Diagnosis Of Invasive Aspergillosis Research Articles

Evaluation of Serum Galactomannan Enzyme Immunoassay at Two Different Cut-offs for the Diagnosis of Invasive Aspergillosis in Patients with Febrile Neutropenia

Background: Invasive aspergillosis (IA) is an increasingly common and fatal opportunistic fungal infection in patients with haematological diseases. Early diagnosis is difficult as mycological culture techniques have low sensitivity and the radiological tools have low specificity. Galactomannan enzyme immunoassay (GEI) detects galactomannan in the human serum with a reported sensitivity and specificity between 30% and 100%. Aims: The aim of this study was to analyse the role of GEI in diagnosis of IA in patients with febrile neutropenia and to evaluate the role of GEI in the diagnosis of IA as per the revised (2008) European Organization for Research and Treatment of Cancer–Mycoses Study Group (EORTC–MSG) criteria at two different optical density (OD) cut-offs of 0.5 and 1.0. Setting: This prospective study was conducted in Safdarjung Hospital, New Delhi, India. Methods: GEI testing was performed in adult patients of febrile neutropenia with evidence of IA. Results at two different OD indices (ODIs) of 0.5 and 1.0 were analysed. The evaluation of the diagnostic parameter, that is, GEI was measured in terms of sensitivity, specificity and positive and negative predictive value and was validated with the revised (2008) EORTC–MSG diagnostic criteria of IA. Results: One hundred and eleven patients had evidence of IA, of which 79 patients were GEI positive when cut-off ODI was 0.5, whereas with cut-off ODI 1.0, 55 patients were GEI positive. Conclusion: ODI of 1.0 should be considered as positive while in patients with OD between 0.5 and 1.0, repeat sampling from the patient is recommended.

Long-term follow up of invasive aspergillosis in allogeneic stem cell transplantation recipients and leukemia patients: Differences in risk factors and outcomes

Antifungal prophylaxis (AP) has dramatically changed the epidemiology of invasive aspergillosis (IA). To better understand the differences in terms of clinical significance of IA between allogeneic stem cell transplantation (allo-SCT) recipients and patients treated for leukemia, we report a single-center study of 735 unselected consecutive patients treated between 2000 and 2004, before the era of systematic AP. Probable or confirmed IA were observed in 29 patients (2008 EORTC/MSG criteria), including 7/235 undergoing allo-SCT (5.2%), 19/380 treated for acute leukemia (5.0%), 1/116 for chronic lymphocytic leukemia (0.9%) and 2/104 for myelodysplastic syndrome (1.9%). In allo-SCT recipients, IA occurred later than in leukemia patients, after the neutropenic period. The median time between the last treatment and the diagnosis of IA was 231 days (range, 68–341) in allo-SCT recipients and 17 days (6–57) in leukemia patients (P<0.001). Importantly, the 7 cases of IA after allo- SCT occurred only in patients treated with corticosteroids for graft-versus-host disease (GVHD). Mortality directly related to IA was 24%. The 100-day, 2-year and 10-year overall survival were 42.9%, 0%, 0% in allo-SCT recipients compared to 68.1%, 18.2%, 13.6% in leukemia patients, respectively (P≥0.05). These poor outcomes were mainly attributable to non-relapse mortality (NRM). In conclusion, our data allows distinguishing 2 types of IA occurring at different time in the treatment course. In both cases, the NRM is very high and treatment remains challenging. Thus, systematic broad-spectrum AP against Aspergillus should be considered in acute leukemia patients during the neutropenic phase and in all patients undergoing allo-SCT who develop GVHD.

