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Abstract The debates over the requirement of the International Code of Nomenclature for algae, fungi, and plants (ICNafp) for a viable specimen to represent the name-bearing type material for a species or infraspecific taxon have a long history. Taxonomy of fungi commonly studied as living cultures exemplified by yeasts and moulds, strongly depend on viable reference material. The availability of viable cultures is also particularly useful for several groups of filamentous and dimorphic fungi. While the preservation of metabolically inactive cultures is permitted and recommended by the ICNafp, there is room for improvement. Below, we review the history and current status of cultures as the name-bearing type material under the Code. We also present a roadmap with tasks to be achieved in order to establish a stable nomenclatural system that properly manages taxa typified by viable specimens. Furthermore, we propose setting up rules and defining the nomenclatural status of ex-type cultures under Chapter F, the section of the ICNafp that includes provisions specific to names of fungi.

ABSTRACT Candida albicans causes life-threatening invasive infections that are hard to diagnose and treat, with drug resistance leading to treatment failure. The goal of this study was to develop VHH (single variable domain on a heavy chain) nanobodies to detect drug-resistant infections. Llamas were immunized with a mixture of heat killed and fixed C. albicans cells of different morphologies. Llama lymphocyte RNA was used to generate phage display libraries that were tested for binding to C. albicans cells or cell wall fractions, and single antibody domains were isolated. The libraries were panned against echinocandin-resistant C. albicans isolates and counter-selected against echinocandin-susceptible isolates with the aim of isolating binding domains specific for antigens on drug-resistant cells. Thirty diverse VHH nanobodies were selected, and binding characteristics were assessed via dose-response ELISA. Binding was tested against a variety of C. albicans isolates and other Candida species, indicating that the VHHs were specific for C. albicans. The VHH nanobodies were sorted into four distinct groups based on their binding patterns. Two of the groups bound preferentially to the yeast cell poles and hyphae, respectively. Nanobody binding to C. albicans deletion mutants was tested by fluorescence microscopy and ELISA to identify the antigen targets. VHH19 nanobody, belonging to the largest group, recognized the Als4 adhesin. VHH14 antibody in the hyphae-specific group recognized Als3. None of the isolated VHH nanobodies was selective for drug-resistant clinical isolates. Our data indicate that this approach can generate valuable single-domain antibodies specific to C. albicans proteins.IMPORTANCEThe human fungal pathogen Candida albicans causes a range of diseases from superficial mucosal infections such as oral and vaginal thrush to life-threatening, systemic infections. Accurate and rapid diagnosis of these infections remains challenging, and currently, there are no rapid ways to diagnose drug-resistant infections without performing drug susceptibility testing from blood culture, which can take several days. In this proof-of-concept study, we have generated a diverse set of single domain VHH antibodies (nanobodies) from llamas that recognize and bind specifically to C. albicans cell surface. The nanobodies were classified into four groups based on their binding patterns, for example, cell poles or hyphae. Specific nanobodies were verified as recognizing the important adhesin Als4 or the hyphae associated invasin Als3, respectively. The data validate the approach that small VHH antibody domains hold future promise for diagnostic applications and as probes to study the fungal cell surface.

ABSTRACT Malassezia globosa is a lipophilic basidiomycetous yeast that occurs abundantly in breast tumors and that may contribute to a shortened overall survival of breast cancer (BRAC) patients, suggesting that the yeast may participate in the carcinogenesis of BRAC. However, the mechanisms involved in the M. globosa-based acceleration of BRAC are unknown. Here, we show that M. globosa can colonize mammary tissue in 7,12-dimethylbenz[a] anthracene-induced mice. The abundance of M. globosa shortened the overall survival and increased the tumor incidence. Transcriptome data illustrated that IL-17A plays a key role in tumor growth due to M. globosa colonization, and tumor-associated macrophage infiltration was elevated during M. globosa colonization which triggers M2 polarization of macrophages via toll-like receptors 4/nuclear factor kappa-B (Nf-κB) signaling. Our results show that the expression of sphingosine kinase 1 (Sphk1) is increased in breast tumors after inoculation with M. globosa. Moreover, we discovered that Sphk1-specific small interfering RNA blocked the formation of lipid droplets, which can effectively alleviate the expression of the signal transducer and activator of the transcription 3 (STAT3)/Nf-κB pathway. Taken together, our results demonstrate that M. globosa could be a possible factor for the progression of BRAC. The mechanisms by which M. globosa promotes BRAC development involve the IL-17A/macrophage axis. Meanwhile, Sphk1 overexpression was induced by M. globosa infection, which also promoted the proliferation of MCF-7 cells.IMPORTANCELiterature has suggested that Malassezia globosa is associated with breast tumors; however, this association has not been confirmed. Here, we found that M. globosa colonizes in breast fat pads leading to tumor growth. As a lipophilic yeast, the expression of sphingosine kinase 1 (Sphk1) was upregulated to promote tumor growth after M. globosa colonization. Moreover, the IL-17A/macrophages axis plays a key role in mechanisms involved in the M. globosa-induced breast cancer acceleration from the tumor immune microenvironment perspective.

