Course Of Invasive Aspergillosis Research Articles

Clinical insights into Invasive Aspergillosis in Mexican children with cancer

Abstract Background In the last years Invasive Aspergillosis (IA) has become the most frequent invasive fungal infection (IFI) affecting children with hematological malignancies. This complication results in the delay of cancer treatment and/or poor prognosis mainly due to late diagnosis and late start of antifungal treatment. Therefore, antifungal prophylaxis and preemptive approaches have been proposed for these patients based on risk factors and local epidemiological data. For this reason, we performed a unicenter, descriptive and prospective study to determine the frequency and clinical course of IA in hospitalized cancer patients with fever and neutropenia. Methods we included events of fever and neutropenia of hospitalized patients with cancer and either with persistent febrile neutropenia (PFN: fever for >96h with total neutrophil count of <500cells/mm3) or profound prolonged neutropenia (PPN: total neutrophil count of <500cells/mm3 for >7 days). Informed consent was requested prior inclusion and follow up of these events was performed with weekly measurement of serum galactomannan antigen (until the recovery of neutrophil count), lung computed tomography (CT) and/or sinus and/or central nervous system (CNS) imaging, depending on clinical findings, and if feasible bronchoscopy to obtain bronchioalveolar lavage (BAL) samples. Clinical and laboratory data were obtained; with this information diagnosis of IA was classified as possible, probable, and proven, according to the EORTC/MSG guidelines. This protocol was approved by the hospital’s research and ethics committees (INP 041/2021). For data analysis categorical variables were presented as frequencies and numerical variables as median and range. Statistical analysis was made with X2 and U Mann Whitney tests. Results Seventy events were included, 14 (20%) had IA diagnosis; 2 possible, 11 probable and 1 proved. Median of age was 10.2y (5.5 – 17) and 64.3% were male. Acute lymphoblastic leukemia was the main oncologic diagnosis (71.4%), 35.7% during induction phase; followed by acute myeloblastic leukemia (21.4%). 92.9% of events with IA presented as PFN (p=0.001) with a fever duration of 3.5 days (1-8) (p=0.004). Pulmonary disease was evident in all events with IA, two had disseminated infection. In 5/14 events serum galactomannan antigen was positive in the first measurement. Only 8 events underwent bronchoscopy: BAL was positive in 6 and 4 had positive culture with Aspergillus spp. Lung CT scan showed different findings, lobar/segmental consolidation and nodule with halo sign were the most common; 85.7% of cases presented with more than one radiological sign (p=0.003). Voriconazole monotherapy was given to 78.5% of IA cases and 21.4% required combined antifungal therapy (added caspofungine or liposomal amphotericin B) due to disseminated disease or severe clinical course. Intensive care admission resulted with statistical significance for the events with diagnosis of IA (64.3% p= 0.019). Within the IA events survival at 60 days was of 78.6%. Conclusion Frequency of IA was high among our high risk cancer patients. This study demonstrates the importance of a complete work up for invasive fungal infection in high-risk patients with persistent febrile neutropenia. The information obtained is of great use for the implementation of pre-emptive approaches and antifungal prophylaxis in this patient population.

Baseline predictors influencing the prognosis of invasive aspergillosis in adults.

Invasive aspergillosis (IA) is a serious hazard to haematological and critical care patients. Impactful risk factors for developing IA have been characterised; however, systematic analysis of baseline prognostic factors for treatment course of IA is missing. To understand prognostic variables, we analysed original articles identifying baseline factors that predict treatment outcome in patients with IA. PubMed database was searched for publications since database inception until May 2018. Inclusion criteria were published baseline prognostic factors present at the diagnosis of IA. In total, 58 studies from 267 centres reported 7320 patients with IA and 40 different predictors. Unfavourable predictors in medical history were kidney (7.4%, 10/136) and liver failure (3.7%, 5/136), ICU admission (3.7%, 5/136) and uncontrolled underlying disease (3.7%, 5/136). Regarding state of immunosuppression, negative outcome predictors were prolonged neutropenia (12.5%, 17/136), corticosteroid treatment (8.1%, 11/136) and graft-vs-host disease (3.7%, 5/136). On the pathogen side, relevant predictors were galactomannan positivity (8.1%, 11/136), Aspergillus terreus infection (2.2%, 3/136) and lack of amphotericin B susceptibility (1.5%, 2/136). IA-specific predictors were disseminated disease (5.1%, 7/136) and CNS involvement (2.9%, 4/136). Imaging results associated with negative outcome were multiple consolidations (2.9%, 4/136), bipulmonary lesions (2.2%, 3/136) and pleural effusion (2.2%, 3/136). At diagnosis of IA, most frequently identified predictors of outcome were neutropenia, corticosteroid use, elevated galactomannan, renal failure and disseminated disease. The predictors may be used to identify patients at high risk for treatment failure and to stratify neglected patient groups for clinical trials.