(40) - Aspergillus Galactomannan Detection in Exhaled Breath Condensate for the Diagnosis of Invasive Aspergillosis in Lung Transplant Recipients

(40) - Aspergillus Galactomannan Detection in Exhaled Breath Condensate for the Diagnosis of Invasive Aspergillosis in Lung Transplant Recipients

Challenges in microbiological diagnosis of invasive Aspergillus infections

Invasive aspergillosis (IA) has been increasingly reported in populations other than the historical hematology patients and there are new questions about the performance of microbiological tools. Microscopy and culture have been completed by biomarkers, either antigens or DNA, and in blood or respiratory specimens or both. First studied in hematology, the antigen galactomannan performance in serum is low in other patient populations where the pathophysiology of the infection can be different and the prevalence of IA is much lower. DNA detection with polymerase chain reaction (PCR) in blood or serum (or both) has reached a certain level of acceptance thanks to consensus methods based on real-time quantitative PCR (qPCR). When used on respiratory specimens, galactomannan and qPCR depend on standardization of the sampling and the diverse mycological procedures. Thus, culture remains the main diagnostic criterion in critically ill patients. The current trend toward more effective anti-mold prophylaxis in hematology hampers the yield of a screening strategy, as is usually performed in hematology. Therefore, circulating biomarkers as confirmatory tests should be considered and their performance should be reappraised in each new setting. The use of azole prophylaxis also raises the issue of selecting azole-resistance Aspergillus fumigatus isolates. Ideally, the biomarkers will be more efficient when individual genetic risks of IA are defined. Culture, though not standardized, remains a key element for the diagnosis of IA and has the advantage to easily detect molds other than A. fumigatus. It is still unclear whether next-generation sequencing will replace culture in the future.

Prognostic value of galactomannan: current evidence for monitoring response to antifungal therapy in patients with invasive aspergillosis.

Galactomannan (GM) is a polysaccharide present in the cell wall of Aspergillus spp. that is released during growth of the organism. It has been successfully used to aide in the diagnosis of invasive aspergillosis allowing for earlier recognition of disease compared to conventional methods. Since its implementation in the clinic as a diagnostic tool, GM has been used in experimental models to measure therapeutic response. Several clinical studies describe the prognostic value of GM. Herein, we review the evidence supporting the utilization of GM antigen as a biomarker to measure response to systemic antifungal therapy.

Comparison of serum PCR assay and histopathology for the diagnosis of invasive aspergillosis and mucormycosis in immunocompromised patients with sinus involvement.

Background and Purpose:Invasive fungal infections cause morbidity and mortality in patients with hematologic malignancies and immunosuppression. Although these infections are commonly caused by Candida and Aspergillus species, infections caused by Mucoralean fungi are also on a growing trend. The definitive diagnosis of mucormycosis includes visualization of non-septate hyphae on pathology or growth of Mucoralean fungi culture. Polymerase chain reaction (PCR) is used to diagnose mucormycosis from paraffin blocks; however, it yields discrepant results in diagnosis of mucormycosis from blood samples. In the current study, we sought to examine the efficiency of PCR test for the diagnosis of mucormycosis and aspergillosis.Materials and Methods:Thirty-one patients with suspected fungal sinus infection were recruited from the Hematology-Oncology unit in Taleghani Hospital, Tehran, Iran. DNA was extracted and semi-nested PCR was performed.Results:PCR was reported negative for all the 31 serum samples. Our assay had a sensitivity of 1.3 ng and 12 pg for Mucoralean and Aspergillus species, respectively.Conclusion:Using serum PCR, we detected Aspergillus and Mucoralean species in patients with suspected fungal sinus infection. While this test may have utility in diagnosis directly from biopsy site, it appears unreliable for use as a noninvasive blood test.

Impact of pre-analytical variables in the determination of serum galactomannan.

Invasive aspergillosis (IA) is associated with a high morbidity and mortality. Since Aspergillus species are usually not cultured in these patients, presumptive diagnosis of IA is more commonly based on galactomannan (GM) detection. Several factors are known to cause false-positive results in the GM test, but little is known on the influence of pre-analytical variables interfering on the test. Here we studied the influence of temperature and sample storage duration in GM results, using samples known to be negative and positive (spiked) for GM. We also evaluated the effect of hemolysis and hyperbilirubinemia on GM optical indexes. We found no influence of storage time (up to 96 h) and temperatures (refrigerated vs. RT) on GM results. However, bilirubin (P = .022) and haemoglobin (P = .003) content influenced GM readings in samples known for being GM positive and negative at baseline, respectively. We conclude that the Platelia GM test does not suffer major influence of pre-analytical variables such as storage conditions, and low levels of hemolysis and hyperbilirubinemia. Nonetheless, massive haemolysis seems to interfere with GM readings in GM-negative samples, and high levels of bilirubin can affect GM readings in samples that are positive for GM at baseline. These findings may facilitate logistics and the implementation of standard operational procedures in clinical laboratories.

