Use and limits of (1-3)-ß-d-glucan assay (Fungitell), compared to galactomannan determination (Platelia Aspergillus), for diagnosis of invasive aspergillosis.

This study was undertaken to examine the performance of the Fungitell ß-glucan (BG) assay, to compare it with that of the galactomannan (GM) test for the diagnosis of invasive aspergillosis (IA) in patients with hematological malignancies, and to examine the rates of false-positive BG and GM test results due to ß-lactam antibiotics among sera of patients with Gram-positive or Gram-negative bacteremia and selected sera with false-positive results from the GM test. Serum samples from 105 patients with proven (n = 14) or probable (n = 91) IA, 97 hematology patients at risk for invasive fungal infections, 50 healthy blood donors, and 60 patients with bacteremia were used to study the sensitivities and specificities of the assays. The GM test was more specific than the BG assay (97% versus 82%, respectively; P = 0.0001) and the BG assay was more sensitive than the GM test (81% versus 49%, respectively; P < 0.0001) for IA diagnosis. The study of 49 separate batches of ß-lactam antibiotics showed high and very similar rates of false-positive results for the GM and BG assays (29 and 33%, respectively; P = 0.82) but with an almost complete lack of concordance between the 2 assays. For patients with bacteremia, the rate of false-positive results was much higher with the BG test than with the GM test (37% versus 2%, respectively; P < 0.0001), with no significant difference between Gram-positive and Gram-negative bacteremia. In conclusion, the BG test may be useful for the diagnosis of IA because of its high sensitivity in comparison with the GM test, but the overall benefit of this assay remains limited because of its inadequate specificity and its cost.

Bayesian development of a dose-response model for Aspergillus fumigatus and invasive aspergillosis.

Invasive aspergillosis (IA) is a major cause of mortality in immunocompromized hosts, most often consecutive to the inhalation of spores of Aspergillus. However, the relationship between Aspergillus concentration in the air and probability of IA is not quantitatively known. In this study, this relationship was examined in a murine model of IA. Immunosuppressed Balb/c mice were exposed for 60 minutes at day 0 to an aerosol of A. fumigatus spores (Af293 strain). At day 10, IA was assessed in mice by quantitative culture of the lungs and galactomannan dosage. Fifteen separate nebulizations with varying spore concentrations were performed. Rates of IA ranged from 0% to 100% according to spore concentrations. The dose-response relationship between probability of infection and spore exposure was approximated using the exponential model and the more flexible beta-Poisson model. Prior distributions of the parameters of the models were proposed then updated with data in a Bayesian framework. Both models yielded close median dose-responses of the posterior distributions for the main parameter of the model, but with different dispersions, either when the exposure dose was the concentration in the nebulized suspension or was the estimated quantity of spores inhaled by a mouse during the experiment. The median quantity of inhaled spores that infected 50% of mice was estimated at 1.8 × 10(4) and 3.2 × 10(4) viable spores in the exponential and beta-Poisson models, respectively. This study provides dose-response parameters for quantitative assessment of the relationship between airborne exposure to the reference A. fumigatus strain and probability of IA in immunocompromized hosts.

Efficacy of liposomal amphotericin B for prophylaxis of acute or reactivation models of invasive pulmonary aspergillosis.

The efficacy of antifungal prophylaxis for prevention of invasive aspergillosis (IA) may depend on whether IA results from recent inhalation of spores or reactivation of latent colonisation. Compare the efficacy of liposomal amphotericin B (LAmB) for prophylaxis in acute and reactivation models of IA. In the acute model, mice immunosuppressed from day 0 were challenged at day 3 with an aerosol of Aspergillus fumigatus. LAmB (15 mg kg(-1) ) was administered at day 0 or at challenge. In the reactivation model, naïve mice exposed to A. fumigatus remained untreated until clearance of spores from the lungs, then immunosuppressed to induce reactivation. A single LAmB dose was administered at start of immunosuppression. In the acute model, a single administration of LAmB at start of immunosuppression was not effective, but an additional administration resulted in a significant decrease in lung fungal burden (P < 0.05 vs. controls). A significant prophylactic efficacy was observed when LAmB was administered once at challenge (P < 0.01). In the reactivation model, a single LAmB administration at start of immunosuppression significantly reduced both reactivation rate and fungal burden vs. controls (P < 0.01). Our results show that the conditions under which IA develop and timing of administration of LAmB were determinant variables for prophylactic efficacy.

