High Risk For Invasive Aspergillosis Research Articles

Identification and validation of diagnostic cut-offs of the ELISpot assay for the diagnosis of invasive aspergillosis in high-risk patients.

We investigated the performance of enzyme linked immunospot (ELISpot) assay for the diagnosis of invasive aspergillosis (IA) in high-risk patients with hematologic malignancies. We prospectively enrolled two cohorts of patients undergoing intensive myelosuppressive or immunosuppressive treatments at high risk for IA. ELISpot was performed to detect Aspergillus-specific T cells producing Interleukin-10. In the discovery cohort, a derived cut-off of 40 spot forming cells (SFCs)/106 PBMCs has shown to correctly classify IA cases with a sensitivity and specificity of 89.5% and 88.6%, respectively. This cut-off is lowered to 25 SFC when considering the subset of possible IA patients, with sensitivity and specificity of 76% and 93%, respectively. The application of the 40 SFCs cut-off to the validation cohort resulted in a positivity rate of 83.3% in proven/probable cases and a negativity rate of 92.5% in possible/non-IA cases. Adopting the 25 SCFs cut-off, the assay resulted positive in 83.3% of proven/probable cases while it resulted negative in 66.7% of possible/non-IA cases. ELISpot shows promises in the diagnosis of IA and the possibility to use two distinct cut-offs with similar diagnostic performances according to patients' different pre-test probability of infection can widen its use in patients at risk.

Is there still a place for serum galactomannan in the diagnosis of invasive aspergillosis in children at high risk and under antifungal prophylaxis?

The performance of serum galactomannan (GM) for the diagnosis of invasive aspergillosis (IA) has been studied mainly in adults. Paediatric data are scarce and based on small and heterogeneous cohorts. To evaluate the performance of serum GM for the diagnosis of IA in a paediatric oncologic population at high risk of IA and to clarify the impact of antifungal prophylaxis on this test. We performed a retrospective study from January 2014 to December 2020 in the paediatric oncologic haematologic department of the University Hospital of Bordeaux. The diagnosis of IA was made using the recommendations of the EORTC and the MSGERC. Among the 329 periods at high risk of IA in 222 patients, the prevalence of IA was 1.8% (3 proven and 3 probable IA). In the total population, the sensitivity, and the positive predictive value (PPV) were respectively 50% and 17.6%. Under antifungal prophylaxis, the sensitivity and PPV dropped, respectively, to 33.3% and 14.3%. In this group, the post-test probability of IA was 2% for a negative serum GM and only 14%. In this large cohort of children at high risk of IA, the incidence of IA is low and the diagnostic performance of GM is poor, especially in the case of mould-active prophylaxis. Screening should be targeted rather than systematic and should be reserved for patients at highest risk for IA without mould-active prophylaxis. Combination with other tests such as Aspergillus PCR would increase the accuracy of GM in screening setting.

Identification and validation of diagnostic cut-offs of the ELISpot assay for the diagnosis of invasive aspergillosis in high-risk patients.

We investigated the performance of enzyme linked immunospot (ELISpot) assay for the diagnosis of invasive aspergillosis (IA) in high-risk patients with hematologic malignancies. We prospectively enrolled two cohorts of patients undergoing intensive myelosuppressive or immunosuppressive treatments at high risk for IA. ELISpot was performed to detect Aspergillus-specific T cells producing Interleukin-10. In the discovery cohort, a derived cut-off of 40 spot forming cells (SFCs)/106 PBMCs has shown to correctly classify IA cases with a sensitivity and specificity of 89.5% and 88.6%, respectively. This cut-off is lowered to 25 SFC when considering the subset of possible IA patients, with sensitivity and specificity of 76% and 93%, respectively. The application of the 40 SFCs cut-off to the validation cohort resulted in a positivity rate of 83.3% in proven/probable cases and a negativity rate of 92.5% in possible/non-IA cases. Adopting the 25 SCFs cut-off, the assay resulted positive in 83.3% of proven/probable cases while it resulted negative in 66.7% of possible/non-IA cases. ELISpot shows promises in the diagnosis of IA and the possibility to use two distinct cut-offs with similar diagnostic performances according to patients' different pre-test probability of infection can widen its use in patients at risk.

82. Incidence rates of invasive aspergillosis among lung transplant recipients in timeperiods with universal and targeted antifungal prophylaxis - a nationwide cohort study