Fever-range temperature modulates activation and function of human dendritic cells stimulated with the pathogenic mould Aspergillus fumigatus

In immunocompromised patients, invasive aspergillosis (IA) is the most frequent disease caused by the pathogenic mould Aspergillus fumigatus. Fever is one of the most common yet nonspecific clinical symptoms of IA. To evaluate the role of hyperthermia in the innate immune response to A. fumigatus in vitro, human monocyte-derived dendritic cells (DCs) were stimulated with germ tubes of A. fumigatus or the fungal cell wall component zymosan at 37°C or 40°C, followed by characterization of specific DC functions. While maturation of DCs was enhanced and DC phagocytic capacity was reduced at 40°C, we observed that DC viability and cytokine release were unaffected. Thus, our results suggest that hyperthermia has substantial impacts on DC function in vitro, which might also influence the course and outcome of IA in immunocompromised patients.

Characterization of Specific Immune Responses to Different Aspergillus Antigens during the Course of Invasive Aspergillosis in Hematologic Patients

Several studies in mouse model of invasive aspergillosis (IA) and in healthy donors have shown that different Aspergillus antigens may stimulate different adaptive immune responses. However, the occurrence of Aspergillus-specific T cells have not yet been reported in patients with the disease. In patients with IA, we have investigated during the infection: a) whether and how specific T-cell responses to different Aspergillus antigens occur and develop; b) which antigens elicit the highest frequencies of protective immune responses and, c) whether such protective T cells could be expanded ex-vivo. Forty hematologic patients have been studied, including 22 patients with IA and 18 controls. Specific T cells producing IL-10, IFN-γ, IL-4 and IL-17A have been characterized through enzyme linked immunospot and cytokine secretion assays on 88 peripheral blood (PB) samples, by using the following recombinant antigens: GEL1p, CRF1p, PEP1p, SOD1p, α1–3glucan, β1–3glucan, galactomannan. Specific T cells were expanded through short term culture. Aspergillus-specific T cells producing non-protective interleukin-10 (IL-10) and protective interferon-gamma (IFN-γ) have been detected to all the antigens only in IA patients. Lower numbers of specific T cells producing IL-4 and IL-17A have also been shown. Protective T cells targeted predominantly Aspergillus cell wall antigens, tended to increase during the IA course and to be associated with a better clinical outcome. Aspergillus-specific T cells could be successfully generated from the PB of 8 out of 8 patients with IA and included cytotoxic subsets able to lyse Aspergillus hyphae. Aspergillus specific T-cell responses contribute to the clearance of the pathogen in immunosuppressed patients with IA and Aspergillus cell wall antigens are those mainly targeted by protective immune responses. Cytotoxic specific T cells can be expanded from immunosuppressed patients even during the infection by using the above mentioned antigens. These findings may be exploited for immunotherapeutic purposes in patients with IA.

Detection of Urinary Excreted Fungal Galactomannan-like Antigens for Diagnosis of Invasive Aspergillosis

Mortality associated with invasive aspergillosis (IA) remains high, partly because of delayed diagnosis. Detection of microbial exoantigens, released in serum and other body fluids during infection, may help timely diagnosis. In course of IA, Aspergillus galactomannan (GM), a well established polysaccharide biomarker, is released in body fluids including urine. Urine is an abundant, safely collected specimen, well-suited for point-of-care (POC) testing, which could play an increasing role in screening for early disease. Our main objective was to demonstrate GM antigenuria as a clinically relevant biological phenomenon in IA and establish proof-of-concept that it could be translated to POC diagnosis. Utilizing a novel IgM monoclonal antibody (MAb476) that recognizes GM-like antigens from Aspergillus and other molds, we demonstrated antigenuria in an experimental animal IA model (guinea pig), as well as in human patients. In addition, we investigated the chemical nature of the urinary excreted antigen in human samples, characterized antigen detection in urine by immunoassays, described a putative assay inhibitor in urine, and indicated means of alleviation of the inhibition. We also designed and used a lateral flow immunochromatographic assay to detect urinary excreted antigen in a limited number of IA patient urine samples. In this study, we establish that POC diagnosis of IA based on urinary GM detection is feasible. Prospective studies will be necessary to establish the performance characteristics of an optimized device and define its optimal clinical use.

