Prevalence and characterization of azole-resistant Aspergillus fumigatus in patients with cystic fibrosis: a prospective multicentre study in Germany.

Objectives: Aspergillus fumigatus is the most prevalent filamentous fungus in the respiratory tract of patients with cystic fibrosis (CF). The aim of this prospective multicentre study was to investigate the prevalence of azole-resistant A. fumigatus (ARAF) in respiratory secretions from CF patients across Germany and to characterize ARAF isolates by phenotypic and molecular methods. Methods: Twelve tertiary care centres from Germany participated in the study. In total, 2888 A. fumigatus isolates from 961 CF patients were screened for ARAF by using azole-containing agar plates. Antifungal susceptibility testing of isolates was performed by broth microdilution according to EUCAST guidelines. Analysis of mutations mediating resistance was performed using PCR and sequencing of the cyp51A gene. Furthermore, genotyping by microsatellite PCR was performed. Results: Of a total of 2888 A. fumigatus isolates, 101 isolates from 51 CF patients were found to be azole resistant (prevalence per patient 5.3%). The Essen centre had the highest prevalence (9.1%) followed by Munich (7.8%), Münster (6.0%) and Hannover (5.2%). Most ARAF isolates (n = 89) carried the TR34/L98H mutation followed by eight G54E/R, one TR46/Y121F/T289A and one F219S mutation. In two isolates no mutation was found. Genotyping results showed no major clustering. Forty-five percent of CF patients with ARAF had previously received azole therapy. Conclusions: This is the first multicentre study analysing the prevalence of ARAF isolates in German CF patients. Because of a resistance rate of up to 9%, susceptibility testing of A. fumigatus isolates from CF patients receiving antifungal treatment should be part of standard diagnostic work-up.

Actions of pyrethroid insecticides on voltage-gated chloride channels in neuroblastoma cells.

Pyrethroid insecticides have the potential to act at several sites in excitable tissues in addition to their primary effect on sodium channels. We have assessed one such site, the voltage gated chloride channel. Following our previous demonstration of actions of deltamethrin on the voltage-gated chloride channel in rats and in neuroblastoma cells, we examined the dose-response relationship for deltamethrin by patch clamp analysis of voltage-gated channels in partially differentiated NIE115 cells. Open channel probability (P0) was assessed in channels stepped from 0 to +20 mV for 10 seconds. Deltamethrin significantly decreased P0 by 0.237+/-0.070 at 10(-10) M and, although construction of a dose-response relationship was rendered difficult by the very low water solubility of the pyrethroids, a decrease of 0.968+/-0.140 was reached at the highest concentration tested, 10(-4) M. A second Type II pyrethroid cypermethrin also decreased P0, by 0.430+/-0.09 at 5 x 10(-6) M, but the Type I pyrethroid cismethrin produced a decrease of only 0.188+/-0.08 at 2 x 10(-5) M. These effects were seen in 340 pS conductance, calcium-independent channels. A sample of 9 calcium-dependent channels with a conductance of 225 pS failed to show any response to deltamethrin. We conclude that actions on calcium-independent voltage-gated chloride channels are likely to contribute significantly to type II pyrethroid toxicity.