Multicenter, International Study of MIC/MEC Distributions for Definition of Epidemiological Cutoff Values for Sporothrix Species Identified by Molecular Methods.

Clinical and Laboratory Standards Institute (CLSI) conditions for testing the susceptibilities of pathogenic Sporothrix species to antifungal agents are based on a collaborative study that evaluated five clinically relevant isolates of Sporothrixschenckii sensu lato and some antifungal agents. With the advent of molecular identification, there are two basic needs: to confirm the suitability of these testing conditions for all agents and Sporothrix species and to establish species-specific epidemiologic cutoff values (ECVs) or breakpoints (BPs) for the species. We collected available CLSI MICs/minimal effective concentrations (MECs) of amphotericin B, five triazoles, terbinafine, flucytosine, and caspofungin for 301 Sporothrix schenckii sensu stricto, 486 S. brasiliensis, 75 S. globosa, and 13 S. mexicana molecularly identified isolates. Data were obtained in 17 independent laboratories (Australia, Europe, India, South Africa, and South and North America) using conidial inoculum suspensions and 48 to 72 h of incubation at 35°C. Sufficient and suitable data (modal MICs within 2-fold concentrations) allowed the proposal of the following ECVs for S. schenckii and S. brasiliensis, respectively: amphotericin B, 4 and 4 µg/ml; itraconazole, 2 and 2 µg/ml; posaconazole, 2 and 2 µg/ml; and voriconazole, 64 and 32 µg/ml. Ketoconazole and terbinafine ECVs for S. brasiliensis were 2 and 0.12 µg/ml, respectively. Insufficient or unsuitable data precluded the calculation of ketoconazole and terbinafine (or any other antifungal agent) ECVs for S. schenckii, as well as ECVs for S. globosa and S. mexicana These ECVs could aid the clinician in identifying potentially resistant isolates (non-wild type) less likely to respond to therapy.

Phylogeography and evolutionary patterns in Sporothrix spanning more than 14 000 human and animal case reports.

Pathology to vertebrate hosts has emerged repeatedly in the order Ophiostomatales. Occasional infections have been observed in Sporothrix mexicana at a low level of virulence, while the main pathogenic species cluster in a derived clade around S. schenckii s.str. In this paper, phylogeny and epidemiology of the members of this clade were investigated for 99 clinical and 36 environmental strains using four genetic loci, viz. rDNA ITS and partial CAL, TEF1, and TEF3; data are compared with amplified fragment length polymorphism (AFLP) genotyping. The four main species of the pathogenic clade were recognised. The species proved to show high degrees of endemicity, which enabled interpretation of literature data where live material or genetic information is lacking. The clade of four species comprised nine subclusters, which often had limited geographic distribution and were separate from each other in all partitions, suggesting low degrees of interbreeding between populations. In contrast, S. globosa exhibited consistent global distribution of identical AFLP types, suggesting another type of dispersal. Sporothrix brasiliensis is known to be involved in an expanding zoonosis and transmitted by cats, whereas S. globosa infections originated from putrid plant material, causing a sapronosis. Sporothrix schenckii s.str., the most variable species within the clade, also had a plant origin, with ecological similarities to that of S. globosa. A hypothesis was put forward that highly specific conditions in the plant material are required to promote the growth of Sporothrix. Fermented, self-heated plant debris may stimulate the thermodependent yeast-like invasive form of the fungus, which facilitates repeated infection of mammals.

Rapid diagnosis of coccidioidomycosis by nested PCR assay of sputum.

Coccidioidomycosis is a deep infection caused by two dimorphic fungi, Coccidioides immitis and Coccidioides posadasii. Diagnosis of the disease requires culture of suspicious clinical samples on mycological media. However, as these species are virulent pathogens, handling of their cultures is a high-risk activity, and is limited to Biosafety Level 3 laboratories. This study describes the direct detection of C. posadasii DNA in an inappropriate sputum sample by PCR amplification of the highly specific Ag2/PRA antigen gene. The results obtained suggest that direct detection of the Ag2/PRA sequence in sputum is an excellent method for rapid and specific diagnosis of coccidioidomycosis.

Comparison between magnetic enzyme-linked immunosorbent assay (MELISA) and complement fixation test (CF) in the diagnosis of paracoccidioidomycosis.

MELISA and CF were compared using sera from paracoccidioidomycosis patients before treatment and patients undergoing antimycotic treatment. With MELISA it was possible to distinguish different antibody levels in both groups of patients whereas such distinction was not observed by using CF tests. MELISA is thus an advantageous alternative to CF in the diagnosis of paracoccidioidomycosis, including the possibility of testing sera with anticomplementary activity.

Use of morpho-physiological characteristics for differentiation of the species of Prototheca.

Fifty-two cultures of Prototheca spp. isolated from water and 7 isolates received from a culture collection were tested for their ability to assimilate carbon and nitrogen sources. Based upon these findings and on micromorphological features of the isolates a rapid method allowing differentiation of Prototheca spp. in culture is presented.