Molecular phylogenetic diversity of the emerging mucoralean fungus Apophysomyces: proposal of three new species.

BACKGROUND: Apophysomyces is a monotypic genus belonging to the order Mucorales. The species Apophysomyces elegans has been reported to cause severe infections in immunocompromised and immunocompetent people. In a previous study of Alvarez et al.(3) [J Clin Microbiol 2009;47:1650-6], we demonstrated a high variability among the 5.8S rRNA gene sequences of clinical strains of A. elegans. MATERIAL AND METHODS: We performed a polyphasic study based on the analysis of the sequences of the histone 3 gene, the internal transcribed spacer region of the rDNA gene, and domains D1 and D2 of the 28S rRNA gene, as well as by evaluation of some relevant morphological and physiological characteristics of a set of clinical and environmental strains of A. elegans. RESULTS AND CONCLUSIONS: We have demonstrated that A. elegans is a complex of species. We propose as new species Apophysomyces ossiformis, characterised by bone-shaped sporangiospores, Apophysomyces trapeziformis, with trapezoid-shaped sporangiospores, and Apophysomyces variabilis, with variable-shaped sporangiospores. These species failed to assimilate esculin, whereas A. elegans was able to assimilate that glycoside. Amphotericin B and posaconazole are the most active in vitro drugs against Apophysomyces.

Micafungin concentrations from brain tissue and pancreatic pseudocyst fluid.

We report the attainment of micafungin concentrations from brain tissue and pancreatic pseudocyst fluid from two patients with invasive candidiasis. Micafungin was present in low levels at both body sites, indicating limited penetration into central nervous system (CNS) tissue and pancreatic fluid. Further studies are needed to fully characterize its pharmacokinetics at these locations, as micafungin may potentially serve as an alternative antifungal therapy for CNS or pancreatic candidal infections for which the currently recommended first-line therapy fails.

Trichosporon mycotoxinivorans, a novel respiratory pathogen in patients with cystic fibrosis.

This report describes the molecular epidemiology, in vitro susceptibility, colonial and microscopic morphologies, and biochemical features of Trichosporon mycotoxinivorans, a newly recognized pathogen that appears to have a propensity for patients with cystic fibrosis. The index patient died with histologically documented Trichosporon pneumonia complicating cystic fibrosis. This is also the first report of disease caused by a Trichosporon species in a nontransplant patient with cystic fibrosis. As T. mycotoxinivorans has not previously been recognized as a respiratory pathogen, the significance of its recovery from sputum samples was not initially appreciated. Genetic analysis of archived clinical samples found three additional cases of T. mycotoxinivorans infection which had previously been identified as other members of the genus. An additional isolate of T. mycotoxinivorans was identified from a clinical sample on initial testing. Three of these four cases were also patients with cystic fibrosis. All isolates had MICs at 48 h of amphotericin B of > or = 1 microg/ml and of echinocandins of > or = 16 microg/ml, but they displayed various susceptibilities to the triazoles. In summary, Trichosporon mycotoxinivorans is a newly recognized human pathogen that is associated with cystic fibrosis.

Cutaneous infection caused by Macrophomina phaseolina in a child with acute myeloid leukemia.

We report a case of Macrophomina phaseolina skin infection in an immunocompromised child with acute myeloid leukemia, which was treated successfully with posaconazole without recurrence after a hematopoietic stem cell transplant. The fungus was identified by DNA sequencing using both the internal transcribed spacer and D1/D2 region of the 28S ribosomal DNA gene.

Studies on peptidyl nucleoside antibiotics: synthesis and antifungal evaluation of pyranosyl nucleoside analogs of nikkomycin.

BACKGROUND: The Streptomyces-derived nikkomycins are a unique class of peptidyl nucleoside natural products, with potent antifungal activity against a variety of pathogenic fungi. RESULTS: In continuation of our structure-activity relationship studies on the nikkomycins, this paper describes the strategic design, synthesis and biological evaluation of a 'doubly modified' generation of nikkomycin analogs. The structural modifications included a ring-expanded carbohydrate core and a simplified peptidyl side chain. Biological screening of these novel analogs against clinical isolates of various human pathogenic fungi indicated that the described modifications of the structural features of nikkomycin could be a potentially beneficial strategy towards optimizing the antifungal potency of this class of peptidyl nucleoside antibiotics. CONCLUSION: Continued investigation of the pyranosyl nikkomycin analogs is warranted to fully explore and optimize the structural features of this novel lead for the desired development of a new class of therapeutically useful antifungal drugs.