Abstract Purpose Co-culturing is a widely used method to improve bioethanol production from biomass enriched in fermentable sugars. This study aims to produce bioethanol from sugarcane molasses by simultaneous co-fermentation of S. cerevisiae isolate TA2 and W. anomalus isolate HCJ2F-19. Methods Response surface methodology (RSM) based on the central composite design (CCD) was employed to optimize fermentation conditions, including mixing rate (110–150 rpm), temperature (25–35 °C), molasses concentration (25–35 obrix), and incubation time (36–72 h). The ethanol concentration was analyzed using HPLC equipped with a UV detector. Results The monocultureS. cerevisiae isolate TA2 produced 17.2 g.L−1 of ethanol, 0.33 g.g−1 of ethanol yield, and 0.36 g.L−1.h−1 of productivity compared to W. anomalus isolate HCJ2F that produced 14.5 g.L−1, 0.30 g.g−1 and 0.28 g.L−1.h−1 ethanol, ethanol yield, and productivity under laboratory conditions, respectively. In comparison to single cultures of S. cerevisiae TA2 and W. anomalus HCJ2F, the co-fermentation using both isolates showed an increased ethanol yield of 29% and 53% compared to the single species fermentations, respectively. The results showed that the growth of W. anomalus HCJ2F-19 and S. cerevisiae TA2 was not influenced by each other during the co-fermentation process. The one variable at a time optimization (OVAT) analysis resulted in an ethanol concentration of 26.5 g.L−1 with a specific yield and productivity of 0.46 g.g−1, 0.55 g.L−1.h−1, respectively, at pH 5.5, 25 obrix, 48 h, 150 rpm, 30 °C, 60:40 inoculum ratio, and 10% overall inoculum size. The maximum ethanol concentration of 35.5 g.L−1 was obtained by co-fermentation using the RSM-CCD tool at 30 obrix, 30 °C, 54 h, and 130 rpm. Conclusion The results suggested that the co-fermentation of S. cerevisiae isolate TA2 and W. anomalus isolate HCJ2F improves bioethanol production from sugar cane molasses under optimum fermentation conditions.

Abstract Hybridisation is a common event in yeasts often leading to genomic variability and adaptation. The yeast Candida orthopsilosis is a human-associated opportunistic pathogen belonging to the Candida parapsilosis species complex. Most C. orthopsilosis clinical isolates are hybrids resulting from at least four independent crosses between two parental lineages, of which only one has been identified. The rare presence or total absence of parentals amongst clinical isolates is hypothesised to be a consequence of a reduced pathogenicity with respect to their hybrids. Here, we sequence and analyse the genomes of environmental C. orthopsilosis strains isolated from warm marine ecosystems. We find that a majority of environmental isolates are hybrids, phylogenetically closely related to hybrid clinical isolates. Furthermore, we identify the missing parental lineage, thus providing a more complete overview of the genomic evolution of this species. Additionally, we discover phenotypic differences between the two parental lineages, as well as between parents and hybrids, under conditions relevant for pathogenesis. Our results suggest a marine origin of C. orthopsilosis hybrids, with intrinsic pathogenic potential, and pave the way to identify pre-existing environmental adaptations that rendered hybrids more prone than parental lineages to colonise and infect the mammalian host.

Abstract Background Stress-tolerant yeasts are highly desirable for cost-effective bioprocessing. Several strategies have been documented to develop robust yeasts, such as genetic and metabolic engineering, artificial selection, and natural selection strategies, among others. However, the significant drawbacks of such techniques have motivated the exploration of naturally occurring stress-tolerant yeasts. We previously explored the biodiversity of non-conventional dung beetle-associated yeasts from extremophilic and pristine environments in Botswana (Nwaefuna AE et.al., Yeast, 2023). Here, we assessed their tolerance to industrially relevant stressors individually, such as elevated concentrations of osmolytes, organic acids, ethanol, and oxidizing agents, as well as elevated temperatures. Results Our findings suggest that these dung beetle-associated yeasts tolerate various stresses comparable to those of the robust bioethanol yeast strain, Saccharomyces cerevisiae (Ethanol Red™). Fifty-six percent of the yeast isolates were tolerant of temperatures up to 42 °C, 12.4% of them could tolerate ethanol concentrations up to 9% (v/v), 43.2% of them were tolerant to formic acid concentrations up to 20 mM, 22.7% were tolerant to acetic acid concentrations up to 45 mM, 34.0% of them could tolerate hydrogen peroxide up to 7 mM, and 44.3% of the yeasts could tolerate osmotic stress up to 1.5 M. Conclusion The ability to tolerate multiple stresses is a desirable trait in the selection of novel production strains for diverse biotechnological applications, such as bioethanol production. Our study shows that the exploration of natural diversity in the search for stress-tolerant yeasts is an appealing approach for the development of robust yeasts.

