside (1-3)-ß-D-Glucan testing provided high probabilities of PcP (up to 85.2%) when both were positive and very low probabilities (<1%) when both were negative. CONCLUSIONS: The OLM PneumID qPCR provides a commercial option for the accurate diagnosis of PcP, generating excellent sensitivity and specificity, particularly when testing respiratory specimens. The combination of PcP PCR with serum (1-3)-ß-D-Glucan provides excellent clinical utility for diagnosing PcP.

Boyle's Law ignores dynamic processes in governing barotrauma in fish.

The expansion and potential rupture of the swim bladder due to rapid decompression, a major cause of barotrauma injury in fish that pass through turbines and pumps, is generally assumed to be governed by Boyle's Law. In this study, two swim bladder expansion models are presented and tested in silico. One based on the quasi-static Boyle's Law, and a Modified Rayleigh Plesset Model (MRPM), which includes both inertial and pressure functions and was parametrised to be representative of a fish swim bladder. The two models were tested using a range of: (1) simulated and (2) empirically derived pressure profiles. Our results highlight a range of conditions where the Boyle's Law model (BLM) is inappropriate for predicting swim bladder size in response to pressure change and that these conditions occur in situ, indicating that this is an applied and not just theoretical issue. Specifically, these conditions include any one, or any combination, of the following factors: (1) when rate of pressure change is anything but very slow compared to the resonant frequency of the swim bladder; (2) when the nadir pressure is near or at absolute zero; and (3) when a fish experiences liquid tensions (i.e. negative absolute pressures). Under each of these conditions, the MRPM is more appropriate tool for predicting swim bladder size in response to pressure change and hence it is a better model for quantifying barotrauma in fish.

Co-design with Integrated Care Teams: Establishing Information Needs.

Introduction: Co-design has been cited as playing a major role in the future of effective integrated care, however, there is a lack of reporting and reflection on the methods used. Information sharing is fundamental when working in integrated care, however sharing across professions, service settings and localities can be complex. Through co-design, we seek to establish a shared understanding of information needs within a newly formed integrated care team. In doing so we aim to inform future practice in the understanding of co-design. Description: Co-design Workshop 1 (N = 24 participants, plus 6 facilitators), collected 'Current Position' understanding of service information needs. Co-design Workshop 2 (N = 18 participants, plus 6 facilitators) sought a 'Future Position' understanding, identifying solutions and next steps for establishing information-need solutions. Reflection on the co-design process was conducted to inform future co-design practices. Conclusion: Identified was a wide range of future service information needs under the themes of Culture Building, Health System Needs, and Processes. We conclude with 4 key learning points on co-designing. 1. Ensure simplicity in format. 2. Interdisciplinary co-design and co-facilitation of workshops are beneficial. 3. Planning and preparation are key. 4. Co-designing can enhance communication for service improvement.

An Overview of Systematic Reviews of Polymerase Chain Reaction (PCR) for the Diagnosis of Invasive Aspergillosis in Immunocompromised People: A Report of the Fungal PCR Initiative (FPCRI)-An ISHAM Working Group.

This overview of reviews (i.e., an umbrella review) is designed to reappraise the validity of systematic reviews (SRs) and meta-analyses related to the performance of Aspergillus PCR tests for the diagnosis of invasive aspergillosis in immunocompromised patients. The methodological quality of the SRs was assessed using the AMSTAR-2 checklist; the quality of the evidence (QOE) within each SR was appraised following the GRADE approach. Eight out of 12 SRs were evaluated for qualitative and quantitative assessment. Five SRs evaluated Aspergillus PCR on bronchoalveolar lavage fluid (BAL) and three on blood specimens. The eight SRs included 167 overlapping reports (59 evaluating PCR in blood specimens, and 108 in BAL), based on 107 individual primary studies (98 trials with a cohort design, and 19 with a case-control design). In BAL specimens, the mean sensitivity and specificity ranged from 0.57 to 0.91, and from 0.92 to 0.97, respectively (QOE: very low to low). In blood specimens (whole blood or serum), the mean sensitivity ranged from 0.57 to 0.84, and the mean specificity from 0.58 to 0.95 (QOE: low to moderate). Across studies, only a low proportion of AMSTAR-2 critical domains were unmet (1.8%), demonstrating a high quality of methodological assessment. Conclusions. Based on the overall methodological assessment of the reviews included, on average we can have high confidence in the quality of results generated by the SRs.