Infections caused by Scedosporium spp.

Scedosporium spp. are increasingly recognized as causes of resistant life-threatening infections in immunocompromised patients. Scedosporium spp. also cause a wide spectrum of conditions, including mycetoma, saprobic involvement and colonization of the airways, sinopulmonary infections, extrapulmonary localized infections, and disseminated infections. Invasive scedosporium infections are also associated with central nervous infection following near-drowning accidents. The most common sites of infection are the lungs, sinuses, bones, joints, eyes, and brain. Scedosporium apiospermum and Scedosporium prolificans are the two principal medically important species of this genus. Pseudallescheria boydii, the teleomorph of S. apiospermum, is recognized by the presence of cleistothecia. Recent advances in molecular taxonomy have advanced the understanding of the genus Scedosporium and have demonstrated a wider range of species than heretofore recognized. Studies of the pathogenesis of and immune response to Scedosporium spp. underscore the importance of innate host defenses in protection against these organisms. Microbiological diagnosis of Scedosporium spp. currently depends upon culture and morphological characterization. Molecular tools for clinical microbiological detection of Scedosporium spp. are currently investigational. Infections caused by S. apiospermum and P. boydii in patients and animals may respond to antifungal triazoles. By comparison, infections caused by S. prolificans seldom respond to medical therapy alone. Surgery and reversal of immunosuppression may be the only effective therapeutic options for infections caused by S. prolificans.

Scedosporium prolificans brain abscess in a patient with chronic granulomatous disease: successful combination therapy with voriconazole and terbinafine.

A patient with chronic granulomatous disease developed brain abscesses with Scedosporium prolificans. In vitro susceptibility revealed a synergistic effect of terbinafine and voriconazole. He received therapy with both these antifungals which resulted in disappearance of the brain abscesses. This is the first reported cure of a CNS S. prolificans infection in an immunocompromised host.

Interlaboratory comparison of results of susceptibility testing with caspofungin against Candida and Aspergillus species.

Seventeen laboratories participated in a study of interlaboratory reproducibility with caspofungin microdilution susceptibility testing against panels comprising 30 isolates of Candida spp. and 20 isolates of Aspergillus spp. The laboratories used materials supplied from a single source to determine the influence of growth medium (RPMI 1640 with or without glucose additions and antibiotic medium 3 [AM3]), the same incubation times (24 h and 48 h), and the same end point definition (partial or complete inhibition of growth) for the MIC of caspofungin. All tests were run in duplicate, and end points were determined both spectrophotometrically and visually. The results from almost all of the laboratories for quality control and reference Candida and Aspergillus isolates tested with fluconazole and itraconazole matched the NCCLS published values. However, considerable interlaboratory variability was seen in the results of the caspofungin tests. For Candida spp. the most consistent MIC data were generated with visual "prominent growth reduction" (MIC(2)) end points measured at 24 h in RPMI 1640, where 73.3% of results for the 30 isolates tested fell within a mode +/- one dilution range across all 17 laboratories. MIC(2) at 24 h in RPMI 1640 or AM3 also gave the best interlaboratory separation of Candida isolates of known high and low susceptibility to caspofungin. Reproducibility of MIC data was problematic for caspofungin tests with Aspergillus spp. under all conditions, but the minimal effective concentration end point, defined as the lowest caspofungin concentration yielding conspicuously aberrant hyphal growth, gave excellent reproducibility for data from 14 of the 17 participating laboratories.

Candida glabrata is an emerging cause of oropharyngeal candidiasis in patients receiving radiation for head and neck cancer.

Oropharyngeal candidiasis (OPC) is relatively common in patients receiving radiation for head and neck cancer occurring in approximately 25% of patients. Candida albicans has been described as the primary infecting organism. Recently, other organisms, particularly Candida glabrata, have emerged as causative agents of OPC among immunocompromised patients. This study describes the characteristics of 6 patients with head and neck cancer treated with radiotherapy at our institution, who were found to have Candida glabrata-associated OPC and their responses to oral fluconazole. All 6 patients were successfully treated with oral fluconazole. However, most did not respond to the usual dose of 100 mg/day necessitating doses ranging from 200 to 800 mg/day to achieve clinical cure. All 3 patients receiving radiation only were successfully treated with up to 200 mg/day; 2 of 3 patients receiving concomitant chemoradiation required doses ranging from 400 to 800 mg/day. As with systemic infection, previous fluconazole use appears to be a risk factor for this infection, but not with all patients.

