Multiple species of Trichosporon produce biofilms highly resistant to triazoles and amphotericin B.

Isabel Antonieta Iturrieta-González,Ana Carolina Barbosa Padovan,Arnaldo Lopes Colombo,Rosane Christine Hahn,Fernando César Bizerra,David R Andes

doi:10.1371/journal.pone.0109553

Isabel Antonieta Iturrieta-González, Ana Carolina Barbosa Padovan + Show 4 more

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https://doi.org/10.1371/journal.pone.0109553

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Abstract

Invasive infections caused by Trichosporon spp. have increased considerably in recent years, especially in neutropenic and critically ill patients using catheters and antibiotics. The genus presents limited sensitivity to different antifungal agents, but triazoles are the first choice for treatment. Here, we investigated the biofilm production and antifungal susceptibility to triazoles and amphotericin B of 54 Trichosporon spp. isolates obtained from blood samples (19), urine (20) and superficial mycosis (15). All isolates and 7 reference strains were identified by sequence analysis and phylogenetic inferences of the IGS1 region of the rDNA. Biofilms were grown on 96-well plates and quantitation was performed using crystal violet staining, complemented with Scanning Electron Microscopy (SEM). Susceptibility tests for fluconazole, itraconazole, voriconazole and amphotericin B were processed using the microdilution broth method (CLSI) for planktonic cells and XTT reduction assay for biofilm-forming cells. Our results showed that T. asahii was the most frequent species identified (66.7%), followed by T. faecale (11.1%), T. asteroides (9.3%), T. inkin (7.4%), T. dermatis (3.7%) and one T. coremiiforme (1.8%). We identified 4 genotypes within T. asahii isolates (G1, G3, G4 and G5) and 2 genotypes within T. faecale (G1 and G3). All species exhibited high adhesion and biofilm formation capabilities, mainly T. inkin, T. asteroides and T. faecale. Microscopy images of high biofilm-producing isolates showed that T. asahii presented mainly hyphae and arthroconidia, whereas T. asteroides exhibited mainly short arthroconidia and few filaments. Voriconazole exhibited the best in vitro activity against all species tested. Biofilm-forming cells of isolates and reference strains were highly resistant to all antifungals tested. We concluded that levels of biofilm formation by Trichosporon spp. were similar or even greater than those described for the Candida genus. Biofilm-forming cells were at least 1,000 times more resistant to antifungals than planktonic cells, especially to voriconazole.

Highlights

Trichosporon spp. are basidiomycetous yeast-like organisms that are widely distributed in nature, and can generally be isolated from soil, water, decomposing matter, and bird and bat droppings [1]
Trichosporon sp. isolates are classically recognized as a cause of superficial mycoses but have recently emerged as pathogens in invasive infections, in patients with acute leukemia and those subjected to invasive clinical procedures [10,37]
We showed that 6 Trichosporon species from 54 clinical isolates obtained from different types of human infections were able to form robust biofilms in vitro that were not susceptible to triazoles or amphotericin B

Summary

Introduction

Trichosporon spp. are basidiomycetous yeast-like organisms that are widely distributed in nature, and can generally be isolated from soil, water, decomposing matter, and bird and bat droppings [1]. It can be part of the transient human microbiota, mainly in the skin, nails and mucosa of the respiratory and gastrointestinal tracts [1,2]. According to data provided by the ARTEMIS DISK collection, Trichosporon spp. are considered to be the second or third most commonly isolated yeast species in clinical laboratories, representing 5.5 to 10.6% of all isolates [3,4,5]. The genus contains 51 species, 16 of which are able to infect human hosts [1].

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