Black yeast habitat choices and species spectrum on high altitude creosote-treated railway ties

Polyextremotolerant black yeast-like fungi thrive in moderately hostile environments where they are concomitantly subjected to several types of stress, such as toxicity, scarce nutrient availability, and high or low temperature extremes. Their ability to assimilate alkylbenzenes (toxic environmental pollutants) enhances their growth in harsh conditions, including on railway ties. Samples were collected using cotton swabs, premoistened with physiological saline, from 658 oak and concrete railway ties at six train stations in Turkey at altitudes ranging between 1026 and 1427 m. The samples were inoculated on malt extract agar supplemented with chloramphenicol, and incubated at 26 °C for 4 weeks. Twenty-four samples (3.6 %), 17 from oak and 7 from concrete (5.6 % vs. 2 %; P = 0.02), tested positive for fungi. Exophiala crusticola was found to be the most common species (n = 13), followed by Exophiala phaeomuriformis (n = 7) and Exophiala heteromorpha (n = 4). These results suggest that hydrocarbons, particularly creosote-treated oak woods, support the growth of black yeasts, some of which are opportunists in humans

Investigation of the Mating-Type Gene Locus of Candida albicans

Amaç: Askomiset mantarlarda çiftleşme, MAT lokus adı verilen bir eşey tipi gen bölge- si ile yönetilir. MAT lokus, zıt iki eşey tipinde birbirine benzemeyen hatta tamamen farklı idiomorf dizilimler içerir. Pezizomycotina alt şubesine dâhil olan mantarlarda her bir MAT idiomorfu, bir alfa domain, bir de HMG (High Mobility Group) domain trans- kripsiyon faktörünü kodlar. Alfa domain ve HMG domain transkripsiyon faktör genleri MAT lokusun olmazsa olmaz parçaları olsa da her bir mantarın MAT lokus yapısı, yani uzunluğu, gen içeriği ve genlerin transkripsiyon yönleri kendilerine özgüdür. Bu çalış- mada, laboratuvarlarımızın kültür koleksiyonunda bulunan ve kan kültürlerinden izole edilen Candida albicans’ın eşey tipi gen bölgesinin karakterizasyonunun belirlenmesi amaçlanmıştır. Yöntem: Candida albicans genomunda yer alan MTLa (Mating Type Locus)ve MTL? primerlerinin 485 bp ve 515 bp’lik DNA bölgesi multiplex PCR tekniği ile belirlendi. Bulgular: Toplam, 156 C. albicans kökeninin 155 (%99.4)’i MTLa/?, 1 (%0.6) köken ise MTL?/? olarak bulundu. Sonuç: MTL karakterizasyonun belirlenmesi çalışmaları hem moleküler genetik çalış- malarının daha kolay ve çabuk yapılabilmesi hem de bu patojen mantarın doğasının anlaşılması yönünden önemlidirObjective: Mating in Ascomycete fungi is governed by mating type gene region called the MAT locus. the MAT locus contains idiomorphic sequences that are dissimilar in two opposite mating types.In fungi belonging to the Pezizomycotina subphylum, each MAT idiomorph encodes an alpha domain and an HMG (High Mobility Group) domain transcription factor. Although the alpha domain and the HMG domain transcription factor genes are indispensable components, the structure of the MAT locus (length), the gene content, and the transcriptional directions of the genes differ among fungi. This study aimed to evaluate the characterization of mating type of Candida albicans isolated from blood samples in our labouratory culture collection. Method: Using the multiplex PCR technique, 486 bp and 515 bp of the DNA region of MTLa and MTL? primers in the C. albicans genome were determined, respectively. Results: 156 C. albicans isolates showed 155 MTLa/? (99.4%) and 1 MTL?/? (0.6%). Conclusion: Determination of MTL is essential both for making molecular genetic studies more efficient and for understanding the nature of this pathogenic fungus

In vitro activities of antifungal drugs against environmental Exophiala isolates and review of the literature

