Phylogenomic delineation of two new species of ascomycetous yeasts, Wickerhamiella koratensis sp. nov. and Wickerhamiella limtongiae sp. nov., and proposal of two synonyms, Wickerhamiella infanticola and Wickerhamiella tropicalis.

Two novel ascomycetous yeast species of the genus Wickerhamiella are proposed based on isolates obtained in Thailand from food waste and the fruiting body of a polypore fungus, and on a combination of conventional DNA-barcode sequence analyses and whole-genome phylogenies. We focus on a particular subclade of the genus Wickerhamiella that contains species found in anthropic environments and describe Wickerhamiella limtongiae sp. nov. (DMKU-FW31-5T=PYCC 9022T=TBRC 15055T), found on food waste samples. In an adjacent clade, we describe Wickerhamiella koratensis sp. nov. (DMKU-KO16T=PYCC 8908T=TBRC 14869T), which represents the closest relative of Wickerhamiella slavikovae and was isolated from the fruiting body of Bjerkandera sp. In the subclade of W. limtongiae sp. nov., we propose that Wickerhamiella infanticola should be regarded as a synonym of Wickerhamiella sorbophila and that Wickerhamiella tropicalis should be regarded as a synonym of Wickerhamiella verensis.

Cystofilobasidium josepaulonis sp. nov., a novel basidiomycetous yeast species.

A yeast strain belonging to the basidiomycetous yeast genus Cystofilobasidium was isolated from a marine sediment sample collected in an intertidal zone in Shandong province, PR China. The results of phylogenetic analyses based on sequences of the D1/D2 domain of the 26S ribosomal RNA gene and the internal transcribed spacer (ITS) region indicate that this strain, together with three other strains isolated from basal ice collected in Norway, the gut of an insect and an alga collected in Russia, represent a novel species of the genus, for which the name Cystofilobasidium josepaulonis sp. nov. (holotype strain CGMCC 2.6672T) is proposed. The novel species differs from the known species of the genus Cystofilobasidium by 1.7 %-4.1 and 11.3 %-17.1 % mismatches in the D1/D2 domain and the ITS region, respectively. This species forms teliospores on potato dextrose agar (PDA) and 10 % V8 juice agar, but teliospore germination with basidia was not observed.

Pichia bovicola sp. nov., a yeast species associated with small-intestinal content of cattle.

Three yeast strains, DMKU-MP6-4T, DMKU-MP2-6 and DMKU-MP5-1, were isolated from the small-intestinal content or Pia of cattle in Thailand during the investigation of yeast diversity in this habitat. According to the D1/D2 domain of the large subunit (LSU) rRNA gene and internal transcribed spacer (ITS) region sequence analysis, these strains represent a novel yeast species in the genus Pichia. The species produced one to four ascospores per ascus with spherical to ellipsoidal shape and heterogenous in terms of size. These three strains were identical and differed from their closely related species, Pichia exigua NRRL Y-10920T by 2% (six nucleotide substitutions and five gaps) in the D1/D2 domain of the large subunit rRNA gene, while ITS region sequences differed by 3.1% (16 nucleotide substitutions and 27 gaps), 3.7% (19 nucleotide substitutions and 28 gaps) and 3.1% (16 nucleotide substitutions and 27 gaps) for DMKU-MP6-4T, DMKU-MP2-6 and DMKU-MP5-1, respectively. The name, Pichia bovicola, is proposed to accommodate these species. The holotype is DMKU-MP6-4T (TBRC 15616T=PYCC 8905T).

Evaluation of various phenotypic methods for differentiation of Candida dubliniensis from Candida albicans.