Utility of the serum galactomannan assay for the diagnosis of invasive aspergillosis in children with acute lymphoblastic leukemia

Invasive aspergillosis (IA) is an important cause of mortality and morbidity in children with hematological malignancies. The monitoring of serum galactomannan (GM) antigen is considered useful in the diagnosis of IA . The aim of this study was to determine the utility of serum GM monitoring in the early diagnosis of IA and the role of positive antigenemia in the management of children with acute lymphoblastic leukemia (ALL). The cases of 141 children who were being treated for ALL in the Division of Pediatric Hematology of the Medical School of Ege University between January 2006 and February 2015 were reviewed retrospectively. Cases of proven and probable IA were defined according to the European Organization for the Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG) criteria. The incidence of proven and probable IA was 3.5% (5/141). The incidence of positive GM antigenemia among 3264 serum samples was 5.5% (n=179). Of the cases detected, 21.7% were true-positive, 52.1% were false-positive, and the remaining 26.1% were classified as 'undetermined.' An increase in the incidence of true-positive tests and induction of antifungal therapy was determined through multiple consecutive positive tests. GM may be detected in the serum before the clinical signs of IA appear, but its sensitivity and specificity are variable. False-positivity is a significant disadvantage, and consecutive positive GM must be taken into account in the case of clinical and imaging findings that are relevant to IA.

Galactomannan testing and Aspergillus PCR in same-day bronchoalveolar lavage and blood samples for diagnosis of invasive aspergillosis.

In recent years galactomannan antigen testing (GM) and also Aspergillus PCR have become increasingly important for diagnosis of invasive aspergillosis (IA). Whether or not these tests need to be performed with bronchoalveolar lavage fluid (BALF; i.e., primary site of infection), or testing of blood samples is sufficient, remains, however, a matter of debate. We evaluated the diagnostic performance of GM ELISA, and Aspergillus PCR by using BALF samples and blood samples obtained at the same day from a total of 53 immunocompromised patients (16 with probable/proven IA and 37 with no evidence of IA according to the revised EORTC/MSG criteria; 38 patients with hematological malignancies were prospectively enrolled at the Medical University of Graz, Austria, 15 patients with mixed underlying diseases at the Mannheim University Hospital). Patients with possible IA were excluded from this analysis. A total of 34/53 (64%) of all patients and 12/16 (75%) of patients with probable/proven IA received mold-active antifungal prophylaxis/therapy at the time of the BALF procedure. Sensitivities of GM and Aspergillus PCR were 38% and 44% in BALF, and 31% and 0% in blood, respectively. Best sensitivity (75%) for detecting proven/probable IA was achieved when BALF Aspergillus PCR, BALF GM (>1.0 ODI), BALF-culture and serum-GM (>0.5 ODI) were combined (specificity 95%). In conclusion, sensitivities of the evaluated diagnostic tests-when interpreted on their own-were low in BALF and even lower in blood, sensitivities increased markedly when diagnostic tests were combined.

Determining the analytical specificity of PCR-based assays for the diagnosis of IA: What is Aspergillus?

A wide array of PCR tests has been developed to aid the diagnosis of invasive aspergillosis (IA), providing technical diversity but limiting standardisation and acceptance. Methodological recommendations for testing blood samples using PCR exist, based on achieving optimal assay sensitivity to help exclude IA. Conversely, when testing more invasive samples (BAL, biopsy, CSF) emphasis is placed on confirming disease, so analytical specificity is paramount. This multicenter study examined the analytical specificity of PCR methods for detecting IA by blind testing a panel of DNA extracted from a various fungal species to explore the range of Aspergillus species that could be detected, but also potential cross reactivity with other fungal species. Positivity rates were calculated and regression analysis was performed to determine any associations between technical specifications and performance. The accuracy of Aspergillus genus specific assays was 71.8%, significantly greater (P < .0001) than assays specific for individual Aspergillus species (47.2%). For genus specific assays the most often missed species were A. lentulus (25.0%), A. versicolor (24.1%), A. terreus (16.1%), A. flavus (15.2%), A. niger (13.4%), and A. fumigatus (6.2%). There was a significant positive association between accuracy and using an Aspergillus genus PCR assay targeting the rRNA genes (P = .0011). Conversely, there was a significant association between rRNA PCR targets and false positivity (P = .0032). To conclude current Aspergillus PCR assays are better suited for detecting A. fumigatus, with inferior detection of most other Aspergillus species. The use of an Aspergillus genus specific PCR assay targeting the rRNA genes is preferential.

