Fungal CYP51 Inhibitors VT-1161 and VT-1129 Exhibit Strong In Vitro Activity against Candida glabrata and C. krusei Isolates Clinically Resistant to Azole and Echinocandin Antifungal Compounds.

The in vitro activities of fungal CYP51 inhibitors VT-1161 and VT-1129 were determined for Candida glabrata (n = 34) and C. krusei (n = 50). C. glabrata isolates were screened for FKS gene mutations. All isolates were resistant clinically and/or in vitro to at least one standard antifungal compound. VT-1161 and VT-1129 MICs for all isolates were at least 5-fold below achievable human plasma levels for VT-1161. VT-1161 and VT-1129 are promising for the treatment of resistant C. glabrata and C. krusei infections.

Cryptococcus neoformans-Cryptococcus gattii species complex: an international study of wild-type susceptibility endpoint distributions and epidemiological cutoff values for fluconazole, itraconazole, posaconazole, and voriconazole.

Epidemiological cutoff values (ECVs) for the Cryptococcus neoformans-Cryptococcus gattii species complex versus fluconazole, itraconazole, posaconazole, and voriconazole are not available. We established ECVs for these species and agents based on wild-type (WT) MIC distributions. A total of 2,985 to 5,733 CLSI MICs for C. neoformans (including isolates of molecular type VNI [MICs for 759 to 1,137 isolates] and VNII, VNIII, and VNIV [MICs for 24 to 57 isolates]) and 705 to 975 MICs for C. gattii (including 42 to 260 for VGI, VGII, VGIII, and VGIV isolates) were gathered in 15 to 24 laboratories (Europe, United States, Argentina, Australia, Brazil, Canada, Cuba, India, Mexico, and South Africa) and were aggregated for analysis. Additionally, 220 to 359 MICs measured using CLSI yeast nitrogen base (YNB) medium instead of CLSI RPMI medium for C. neoformans were evaluated. CLSI RPMI medium ECVs for distributions originating from at least three laboratories, which included ≥95% of the modeled WT population, were as follows: fluconazole, 8 µg/ml (VNI, C. gattii nontyped, VGI, VGIIa, and VGIII), 16 µg/ml (C. neoformans nontyped, VNIII, and VGIV), and 32 µg/ml (VGII); itraconazole, 0.25 µg/ml (VNI), 0.5 µg/ml (C. neoformans and C. gattii nontyped and VGI to VGIII), and 1 µg/ml (VGIV); posaconazole, 0.25 µg/ml (C. neoformans nontyped and VNI) and 0.5 µg/ml (C. gattii nontyped and VGI); and voriconazole, 0.12 µg/ml (VNIV), 0.25 µg/ml (C. neoformans and C. gattii nontyped, VNI, VNIII, VGII, and VGIIa,), and 0.5 µg/ml (VGI). The number of laboratories contributing data for other molecular types was too low to ascertain that the differences were due to factors other than assay variation. In the absence of clinical breakpoints, our ECVs may aid in the detection of isolates with acquired resistance mechanisms and should be listed in the revised CLSI M27-A3 and CLSI M27-S3 documents.

Cryptococcus neoformans-Cryptococcus gattii species complex: an international study of wild-type susceptibility endpoint distributions and epidemiological cutoff values for amphotericin B and flucytosine.

Clinical breakpoints (CBPs) are not available for the Cryptococcus neoformans-Cryptococcus gattii species complex. MIC distributions were constructed for the wild type (WT) to establish epidemiologic cutoff values (ECVs) for C. neoformans and C. gattii versus amphotericin B and flucytosine. A total of 3,590 amphotericin B and 3,045 flucytosine CLSI MICs for C. neoformans (including 1,002 VNI isolates and 8 to 39 VNII, VNIII, and VNIV isolates) and 985 and 853 MICs for C. gattii, respectively (including 42 to 259 VGI, VGII, VGIII, and VGIV isolates), were gathered in 9 to 16 (amphotericin B) and 8 to 13 (flucytosine) laboratories (Europe, United States, Australia, Brazil, Canada, India, and South Africa) and aggregated for the analyses. Additionally, 442 amphotericin B and 313 flucytosine MICs measured by using CLSI-YNB medium instead of CLSI-RPMI medium and 237 Etest amphotericin B MICs for C. neoformans were evaluated. CLSI-RPMI ECVs for distributions originating in ≥3 laboratories (with the percentages of isolates for which MICs were less than or equal to ECVs given in parentheses) were as follows: for amphotericin B, 0.5 µg/ml for C. neoformans VNI (97.2%) and C. gattii VGI and VGIIa (99.2 and 97.5%, respectively) and 1 µg/ml for C. neoformans (98.5%) and C. gattii nontyped (100%) and VGII (99.2%) isolates; for flucytosine, 4 µg/ml for C. gattii nontyped (96.4%) and VGI (95.7%) isolates, 8 µg/ml for VNI (96.6%) isolates, and 16 µg/ml for C. neoformans nontyped (98.6%) and C. gattii VGII (97.1%) isolates. Other molecular types had apparent variations in MIC distributions, but the number of laboratories contributing data was too low to allow us to ascertain that the differences were due to factors other than assay variation. ECVs may aid in the detection of isolates with acquired resistance mechanisms.

