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We report the first well documented case of subcutaneous phaeohyphomycotic infection caused by Phialophora verrucosa in India. Examination of the biopsied tissue from an ulcerating lesion on the shin of the left leg of a 45-year-old woman from Bombay, India, showed numerous dematiaceous, septate, branching hyphal elements and thick-walled cells characteristic of phaeohyphomycosis. Cultures of the scrapings from the lesion and of the biopsied tissue yielded a pigmented fungus that was identified as P. verrucosa. Initial treatment with fluconazole followed by oral itraconazole for 30 days and local application of a copper sulphate solution resulted in complete resolution of the lesion. Treatment with itraconazole was continued for an additional 3 months after complete healing. No new lesions developed and the patient did not show jaundice, hepatosplenomegaly or any other signs of toxicity.

Sixteen isolates belonging to Fusarium chlamydosporum (n = 4), Fusarium equiseti (n = 1), Fusarium moniliforme (n = 2), Fusarium oxysporum (n = 3), Fusarium proliferatum (n = 1), and Fusarium solani (n = 5) were tested against amphotericin B, 5-fluorocytosine, fluconazole, itraconazole, ketoconazole, JAI-amphotericin B (water-soluble compound), hamycin and amphotericin B combined with 5-fluorocytosine, using antibiotic medium M3, high-resolution broth (pH 7.1), Sabouraud's dextrose, and yeast-nitrogen broth media (1 ml/tube). The minimal inhibitory and minimal fungicidal concentrations of 5-fluorocytosine and fluconazole for all species were > 100 micrograms/ml. All Fusarium isolates, except F. equiseti (3.125 micrograms), gave minimal inhibitory concentrations of 12.5-100 micrograms/ml for hamycin. The values for amphotericin B, itraconazole, ketoconazole, JAI-amphotericin B, and amphotericin B combined with 5-fluorocytosine were 1.56-100, 0.78-50, 3.125-100,50-100, and 1.56 to > 100 micrograms/ml, respectively. Although a wide range of minimal inhibitory concentrations was recorded for most of the isolates studied, it appears that some--F. solani, F. oxysporum, F. chlamydosporum, F. equiseti, and F. moliniforme--were more susceptible to amphotericin B, itraconazole, ketoconazole, hamycin, and amphotericin B in the presence of 5-fluorocytosine. All isolates showed resistance to 5-fluorocytosine and fluconazole. The minimal fungicidal concentrations were either the same or several times higher than the minimal inhibitory concentrations.

The mycelial (25 degrees C) and yeast-like (37 degrees C) forms of Penicillium marneffei clinical and type strains were investigated for their in vitro susceptibility to amphotericin B (AmB), 5-fluorocytosine (5-FC), fluconazole (FLU) and itraconazole (ITZ), using Bacto antibiotic medium 3, yeast-nitrogen, Sabouraud's dextrose (pH 5.7) and high resolution (pH 7.1) broth media (1ml/tube), respectively. Results indicated that the minimal inhibitory and minimal fungicidal concentrations (MICs and MFCs) for the mycelial cultures of P. marneffei to AmB were in the range 0.78-1.56 and 0.78-3.125 micrograms/ml, respectively, as against 3.125-25 micrograms (MICs) for the yeast form cultures. The MFCs to AmB for the yeast form were one dilution higher. The MICs to FLU were generally lower for the yeast form (6.25-25 micrograms) than the mycelial form (25-50 micrograms/ml), whereas MFCs for the mycelial cultures were > 100 micrograms as compared to 6.25-100 micrograms for their yeast form. The MICs for the mycelial form to 5-FC ranged from < 0.195-0.39 microgram. Higher MICs (6.25 micrograms) were recorded for their yeast form. The MFCs to 5-FC for the yeast form were 25-100 micrograms/ml. The MICs for the mycelial form to ITZ ranged from < 0.195 to 3.125 micrograms/ml. Higher values (< 0.195-50 micrograms) were recorded for their yeast-like form. The MFCs to ITZ for mycelial and yeast forms ranged from < 0.195-0.39 and 25-100 micrograms/ml, respectively. Results indicate that P. marneffei's yeast form is more sensitive to FLU and ITZ (8 of 10 strains) while the mycelial form displayed greater susceptibility to AmB and 5-FC. The MICs for ITZ remained steady in SD medium, pH 5.7 to 7.1. However, some strains gave higher MIC values (0.39-1.56 micrograms/ml) when tested in the HR.

Ten isolates of Penicillium marneffei and eight of Pythium insidiosum were tested for their in vitro sensitivity to amphotericin B, hamycin (a polyene heptaene), two water-soluble analogs of amphotericin B and hamycin, namely, JAI-Amb, and JAI-hamycin,5-fluorocytosine, fluconazole, itraconazole,ketoconazole and miconazole. Itraconazole manifested the strongest activity against all of the 10 isolates of P. marneffei and would be the drug of choice in the treatment of penicilliosis due to P. marneffei. The polyene antibiotics amphotericin B and hamycin and their water-soluble analogs showed no appreciable activity against P. insidiosum. Pytium insidiosum isolates were sensitive to fluconazole, ketoconazole, and miconazole. Miconazole exhibited the strongest in vitro activity against all of the 8 isolates of P. insidiosum, followed by ketoconazole.