Abstract Background The best stategy for prevention of invasive aspergillosis (IA) after lung transplantation (LTX) has not been determined. In 2016, Danish guidelines for IA prophylaxis was changed from 12 weeks of universal prophylaxis with voriconazole to targeted prophylaxis with posaconazole and inhaled amphotericin B for high risk patients. In a previous study we found significantly higher rates of adverse events in the period with universal prophylaxis. The objectives of this study were to compare rates of IA 1) in time periods before and after change of guidelines; 2) during persontime on vs. persontime without antifungal prophylaxis in the total study period. Methods In a Danish nationwide study we included all adult lung transplant recipients (LTXr), 2010–2019. Proven and probable IA were defined using ISHLT criteria. Patients were censored at the first of following events: death, retransplantation, end of 2019, or one year after LTX. The cumulative hazard of IA was calculated using the Nelson Aalen estimator. Rates of IA were analyzed in bi- and multivariate Poisson regression models adjusted for the time period after LTX (0–6 and 6–12 months). Persontime on protocol prophylaxis (start to end + 14 days) was treated as time-updated variable. Results A total of 295 LTXr were followed for 245 person-years, of whom 183/193 vs. 6/102 initiated antifungal prophylaxis during the universal vs. the targeted prophylaxis period (Table 1). IA was diagnosed in 34/193 (17.6%) vs. 15/102 (14.7%) LTXr the first year after transplantation in the universal vs. targeted prophylaxis period. Overall, similar cumulative hazards of IA were observed between the universal and targeted prophylaxis period (p=0.59, Figure 1). The adjusted incidence rate ratio (aIRR) of IA during the universal vs. targeted prophylaxis period was 1.30 (95% CI 0.69–2.44), (Table 2). In the total study period, the aIRR of IA during persontime on vs. persontime without antifungal prophylaxis was 0.87 (95% CI 0.40–1.88), (Table 2). Characteristics of lung transplant recipients in time periods with universal voriconazole and targeted posaconazole and inhaled amphotericin B prophylaxis. Proven and probable invasive aspergillosis including anastomosis infection, trakeobronkitis and pneumonia according to ISHLT criteria. *) Underlying diseases categorized as high risk of invasive aspergillosis. Incidence rates (IR) and incidence rate ratios (IRR) of invasive aspergillosis in lung transplant recipients in two multivariate models. Poisson regression models comparing rates of IA in model 1) during universal prophylaxis time period versus targeted prophylaxis time period and in model 2) during persontime on prophylaxis (start to end + 14 days) versus persontime without prophylaxis in the total study period 2010–2019. Adjusted for time after LTX IRR: Bivariable model including the listed variables in bold and adjustment for time after transplantation (time-updated). Fully adjusted IRR: Multivariable model with adjustment for sex, age, high risk underlying disease, single/double lung transplant, Aspergillus prior to transplantation and time after transplantation (time-updated). Abbreviations: LTX = lung transplant recipients; CI = confidence interval; IR = incidence rates per 100 person-years; IRR = incidence rate ratio; Ref. = reference. Cumulative hazards of invasive aspergillosis the first year after lung transplantation in time periods with universal voriconazole versus targeted posaconazole and inhaled amphotericin B prophylaxis. Conclusion A strategy of targeted prophylaxis, for high risk patients only, may be preferred over universal voriconazole prophylaxis after LTX due to similar rates of IA and lower rates of adverse events. Disclosures Maiken C. Arendrup, DMSci, PhD, MD, Chiesi, Gilead: Honoraria|F2G, Cidara, Scynexis: Grant/Research Support Michael Perch, MD, Boeringer-Ingelheim: Travel grant|PulmonX: Expert Testimony|Roche: Grant/Research Support|Takeda: Expert Testimony|Therakos: Honoraria|Zambon: Advisor/Consultant|Zambon: Expert Testimony Marie Helleberg, PhD, DMSc, AstraZeneca: Advisor/Consultant|Gilead: Advisor/Consultant|Gilead: Honoraria|GSK: Advisor/Consultant|GSK: Honoraria|Janssen: Advisor/Consultant|Roche: Advisor/Consultant|Sobi: Advisor/Consultant.

2095. Associations between Invasive Aspergillosis and Cytomegalovirus Infections in Lung Transplant Recipients

Abstract Background Cytomegalovirus infection (CMV) and invasive aspergillosis (IA) are important causes of morbidity and mortality among lung transplant recipients (LTXr). These opportunistic infections share risk factors, but their interrelationship need further evaluation. Early diagnosis and treatment may improve outcomes. We examined incidence rates of CMV after IA and vice versa to assess whether screening for one of these infections may be indicated when the other is diagnosed. Methods All adults receiving a lung transplant in Denmark from 2010 to 2019 were included and followed for 2 years after lung transplantation. IA was defined using the ISHLT criteria. Standardized screening for CMV and IA was performed during the study period (Figure 1). Incidence rates (IR) and incidence rate ratios (IRR) of CMV and IA was estimated by multivariate Poisson regression adjusted for time after transplantation (time updated), sex, age and high-risk CMV serostatus/IA high-risk condition. In analyses of CMV, we included IA as a time updated variable, and in analyses of IA, we included CMV as a time updated variable. Figure 1Screening protocol for cytomegalovirus (CMV) and invasive aspergillosis following lung transplantation Valganciclovir prophylaxis was administered to patients with CMV serostatus D+/R−, D+/R+, and D−/R+ for three months after transplantation. In 2010-2016 voriconazole was administered as antifungal prophylaxis for all LTXr three months after transplantation. In 2016-2019 posaconazole and inhaled amphotericin B was administered for three months after transplantation for patients at high risk of IA. D, Donor; R, Recipient; PCR, Polymerase Chain Reaction; CMV, cytomegalovirus; BAL, bronchoalveolar lavage; Lung biopsy, transbronchial biopsy Results A total of 295 LTXr with 351 person-years of follow-up (PYFU) for CMV and 440 PYFU for IA were included (Table 1). CMV and IA were diagnosed in 122 (41.4%) and 57 (19.3%) LTXr, respectively (Figure 2). IA diagnosis was proven in 20 and probable in 37 patients (Table 1). Among LTXr diagnosed with IA, 15.8% developed CMV within 3 months, IR 109 per 100 PYFU (95% CI 57-210) (Table 2). The median viral load at CMV diagnosis after IA was 2900 copies/mL (IQR 525-19000). The first 3 months following IA diagnosis there was an increased risk of CMV, although not statistically significant after adjustment for time after LTX with aIRR 1.60 (95% CI 0.80-3.20). Among LTXr diagnosed with CMV, 8.2% developed IA within 3 months, IR 40 per 100 PYFU (95% CI 22-75) (Table 2). In the first 3 months following CMV diagnosis, there was a significantly increased risk of IA, aIRR 2.97 (95% CI 1.47-6.00). Numbers needed to screen to diagnose one case of CMV following IA, and one case of IA following CMV were approximately 6 and 12, respectively. Table 1:Characteristics of lung transplant recipient and infectious outcomes, n (%)*IA high-risk conditions; **Number of patients negative in blood samples but positive in BAL samples (minimum >3000 CMV copies/mL). Invasive aspergillosis defined by the ISHLT criteria including anastomosis infection, tracheobronchitis and pneumonia.TX, Transplantation; COPD, Chronic obstructive pulmonary disease; CMV, cytomegalovirus; D, Donor; R, Recipient; BAL, Bronchoalveolar lavage.Table 2:Incidence rates and incidence rate ratios of cytomegalovirus (CMV) and invasive aspergillosis (IA)1Adjusted for age >50 years, sex, and high-risk CMV serostatus. 2Adjusted for time after transplantation, age >50 years, sex, and high-risk CMV serostatus. 3Adjusted for age >50 years, sex, and IA high-risk underlying condition. 4Adjusted for time after transplantation, age >50 years, sex, and IA high-risk underlying condition.CMV, cytomegalovirus; IR, incidence rate; PYFU, Person-years of follow-up; IRR, incidence rate ratios; aIRR, adjusted incidence rate ratio; CI, confidence interval; TX, transplantation; IA, invasive aspergillosis.Figure 2:Kaplan-Meier survival estimates for CMV infection (A) and invasive aspergillosis (B) after lung transplantation Conclusion Systematic screening for CMV following diagnosis of IA, and vice versa, may improve the timeliness of treatment and thereby outcomes for LTX recipients. Disclosures Maiken C. Arendrup, DMSci, PhD, MD, Chiesi, Gilead: Honoraria|F2G, Cidara, Scynexis: Grant/Research Support Marie Helleberg, PhD, DMSc, AstraZeneca: Advisor/Consultant|Gilead: Advisor/Consultant|Gilead: Honoraria|GSK: Advisor/Consultant|GSK: Honoraria|Janssen: Advisor/Consultant|Roche: Advisor/Consultant|Sobi: Advisor/Consultant.