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.

Performance evaluation of multiplex PCR including Aspergillus—not so simple!: Table 1.

Multiplex PCRs have been designed for including species other than Aspergillus fumigatus for the diagnosis of invasive aspergillosis, such as microarrays, liquid-phase array, and electrospray-ionization mass spectrometry (PCR/ESI MS). These methods are based on the selection of multiple primers to amplify different species with the specificity checked by hybridization to a probe or by base composition of the amplicon for the PCR/ESI MS. When testing complex samples such as respiratory specimens, some clinically relevant species can be missed. Indeed, it is impossible to design primers able to amplify all the known fungal species with the same efficiency. Therefore, the best amplified species may not be the most clinically relevant. Multiplex assays have also been proposed to detect A. fumigatus DNA and azole resistance. Since the gene responsible for azole resistance is single copy and the gene used for detection is multicopy, only the high fungal loads can be evaluated. Thus, although interesting for investigating mycobiome, the multiplex assays should be used with cautious for the diagnosis of IA or the detection of resistance. For the diagnosis of invasive aspergillosis, validated quantitative PCRs specifically targeting A. fumigatus or a limited set of species to increase sensitivity is a safer option.

Aspergillus nodules; another presentation of Chronic Pulmonary Aspergillosis.

BackgroundThere are a number of different manifestations of pulmonary aspergillosis. This study aims to review the radiology, presentation, and histological features of lung nodules caused by Aspergillus spp.MethodsPatients were identified from a cohort attending our specialist Chronic Pulmonary Aspergillosis clinic. Patients with cavitating lung lesions, with or without fibrosis and those with aspergillomas or a diagnosis of invasive aspergillosis were excluded. Demographic, laboratory, and clinical data and radiologic findings were recorded.ResultsThirty-three patients with pulmonary nodules and diagnostic features of aspergillosis (histology and/or laboratory findings) were identified. Eighteen (54.5 %) were male, mean age 58 years (range 27–80 years). 19 (57.6 %) were former or current smokers. The median Charleston co-morbidity index was 3 (range 0–7). All complained of a least one of; dyspnoea, cough, haemoptysis, or weight loss. None reported fever. Ten patients (31 %) did not have an elevated Aspergillus IgG, and only 4 patients had elevated Aspergillus precipitins. Twelve patients (36 %) had a single nodule, six patients (18 %) had between 2 and 5 nodules, 2 (6 %) between 6 and 10 nodules and 13 (39 %) had more than 10 nodules. The mean size of the nodules was 21 mm, with a maximum size ranging between 5–50 mm. No nodules had cavitation radiographically. The upper lobes were most commonly involved. Histology was available for 18 patients and showed evidence of granulation tissue, fibrosis, and visualisation of fungal hyphae.ConclusionPulmonary nodules are a less common manifestation of aspergillosis in immunocompetent patients. Distinguishing these nodules from other lung pathology may be difficult on CT findings alone.

PCR Technology for Detection of Invasive Aspergillosis.

The application of molecular technologies to aid diagnosis and management of infectious diseases has had a major impact and many assays are in routine use. Diagnosis of aspergillosis has lagged behind. Lack of standardization and limited commercial interest have meant that PCR was not included in consensus diagnostic criteria for invasive fungal disease. In the last ten years careful evaluation and validation by the Aspergillus European PCR initiative with the development of standardized extraction, amplification and detection protocols for various specimen types, has provided the opportunity for clinical utility to be investigated. PCR has the potential to not only exclude a diagnosis of invasive aspergillosis but in combination with antigen testing may offer an approach for the early diagnosis and treatment of invasive aspergillosis in high-risk populations, with the added benefit of detection of genetic markers associated with antifungal resistance.