INTRODUCTION: Candida dubliniensis was first identified by Sullivan et al. (1995) in Dublin, Ireland. Its clinical significance is associated with development of fluconazole-resistance and invasive diseases in immunocompromised hosts. C. dubliniensis share many features with C. albicans so has been overlooked and misidentified for a long time. AIMS: Evaluation of various phenotypic tests with polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) as a gold standard to find out the best method/methods for identifying C. dubliniensis. MATERIALS AND METHODS: First PCR-RFLP was performed on 186C. albicans and 14C. dubliniensis strains and then five phenotypic tests were performed simultaneously on all the strains. RESULTS: The results of salt tolerance test at 48 h, colony color on HiCrome candida differential agar (HCDA) at 72 h, heat tolerance test at 48 h, xylose assimilation using discs at 72 h and growth on xylose based agar medium (XAM) at 48 h are completely concordant with PCR-RFLP. Colony color on Tobacco agar could differentiate accurately 100% test strains while peripheral hyphal fringes and chlamydosporulation on this agar was seen in only 86% and 87% respectively. Our routine methods proved to be cost effective than PCR-RFLP but the turnaround time was same or more than PCR-RFLP. CONCLUSION: For routine identification of C. dubliniensis we recommend use of colony color on HCDA and growth on XAM as simple, reliable and inexpensive method.

Perception of fungi by farmers in the Cerrado / Percepção de agricultores do Cerrado sobre os fungos

Fungi are important in several aspects of human life. In particular, to agriculture, pathogenic fungi are of great importance, as they are responsible for production losses of the most diverse types. Because of this, knowledge about pathogenic fungus is of extreme importance for farmers and professionals working in agricultural areas. Among farmers who use specific agroecological practices, this knowledge is even more valuable, since by not adopting conventional methods of production, they resort to non-invasive alternatives that are less or not harmful at all to the environment in consideration of production management methods. This study aimed to assess farmer perception in the Cerrado biome in the city of Goiás (GO), Brazil, in order to understand their ethnomycological perceptions to verify historical management practices, their knowledge about phytopathogenic fungi, and how these producers perceive fungi. We used the theoretical reference method "From peasant to peasant" formulated by ANPA - National Association of Small Farmers. Some aspects of farmers' ethnomycological knowledge are discussed. These ease identification the representatives of the Fungi Kingdom is associated with organisms that present easily recognizable characteristics, such as wood-ears or disease-causing fungi. In general, farmers are able to identify representatives of the Fungi Kingdom that are found in their daily lives. The perception of farmers about fungi, a group still much unknown by society, is very relevant for future actions of ethnomycology.

Percepção dos fungos por agricultores do Cerrado – Fungos são organismos importantes em vários aspectos da vida humana. Em particular, para a agricultura, fungos patogênicos são de grande importância, pois são responsáveis por perdas de produção dos mais diversos tipos. Por isso, o conhecimento sobre fungos patogênicos é de extrema importância para agricultores e profissionais que trabalham em áreas agrícolas. Entre os agricultores que utilizam práticas agroecológicas, esse conhecimento é ainda mais valioso, pois, ao não adotar métodos convencionais de produção, recorrem a alternativas não invasivas que são menos ou não prejudiciais ao meio ambiente, considerando os métodos de gerenciamento da produção. Este estudo teve como objetivo avaliar a percepção de um grupo de agricultores do bioma Cerrado, na cidade de Goiás (GO), Brasil, a fim de compreender suas percepções etnomicológicas, de modo a verificar práticas históricas de manejo, seu conhecimento sobre fungos fitopatogênicos e como esses produtores percebem os fungos, de modo geral. Utilizamos o método "de camponês para camponês", formulado pela ANPA – Associação Nacional de Pequenos Agricultores. Discutimos alguns aspectos do conhecimento etnomicológico destes agricultores. A facilidade de identificação de representantes fúngicos está associada a organismos que apresentam características facilmente reconhecíveis, como estereótipo orelha-de-pau ou fungos causadores de doenças em cultivares. Em geral, os agricultores do estudo são capazes de identificar representantes do Reino Fungi que são encontrados em suas vidas cotidianas. A percepção dos agricultores sobre os fungos, um grupo ainda muito desconhecido pela sociedade, é muito relevante para ações futuras dentro da etnomicologia.

Optimized methodology for obtention of high-yield and -quality RNA from the mycelium of the bioluminescent fungus Neonothopanus gardneri.

