High resolution genotyping of clinical Aspergillus flavus isolates from India using microsatellites.

Background: Worldwide, Aspergillus flavus is the second leading cause of allergic, invasive and colonizing fungal diseases in humans. However, it is the most common species causing fungal rhinosinusitis and eye infections in tropical countries. Despite the growing challenges due to A.

[Melioidosis: the importance of a detailed medical history, including recent travels].

A 70-year-old woman was admitted to hospital with fever and signs of bronchopneumonia following a recent visit to Southeast Asia. She was diagnosed with melioidosis and treated with ceftazidime i.v.

Utility of CSP typing to sub-type clinical Aspergillus fumigatus isolates and proposal for a new CSP type nomenclature.

CSP typing is a newly developed sub-typing strategy that employs comparative DNA sequence analysis of the 12-mer tandem repeat region of the AFUA_3G08890 gene. In order to allow standardization of analysis and exchange of results between laboratories, we propose a new nomenclature for individual CSP repeats as well as for CSP types. A collection of 209 clinical isolates of Aspergillus fumigatus recovered from various hospitals throughout The Netherlands was analyzed by using CSP typing and this newly proposed nomenclature.

In vitro activities at pH 5.0 and pH 7.0 and in vivo efficacy of flucytosine against Aspergillus fumigatus.

The antifungal agent flucytosine was found to be active in vitro against Aspergillus fumigatus isolates when the MIC was determined at pH 5.0 instead of pH 7.0.

Utility of a microsatellite assay for identifying clonally related outbreak isolates of Aspergillus fumigatus.

A microsatellite assay based on short tandem repeats (STRAf) has been recently described as a discriminatory, high throughput assay for fingerprinting Aspergillus fumigatus isolates. However, the STRAf assay has not been tested for its utility in outbreak settings where it is critical to distinguish clonal clusters from genetically unrelated genotypes. In the present study, employing a panel of epidemiologically linked A.

Retrotransposon insertion-site context (RISC) typing: a novel typing method for Aspergillus fumigatus and a convenient PCR alternative to restriction fragment length polymorphism analysis.

Retrotransposon(-like) sequences in Aspergillus fumigatus have been used as typing targets through restriction fragment length polymorphism (RFLP)/Southern blotting approaches. Differences in fingerprints between unrelated isolates are the result of variations in copy-number and differences in the regions flanking the retrotransposon elements. Here, we present retrotransposon insertion-site context (RISC) typing as a novel and convenient PCR-based typing alternative to the RFLP approach.

Activity and post antifungal effect of chlorpromazine and trifluopherazine against Aspergillus, Scedosporium and zygomycetes.

The phenothiazine compounds chlorpromazine and trifluopherazine are antipsychotic agents that exhibit antimicrobial activity against bacteria, some protozoa and yeasts. Data of activity against filamentous fungi are lacking. The in vitro activity and postantifungal effect (PAFE) of chlorpromazine and trifluopherazine was determined against Aspergillus species, zygomycetes and Scedosporium species.

Comparison of two highly discriminatory molecular fingerprinting assays for analysis of multiple Aspergillus fumigatus isolates from patients with invasive aspergillosis.

Two highly discriminatory fingerprinting assays, short tandem repeat typing and amplified fragment length polymorphism (AFLP), were compared to determine the genetic relatedness between 55 isolates of Aspergillus fumigatus obtained from 15 different patients suffering from proven invasive aspergillosis. Both techniques showed that interpatient isolates belonged to different genotypes and that intrapatient isolates from deep sites were all of the same genotype. By contrast, multiple genotypes were found among isolates originating from respiratory samples.

Microsatellite based typing of Aspergillus fumigatus: strengths, pitfalls and solutions.

Microsatellites, or short tandem repeats (STR's), are popular tools to discriminate between microbial isolates. Here, we report on the robustness of a microsatellite panel for discrimination of Aspergillus fumigatus isolates. Two major PCR artefacts (stutter peaks and minus-A peaks) can complicate correct interpretation of STR data.

Clostridium cadaveris bacteraemia: two cases and review.

Clostridium cadaveris is a strict anaerobic Gram-positive rod that is the most prominent bacterium during the decay of dead bodies. We present 2 rare cases of bacteraemia with C. cadaveris.

Use of a novel panel of nine short tandem repeats for exact and high-resolution fingerprinting of Aspergillus fumigatus isolates.

Here we describe a new panel of short tandem repeats (STRs) for a novel exact typing assay that can be used to discriminate between Aspergillus fumigatus isolates. A total of nine STR markers were selected from available genomic A. fumigatus sequences and were divided into three multicolor multiplex PCRs.

Assessing in vitro combinations of antifungal drugs against yeasts and filamentous fungi: comparison of different drug interaction models.

Non-parametric and parametric approaches of two competing zero-interaction theories--the Loewe additivity and the Bliss independence - were evaluated for analyzing the in vitro interactions of various antifungal drugs. Fifty-one data sets, derived from three drug combinations, tested in triplicate against 17 clinical yeast and mold isolates with a two-dimensional checkerboard microdilution technique, were selected to span from strong synergy to strong antagonism. These were analyzed with the standard FIC index model and modern concentration-effect response surface models: the fully parametric model developed by Greco et al.