Galactomannan and 1,3-β-d-Glucan Testing for the Diagnosis of Invasive Aspergillosis

Invasive aspergillosis (IA) is a severe complication among hematopoietic stem cell transplant recipients or patients with hematological malignancies and neutropenia following anti-cancer therapy. Moreover, IA is increasingly observed in other populations, such as solid-organ transplant recipients, patients with solid tumors or auto-immune diseases, and among intensive care unit patients. Frequent delay in diagnosis is associated with high mortality rates. Cultures from clinical specimens remain sterile in many cases and the diagnosis of IA often only relies on non-specific radiological signs in the presence of host risk factors. Tests for detection of galactomannan- (GM) and 1,3-β-d-glucan (BDG) are useful adjunctive tools for the early diagnosis of IA and may have a role in monitoring response to therapy. However, the sensitivity and specificity of these fungal biomarkers are not optimal and variations between patient populations are observed. This review discusses the role and interpretation of GM and BDG testing for the diagnosis of IA in different clinical samples (serum, bronchoalveolar lavage fluid, cerebrospinal fluid) and different groups of patients (onco-hematological patients, solid-organ transplant recipients, other patients at risk of IA).

Pediatric Invasive Aspergillosis.

Invasive aspergillosis (IA) is a disease of increasing importance in pediatrics due to growth of the immunocompromised populations at risk and improvements in long-term survival for many of these groups. While general principles of diagnosis and therapy apply similarly across the age spectrum, there are unique considerations for clinicians who care for children and adolescents with IA. This review will highlight important differences in the epidemiology, clinical manifestations, diagnosis, and therapy of pediatric IA.

Anti-Aspergillus immunoglobulin-G testing in serum of hematopoietic stem cell transplant recipients.

Pulmonary invasive aspergillosis (IA) is a major clinical problem in patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT). Acquisition of IA during allo-HSCT by inhalation of spores is the rationale for the widespread use of air filtration systems. Recent data suggest that activation of fungal growth in already colonized patients is a relevant factor, and a recent study found a positive correlation of serum immunoglobulin responses against purified recombinant Aspergillus fumigatus proteins before allo-HSCT with the incidence of IA after allo-HSCT. To investigate the clinical utility of this approach, we performed a prospective study. We used a commercially available and standardized assay for detection of anti-Aspergillus immunoglobulin-G (aA-IgG) in serum (Platelia(™) Aspergillus IgG) that has previously demonstrated high sensitivity and specificity. In a cohort of 104 allo-HSCT recipients, we measured aA-IgG and Aspergillus antigen serum levels before allo-HSCT, and weekly during hospital stay. Overall prevalence of possible, probable, and proven IA during hospital stay was 10%, 6%, and 0%. We found no correlation between aA-IgG levels before allo-HSCT, or after allo-HSCT, and the prevalence of IA during hospital stay. Furthermore, median aA-IgG levels did not differ between patients with history of probable or proven IA, as compared to patients without history of IA. Taken together, our data argue against the clinical utility of measuring aA-IgG levels for diagnosis or prediction of IA in patients undergoing allo-HSCT.

Galactomannan in bronchoalveolar lavage fluid for diagnosis of invasive aspergillosis in non-hematological patients