Oropharyngeal candidiasis caused by non-albicans yeast in patients receiving external beam radiotherapy for head-and-neck cancer.

PURPOSE: To characterize non-albicans Candida oral infections in patients with head-and-neck cancer receiving external beam radiotherapy (EBRT) with or without concurrent chemotherapy. METHODS AND MATERIALS: Thirty-seven patients with head-and-neck cancer received EBRT in 2.0-Gy daily fractions to a median dose of 60.4 Gy (range 38-82.8, mean 64.6). They were followed for oropharyngeal candidiasis (OPC) confirmed by positive examination, positive KOH smear, and/or positive swab or swish culture. Samples were identified and plated on chromogenic media to identify non-albicans yeasts. Colonies were plated on Sabouraud dextrose slants for microdilution antifungal susceptibility testing to fluconazole. DNA typing, including karyotyping, restriction fragment length polymorphism analysis, and Southern blot hybridization with the moderately repetitive Ca3 probe, was performed on selected isolates to confirm individual species. RESULTS: Of the 37 patients, 10 (27%) developed OPC, and 26 (70.3%) displayed Candida carriage state. The median EBRT dose at time of positive culture was 22.5 Gy and at time of OPC was 28.6 Gy. Of the 6 patients receiving chemotherapy and EBRT, 4 (66%) developed OPC at median dose of 27.6 Gy. Three (8%) of 37 patients were infected with non-albicans Candida, and 3 (30%) of all 10 infections were caused by these organisms. CONCLUSION: Non-albicans Candida is emerging as a relatively common cause of OPC in head-and-neck cancer patients. Chromogenic media are helpful to screen these infections. Our data also suggest a greater likelihood of developing OPC in patients receiving concomitant chemotherapy and EBRT.

Pseudallescheria boydii (Anamorph Scedosporium apiospermum). Infection in solid organ transplant recipients in a tertiary medical center and review of the literature.

(Sca) is a ubiquitous filamentous fungus capable of causing invasive disease. We reviewed our electronic microbiology records and the English-language literature. Between 1976 and December 1999 we identified 23 solid organ transplant recipients with Sca infection, 7 of which occurred between December 1987 and December 1999 at our institution. Overall incidence was 1 per 1,000 patients, with a trend of higher incidence in patients receiving lung transplants compared with other transplant organs (p = 0.06). The 23 patients included liver (4), kidney (8), heart (8), lung (2), and heart/lung (1) recipients. Male to female ratio was 19:4, and the mean age was 46 +/- 12 (SD) years. Fungal infection was diagnosed at a median of 4 months (range, 0.4-156 mo) after transplant. The clinical presentation included disseminated disease ( 8), skin lesions (3), lung disease (5), endophthalmitis (1), meningitis (1), brain abscess with or without extension to eye (3), fungal mycotic aneurysm (1), and sinusitis (1). Seven (30%) patients had intravascular infection, and 11 (48%) patients had central nervous system involvement. Antifungal therapy was accompanied by surgical debridement in 9 cases. Three additional patients were found to have airway colonization only and received itraconazole prophylaxis, without evidence of disease. Of 22 patients with known outcome, 16 (72.7%) died. Five of 6 patients who survived had localized infections: skin lesions (n = 3), sinus fungus ball (n = 1), and solitary lung nodule (n = 1). All patients with disseminated disease and 10 of 11 patients with central nervous system disease died. An exception was 1 patient with a brain abscess, successfully treated with voriconazole and surgical drainage. Sca infection is rare but is associated with high mortality. Early diagnosis by culture is important because Sca is resistant to amphotericin B, routinely used in the empiric therapy of invasive fungal infections. Treatment with the combination of an antifungal and surgery may have a better outcome. Voriconazole promises to be an effective antifungal agent. Cultures positive for Sca should not be ignored, and long-term antifungal prophylaxis in candidates and transplant recipients should be considered.

In vitro interaction of caspofungin acetate with voriconazole against clinical isolates of Aspergillus spp.