Evaluation of a digital microfluidic real-time PCR platform to detect DNA of Candida albicans in blood.

Species of Candida frequently cause life-threatening infections in neonates, transplant and intensive care unit (ICU) patients, and others with compromised host defenses. The successful management of systemic candidiasis depends upon early, rapid diagnosis. Blood cultures are the standard diagnostic method, but identification requires days and less than half of the patients are positive. These limitations may be eliminated by using real-time polymerase chain reaction (PCR) to detect Candida DNA in the blood specimens of patients at risk. Here, we optimized a PCR protocol to detect 5-10 yeasts in low volumes of simulated and clinical specimens. We also used a mouse model of systemic candidiasis and determined that candidemia is optimally detectable during the first few days after infection. However, PCR tests are often costly, labor-intensive, and inconvenient for routine use. To address these obstacles, we evaluated the innovative microfluidic real-time PCR platform (Advanced Liquid Logic, Inc.), which has the potential for full automation and rapid turnaround. Eleven and nine of 16 specimens from individual patients with culture-proven candidemia tested positive for C. albicans DNA by conventional and microfluidic real-time PCR, respectively, for a combined sensitivity of 94%. The microfluidic platform offers a significant technical advance in the detection of microbial DNA in clinical specimens.

Fatal Apophysomyces elegans infection transmitted by deceased donor renal allografts.

Two patients developed renal mucormycosis following transplantation of kidneys from the same donor, a near-drowning victim in a motor vehicle crash. Genotypically, indistinguishable strains of Apophysomyces elegans were recovered from both recipients. We investigated the source of the infection including review of medical records, environmental sampling at possible locations of contamination and query for additional cases at other centers. Histopathology of the explanted kidneys revealed extensive vascular invasion by aseptate, fungal hyphae with relative sparing of the renal capsules suggesting a vascular route of contamination. Disseminated infection in the donor could not be definitively established. A. elegans was not recovered from the same lots of reagents used for organ recovery or environmental samples and no other organ transplant-related cases were identified. This investigation suggests either isolated contamination of the organs during recovery or undiagnosed disseminated donor infection following a near-drowning event. Although no changes to current organ recovery or transplant procedures are recommended, public health officials and transplant physicians should consider the possibility of mucormycosis transmitted via organs in the future, particularly for near-drowning events. Attention to aseptic technique during organ recovery and processing is re-emphasized.

Caspofungin for the treatment of azole resistant candidemia in a premature infant.

Candidemia is common in extremely low birth weight infants and is associated with substantial mortality and morbidity. Treatment options have traditionally been limited to amphotericin B deoxycholate or fluconazole. We present a case of a premature infant with persistent candidemia despite antifungal treatment that responded to therapy with caspofungin, an echinocandin antifungal. The infant's Candida isolate developed resistance to azoles during fluconazole administration and also suffered from severe hypercalcemia during the initiation of caspofungin therapy.

Native valve endocarditis due to Candida glabrata treated without valvular replacement: a potential role for caspofungin in the induction and maintenance treatment.

Conventional antifungal therapy for fungal endocarditis has been associated with a poor cure rate. Therefore, combined medical and surgical therapy has been recommended. However, new potent antifungal agents, such as echinocandins, could increase the medical options and, in some cases, avoid the need for surgery. We report a case of Candida endocarditis treated successfully without valve replacement with intravenous liposomal amphotericin B (total dose, 4 g) and intravenous caspofungin (a 100-mg loading dose followed by 50 mg per day for 8 weeks) as induction therapy and intravenous caspofungin (100 mg 3 times per week for 12 weeks) as maintenance therapy.

In vitro and in vivo efficacies of the new triazole albaconazole against Cryptococcus neoformans.

The activity of albaconazole (UR-9825; J. Uriach & Cía. S.A., Barcelona, Spain) was compared to that of fluconazole against 12 isolates of Cryptococcus neoformans in vitro and against 1 isolate in vivo in a rabbit model of cryptococcal meningitis. Albaconazole was 100-fold more potent in vitro than fluconazole on a per-weight basis and was fungicidal at potentially relevant concentrations for two isolates. MICs ranged from

Diguanidino and "reversed" diamidino 2,5-diarylfurans as antimicrobial agents.