Influenza vaccination is associated with a reduced risk of invasive aspergillosis in high-risk individuals in Taiwan: a population-based cohort study

ABSTRACT Invasive aspergillosis (IA) has become the emerging life-threatening disease in recent years. Influenza has been identified as an independent risk factor for IA. Vaccination is the most effective way to prevent influenza, while whether it can reduce IA in high-risk population still uncertain. We aimed to investigate the association between influenza vaccination and the risk of IA in high-risk population. We performed a population-based cohort study of people who qualified for government-funded influenza vaccination and were at high risk for IA at the start of the influenza season each year between 2016 and 2019. We utilized Taiwan’s National Health Insurance Research Database to identify the influenza vaccination status and IA diagnosis during the follow-up period. We compared the risk of IA between people with and without vaccination using multivariable logistic regression analysis. Out of total 8,544,451 people who were eligible during the 3 influenza seasons, 3,136,477 (36.7%) were vaccinated. A total of 1179 IA cases with the incidence of 13.8 cases per 100,000 high-risk individuals were identified during the follow-up. Compared to non-vaccinated group, vaccinated individuals had a 21% risk reduction of IA (adjusted odds ratio 0.79, 95% confidence interval 0.70–0.90). Influenza vaccination was associated with a lower risk of IA among males, immunosuppressive conditions, malignancy, diabetes, and those having host factors according to the European Organization for Research and Treatment of Cancer and the Mycoses Study Group Education and Research Consortium. Influenza vaccination is recommended for high-risk population to reduce the risk of IA.

Targeted caspofungin prophylaxis for invasive aspergillosis in high-risk liver transplant recipients, a single-center experience.

Invasive aspergillosis (IA) is a rare but highly lethal complication after orthotopic liver transplantation (OLT). Targeted antifungal prophylaxis has been proposed as a strategy to prevent IA among orthotopic liver transplant recipient (OLTr), but limited data are available to support its efficacy. We conducted a single-center, retrospective, before and after cohort study, comparing IA incidences among OLTr who did not receive antifungal prophylaxis after transplantation (cohort 1) to OLTr who received targeted antifungal prophylaxis after liver transplantation (cohort 2). Patients in cohort 2 received caspofungin prophylaxis if they presented one of the following risk factors: retransplantation, acute liver failure, dialysis, or Aspergillus colonization prior to transplantation. The primary outcome was IA at 90days after transplantation. A total of 391 OLTr were included in the study; 181 patients in the cohort 1 (no prophylaxis) and 210 patients in the cohort 2 (targeted prophylaxis). Among patients in cohort 2, 19% (40/ 210) were considered at high risk for IA and 85% (34/40) of those received caspofungin prophylaxis. The incidence of IA at 90days was 3.3% (6/ 181) and 0.5% (1/ 210), in cohort 1 and 2, respectively (OR 0.14; 95%CI 0.01-0.83; P=.03). Ninety-day mortality was similar among the two cohorts (3.9% (7/181) and 2.4% (5/210) in cohort 1 and 2, respectively (OR 0.61; 95% 0.18-1.93; P=.40)). The 90-day mortality among the OLTs with IA was 71% (5/7). Targeted caspofungin prophylaxis was associated with lower rate of IA.

Impact of invasive aspergillosis occurring during first induction therapy on outcome of acute myeloid leukaemia (SEIFEM-12B study).