Persistent poor long-term prognosis of allogeneic hematopoietic stem cell transplant recipients surviving invasive aspergillosis.

BACKGROUND: Voriconazole treatment increases early survival of allogeneic hematopoietic stem cell transplant recipients with invasive aspergillosis. We investigated whether this survival advantage translates into an increased long-term survival. DESIGN AND METHODS: This retrospective study involved all patients with an invasive aspergillosis diagnosis transplanted between September 1997 and December 2008, at the Saint-Louis Hospital, Paris, France. The primary end point was survival up to 36 months. Survival analysis before and after 12 weeks, as well as cumulative incidence analysis in a competing risk framework, were used to assess the effect of voriconazole treatment and other factors on mortality. RESULTS: Among 87 patients, 42 received first-line voriconazole and 45 received another antifungal agent. Median survival time was 2.6 months and survival rate at 36 months was 18%. Overall, there was a significant difference in the survival rates of the two groups. Specifically, there was a dramatic difference in survival rates up to ten months post-aspergillosis diagnosis but no significant difference after this time. Over the first 36 months as a whole, no significant difference in survival rate was observed between the two groups. First-line voriconazole significantly reduced aspergillosis-attributable mortality. However, first-line voriconazole patients experienced a significantly higher probability of death from a non-aspergillosis-attributable cause. CONCLUSIONS: Although the prognosis for invasive aspergillosis after stem cell transplantation has dramatically improved with the use of voriconazole, this major advance in care does not translate into increased long-term survival for these severely immunocompromised patients.

Prospective evaluation of clinical and biological markers to predict the outcome of invasive pulmonary aspergillosis in hematological patients.

Early evaluation of treatment efficacy in invasive aspergillosis (IA), a leading cause of morbidity and mortality in hematological patients, remains a challenge. We conducted a prospective study to evaluate the performance of different markers in predicting the outcome of patients with IA. Both clinical and biological criteria were assessed 7, 14, 21, and 45 days after inclusion in the study, and mortality was assessed at day 60. The association between baseline data and their evolution and the day 45 response to treatment was analyzed. A total of 57 patients (4 with proven, 44 with probable, and 9 with possible aspergillosis according to the revised EORTC/MSG [European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and National Institute of Allergy and Infectious Diseases Mycoses Study Group] definitions) were included. At day 45, 30 patients (53%) were determined to be responders, 25 (44%) were nonresponders, and 2 were not able to be evaluated. Twenty patients died within the 60 days of follow-up. We found that a poor day 45 outcome was associated with patients who had high baseline serum galactomannan (GM) antigen levels and those receiving steroids at the time of IA. A consistently negative serum GM index was associated with a good outcome, and the day 14 clinical evaluation was predictive of the day 45 outcome. No association was found between Aspergillus antibodies or DNA detection and patients' outcome. We conclude that the GM index value at diagnosis of IA, GM index kinetics, and clinical evaluation at day 14 are good markers for predicting the outcome of patients with IA and should be taken into account for adapting antifungal treatment.

The strategy for the diagnosis of invasive pulmonary aspergillosis should depend on both the underlying condition and the leukocyte count of patients with hematologic malignancies.

The identification of the causative organism in invasive pulmonary aspergillosis (IPA) is recommended. We investigated whether a mycologic diagnostic strategy could be optimized based on patient characteristics. Fifty-five patients were enrolled in a prospective study. The presence of Aspergillus in respiratory samples occurred more frequently in non-acute leukemia (AL) patients than in AL patients (P = .0003), and in patients with leukocyte counts more than 100/mm(3) (P = .002). In a logistic regression model, these 2 factors appeared to be independent, with an adjusted odds ratio of 7.14 (95% confidence interval, 1.40-36.5) for non-AL patients and an adjusted odds ratio of 6.97 (95% confidence interval, 1.33-36.5) for patients with leukocyte counts more than 100/mm(3). A positive mycologic result was also more frequent among patients with lung CT scan signs of airway-invasive disease than among other patients (P = .043). Airway-invasive signs were more frequent among non-AL patients (P = .049), whereas angioinvasive disease was more frequent among both AL patients (P = .01) and patients with leukocyte counts less than 100/mm(3) (P = .001). A concomitant pulmonary infection was identified more frequently among non-AL patients (P = .005 vs allogeneic hematopoietic stem cell transplant and P = .048 vs others). Our results suggest that different strategies for diagnosing IPA should be considered based on the underlying condition.