Baseline Platelet Count and Creatinine Clearance Rate Predict the Outcome of Neutropenia-Related Invasive Aspergillosis

Invasive aspergillosis (IA) is a life-threatening infection for immunocompromised patients. Improvement in IA outcome has been hampered by lack of early prognostic factors, namely, those available before starting chemotherapy (baseline) or early in the course of IA (nonbaseline). We hypothesized that prognostic factors can be identified before chemotherapy, ≤7 days from the first positive serum Aspergillus galactomannan index (s-GMI). We analyzed 98 patients with multiple myeloma who developed neutropenia-related IA and had a positive s-GMI. Three response criteria were used: kinetics of s-GMI, European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) definitions, and 6-week survival. Baseline and nonbaseline variables were analyzed separately. Independent response predictors at baseline were a platelet count ≥65,000 platelets/mm(3) (odds ratio [OR], 1.009; 95% confidence interval [CI], 1.001-1.017; P = .03) by s-GMI kinetics, and a platelet count ≥65,000 platelets/mm(3) (OR, 1.009; 95% CI, 1.002-1.017; P = .01) and a creatinine clearance rate ≥53 mL/min (OR, 1.024; 95% CI, 1.006-1.042; P = .009) by EORTC/MSG criteria, with response rates of 83% and 28% when both variables were above or below these cutoffs, respectively (P < .001). Only baseline creatinine clearance rate ≥53 mL/min predicted 6-week survival (P = .003). Normalization of the s-GMI ≤7 days after the first positive s-GMI and neutrophil recovery were the nonbaseline factors associated with positive outcomes. Two simple, inexpensive to measure, widely available, and routinely collected prechemotherapy values, platelet count and creatinine clearance rate, predict IA outcome and stratify patients into low-, intermediate-, and high-risk categories, while early evaluation of s-GMI allows timely treatment modification. These findings may improve patient outcomes by optimizing management strategies for this serious infection and may prove valuable in designing clinical trials of interventions to improve IA outcomes.

Immune reconstitution syndrome and fungal infections

Fungal infections-related immune reconstitution syndrome (IRS) poses challenging diagnostic and management issues in immunocompromised hosts. This review summarizes the current state of knowledge regarding its pathophysiologic basis, presentation, and treatment strategies. Existing evidence suggests that IRS is a state of an imbalance between protective immunity and inflammatory pathology versus anti-inflammatory responses that restrain inflammation. IRS has been observed in diverse hosts including HIV-infected patients initiating potent antiretroviral therapy, transplant recipients, pregnant women, and recipients of iatrogenic biologic agents. Among the most common fungal infections associated with IRS is cryptococcosis, although this entity has been documented during the course of invasive aspergillosis, histoplasmosis, and candidiasis. Unique risk factors for IRS have been recognized in specific hosts. IRS is a culmination of immunologic sequelae of host-pathogen interaction during evolution of an opportunistic infection, and as such the development of biomarkers that differentiate it from progressive disease would represent an important advance. Optimal management of immunosuppression and immunomodulatory approaches that target precise regulatory pathways for IRS warrant future investigations.

Protective T-Cell Responses to Several Recombinant Aspergillus Antigens May Be Detected Since the Onset of the Infection in Patients with Invasive Aspergillosis, and May Be Exploited for Therapeutic Purposes,