Neonothopanus gardneri, also known as coconut flower mushroom (flor-de-coco), is a Brazilian bioluminescent basidiomycete found in Palm Forest, a transitional biome between the Amazonian Forest and Caatinga (Savanna-like vegetation) in Northeast Brazil, especially in Piauí State. Recent advances toward the elucidation of fungal bioluminescence have contributed to the discovery of four genes (hisps, h3h, luz and cph) involved with the bioluminescence process, the so-called Caffeic Acid Cycle (CAC) and to develop biotechnological applications such autoluminescent tobacco plants and luciferase-based reporter genes. High-yield and -quality RNA-extraction methods are required for most of these purposes. Herein, four methods for RNA isolation from the mycelium of N. gardneri were evaluated: RNeasy® kit (QIAGEN), TRI+, TRI18G+, and TRI26G+. Highest RNA yield was observed for TRI18G+ and TRI26G+ methods, an increase of ~130% in comparison to the RNeasy® method and of ~40% to the TRI+ protocol. All the RNA samples showed good purity and integrity, except by gDNA contamination in RNA samples produced with the RNeasy® method. High quality of RNA samples was confirmed by successful cDNA synthesis and PCR amplification of the coding sequence of h3h gene, responsible for the hydroxylation of the precursor of fungal luciferin (3-hydroxyhispidin). Similarly, RT-qPCR amplification of ef-tu gene, related to the protein biosynthesis in the cell, was demonstrated from RNA samples. This is the first report of a reproducible, time-saving and low-cost optimized method for isolation of high-quality and -yield, DNA-free RNA from a bioluminescent fungus, but that can also be useful for other basidiomycetes.

Early diagnosis of Cryptococcus neoformans var. grubii meningitis using multiplex PCR assay in an immunocompetent patient.

Cryptococcosis is an invasive fungal infection that mainly affects the lungs and central nervous system. While patients with cell-mediated immunodeficiency are at high risk of developing cryptococcosis, there have been increasing reports of cryptococcosis in immunocompetent individuals with no underlying conditions. Herein, we report a case of cryptococcal meningitis in a 55-year-old apparently immunocompetent man with a history of heavy alcohol consumption. Although the patient was initially treated for tuberculous meningitis and varicella-zoster virus induced vasculopathy due to a history of exposure to tuberculosis and a presence of stroke, a multiplex polymerase chain reaction (PCR) assay of cerebrospinal fluid (CSF) identified Cryptococcus species unexpectedly, enabling swift treatment and a favorable clinical outcome. The multiplex PCR assay, which can identify multiple pathogens simultaneously and instantly, may lead to early diagnosis and treatment by detecting unanticipated pathogens. Furthermore, the strain was identified through multilocus sequence typing (MLST) analysis as Cryptococcus neoformans var. grubii, Sequence Type 5, molecular type: VNI. Although simplified microbial identification techniques such as mass spectrometry have recently been developed, molecular biological assays are still essential for the accurate identification of infectious strains.

The Role of Corneal Donor Rim Fungal Cultures: Pro Side.

ABSTRACT: Fungal infection after corneal transplantation is a rare but potentially devastating complication. It is of paramount concern for transplant surgeons and the eye banking community. The value of universal corneal rim cultures for keratoplasty remains controversial. In 2016, The Eye Bank Association for America reported an increasing trend in the incidence of post keratoplasty fungal infections and a higher incidence of post keratoplasty [penetrating keratoplasty and endothelial keratoplasty (EK)] fungal endophthalmitis cases. This increasing trend in rate over time from previous Eye Bank Association for America reports was disproportionately associated with EK and Candida species. Additionally, several studies confirmed a high correlation between positive corneoscleral donor rim fungal cultures and postoperative infections, and a higher risk to the mate eye of a cornea that had the positive fungal corneal rim culture and developed an infection. Positive fungal donor rim cultures-especially in the setting of interface keratitis after EK surgery-can raise the index of suspicion for a fungal cause and may help direct therapy, especially in the early stages, where the symptoms and signs of spread may not be obvious and obtaining direct cultures is inherently difficult without surgical intervention.

