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Active surveillance for laboratory-confirmed Salmonella serotype Enteritidis (SE) infection revealed a decline in incidence in the 1990s, followed by an increase starting in 2000. We sought to determine if the fluctuation in SE incidence could be explained by changes in foodborne sources of infection. We conducted a population-based case-control study of sporadic SE infection in five of the Foodborne Diseases Active Surveillance Network (FoodNet) sites during a 12-month period in 2002-2003. A total of 218 cases and 742 controls were enrolled. Sixty-seven (31%) of the 218 case-patients and six (1%) of the 742 controls reported travel outside the United States during the 5 days before the case's illness onset (OR 53, 95% CI 23-125). Eighty-one percent of cases with SE phage type 4 travelled internationally. Among persons who did not travel internationally, eating chicken prepared outside the home and undercooked eggs inside the home were associated with SE infections. Contact with birds and reptiles was also associated with SE infections. This study supports the findings of previous case-control studies and identifies risk factors associated with specific phage types and molecular subtypes. [ABSTRACT FROM AUTHOR]

Introduction: While Pneumocystis pneumonia (PcP) remains a major AIDS-defining disease, the majority of cases of PcP now present in the HIV-negative cohort, causing significant mortality. PcP PCR diagnosis is not novel, and the optimal route of diagnosis remains unclear, with an imperfect reference method and complexity in result interpretation for alternative tests., Areas Covered: This extensive review utilizing a literature search underpinning a recent systematic review/meta-analysis discusses the technical and clinical performance of PcP PCR, the added benefits of PCR testing, future aspects/considerations, and how PCR may be best used in clinical algorithms to provide a probability of PcP., Expert Opinion: Given the current imperfect reference test for PcP, an alternative would be beneficial. Concerns over PcP PCR generating false positive results are valid but can be resolved by using positivity thresholds that drive specificity. Unfortunately, the extensive range of PCR assays complicates the provision of a PCR reference method. Combination testing incorporating PCR and B-D-Glucan, along with clinical and host risk factors, is key to understanding the individual probability of PcP. It is critical that access to PcP PCR testing is improved through technical and logistical development. Conversely, syndromic approaches including PcP need to be fully evaluated.

Medically important pathogenic fungi invade vertebrate tissue and are considered primary when part of their nature life cycle is associated with an animal host and are usually able to infect immunocompetent hosts. Opportunistic fungal pathogens complete their life cycle in environmental habitats or occur as commensals within or on the vertebrate body, but under certain conditions can thrive upon infecting humans. The extent of host damage in opportunistic infections largely depends on the portal and modality of entry as well as on the host's immune and metabolic status. Diseases caused by primary pathogens and common opportunists, causing the top approximately 80% of fungal diseases [D. W. Denning, Lancet Infect Dis, 24:e428-e438, 2024, https://doi.org/10.1016/S1473-3099(23)00692-8], tend to follow a predictive pattern, while those by occasional opportunists are more variable. For this reason, it is recommended that diseases caused by primary pathogens and the common opportunists are named after the etiologic agent, for example, histoplasmosis and aspergillosis, while this should not be done for occasional opportunists that should be named as [causative fungus] [clinical syndrome], for example, Alternaria alternata cutaneous infection. The addition of a descriptor that identifies the location or clinical type of infection is required, as the general name alone may cover widely different clinical syndromes, for example, "rhinocerebral mucormycosis." A list of major recommended human and animal disease entities (nomenclature) is provided in alignment with their causative agents. Fungal disease names may encompass several genera of etiologic agents, consequently being less susceptible to taxonomic changes of the causative species, for example, mucormycosis covers numerous mucormycetous molds.