Objective: The increasing resistance of Malassezia yeasts against commonly used antifungal drugs dictates the need for novel antifungal compounds. Human lactoferrin-based peptides show a broad spectrum of antimicrobial activities. Various assays were performed to find the optimal growth conditions of the yeasts and to assess cell viability, using media with low lipid content to avoid peptide binding to medium components. Methods: In the current study, we tested the antimicrobial susceptibility of 30 strains of M. furfur that cover the known IGS1 genotypic variation. Results: hLF(1-11) inhibited the growth of all species tested, resulting in minimum inhibitory concentrations (MIC) values ranging from 12.5 to 100 μg/mL. In the combinatory tests, the majority of fractional inhibitory concentration indexes (FIC) for the tested strains of M. furfur were up to 1.0, showing that there is a synergistic or additive effect on the efficacy of the antifungal drugs when used in combination with hLF(1-11). Conclusion: Results showed that hLF(1-11) could be combined with fluconazole or amphotericin for the antimicrobial treatment of resistant strains, enhancing the potency of these antifungal drugs, resulting in an improved outcome for the patient.

In this study, we conducted a comprehensive survey aimed at assessing the diversity of yeast species inhabiting the guts of various insect species collected mainly from two Bulgarian National Parks, namely, Rila, and Pirin. The insect specimens encompass a broad taxonomic spectrum, including representatives from Coleoptera, Orthoptera, Lepidoptera, Hymenoptera, Dermaptera, Isopoda, and Collembola. Yeast strains were identified with DNA barcoding using the ribosomal markers, specifically, the D1/D2 domains of the ribosomal large subunit (LSU) and the internal transcribed spacers regions ITS 1 + 2 (ITS). The analysis unveiled the presence of 89 ascomycetous and 18 basidiomycetous yeast isolates associated with the insect specimens. Furthermore, our study identified 18 hitherto unknown yeast species.

Abstract Background The Ustilaginales comprise hundreds of plant-parasitic fungi with a characteristic life cycle that directly links sexual reproduction and parasitism: One of the two mating-type loci codes for a transcription factor that not only facilitates mating, but also initiates the infection process. However, several species within the Ustilaginales have no described parasitic stage and were historically assigned to the genus Pseudozyma. Molecular studies have shown that the group is polyphyletic, with members being scattered in various lineages of the Ustilaginales. Together with recent findings of conserved fungal effectors in these non-parasitic species, this raises the question if parasitism has been lost recently and in multiple independent events or if there are hitherto undescribed parasitic stages of these fungi. Results In this study, we sequenced genomes of five Pseudozyma species together with six parasitic species from the Ustilaginales to compare their genomic capability to perform two central functions in sexual reproduction: mating and meiosis. While the loss of sexual capability is assumed in certain lineages and asexual species are common in Asco- and Basidiomycota, we were able to successfully annotate potentially functional mating and meiosis genes that are conserved throughout the whole group. Conclusion Our data suggest that at least the key functions of a sexual lifestyle are maintained in the analyzed genomes, challenging the current understanding of the so-called asexual species with respect to their evolution and ecological role.

Abstract Background Hybrids are chimeric organisms with highly plastic heterozygous genomes that may confer unique traits enabling the adaptation to new environments. However, most evolutionary theory frameworks predict that the high levels of genetic heterozygosity present in hybrids from divergent parents are likely to result in numerous deleterious epistatic interactions. Under this scenario, selection is expected to favor recombination events resulting in loss of heterozygosity (LOH) affecting genes involved in such negative interactions. Nevertheless, it is so far unknown whether this phenomenon actually drives genomic evolution in natural populations of hybrids. To determine the balance between selection and drift in the evolution of LOH patterns in natural yeast hybrids, we analyzed the genomic sequences from fifty-five hybrid strains of the pathogenic yeasts Candida orthopsilosis and Candida metapsilosis, which derived from at least six distinct natural hybridization events. Results We found that, although LOH patterns in independent hybrid clades share some level of convergence that would not be expected from random occurrence, there is an apparent lack of strong functional selection. Moreover, while mitosis is associated with a limited number of inter-homeologous chromosome recombinations in these genomes, induced DNA breaks seem to increase the LOH rate. We also found that LOH does not accumulate linearly with time in these hybrids. Furthermore, some C. orthopsilosis hybrids present LOH patterns compatible with footprints of meiotic recombination. These meiotic-like patterns are at odds with a lack of evidence of sexual recombination and with our inability to experimentally induce sporulation in these hybrids. Conclusions Our results suggest that genetic drift is the prevailing force shaping LOH patterns in these hybrid genomes. Moreover, the observed LOH patterns suggest that these are likely not the result of continuous accumulation of sporadic events—as expected by mitotic repair of rare chromosomal breaks—but rather of acute episodes involving many LOH events in a short period of time.

Abstract Mitogenomes are essential due to their contribution to cell respiration. Recently they have also been implicated in fungal pathogenicity mechanisms. Members of the basidiomycetous yeast genus Malassezia are an important fungal component of the human skin microbiome, linked to various skin diseases, bloodstream infections, and they are increasingly implicated in gut diseases and certain cancers. In this study, the comparative analysis of Malassezia mitogenomes contributed to phylogenetic tree construction for all species. The mitogenomes presented significant size and gene order diversity which correlates to their phylogeny. Most importantly, they showed the inclusion of large inverted repeats (LIRs) and G-quadruplex (G4) DNA elements, rendering Malassezia mitogenomes a valuable test case for elucidating the evolutionary mechanisms responsible for this genome diversity. Both LIRs and G4s coexist and convergently evolved to provide genome stability through recombination. This mechanism is common in chloroplasts but, hitherto, rarely found in mitogenomes.