Acute myeloid leukaemia (AML) patients are at high risk of invasive aspergillosis (IA) after first induction chemotherapy (CHT). Although IA risk factors have been identified, few data are available on impact of IA, occurring during induction phase, on overall AML outcome. The end point of this multicentre, case-control, study was to evaluate whether IA, occurring after first induction CHT, can affect treatment schedule and patient's outcome. We identified 40 AML patients (cases) who developed IA during first induction phase, 31 probable (77.5%) and 9 proven (22.5%). These cases were matched with a control group (80 AML) without IA, balanced according to age, type of CHT, AML characteristics and cytogenetic-molecular risk factors. The overall response rate to induction CHT was the same in the 2 groups. In the 40 cases with IA, the overall response rate to antifungal treatment was favourable (80%) but it was significantly affected by the achievement of leukaemia complete remission (CR) with induction CHT. In fact, in cases with AML responsive to induction CHT, responses of IA to antifungal therapy were 96% compared to 21% in cases of AML not responsive to induction treatment (P<.0001). The adherence to the schedule and full doses of CHT were reported in 35% of cases (14/40) and in 76% of controls (61/80) (P=.0001; OR 6.7; 95% CI 2.7-16.6). After first induction CHT, a significant higher number of cases (15/40; 37.5%) compared to controls (9/80; 11%) could not receive additional cycles of CHT (P=.0011, OR 4.8; 95% CI 1.9-12.3). The IA-related mortality was 22.5%. The median OS of cases was significantly worse than OS of controls with a difference of 12.3months (12.1 vs 24.4months, P=.04). However, the occurrence of IA during first induction phase did not have a significant impact on the OS of cases who achieved a CR of AML with induction CHT which are able to proceed, despite the IA, with their therapeutic program, achieving the same OS as the control group with AML in CR (P=ns). These data show that IA during first induction CHT can delay the subsequent therapeutic program and has a significant impact on OS, specifically in AML patients who did not achieved a CR of AML with the first course of CHT.

Polymerase chain reaction blood tests for the diagnosis of invasive aspergillosis in immunocompromised people

This is an update of the original review published in the Cochrane Database of Systematic Reviews Issue 10, 2015.Invasive aspergillosis (IA) is the most common life-threatening opportunistic invasive mould infection in immunocompromised people. Early diagnosis of IA and prompt administration of appropriate antifungal treatment are critical to the survival of people with IA. Antifungal drugs can be given as prophylaxis or empirical therapy, instigated on the basis of a diagnostic strategy (the pre-emptive approach) or for treating established disease. Consequently, there is an urgent need for research into both new diagnostic tools and drug treatment strategies. Increasingly, newer methods such as polymerase chain reaction (PCR) to detect fungal nucleic acids are being investigated. To provide an overall summary of the diagnostic accuracy of PCR-based tests on blood specimens for the diagnosis of IA in immunocompromised people. We searched MEDLINE (1946 to June 2015) and Embase (1980 to June 2015). We also searched LILACS, DARE, Health Technology Assessment, Web of Science and Scopus to June 2015. We checked the reference lists of all the studies identified by the above methods and contacted relevant authors and researchers in the field. For this review update we updated electronic searches of the Cochrane Central Register of Controlled Trials (CENTRAL; 2018, Issue 3) in the Cochrane Library; MEDLINE via Ovid (June 2015 to March week 2 2018); and Embase via Ovid (June 2015 to 2018 week 12). We included studies that: i) compared the results of blood PCR tests with the reference standard published by the European Organisation for Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG); ii) reported data on false-positive, true-positive, false-negative and true-negative results of the diagnostic tests under investigation separately; and iii) evaluated the test(s) prospectively in cohorts of people from a relevant clinical population, defined as a group of individuals at high risk for invasive aspergillosis. Case-control and retrospective studies were excluded from the analysis. Authors independently assessed quality and extracted data. For PCR assays, we evaluated the requirement for either one or two consecutive samples to be positive for diagnostic accuracy. We investigated heterogeneity by subgroup analyses. We plotted estimates of sensitivity and specificity from each study in receiver operating characteristics (ROC) space and constructed forest plots for visual examination of variation in test accuracy. We performed meta-analyses using the bivariate model to produce summary estimates of sensitivity and specificity. We included 29 primary studies (18 from the original review and 11 from this update), corresponding to 34 data sets, published between 2000 and 2018 in the meta-analyses, with a mean prevalence of proven or probable IA of 16.3 (median prevalence 11.1% , range 2.5% to 57.1%). Most patients had received chemotherapy for haematological malignancy or had undergone hematopoietic stem cell transplantation. Several PCR techniques were used among the included studies. The sensitivity and specificity of PCR for the diagnosis of IA varied according to the interpretative criteria used to define a test as positive. The summary estimates of sensitivity and specificity were 79.2% (95% confidence interval (CI) 71.0 to 85.5) and 79.6% (95% CI 69.9 to 86.6) for a single positive test result, and 59.6% (95% CI 40.7 to 76.0) and 95.1% (95% CI 87.0 to 98.2) for two consecutive positive test results. PCR shows moderate diagnostic accuracy when used as screening tests for IA in high-risk patient groups. Importantly the sensitivity of the test confers a high negative predictive value (NPV) such that a negative test allows the diagnosis to be excluded. Consecutive positives show good specificity in diagnosis of IA and could be used to trigger radiological and other investigations or for pre-emptive therapy in the absence of specific radiological signs when the clinical suspicion of infection is high. When a single PCR positive test is used as the diagnostic criterion for IA in a population of 100 people with a disease prevalence of 16.3% (overall mean prevalence), three people with IA would be missed (sensitivity 79.2%, 20.8% false negatives), and 17 people would be unnecessarily treated or referred for further tests (specificity of 79.6%, 21.4% false positives). If we use the two positive test requirement in a population with the same disease prevalence, it would mean that nine IA people would be missed (sensitivity 59.6%, 40.4% false negatives) and four people would be unnecessarily treated or referred for further tests (specificity of 95.1%, 4.9% false positives). Like galactomannan, PCR has good NPV for excluding disease, but the low prevalence of disease limits the ability to rule in a diagnosis. As these biomarkers detect different markers of disease, combining them is likely to prove more useful.