Exophiala is a genus of black fungi isolated worldwide from environmental and clinical specimens. Data on antifungal susceptibility of Exophiala isolates are limited and the methodology on susceptibility testing is not yet standardised. In this study, we investigated in vitro antifungal susceptibilities of environmental Exophiala isolates. A total of 87 Exophiala isolated from dishwashers or railway ties were included. A CLSI M38-A2 microdilution method with modifications was used to determine antifungal susceptibility for fluconazole, voriconazole, posaconazole, itraconazole, amphotericin B and terbinafine. Minimum inhibitory concentration (MIC) values were determined visually at 48 hours, 72 hours and 96 hours. MIC-0 endpoint (complete inhibition of growth) was used for amphotericin B and azoles, except fluconazole, for which MIC-2 endpoint (~50% inhibition compared to growth control) was used. Both MIC-0 and MIC-1 (~80% inhibition compared to growth control) results were analysed for terbinafine to enable comparison with previous studies. Fungal growth was sufficient for determination of MICs at 48 hours for all isolates except two Exophiala dermatitidis strains. At 72 hours, most active antifungal agents according to GM MIC were voriconazole and terbinafine, followed by posaconazole, itraconazole and amphotericin B in rank order of decreasing activity. While amphotericin B displayed adequate in vitro activity despite relatively high MICs, fluconazole showed no meaningful antifungal activity against Exophiala

<i>MTL</i> genotypes, phenotypic switching, and susceptibility profiles of <i>Candida parapsilosis</i> species group compared to <i>Lodderomyces elongisporus</i>

<div><p>Reference isolates of <i>Candida parapsilosis</i> (n = 8), <i>Candida metapsilosis</i> (n = 6), <i>Candida orthopsilosis</i> (n = 7), and <i>Lodderomyces elongisporus</i> (n = 11) were analyzed to gain insight into their pathobiology and virulence mechanisms. Initial evaluation using BBL Chromagar Candida medium misidentified <i>L</i>. <i>elongisporus</i> isolates as <i>C</i>. <i>albicans</i>. Polymerase chain reaction analysis of isolate <i>MTL</i> idiomorphs revealed that all <i>C</i>. <i>parapsilosis</i> isolates were <i>MTL</i><b>a</b> homozygous and no <i>MTL α</i>1, <i>α</i>2, <b>a</b>1, or <b>a</b>2 gene was detected in <i>L</i>. <i>elongisporus</i> isolates. For <i>C</i>. <i>orthopsilosis</i>, two isolates were <i>MTL</i><b>a</b> homozygous and five were <i>MTL</i>-heterozygous. Similarly, one <i>C</i>. <i>metapsilosis</i> isolate was <i>MTLα</i> homozygous whereas five were <i>MTL</i>-heterozygous. Isolate phenotypic switching analysis revealed potential phenotypic switching in the <i>MTLα</i> homozygous <i>C</i>. <i>metapsilosis</i> isolate, resulting in concomitant elongated cell formation. Minimum inhibitory concentrations of fluconazole (FLC) and FK506, alone or in combination, were determined by checkerboard assay, with data analyzed using the fractional inhibitory concentration index model. Synergistic or additive effects of these compounds were commonly observed in <i>C</i>. <i>parapsilosis</i> and <i>L</i>. <i>elongisporu</i>s isolates. No killer activity was observed in the studied isolates, as determined phenotypically. No significant difference in virulence was seen for the four species in a <i>Galleria mellonella</i> model (<i>P</i> > 0.05). In conclusion, our results demonstrated phenotypic switching of <i>C</i>. <i>metapsilosis</i> CBS 2315 and that FLC and FK506 represent a promising drug combination against <i>C</i>. <i>parapsilosis</i> and <i>L</i>. <i>elongisporu</i>s. The findings of the present study contribute to our understanding of the biology, diagnosis, and new possible treatments of the <i>C</i>. <i>parapsilosis</i> species group and <i>L</i>. <i>elongisporu</i>s.</p></div

Phenotypic switching test with <i>Candida metapsilosis</i> CBS 2315 (MAT <i>α</i>/<i>α</i>).

<p><b>(a)</b> Pink colonies indicating phenotypic switching (red box), grown at 30°C for 7 d on SC medium containing 5 μg/mL phloxine B; <b>(b)</b> elongated cells analyzed by scanning electron microscopy (Scale bar, 10 μm).</p

<i>MTL</i> genotypes, phenotypic switching, and susceptibility profiles of <i>Candida parapsilosis</i> species group compared to <i>Lodderomyces elongisporus</i> - Fig 2

