Clinical Isolates Of Aspergillus Research Articles

Short Communication - Antifungal properties of 5-hydroxytryptamine (serotonin) against Aspergillus spp. in vitro

Abstract This study shows that 5-hydroxytrypatmine (5-HT, serotonin) is fungicidal towards conidia and hyphae of clinical isolates of Aspergillus (A.) fumigatus, A. flavus and A. terreus. The minimal fungicidal concentrations for Aspergillus conidia and hyphae ranged between 14.68 to 117.5 mM and 29.37 to 235 mM during 24 and 48 h of incubation. Several serotonin receptor antagonists (5-HT 2, 5-HT 3) studied in vitro did not influence antifungal activity.

In vitro synergy of caspofungin and amphotericin B against Aspergillus and Fusarium spp.

We investigated the in vitro interaction of caspofungin and amphotericin B for clinical isolates of Aspergillus and FUSARIUM: Synergy tests were performed using the checkerboard method and following the NCCLS M38-P guidelines in Antibiotic Medium 3 broth supplemented to 2% glucose. Antagonism was not observed for any of the isolates tested. Caspofungin and amphotericin B were synergistic or synergistic to additive for at least half of the isolates.

Antifungal activity of a new triazole, voriconazole (UK-109496), against clinical isolates of Aspergillus spp.

Voriconazole is a new triazole antifungal agent with potent activity against yeast and molds. We investigated the in-vitro activity of voriconazole compared with that of other antifungal agents against 50 clinical isolates of Aspergillus spp., measured by the National Committee for Clinical Laboratory Standards (NCCLS) reference method described in the M27-A document, and by an alamar blue colorimetric method. Voriconazole was the most potent agent against Aspergillus fumigatus (minimum inhibitory concentration [MIC]90, 0.5 mg/l) and Aspergillus niger (MIC90, 1.0 mg/l). Voriconazole was less active (MIC90, 1.0 mg/l) against Aspergillus flavus than itraconazole (MIC90, 0.5 mg/l). Voriconazole was more active than itraconazole against Aspergillus fumigatus and Aspergillus flavus by the alamar blue indicator method for the measurement of MIC. Based on these results, voriconazole has promising activity against commonly encountered isolates of Aspergillus spp., and its clinical usefulness should be established by further studies.

In vitro susceptibilities of Aspergillus species to voriconazole, itraconazole, and amphotericin B

We studied the in vitro activity of voriconazole (VCZ) itraconazole (ITZ) and amphotericin B (AMB) against 216 clinical isolates of Aspergillus spp. (142 Aspergillus fumigatus and 74 nonfumigatus Aspergillus spp. isolates) using a broth macrodilution method. The MICs (μg/mL) (mean, range) for A. fumigatus were: VCZ 0.88, 0.25–4; ITZ 0.54, 0.25–4; AMB 2.16, 0.5–8. MIC 90s were: VCZ 2, ITZ 1, AMB 4. MICs for nonfumigatus Aspergillus spp. were: VCZ 1.57, 0.25–4; ITZ 1.74, 0.25–4; AMB 2.88, 0.5–8. MIC 90s for this group were: VCZ 4, ITZ 4, AMB 4. We also studied the susceptibility to VCZ of 18 AMB-resistant (mean, MIC 6.0 μg/mL) and 28 ITZ-resistant (mean, MIC 13.28 μg/mL) A. fumigatus isolates selected in the laboratory. The mean MICs of VCZ were 0.59 μg/mL for AMB-resistant and 1.32 μg/mL for ITZ-resistant isolates. Our study showed that VCZ and ITZ had comparable in vitro activity against the isolates studied, except against A. fumigatus, where the MIC of ITZ was lower. The azoles had better in vitro activity than AMB against A. fumigatus and non- fumigatus spp. The non- fumigatus Aspergillus spp. were less susceptible to all three antifungals evaluated. When tested against ITZ- or AMB-resistant A. fumigatus strains, VCZ retained good activity, showing only a modest rise in the MIC against ITZ-resistant strains.

Potentiation of antifungal activity of amphotericin B by azithromycin against Aspergillus species.