Abstract 3229 Introduction:Several studies have reported that different components of fungi of the genera Aspergillus spp may induce protective T-cell responses in either mouse models of invasive aspergillosis (IA) or in human healthy subjects.We evaluated the occurrence of Aspergillus-specific T-cell responses to different Aspergillus recombinant antigens in patients with proven IA, during the course of the IA, to identify the antigens most frequently targeted by protective immune responses. We characterized phenotypically and functionally such specific T cells. Finally, from peripheral blood (PB) samples of the same IA proven patients, also collected during the active infection phase, we sought to expand such Aspergillus-specific T cells. Methods:15 patients with proven IA, according to the EORTC/MSG criteria, have been enrolled into the study. Specific immune responses producing interleukin-10 (IL-10), interferon-gamma (IFN-γ), IL-4 and IL-17A were detected and characterized by enzyme linked immunospot (ELISpot) assay and cytokine secretion assay (CSA), in all the patients, during the entire course of the IA. The recombinant antigens used were GEL1p, CRF1p, PEP1p, SOD1p, α1–3glucan, β1–3glucan, and galactomannan (GM). Cytotoxicity has been investigated by means of the colorimetric assay with (2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]2H-tetrazolium-5-carboxyanilide) sodium salt plus coenzyme Q0 (XTT assay). Aspergillus-specific T cells were obtained by culturing PB mononuclear cells with a mixture containing PEP1p, GEL1p, α1–3 glucan and β1–3 glucan. The infection course were divided into 4 phases, defined from t1 to t4, each of fifteen days interval, starting from the radiological diagnosis of IA. Results:Aspergillus-specific T cells producing either IL-10 or IFN-γ were detected to all the antigens, but GM. The number of antigens targeted by IFN-γ producing specific T cells progressively increased along the course of IA, being such protective responses to 3 out of 7 antigens at t1 and to 6 out of 7 antigens at t4. At t1, IFN-γ producing specific T cells were only detected to GEL1p, α1–3 glucan and β1–3 glucan. GEL1p and α1–3 glucan resulted the antigens most constantly targeted by IFN-γ producing specific T cells, persisting the responses to these antigens in all the phases of IA. No Aspergillus-specific T cells producing IL-4 to any antigens were detected by the ELISpot assay. Aspergillus-specific T cells producing IL-17A were detected in only one out of 15 patients, and targeted CRF1p. Specific T cells to GM and specific T cells producing IL-4 to all the antigens could be shown only by CSA, suggesting that they are present only at very low frequencies during the infection. After 13-day cultures, Aspergillus-specific T cells were expanded from five out of five patients. The specific T cells tested for lytic activity included a median of 95.8% CD3+ cells, either CD4+ or CD8+ T cells, and showed a median lytic activity of 9.45% either at 3/1 or at 5/1 effector/target cells ratios. Conclusions:In patients with IA, protective immune responses may be detected since the onset and increase during the infection. At the onset of IA, specific T cells producing IFN-γ target antigens involved in the cell wall biosynthesis of Aspergillus. On the other hand, at the same phase, specific protective immune responses to CRF1p, PEP1p, and SOD1p, which are all putative virulence factors for Aspergillus, are absent. Aspergillus-specific T cells may be expanded by a mixture of antigens, even in the course of IA and are able to directly kill fungal hyphae. The above mentioned antigens and the corresponding protective T cells may be exploited for therapeutic strategies of either vaccine or autologous cytotoxic cell infusions in patients at high risk for IA. Disclosures:Luppi:MSD; GILEAD: Research Funding.