An update on current and novel molecular diagnostics for the diagnosis of invasive fungal infections.

BACKGROUND: Invasive fungal infections cause millions of infections annually, but diagnosis remains challenging. There is an increased need for low-cost, easy to use, highly sensitive and specific molecular assays that can differentiate between colonized and pathogenic organisms from different clinical specimens. AREAS COVERED: We reviewed the literature evaluating the current state of molecular diagnostics for invasive fungal infections, focusing on current and novel molecular tests such as polymerase chain reaction (PCR), digital PCR, high-resolution melt (HRM), and metagenomics/next generation sequencing (mNGS). EXPERT OPINION: PCR is highly sensitive and specific, although performance can be impacted by prior/concurrent antifungal use. PCR assays can identify mutations associated with antifungal resistance, non-Aspergillus mold infections, and infections from endemic fungi. HRM is a rapid and highly sensitive diagnostic modality that can identify a wide range of fungal pathogens, including down to the species level, but multiplex assays are limited and HRM is currently unavailable in most healthcare settings, although universal HRM is working to overcome this limitation. mNGS offers a promising approach for rapid and hypothesis-free diagnosis of a wide range of fungal pathogens, although some drawbacks include limited access, variable performance across platforms, the expertise and costs associated with this method, and long turnaround times in real-world settings.

COVID-19-associated pulmonary aspergillosis in mechanically ventilated patients: a prospective, multicentre UK study.

BACKGROUND: Invasive pulmonary aspergillosis is a complication of severe COVID-19, with regional variation in reported incidence and mortality. We describe the incidence, risk factors and mortality associated with COVID-19-associated pulmonary aspergillosis (CAPA) in a prospective, multicentre UK cohort. METHODS: From March 2020 to March 2021, 266 mechanically ventilated adults with COVID-19 were enrolled across 5 UK hospital intensive care units (ICUs). CAPA was defined using European Confederation for Medical Mycology and the International Society for Human and Animal Mycology criteria and fungal diagnostics performed on respiratory and serum samples. RESULTS: Twenty-nine of 266 patients (10.9%) had probable CAPA, 14 (5.2%) possible CAPA and none proven CAPA. Probable CAPA was diagnosed a median of 9 (IQR 7-16) days after ICU admission. Factors associated with probable CAPA after multivariable logistic regression were cumulative steroid dose given within 28 days prior to ICU admission (adjusted OR (aOR) 1.16; 95% CI 1.01 to 1.43 per 100 mg prednisolone-equivalent), receipt of an interleukin (IL)-6 inhibitor (aOR 2.79; 95% CI 1.22 to 6.48) and chronic obstructive pulmonary disease (COPD) (aOR 4.78; 95% CI 1.13 to 18.13). Mortality in patients with probable CAPA was 55%, vs 46% in those without. After adjustment for immortal time bias, CAPA was associated with an increased risk of 90-day mortality (HR 1.85; 95% CI 1.07 to 3.19); however, this association did not remain statistically significant after further adjustment for confounders (adjusted HR 1.57; 95% CI 0.88 to 2.80). There was no difference in mortality between patients with CAPA prescribed antifungals (9 of 17; 53%) and those who were not (7 of 12; 58%) (p=0.77). INTERPRETATION: In this first prospective UK study, probable CAPA was associated with corticosteroid use, receipt of IL-6 inhibitors and pre-existing COPD. CAPA did not impact mortality following adjustment for prognostic variables.

Predictors for Prolonged Hospital Stay Solely to Complete Intravenous Antifungal Treatment in Patients with Candidemia: Results from the ECMM Candida III Multinational European Observational Cohort Study.