Abstract Purpose Yeast strains tolerant to a wide range of stress conditions are needed for the production of bioethanol from substrates rich in sugar. In our earlier research findings, Meyerozyma caribbica isolate MJTm3 (OM329077) demonstrated remarkable stress tolerance and fermentative activity. The present study aimed to optimize six fermentation parameters to generate conducive fermentation conditions for ethanol production by M. caribbica isolate MJTm3. Method The response surface method (RSM) based on central composite design (CCD) was employed to optimize process conditions for higher bioethanol yield. The optimization process was carried out based on six independent parameters, namely temperature (25–35 °C), pH (5.5–6.5), inoculum size (10–20% (v/v)), molasses concentration (25–35 (w/v)), mixing rate (110–150 rpm), and incubation period (48–72-h). Analysis of ethanol concentration was done by HPLC equipped with a UV detector. Result The optimal conditions of the parameters resulting in a maximum predicted ethanol yield were as follows: pH 5.5, an inoculum size of 20%, a molasses concentration of 25 °Bx, a temperature of 30 °C, an incubation period of 72-h, and a mixing rate of 160 revolutions per minute (rpm). Using the above optimum conditions, the model predicted a bioethanol yield of 79%, 92% of the theoretical yield, a bioethanol concentration of 49 g L−1, and a productivity of 0.68 g L−1 h−1. A batch fermentation experiment was carried out to validate the predicted values and resulted in a bioethanol yield of 86%, 95% of theoretical yield, a bioethanol concentration of 56 g L−1, and productivity of 0.78 g L−1 h−1. On the other hand, the surface plot analysis revealed that the synergistic effect of the molasses concentration and the mixing rate were vital to achieving the highest bioethanol yield. These values suggested that the RSM with CCD was an effective method in producing the highest possible output of bioethanol from molasses in actual operation. Conclusion The study confirmed the potential of using M. caribbica isolate MJTm3 for bioethanol production from sugarcane molasses under the abovementioned optimal fermentation conditions.

Abstract Purpose Yeasts are widely used for the production of bioethanol from biomasses rich in sugar. The present study was aimed at isolating, screening, and characterizing fermentative wild yeast recovered from bio-waste and co-products of Ethiopian sugar factories for bioethanol production using sugarcane molasses as a substrate. Method The wild yeasts were identified according to their cellular morphology and D1/D2 and ITS1-5.8S-ITS2 rDNA sequencing. Analysis of ethanol and by-product concentration was done by HPLC equipped with a UV detector. Higher alcohols, acetaldehyde, and methanol were analyzed using GC-MS equipped with a flame ionization detector (FID). Result Seven strains (Meyerozyma caribbica MJTm3, Meyerozyma caribbica MJTPm4, Meyerozyma caribbica SHJF, Saccharomyces cerevisiae TA2, Wickerhamomyces anomalus MJTPm2, Wickerhamomyces anomalus 4m10, and Wickerhamomyces anomalus HCJ2F) were found tolerant to 18% (v/v) ethanol, whereas one strain Meyerozyma caribbica MJTm3 tolerated 20%. These strains also showed tolerance to 45°C, 50% of sugar, and pH 2–10. Meyerozyma caribbica MJTm3 produced 12.7% (v/v) of alcohol with an actual ethanol concentration of 26 g L−1, an ethanol yield of 47%, 78% of theoretical yield, and a productivity of 0.54 g L−1 h−1 from 30 °Brix of molasses at 48 h incubation under laboratory scale. Based on the one variable at a time optimization (OVAT), the optimal parameters for maximum bioethanol production were at initial pH 5.5, 35 °Brix, 30°C, 15% inoculum size, 150 rpm, 4 g L−1 di-ammonium phosphate supplement, and 48 h incubation. Under these optimum conditions, 14% (v/v) alcohol, 42 g L−1 actual ethanol concentration, 69% ethanol yield, 89% of theoretical yield, and productivity of 0.88 g L−1 h−1 were obtained. Conclusion These results indicated that M. caribbica MJTm3 should further be evaluated, optimized, and improved for industrial bioethanol production due to its fermentation potential.

Annual surveys of Irish soil samples identified three isolates, CBS 16921 (UCD88), CBS 18246 (UCD443), and CBS 18247 (UCD483), of an apiculate yeast species within the Hanseniaspora genus. The internal transcribed spacer (ITS) and D1/D2 region of the large subunit (LSU) rRNA sequences showed that these are isolates of the recently described species Hanseniaspora menglaensis, first isolated from Southwest China. No genome sequence for H. menglaensis is currently available. The genome sequences of the three Irish isolates were determined using short-read (Illumina) sequencing, and the sequence of one isolate (CBS 16921) was assembled to chromosome level using long-read sequencing (Oxford Nanopore Technologies). Phylogenomic analysis shows that H. menglaensis belongs to the fast-evolving lineage (FEL) of Hanseniaspora. Only one MAT idiomorph (encoding MATα1) was identified in all three sequenced H. menglaensis isolates, consistent with one mating type of a heterothallic species. Genome comparisons showed that there has been a rearrangement near MATα of FEL species compared to isolates from the slowly evolving lineage (SEL).

