Azole resistance in Candida spp. isolated from Catú Lake, Ceará, Brazil: an efflux-pump-mediated mechanism

Abstract Since, there is no study reporting the mechanism of azole resistance among yeasts isolated from aquatic environments; the present study aims to investigate the occurrence of antifungal resistance among yeasts isolated from an aquatic environment, and assess the efflux-pump activity of the azole-resistant strains to better understand the mechanism of resistance for this group of drugs. For this purpose, monthly water and sediment samples were collected from Catú Lake, Ceará, Brazil, from March 2011 to February 2012. The obtained yeasts were identified based on morphological and biochemical characteristics. Of the 46 isolates, 37 were Candida spp., 4 were Trichosporon asahii, 3 were Cryptococcus laurentii, 1 Rhodotorula mucilaginosa, and 1 was Kodamaea ohmeri. These isolates were subjected to broth microdilution assay with amphotericin B, itraconazole, and fluconazole, according to the methodology standardized by the Clinical and Laboratory Standards Institute (CLSI). The minimum inhibitory concentrations (MICs) of amphotericin B, itraconazole, and fluconazole were 0.03125–2 µg/mL, 0.0625 to ≥16 µg/mL, and 0.5 to ≥64 µg/mL, respectively, and 13 resistant azole-resistant Candida isolates were detected. A reduction in the azole MICs leading to the phenotypical reversal of the azole resistance was observed upon addition of efflux-pump inhibitors. These findings suggest that the azole resistance among environmental Candida spp. is most likely associated with the overexpression of efflux-pumps.

Azole resistance in Candida spp. isolated from Catú Lake, Ceará, Brazil: an efflux-pump-mediated mechanism.

Since, there is no study reporting the mechanism of azole resistance among yeasts isolated from aquatic environments; the present study aims to investigate the occurrence of antifungal resistance among yeasts isolated from an aquatic environment, and assess the efflux-pump activity of the azole-resistant strains to better understand the mechanism of resistance for this group of drugs. For this purpose, monthly water and sediment samples were collected from Catú Lake, Ceará, Brazil, from March 2011 to February 2012. The obtained yeasts were identified based on morphological and biochemical characteristics. Of the 46 isolates, 37 were Candida spp., 4 were Trichosporon asahii, 3 were Cryptococcus laurentii, 1 Rhodotorula mucilaginosa, and 1 was Kodamaea ohmeri. These isolates were subjected to broth microdilution assay with amphotericin B, itraconazole, and fluconazole, according to the methodology standardized by the Clinical and Laboratory Standards Institute (CLSI). The minimum inhibitory concentrations (MICs) of amphotericin B, itraconazole, and fluconazole were 0.03125-2µg/mL, 0.0625 to ≥16µg/mL, and 0.5 to ≥64µg/mL, respectively, and 13 resistant azole-resistant Candida isolates were detected. A reduction in the azole MICs leading to the phenotypical reversal of the azole resistance was observed upon addition of efflux-pump inhibitors. These findings suggest that the azole resistance among environmental Candida spp. is most likely associated with the overexpression of efflux-pumps.

Minimum inhibitory concentrations of amphotericin B, azoles and caspofungin against Candida species are reduced by farnesol.

The objective of this study was to evaluate the antifungal activity of farnesol and its interaction with traditional antifungals against drug-resistant strains of Candida species. To do so, we studied the minimum in vitro inhibitory concentration (MIC) of amphotericin B (AMB), fluconazole (FLC), itraconazole (ITC), caspofungin (CAS) and farnesol against 45 isolates of Candida spp., i.e., 24 C. albicans, 16 C. parapsilosis and 5 C. tropicalis through the use of the broth microdilution method. Then, the isolates were tested with the combination of farnesol plus drugs to which they were previously found to be resistant. Additionally, the strains were pre-incubated at sub-inhibitory farnesol concentrations and their antifungal susceptibilities were re-evaluated. We found the MIC values for farnesol varied from 4.68-150 µM for Candida spp., with 19 isolates having a MIC > 1 mg/l, 18 a MIC ≥ 64 mg/l, 35 having a MIC ≥ 1 mg/l and 6 isolates a MIC ≥ 2 mg/l or were resistant to AMB, FLC, ITC and CAS, respectively. Significant MIC reductions were observed when farnesol and antifungal drugs were combined (P < 0.05) and when Candida strains were incubated with farnesol (P < 0.05). We conclude that the in vitro effects of farnesol improved the activity of traditional antifungals to which the Candida spp. isolates were resistant. These results support further investigation of the role of farnesol in the balance of the sterol biosynthetic pathway and how it interferes with cell viability.

Yeast microbiota of raptors: a possible tool for environmental monitoring.

Twenty-two raptors from a rehabilitation centre were evaluated for the presence of yeasts prior to returning them to the wild, and the recovered Candida isolates were tested for in vitro antifungal susceptibility and phospholipase production. Samples were collected from the crop/lower esophagus and cloaca. In vitro antifungal susceptibility and phospholipase production of 21 Candida strains were assessed through broth microdilution and growth on egg yolk agar respectively. Twenty-seven isolates, belonging to seven species, were recovered from 16 tested birds, with C. albicans and C. famata as the most prevalent species. Three out of 21 isolates (2 C. albicans and 1 C. tropicalis) were simultaneously resistant to fluconazole and itraconazole. As for phospholipase production, 8 (8/21) isolates (6 C. albicans, 1 C. famata and 1 C. parapsilosis) showed enzymatic activity. The most relevant finding in this study was the isolation of resistant Candida spp. from wild raptors that had never been submitted to antifungal therapy, which suggests exposure to environmental contaminants. Based on this, we propose the assessment of Candida spp. from the gastrointestinal tract of raptors as a tool for environmental monitoring.

Yeasts from Macrobrachium amazonicum: a focus on antifungal susceptibility and virulence factors of Candida spp.

In the present study, it was sought to compare yeast microbiota of wild and captive Macrobrachium amazonicum and evaluate the antifungal susceptibility and production of virulence factors by the recovered isolates of Candida spp. Additionally, cultivation water was monitored for the presence of fungi. Overall, 26 yeast isolates belonging to three genera and seven species were obtained, out of which 24 were Candida spp., with Candida famata as the most prevalent species for both wild and captive prawns. From cultivation water, 28 isolates of filamentous fungi were obtained, with Penicillium spp., Cladosporium spp. and Aspergillus spp. as the most frequent genera. Eight out of 24 Candida spp. isolates were resistant to azole derivatives, out of which four were recovered from wild-harvested prawns. As for production of virulence factors, three (12.5%) and eight (33.3%) isolates presented phospholipase and protease activity, respectively. This is the first comparative study between wild and captive prawns and the first report on yeast microbiota of M. amazonicum. The most relevant finding was the high percentage of resistant Candida spp., including from wild individuals, which suggests the occurrence of an environmental imbalance in the area where these prawns were captured.