Diagnosis of Fungal Infections. A Systematic Review and Meta-Analysis Supporting American Thoracic Society Practice Guideline

Rationale: Prompt diagnosis of invasive fungal infections is important because of the associated morbidity and mortality; however, diagnosis is challenging because of the nonspecific symptoms and radiographic findings.Objectives: To conduct a systematic review and meta-analysis of studies that evaluated the diagnostic accuracy of serum and bronchoalveolar lavage (BAL) galactomannan (GM) and serum or BAL polymerase chain reaction (PCR) in patients with suspected invasive aspergillosis (IA), β-d-glucan in critically ill patients at risk for candidiasis or candidemia, and serology testing and antigen detection in patients with endemic mycoses (histoplasmosis, blastomycosis, and coccidioidomycosis).Methods: Studies were selected and appraised by pairs of reviewers. Bivariate random effects meta-analysis was used to generate pooled sensitivity, specificity, and diagnostic likelihood ratios.Results: Serum GM in patients with impaired immunity suspected of having IA had sensitivity of 0.71 (95% confidence interval [CI], 0.64-0.78) and specificity of 0.89 (95% CI, 0.84-0.92). A cutoff of 1 optical density index yielded optimal sensitivity and specificity. BAL GM in patients with impaired immunity suspected of having IA had sensitivity of 0.84 (95% CI, 0.73-0.91) and specificity of 0.88 (95% CI, 0.81-0.91). Serum or whole-blood PCR in immunocompromised patients with suspected IA had sensitivity of 0.81 (95% CI, 0.73-0.86) and specificity of 0.79 (95% CI, 0.68-0.86). BAL PCR in patients at high risk for IA had high sensitivity of 0.90 (95% CI, 0.77-0.96) and specificity of 0.96 (95% CI, 0.93-0.98) for diagnosing IA. β-d-glucan assay in patients in the intensive care unit at risk for invasive candidiasis or candidemia had sensitivity of 0.81 (95% CI, 0.74-0.86) and specificity of 0.60 (95% CI, 0.49-0.71). Data on diagnostic accuracy of antigen detection and serology testing for endemic mycoses were limited and heterogeneous (varied according to test, patient immunity, and suspected endemic disease).Conclusions: The diagnosis of invasive fungal infections remains a challenge. Various serum and BAL markers can aid in diagnosis. This evidence supports the development of clinical practice recommendations by the American Thoracic Society.

Voriconazole Increases Risk for Skin Cancer in Lung Transplant Recipients

Because lung transplant recipients are at high risk for invasive aspergillosis (IA), many centers employ the antifungal medication voriconazole as

Evaluation of Hirst-type spore traps in outdoor Aspergillaceae monitoring during large demolition work in hospital.

Demolition can generate fungal spore suspensions in association with various adverse health effects, such as high risk of invasive aspergillosis in immunocompromised patients. One block of Edouard Herriot Hospital was entirely demolished. The aim of the present study was to evaluate Hirst-type spore traps utility in monitoring outdoor Aspergillaceae (Aspergillus spp. + Penicillium spp.) spores in part of Edouard Herriot Hospital (Lyon, France) undergoing major demolition. Three periods were scheduled in 2015: (A) Gutting of building and asbestos removal, (B) Demolition of floors, (C) Excavation and earthwork. Outdoor Aspergillaceae fungal load was monitored by cultivable (Air Ideal®, bioMérieux) and non-cultivable methods (Lanzoni VPPS-2000, Analyzair®, Bologna, Italy). Differences of Aspergillaceae recorded with Hirst-type spore traps were observed between Gerland and Edouard Herriot Hospital. Differences between Aspergillaceae were recorded between day time and night time at Gerland and Edouard Herriot Hospital. Daily paired differences between Aspergillaceae recorded with non-cultivable methodology at Edouard Herriot Hospital and in an area without demolition work were significant in Period A vs Period B (p = 10–4) and Period A vs Period C (p = 10–4). Weak correlation of daily Aspergillaceae recorded by both methods at Edouard Herriot Hospital was significant only for Period C (r = 0.26, p = 0.048, n = 58). Meteorological parameters and type of demolition works were found to heavily influenced Aspergillaceae dispersion. Non-cultivable methodology is a promising tool for outdoor Aspergillaceae scrutiny during major demolition work in hospital, helping infection control staff to rapidly implement control measures.

Is a biomarker-based diagnostic strategy for invasive aspergillosis cost effective in high-risk haematology patients?

Empirical antifungal therapy is frequently used in hematology patients at high risk of invasive aspergillosis (IA), with substantial cost and toxicity. Biomarkers for IA aim for earlier and more accurate diagnosis and targeted treatment. However, data on the cost-effectiveness of a biomarker-based diagnostic strategy (BDS) are limited. We evaluated the cost effectiveness of BDS using results from a randomized controlled trial (RCT) and individual patient costing data. Data inputs derived from a published RCT were used to construct a decision-analytic model to compare BDS (Aspergillus galactomannan and PCR on blood) with standard diagnostic strategy (SDS) of culture and histology in terms of total costs, length of stay, IA incidence, mortality, and years of life saved. Costs were estimated for each patient using hospital costing data to day 180 and follow-up for survival was modeled to five years using a Gompertz survival model. Treatment costs were determined for 137 adults undergoing allogeneic hematopoietic stem cell transplant or receiving chemotherapy for acute leukemia in four Australian centers (2005-2009). Median total costs at 180 days were similar between groups (US$78,774 for SDS [IQR US$50,808-123,476] and US$81,279 for BDS [IQR US$59,221-123,242], P = .49). All-cause mortality was 14.7% (10/68) for SDS and 10.1% (7/69) for BDS, (P = .573). The costs per life-year saved were US$325,448, US$81,966, and US$3,670 at 180 days, one year and five years, respectively. BDS is not cost-sparing but is cost-effective if a survival benefit is maintained over several years. An individualized institutional approach to diagnostic strategies may maximize utility and cost-effectiveness.