Purpose of Review: This review describes the current status of diagnosing invasive mould disease and Pneumocystis pneumonia using nonconventional diagnostics methods., Recent Findings: There has been significant development in the range of nonculture mycological tests. Lateral flow tests (LFTs) for diagnosing aspergillosis complement galactomannan ELISA testing, and LFTs for other fungal diseases are in development. Rapid and low through-put B-D-Glucan assays increase access to testing and there has been significant progress in the standardization/development of molecular tests. Despite this, no single perfect test exists and combining tests (e.g., antigen and molecular testing) is likely required for the optimal diagnosis of most fungal diseases., Summary: Based on established clinical performance few mycological tests can be used alone for optimal diagnosis of fungal disease (FD) and combining tests, including classical approaches is the preferred route for confirming and excluding disease. Next-generation sequencing will likely play an increasing role in how we diagnose disease, but optimization, standardization and validation of the entire molecular process is needed and we must consider how host biomarkers can stratify risk. Given the burden of FD in low- and medium-income countries, improved access to novel but more so existing diagnostic testing is critical along with simplification of testing processes., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Background: This meta-analysis examines the comparative diagnostic performance of polymerase chain reaction (PCR) for the diagnosis of Pneumocystis pneumonia (PCP) on different respiratory tract samples, in both human immunodeficiency virus (HIV) and non-HIV populations., Methods: A total of 55 articles met inclusion criteria, including 11 434 PCR assays on respiratory specimens from 7835 patients at risk of PCP. QUADAS-2 tool indicated low risk of bias across all studies. Using a bivariate and random-effects meta-regression analysis, the diagnostic performance of PCR against the European Organisation for Research and Treatment of Cancer-Mycoses Study Group definition of proven PCP was examined., Results: Quantitative PCR (qPCR) on bronchoalveolar lavage fluid provided the highest pooled sensitivity of 98.7% (95% confidence interval [CI], 96.8%-99.5%), adequate specificity of 89.3% (95% CI, 84.4%-92.7%), negative likelihood ratio (LR-) of 0.014, and positive likelihood ratio (LR+) of 9.19. qPCR on induced sputum provided similarly high sensitivity of 99.0% (95% CI, 94.4%-99.3%) but a reduced specificity of 81.5% (95% CI, 72.1%-88.3%), LR- of 0.024, and LR+ of 5.30. qPCR on upper respiratory tract samples provided lower sensitivity of 89.2% (95% CI, 71.0%-96.5%), high specificity of 90.5% (95% CI, 80.9%-95.5%), LR- of 0.120, and LR+ of 9.34. There was no significant difference in sensitivity and specificity of PCR according to HIV status of patients., Conclusions: On deeper respiratory tract specimens, PCR negativity can be used to confidently exclude PCP, but PCR positivity will likely require clinical interpretation to distinguish between colonization and active infection, partially dependent on the strength of the PCR signal (indicative of fungal burden), the specimen type, and patient population tested., Competing Interests: Potential conflicts of interest. A. A. has received honoraria from Gilead Sciences. S. C.-A. A. has received united educational grants from F2G Ltd and consulted for MSD Australia. T. R. R. has received grants from Pfizer and the Irish Department of Agriculture, Food and the Marine and honoraria from Gilead and Menarini Pharma. P. L. W. has performed diagnostic evaluations and received meeting sponsorship from Associates of Cape Cod, Bruker, Dynamiker, and Launch Diagnostics; speaker fees, expert advice fees, and meeting sponsorship from Gilead; and speaker and expert advice fees from F2G. J. P. D. has received personal fees from F2G, Gilead Sciences, and Pfizer. W. J. H. has received lecture honoraria from AbbVie, Amgen, AstraZeneca, Celgene/BMS, Gilead Sciences, Janssen, MSD, and Pfizer; has received support to attend meetings and travel support from AbbVie, Alexion, Astellas, Janssen, Lilly, MSD, Novartis, and Pfizer; and has participated on advisory boards for AbbVie, Amgen, AstraZeneca, Basilea, Celgene/BMS, Gilead Sciences, Janssen, and Sanofi-Aventis. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Fungal diseases represent a considerable global health concern, affecting >1 billion people annually. In response to this growing challenge, the World Health Organization introduced the pivotal fungal priority pathogens list (FPPL) in late 2022. The FPPL highlights the challenges in estimating the global burden of fungal diseases and antifungal resistance (AFR), as well as limited surveillance capabilities and lack of routine AFR testing. Furthermore, training programs should incorporate sufficient information on fungal diseases, necessitating global advocacy to educate health care professionals and scientists. Established international guidelines and the FPPL are vital in strengthening local guidance on tackling fungal diseases. Future iterations of the FPPL have the potential to refine the list further, addressing its limitations and advancing our collective ability to combat fungal diseases effectively. Napp Pharmaceuticals Limited (Mundipharma UK) organized a workshop with key experts from Northern Europe to discuss the impact of the FPPL on regional clinical practice., Competing Interests: Potential conflicts of interest. M.C.A. has, over the past 5 years, received research grants/contract work (paid to the Statens Serum Institute) from Cidara, F2G, Gilead, and Scynexis and speaker honoraria (personal fees) from Astellas, Chiesi, Gilead, and F2G; she is also the current Chair of EUCAST-AFST. D.A.J. has share options in pulmocide. A.B. has no conflicts of interest. D.D. holds founder shares in F2G Ltd., an antifungal discovery company spun out from the University of Manchester, and share options in TFF Pharma; acts or has recently acted as a consultant to Biosergen, Lifemine Therapeutics, Mucpharm PTY, Pulmatrix, Pulmocide, Rostra Therapeutics, and TFF Pharmaceuticals; has received payments for talks on behalf of Avir, Basilea, BioRad, Gilead, Mundipharma, and Pfizer in the past 3 years; and has been involved in multiple guideline groups, primarily focused on diagnostics and aspergillosis. M.F. has received speaker fees and funding from Gilead Scientific and received funding from the Leverhulme Trust and the Welcome Trust. R.G. has no conflicts of interest. J.M. reports support consulting fees from Amplyx, Basilea, Cidara, F2G, Gilead, Mundipharma, Pfizer, Scynexis, and Takeda; honoraria for lectures from Astellas, Basilea, Gilead, MedScape, Mundipharma, Pfizer, Shionogi, and Takeda; payment for expert testimony from Cidara; and participation on advisory boards with Basilea, Cidara, Pulmocide, Sfunga, and Shionogi. I.M.L. has received lecturer honoraria and has participated on advisory boards with Accelerate, bioMérieux, Biotest, Clinigen, Fresenius Kabi, Gilead, MSD, Mundipharma, Polyphor, and ThermoFisher. V.M. has received speaker honoraria and participated on advisory boards and received research funding from different industry partners including Gilead, Pfizer, Mundipharma, MSD, Novartis, Jazz Pharma, Therakos, Sanofi. T.M. reports consultancy fees from AstraZeneca, Gilead Sciences, Pfizer, and Roche; research grants from Gilead Sciences; and travel support from AstraZeneca, Amgen, and Pfizer. J.P. has given a presentation for Pfizer. R.R. has received lecturer honoraria from Astellas, Mundipharma, Basilea, Gilead, and Pfizer and has participated as a clinical investigator for a Phase 2 clinical trial conducted by Scynexis and as a principal investigator for a Phase 2 clinical trial conducted by F2G. R.W. has no conflicts of interest. N.A. works at Napp Pharmaceuticals Ltd. which is a member of the Mundipharma network of independent associated companies. L.W. performed diagnostic evaluations and received meeting sponsorship from Associates of Cape Cod, Bruker, Dynamiker, and Launch Diagnostics; speaker's fees, expert advice fees, and meeting sponsorship from Gilead; speaker and expert advice fees from Pfizer and Mundipharma; and expert advice fees from F2G., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