Dicationic 2,5-bis(4-guanidinophenyl)furans 5a-5f, 2,5-bis[4-(arylimino)aminophenyl]furans 6a-6b and 6e-6k, and 2,5-bis[4-(alkylimino)aminophenyl]furans 6c-6d have been synthesized starting from 2,5-bis[tri-n-butylstannyl]furan. Thermal melting studies with poly dA*dT and the duplex oligomer d(CGCGAATTCGCG)2 demonstrated high DNA binding affinities for a number of the compounds. The binding affinities are highly dependent on structure and are significantly affected by substituents both on the phenyl rings of the 2,5-diphenylfuran nucleus and on the cationic centers. Of the 17 novel dicationic compounds synthesized, six (6a, 6b, 5b, 6f, 6h, 6i) exhibited MICs of 2 microg/mL or less versus Mycobacterium tuberculosis. Of the compounds screened against Candida albicans, three gave MICs of 2 microg/mL or less (5b, 6h, 6i), and two (5b, 6i) were fungicidal, unlike a standard antifungal drug fluconazole, which was fungistatic. In addition, one of the tested compounds (6i) exhibited a MIC of <1 microg/mL against Aspergillus fumigatus, while also being a fungicidal against this organism. Finally, when evaluated against an expanded fungal panel, compound 6h showed good activity against Cryptococcus neoformans and Rhizopus arrhizus.

Histopathology of fungal rhinosinusitis.

Treatments for the various kinds of fungal rhinosinusitis are fundamentally different, and it is essential to obtain an accurate diagnosis of the particular syndrome in a given case. Microscopic examination of specimens is the most definite and rapid means of establishing a diagnosis of fungal rhinosinusitis. This article compares the utility of various staining and microscopy methods, presents two keys for the algorithmic evaluation of fungal cells as seen in clinical specimens, discusses practical aspects of fungal spore formation within clinical specimens, and reevaluates the role of Candida albicans in rhinosinus disease.

Unusual fungal pathogens in fungal rhinosinusitis.

Approximately 300 fungal species are known to cause mycotic disease in humans and other animals. More than 50 of these species are documented as agents of rhinosinusitis. Most such infections are caused by species of Aspergillus, Rhizopus, Alternaria, Bipolaris, and Curvularia. A growing number, however, has been attributed to lesser known fungi. Here, 38 fungi that are unusual causes of rhinosinusitis are tabulated and referenced in conjunction with their associated symptoms.

Structure-in vitro activity relationships of pentamidine analogues and dication-substituted bis-benzimidazoles as new antifungal agents.

Twenty analogues of pentamidine, 7 primary metabolites of pentamidine, and 30 dicationic substituted bis-benzimidazoles were screened for their inhibitory and fungicidal activities against Candida albicans and Cryptococcus neoformans. A majority of the compounds had MICs at which 80% of the strains were inhibited (MIC80s) comparable to those of amphotericin B and fluconazole. Unlike fluconazole, many of these compounds were found to have potent fungicidal activity. The most potent compound against C. albicans had an MIC80 of

In vitro antifungal activities of a series of dication-substituted carbazoles, furans, and benzimidazoles.

Aromatic dicationic compounds possess antimicrobial activity against a wide range of eucaryotic pathogens, and in the present study an examination of the structures-functions of a series of compounds against fungi was performed. Sixty-seven dicationic molecules were screened for their inhibitory and fungicidal activities against Candida albicans and Cryptococcus neoformans. The MICs of a large number of compounds were comparable to those of the standard antifungal drugs amphotericin B and fluconazole. Unlike fluconazole, potent inhibitory compounds in this series were found to have excellent fungicidal activities. The MIC of one of the most potent compounds against C. albicans was 0.39 microg/ml, and it was the most potent compound against C. neoformans (MIC,

In vitro and in vivo efficacy of the triazole TAK-187 against Cryptococcus neoformans.

Multiple isolates of Cryptococcus neoformans, including those with fluconazole resistance, were tested to assess the in vitro activity of the new triazole TAK-187. MICs of TAK-187 were at least eightfold lower than those of fluconazole, and fungicidal concentrations for most isolates were 4 microg/ml or less. TAK-187 also was evaluated as intermittent therapy using two dosages in a rabbit model of experimental cryptococcal meningitis. Compared to daily treatment with fluconazole, as little as two doses of TAK-187 given 7 days apart were found to be effective. Plasma and cerebrospinal fluid TAK-187 concentrations were many times higher than MICs and fungicidal concentrations. Based upon its therapeutic efficacy and long half-life in the rabbit model, TAK-187 should be investigated for intermittent dosing in treatment or suppression of cryptococcal infections in humans.

Morphologic criteria for the preliminary identification of Fusarium, Paecilomyces, and Acremonium species by histopathology.