Development and evaluation of enzyme-linked immunosorbent assay of nucleic acid sequence-based amplification for diagnosis of invasive aspergillosis.

Invasive aspergillosis (IA) is a life-threatening infection in immunocompromised patients, rapid and sensitive detection of Aspergillus from clinical samples has been a major challenge in the early diagnosis of IA. An enzyme-linked immunosorbent assay of nucleic acid sequence-based amplification (NASBA-ELISA) was developed to fulfil the need for the efficient diagnosis of these infections. The primers targeting 18S rRNA were selected for the amplification of Aspergillus RNA by the isothermal digoxigenin (DIG)-labeling NASBA process. The DIG-labeled RNA amplicons were hybridized with a specific biotinylated DNA probe immobilized on streptavidin-coated microtiter plate. The hybrids were colorimetrically detected by the addition of an anti-DIG antibodies linked to ALP and substrate (disodium 4-nitrophenyl phosphate). The detection limit of the Aspergillus NABSA-ELISA system was 1 CFU and the RNA in non-target bacteria or fungus was not amplified. The performance of this NASBA-ELISA compared to RT-PCR and galactomannan (GM) was evaluated by testing blood samples from 86 patients at high risk for IA. The sensitivity of NASBA-ELISA, RT-PCR and GM-ELISA was 80.56 % (95 % CI 63.98–91.81), 72.22 % (95 % CI 54.81–85.80), 58.33 % (95 % CI 40.76–74.49), respectively, and the specificity was 80.00 % (95 % CI 66.28–89.97), 84.00 % (95 % CI 70.89–92.83), 82.00 % (95 % CI 68.56–91.42). The efficiency of the three methods in various combinations was also evaluated. Combination of NASBA-ELISA and GM-ELISA testing achieved perfect specificity (100 %; 95 % CI 92.89–100) and perfect positive predictive value (100 %; 95 % CI 83.16–100). The best sensitivity (97.22 %; 95 % CI 85.47–99.93) and the highest Youden index (0.652) were obtained by testing with both NASBA and RT-PCR in parallel. In conclusion, the NASBA-ELISA assay consists of an alternative process for large-scale samples detection with semi-quantitative results and provides good clinical performance without resorting to expensive equipment. This assay makes it possible for the NASBA based RNA diagnosis to become a routine work in laboratories in less developed countries with fewer resources.Electronic supplementary materialThe online version of this article (doi:10.1186/s13568-016-0266-0) contains supplementary material, which is available to authorized users.

Polymerase chain reaction blood tests for the diagnosis of invasive aspergillosis in immunocompromised people.

Invasive aspergillosis (IA) is the most common life-threatening opportunistic invasive mould infection in immunocompromised people. Early diagnosis of IA and prompt administration of appropriate antifungal treatment are critical to the survival of people with IA. Antifungal drugs can be given as prophylaxis or empirical therapy, instigated on the basis of a diagnostic strategy (the pre-emptive approach) or for treating established disease. Consequently there is an urgent need for research into both new diagnostic tools and drug treatment strategies. Newer methods such as polymerase chain reaction (PCR) to detect fungal nucleic acids are increasingly being investigated. To provide an overall summary of the diagnostic accuracy of PCR-based tests on blood specimens for the diagnosis of IA in immunocompromised people. We searched MEDLINE (1946 to June 2015) and EMBASE (1980 to June 2015). We also searched LILACS, DARE, Health Technology Assessment, Web of Science and Scopus to June 2015. We checked the reference lists of all the studies identified by the above methods and contacted relevant authors and researchers in the field. We included studies that: i) compared the results of blood PCR tests with the reference standard published by the European Organisation for Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG); ii) reported data on false-positive, true-positive, false-negative and true-negative results of the diagnostic tests under investigation separately; and iii) evaluated the test(s) prospectively in cohorts of people from a relevant clinical population, defined as a group of individuals at high risk for invasive aspergillosis. Case-control studies were excluded from the analysis. Authors independently assessed quality and extracted data. For PCR assays, we evaluated the requirement for either one or two consecutive samples to be positive for diagnostic accuracy. We investigated heterogeneity by subgroup analyses. We plotted estimates of sensitivity and specificity from each study in receiver operating characteristics (ROC) space and constructed forest plots for visual examination of variation in test accuracy. We performed meta-analyses using the bivariate model to produce summary estimates of sensitivity and specificity. Eighteen primary studies, corresponding to 19 cohorts and 22 data sets, published between 2000 and 2013 were included in the meta-analyses, with a median prevalence of IA (proven or probable) of 12.0% (range 2.5 to 30.8 %). The majority of people had received chemotherapy for a haematological malignancy or had undergone a hematopoietic stem cell transplant. Several PCR techniques were used among the included studies. The sensitivity and specificity of PCR for the diagnosis of IA varied according to the interpretative criteria used to define a test as positive. The mean sensitivity and specificity were 80.5% (95% CI; 73.0 to 86.3) and 78.5% (67.8 to 86.4) for a single positive test result, and 58.0% (36.5 to 76.8) and 96.2% (89.6 to 98.6) for two consecutive positive test results. PCR shows moderate diagnostic accuracy when used as screening tests for IA in high-risk patient groups. Importantly the sensitivity of the test confers a high negative predictive value (NPV) such that a negative test allows the diagnosis to be excluded. Consecutive positives show good specificity in diagnosis of IA and could be used to trigger radiological and other investigations or for pre-emptive therapy in the absence of specific radiological signs when the clinical suspicion of infection is high. When a single PCR positive test is used as diagnostic criterion for IA in a population of 100 people with a disease prevalence of 13.0% (overall mean prevalence), three people with IA would be missed (sensitivity 80.5%, 19.5% false negatives), and 19 people would be unnecessarily treated or referred for further tests (specificity of 78.5%, 21.5% false positives). If we use the two positive test requirement in a population with the same disease prevalence, it would mean that six IA people would be missed (sensitivity 58.0%, 42.1% false negatives) and three people would be unnecessarily treated or referred for further tests (specificity of 96.2%, 3.8% false positives). Galactomannan and PCR have good NPV for excluding disease but the low prevalence of disease limits the ability to rule in a diagnosis. The biomarkers are detecting different aspects of disease and the combination of both together is likely to be more useful.