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., Competing Interests: M.H. and S.I.F. received research funding from Astellas for a portion of this work. M.H. also received research funding from Gilead, MSD, Euroimmune, IMMY, Scynexis, Pulmocide, F2G, and Pfizer, all outside the submitted work. S.I.F. is a scientific cofounder, director, and advisor of MelioLabs Inc. and has an equity interest in the company. J.D.J. received research funding from Astellas, F2G, and Pfizer, all outside of the submitted work. M.S. is co-founder and CEO of Melio and has equity interest. R.S., A.K., and A.S. are employees of Melio. NICHD award number R41HD107895 and NIAID award number R01AI134982 have been identified for conflict of interest management based on the overall scope of the project and its potential benefit to MelioLabs Inc.; however, the research findings included in this particular publication may not necessarily relate to the interests of MelioLabs Inc. The terms of this arrangement have been reviewed and approved by the University of California, San Diego, in accordance with its conflict of interest policies. All other authors declare no conflicts of interests related to this work.

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; I 2  = 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., (© 2024. The Author(s).)

The emergence of fungal antimicrobial resistance-fAMR-is having a growing impact on human and animal health, and food security. This roadmap charts inter-related actions that will enhance our ability to mitigate the risk of fAMR. As humanity's reliance on antifungal chemicals escalates, our understanding of their one-health consequences needs to scale accordingly if we are to protect our ability to manage the global spectrum of fungal disease sustainably., Competing Interests: Competing interestsM.C.F. is an editor for npj Antimicrobials and Resistance. The remaining authors declare no competing interests, (© The Author(s) 2024.)

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., Competing Interests: Competing interests: MPW, WH and TB have received speaker fees from Gilead Sciences. JY has received honoraria from Pfizer for contributing to a CAPA working group. TB has received Advisory Board fees from Gilead Sciences and Mundipharma and funding from MSD and Pfizer. DRJ is the president of the British Society for Antimicrobial Chemotherapy and has received honoraria from Pfizer, Shionogi, Menarini and Tillots. SS has received honoraria from Pfizer and Gilead for educational purposes. PLW performed diagnostic evaluations and received meeting sponsorship from Associates of Cape Cod, Bruker, Dynamiker and Launch Diagnostics; speaker fees, expert advice fees and meeting sponsorship from Gilead; and speaker and expert advice fees from Pfizer and expert advice fees from F2G., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

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., (© 2023. The Author(s).)

Global changes in transcriptional regulation and RNA metabolism are crucial features of cancer development. However, little is known about the role of the core promoter in defining transcript identity and post-transcriptional fates, a potentially crucial layer of transcriptional regulation in cancer. In this study, we use CAGE-seq analysis to uncover widespread use of dual-initiation promoters in which non-canonical, first-base-cytosine (C) transcription initiation occurs alongside first-base-purine initiation across 59 human cancers and healthy tissues. C-initiation is often followed by a 5' terminal oligopyrimidine (5'TOP) sequence, dramatically increasing the range of genes potentially subjected to 5'TOP-associated post-transcriptional regulation. We show selective, dynamic switching between purine and C-initiation site usage, indicating transcription initiation-level regulation in cancers. We additionally detail global metabolic changes in C-initiation transcripts that mark differentiation status, proliferative capacity, radiosensitivity, and response to irradiation and to PI3K-Akt-mTOR and DNA damage pathway-targeted radiosensitization therapies in colorectal cancer organoids and cancer cell lines and tissues., (© 2023. The Author(s).)

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 Pneum ID 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 Pneum ID 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 Pneum ID 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 Pneum ID assay into diagnostic algorithms alongside (1-3)-β-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 Pneum ID 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)-β-D-Glucan provides excellent clinical utility for diagnosing PcP.

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.

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.

Pneumocystis pneumonia (PcP) is a serious complication of many significant immunocompromising conditions. Prior incidence estimates in Wales are based on PcP's presentation in the HIV and transplant populations. The objectives were to describe the incidence of PcP in Wales using laboratory reporting measures and assess the impact of underlying immunosuppression cause on mortality. All positive PCR results for PcP between 2015 and 2018 were identified. The total number of unique positives with clinical and radiological correlation was 159 patients, a mean of 39.75 annually. The healthcare records of these patients were reviewed. The mortality at one month was 35.2% and 49.1% at one year. HIV remains the commonest cause of immunosuppression but has lower mortality than non-HIV conditions (12% vs. 59% at one year, p < 0.00001). Non-HIV conditions were categorised as life-threatening and non-life threatening but had a non-significant mortality (66% vs. 54%; p = 0.149), highlighting the negative impact of PcP. An incidence of PcP in Wales of 1.23-1.26 cases per 100,000 has been identified, 32-35% greater than the upper limit previously estimated. There is high mortality in non-HIV patients regardless of immunosuppression cause. A heightened awareness of PcP in these groups will hasten diagnosis and potentially improve mortality.