Contribution of galactomannan antigen detection in BAL to the diagnosis of invasive pulmonary aspergillosis in patients with hematologic malignancies.

BACKGROUND: Invasive pulmonary aspergillosis (IPA) is difficult to diagnose. The detection of galactomannan (GM) in serum samples is useful for diagnosing IPA. A positive test for GM antigen in BAL has also been proposed as a criterion of IPA, although it has not been fully validated. The aim of our study was to evaluate the contribution of GM antigen detection in BAL to the diagnosis of IPA in hematologic patients. METHODS: One hundred one consecutive patients treated for hematologic malignancy were explored by bronchoscopy and BAL for new pulmonary infiltrates. Both BAL fluid and serum samples were evaluated for GM using an enzyme-linked immunosorbent assay test, with an optical density index >or= 0.5 considered positive. Respiratory samples were also examined for the presence of fungi. RESULTS: IPA was diagnosed in 33 patients according to European Organization for Research and Treatment of Cancer and Mycoses Study Group consensus group criteria (six proven, 23 probable, four possible). Nineteen of these 33 patients had a positive BAL GM test, whereas three patients without IPA had false-positive results. GM detection in BAL had a sensitivity of 57.6% (95% CI, 40.8%-72.8%) and a specificity of 95.6% (95% CI, 87.8%-98.5%). Among the 19 patients with IPA whose BAL was positive for GM, 15 also had a positive serum GM test. In 11 of these 19 patients, Aspergillus was identified in the respiratory samples. CONCLUSION: Detection of GM in BAL is complementary of serum GM testing and mycologic evaluation of the respiratory samples for the diagnosis of IPA. Positive GM BAL was the sole microbiologic criterion in two of 33 patients studied.

Contribution of the (1-->3)-beta-D-glucan assay for diagnosis of invasive fungal infections.

Diagnosis of invasive fungal infection (IFI) remains a challenge. A retrospective study was performed on 279 patients at three French university hospitals to evaluate the performance of the (1-->3)-beta-D-glucan assay (BG assay; Fungitell; Associates of Cape Cod, Inc.) for the diagnosis of IFI. The results of one serum per subject were analyzed for 117 patients who had probable or proven IFI according to the European Organization for Research and Treatment of Cancer criteria (70 invasive pulmonary aspergilloses [IPA], 27 fungal bloodstream infections, and 20 Pneumocystis jiroveci pneumonias), 40 blood donors, and 122 patients who were hospitalized in hematology wards or intensive care units and were at risk for IFI but in whom IFI had not been diagnosed. For the overall IFI diagnosis, the BG assay had 77.8% sensitivity and specificities of 92.5 and 70.5% for blood donors and patients at risk, respectively. The assay was positive in 48 patients with IPA (68%), in 23 with bloodstream infections (85.2%), and in all who had P. jiroveci pneumonias (100%), and the false-positive rate varied depending on the controls used. It allowed a higher rate of detection among IPA patients compared to the galactomannan enzyme-linked immunosorbent assay (ELISA) (48 versus 39 patients, respectively) and among candidemia patients compared to the mannan ELISA (20 versus 11 patients, respectively). This assay therefore appears to be useful in the diagnosis of IFI, particularly for serum analysis of pneumocystosis pneumonia patients, but further studies are needed to evaluate false-positive rates and its future role in IFI diagnosis.

Recombinant antigens as diagnostic markers for aspergillosis.

Eight recombinant proteins and purified galactomannan of Aspergillus fumigatus were tested by enzyme-linked immunosorbent assay to quantify the anti-Aspergillus antibodies in sera of patients with aspergilloma, allergic bronchopulmonary aspergillosis (ABPA), and invasive aspergillosis (IA). In spite of the variability observed in the immune responses of individual patients, quantification of the antibody titers against the 18-kDa ribonuclease (RNU), the 360-kDa catalase (CAT), and the 88-kDa dipeptidylpeptidase V (DPPV) was useful for the diagnosis of aspergilloma and ABPA. Differential diagnosis of ABPA was even possible among cystic fibrosis as well as noncystic fibrosis patients. In the group of immunocompromised patients with IA, no antibody response was mounted in response to the Aspergillus infection in any of the patients. Interestingly, about half of the patients with proven IA came to the hospital with high titers of anti-Aspergillus antibodies, suggesting that they were infected upon entry to the hospital. These results suggest that recombinant RNU, CAT, and DPPV have a great potential in the serodiagnosis of all forms of aspergillosis in the immunocompromised and immunocompetent patient.