Clinical characteristics, Cryptococcus neoformans genotypes, antifungal susceptibility, and outcomes in human immunodeficiency virus-positive patients in Beijing, China.

Cryptococcus neoformans is an environmental fungal pathogen that causes opportunistic infections and severe disseminated meningoencephalitis, mainly in immunocompromised patients such as those with acquired immunodeficiency syndrome (AIDS). In this study, the clinical characteristics, treatment protocols, and outcomes of 70 patients with AIDS and Cryptococcus neoformans infection at Beijing Ditan Hospital were retrospectively analyzed. We performed antimicrobial sensitivity tests and multilocus sequence typing (MLST) on C. neoformans isolates from these patients. The most common symptoms were headache (58.6%), fever (54.3%), and high cerebrospinal fluid pressure (≥200 mm H2O) (71.4%). All patients were positive for C. neoformans antigen in blood or cerebrospinal fluid. The CD4 cell counts of 92.8% (65/70) of patients were <100 cells/µL. In total, 74 C. neoformans isolates were obtained from the 70 patients. The 65 isolates that could be typed fell into 12 sequence types (STs) by MLST: ST5, ST31, ST63, ST202, ST237, ST289, ST295, ST296, ST298, ST324, ST337, and ST359. ST5 was the major type, accounting for 78.5% of isolates (51/65). This study comprehensively assessed the clinical and molecular epidemiology of C. neoformans in patients with AIDS and may inform the development of targeted prevention and treatment strategies for immunocompromised patients with C. neoformans infection.

Identification patterns of Trichoderma strains using morphological characteristics, phylogenetic analyses and lignocellulolytic activities.

Trichoderma is a genus of soil-borne fungus with an abundance of reports of its economic importance in the agriculture industry. Thus, the correct identification of Trichoderma species is necessary for its commercial purposes. Globally, Trichoderma species are routinely identified from micro-morphological descriptions which can be tedious and prone to errors. Thus, we emphasize that the accurate identification of Trichoderma strains requires a three-pronged approach i.e. based on its morphological characteristics, multilocus gene sequences of the rDNA [internal transcribed spacer (ITS) 1 and 2 regions], translation elongation factor 1-α (TEF-1α), Calmodulin (CAL) and its lignocellulolytic activities. We used this approach to identify a total of 53 Trichoderma strains which were isolated from a wet paddy field located at Tuaran, Sabah, Malaysia. The 53 strains were positively identified as belonging to three Trichoderma species, namely T. asperellum (43 strains), T. harzianum (9 strains), and T. reesei (one strain) on the basis of its morphological characteristics and multilocus gene sequences. Phylogenetic trees constructed based on the UPGMA method of the ITS 1 and 2 regions of the rDNA, TEF-1α and CAL revealed three distinct groups with the T. asperellum, T. harzianum and T. reesei strains placed under the section of Trichoderma, Pachybasium and Longibrachiatum, respectively. In addition, the lignocellulolytic activities of the isolates were measured based on the diameters of the halo zones produced when degrading cellulose, lignin, and starch, respectively. This diagnostic assay can be used to identify Trichoderma as it produces polyphenol oxidase when Tannic Acid Media is used for the lignin test, endoglucanases when Jensen media is used for cellulose, and it hydrolyzes starch to glucose when the modified Melin-Nokrans media is used for the starch test. Accurate identification of Trichoderma species is needed as these strains can potentially be used as a biocontrol agent to prevent diseases and to increase yield in agriculture crops.

Zygotorulaspora dagestanica sp. nov., a novel ascomycetous yeast species associated with the Georgian honeysuckle (Lonicera iberica M. Bieb.).

