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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

Scanning electron microscopy of 4 <i>Trichosporon</i> spp. strains grown on catheter surfaces.

<p><b>A</b> and <b>B</b>: Low biofilm producer <i>T. asahii</i> 05-001 (CV-A₅₇₀  =  0.287); <b>C</b> and <b>D</b>: Medium biofilm producer <i>T. asahii</i> 18-001 (CV-A₅₇₀  =  1.557); <b>E</b> and <b>F</b>: High biofilm producer <i>T. asahii</i> 07-001A (CV-A₅₇₀  =  3.337); and <b>G</b> and <b>H</b>: High biofilm producer <i>T. asteroides</i> 13-001 (CV-A₅₇₀  =  2.755).</p

Inter and intra species variation in the biofilm production of 54 <i>Trichosporon</i> spp. clinical isolates and 7 reference strains.

<p><b>A-</b> 19 isolates from blood identified as <i>T. asahii, T. asteroides, T. coremiiforme</i> and <i>T. dermatis</i>. <b>B-</b> 20 isolates from urine identified as <i>T. asahii</i>. <b>C-</b> 15 isolates from superficial mycosis/skin colonization identified as <i>T. asahii, T. dermatis, T. faecale</i> and <i>T. inkin</i>. <b>D-</b> 7 reference strains from CBS obtained from different sources.</p

<i>In vitro</i> activity of 4 antifungal drugs against planktonic cells of 54 clinical isolates of <i>Trichosporon</i> spp. according to the isolation site.

<p>MIC₅₀: Minimal inhibitory concentration capable of inhibiting growth of 50% of isolates.</p><p>MIC₉₀: Minimal inhibitory concentration capable of inhibiting growth of 90% of isolates.</p><p>GM: Geometric Mean.</p><p><i>In vitro</i> activity of 4 antifungal drugs against planktonic cells of 54 clinical isolates of <i>Trichosporon</i> spp. according to the isolation site.</p

Species distribution of 54 <i>Trichosporon</i> spp. clinical isolates according to the site of infection or colonization identifiyed by IGS1 rDNA sequencing.

<p>Species distribution of 54 <i>Trichosporon</i> spp. clinical isolates according to the site of infection or colonization identifiyed by IGS1 rDNA sequencing.</p

Comparative analysis of MIC values obtained against planktonic cells of <i>T. asahii</i> (36 isolates) <i>versus</i> non-<i>T. asahii</i> (18) isolates for all antifungals tested.

<p><b>A-</b> Fluconazole; <b>B-</b> Itraconazole<b>; C-</b> Voriconazole and D- Amphotericin B.</p

<i>In vitro</i> activity of 4 antifungal drugs against biofilm forming cells of 54 clinical isolates of <i>Trichosporon</i> spp.

<p>MIC₅₀: Minimal inhibitory concentration capable of inhibiting growth of 50% of isolates.</p><p>MIC₉₀: Minimal inhibitory concentration capable of inhibiting growth of 90% of isolates.</p><p><i>In vitro</i> activity of 4 antifungal drugs against biofilm forming cells of 54 clinical isolates of <i>Trichosporon</i> spp.</p