Polymerase chain reaction blood tests for the diagnosis of invasive aspergillosis in immunocompromised people.

Invasive aspergillosis (IA) is the most common life-threatening opportunistic invasive mould infection in immunocompromised people. Early diagnosis of IA and prompt administration of appropriate antifungal treatment are critical to the survival of people with IA. Antifungal drugs can be given as prophylaxis or empirical therapy, instigated on the basis of a diagnostic strategy (the pre-emptive approach) or for treating established disease. Consequently there is an urgent need for research into both new diagnostic tools and drug treatment strategies. Newer methods such as polymerase chain reaction (PCR) to detect fungal nucleic acids are increasingly being investigated. To provide an overall summary of the diagnostic accuracy of PCR-based tests on blood specimens for the diagnosis of IA in immunocompromised people. We searched MEDLINE (1946 to June 2015) and EMBASE (1980 to June 2015). We also searched LILACS, DARE, Health Technology Assessment, Web of Science and Scopus to June 2015. We checked the reference lists of all the studies identified by the above methods and contacted relevant authors and researchers in the field. We included studies that: i) compared the results of blood PCR tests with the reference standard published by the European Organisation for Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG); ii) reported data on false-positive, true-positive, false-negative and true-negative results of the diagnostic tests under investigation separately; and iii) evaluated the test(s) prospectively in cohorts of people from a relevant clinical population, defined as a group of individuals at high risk for invasive aspergillosis. Case-control studies were excluded from the analysis. Authors independently assessed quality and extracted data. For PCR assays, we evaluated the requirement for either one or two consecutive samples to be positive for diagnostic accuracy. We investigated heterogeneity by subgroup analyses. We plotted estimates of sensitivity and specificity from each study in receiver operating characteristics (ROC) space and constructed forest plots for visual examination of variation in test accuracy. We performed meta-analyses using the bivariate model to produce summary estimates of sensitivity and specificity. Eighteen primary studies, corresponding to 19 cohorts and 22 data sets, published between 2000 and 2013 were included in the meta-analyses, with a median prevalence of IA (proven or probable) of 12.0% (range 2.5 to 30.8 %). The majority of people had received chemotherapy for a haematological malignancy or had undergone a hematopoietic stem cell transplant. Several PCR techniques were used among the included studies. The sensitivity and specificity of PCR for the diagnosis of IA varied according to the interpretative criteria used to define a test as positive. The mean sensitivity and specificity were 80.5% (95% CI; 73.0 to 86.3) and 78.5% (67.8 to 86.4) for a single positive test result, and 58.0% (36.5 to 76.8) and 96.2% (89.6 to 98.6) for two consecutive positive test results. PCR shows moderate diagnostic accuracy when used as screening tests for IA in high-risk patient groups. Importantly the sensitivity of the test confers a high negative predictive value (NPV) such that a negative test allows the diagnosis to be excluded. Consecutive positives show good specificity in diagnosis of IA and could be used to trigger radiological and other investigations or for pre-emptive therapy in the absence of specific radiological signs when the clinical suspicion of infection is high. When a single PCR positive test is used as diagnostic criterion for IA in a population of 100 people with a disease prevalence of 13.0% (overall mean prevalence), three people with IA would be missed (sensitivity 80.5%, 19.5% false negatives), and 19 people would be unnecessarily treated or referred for further tests (specificity of 78.5%, 21.5% false negatives). If we use the two positive test requirement in a population with the same disease prevalence, it would mean that six IA people would be missed (sensitivity 58.0%, 42.1% false negatives) and three people would be unnecessarily treated or referred for further tests (specificity of 96.2%, 3.8% false negatives). Galactamannan and PCR have good NPV for excluding disease but the low prevalence of disease limits the ability to rule in a diagnosis. The biomarkers are detecting different aspects of disease and the combination of both together is likely to be more useful.

