a Laboratori de Microbiologia, Hospital Universitari de Sant Joan de Reus; b Unitat de Microbiologia, Facultat de Medicina i Ciències de la Salut, 43201 Reus; c Institut d'Estudis Avançats, Universitat Rovira i Virgili, Tarragona, Spain
Sir,
Isolates of the fungal genus Alternaria are usually found as soil saprophytes and plant pathogens, although some species can play an important role in causing disease in both healthy and immunocompromised patients. The most common clinical manifestations associated with Alternaria spp. infections include keratitis, peritonitis, osteomyelitis, pulmonary and cutaneous infections, and subcutaneous infections in the case of Ulocladium,1 a morphologically closely related genus. There are scarce data about the in vitro antifungal activity against these fungi, probably because of the lack of reference methods for in vitro antifungal susceptibility testing of dematiaceous fungi.
In this study, the in vitro antifungal activity of amphotericin B, flucytosine, fluconazole, itraconazole, ketoconazole and miconazole was evaluated against 20 isolates of Alternaria spp. (four Alternaria alternata, two Alternaria chlamydospora, two Alternaria dianthicola, one Alternaria geophila, two Alternaria infectoria, three Alternaria longipes and six Alternaria tenuissima) and seven Ulocladium spp. (four Ulocladium chartarum and three Ulocladium botrytis) from very different sources. The antifungal agents were provided by the manufacturer in standard powder form, with the exception of amphotericin B and fluconazole, which were commercial intravenous preparations (Fungizone and Diflucan, respectively). Paecilomyces variotti ATCC 36257 was included as the control.
MICs of antifungal agents were obtained by the broth microdilution method according to the recommendations of the NCCLS (M38-P)2 with some modifications. Conidial inoculum suspension was standardized by the haemocytometer method and it was tested at a final concentration of 15 x 104 conidia/mL. The final test drug dilutions were 0.12 to 64 mg/L of fluconazole, 0.25 to 128 mg/L of flucytosine, and 0.03 to 16 mg/L of amphotericin B, ketoconazole, miconazole and itraconazole. Incubation was at 30°C for 48 h. Flucytosine and azole MICs were defined as the lowest drug dilution that resulted in slight turbidity (approximately 25%) in comparison with the control growth. The MIC of amphotericin B was defined as the lowest drug dilution that inhibited fungal growth completely.
The results of the antifungal susceptibility testing are summarized in the Table. For purposes of comparison, we have considered the breakpoints for defining resistance published by Sutton et al.3 (
2 mg/L of amphotericin B,
1 mg/L of itraconazole,
16 mg/L of miconazole and ketoconazole,
64 mg/L of fluconazole and
32 mg/L of flucytosine). According to these criteria, all isolates of Alternaria spp. were susceptible to miconazole and ketoconazole, 20% of strains were resistant to itraconazole and 45% of strains were resistant to amphotericin B and fluconazole. All isolates were resistant to flucytosine. In contrast, Ulocladium spp. showed a higher degree of resistance to all antifungals: 14.3% of strains were resistant to miconazole, 28.6% to ketoconazole, 71.4% to itraconazole and 85.7% to amphotericin B and fluconazole. All isolates were resistant to flucytosine.
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Our results also agree with those reported from the clinical setting, particularly in the case of itraconazole, i.e. in 11 out of 12 clinical cases treated with this drug the outcome was positive. The clinical efficacy of ketoconazole and amphotericin B has been more variable. Six out of 12 cases were resolved with ketoconazole and eight out of 16 with amphotericin B. Treatment with flucytosine was always in combination with amphotericin B and successful results were obtained in four cases out of six. In contrast to our results, treatment with fluconazole brought a positive outcome in all five clinical cases in which it was used. Only one successful case of treatment with miconazole has been reported, which was in combination with fluconazole. In some cases alternariosis was resolved by combining antifungal drugs with surgical resection, whereas in others only by surgical resection or by stopping the corticosteroid therapy.
In conclusion, our in vitro results generally agree with the clinical data reported in the literature and confirm itraconazole as a promising drug for the treatment of Alternaria spp. infections. Voriconazole, the new triazole derivative, has also demonstrated good in vitro efficacy against A. alternata7 but its possible clinical application should be established by clinical use.
Acknowledgments
This work was supported by the Fundació Ciència i Salut, Reus, Spain and by theCICYT grant PM98-0059.
Notes
J Antimicrob Chemother 2000; 46: 337338
* Corresponding author. Tel: +34-977-759359; Fax: +34-977-759322.
References
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3 . Sutton, D. A., Fothergill, A. W. & Rinaldi, M. G. (1998). Guide to Clinically Significant Fungi. Williams & Wilkins, Baltimore, MD.
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