During an investigation of the yeast communities associated with wild fruit shrubs in Dagestan (Caucasus, Russia), four fermenting ascospore-producing yeast strains were isolated from leaves of the Georgian honeysuckle (Lonicera iberica M. Bieb.) and from soil underneath this plant. Phylogenetic analyses based on concatenated sequences of the ITS region and D1/D2 domains of the large subunit rRNA gene and concatenated sequences of the ribosomal DNA cystron, RPB2 and TEF1 genes showed that the isolated strains represented a new species of the genus Zygotorulaspora. The new species was placed in the basal position to other species of the clade and close to Zygotorulaspora mrakii. Based on the results of phylogenetic analyses and the phenotypic characteristics of the four studied strains, a novel species is described, for which the name Zygotorulaspora dagestanica sp. nov. is proposed. The holotype is KBP Y-4591T, three metabolically inactive cryopreserved isotype cultures are DSM 100088, VKM Y-3060 and VKPM Y-4318. The MycoBank number is MB 838285.

Cyberlindnera sylvatica sp. nov., a yeast species isolated from forest habitats.

Five yeast strains isolated from forest habitats in Hungary and Germany were characterized phenotypically and by sequencing of the D1/D2 domain of the large subunit rRNA gene and the ITS1-5.8S-ITS2 (ITS) region of the rRNA gene. The strains have identical D1/D2 domain and ITS region sequences. By sequence comparisons, Candida mycetangii and Candida maritima were identified as the closest relatives among the currently recognized yeast species. The DNA sequences of the investigated strains differ by 1.2 % (six substitutions) in the D1/D2 domain and by 3.5 % (12 substitutions and eight indels) in the ITS region from the type strain of C. mycetangii (CBS 8675T) while by 1.2 % (six substitutions and one indel) in the D1/D2 domain and by 7 % (32 substitutions and seven indels) in the ITS region from the type strain of C. maritima (CBS 5107T). Because the intraspecies heterogeneity seems to be very low and the distance to the most closely related species is above the commonly expected level for intraspecies variability Cyberlindnera sylvatica sp. nov. (holotype, CBS 16335T; isotype, NCAIM Y.02233T; MycoBank no., MB 835268) is proposed to accommodate the above-noted five yeast strains. Phenotypically the novel species can be distinguished from C. mycetangii and C. maritima by the formation of ascospores. Cyberlindnera sylvatica forms one or two hat-shaped ascospores per ascus on many different media as well as well-developed pseudohyphae and true hyphae. Additionally, we propose the transfer of three anamorphic members of the Cyberlindnera americana sub-clade to the genus Cyberlindnera as the following new taxonomic combinations Cyberlindnera maritima f.a., comb. nov., Cyberlindnera mycetangii f.a., comb. nov. and Cyberlindnera nakhonratchasimensis f.a., comb. nov.

Scedosporiosis/lomentosporiosis observational study (SOS): Clinical significance of Scedosporium species identification.

Scedosporiosis/lomentosporiosis is a devastating emerging fungal infection. Our objective was to describe the clinical pattern and to analyze whether taxonomic grouping of the species involved was supported by differences in terms of clinical presentations or outcomes. We retrospectively studied cases of invasive scedosporiosis in France from 2005 through 2017 based on isolates characterized by polyphasic approach. We recorded 90 cases, mainly related to Scedosporium apiospermum (n = 48), S. boydii/S. ellipsoideum (n = 20), and Lomentospora prolificans (n = 14). One-third of infections were disseminated, with unexpectedly high rates of cerebral (41%) and cardiovascular (31%) involvement. In light of recent Scedosporium taxonomic revisions, we aimed to study the clinical significance of Scedosporium species identification and report for the first time contrasting clinical presentations between infections caused S. apiospermum, which were associated with malignancies and cutaneous involvement in disseminated infections, and infections caused by S. boydii, which were associated with solid organ transplantation, cerebral infections, fungemia, and early death. The clinical presentation of L. prolificans also differed from that of other species, involving more neutropenic patients, breakthrough infections, fungemia, and disseminated infections. Neutropenia, dissemination, and lack of antifungal prescription were all associated with 3-month mortality. Our data support the distinction between S. apiospermum and S. boydii and between L. prolificans and Scedosporium sp. Our results also underline the importance of the workup to assess dissemination, including cardiovascular system and brain.

