Current knowledge on the strain typing of the pathogenic fungus Histoplasma capsulatum var. capsulatum: a review of the findings

The classification of microbial strains is currently based on different typing methods, which must meet certain criteria in order to be widely used. Phenotypic and genotypic methods are being employed in the epidemiology of several fungal diseases. However, some problems associated to the phenotypic methods have fostered genotyping procedures, from DNA polymorphic diversity to gene sequencing studies, all aiming to differentiate and to relate fungal isolates or strains. Through these studies, it is possible to identify outbreaks, to detect nosocomial infection transmission, and to determine the source of infection, as well as to recognize virulent isolates. This paper is aimed at analyzing the methods recently used to type Histoplasma capsulatum, causative agent of the systemic mycosis known as histoplasmosis, in order to recommend those that yield reproducible and accurate results.     

Antifungal Efficacy during Candida krusei Infection in Non-Conventional Models Correlates with the Yeast In Vitro Susceptibility Profile

The incidence of opportunistic fungal infections has increased in recent decades due to the growing proportion of immunocompromised patients in our society. Candida krusei has been described as a causative agent of disseminated fungal infections in susceptible patients. Although its prevalence remains low among yeast infections (2–5%), its intrinsic resistance to fluconazole makes this yeast important from epidemiologic aspects. Non mammalian organisms are feasible models to study fungal virulence and drug efficacy. In this work we have used the lepidopteran Galleria mellonella and the nematode Caenorhabditis elegans as models to assess antifungal efficacy during infection by C. krusei. This yeast killed G. mellonella at 25, 30 and 37°C and reduced haemocytic density. Infected larvae melanized in a dose-dependent manner. Fluconazole did not protect against C. krusei infection, in contrast to amphotericin B, voriconazole or caspofungin. However, the doses of these antifungals required to obtain larvae protection were always higher during C. krusei infection than during C. albicans infection. Similar results were found in the model host C. elegans. Our work demonstrates that non mammalian models are useful tools to investigate in vivo antifungal efficacy and virulence of C. krusei.     

Amphotericin B mediates killing in Cryptococcus neoformans through the induction of a strong oxidative burst

We studied the effects of Amphotericin B (AmB) on Cryptococcus neoformans using different viability methods (CFUs enumeration, XTT assay and propidium iodide permeability). After 1h of incubation, there were no viable colonies when the cells were exposed to AmB concentrations ≥ 1 mg/L. In the same conditions, the cells did not become permeable to propidium iodide, a phenomenon that was not observed until 3h of incubation. When viability was measured in parallel using XTT assay, a result consistent with the CFUs was obtained, although we also observed a paradoxical effect in which at high AmB concentrations, a higher XTT reduction was measured than at intermediate AmB concentrations. This paradoxical effect was not observed after 3h of incubation with AmB, and lack of XTT reduction was observed at AmB concentrations higher than 1mg/L. When stained with dihydrofluorescein, AmB induced a strong intracellular oxidative burst. Consistent with oxidative damage, AmB induced protein carbonylation. Our results indicate that in C. neoformans, Amphotericin B causes intracellular damage mediated through the production of free radicals before damage on the cell membrane, measured by propidium iodide uptake.     

Actividad antimicótica y citotóxica de aceites esenciales de plantas de la familia Asteraceae

BackgroundThe plants of the Asteraceae family have been used for medicinal purposes,in traditional Colombian medicine.AimTo evaluate the antifungal activity and the cytotoxic effects of 15 essential oils from plants of the Asteraceae family.MethodsAntifungal activity was evaluated against Candida parapsilosis ATCC 22019, Candida krusei ATCC 6258, Aspergillus flavus ATCC 204304 and Aspergillus fumigatus ATCC 204305 following EUCAST and CLSI M38-A standard methods, for yeast and filamentous fungi, respectively. Cytotoxic effect was evaluated on Vero cell line by MTT assay.ResultsThe oils from the plants Achyrocline alata and Baccharis latifolia were the only ones active against A. fumigatus (GM-MIC=78.7 and 157.4 μg/ml, respectively). In contrast, there was no evidence of oils active against Candida species. In addition, these oils were not cytotoxic on Vero cells. The oils of A. alata and Baccharis latifolia could be candidates for disinfecting hospital environments and for inhibiting biofilm formation by A. fumigatusConclusionsThe oils of A. alata and B. latifolia could be candidates for disinfecting hospital environments and for inhibiting biofilm formation by A. fumigatus     

Molecular Identification and Antifungal Susceptibility of Clinical Isolates of Sporothrix schenckii Complex in Medellin,Colombia

Background

Sporotrichosis is a subcutaneous mycosis that affects humans and other animals. Infection prevails in tropical and subtropical countries. Until a few years ago, it was considered that two varieties of Sporothrix schenckii caused this mycosis, but by applying molecular taxonomic markers, it has been demonstrated that there are several cryptic species within S. schenckii complex which varies in susceptibility, virulence, and geographic distribution.

Objective

This study aimed to identify the clinical isolates of Sporothrix spp. from patients with sporotrichosis in Medellin, Colombia, using two markers and to evaluate the in vitro susceptibility to itraconazole.

Methods

Thirty-four clinical isolates of Sporothrix spp. from Colombia, three from Mexico, and one from Guatemala were identified through sequencing of the noncoding region ITS-1?+?5.8SDNAr?+?ITS-2 and of the fragment containing exons 3 and 4 of the β-tubulin gene. Clinical isolate sequences were compared with GenBank reference sequences using the BLASTN tool, and then, phylogenetic analysis was performed. Besides, the in vitro susceptibility to itraconazole was evaluated by determining the minimum inhibitory concentrations according to the CLSI M38-A2 method.

Results

Clinical isolates were identified by morphology as Sporothrix spp. Using the molecular markers, ITS and β-tubulin, isolates were identified as S. schenckii sensu stricto (25) and Sporothrix globosa (13). Susceptibility to itraconazole was variable among clinical isolates.

Conclusion

This is the first scientific publication that identifies species that cause sporotrichosis in Colombia, along with the antifungal susceptibility to itraconazole.