BACKGROUND: To date, azoles represent the only viable option for oral treatment of invasive Candida infections, while rates of azole resistance among non-albicans Candida spp. continue to increase. The objective of this sub-analysis of the European multicenter observational cohort study Candida III was to describe demographical and clinical characteristics of the cohort requiring prolonged hospitalization solely to complete intravenous (iv) antifungal treatment (AF Tx). METHODS: Each participating hospital (number of eligible hospitals per country determined by population size) included the first ~ 10 blood culture proven adult candidemia cases occurring consecutively after July 1st, 2018, and treating physicians answered the question on whether hospital stay was prolonged only for completion of intravenous antifungal therapy. Descriptive analyses as well as binary logistic regression was used to assess for predictors of prolonged hospitalization solely to complete iv AF Tx. FINDINGS: Hospital stay was prolonged solely for the completion of iv AF Tx in 16% (100/621) of candidemia cases by a median of 16 days (IQR 8 - 28). In the multivariable model, initial echinocandin treatment was a positive predictor for prolonged hospitalization to complete iv AF Tx (aOR 2.87, 95% CI 1.55 - 5.32, p < 0.001), while (i) neutropenia, (ii) intensive care unit admission, (iii) catheter related candidemia, (iv) total parenteral nutrition, and (v) C. parapsilosis as causative pathogen were found to be negative predictors (aOR 0.22 - 0.45; p < 0.03). INTERPRETATION: Hospital stays were prolonged due to need of iv AF Tx in 16% of patients with candidemia. Those patients were more likely to receive echinocandins as initial treatment and were less severely ill and less likely infected with C. parapsilosis.

Pneumocystis jirovecii pneumonia in intensive care units: a multicenter study by ESGCIP and EFISG.

BACKGROUND: Pneumocystis jirovecii pneumonia (PJP) is an opportunistic, life-threatening disease commonly affecting immunocompromised patients. The distribution of predisposing diseases or conditions in critically ill patients admitted to intensive care unit (ICU) and subjected to diagnostic work-up for PJP has seldom been explored. MATERIALS AND METHODS: The primary objective of the study was to describe the characteristics of ICU patients subjected to diagnostic workup for PJP. The secondary objectives were: (i) to assess demographic and clinical variables associated with PJP; (ii) to assess the performance of Pneumocystis PCR on respiratory specimens and serum BDG for the diagnosis of PJP; (iii) to describe 30-day and 90-day mortality in the study population. RESULTS: Overall, 600 patients were included in the study, of whom 115 had presumptive/proven PJP (19.2%). Only 8.8% of ICU patients subjected to diagnostic workup for PJP had HIV infection, whereas hematological malignancy, solid tumor, inflammatory diseases, and solid organ transplants were present in 23.2%, 16.2%, 15.5%, and 10.0% of tested patients, respectively. In multivariable analysis, AIDS (odds ratio [OR] 3.31; 95% confidence interval [CI] 1.13-9.64, p = 0.029), non-Hodgkin lymphoma (OR 3.71; 95% CI 1.23-11.18, p = 0.020), vasculitis (OR 5.95; 95% CI 1.07-33.22, p = 0.042), metastatic solid tumor (OR 4.31; 95% CI 1.76-10.53, p = 0.001), and bilateral ground glass on CT scan (OR 2.19; 95% CI 1.01-4.78, p = 0.048) were associated with PJP, whereas an inverse association was observed for increasing lymphocyte cell count (OR 0.64; 95% CI 0.42-1.00, p = 0.049). For the diagnosis of PJP, higher positive predictive value (PPV) was observed when both respiratory Pneumocystis PCR and serum BDG were positive compared to individual assay positivity (72% for the combination vs. 63% for PCR and 39% for BDG). Cumulative 30-day mortality and 90-day mortality in patients with presumptive/proven PJP were 52% and 67%, respectively. CONCLUSION: PJP in critically ill patients admitted to ICU is nowadays most encountered in non-HIV patients. Serum BDG when used in combination with respiratory Pneumocystis PCR could help improve the certainty of PJP diagnosis.

Focused Ultrasound Thalamotomy for Tremor in Parkinson's Disease: Outcomes in a Large, Prospective Cohort.