Scedosporiosis/lomentosporiosis is a devastating emerging fungal infection. Our objective was to describe the clinical pattern and to analyze whether taxonomic grouping of the species involved was supported by differences in terms of clinical presentations or outcomes.

Candida spp. infective endocarditis: Characteristics and outcomes of twenty patients with a focus on injection drug use as a predisposing risk factor.

BACKGROUND: Candida spp. infective endocarditis (CIE) although rare is associated with high morbidity and mortality. Risk factors include prosthetic heart valves and injection drug use (IDU). We reviewed all cases of CIE at our institution to describe the microbiology, treatment and outcomes of patients focusing on IDU as a predisposing factor. METHODS: Retrospective cohort of patients with definite CIE between 2013 and 2019 at a university hospital was analysed. Demographic data collected included IDU, microbiologic, treatment and mortality. The primary outcome of interest was 12-month, all-cause mortality. RESULTS: Twenty patients were included (one had two separate episodes of CIE); CIE accounted for 4% of total infective endocarditis (IE) cases during the study period. The median (IQR) age was 38 (30-58) years, 10 (50%) had a previous history of IE, and 4 (20%) patients had prosthetic heart valves or an implanted cardiac device. Thirteen (65%) patients were IDU. The tricuspid valve was the primary valve involved (8/18, 44%), and C albicans was the most frequently isolated organism (8, 36%). Echinocandin was the most common treatment strategy (8, 40%). Only three (15%) patients underwent valve replacement during hospitalisation. There were no in-hospital fatalities, and 5 (25%) patients died at one year; all were IDU (39% to 0%, p = .11). CONCLUSION: CIE is a rare infectious disease seen more commonly in the IDU population. Cardiac surgery was rarely performed, and long-term mortality was 25%. Additional data are needed to identify ideal management strategies in this population.

Cryptococcal meningitis in non-HIV patients in the State of Amazonas, Northern Brazil.

Cryptococcosis is a life-threatening fungal infection caused by the Cryptococcus neoformans/Cryptococcus gattii species complex. Most cases are recorded in patients suffering from HIV/AIDS (human immunodeficiency virus/acquired immunodeficiency syndrome). However, this infection also occurs in non-HIV patients with a proportion of 10-30% of all cases. The study aimed at the clinical and molecular characterization of non-HIV patients diagnosed with cryptococcosis at the Tropical Medicine Foundation (FMT-HVD) from July 2016 to June 2019. Medical records of respective patients were analyzed to describe the course of cryptococcosis in non-HIV patients. In addition, multi-locus sequence typing (MLST) was applied to identify the sequence types of the isolated Cryptococcus strains, to perform phylogenetic analysis, and to evaluate the isolates' genetic relationship to global reference strains. Antifungal susceptibility profiles to amphotericin B, fluconazole, and itraconazole were assessed by broth microdilution. From a total of 7 patients, 4 were female, the age range varied between 10 and 53 years (median of 36.3 years). Cryptococcal meningitis was the common clinical manifestation (100%). The period between onset of symptoms and confirmed diagnosis ranged from 15 to 730 days (mean value of 172.9 days), and the observed mortality was 57.1%. Of note, comorbidities of the assessed cryptococcosis patients comprised hypertension, diabetes mellitus, and intestinal tuberculosis. Genotyping applying PCR-RFLP of the URA5 gene identified all clinical isolates as C. gattii genotype VGII. Using MLST, it was possible to discriminate the sequence types ST20 (n = 4), ST5 (n = 3), and the newly identified sequence type ST560 (n = 1). The antifungals amphotericin B, fluconazole, and itraconazole showed satisfactory inhibitory activity (microdilution test) against all C. gattii VGII strains.

Monitoring Inflammasome Priming and Activation in Response to Candida albicans.