Abstract Irritable bowel syndrome (IBS) is a common disorder characterized by chronic abdominal pain and changes in bowel movements. Visceral hypersensitivity is thought to be responsible for pain complaints in a subset of patients. In an IBS-like animal model, visceral hypersensitivity was triggered by intestinal fungi, and lower mycobiota α-diversity in IBS patients was accompanied by a shift toward increased presence of Candida albicans and Saccharomyces cerevisiae. Yet, this shift was observed in hypersensitive as well as normosensitive patients and diversity did not differ between IBS subgroups. The latter suggests that, when a patient changes from hyper- to normosensitivity, the relevance of intestinal fungi is not necessarily reflected in compositional mycobiota changes. We now confirmed this notion by performing ITS1 sequencing on an existing longitudinal set of fecal samples. Since ITS1 methodology does not recognize variations within species, we next focused on heterogeneity within cultured healthy volunteer and IBS-derived C. albicans strains. We observed inter- and intra-individual genomic variation and partial clustering of strains from hypersensitive patients. Phenotyping showed differences related to growth, yeast-to-hyphae morphogenesis and gene expression, specifically of the gene encoding fungal toxin candidalysin. Our investigations emphasize the need for strain-specific cause-and-effect studies within the realm of IBS research.

ABSTRACT In recent decades, the incidence of Candida infections has increased in immunocompromised patients. This multicenter study aimed to evaluate in vitro antifungal activities of 8 antifungal agents against the Candida species isolated from 10 university hospitals in Iran. During the period from Dec 2019 to Dec 2021, Candida species were collected from clinical samples of patients. The isolates were identified by PCR restriction fragment length polymorphism and sequencing methods. The antifungal susceptibility tests of each isolate to eight antifungal agents were performed according to the microdilution CLSI M27, M59, and M60 standard methods. A total of 598 Candida strains were isolated from clinical samples. The most commonly isolated Candida species was C. albicans, followed by C. glabrata, C. parapsilosis, Debaryomyces hansenii (Candida famata), C. tropicalis, Pichia kudriavzevii (Candida krusei), C. orthopsilosis, Meyerozyma guilliermondii (Candida guilliermondii), Kluyveromyces marxianus (Candida kefyr), and Clavispora lusitaniae (Candida lusitaniae). MIC90 values in all Candida species were as follows: 0.25 μg/mL for caspofungin and voriconazole; 0.5 μg/mL for amphotericin B and isavuconazole; 2 μg/mL for itraconazole, luliconazole, and posaconazole; and 16 μg/mL for fluconazole. Although 30/285 C. albicans, 15/31 C. hansenii, 3/12 M. guilliermondii, 67/125 C. glabrata, 5/15 P. kudriavzevii, 6/60 C. parapsilosis, and 5/23 C. tropicalis isolates were multiazole resistant with resistance to 2 to 4 azoles, pan-azole resistance was not observed. According to our data, Candida albicans and C. glabrata were the most frequent species isolated from clinical samples in Iran. Caspofungin and voriconazole, with lower MIC90 values, are the most effective than other antifungal agents for the treatment of Candida infections in this region. IMPORTANCE Candida species cause severe invasive infections of the heart, brain, eyes, bones, and other parts of the body. Knowledge of regional distributions of causative Candida agents and their antifungal susceptibility patterns can help to monitor resistance to antifungal agents of various species and support local and national surveillance programs. In the present study, C. albicans and C. glabrata were the most frequently isolated species from clinical samples in Iran. Increasing rates of non-albicans Candida isolates from the Iranian population should be looked at as alarming due to various levels of intrinsic MIC values or resistance to various antifungal drugs. Caspofungin and voriconazole are recommended over fluconazole for the treatment of Candida infections in the study region. However, amphotericin B and isavuconazole are also active against the most common Candida species isolated from patients. Pan azole-resistant Candida species were not observed in the present study.

ABSTRACT The unambiguous application of fungal names is important to communicate scientific findings. Names are critical for (clinical) diagnostics, legal compliance, and regulatory controls, such as biosafety, food security, quarantine regulations, and industrial applications. Consequently, the stability of the taxonomic system and the traceability of nomenclatural changes is crucial for a broad range of users and taxonomists. The unambiguous application of names is assured by the preservation of nomenclatural history and the physical organisms representing a name. Fungi are extremely diverse in terms of ecology, lifestyle, and methods of study. Predominantly unicellular fungi known as yeasts are usually investigated as living cultures. Methods to characterize yeasts include physiological (growth) tests and experiments to induce a sexual morph; both methods require viable cultures. Thus, the preservation and availability of viable reference cultures are important, and cultures representing reference material are cited in species descriptions. Historical surveys revealed drawbacks and inconsistencies between past practices and modern requirements as stated in the International Code of Nomenclature for Algae, Fungi, and Plants (ICNafp). Improper typification of yeasts is a common problem, resulting in a large number invalid yeast species names. With this opinion letter, we address the problem that culturable microorganisms, notably some fungi and algae, require specific provisions under the ICNafp. We use yeasts as a prominent example of fungi known from cultures. But viable type material is important not only for yeasts, but also for other cultivable Fungi that are characterized by particular morphological structures (a specific type of spores), growth properties, and secondary metabolites. We summarize potential proposals which, in our opinion, will improve the stability of fungal names, in particular by protecting those names for which the reference material can be traced back to the original isolate.