Reply to Kontoyiannis

To the Editor We thank Dr Kontoyiannis for his interest and we appreciate his useful insights regarding our study. As highlighted by Dr Kontoyiannis, invasive aspergillosis (IA) invariably occurs in a complex clinical setting in which underlying hematologic disease, administered treatment, and concomitant disease processes can all influence the resultant host response. We had found that a persistence of elevated proinflammatory interleukin-6 and C-reactive protein at 1 week after initiation of antifungal therapy was associated with adverse outcome in a cohort of patients with IA recruited in a clinical trial [1]. While the intricacies of immune pathogenesis in IA remain to be fully understood, we cannot exclude that they are due to the evolution of underlying hematologic disease. One should also appreciate the fact that these cytokines are often associated with an infectious process and, in the context of our study, may reasonably be explained by the clinical course of IA. We have alluded to both aspects of infection and hematologic malignancy by discussing that: (1) the cytokine trends may be indicative of the host's incapacity to effect an appropriate antimicrobial immune response against the pathogen, while also making reference to a recent proposition that an uncurbed proinflammatory response may paradoxically facilitate the pathogenesis of IA [2]; and (2) similar biomarker trends were incidentally also observed in hematopoietic stem cell transplant recipients (HSCT) [3] and patients with sepsis [4]. Likewise, it is certainly plausible that neutrophil levels in the host could have an influence on the distinct results between our study and that of Roillides et al [5], as indicated. We thank Dr Kontoyiannis for highlighting these points as discussed in the article. The significance of these findings, however, lies in that these distinct biomarker trends were observed de novo during the course of managing the disease in a sizable cohort of patients with IA who primarily had pulmonary IA, as would have been encountered by clinicians at the bedside. The underlying diseases in the study cohort leading to IA was diverse: patients recruited had the various types of leukemias and lymphomas, had undergone autologous and allogenic HSCTs requiring appropriate chemotherapeutic regimens, and were at different stages of their hematologic diseases [1]. Approximately one-quarter of the patients had documented coinfections at various points in their illness, although the rate of mortality attributable to non-IA causes was not high. Given these mitigating circumstances, it would not have been easy to correct for all of the aforementioned complex variables during analysis. Nonetheless, to facilitate objective assessment of the utility of these biomarkers, we had depicted both the receiver operating curve diagrams with areas under the curve, as well as sensitivity and specificity values for the various cutoff levels that we deemed appropriate. We agree that a well-designed, prospective controlled trial examining cytokine and/or chemokine response in a large cohort of patients with IA would be ideal, and we look forward to the challenge of conducting such a study. Until then, it is hoped that our current study has provided the clinical basis for the potential utility of biomarkers, especially C-reactive protein, as surrogate indicators of treatment response and lays the foundation for additional research on understanding host immunes response during IA.

The Y238X Stop Codon Polymorphism in the Human β-Glucan Receptor Dectin-1 and Susceptibility to Invasive Aspergillosis

Dectin-1 is the major receptor for fungal β-glucans on myeloid cells. We investigated whether defective Dectin-1 receptor function, because of the early stop codon polymorphism Y238X, enhances susceptibility to invasive aspergillosis (IA) in at-risk patients. Association of Dectin-1 Y238X polymorphism with occurrence and clinical course of IA was evaluated in 71 patients who developed IA post hematopoietic stem cell transplantation (HSCT) and in another 21 non-HSCT patients with IA. The control group consisted of 108 patients who underwent HSCT. Functional studies were performed to investigate consequences of the Y238X Dectin-1 polymorphism. The Y238X allele frequency was higher in non-HSCT patients with IA (19.0% vs 6.9%-7.7%; P < .05). Heterozygosity for Y238X polymorphism in HSCT recipients showed a trend toward IA susceptibility (odds ratio, 1.79; 95% CI, .77-4.19; P = .17) but did not influence clinical course of IA. Functional assays revealed that although peripheral blood mononuclear cells with defective Dectin-1 function due to Y238X responded less efficiently to Aspergillus, corresponding macrophages showed adequate response to Aspergillus. Dectin-1 Y238X heterozygosity has a limited influence on susceptibility to IA and may be important in susceptible non-HSCT patients. This is partly attributable to redundancy inherent in the innate immune system. Larger studies are needed to confirm these findings.

Invasive pulmonary aspergillosis due to Aspergillus terreus: value of DNA, galactomannan and (1→3)-β-d-glucan detection in serum samples as an adjunct to diagnosis

A case of invasive pulmonary aspergillosis caused by Aspergillus terreus is described. The diagnosis was based on demonstration of branched septate hyphae in a sputum specimen and isolation of the fungus in culture. The diagnosis was further supported by detection of A. terreus-specific DNA, galactomannan (GM) and (1→3)-β-D-glucan (BDG) in consecutive serum specimens. The patient was treated for about 10 weeks with voriconazole. The decreasing levels of GM and BDG in serum samples were accompanied by symptomatic and radiological improvement. The report highlights the value of surrogate markers in the diagnosis and for monitoring the course of invasive aspergillosis during therapy.