Candida albicans is a common mucosal colonizer, as well as a cause of lethal invasive fungal infections. The major predisposing factor for invasive fungal disease is a compromised immune system. One component of the host immune response to fungal infection is the activation of the inflammasome, a multimeric protein complex that is critical for regulating host pro-inflammatory responses. Here, we describe methods for investigating the interactions between C. albicans and host macrophages, with a focus on the inflammasome. C. albicans isolates differ in the degree to which they activate the inflammasome due to differences in internalization, morphogenic switching, and inflammasome priming. Therefore, we include protocols for identifying these factors. This simple in vitro model can be used to elucidate the contributions of specific C. albicans strains or mutants to different aspects of interactions with macrophages. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Measuring inflammasome priming in response to Candida albicans Basic Protocol 2: Measuring inflammasome activation in response to Candida albicans Support Protocol: Controlling for phagocytosis.

Candida nivariensis: Identification strategy in mycological laboratories.

Candida nivariensis is a cryptic fungal species classified within the Candida glabrata complex. It was described for the first time in 2005 by the means of DNA sequencing. We report a rare case of C. nivariensis deep-seated infection occurring in a 77-year-old man hospitalized for cysto-prostatectomy. Phenotypic testing based on the direct examination and the macroscopic features of the in vitro culture initially suggested C. glabrata species, while MALDI-TOF mass spectrometry enables correct identification. The isolate was found resistant to fluconazole, like in almost 20% of the reported cases. Herein, we present our practical strategy to reliably characterize this rare cryptic species. To date, MALDI-TOF mass spectrometry-based analysis showed very good results for such a purpose.

Frequency and geographical distribution of genotypes and mating types of Cryptococcus neoformans and Cryptococcus gattii species complexes in Argentina / Frecuencia y distribución geográfica de genotipos y tipos sexuales de los complejos de especies Cryptococcus neoformans y Cryptococcus gattii en Argentina

Abstract The aim of this work was to know the frequency and geographical distribution of genotypes and mating types of Cryptococcus neoformans and Cryptococcus gattii species complexes isolated from human infections in Argentina during the period from April 2009 to April 2011. A multicenter study was conducted, in which 372 isolates were obtained from 61 laboratories throughout the country. Of those, 98.8% of the isolates belonged to the C. neoformans species complex and 1.1% to the C. gattii species complex. Genotype VNI (MATa) was the most frequently isolated (n = 326, 87.6%), followed by VNII (MATa) (n = 22, 5.9%), the recently described VNII-VNIV (aADa) hybrid (n = 14, 3.8%), VNIV (MATa) (n=4, 1.1%), VNIII (aADa) hybrid (n = 1, 0.3%), and VNIII (aADa) hybrid (n = 1, 0.3%). The Argentine Central region showed the greatest number of cases and genotype diversity. Interestingly, a relative high frequency was observed in genotype VNII (MATa) in the Cuyo, Northeast and Northwest regions and, also in VNII-VNIV (aADa) hybrids in the Northwest region. C. gattii species complex was isolated at a low rate; 3 VGI (MATa) and 1 VGII (MATa) isolates were obtained from the Northwest and Central regions. In conclusion, this study shows that genotype frequencies seem to vary among regions in Argentina and reveals a relatively high frequency of rare hybrids in the Northwest region. Further regional clinical and environmental studies may help to elucidate if those varia-tions in frequencies are associated with the existence of regional ecological niches or any other regional factors.