Purpose of Review: The true incidence of fungal disease is hampered by conventionally poor diagnostic tests, limited access to advanced diagnostics, and limited surveillance. The availability of serological testing has been available for over two decades and generally underpins the modern diagnosis of the most common forms of fungal disease. This review will focus on technical developments of serological tests for the diagnosis of fungal disease, describing advances in clinical performance when available., Recent Findings: Despite their longevity, technical, clinical, and performance limitations remain, and tests specific for fungal pathogens outside the main pathogens are lacking. The availability of LFA and automated systems, capable of running multiple different tests, represents significant developments, but clinical performance data is variable and limited., Summary: Fungal serology has significantly advanced the diagnosis of the main fungal infections, with LFA availability increasing accessibility to testing. Combination testing has the potential to overcome performance limitations., Competing Interests: Conflict of InterestPLW performed diagnostic evaluations and received meeting sponsorship from Bruker; speakers fees, expert advice fees, and meeting sponsorship from Gilead; expert advice fees from F2G and speaker fees from Pfizer; speakers fees and performed diagnostic evaluations for Associates of Cape Cod and IMMY., (© The Author(s) 2023.)

Timely diagnosis remains an unmet need in non-neutropenic patients at risk for aspergillosis, including those with COVID-19-associated pulmonary aspergillosis (CAPA), which in its early stages is characterized by tissue-invasive growth of the lungs with limited angioinvasion. Currently available mycological tests show limited sensitivity when testing blood specimens. Metagenomic next-generation sequencing (mNGS) to detect microbial cell-free DNA (mcfDNA) in plasma might overcome some of the limitations of conventional diagnostics. A two-center cohort study involving 114 COVID-19 intensive care unit patients evaluated the performance of plasma mcfDNA sequencing for the diagnosis of CAPA. Classification of CAPA was performed using the European Confederation for Medical Mycology (ECMM)/International Society for Human and Animal Mycoses (ISHAM) criteria. A total of 218 plasma samples were collected between April 2020 and June 2021 and tested for mcfDNA (Karius test). Only 6 patients were classified as probable CAPA, and 2 were classified as possible, while 106 patients did not fulfill CAPA criteria. The Karius test detected DNA of mold pathogens in 12 samples from 8 patients, including Aspergillus fumigatus in 10 samples from 6 patients. Mold pathogen DNA was detected in 5 of 6 (83% sensitivity) cases with probable CAPA (A. fumigatus in 8 samples from 4 patients and Rhizopus microsporus in 1 sample), while the test did not detect molds in 103 of 106 (97% specificity) cases without CAPA. The Karius test showed promising performance for diagnosis of CAPA when testing plasma, being highly specific. The test detected molds in all but one patient with probable CAPA, including cases where other mycological tests from blood resulted continuously negative, outlining the need for validation in larger studies.

Invasive fungal disease (IFD) causes severe morbidity and mortality, and the number of IFD cases is increasing. Exposure to opportunistic fungal pathogens is inevitable, but not all patients with underlying diseases increasing susceptibility to IFD, develop it. IFD diagnosis currently uses fungal biomarkers and clinical risk/presentation to stratify high-risk patients and classifies them into possible, probable, and proven IFD. However, the fungal species responsible for IFD are highly diverse and present numerous diagnostic challenges, which culminates in the empirical anti-fungal treatment of patients at risk of IFD. Recent studies have focussed on host-derived biomarkers that may mediate IFD risk and can be used to predict, and even identify IFD. The identification of novel host genetic variants, host gene expression changes, and host protein expression (cytokines and chemokines) associated with increased risk of IFD has enhanced our understanding of why only some patients at risk of IFD actually develop disease. Furthermore, these host biomarkers when incorporated into predictive models alongside conventional diagnostic techniques enhance predictive and diagnostic results. Once validated in larger studies, host biomarkers associated with IFD may optimize the clinical management of populations at risk of IFD. This review will summarise the latest developments in the identification of host biomarkers for IFD, their use in predictive modelling and their potential application/usefulness for informing clinical decisions.

Azole resistance in Aspergillus fumigatus is a One Health resistance threat, where azole fungicide exposure compromises the efficacy of medical azoles. The use of the recently authorized fungicide ipflufenoquin, which shares its mode-of-action with a new antifungal olorofim, underscores the need for risk assessment for dual use of antifungals., Competing Interests: Declaration of Competing Interest Paul E Verweij declares grants from F2G and Gilead Sciences and payments for lectures and advisory boards by F2G, Gilead Sciences, Pfizer and Mundipharma. All payments to institute. Maiken C Arendrup, Ana Alastruey-Izquierdo, Jeremy A W Gold, Shawn R Lockhart, and Tom Chiller declare no competing interests. P Lewis White declares grants from F2G, CERBA, IHMA, payments for lectures and advisory boards by F2G, Gilead Sciences, and Pfizer, and support for travel from OLM., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The COVID-19 pandemic has placed a huge strain on global healthcare and been a significant cause of increased morbidity and mortality, particularly in at-risk populations. This disease attacks the respiratory systems and causes significant immune dysregulation in affected patients creating a perfect opportunity for the development of invasive fungal disease (IFD). COVID-19 infection can instill a significant, poorly regulated pro-inflammatory response. Clinically induced immunosuppression or pro-inflammatory damage to mucosa facilitate the development of IFD and Aspergillus , Mucorales, and Candida infections have been regularly reported throughout the COVID-19 pandemic. Corticosteroids and immune modulators are used in the treatment of COVID-19. Corticosteroid use is also a risk factor for IFD, but not the only reason for IFD in COVID -19 patients. Specific dysregulation of the immune system through functional exhaustion of Natural killer (NK) cells and T cells has been observed in COVID-19 through the expression of the exhaustion markers NK-G2A and PD-1. Reduced fungicidal activity of neutrophils from COVID-19 patients indicates that immune dysfunction/imbalance are important risk factors for IFD. The COVID-19 pandemic has significantly increased the at-risk population for IFD. Even if the incidence of IFD is relatively low, the size of this new at-risk population will result in a substantial increase in the overall, annual number of IFD cases. It is important to understand how and why certain patients with COVID-19 developed increased susceptibility to IFD, as this will improve our understanding of risk of IFD in the face of future pandemics but also in a clinical era of increased clinical immuno-suppression/modulation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The handling editor SW declared a past collaboration with the authors CM, JL, LW., (Copyright © 2022 Morton, Griffiths, Loeffler, Orr and White.)