ABSTRACT Aspergillus species are a major cause of life-threatening invasive infections and noninvasive diseases. This study seeks to investigate the frequency of Aspergillus species among Iranian patients and their susceptibility to seven antifungals. In a cross-sectional study, 233 Aspergillus isolates were collected from 11 university hospitals in Iran between 2018 and 2021. Aspergillus isolates were identified based on colony morphology, microscopic characteristics, PCR-restriction fragment length polymorphism (RFLP), and sequencing of the beta-tubulin gene. The CLSI M38-A2 reference methodology was used for antifungal susceptibility testing of amphotericin B, voriconazole, posaconazole, itraconazole, luliconazole, isavuconazole, and caspofungin. Members of Aspergillus section Flavi (117/233, 50.2%), Aspergillus section Nigri (77/233, 33.1%), Aspergillus section Fumigati (21/233, 9%), Aspergillus section Terrei (14/233, 6%), Aspergillus pseudodeflectus (2/233, 0.85%), and Aspergillus melleus (2/233, 0.85%) were isolated from the samples. The lowest 0.25 MIC90 values for all isolates tested were for luliconazole (0.016 μg/mL) and isavuconazole (0.250 μg/mL), and the highest value was observed for itraconazole (≥ 8μg/mL). The 90% minimum effective concentration (MEC90) value for caspofungin was 0.125 μg/mL. MIC90 values for voriconazole, amphotericin B, and posaconazole were 1, 2, and 2 μg/mL, respectively. The non-wild-type species were presented for amphotericin B (3%), voriconazole (1.3%), posaconazole (2.6%), luliconazole (1.3%), isavuconazole (1.7%), and caspofungin (4.7%). Positive correlations in the MIC values of azole antifungals were observed, and using one azole increases the MIC value rates of other ones. None of the species were pan-azole resistant. Species of Aspergillus section Flavi were the most common Aspergillus species isolated from Iranian samples. Luliconazole, caspofungin, and isavuconazole present the most effective antifungal agents for treatment of infection due to Aspergillus species. Susceptibility tests should be performed frequently in each region for the best management of patients. IMPORTANCE Aspergillus species are the leading cause of invasive aspergillosis in immunocompromised hosts. The susceptibility of Aspergillus species to antifungal agents might be different. Azole-resistant species have emerged worldwide. Performing susceptibility testing in each region can help in the best management of patients. Here, we show the epidemiology and distribution of Aspergillus species in Iran and their susceptibility patterns for seven antifungal agents. The significant points of the present study are that species of Aspergillus section Flavi are the most prevalent Aspergillus species isolated from 11 university hospitals. Luliconazole, caspofungin, and isavuconazole were effective antifungal agents against all Aspergillus species.

Sexual reproduction is a ubiquitous, ancient eukaryotic trait. While most sexual organisms have to find a mating partner, species as diverse as animals, plants, and fungi have evolved the ability to reproduce sexually without requiring another individual. Here, we uncovered the mechanism of self-compatibility (homothallism) in Cryptococcus depauperatus, a fungal species closely related to the human fungal pathogens Cryptococcus neoformans and Cryptococcus gattii. In contrast to C. neoformans or C. gattii, which grow as a yeast asexually, and produce hyphae, basidia, and infectious spores during sexual reproduction, C. depauperatus grows exclusively as hyphae decorated with basidia and abundant spores, thus continuously engaged in sexual reproduction. Through comparative genomics and analyses of mutants defective in key mating/meiosis genes, we demonstrate the C. depauperatus sexual cycle involves meiosis and that self-compatibility is orchestrated by an unlinked mating receptor (Ste3a) and pheromone ligand (MFα) pair derived from opposite mating types of a heterothallic (self-sterile) ancestor. We identified a putative mating-type (MAT) determining region containing genes phylogenetically aligned with MATa alleles of other species, and a few MATα gene alleles scattered throughout the genome, but no homologs of the mating-type homeodomain genes SXI1 (HD1) and SXI2 (HD2). Comparative analyses suggest a dramatic remodeling of the MAT locus possibly owing to reduced selective constraints to maintain mating-type genes in tight linkage, associated with a transition to self-fertility. Our findings support C. depauperatus as an obligately sexual, homothallic fungus and provide insight into repeated transitions between sexual reproduction modes that have occurred throughout the fungal kingdom.

Abstract Bioethanol from abundant and inexpensive agricultural and industrial wastes possesses the potential to reduce greenhouse gas emissions. Bioethanol as renewable fuel addresses elevated production costs, as well as food security concerns. Although technical advancements in simultaneous saccharification and fermentation have reduced the cost of production, one major drawback of this technology is that the pre-treatment process creates environmental stressors inhibitory to fermentative yeasts subsequently reducing bioethanol productivity. Robust fermentative yeasts with extreme stress tolerance remain limited. This review presents the potential of dung beetles from pristine and unexplored environments as an attractive source of extremophilic bioethanolic yeasts. Dung beetles survive on a recalcitrant lignocellulose-rich diet suggesting the presence of symbiotic yeasts with a cellulolytic potential. Dung beetles inhabiting extreme stress environments have the potential to harbour yeasts with the ability to withstand inhibitory environmental stresses typically associated with bioethanol production. The review further discusses established methods used to isolate bioethanolic yeasts, from dung beetles.

