This study was undertaken to examine the performance of the Fungitell ?-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 ?-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 ?-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.
The BioPlex 2200 automated analyzer (Bio-Rad Laboratories, Hercules, CA) is a recently developed multiplex analyzer that enables the detection of anti-Toxoplasma, -rubella, and -cytomegalovirus antibodies in the same assay. The aim of this study was to compare this new technology (using the BioPlex 2200 ToRC IgG/IgM kit) in critical cases of serodiagnosis of toxoplasmosis (acute, chronic, or congenital infections and cases with discrepant results) to the technologies used in our routine practice, i.e., the Platelia IgG/IgM enzyme-linked immunosorbent assays (ELISAs) (Bio-Rad Laboratories) and the Toxo-Screen direct agglutination assay (bioMérieux, Lyon, France). Overall, most cases of false-positive/negative results obtained with the Platelia IgG or Toxo-Screen assay were corrected by the BioPlex 2200 ToRC IgG (87.5%). Furthermore, the analysis of 35 sequences of sera showed a trend toward a more rapid decrease of IgM titers by BioPlex 2200 than by Platelia. These results for IgM detection can be explained by a weaker detection of residual IgM. Indeed, among 23 serum samples from patients with probable past infection with long-lasting IgM (Platelia M positive and IgG avidity index, ?0.5), the BioPlex 2200 Toxoplasma IgM assay was positive for only 11 serum samples. In our panel of critical cases comprising 156 serum and 6 cord blood samples from 103 patients with acute, chronic, or congenital infection, the BioPlex 2200 IgG assay was a sensitive (97.8%) and specific (91.3%) method for IgG detection. The high specificity (97.4%) of IgM detection combined with the shorter kinetics of IgM titers may considerably reduce the number of residual IgM detections, thus yielding more precise diagnoses of acute infections.
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.
To better understand the role of immunocompetent hosts in the diffusion of Pneumocystis in the environment, airborne shedding of Pneumocystis carinii in the surrounding air of experimentally infected Sprague Dawley rats was quantified by means of a real-time PCR assay, in parallel with the kinetics of P. carinii loads in lungs and specific serum antibody titres. Pneumocystis-free Sprague Dawley rats were intratracheally inoculated at day 0 (d0) and then followed for 60 days. P. carinii DNA was detected in lungs until d29 in two separate experiments and thereafter remained undetectable. A transient air excretion of Pneumocystis DNA was observed between d14 and d22 in the first experiment and between d9 and d19 in the second experiment; it was related to the peak of infection in lungs. IgM and IgG anti-P. carinii antibody increase preceded clearance of P. carinii in the lungs and cessation of airborne excretion. In rats receiving a second challenge 3 months after the first inoculation, Pneumocystis was only detected at a low level in the lungs of 2 of 3 rats at d2 post challenge and was never detected in air samples. Anti-Pneumocystis antibody determinations showed a typical secondary IgG antibody response. This study provides the first direct evidence that immunocompetent hosts can excrete Pneumocystis following a primary acquired infection. Lung infection was apparently controlled by the immune response since fungal burdens decreased to become undetectable as specific antibodies reached high titres in serum. This immune response was apparently protective against reinfection 3 months later.
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 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.
Although indoor air has wide ranging effects on human health, the effects of environmental, chemical, and biological pollutants on the respiratory system are not fully understood. In order to clarify the health effects of airborne pollutant exposure, it would appear that toxicological evidence is needed to complement epidemiological observations to support by providing biological plausibility. The aim of this study is to manage air-liquid successive exposures to different pollutants such as a chemical pollutant (formaldehyde--FA), and a biological contaminant (Aspergillus fumigatus--Asp) using our in vitro model. Human alveolar cells (A549) were exposed at the air-liquid interface in an exposure module, firstly to an environmental level of FA (50 ?g/m³) (or air) for 30 min, and 14 h later to Asp (7×10? spores/m³) (or air) for 30 min. After 10 h post-incubation, cellular viability was assessed. Inflammation biomarkers (IL-8, MCP-1) were assayed by ELISA and by RT-PCR. Whatever the conditions, no cytotoxic effect was observed. FA followed by air exposure did not induce modification of production and expression of cytokines, confirming results with a unique FA exposure. Air followed by Asp exposure tended to induce IL-8 expression whereas IL-8 production tended to increase after FA and Asp exposure compared to FA and air exposure. The reaction of cells to sequential exposure to FA and Asp was moderate. These results show the feasibility of our model for sequential exposures to different types of environmental pollutants, allowing using it for preliminary assessment of cellular activity modification induced by airborne contaminants.
To better understand the diffusion of Pneumocystis in the environment, airborne shedding of Pneumocystis carinii in the surrounding air of experimentally infected rats was quantified by means of a real-time polymerase chain reaction assay, in parallel with the kinetics of P. carinii loads in their lungs. P. carinii DNA was detected in the air 1 week after infection and increased until 4-5 weeks after infection before stabilizing. A significant correlation was shown between lung burdens and the corresponding airborne levels, suggesting the possibility of estimating the fungal lung involvement through quantification of Pneumocystis in the exhaled air.
Controlling airborne contamination is of major importance in burn units because of the high susceptibility of burned patients to infections and the unique environmental conditions that can accentuate the infection risk. In particular the required elevated temperatures in the patient room can create thermal convection flows which can transport airborne contaminates throughout the unit. In order to estimate this risk and optimize the design of an intensive care room intended to host severely burned patients, we have relied on a computational fluid dynamic methodology (CFD).
Airborne transmission of Pneumocystis has been demonstrated in animal models and is highly probable in humans. However, information concerning burdens of Pneumocystis jirovecii (human-derived Pneumocystis) in exhaled air from infected patients is lacking. Our objective is to evaluate P. jirovecii air diffusion in patients with Pneumocystis pneumonia.
Disseminated microsporidiosis is a life-threatening opportunistic infection. Here, we report about a previously undescribed genovar of Encephalitozoon cuniculi causing disseminated infection in a non-HIV-infected renal transplant recipient. Disseminated microsporidiosis must be considered in the differential diagnosis of chronic fever in renal allograft recipients, even those without urinary symptoms.
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.
The use of combination antiretroviral therapy (cART) has dramatically reduced the prevalence of opportunistic infections, however data on the prevalence of intestinal parasitic infections in HIV-infected patients with low CD4 cell counts in the cART era are scarce.
Related JoVE Video
Journal of Visualized Experiments
What is Visualize?
JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.
How does it work?
We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.
Video X seems to be unrelated to Abstract Y...
In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.