Purpose: Common CT abnormalities of pulmonary aspergillosis represent a cavity with air-meniscus sign, nodule, mass, and consolidation having an angio-invasive pattern. This study aims to conduct a systematic review and an individual patient-level image analysis of CT findings of COVID-19-associated pulmonary aspergillosis (CAPA)., Methods: A systematic literature search was conducted to identify studies reporting CT findings of CAPA as of January 7, 2021. We summarized study-level clinical and CT findings of CAPA and collected individual patient CT images by inviting corresponding authors. The CT findings were categorized into four groups: group 1, typical appearance of COVID-19; group 2, indeterminate appearance of COVID-19; group 3, atypical for COVID-19 without cavities; and group 4, atypical for COVID-19 with cavities. In group 2, cases had only minor discrepant findings including solid nodules, isolated airspace consolidation with negligible ground-glass opacities, centrilobular micronodules, bronchial abnormalities, and cavities., Results: The literature search identified 89 patients from 25 studies, and we collected CT images from 35 CAPA patients (mean age 62.4 ± 14.6 years; 21 men): group 1, thirteen patients (37.1%); group 2, eight patients (22.9%); group 3, six patients (17.1%); and group 4, eight patients (22.9%). Eight of the 14 patients (57.1%) with an atypical appearance had bronchial abnormalities, whereas only one (7.1%) had an angio-invasive fungal pattern. In the study-level analysis, cavities were reported in 12 of 54 patients (22.2%)., Conclusion: CAPA can frequently manifest as COVID-19 pneumonia without common CT abnormalities of pulmonary aspergillosis. If abnormalities exist on CT images, CAPA may frequently accompany bronchial abnormalities., Competing Interests: Declaration of competing interest This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors. Soon Ho Yoon works in the MEDICALIP as a chief medical officer. Philipp Koehler is supported by the German Federal Ministry of Research and Education and the State of North Rhine-Westphalia, Germany and has received non-financial scientific grants from Miltenyi Biotec GmbH, Bergisch Gladbach, Germany, and the Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases, University of Cologne, Cologne, Germany, and received lecture honoraria from and/or is advisor to Akademie für Infektionsmedizin e.V., Ambu GmbH, Astellas Pharma, European Confederation of Medical Mycology, Gilead Sciences, GPR Academy Ruesselsheim, MSD Sharp & Dohme GmbH, Noxxon N.V., and University Hospital, LMU Munich outside the submitted work. Other authors have no conflicts of interest to declare for this article., (Copyright © 2022. Published by Elsevier Inc.)

Aspergillus fumigatus is a ubiquitous mold that can cause severe infections in immunocompromised patients, typically manifesting as invasive pulmonary aspergillosis (IPA). Adaptive and innate immune cells that respond to A. fumigatus are present in the endogenous repertoire of patients with IPA but are infrequent and cannot be consistently isolated and expanded for adoptive immunotherapy. Therefore, we gene-engineered A. fumigatus -specific chimeric antigen receptor (Af-CAR) T cells and demonstrate their ability to confer antifungal reactivity in preclinical models in vitro and in vivo. We generated a CAR targeting domain AB90-E8 that recognizes a conserved protein antigen in the cell wall of A. fumigatus hyphae. T cells expressing the Af-CAR recognized A. fumigatus strains and clinical isolates and exerted a direct antifungal effect against A. fumigatus hyphae. In particular, CD8 + Af-CAR T cells released perforin and granzyme B and damaged A. fumigatus hyphae. CD8 + and CD4 + Af-CAR T cells produced cytokines that activated macrophages to potentiate the antifungal effect. In an in vivo model of IPA in immunodeficient mice, CD8 + Af-CAR T cells localized to the site of infection, engaged innate immune cells, and reduced fungal burden in the lung. Adoptive transfer of CD8 + Af-CAR T cells conferred greater antifungal efficacy compared to CD4 + Af-CAR T cells and an improvement in overall survival. Together, our study illustrates the potential of gene-engineered T cells to treat aggressive infectious diseases that are difficult to control with conventional antimicrobial therapy and support the clinical development of Af-CAR T cell therapy to treat IPA.