<p>Determination of the <i>MTL</i> genotypes by PCR, <b>(a)</b> <i>Candida parapsilosis</i> isolates (1–8 referring to CBS 8836, CBS 7248, CBS 2915, CBS 604, CBS 2216, CBS 8181, CBS 125.41, and CBS 1954), the 425 bp <i>MTL</i><b>a</b>1 and the 495 bp <i>MTL</i><b>a</b>2 products were obtained for all isolates; <b>(b1</b> and <b>b2)</b> <i>Lodderomyces elongisporus</i> isolates (1–11 referring to the isolates 7660, 7661, 7663, 7665, 7666, 7668, 7669, 7670, 7672, 7673, and 7675), the 933 bp PCR product obtained for all isolates indicates the absence of any <i>MTL</i> transcription factor gene in between <i>PIK</i><b>a</b> and orf19.3202; <b>(c)</b> <i>Candida orthopsilosis</i> isolates (1–7 referring to CBS 107.41, CBS 107.42, CBS 109.06, CBS 8825, CBS 107.43, CBS 9894, and CBS 2212) were screened with primers designed based on <i>C</i>. <i>orthopsilosis</i> type 1 <b>(c1)</b> and type 2 <i>MTL</i> sequences <b>(c2)</b>; both primer pairs worked for the isolates although band brightness’ varied owing to differences among type 1 and type 2 sequences. With CBS 107.41 and CBS 107.42, only <i>MTL</i><b>a</b>1 and <i>MTL</i><b>a</b>2 PCR products were obtained indicating that these isolates are <i>MTL</i><b>a</b> homozygotes; the other isolates were found to be heterozygous for <i>MTL</i>; <b>(d)</b> <i>Candida metapsilosis</i> isolates (1–6 referring to CBS 2315, CBS 107.47, CBS 109.07, CBS 111.27, CBS 1046, and CBS 2916), whereas only <i>MTLα</i>1 (150 bp) and <i>MTLα</i>2 (188 bp) PCR products were obtained with CBS 2315 indicating <i>MTLα</i> homozygosity; all <i>MTL</i> gene products were obtained for the other isolates showing that they are <i>MTL</i> heterozygous. M, Marker; NC, Negative control.</p

<i>Galleria mellonella</i> survival curves after larval infection with the indicated <i>Candida parapsilosis</i> (blue line), <i>Candida orthopsilosis</i> (green line), <i>Candida metapsilosis</i> (purple line), and <i>Lodderomyces elongisporus</i> (turquoise line) species, or with PBS as a control (red line).

<p>A total of 1 × 10⁶ cells were used to infect the larvae and animal survival was observed at 37°C. All study isolates were tested.</p

The effect of Tween-80 on the differentiation of Trichophyton mentagrophytes and Trichophyton rubrum strains with FT-IR spectroscopy