Occurrence and kinetics of false-positive Aspergillus galactomannan test results following treatment with beta-lactam antibiotics in patients with hematological disorders.

Several reports have described a high rate of false-positive Aspergillus galactomannan (GM) test results for patients treated with piperacillin-tazobactam. In this retrospective study, we first examined the relationships between intravenous administration of three beta-lactam antibiotics and the occurrence of false-positive GM test results in hematology patients. We then estimated the kinetics of clearance of GM after the cessation of treatment. Sequential serum samples from 69 patients that had received beta-lactams were analyzed by using a Platelia Aspergillus test. A significant association was found between GM positivity (>/=0.5) and the administration of beta-lactams (P < 0.0001). The direct role of beta-lactams in patients' serum positivity was assessed by testing 39 batches of beta-lactams, of which 27 were positive for GM. None of the latter were positive according to a fungus- and Aspergillus-specific PCR. The kinetics of the decrease of GM was analyzed on sequential serum samples obtained after treatment. By use of a nonlinear regression model, the average time to negative antigen was assessed to be 5.5 days (95% confidence interval [CI], 4.1 to [7.0]), with a half-life of elimination of GM of 2.4 days (95% CI, 1.8 to 3.0). This study confirms that the administration of beta-lactams containing GM is responsible for false-positive diagnostic results, even up to 5 days after the cessation of treatment.

Development and evaluation of a Western blot kit for diagnosis of schistosomiasis.

We evaluated the performance of Western blot (WB) analysis using commercially available antigen strips and compared the results with those of indirect hemagglutination (IHA) and indirect immunofluorescence (IFAT) for the serodiagnosis of human schistosomiasis. The antigen preparation was a crude extract of Schistosoma mansoni. The WB profile characteristics of schistosomiasis were characterized by comparing the results for 58 serum samples from patients with parasitologically proven S. mansoni (n = 12) and S. haematobium (n = 46) infections and 37 individuals with probable cases of schistosomiasis but with only positive serology results. The specificity of WB analysis was assessed by testing 12 serum samples from healthy subjects, 67 serum samples from patients with other proven helminthic and protozoan infections, and 16 serum samples from patients with autoantibodies. Six immunodominant bands (65, 70, 80, 95, 110, and 120 kDa) were revealed with sera from patients with schistosomiasis. The presence of three or more bands in the range 65 to 120 kDa, with the exception of the 100-kDa band, was considered diagnostic for Schistosoma infection and had a specificity of 100% in our series. In patients with proven schistosomiasis, the sensitivity of WB analysis was 84.5%, whereas those of IFAT and IHA were 65.5 and 72.9%, respectively. For serologically proven cases, the sensitivity of WB analysis was 97.3%. The overall sensitivity and specificity for both groups of patients were 89.5 and 100%, respectively, with positive and negative predictive values of 100 and 91.3%, respectively. We conclude that WB analysis is a useful technique for the immunological diagnosis of schistosomiasis.

Invasive aspergillosis in allogeneic stem cell transplant recipients: increasing antigenemia is associated with progressive disease.

The kinetics of serum Aspergillus galactomannan, as determined by enzyme-linked immunosorbent assay, was examined in 37 allogeneic stem cell transplant (SCT) recipients treated for invasive aspergillosis (IA). Fifty-eight periods of response ("response episodes") were evaluated. There were 42 response episodes that were considered "treatment failures" and 16 that were considered "good" (that is, complete or partial) responses. At baseline (the first day of each new response episode), the patients who experienced treatment failure and those who had good responses did not differ significantly with regard to median galactomannan index (GMI) value. Thereafter, GMI values significantly increased in the treatment failure group, whereas no significant changes were observed in the good response group (P=.002). An increase in the GMI value of 1.0 over the baseline value during the first week of observation was predictive of treatment failure with a sensitivity of 44%, a specificity of 87%, and a positive predictive value of 94%. We conclude that serial determination of serum GMI values is a useful tool for assessing prognosis of IA in allogeneic SCT recipients during treatment.