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

Iturrieta-González IA, Padovan AC, Bizerra FC, Hahn RC, Colombo AL - PLoS ONE (2014)

Bottom Line: Voriconazole exhibited the best in vitro activity against all species 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.

View Article: PubMed Central - PubMed

Affiliation: Laboratório Especial de Micologia, Disciplina de Infectologia, Universidade Federal de São Paulo, São Paulo, SP, Brazil.

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.

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Inter and intra species variation in the biofilm production of 54 Trichosporon spp. clinical isolates and 7 reference strains.A- 19 isolates from blood identified as T. asahii, T. asteroides, T. coremiiforme and T. dermatis. B- 20 isolates from urine identified as T. asahii. C- 15 isolates from superficial mycosis/skin colonization identified as T. asahii, T. dermatis, T. faecale and T. inkin. D- 7 reference strains from CBS obtained from different sources.
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pone-0109553-g002: Inter and intra species variation in the biofilm production of 54 Trichosporon spp. clinical isolates and 7 reference strains.A- 19 isolates from blood identified as T. asahii, T. asteroides, T. coremiiforme and T. dermatis. B- 20 isolates from urine identified as T. asahii. C- 15 isolates from superficial mycosis/skin colonization identified as T. asahii, T. dermatis, T. faecale and T. inkin. D- 7 reference strains from CBS obtained from different sources.

Mentions: The total biofilm mass of 54 clinical isolates and 7 reference strains of Trichosporon spp. was then assessed by CV staining after 48 h growth. Absorbance values ranged from 0.109 to 3.337. To correlate the capability for biofilm production with the different species tested and isolation sites, we arbitrarily established three categories (33.3 percentiles) of biofilm production based on the total range of the CV quantifications exhibited by the 61 isolates, categorizing the producers as follows: low: A570<1, medium: A570≥1.01 to 2.499 or high: A570≥2.5. Figure 2 illustrates the biofilm production by all strains tested according to the isolation source and species distribution. Trichosporon sp. strains were scaled from the highest to the lowest biofilm producer as T. inkin> T. asteroides ≈ T. faecale> T. asahii. The two T. dermatis isolates (classified as medium and high biofilm producers) and the single T. coremiiforme isolate (high biofilm producer) were not included in this rank scale due to the low number of isolates tested. When considering the source of isolation, 93.3% of the Trichosporon spp. strains isolated from skin were categorized as high or medium biofilm producers. Otherwise, 65% to 73.7% of the strains isolated from blood and urine, respectively, were considered high or medium biofilm producers. The reference strains were characterized as low (T. asahii CBS2479, T. mucoides CBS7625 and T. faecale CBS4828), medium (T. asahii CBS7631 and T. ovoides CBS7556) or high (T. dermatis CBS2043 and T. inkin CBS5585) biofilm producers.


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

Iturrieta-González IA, Padovan AC, Bizerra FC, Hahn RC, Colombo AL - PLoS ONE (2014)

Inter and intra species variation in the biofilm production of 54 Trichosporon spp. clinical isolates and 7 reference strains.A- 19 isolates from blood identified as T. asahii, T. asteroides, T. coremiiforme and T. dermatis. B- 20 isolates from urine identified as T. asahii. C- 15 isolates from superficial mycosis/skin colonization identified as T. asahii, T. dermatis, T. faecale and T. inkin. D- 7 reference strains from CBS obtained from different sources.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4215839&req=5

pone-0109553-g002: Inter and intra species variation in the biofilm production of 54 Trichosporon spp. clinical isolates and 7 reference strains.A- 19 isolates from blood identified as T. asahii, T. asteroides, T. coremiiforme and T. dermatis. B- 20 isolates from urine identified as T. asahii. C- 15 isolates from superficial mycosis/skin colonization identified as T. asahii, T. dermatis, T. faecale and T. inkin. D- 7 reference strains from CBS obtained from different sources.
Mentions: The total biofilm mass of 54 clinical isolates and 7 reference strains of Trichosporon spp. was then assessed by CV staining after 48 h growth. Absorbance values ranged from 0.109 to 3.337. To correlate the capability for biofilm production with the different species tested and isolation sites, we arbitrarily established three categories (33.3 percentiles) of biofilm production based on the total range of the CV quantifications exhibited by the 61 isolates, categorizing the producers as follows: low: A570<1, medium: A570≥1.01 to 2.499 or high: A570≥2.5. Figure 2 illustrates the biofilm production by all strains tested according to the isolation source and species distribution. Trichosporon sp. strains were scaled from the highest to the lowest biofilm producer as T. inkin> T. asteroides ≈ T. faecale> T. asahii. The two T. dermatis isolates (classified as medium and high biofilm producers) and the single T. coremiiforme isolate (high biofilm producer) were not included in this rank scale due to the low number of isolates tested. When considering the source of isolation, 93.3% of the Trichosporon spp. strains isolated from skin were categorized as high or medium biofilm producers. Otherwise, 65% to 73.7% of the strains isolated from blood and urine, respectively, were considered high or medium biofilm producers. The reference strains were characterized as low (T. asahii CBS2479, T. mucoides CBS7625 and T. faecale CBS4828), medium (T. asahii CBS7631 and T. ovoides CBS7556) or high (T. dermatis CBS2043 and T. inkin CBS5585) biofilm producers.

Bottom Line: Voriconazole exhibited the best in vitro activity against all species 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.

View Article: PubMed Central - PubMed

Affiliation: Laboratório Especial de Micologia, Disciplina de Infectologia, Universidade Federal de São Paulo, São Paulo, SP, Brazil.

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.

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Related in: MedlinePlus