Purulent Pericardial Effusion in a 14-Year-Old Girl

Purulent Pericardial Effusion in a 14-Year-Old Girl

Invasive aspergillosis and allogeneic hematopoietic stem cell transplantation in children: a 15‐year experience

Despite progress in diagnosis and treatment, invasive aspergillosis (IA) remains a principal cause of mortality due to infection after allogeneic hematopoietic stem cell transplantation (AHSCT). In order to clarify the course of IA among children receiving an AHSCT before the advent of new drugs such as voriconazole or caspofungin, we retrospectively reviewed the medical records of all proven and probable IA between January 1986 and December 2000. 1) Ten children developed IA after AHSCT, mostly long after transplantation. Overall incidence was 2.7%. Seven of those children experienced 1 or more complications after AHSCT and before IA. Mortality was 90% with a median survival of 23 days (2-90). 2) Five children underwent AHSCT after a previous episode of IA. All patients were treated with systemic antifungal therapy combined with surgery. Median time between IA and AHSCT was 110 days (73-370). Two children were diagnosed with IA relapse after transplantation. One child was cured while the other died of IA and AHSCT complications. AHSCT could be considered even in the setting of previous IA, but established strategies implementing newer less toxic antifungal agents as treatment or prophylaxis in high-risk patients are needed.

A Murine Model of Invasive Aspergillosis: Variable Benefit of Interferon-Gamma Administration under In Vitro and In Vivo Conditions

Interferon-gamma modulates host defense in a number of infectious diseases. Previous studies have shown that systemic administration of interferon-gamma (IFN-gamma) can enhance survival in experimental invasive aspergillosis (IA). Using a novel model of murine IA that is characterized by primary pulmonary infection, we investigated the role of IFN-gamma in the phagocytosis and killing of Aspergillus fumigatus by murine neutrophils and pulmonary alveolar macrophages in vitro and the impact of systemic and regional administration of IFN-gamma on the course of IA in glucocorticoid-treated mice. In vitro, IFN-gamma significantly enhanced phagocytosis and killing function of both neutrophils and alveolar macrophages from normal animals, but not cortisone-treated animals. In vivo, intravenous administration of IFN-gamma did not improve phagocyte recruitment, in vivo killing, or mortality from IA. Regional (intranasal) administration of IFN-gamma to the lungs enhanced recruitment of phagocytic cells to the lungs and improved in vivo killing, but did not alter (and actually worsened) mortality from IA. The in vitro and in vivo effects of IFN-gamma in IA are contingent on many variables, including the route of administration and the specific pathogenesis of infection.

Galactomannan and advances in fungal diagnostics

Purpose of review Invasive fungal infections remain a predominant cause of morbidity and mortality in the immunocompromised population. Fungal diagnostics, namely the Platelia Aspergillus enzyme immunoassay (Bio-Rad, Redmond, WA) and the Fungitell 1→3-β-D-glucan assay (Associates of Cape Cod, East Falmouth, MA), are now commercially available to assist clinicians in the early diagnosis of fungal infections. This review highlights the performance characteristics of these tests and introduces the reader to other advances in fungal diagnostics. Recent findings Early diagnosis of invasive aspergillosis in patients with leukemia and hematopoietic stem cell transplants is facilitated by the use of the Platelia Aspergillus enzyme immunoassay. This test may also be useful for monitoring the course of invasive aspergillosis. The Fungitell1→3-β-D-glucan assay can be used for early diagnosis of fungal infections due to multiple different organisms, including certain non-Aspergillus mould infections and invasive candidiasis. Physicians utilizing these fungal diagnostic tests should be aware of important issues regarding test performance. Summary Advances in fungal diagnostics are an important addition to the diagnosis and management of invasive fungal infections. Additional tests such as fungal polymerase chain reaction are on the horizon and are expected to be helpful adjuncts to the diagnosis and management of invasive fungal infections.

Fungal CNS infections in patients with hematologic malignancy

Various fungal agents can cause CNS infections. CNS fungal infections may present as a mass (i.e., brain abscess) typically in the course of aspergillosis or zygomycosis, or may primarily involve the meninges (i.e., meningitis), as can be observed in patients with candidiasis or cryptococcosis. Most commonly, fungal brain abscesses are due to aspergillosis. CNS aspergillosis is observed particularly in acute leukemia and allogeneic hemopoietic stem cell transplantation patients. Usually, aspergillosis is localized in the lungs and secondarily spreads to the brain; only in few cases does it develop as solitary localization of CNS. In these conditions, diagnosis is very difficult because signs and symptoms can be completely aspecific. Diagnosis can often be performed only through aggressive procedures (i.e., stereotactic puncture). Zygomycetes are the second most frequent cause of brain abscesses. CNS involvement is higher than in the course of invasive aspergillosis, and this fungal complication is also characterized by a high mortality rate. In vitro and in vivo studies demonstrated that only posaconazole and lipid formulations of amphotericin B present some possibility of success in the treatment of zygomycosis, but the pharmacologic approach should always be associated with surgery. Among molds, other agents (i.e., Fusarium and Scedosporium) may also be responsible for fungal abscess. More rarely during the course of a hematologic malignancy, a meningeal candidiasis or cryptococcosis may be observed. This review mainly focuses on the epidemiology, clinical manifestations, diagnosis and management strategies of all cases of CNS fungal infections in hematologic patients.