Background: Treatment for invasive candidiasis (IC) is time-critical, and culture-based tests can limit clinical utility. Nonculture-based methods such as Candida PCR represent a promising approach to improving patient management but require further evaluation to understand their optimal role and incorporation into clinical algorithms. This study determined the performance of the commercially available OLM CandID real-time PCR when testing serum and developed a diagnostic algorithm for IC. Methods: The study comprised a retrospective performance evaluation of the CandID real-time PCR assay when testing surplus serum (n = 83 patients, 38 with IC), followed by a prospective consecutive cohort evaluation (n = 103 patients, 24 with IC) post incorporation into routine service. A combined diagnostic algorithm, also including (1-3)-β-D-Glucan testing, was generated. Results: Prospective CandID testing generated a sensitivity/specificity of 88%/82%, respectively. Specificity was improved (>95%) when both PCR replicates were positive and/or the patient had multiple positive samples. When combining CandID with (1-3)-β-D-Glucan testing, the probability of IC when both were positive or negative was >69% or <1%, respectively. Conclusions: The CandID provides excellent performance and a rapid time-to-result using methods widely available in generic molecular diagnostic laboratories. By combining nonculture diagnostics, it may be possible to accurately confirm or exclude IC.

Invasive fungal infections pose an important threat to public health and are an under-recognized component of antimicrobial resistance, an emerging crisis worldwide. Across a period of profound global environmental change and expanding at-risk populations, human-infecting pathogenic fungi are evolving resistance to all licensed systemic antifungal drugs. In this Review, we highlight the main mechanisms of antifungal resistance and explore the similarities and differences between bacterial and fungal resistance to antimicrobial control. We discuss the research and innovation topics that are needed for risk reduction strategies aimed at minimizing the emergence of resistance in pathogenic fungi. These topics include links between the environment and One Health, surveillance, diagnostics, routes of transmission, novel therapeutics and methods to mitigate hotspots for fungal adaptation. We emphasize the global efforts required to steward our existing antifungal armamentarium, and to direct the research and development of future therapies and interventions., (© 2022. Springer Nature Limited.)

Patients suffering from coronavirus disease-2019 (COVID-19) are susceptible to deadly secondary fungal infections such as COVID-19-associated pulmonary aspergillosis and COVID-19-associated mucormycosis. Despite this clinical observation, direct experimental evidence for severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2)-driven alterations of antifungal immunity is scarce. Using an ex-vivo whole blood stimulation assay, we challenged blood from twelve COVID-19 patients with Aspergillus fumigatus and Rhizopus arrhizus antigens and studied the expression of activation, maturation, and exhaustion markers, as well as cytokine secretion. Compared to healthy controls, T-helper cells from COVID-19 patients displayed increased expression levels of the exhaustion marker PD-1 and weakened A. fumigatus - and R. arrhizus -induced activation. While baseline secretion of proinflammatory cytokines was massively elevated, whole blood from COVID-19 patients elicited diminished release of T-cellular (e.g., IFN-γ, IL-2) and innate immune cell-derived (e.g., CXCL9, CXCL10) cytokines in response to A. fumigatus and R. arrhizus antigens. Additionally, samples from COVID-19 patients showed deficient granulocyte activation by mold antigens and reduced fungal killing capacity of neutrophils. These features of weakened anti-mold immune responses were largely decoupled from COVID-19 severity, the time elapsed since diagnosis of COVID-19, and recent corticosteroid uptake, suggesting that impaired anti-mold defense is a common denominator of the underlying SARS-CoV-2 infection. Taken together, these results expand our understanding of the immune predisposition to post-viral mold infections and could inform future studies of immunotherapeutic strategies to prevent and treat fungal superinfections in COVID-19 patients., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tappe, Lauruschkat, Strobel, Pantaleón García, Kurzai, Rebhan, Kraus, Pfeuffer-Jovic, Bussemer, Possler, Held, Hünniger, Kniemeyer, Schäuble, Brakhage, Panagiotou, White, Einsele, Löffler and Wurster.)

The increasing incidence and changing epidemiology of invasive fungal infections continue to present many challenges to their effective management. The repertoire of antifungal drugs available for treatment is still limited although there are new antifungals on the horizon. Successful treatment of invasive mycoses is dependent on a mix of pathogen-, host- and antifungal drug-related factors. Laboratories need to be adept at detection of fungal pathogens in clinical samples in order to effectively guide treatment by identifying isolates with acquired drug resistance. While there are international guidelines on how to conduct in vitro antifungal susceptibility testing, these are not performed as widely as for bacterial pathogens. Furthermore, fungi generally are recovered in cultures more slowly than bacteria, and often cannot be cultured in the laboratory. Therefore, non-culture-based methods, including molecular tests, to detect fungi in clinical specimens are increasingly important in patient management and are becoming more reliable as technology improves. Molecular methods can also be used for detection of target gene mutations or other mechanisms that predict antifungal drug resistance. This review addresses acquired antifungal drug resistance in the principal human fungal pathogens and describes known resistance mechanisms and what in-house and commercial tools are available for their detection. It is emphasized that this approach should be complementary to culture-based susceptibility testing, given the range of mutations, resistance mechanisms and target genes that may be present in clinical isolates, but may not be included in current molecular assays., (© The Author(s) 2022. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy.)