The interaction between caspofungin acetate and voriconazole was studied in vitro by using 48 clinical Aspergillus spp. isolates obtained from patients with invasive aspergillosis. MICs were determined by the NCCLS broth microdilution method. Synergy, defined as a fractional inhibitory concentration (FIC) index of <1, was detected in 87.5% of the interactions; an additive effect, defined as an FIC index of 1.0, was observed in 4.2% of the interactions; and a subadditive effect, defined as an FIC index of 1.0 to 2.0, was found in 8.3% of the interactions. No antagonism was observed. Animal models are required to validate the in vivo significance of these in vitro data presented for the combination of caspofungin and voriconazole.

Enteral fluconazole is well absorbed in critically ill surgical patients.

BACKGROUND: Enteral fluconazole, a triazole antifungal agent with an excellent oral bioavailability, has not been widely studied in critically ill surgical patients. METHODS: During a randomized placebo-controlled trial of enteral fluconazole (N = 130) versus placebo (N = 130) for the prevention of fungal infections in critically ill surgical patients, trough fluconazole levels were measured after the loading dose and 3 times weekly during intensive care unit stay. Minimum inhibitory concentrations (MICs) for fluconazole were measured on all infecting Candida isolates. RESULTS: Four hundred sixty-seven serum samples were assayed for fluconazole levels in 121 patients. The most common infecting fungal species was Candida albicans, isolated in 14 of 31 infections (45%). Other infecting species were C glabrata, C tropicalis, and C parapsilosis. Mean fluconazole levels were above the highest MIC for C albicans and C parapsilosis in all but 5 patients (4%). Mean fluconazole levels were below the median MIC for C glabrata in 93 of 121 patients (77%). No significant relationship was seen between fluconazole levels and risk for fungal infection. CONCLUSIONS: Serum fluconazole levels are above the MIC of most yeast species found in these patients. These levels may not be above the MIC of C glabrata, the second most common Candida isolate causing infection in this study.

Candida glabrata oropharyngeal candidiasis in patients receiving radiation treatment for head and neck cancer.

Candida glabrata colonization is common in patients receiving radiation treatment for head and neck cancer, but to our knowledge has never been described as the infecting organism with oropharyngeal candidiasis (OPC). This study presents the first three patients described with C. glabrata OPC in this patient population. Patient 1 developed C. glabrata OPC and required fluconazole, 800 mg/day, for clinical resolution. Antifungal susceptibility testing revealed a MIC of fluconazole of >64 microg/ml. Elapsed time from initial culturing to treatment decision was 7 days. Patients 2 and 3 developed C. glabrata OPC. They were patients in a study evaluating OPC infections, and cultures were taken immediately. CHROMagar Candida plates with 0, 8, and 16 microg of fluconazole/ml were employed for these cultures. Lavender colonies, consistent with C. glabrata, grew on the 0- and 8-microg plates but not on the 16-microg plate from patient 2 and grew on all three plates from patient 3. Based on these data, a fluconazole dose of 200 mg/day was chosen for patient 2 and a dose of 400 mg/day was chosen for patient 3, with clinical resolution in both. Elapsed time from initial culturing to treatment decision was 2 days. C. glabrata does cause OPC in head and neck radiation treatment patients, and the use of fluconazole-impregnated chromogenic agar may significantly reduce treatment decision time compared to that with conventional culturing and antifungal susceptibility testing.

Experimental murine chromomycosis mimicking chronic progressive human disease

Congenitally athymic (nu/nu) mice, mice defective in NK cell and macrophage function (bg/bg) and normal BALB/c mice were inoculated sc with 10 5-6 conidia of Fonsecae pedrosoi (FP). In immunologically intact and immunodeficient mice, a local infection developed approximately 2 weeks post-inoculation and enlarged over 1-2 weeks. In bg/bg and normal nu/+ mice, lesions resolved within 5-6 weeks. However, nu/nu mice continued to have enlarging cs lesions during >4-6 months od observation. These eventually metastasized. Lesions contained, few hyphal elements and massive numbers of sclerotic bodies. Five weeks after inoculation. 10 4-6 conidia froming units/gm of tissue were recovered from lesions. Delayed type hypersensitivity and serum antibody to FP antigens were demonstrated. Adoptive transfer of lymphocytes from nu/+ mice was followed in months bu the resolution of the lesions