The sensitivity of 23 isolates of opportunistic fungi, Aspergillus fumigatus (5), A. flavus (5), A. niger (5), Pseudallescheria boydii (5), Alternaria alternata (2) and Xylohypha bantiana (1), was investigated against fluconazole and itraconazole, using Sabouraud's dextrose broth (SD) and a high-resolution (HR) medium (Pfizer, Inc.). The procedure followed was a standard tube dilution (1 ml/tube) method. Candida albicans Y01 09 was included as reference strain to monitor quality and reproducibility. Results indicated that the minimal inhibitory concentrations (MICs) of fluconazole for all Aspergillus spp. and C. albicans were greater than or equal to 100 micrograms/ml in SD medium, whereas, for P. boydi, A. alternata and X. bantiana, the MICs were 50 micrograms/l. The MICs of itraconazole in SD medium were less than 0.195-1.56 micrograms/ml for the fungi tested. In HR medium, the MICs of fluconazole for the Aspergillus spp, were greater than or equal to 100 micrograms/ml, and those for P. boydii, A. alternata and X. bantiana were 0.78, 50 and 50 micrograms/ml, respectively. The MICs of itraconazole for all fungi ranged from less than 0.198 to 0.78 micrograms/ml in the HR medium. The values for the reference strain were 1.56 and 100 micrograms/ml in the HR medium for fluconazole and itraconazole, respectively. The HR medium was more suitable for testing P. boydii against fluconazole. This culture medium did not appear to significantly affect the MICs of itraconazole as compared to those of fluconazole, for the fungi investigated.

Twenty-eight pathogenic isolates, 4 each of Candida albicans, C. lusitaniae, C. parapsilosis, Cryptococcus neoformans, Torulopsis glabrata and Trichosporon beigelii were tested for their in vitro sensitivity to amphotericin B (AmB), hamycin (HA) and their novel water-soluble compounds, namely JAI-AmB (oral and injectable; patents pending) and JAI-HA (Jaimycin Inc., Walnut Creek, Calif., USA), using a standard double-dilution broth (1 ml/tube) procedure. The 2 novel compounds, namely JAI-AmB and JAI-HA, contain one twenty-fifth (w/w) of the AmB and HA, respectively. Results showed that the minimal inhibitory concentrations (MICs) of AmB for all species, except C. lusitaniae and Tr. beigelii (3.125-6.25 micrograms/ml), were 0.195-1.56 micrograms/ml. The values of JAI-AmB (oral) for C. albicans, C. parapsilosis, Cr. neoformans and To. glabrata ranged from 0.78 to 25 micrograms/ml. The MICs of JAI-AmB (oral) for the other yeasts were 100 micrograms/ml. All of the yeasts yielded higher MICs (3.125-100 micrograms/ml) against JAI-AmB (injectable) than the JAI-AmB (oral) preparation. Except for C. parapsilosis (MICs 25-100 micrograms/ml), all of the other species showed greater sensitivity to the parent HA (0.195-100 micrograms/ml) than AmB. The values for JAI-HA ranged from 0.195 to 100 micrograms/ml for all isolates, except C. tropicalis (100 micrograms/ml). Based on our in vitro findings, in vivo efficacies of JAI-AmB and JAI-HA should be carried out.

Twenty isolates of Prototheca filamenta, Prototheca moriformis, Prototheca stagnora, Prototheca wickerhamii, and Prototheca zopfii were tested for in vitro susceptibility to five commonly used antifungal agents: amphotericin B, 5-fluorocytosine, griseofulvin, miconazole, and nystatin. The results revealed resistance to griseofulvin of all the Prototheca isolates tested and an inhibitory effect on P. filamenta by high 5-fluorocytosine concentrations (minimal inhibitory concentration [MIC] = 12.5 to 100 mug/ml; minimal fungicidal or algacidal concentration [MFC/MAC] = 50 to 100 mug/ml). P. filamenta isolates were also susceptible to miconazole (MIC = 0.1 to 0.5 mug/ml, MFC/MAC = 0.5 to 1 mug/ml); isolates of the other Prototheca species varied in regard to miconazole activity from susceptible to resistant (MIC = 1 - >100 mug/ml, MFC/MAC = 5 - >100 mug/ml). The Prototheca isolates revealed an in vitro susceptibility to the polyene antifungal agents, amphotericin B, and nystatin (MIC = 0.09 to 3.12 mug/ml and 0.19 to 12.5 mug/ml, respectively; MFC/MAC = 0.19 to 25 mug/ml and 0.75 to 25 mug/ml, respectively).