BACKGROUND: Magnetic resonance guided focused ultrasound (MRgFUS) is United States Food and Drug Administration approved for the treatment of tremor-dominant Parkinson's disease (TdPD), but only limited studies have been described in practice. OBJECTIVES: To report the largest prospective experience of unilateral MRgFUS thalamotomy for the treatment of medically refractory TdPD. METHODS: Clinical outcomes of 48 patients with medically refractory TdPD who underwent MRgFUS thalamotomy were evaluated. Tremor outcomes were assessed using the Fahn-Tolosa-Marin scale and adverse effects were categorized using a structured questionnaire and clinical exam at 1 month (n = 44), 3 months (n = 34), 1 year (n = 22), 2 years (n = 5), and 3 years (n = 2). Patients underwent magnetic resonance imaging <24 hours post-procedure. RESULTS: Significant tremor control persisted at all follow-ups (P < 0.001). All side effects were mild. At 3 months, these included gait imbalance (38.24%), sensory deficits (26.47%), motor weakness (17.65%), dysgeusia (5.88%), and dysarthria (5.88%), with some persisting at 1 year. CONCLUSIONS: MRgFUS thalamotomy is an effective treatment for sustained tremor control in patients with TdPD. © 2023 International Parkinson and Movement Disorder Society.

European candidaemia is characterised by notable differential epidemiology and susceptibility pattern: Results from the ECMM Candida III study.

The objectives of this study were to assess Candida spp. distribution and antifungal resistance of candidaemia across Europe. Isolates were collected as part of the third ECMM Candida European multicentre observational study, conducted from 01 to 07-07-2018 to 31-03-2022. Each centre (maximum number/country determined by population size) included ∼10 consecutive cases. Isolates were referred to central laboratories and identified by morphology and MALDI-TOF, supplemented by ITS-sequencing when needed. EUCAST MICs were determined for five antifungals. fks sequencing was performed for echinocandin resistant isolates. The 399 isolates from 41 centres in 17 countries included C. albicans (47.1%), C. glabrata (22.3%), C. parapsilosis (15.0%), C. tropicalis (6.3%), C. dubliniensis and C. krusei (2.3% each) and other species (4.8%). Austria had the highest C. albicans proportion (77%), Czech Republic, France and UK the highest C. glabrata proportions (25-33%) while Italy and Turkey had the highest C. parapsilosis proportions (24-26%). All isolates were amphotericin B susceptible. Fluconazole resistance was found in 4% C. tropicalis, 12% C. glabrata (from six countries across Europe), 17% C. parapsilosis (from Greece, Italy, and Turkey) and 20% other Candida spp. Four isolates were anidulafungin and micafungin resistant/non-wild-type and five resistant to micafungin only. Three/3 and 2/5 of these were sequenced and harboured fks-alterations including a novel L657W in C. parapsilosis. The epidemiology varied among centres and countries. Acquired echinocandin resistance was rare but included differential susceptibility to anidulafungin and micafungin, and resistant C. parapsilosis. Fluconazole and voriconazole cross-resistance was common in C. glabrata and C. parapsilosis but with different geographical prevalence.

The co-creation, initial piloting, and protocol for a cluster randomised controlled trial of a coach-led positive body image intervention for girls in sport.

BACKGROUND: Globally, girls disengage from sports at an earlier age and higher rate than boys. This is, in part, due to the unique body image challenges that girls face, relative to their male peers. Existing intervention efforts that aim to reduce girls' negative body image and movement experiences have proven marginally effective, if not ineffective. This paper outlines the co-creation, initial piloting and protocol for a cluster randomised controlled trial of Body Confident Athletes (BCA); an in-person, coach-led intervention that aims to foster positive body image and sports enjoyment among girls. METHODS: Following co-creation and an initial pilot, a two-armed cluster randomised controlled trial will assess the immediate (post-intervention) and short-term (1-month and 3-month follow-up) impact of BCA on girls' (N = 1,036; 11-17 years old) body image, sports enjoyment, and affect. Sport organisations will be randomly allocated (1:1) into either an intervention or waitlist control condition. Girls and coaches in the intervention condition will complete three 60-minute sessions over three consecutive weeks. The primary outcome will be the immediate change in girls' body esteem, with secondary outcomes assessing the immediate and short-term changes in girls' body appreciation, self-objectification, attuned self-care, sports enjoyment, and affect. DISCUSSION: This research is the first to utilise an international multi-stakeholder partnership to co-create and evaluate an intervention that addresses the intersection of girls' body image and sport experiences. The theoretical and methodological considerations of this research have led to a feasible intervention and trial protocol, and if proven effective, BCA may assist in reducing the global gender disparity in sports participation. TRIAL REGISTRATION NUMBER: NCT05594524 , registered 25th October 2022.