Reports of COVID-19-associated mucormycosis have been increasing in frequency since early 2021, particularly among patients with uncontrolled diabetes. Patients with diabetes and hyperglycaemia often have an inflammatory state that could be potentiated by the activation of antiviral immunity to SARS-CoV2, which might favour secondary infections. In this Review, we analysed 80 published and unpublished cases of COVID-19-associated mucormycosis. Uncontrolled diabetes, as well as systemic corticosteroid treatment, were present in most patients with COVID-19-associated mucormycosis, and rhino-orbital cerebral mucormycosis was the most frequent disease. Mortality was high at 49%, which was particularly due to patients with pulmonary or disseminated mucormycosis or cerebral involvement. Furthermore, a substantial proportion of patients who survived had life-changing morbidities (eg, loss of vision in 46% of survivors). Our Review indicates that COVID-19-associated mucormycosis is associated with high morbidity and mortality. Diagnosis of pulmonary mucormycosis is particularly challenging, and might be frequently missed in India., Competing Interests: MH received research funding from Gilead, Pfizer, Astellas, Scynexis, and US National Institutes of Health. J-PG received speaker and expert advice fees from Pfizer and Gilead. NK has received research grants or honoraria as a speaker or advisor from Astellas, Gilead, MSD, and Pfizer, outside the submitted work. KL received consultancy fees from SMB Laboratories Brussels, MSD, and Gilead; travel support from Pfizer; speaker fees from FUJIFILM Wako, Pfizer, and Gilead; and a service fee from Thermo Fisher Scientific. OAC reports grants or contracts from Amplyx, Basilea, BMBF, Cidara, DZIF, EU-DG RTD (101037867), F2G, Gilead, Matinas, MedPace, MSD, Mundipharma, Octapharma, Pfizer, and Scynexis; Consulting fees from Amplyx, Biocon, Biosys, Cidara, Da Volterra, Gilead, Matinas, MedPace, Menarini, Molecular Partners, MSG-ERC, Noxxon, Octapharma, PSI, Scynexis, and Seres; honoraria for lectures from Abbott, Al-Jazeera Pharmaceuticals, Astellas, Grupo Biotoscana/United Medical/Knight, Hikma, MedScape, MedUpdate, Merck/MSD, Mylan, and Pfizer; payment for expert testimony from Cidara; participation on a data safety monitoring board or advisory board from Actelion, Allecra, Cidara, Entasis, IQVIA, Jannsen, MedPace, Paratek, PSI, and Shionogi; a patent at the German Patent and Trade Mark Office (DE 10 2021 113 0077); and other interests from DGHO, DGI, ECMM, ISHAM, MSG-ERC, and Wiley. JRP received research support from Amplyx, Appili, and Astellas; and honoraria and consulting fees from Pfizer, Appili, and Matinas. PLW has done diagnostic evaluations and received meeting sponsorship from Bruker, Dynamiker, and Launch Diagnostics; speakers fees, expert advice fees, and meeting sponsorship from Gilead; speaker and expert advice fees from F2G; speaker fees MSD and Pfizer; and is a founding member of the European Aspergillus PCR Initiative. ACh received funding support from an educational grant from Pfizer, MSD, and Gilead., (© 2022 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license.)

Exposure to fungi is inevitable, yet only a small number of patients with significant clinical risk develop invasive aspergillosis (IA). While timing of exposure in relation to immune status, environmental and occupational factors will influence the probability of developing IA, factors specific to the individual will likely play a role and variation in the host's genetic code associated with the immunological response to fungi have been linked to increased risk of developing IA. Screening for SNPs in genes significantly associated with IA (e.g. Pentraxin-3, Toll-like receptor 4, Dectin-1, DC-SIGN) could form part of the clinical work-up on admission or post allogeneic stem cell transplantation, to complement fungal biomarker screening. Through the combination of clinical and genetic risk with mycological evidence, we are approaching a time when we should be able to accurately predict the risk of IA in the haematology patient, using predictive modelling to stratifying each individual's management. Understanding the host and their immune responses to infection through genomics, transcriptomics and metabolomics/proteomics is critical to achieving how we manage the individual's risk of IA, underpinning personalized medicine. This review will investigate what is known about the genetic risk associated with developing IA, primarily in haematology patients and whether these strategies are ready to be incorporated into routine clinical practice, and if not what are the remaining hurdles to implementation., Competing Interests: PW: performed diagnostic evaluations and received meeting sponsorship from Bruker, Dynamiker, and Launch Diagnostics; received speaker fees, expert advice fees, and meeting sponsorship from Gilead; received speaker and expert advice fees from F2G and speaker fees MSD and Pfizer; and is a founding member of the European Aspergillus PCR Initiative. The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 White and Price.)

Exophiala dermatitidis is increasingly isolated from cystic fibrosis (CF) respiratory samples. The decision to treat is hampered by limited evidence demonstrating the clinical significance of isolating E. dermatitidis. The objective was to assess the impact of E. dermatitidis isolation on the lung function of CF patients. The rate of lung function decline in the local CF population was calculated using historic lung function data. A control population who had never had E. dermatitidis cultured from the respiratory tract was compared with the E. dermatitidis group, calculating their rate of lung function decline before and after the first isolation of the organism. A total of 1840 lung function measurements were reviewed between the 31 E. dermatitidis group patients and 62 control patients. Their demographics were similar. The control group declined at a rate of −0.824 FEV1%/year. The rate of decline in the E. dermatitidis group prior to infection was −0.337 FEV1%/year (p = 0.2). However, post infection with E. dermatitidis, there was a significant increase in the rate of decline in lung function (−1.824 FEV1%/year, p < 0.01). The results suggest E. dermatitidis has a temporal relationship with accelerated rate of lung function decline. It is not clear if this is a cause or effect, but this accelerated rate of decline indicates a need for further investigation.