The role of galactomannan (GM) in serum or bronchoalveolar lavage fluid (BALF) for the diagnosis of invasive pulmonary aspergillosis (IPA) has been extensively evaluated in hematological patients, however its performance in non-hematological patients is not well established. We performed a multicenter retrospective study in 3 university hospitals in Madrid, Spain between 2010 and 2014. The study population comprised patients with chronic obstructive pulmonary disease (COPD) and patients with immunosuppressive conditions in whom IPA was suspected and for whom BALF GM was available. Patients with hematological disorders were excluded. A total of 188 patients (35 with COPD and 153 with immunosuppressive conditions) were analyzed, and 31 cases of IPA (proven or probable) were identified. The global sensitivity of BALF GM (optical density index [ODI]≥1.0) was 77.4%; sensitivity was higher in patients with immunosuppressive conditions than in patients with COPD (81.8% vs 66.7%; p: 0.38). In COPD patients, the best performance was obtained for BALF GM (ODI≥0.5), although sensitivity (88.9%) was similar to that of BALF fungal culture (88.9%). The sensitivity of GM in serum was very poor in both populations (36.4% and 11.6%, respectively). In the present series, the diagnostic performance of BALF GM was good for IPA in non-hematological patients, especially in patients with immunosuppressive conditions.

Genome-Wide Expression Profiling Reveals S100B as Biomarker for Invasive Aspergillosis.

Invasive aspergillosis (IA) is a devastating opportunistic infection and its treatment constitutes a considerable burden for the health care system. Immunocompromised patients are at an increased risk for IA, which is mainly caused by the species Aspergillus fumigatus. An early and reliable diagnosis is required to initiate the appropriate antifungal therapy. However, diagnostic sensitivity and accuracy still needs to be improved, which can be achieved at least partly by the definition of new biomarkers. Besides the direct detection of the pathogen by the current diagnostic methods, the analysis of the host response is a promising strategy toward this aim. Following this approach, we sought to identify new biomarkers for IA. For this purpose, we analyzed gene expression profiles of hematological patients and compared profiles of patients suffering from IA with non-IA patients. Based on microarray data, we applied a comprehensive feature selection using a random forest classifier. We identified the transcript coding for the S100 calcium-binding protein B (S100B) as a potential new biomarker for the diagnosis of IA. Considering the expression of this gene, we were able to classify samples from patients with IA with 82.3% sensitivity and 74.6% specificity. Moreover, we validated the expression of S100B in a real-time reverse transcription polymerase chain reaction (RT-PCR) assay and we also found a down-regulation of S100B in A. fumigatus stimulated DCs. An influence on the IL1B and CXCL1 downstream levels was demonstrated by this S100B knockdown. In conclusion, this study covers an effective feature selection revealing a key regulator of the human immune response during IA. S100B may represent an additional diagnostic marker that in combination with the established techniques may improve the accuracy of IA diagnosis.

Detection of a Serum Siderophore by LC-MS/MS as a Potential Biomarker of Invasive Aspergillosis.

Invasive aspergillosis (IA) is a life-threatening systemic mycosis caused primarily by Aspergillus fumigatus. Early diagnosis of IA is based, in part, on an immunoassay for circulating fungal cell wall carbohydrate, galactomannan (GM). However, a wide range of sensitivity and specificity rates have been reported for the GM test across various patient populations. To obtain iron in vivo, A. fumigatus secretes the siderophore, N,N',N"-triacetylfusarinine C (TAFC) and we hypothesize that TAFC may represent a possible biomarker for early detection of IA. We developed an ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for TAFC analysis from serum, and measured TAFC in serum samples collected from patients at risk for IA. The method showed lower and upper limits of quantitation (LOQ) of 5 ng/ml and 750 ng/ml, respectively, and complete TAFC recovery from spiked serum. As proof of concept, we evaluated 76 serum samples from 58 patients with suspected IA that were investigated for the presence of GM. Fourteen serum samples obtained from 11 patients diagnosed with probable or proven IA were also analyzed for the presence of TAFC. Control sera (n = 16) were analyzed to establish a TAFC cut-off value (≥6 ng/ml). Of the 36 GM-positive samples (≥0.5 GM index) from suspected IA patients, TAFC was considered positive in 25 (69%). TAFC was also found in 28 additional GM-negative samples. TAFC was detected in 4 of the 14 samples (28%) from patients with proven/probable aspergillosis. Log-transformed TAFC and GM values from patients with proven/probable IA, healthy individuals and SLE patients showed a significant correlation with a Pearson r value of 0.77. In summary, we have developed a method for the detection of TAFC in serum that revealed this fungal product in the sera of patients at risk for invasive aspergillosis. A prospective study is warranted to determine whether this method provides improved early detection of IA.