Resumen El objetivo de! trabajo fue conocer la frecuencia y la distribución geográfica de genotipos y tipos sexuales de aislados pertenecientes a los complejos de especies Cryptococcus neoformansy Cryptococcus gattii obtenidos de infecciones humanas en Argentina. Entre abril de 2009 y abril de 2011 se realizó un estudio multicéntrico del que se obtuvieron 372 aislados de 61 laboratorios de diferentes zonas del país. El 98,8% de los aislados pertenecieron al complejo C. neoformansy el 1,1% al complejo C. gattii. El genotipo VNI (MATa) fue el más frecuente (n = 326; 87,6%), le siguieron VNII (MATa) (n = 22; 5,9%), el híbrido VNII-VNIV (aADa) (n = 14; 3,8%), VNIV (MATa) (n =4; 1,1%) y los híbridos VNIII (aADa) (n = 1; 0,3%) y VNIII (aADa) (n = 1; 0,3%). La región Centro mostró el mayor número de casos y la mayor diversidad de genotipos. Cabe destacar que el genotipo VNII (MATa) tuvo una frecuencia relativamente alta en las regiones de Cuyo, Noreste y Noroeste. En esta última región, también fue alta la frecuencia del híbrido VNII-VNIV (aADa). La frecuencia de aislamiento de miembros del complejo C. gattii fue baja: se obtuvieron 3 aislados VGI (MATa) y 1 VGII (MATa) de las regiones Centro y Noroeste. En conclusión, este estudio muestra que las frecuencias de genotipos varían entre las distintas regiones de Argentina y señala la presencia de híbridos poco comunes en una frecuencia relativamente alta dentro de la región Noroeste. Contar con mayor número de estudios clínicos y ambientales regionales podría ayudar a elucidar si tales variaciones están asociadas a la existencia de nichos ecológicos particulares o a algún otro factor regional.

Phaeotremella camelliae sp. nov. (Phaeotremellaceae, Tremellales), A Novel Yeasts Isolated from Tea-Oil Fruits in Jiangxi Province, China.

Sixty-two isolates among the 65 yeast strains isolated from Jiangxi province, China, were identified into 15 known species based on the sequence analysis of the D1/D2 domains of the LSU rRNA and ITS region. The other three strains, GaoanZ14, GaoanC57, and GaoanC191, isolated from tea-oil fruits, were identified as two undescribed species of Phaeotremella based on the multiple gene sequence analysis, physiological, and biochemical comparisons. Strains GaoanC57 and GaoanC191 had one substitution difference both in the D1/D2 domains of the LSU rRNA and ITS region. They formed a separate branch from the other Phaeotremella species in the D1/D2 and multiple genes trees, and differed from the known species by at least 10 nucleotide substitutions in the D1/D2 domains and more than 6% mismatches in the ITS region. The phylogenetic analysis indicated that those two strains represent a novel species of Phaeotremella, for which the names Phaeotremella camelliae sp. nov. (Holotype CGMCC 2.6141, Mycobank MB832699) is proposed. Only one strain, GaoanZ14, represents the other undescribed species of Phaeotremella, so it will be described in latter when more strains are found.

Epidemiological investigation for grouped cases of Trichosporon asahii using whole genome and IGS1 sequencing.

BACKGROUND: Trichosporonosis is a rare invasive infection in humans mainly due to Trichosporon asahii, and especially recovered from patients having haematological malignancy. Since 2012, IGS1 region sequencing is used as a genotyping method to distinguish isolates, with high frequency of one haplotype worldwide and a geographic specificity for some haplotypes. OBJECTIVES: We compared the IGS1 genotyping method and whole genome sequencing (WGS) to study the relationship between clinical isolates involved in two grouped cases in France. METHODS: IGS1 sequencing and antifungal susceptibility testing were performed for 54 clinical isolates. Clinical data for 28 isolates included in surveillance programs were analysed. Whole genome was sequenced for 32 clinical isolates and the type strain. RESULTS: All isolates were intrinsically resistant to flucytosine, while voriconazole had the most potent in vitro activity. The majority of the isolates was recovered from patients with haematological malignancies (42.86%), with a high proportion of children (<15 yrs-old, 32.14%) and a high mortality rate at three months (46.15%). Based on the WGS analysis, isolates exhibiting IGS1 haplotype 1, 3 and 7 belonged to different clades. Five isolates recovered during the first grouped cases had the same IGS1 haplotype and shared 99% of SNPs similarity. For the second grouped cases, four isolates had 98.7% of SNPs similarity while the isolate recovered 4 years earlier was totally unlinked. CONCLUSIONS: We confirmed the usefulness of IGS1 sequencing for grouped cases infection of T. asahii. We underlined its limitation for the study of population structure and the utility of WGS analysis for the study of epidemiologically unrelated isolates.