Despite available diagnostic tests and recent advances, diagnosis of pulmonary invasive aspergillosis (IPA) remains challenging. We performed a longitudinal case-control pilot study to identify host-specific, novel, and immune-relevant molecular candidates indicating IPA in patients post allogeneic stem cell transplantation (alloSCT). Supported by differential gene expression analysis of six relevant in vitro studies, we conducted RNA sequencing of three alloSCT patients categorized as probable IPA cases and their matched controls without Aspergillus infection (66 samples in total). We additionally performed immunoassay analysis for all patient samples to gain a multi-omics perspective. Profiling analysis suggested LGALS2 , MMP1 , IL-8, and caspase-3 as potential host molecular candidates indicating IPA in investigated alloSCT patients. MMP1, IL-8, and caspase-3 were evaluated further in alloSCT patients for their potential to differentiate possible IPA cases and patients suffering from COVID-19-associated pulmonary aspergillosis (CAPA) and appropriate control patients. Possible IPA cases showed differences in IL-8 and caspase-3 serum levels compared with matched controls. Furthermore, we observed significant differences in IL-8 and caspase-3 levels among CAPA patients compared with control patients. With our conceptual work, we demonstrate the potential value of considering the human immune response during Aspergillus infection to identify immune-relevant molecular candidates indicating IPA in alloSCT patients. These human host candidates together with already established fungal biomarkers might improve the accuracy of IPA diagnostic tools.

Introduction: Fungal PCR has undergone considerable standardization and, together with the availability of commercial assays, external quality assessment schemes, and extensive performance validation data, is ready for widespread use for the screening and diagnosis of invasive fungal disease (IFD)., Areas Covered: Drawing on the experience and knowledge of the leads of the various working parties of the Fungal PCR initiative, this review will address general considerations concerning the use of molecular tests for the diagnosis of IFD, before focusing specifically on the technical and clinical aspects of molecular testing for the main causes of IFD and recent technological developments., Expert Opinion: For infections caused by Aspergillus, Candida , and Pneumocystis jirovecii , PCR testing is recommended, and combination with serological testing will likely enhance the diagnosis. For other IFD (e.g. mucormycosis), molecular diagnostics represent the only non-classical mycological approach toward diagnoses, and continued performance validation and standardization have improved confidence in such testing. The emergence of antifungal resistance can be diagnosed, in part, through molecular testing. Next-generation sequencing has the potential to significantly improve our understanding of fungal phylogeny, epidemiology, pathogenesis, mycobiome/microbiome, and interactions with the host, while identifying novel and existing mechanisms of antifungal resistance and novel diagnostic/therapeutic targets.

This multicenter study evaluated the IMMY Aspergillus Galactomannan Lateral Flow Assay (LFA) with automated reader for diagnosis of pulmonary aspergillosis in patients with COVID-19-associated acute respiratory failure (ARF) requiring intensive care unit (ICU) admission between 03/2020 and 04/2021. A total of 196 respiratory samples and 148 serum samples ( n  = 344) from 238 patients were retrospectively included, with a maximum of one of each sample type per patient. Cases were retrospectively classified for COVID-19-associated pulmonary aspergillosis (CAPA) status following the 2020 consensus criteria, with the exclusion of LFA results as a mycological criterion. At the 1.0 cutoff, sensitivity of LFA for CAPA (proven/probable/possible) was 52%, 80% and 81%, and specificity was 98%, 88% and 67%, for bronchoalveolar lavage fluid (BALF), nondirected bronchoalveolar lavage (NBL), and tracheal aspiration (TA), respectively. At the 0.5 manufacturer's cutoff, sensitivity was 72%, 90% and 100%, and specificity was 79%, 83% and 44%, for BALF, NBL and TA, respectively. When combining all respiratory samples, the receiver operating characteristic (ROC) area under the curve (AUC) was 0.823, versus 0.754, 0.890 and 0.814 for BALF, NBL and TA, respectively. Sensitivity and specificity of serum LFA were 20% and 93%, respectively, at the 0.5 ODI cutoff. Overall, the Aspergillus Galactomannan LFA showed good performances for CAPA diagnosis, when used from respiratory samples at the 1.0 cutoff, while sensitivity from serum was limited, linked to weak invasiveness during CAPA. As some false-positive results can occur, isolated results slightly above the recommended cutoff should lead to further mycological investigations.

Invasive aspergillosis (IA) is an acute infection affecting patients who are immunocompromised, as a result of receiving chemotherapy for malignancy, or immunosuppressant agents for transplantation or autoimmune disease. Whilst criteria exist to define the probability of infection for clinical trials, there is little evidence in the literature or clinical guidelines on when to change antifungal treatment in patients who are receiving prophylaxis or treatment for IA. To try and address this significant gap, an advisory board of experts was convened to develop criteria for the management of IA for use in designing clinical trials, which could also be used in clinical practice. For primary treatment failure, a change in antifungal therapy should be made: (i) when mycological susceptibility testing identifies an organism from a confirmed site of infection, which is resistant to the antifungal given for primary therapy, or a resistance mutation is identified by molecular testing; (ii) at, or after, 8 days of primary antifungal treatment if there is increasing serum galactomannan, or galactomannan positivity in serum, or bronchoalveolar lavage fluid when the antigen was previously undetectable, or there is sudden clinical deterioration, or a new clearly distinct site of infection is detected; and (iii) at, or after, 15 days of primary antifungal treatment if the patient is clinically stable but with ≥2 serum galactomannan measurements persistently elevated compared with baseline or increasing, or if the original lesions on CT or other imaging, show progression by >25% in size in the context of no apparent change in immune status., (© The Author(s) 2021. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.)