Application of Nucleic Acid Sequence-Based Amplification for Diagnosis of and Monitoring the Clinical Course of Invasive Aspergillosis in Patients with Hematologic Diseases

We evaluated nucleic acid sequence-based amplification (NASBA) and a galactomannan enzyme immunosorbent assay (GM-EIA) for the diagnosis of invasive aspergillosis (IA) in neutropenic febrile patients and for monitoring of its clinical course and outcome. Blood samples were collected twice per week from 128 patients with hematologic diseases during periods of neutropenic fever after undergoing chemotherapy or hematopoietic stem cell transplantation. A total of 448 blood samples were tested. There were 14 patients with IA (2 patients with proven IA and 12 with probable IA). The median index of the initial NASBA in the IA group was more than 10-fold higher than that in the non-IA group. Galactomannan antigenemia (index, >0.5) was detected with a sensitivity of 86%. In receiver-operator characteristic analysis, the cutoff index of NASBA for the presumptive diagnosis of IA was determined to be 5.0. Combination of these 2 parameters (either a GM-EIA index of >or=0.5 or a NASBA index of >or=5.0) improved the sensitivity of diagnosis to 100%. There was a close relationship between patient outcome and the kinetics of NASBA values: failure of negative conversion during treatment resulted in death in almost all cases. If either GM-EIA or NASBA results suggest IA, the diagnostic yield for IA could be improved, and NASBA could be a useful marker for predicting the clinical course and outcome of treatment.

Diagnosis of fungal infection: new technologies for the mycology laboratory

The dramatic increase in nosocomial invasive mycoses over the past two decades has led to increased interest in the area of antifungal development. Unfortunately, the infusion of new diagnostic technology into the clinical mycology laboratory has lagged behind. Although newer, automated, continuous-monitoring blood culture systems are as sensitive as the older, manual "gold standard" system, the recovery of fungi from blood, as well as other clinical specimens, remains an insensitive marker for invasive fungal infection. Antigen assays for the rapid diagnosis of invasive fungal infections are in development, and galactomannan and glucan are two such promising antigens. Glucan may be present in the blood of patients with infection secondary to a wide variety of fungal pathogens, including Candida, Aspergillus, Fusarium, Saccharomyces, Trichosporon and Acremonium species. Early data suggest galactomannan may be present in the blood in detectable levels very early in the course of invasive aspergillosis. The galactomannan assay currently undergoing evaluations may actually be positive prior to the clinical suspicion for infection and may be useful in monitoring therapeutic response as well; however, the etiology of false-positive results following cytotoxic chemotherapy still has to be elucidated. PCR assays are also being developed in the research laboratory, however, the PCR assays will require a significant amount of adaptation and validation before they are ready for clinical care. Well-planned studies to evaluate the performance characteristics as well as appropriate clinical and cost-effective application of these new tests are needed.

The significance of secretory and structure-associated proteases of Aspergillus fumigatus for the pathogenesis of invasive aspergillosis

In the course of invasive aspergillosis, Aspergillus fumigatus is capable of penetrating any tissue of the host. Secretory proteinases of the fungus might facilitate the hyphae to grow through fibrillar proteins like elastin and collagen. However, using systemic infection models, no significantly reduced virulence could be shown with fungal mutants deficient for all known secretory proteinases. Thus, secretory proteinases might be of minor relevance for the pathogenesis of invasive aspergillosis. In addition, microscopic examination of aspergilli penetrating vessel walls did not reveal obvious lysis of wall proteins, thus emphasizing a mechanical disruption of fibrillar proteins by the growing hyphae. However, a strictly localized proteolysis at the tips of growing hyphae caused by wall associated proteinases might be involved. Candidates for such a mechanism are the activities of aspartic and serine proteinases which we have discovered in the cell wall fraction of A. fumigatus.