Transcription factor Rme1 regulates meiosis and pseudohyphal growth in baker’s yeast, but its role in the meiosis-defective pathogen Candida albicans is unclear. Here, Hernández-Cervantes et al. show that Rme1 activates the expression of genes required for formation of asexual spores in Candida species.

ABSTRACT Malassezia species are important fungal skin commensals and are part of the normal microbiota of humans and other animals. However, under certain circumstances these fungi can also display a pathogenic behavior. For example, Malassezia furfur is a common commensal of human skin and yet is often responsible for skin disorders but also systemic infections. Comparative genomics analysis of M. furfur revealed that some isolates have a hybrid origin, similar to several other recently described hybrid fungal pathogens. Because hybrid species exhibit genomic plasticity that can impact phenotypes, we sought to elucidate the genomic evolution and phenotypic characteristics of M. furfur hybrids in comparison to their parental lineages. To this end, we performed a comparative genomics analysis between hybrid strains and their presumptive parental lineages and assessed phenotypic characteristics. Our results provide evidence that at least two distinct hybridization events occurred between the same parental lineages and that the parental strains may have originally been hybrids themselves. Analysis of the mating-type locus reveals that M. furfur has a pseudobipolar mating system and provides evidence that after sexual liaisons of mating compatible cells, hybridization involved cell-cell fusion leading to a diploid/aneuploid state. This study provides new insights into the evolutionary trajectory of M. furfur and contributes with valuable genomic resources for future pathogenicity studies. IMPORTANCE Malassezia furfur is a common commensal member of human/animal microbiota that is also associated with several pathogenic states. Recent studies report involvement of Malassezia species in Crohn’s disease, a type of inflammatory bowel disease, pancreatic cancer progression, and exacerbation of cystic fibrosis. A recent genomics analysis of M. furfur revealed the existence of hybrid isolates and identified their putative parental lineages. In this study, we explored the genomic and phenotypic features of these hybrids in comparison to their putative parental lineages. Our results revealed the existence of a pseudobipolar mating system in this species and showed evidence for the occurrence of multiple hybridization events in the evolutionary trajectory of M. furfur. These findings significantly advance our understanding of the evolution of this commensal microbe and are relevant for future studies exploring the role of hybridization in the adaptation to new niches or environments, including the emergence of pathogenicity.

Abstract Background Despite being associated with a high mortality and economic burden, data regarding candidemia are scant in Algeria. The aim of this study was to unveil the epidemiology of candidemia in Algeria, evaluate the antifungal susceptibility pattern of causative agents and understand the molecular mechanisms of antifungal resistance where applicable. Furthermore, by performing environmental screening and microsatellite typing we sought to identify the source of infection. Methods We performed a retrospective epidemiological-based surveillance study and collected available blood yeast isolates recovered from the seven hospitals in Algiers. To identify the source of infection, we performed environmental screening from the hands of healthcare workers (HCWs) and high touch areas. Species identification was performed by API Auxa-Color and MALDI-TOF MS and ITS sequencing was performed for species not reliably identified by MALDI-TOF MS. Antifungal susceptibility testing followed CLSI M27-A3/S4 and included all blood and environmental yeast isolates. ERG11 sequencing was performed for azole-resistant Candida isolates. Microsatellite typing was performed for blood and environmental Candida species, where applicable. Results Candida tropicalis (19/66) was the main cause of candidemia in these seven hospitals, followed by Candida parapsilosis (18/66), Candida albicans (18/66), and Candida glabrata (7/66). The overall mortality rate was 68.6% (35/51) and was 81.2% for C. tropicalis-infected patients (13/16). Fluconazole was the main antifungal drug used (12/51); 41% of the patients (21/51) did not receive any systemic treatment. Candida parapsilosis was isolated mainly from the hands of HCWs (7/28), and various yeasts were collected from high-touch areas (11/47), including Naganishia albida, C. parapsilosis and C. glabrata. Typing data revealed interhospital transmission on two occasions for C. parapsilosis and C. glabrata, and the same clone of C. parapsilosis infected two patients within the same hospital. Resistance was only noted for C. tropicalis against azoles (6/19) and fluconazole-resistant C. tropicalis isolates (≥8 μg/ml) (6/19) contained a novel P56S (5/6) amino acid substitution and a previously reported one (V234F; 1/6) in Erg11p. Conclusions Collectively, our data suggest an urgent need for antifungal stewardship and infection control strategies to improve the clinical outcome of Algerian patients with candidemia. The high prevalence of C. tropicalis joined by fluconazole-resistance may hamper the therapeutic efficacy of fluconazole, the frontline antifungal drug used in Algeria.

