Acquired itraconazole resistance in Aspergillus fumigatus.

The antifungal susceptibility profiles of four ASPERGILLUS: fumigatus isolates, recovered at different times from a patient treated with itraconazole for a pulmonary ASPERGILLUS: infection, were evaluated. Itraconazole MICs against two pre-treatment isolates were 0.5 mg/L, whilst two later isolates, recovered after at least 4 months of itraconazole therapy, had itraconazole MICs of >16 mg/L. In vivo susceptibilities to itraconazole and amphotericin B were tested in a murine model of disseminated aspergillosis. Treatment efficacy was evaluated by examining mortality rates and qualitative cultures of brain and kidneys. Itraconazole therapy significantly prolonged survival of mice infected with the initial isolates as compared with untreated controls. The third isolate was only partially susceptible to itraconazole in vivo, and the fourth isolate was highly resistant. The four isolates were typed by random amplified polymorphic DNA (RAPD) with four different primers. RAPD patterns obtained with each of them were identical, suggesting that the same strain was recovered over time and had acquired resistance to itraconazole.

In-vivo itraconazole resistance of Aspergillus fumigatus in systemic murine aspergillosis. EBGA Network. European research group on Biotypes and Genotypes of Aspergillus fumigatus.

An animal model of disseminated aspergillosis was used to test the in-vivo activity of itraconazole against four isolates of Aspergillus fumigatus. Two reference isolates of A. fumigatus known to be resistant to itraconazole in vitro and in vivo were used as control isolates, and two new isolates were tested under the same conditions. For each isolate MICs for itraconazole and amphotericin B were determined by an NCCLS-based method. Mice infected intravenously were treated either with itraconazole 100 mg/ kg/day or amphotericin B 4.5 mg/kg/day for 10 days. Amphotericin B showed good in-vivo activity against all four isolates. For one strain, which had a low in-vitro MIC for itraconazole, in-vivo therapy with itraconazole prolonged the survival of mice and reduced fungal burdens in organs compared with untreated controls. In mice infected with a strain with a high MIC of >16 mg/L, itraconazole neither prolonged survival nor reduced fungal load in organs compared with controls. It is concluded that there is a relationship between MIC and treatment outcome in mice for A. fumigatus infection.

In-vitro susceptibility of Aspergillus spp. isolates to amphotericin B and itraconazole.

The MICs of amphotericin B and itraconazole for 230 isolates of Aspergillus spp., comprising 156 Aspergillus fumigatus, 20 Aspergillus terreus, 22 Aspergillus flavus, 17 Aspergillus nidulans and 15 Aspergillus niger, were determined by a broth microdilution method with RPMI 1640 medium. No isolate was detected with an MIC of amphotericin B >2 mg/L. Itraconazole MICs >16 mg/L were detected for four Aspergillus fumigatus and one Aspergillus nidulans isolates.

Use of spectrophotometric reading for in vitro antifungal susceptibility testing of Aspergillus spp.

A comparative study of visual and spectrophotometric MIC endpoint determinations for antifungal susceptibility testing of Aspergillus species was performed. A broth microdilution method adapted from the National Committee for Clinical Laboratory Standards (NCCLS) was used for susceptibility testing of 180 clinical isolates of Aspergillus species against amphotericin B and itraconazole. MICs were determined visually and spectrophotometrically at 490 nm after 24, 48, and 72 h of incubation, and MIC pairs were compared. The agreement between the two methods was 99% for amphotericin B and ranged from 95 to 98% for itraconazole. It is concluded that spectrophotometric MIC endpoint determination is a valuable alternative to the visual reference method for susceptibility testing of Aspergillus species.

Fusarium infections in immunocompromised patients: case reports and literature review.

Five cases are reported of Fusarium infection in patients with aplasia following chemotherapy of leukemia. The clinical signs, diagnosis and course of the infection during treatment are outlined and discussed in conjunction with the characteristics of other cases already reported in the literature. Sixty-three cases of Fusarium infection have been reported in immunocompromised patients, 44 cases since 1985. These included patients with hematological malignancies (58 cases), especially acute leukemia (43 cases). The main sites of infection were the skin (46 cases), blood (28 cases) and lungs (13 cases). The infection was mostly diagnosed by means of skin biopsy but also by means of positive blood cultures. Forty-three strains were identified, 19 of which were Fusarium solani. Amphotericin B treatment was given in 55 cases, often combined with other antifungal agents, leukocyte transfusions or granulocyte-macrophage-colony stimulating factor. The outcome was fatal in 36 of the 63 cases reported, often due to resistance of the strain to antifungal agents, particularly amphotericin B (20 of 33 strains tested). The most important risk factor seems to be profound and prolonged aplasia. Deep mycoses due to Fusarium species thus pose an important problem and are occurring in increasing numbers in immunocompromised patients. Treatment of these infections is difficult and the prognosis is poor.

Serum levels of ketoconazole in bone marrow transplanted patients.

Seven patients with acute leukemia were treated by allogenic bone marrow transplantation from HLA matched sibling. The conditioning regimen was classical using cyclophosphamide and Total Body Irradiation, followed by methotrexate. All patients were given ketoconazole (400 mg per day in a single dose) as antifungal prophylaxis for 6 months. Serum ketoconazole levels were measured using the inhibition assay of mycotic culture in gelose, and they were studied at 0, 1, 2, 4 and 6 h after ketoconazole ingestion, and repeated serially after bone marrow transplantation. In these transplanted patients, absorption of ketoconazole could be delayed, with the maximum levels at 4 h or 6 h after ingestion. Most measurements showed appropriate levels (maximum levels greater than 1 mg/l) even after the third week post transplantation. With the exception of severe acute GVH disease (1 patient), the ketoconazole absorption was adequate in minor or mild GVH disease (6 patients) and in chronic GVH disease (2 patients). In four patients ketoconazole absorption was compared with gut absorption tests (Schilling's test, Iron absorption test, xylose test): In all patients the maximum serum levels of ketoconazole were correct, even in three patients with abnormal gut absorption tests. In this series, no life-threatening mycotic infection occurred, and the three deaths observed showed no mycotic infection.