Mikrobiyoloji laboratuvarlarında Trichophyton mentagrophytes ve Trichophyton rubrum en sık tanımlanan iki dermatofit türüdür. Her ne kadar yeni teknolojiler dermatofitlerin tür düzeyinde tanımlanmasına yardımcı olabiliyorsa da, bu yöntemlerin doğrudan uygulanmasında kabul edilebilir sonuçlar için geliştirilmelerine ihtiyaç vardır. FT-IR spektroskopisi ile yapılan önceki araştırmalar, bazı kısıtlılıklarının bulunmasıyla birlikte, bu yöntemin dermatofitlerin tanımlanmasında kullanılabileceğini göstermektedir. Küf mantarlarının ökaryotik karmaşık yapılarından dolayı özellikle FT-IR spektrumdaki organik bağ bölgesi oldukça düzensizlikler göstermektedir. Bu çalışmada, inorganik bir molekül olan Tween-80’in, dermato-fitlerin üreme ortamına ilave edilmesiyle, dermatofitlerin FT-IR spektroskopi analizine etkisi incelenmiştir. Çalışmada, toplam dokuz referans dermatofit kökeni [5 T.mentagrophytes kompleks (T.asteroides CBS 424.63, T.erinacei CBS 344.79, T.erinacei CBS 511.73, T.erinacei CBS 677.86, T.mentagrophytes CBS 110.65) ve farklı morfotipte 4 T.rubrum kompleks (T.fluviomuniense CBS 592.68, T.kuryangei CBS 422.67, T.raubitschekii CBS 102856, T.rubrum CBS 392.58)] incelenmiştir. Tüm kökenler %1 Tween-80 içeren ve içermeyen Sabouraud glukoz agarda üç hafta süreyle çoğaltılmış; inkübasyon sonunda, yüzeyden top- lanan her bir dermatofit kökeni FT-IR spektroskopisi ile incelenmiştir. Tüm ölçümler 4400 ile 400 cm?1 aralığında transmisyon modunda yapılmıştır. Çok sayıda spektral pencere bölgesi, temel bileşenler analizi ve hiyerarşik kümeleme ile incelenerek değerlendirilmiştir. T.mentagrophytes kompleks ve T.rubrum kompleks, beş farklı bölge spektrumlarının ikinci dereceden türevlerin birlikte değerlendirilmesiyle kolaylıkla ayrı olarak gruplandırılmıştır. Çalışmanın diğer bir bulgusu da, Tween-80 ile inkübasyon sonrasında tüm T.mentagrophytes kökenlerinin küf yapılarında lipid bağ bileşimlerinin saptanması olmuş (p= 0.025); bu bağlar T.rubrum kökenlerinde belirlenememiştir (p= 0.269). Sonuç olarak, T.mentagrophytes kompleks kökenlerinin T.rubrum kompleks kökenlerinden FT-IR spektroskopi ile ayrılabilmesi için, kültür ortamına Tween-80 eklenmesinin yeterli olduğu belirlenmiş; bunun nedeninin, kültür ortamından T.mentagrophytes kökenlerinin hücre yapısına geçen lipid bileşikler olduğu düşünülmüştür. Tween-80’in, T.mentagrophytes ve T.rubrum kompleks kökenlerinin FT-IR spektroskopi ile ayrımındaki bu olumlu etkisinin desteklenmesi için, gerek daha fazla sayıda referans köken, gerekse antifungal ilaçlar ve inorganik iyonlar gibi farklı çevresel etkenlerle karşılaşan klinik kökenler ile yapılacak ileri araştırmalara ihtiyaç duyulmaktadır.Trichophyton mentagrophytes and Trichophyton rubrum, are two of the frequently identified dermatophyte species in routine microbiology laboratories. Although newer technologies may assist in species-level identification, direct application of these methods usually require improvement in order to obtain reliable identification of these species. Earlier data have shown that dermatophytes may be identified with FT-IR spectroscopy although there are some limitations. In particular, the organic bond ranges in FT-IR spectra showed more irregularity because of the eucaryotic complexity of the molds. In this study, Tween-80 which is an inorganic molecule, was added to the dermatophyte growth medium in order to investigate its effect on FT-IR spectroscopy analysis of dermatophytes. Nine reference dermatophyte strains [5 T.mentagrophytes complex (T.asteroides CBS 424.63, T.erinacei CBS 344.79, CBS 511.73, CBS 677.86, T.mentagrophytes CBS 110.65) and 4 T.rubrum complex strains with different morphotypes (T.fluviomuniense CBS 592.68, T.kuryangei CBS 422.67, T.raubitschekii CBS 102856, T.rubrum CBS 392.58)] were included in the study. All strains were cultured on Sabouraud glucose agar either with or without 1% Tween-80 for three weeks. After the incubation period, superficial scrapings from each dermatophyte colony were analyzed using FT-IR spectroscopy. All measurements were performed in transmission mode between 4400 and 400 cm&amp;#8722;1. Numerous spectral window data were analyzed by principal component analysis and hierarchical cluste- ring was performed. The second derivations of spectral ranges revealed clear grouping of T.mentagrophytes complex and T.rubrum complex in association over five separate spectral ranges. The findings also showed that while all of the T.mentagrophytes strains contained lipid compounds in their mold structure after Tween-80 incubation (<0.025), T.rubrum strains did not. Based on these results, it was concluded that culture medium containing Tween-80 was sufficient to enable differentiation of T.mentagrophytes complex from T.rubrum complex by FT-IR spectroscopy. This effect might be attributed to the possible transfer of lipid compounds from culture to cell structure during growth. Further studies with the use of large number of reference strains and clinical isolates exposed to different environmental factors, such as antifungal agents and inorganic ions, are needed to support these data indicating favorable effect of Tween-80 on the differentiation of T.mentagrophytes and T.rubrum complexes by FT-IR spectroscopy

Synergy testing of fluconazole (FLU) and FK506 against <i>Candida parapsilosis</i> species group and <i>Lodderomyces elongisporus</i> strains.

<p>Synergy testing of fluconazole (FLU) and FK506 against <i>Candida parapsilosis</i> species group and <i>Lodderomyces elongisporus</i> strains.</p

<i>Candida albicans</i> SC 5314, <i>Candida tropicalis</i> YJM 57, <i>Lodderomyces elongisporus</i> 7660, <i>Candida metapsilosis</i> CBS 2315, <i>Candida orthopsilosis</i> CBS 107.41, and <i>Candida parapsilosis</i> CBS 2315 were grown on BBL Chromagar Candida medium at 37°C for 3 d and photographed.

<p><i>Candida albicans</i> SC 5314, <i>Candida tropicalis</i> YJM 57, <i>Lodderomyces elongisporus</i> 7660, <i>Candida metapsilosis</i> CBS 2315, <i>Candida orthopsilosis</i> CBS 107.41, and <i>Candida parapsilosis</i> CBS 2315 were grown on BBL Chromagar Candida medium at 37°C for 3 d and photographed.</p