Nontraditional human pathogenic fungi, including Fusarium, Paecilomyces, and Acremonium species, have been increasingly documented as agents of infection in immunocompromised patients and, occasionally, in normal hosts. Although definitive identification of these fungi requires culture, they often can be identified provisionally in tissue sections by a combination of histologic features, including hyaline septate hyphae and characteristic reproductive structures known as phialides and phialoconidia. These morphologic characteristics, although familiar to mycologists, are easily overlooked by histopathologists; as a result, Fusarium species and Paecilomyces lilacinus are frequently misidentified in tissue sections as Aspergillus or Candida species. We identified 19 culture-proved cases of infection with species of Fusarium, Paecilomyces, or Acremonium; retrospectively reviewed histologic specimens stained by routine hematoxylin and eosin, Gomori methenamine silver, and/or periodic acid-Schiff stains; and delineated morphologic criteria that will help pathologists make a preliminary identification of these fungi by histopathology. Adventitious sporulation was found in 9 of 9 infections caused by Paecilomyces species, 7 of 10 infections caused by Fusarium species, and in the single case of infection caused by Acremonium strictum. Histologic recognition of these morphologies may help clinicians select appropriate initial antifungal treatment and manage the infection.

In vitro antifungal activity of pneumocandin L-743,872 against a variety of clinically important molds.

The in vitro activity of the new antifungal drug pneumocandin L-743,872 against 55 isolates of clinically important molds was examined by an adapted macrobroth dilution method for yeasts. Pneumocandin L-743,872 exhibited in vitro antifungal activity against Alternaria sp., Aspergillus flavus, Aspergillus fumigatus, Curvularia lunata, Exophiala jeanselmei, Fonsecaea pedrosoi, Paecilomyces variotii, and Scedosporium apiospermum. The drug appeared to lack significant in vitro inhibitory activity against Fusarium oxysporum, Fusarium solani, Rhizopus arrhizus, Paecilomyces lilacinus, and Scedosporium prolificans.

In vitro and in vivo efficacies of the azole SCH56592 against Cryptococcus neoformans.

Multiple isolates of Cryptococcus neoformans were tested to compare the in vitro activity of a new triazole, SCH56592, with those of amphotericin B, fluconazole, and itraconazole, MICs of each drug were determined, and minimum fungicidal concentrations of SCH56592 and amphotericin B were measured. MICs of SCH56592 were lower than those of amphotericin B and fluconazole but not those of itraconazole. Minimum fungicidal concentrations of SCH56592 were lower than those of amphotericin B. SCH56592 in the presence of human serum produces an in vitro fungicidal effect for Cryptococcus neoformans. The data indicate that SCH56592 might exert fungicidal as well as inhibitory properties in vivo. On the basis of these results, SCH56592 was evaluated with a rabbit model of experimental cryptococcal meningitis; SCH56592 treatment was compared with treatment with fluconazole. Despite no detectable drug concentrations in the cerebrospinal fluid, the activity of SCH56592 against C. neoformans infection was equivalent to that of fluconazole. SCH56592 has potent in vitro activity against C. neoformans and compares favorably to treatment with fluconazole for a central nervous system infection. SCH56592 should be studied for use in humans with cryptococcal infections.

Fatal, disseminated Acremonium strictum infection in a neutropenic host.

Disseminated Acremonium strictum infection in a neutropenic patient is reported. Positive fecal cultures preceded positive cutaneous and blood cultures by 18 and 21 days, respectively, which suggests gastrointestinal colonization and invasion as initiating events. Microscopic examination of cutaneous biopsy and pulmonary specimens revealed hyphae, phialides, and phialoconidia in vivo. These adventitious forms also can occur in infections due to other phialidic fungi such as Fusarium and Paecilomyces species and can be misdiagnosed as Candida species. Budding cells also can occur in vivo for species of Fusarium, Paecilomyces, and apparently Acremonium, further adding to the potential for misdiagnosis. The occurrence of adventitious forms in infections caused by species of Acremonium, Fusarium, Paecilomyces, Scedosporium, and Blastoschizomyces is suggested as a mechanism for dissemination of infection and as an explanation of the relatively higher frequency of positive blood cultures in these cases.

The new fungal opportunists are coming.

Frequent and prolonged exposure of immunocompromised patients to a variety of environmental conditions has resulted in the recognition of infections with new fungal opportunists. Infections with fungi such as Fusarium species, Paecilomyces lilacinus, Acremonium species, Trichosporon beigelii, Blastoschizomyces capitatus, Malassezia furfur, Penicillium marneffei, Scedosporium prolificans, and other dematiaceous species have become significant problems in the treatment of immunocompromised hosts. This discussion addresses several issues of pathogenesis, epidemiology, diagnosis, and treatment with regard to these new opportunists, through a review of both general and specific concepts. The growing need for training in clinical mycology is also summarized.