The potential role of azithromycin in combination with amphotericin B against 25 clinical isolates of Aspergillus was assessed. The MIC of amphotericin B was 1 microg/ml for 44% of the isolates, 0.5 microg/ml for 48%, and 0.25 microg/ml for 8%. All isolates were resistant to azithromycin. Synergism, defined as a > or = twofold reduction in the MIC of both drugs upon combination, was demonstrated between amphotericin B and azithromycin for all 25 isolates. To prove that azithromycin exerts its antifungal effect by inhibiting protein synthesis, we studied [35S]-methionine incorporation into protein in one Aspergillus isolate. Neither amphotericin B at 0.125 microg/ml (fourfold below its MIC) nor azithromycin at 16 microg/ml (> or = 16-fold below its MIC) had any effect on protein synthesis when tested alone. Upon combination, however, a 68% inhibition in protein synthesis was evident by the inhibition of [35S]-methionine incorporation.

Targeted Deletion of a GA Tract by S1 Nuclease

Filamentous fungi have a dominant nonhomologous-end joining (NHEJ) DNA repair pathway, which results in the majority of transformed progenies having random heterologous insertion mutagenesis. Thus, lack of a versatile genome-editing tool prevents us from carrying out precise genome editing to explore the mechanism of pathogenesis. Moreover, clinical isolates that have a wild-type ku80 background without any selection nutrition marker especially suffer from low homologous integration efficiency. In this study, we have established a highly efficient CRISPR mutagenesis system to carry out precise and efficient in-frame integration with or without marker insertion with approximately 95–100% accuracy via very short (approximately 35-bp) homology arms in a process referred to as microhomology-mediated end joining (MMEJ). Based on this system, we have successfully achieved an efficient and precise integration of an exogenous GFP tag at the predicted site without marker insertion and edited a conidial melanin gene pksP and a catalytic subunit of calcineurin gene cnaA at multiple predicted sites with or without selection marker insertion. Moreover, we found that MMEJ-mediated CRISPR-Cas9 mutagenesis is independent of the ku80 pathway, indicating that this system can function as a powerful and versatile genome-editing tool in clinical Aspergillus isolates.

Activity of voriconazole (UK-109,496) against clinical isolates of Aspergillus species and its effectiveness in an experimental model of invasive pulmonary aspergillosis.

Voriconazole, a new azole antifungal agent, showed potent activity against clinical isolates of Aspergillus spp. in vitro. For A. fumigatus, the MIC range was < 0.03 to 0.5 microgram/ml and the MIC at which 90% of isolates are inhibited was 0.25 microgram/ml. In an experimental model of invasive pulmonary aspergillosis which mimics infection in humans, oral voriconazole at dosages of 30 mg/kg of body weight per day significantly delayed or prevented mortality.

Prevalence of phthioic acid inAspergillusspecies

We examined fast atom bombardment mass spectra (FAB-MS) of 29 clinical isolates of Aspergillus, from five pathogenic species, for the presence of phthioic acid anions (m/z 395.6) when grown at 37 degrees C. Phthioic acid was detected in only one of 12 A. fumigatus, three of nine A. terreus and one of four A. niger isolates. Phthioic acid is unlikely to be a major pathogenicity determinant of Aspergillus.

Elastase production in clinical isolates of Aspergillus.

Clinical isolates of Aspergillus species were tested, both retrospectively and prospectively, for elastase activity using rose bengal-elastin agar plates. Patient records were then reviewed to determine the clinical diagnosis including the presence or absence of aspergillosis. All isolates that caused invasive aspergillosis produced elastase, but not all isolates producing elastase were associated with invasive disease.

Activity of amphotericin B, 5-fluorocytosine, and rifampin against six clinical isolates of Aspergillus.

Amphotericin B in combination with 5-fluorocytosine was synergistic against three clinical isolates of Aspergillus fumigatus and one of three clinical isolates of A. flavus. Amphotericin B in combination with rifampin was synergistic against all six clinical isolates of Aspergillus tested. The levels of 5-fluorocytosine and rifampin required for synergism were higher than clinically achievable concentrations when measurements of synergism were based on visual turbidity; but when the effects of the drugs were measured by inhibition of ribonucleic acid synthesis or dry-weight increase, much lower concentrations were effective.