Pericarditis as complication of appendicitis.

Pericarditis as a complication of appendicitis is a rare event. In a 25-year period we encountered two pediatric cases with this severe complication due to (a)typical presentation of appendicitis resulting in small bowel obstruction, intraabdominal abscesses, constrictive pericarditis, and purulent pericarditis.

Activity of posaconazole in treatment of experimental disseminated zygomycosis.

Three isolates of zygomycetes were used to produce a disseminated infection in nonimmunocompromised mice. Against all zygomycete strains, amphotericin B significantly prolonged survival. Itraconazole was inactive against Rhizopus microsporus and Rhizopus oryzae but was partially active against Absidia corymbifera.

Molecular epidemiology of Aspergillus fumigatus isolates recovered from water, air, and patients shows two clusters of genetically distinct strains.

There has been an increase in data suggesting that besides air, hospital water is a potential source of transmission of filamentous fungi, and in particular Aspergillus fumigatus. Molecular characterization of environmental and clinical A. fumigatus isolates, collected prospectively during an 18-month period, was performed to establish if waterborne fungi play a role in the pathogenesis of invasive aspergillosis.

Evaluation of the post-antifungal effect (PAFE) of amphotericin B and nystatin against 30 zygomycetes using two different media.

The post-antifungal effect (PAFE) of amphotericin B and nystatin against 30 clinical zygomycetes was evaluated using two different media. PAFE is a suppression of fungal growth after limited drug exposure. The MICs of both drugs were determined using NCCLS M38-P guidelines.

In vitro drug interaction modeling of combinations of azoles with terbinafine against clinical Scedosporium prolificans isolates.

The in vitro interaction between terbinafine and the azoles voriconazole, miconazole, and itraconazole against five clinical Scedosporium prolificans isolates after 48 and 72 h of incubation was tested by a microdilution checkerboard (eight-by-twelve) technique. The antifungal effects of the drugs alone and in combination on the fungal biomass as well as on the metabolic activity of fungi were measured using a spectrophotometric method and two colorimetric methods, based on the lowest drug concentrations showed 75 and 50% growth inhibition (MIC-1 and MIC-2, respectively). The nature and the intensity of the interactions were assessed using a nonparametric approach (fractional inhibitory concentration [FIC] index model) and a fully parametric response surface approach (Greco model) of the Loewe additivity (LA) no-interaction theory as well as a nonparametric (Prichard model) and a semiparametric response surface approaches of the Bliss independence (BI) no-interaction theory.

In vitro susceptibilities of zygomycetes to conventional and new antifungals.

In vitro susceptibilities of 36 zygomycete isolates, belonging to six genera, to itraconazole, posaconazole, voriconazole, terbinafine, amphotericin B and 5-fluorocytosine were determined by using a broth microdilution adaptation of the National Committee for Clinical Laboratory Standards M-38P reference method. The influence of incubation time on MIC values, and the performance of a spectrophotometric method for MIC determination in comparison with the visual reference method, were also evaluated. Amphotericin B was active against most of the isolates.

In vitro synergistic interaction between amphotericin B and pentamidine against Scedosporium prolificans.

To develop new approaches for the treatment of invasive infections caused by Scedosporium prolificans, the in vitro interaction between amphotericin B and pentamidine against 30 clinical isolates was evaluated using a checkerboard microdilution method based on the National Committee for Clinical Laboratory Standards M38-P guidelines. The interaction between the drugs was analyzed using fractional inhibitory concentration index (FICI) analysis and response surface modeling. Amphotericin B alone was inactive against all the isolates.

Comparison of the Etest and the sensititre colorimetric methods with the NCCLS proposed standard for antifungal susceptibility testing of Aspergillus species.

The susceptibilities of 25 clinical isolates of Aspergillus fumigatus, A. flavus, A. terreus, A.

In vitro susceptibilities of zygomycetes to combinations of antimicrobial agents.

Combinations of antimicrobial agents were tested against 35 strains of zygomycetes. The interaction between amphotericin B and rifampin was synergistic or additive. Flucytosine alone was inactive and, upon combination with amphotericin B, synergy was not achieved.

Nationwide survey of in vitro activities of itraconazole and voriconazole against clinical Aspergillus fumigatus isolates cultured between 1945 and 1998.

The isolates in a collection of 170 Aspergillus fumigatus isolates recovered from 114 patients and 21 different medical centers in The Netherlands over a period of 53 years were tested for the presence of resistance to itraconazole and voriconazole according to the guidelines of NCCLS document M38-P and by the E-test. Three isolates were highly resistant to itraconazole, and voriconazole MICs were low for all isolates.

In vitro activities of pentamidine, pyrimethamine, trimethoprim, and sulfonamides against Aspergillus species.

The susceptibilities of 70 strains of Aspergillus species were tested against seven different sulfa drugs and pentamidine by a microdilution method with RPMI 1640 and yeast nitrogen base media. Sulfamethoxazole, sulfadiazine, and pentamidine were active in vitro. The MICs obtained with RPMI 1640 were significantly higher than those with yeast nitrogen base.

Method for measuring postantifungal effect in Aspergillus species.

An in vitro method for determination of postantifungal effect (PAFE) in molds was developed by using three isolates each of Aspergillus fumigatus, A. flavus, A. terreus, A.