Fungal planet description sheets: 716–784

Novel species of fungi described in this study include those from various countries as follows: Australia, Chaetopsina eucalypti on Eucalyptus leaf litter, Colletotrichum cobbittiense from Cordyline stricta × C. australis hybrid, Cyanodermella banksiae on Banksia ericifolia subsp. macrantha, Discosia macrozamiae on Macrozamia miquelii, Elsinoë banksiigena on Banksia marginata, Elsinoë elaeocarpi on Elaeocarpus sp., Elsinoë leucopogonis on Leucopogon sp., Helminthosporium livistonae on Livistona australis, Idriellomyces eucalypti (incl. Idriellomyces gen. nov.) on Eucalyptus obliqua, Lareunionomyces eucalypti on Eucalyptus sp., Myrotheciomyces corymbiae (incl. Myrotheciomyces gen. nov., Myrotheciomycetaceae fam. nov.), Neolauriomyces eucalypti (incl. Neolauriomyces gen. nov., Neolauriomycetaceae fam. nov.) on Eucalyptus sp., Nullicamyces eucalypti (incl. Nullicamyces gen. nov.) on Eucalyptus leaf litter, Oidiodendron eucalypti on Eucalyptus maidenii, Paracladophialophora cyperacearum (incl. Paracladophialophoraceae fam. nov.) and Periconia cyperacearum on leaves of Cyperaceae, Porodiplodia livistonae (incl. Porodiplodia gen. nov., Porodiplodiaceae fam. nov.) on Livistona australis, Sporidesmium melaleucae (incl. Sporidesmiales ord. nov.) on Melaleuca sp., Teratosphaeria sieberi on Eucalyptus sieberi, Thecaphora aus-traliensis in capsules of a variant of Oxalis exilis. Brazil, Aspergillus serratalhadensis from soil, Diaporthe pseudo-inconspicua from Poincianella pyramidalis, Fomitiporella pertenuis on dead wood, Geastrum magnosporum on soil, Marquesius aquaticus (incl. Marquesius gen. nov.) from submerged decaying twig and leaves of unidentified plant, Mastigosporella pigmentata from leaves of Qualea parviflorae, Mucor souzae from soil, Mycocalia aquaphila on decaying wood from tidal detritus, Preussia citrullina as endophyte from leaves of Citrullus lanatus, Queiroziella brasiliensis (incl. Queiroziella gen. nov.) as epiphytic yeast on leaves of Portea leptantha, Quixadomyces cearen-sis (incl. Quixadomyces gen. nov.) on decaying bark, Xylophallus clavatus on rotten wood. Canada, Didymella cari on Carum carvi and Coriandrum sativum. Chile, Araucasphaeria foliorum (incl. Araucasphaeria gen. nov.) on Araucaria araucana, Aspergillus tumidus from soil, Lomentospora valparaisensis from soil. Colombia, Corynespora pseudocassiicola on Byrsonima sp., Eucalyptostroma eucalyptorum on Eucalyptus pellita, Neometulocladosporiella eucalypti (incl. Neometulocladosporiella gen. nov.) on Eucalyptus grandis × urophylla, Tracylla eucalypti (incl. Tracyllaceae fam. nov., Tracyllalales ord. nov.) on Eucalyptus urophylla. Cyprus, Gyromitra anthracobia (incl. Gyromitra subg. Pseudoverpa) on burned soil. Czech Republic, Lecanicillium restrictum from the surface of the wooden barrel, Lecanicillium testudineum from scales of Trachemys scripta elegans. Ecuador, Entoloma yanacolor and Saproamanita quitensis on soil. France, Lentithecium carbonneanum from submerged decorticated Populus branch. Hungary, Pleuromyces hungaricus (incl. Pleuromyces gen. nov.) from a large Fagus sylvatica log. Iran, Zymoseptoria crescenta on Aegilops triuncialis. Malaysia, Ochroconis musicola on Musa sp. Mexico, Cladosporium michoacanense from soil. New Zealand, Acrodontium metrosideri on Metrosideros excelsa, Polynema podocarpi on Podocarpus totara, Pseudoarthrographis phlogis (incl. Pseudoarthrographis gen. nov.) on Phlox subulata. Nigeria, Coprinopsis afrocinerea on soil. Pakistan, Russula mansehraensis on soil under Pinus roxburghii. Russia, Baoran­ gia alexandri on soil in deciduous forests with Quercus mongolica. South Africa, Didymocyrtis brachylaenae on Brachylaena discolor. Spain, Alfaria dactylis from fruit of Phoenix dactylifera, Dothiora infuscans from a blackened wall, Exophiala nidicola from the nest of an unidentified bird, Matsushimaea monilioides from soil, Terfezia morenoi on soil. United Arab Emirates, Tirmania honrubiae on soil. USA, Arxotrichum wyomingense (incl. Arxotrichum gen. nov.) from soil, Hongkongmyces snookiorum from submerged detritus from a fresh water fen, Leratiomyces tesquorum from soil, Talaromyces tabacinus on leaves of Nicotiana tabacum. Vietnam, Afroboletus vietnamensis on soil in an evergreen tropical forest, Colletotrichum condaoense from Ipomoea pes-caprae. Morphological and culture characteristics along with DNA barcodes are provided. © 2018 Naturalis Biodiversity Center & Westerdijk Fungal Biodiversity Institute