High negative predictive value diagnostic strategies for the reevaluation of early antifungal treatment: A multicenter prospective trial in patients at risk for invasive fungal infections

Early antifungal therapeutic strategies are proposed during invasive fungal infection (IFI), but antifungal stewardship programs should institute a systematic reevaluation of prescriptions, particularly in the context of empirical treatment. Here, we aimed to evaluate the performances and particularly the negative predictive value (NPV) of diagnostic strategies, including a whole blood panfungal quantitative PCR assay (PF-qPCR) in a high risk population for IFI. The first step was to standardize and optimize a new PF-qPCR targeting ITS2 region. Then, this method was evaluated in a multicenter prospective study including 313 patients with suspected IFI for whom an early antifungal treatment was prescribed. All patients enrolled at day 0 of their treatment benefited from serum Aspergillus galactomannan (GM) antigen detection twice a week, weekly PF-qPCR assay, and when indicated and feasible, CT-scan and mycological sampling. In total, 125 of 313 patients were diagnosed with IFI: 68 invasive aspergillosis (eight proven, 48 probable and 12 possible), one fusariosis, 47 candidemia, three disseminated candidiasis and six cryptococcosis. Globally, the sensitivity of the PF-qPCR assay was only 40%, but the specificity, PPV and NPV were 96%, 88% and 69%, respectively. In the population of patients at high risk for invasive aspergillosis who also benefited from Aspergillus GM detection, the sensitivity and the NPV of the combined detection reached to 78% and 84%, respectively. Even higher NPV were obtained when combining negative PF-qPCR and CT scan (95%) as well as negative GM and CT scan (93%), thus allowing to rationalize and re-evaluate the prescription of empirical treatment in such highly selected population.

Our 2015 approach to invasive pulmonary aspergillosis.

At the University Hospital of Cologne, in general two patient groups at high risk for invasive aspergillosis receive posaconazole prophylaxis: Acute myelogenous leukaemia patients during remission induction chemotherapy and allogeneic haematopoietic stem cell transplant recipients. Other patients at risk undergo serum galactomannan testing three times weekly. At 72-96h of persisting fever despite broad-spectrum antibiotics, or at onset of lower respiratory tract symptoms a thoracic computed tomography (CT) scan is performed. Without lung infiltrates on CT, IPA is ruled out. In lung infiltrates not suggestive for IPA mycological confirmation is pursued. In patients without posaconazole prophylaxis empiric caspofungin will be considered. CT findings typical for IPA prompt targeted treatment, and mycological confirmation. Bronchoalveolar lavage (BAL) is most important for cultural identification and susceptibility testing, and facilitates diagnosing other pathogens. BAL performance is virtually independent of platelet counts. If despite suggestive infiltrates BAL does not yield the diagnosis, CT-guided biopsy follows as soon as platelet counts allow. Surgery can also be beneficial in diagnosis and treatment of IPA. If the diagnosis of IPA is not established, mucormycosis is a valid concern. In patients with breakthrough IPA during posaconazole prophylaxis liposomal amphotericin B is the drug of choice. If no posaconazole prophylaxis was given, voriconazole is the treatment of choice for IPA.

Retrospective Comparison of Nucleic Acid Sequence–Based Amplification, Real-Time PCR, and Galactomannan Test for Diagnosis of Invasive Aspergillosis

Invasive aspergillosis is a life-threatening infection in immunocompromised patients, and treating these infections at an early stage is often crucial for a favorable outcome. Early diagnosis, however, remains challenging. We performed a retrospective comparison of three methods: real-time quantitative PCR (qPCR), nucleic acid sequence-based amplification (NASBA), and galactomannan enzyme-linked immunosorbent assay (GM-ELISA); these detect circulating Aspergillus DNA, RNA, and galactomannan, respectively. Blood samples from 80 patients at high risk for invasive aspergillosis were tested by each assay. The sensitivity of NASBA, qPCR, and GM-ELISA was 76.47% (95% CI, 58.4-88.6%), 67.65% (95% CI, 49.4-82.0%), and 52.94% (95% CI, 35.4-69.8%), respectively, and the specificity was 80.43% (95% CI, 65.6-90.1%), 89.13% (95% CI, 75.6-95.9%), and 80.43% (95% CI, 65.6-90.1%), respectively. We also evaluated the efficiency of the three tests in various combinations. Perfect specificity (100%; 95% CI, 90.4-100%) and perfect positive predictive value (100%; 95% CI, 77.1-100%) were achieved by combining NASBA and qPCR testing in series. Testing with both NASBA and qPCR in parallel was the most sensitive and had the highest Youden index. Our data support the great potential of NASBA and qPCR, singly or in combination, for diagnosis of invasive aspergillosis in high-risk populations.

Immunosuppressive Compounds Exhibit Particular Effects on Functional Properties of Human Anti-Aspergillus T H 1 Cells

Allogeneic hematopoietic stem cell transplant (HSCT) recipients are at high risk for invasive aspergillosis. Whereas adoptive immunotherapy transferring donor-derived anti-Aspergillus TH1 cells has been shown to be beneficial for HSCT recipients suffering from invasive aspergillosis, little is known about the impact of commonly used immunosuppressants on the functional properties of anti-Aspergillus TH1 cells. Anti-Aspergillus TH1 cells were coincubated with different concentrations of methylprednisolone, cyclosporine (CsA), mycophenolic acid (MPA), the active component of mycophenolate mofetil, and rapamycin. Immunosuppressants were tested in concentrations reflecting common target levels in serum and in significantly lower and higher concentrations. Apoptosis of anti-Aspergillus TH1 cells, as well as proliferation and production of gamma interferon (IFN-γ) and CD154 upon restimulation, was evaluated in the presence and absence of immunosuppressive compounds. All dosages of CsA, MPA, and methylprednisolone significantly decreased the number of viable anti-Aspergillus TH1 cells in the cell culture, which was due partly to an impaired proliferative capacity of the cells and partly to an increased rate of apoptosis. In addition, CsA significantly decreased the number of IFN-γ-producing cells and had the highest impact of all immunosuppressants on IFN-γ levels in the supernatant. CsA also significantly decreased the expression of CD154 by anti-Aspergillus TH1 cells. Variant dosages of immunosuppressants exhibit particular effects on essential functional properties of anti-Aspergillus TH1 cells. Our findings may have an important impact on the design of clinical trials evaluating the therapeutic benefit of anti-Aspergillus TH1 cells in allogeneic HSCT recipients suffering from invasive aspergillosis.