Plastic debris has been accumulating in the marine realm since the start of plastic mass production in the 1950s. Due to the adverse effects on ocean life, the fate of plastics in the marine environment is an increasingly important environmental issue. Microbial degradation, in addition to weathering, has been identified as a potentially relevant breakdown route for marine plastic debris. Although many studies have focused on microbial colonization and the potential role of microorganisms in breaking down marine plastic debris, little is known about fungi-plastic interactions. Marine fungi are a generally understudied group of microorganisms but the ability of terrestrial and lacustrine fungal taxa to metabolize recalcitrant compounds, pollutants, and some plastic types (e.g., lignin, solvents, pesticides, polyaromatic hydrocarbons, polyurethane, and polyethylene) indicates that marine fungi could be important degraders of complex organic matter in the marine realm, too. Indeed, recent studies demonstrated that some fungal strains from the ocean, such as Zalerion maritimum have the ability to degrade polyethylene. This mini-review summarizes the available information on plastic-fungi interactions in marine environments. We address (i) the currently known diversity of fungi colonizing marine plastic debris and provide (ii) an overview of methods applied to investigate the role of fungi in plastic degradation, highlighting their advantages and drawbacks. We also highlight (iii) the underestimated role of fungi as plastic degraders in marine habitats.

Fecal microbiota transplantation (FMT) has the potential to restore (bacterial and fungal) microbial imbalance in ulcerative colitis (UC) patients and contribute to disease remission. Here, we aimed to identify fecal fungal species associated with the induction of clinical remission and endoscopic response to FMT for patients with mild-to-moderate ulcerative colitis. We analyzed the internal transcribed spacer 1 (ITS1)-based mycobiota composition in fecal samples from patients (n = 31) and donors (n = 7) that participated previously in a double-blinded randomized control trial evaluating the efficacy of two infusions of donor FMT compared with autologous FMT. The abundance of the yeast genus Filobasidium in fecal material used for transplantation was shown to correlate with clinical remission following FMT, irrespective of its presence in the material of donor or autologous fecal microbiota transfer. The amplified sequence variants within the genus Filobasidium most closely resembled Filobasidium magnum. Monocyte-derived macrophages and HT29 epithelial cells were stimulated with fungal species. Especially Filobasidium floriforme elicited an IL10 response in monocyte-derived macrophages, along with secretion of other cytokines following stimulation with other Filobasidium species. No effect of Filobasidium spp. was seen on epithelial wound healing in scratch assays. In conclusion, the enriched presence of Filobasidium spp. in donor feces is associated with the positive response to FMT for patients with UC and hence it may serve as a predictive fungal biomarker for successful FMT.

Malassezia spp. are lipid-dependent yeasts, inhabiting the skin and mucosa of humans and animals. They are involved in a variety of skin disorders in humans and animals and may cause bloodstream infections in severely immunocompromised patients. Despite a tremendous increase in scientific knowledge of these yeasts during the last two decades, the epidemiology of Malassezia spp. related to fungemia remains largely underestimated most likely due to the difficulty in the isolation of these yeasts species due to their lipid-dependence. This review summarizes and discusses the most recent literature on Malassezia spp. infection and fungemia, its occurrence, pathogenicity mechanisms, diagnostic methods, in vitro susceptibility testing and therapeutic approaches.

Clonal expansion of fluconazole resistant (FLZ-R) Candida parapsilosis isolates is increasingly being identified in many countries, while there is no study exploring the antifungal susceptibility pattern, genetic diversity, and clinical information for Iranian C. parapsilosis blood isolates. Candida parapsilosis species complex blood isolates (n = 98) were recovered from nine hospitals located in three major cities, identified by MALDI-TOF MS, and their genetic relatedness was examined by AFLP fingerprinting. Antifungal susceptibility testing followed CLSI-M27-A3 and ERG11, MRR1 and hotspots 1/2 (HS1/2) of FKS1 were sequenced to assess the azole and echinocandin resistance mechanisms, respectively. Ninety-four C. parapsilosis and four Candida orthopsilosis isolates were identified from 90 patients. Only 43 patients received systemic antifungal drugs with fluconazole as the main antifungal used. The overall mortality rate was 46.6% (42/90) and death mostly occurred for those receiving systemic antifungals (25/43) relative to those not treated (17/47). Although, antifungal-resistance was rare, one isolate was multidrug-resistant (FLZ = 16 μg/ml and micafungin = 8 μg/ml) and the infected patient showed therapeutic failure to FLZ prophylaxis. Mutations causing azole and echinocandin resistance were not found in the genes studied. AFLP revealed five genotypes (G) and G1 was the main one (59/94; 62.7%). Clinical outcome was significantly associated with city (P = 0.02, α

The genus Madurella comprising four species, M. fahalii, M. mycetomatis, M. pseudomycetomatis, and M. tropicana, represents the prevalent cause of eumycetoma worldwide. The four species are phenotypically similar and cause an invariable clinical picture, but differ markedly in their susceptibility to antifungal drugs, and epidemiological pattern. Therefore, specific identification is required for optimal management of Madurella infection and to reveal proper epidemiology of the species. In this study, a novel multiplex real-time PCR targeting the four Madurella species was developed and standardized. Evaluation of the assay using reference strains of the target and non-target species resulted in 100% specificity, high analytical reproducibility (R2 values >0.99) and a lowest detection limit of 3 pg target DNA. The accuracy of the real-time PCR was further assessed using biopsies from eumycetoma suspected patients. Unlike culture and DNA sequencing as gold standard diagnostic methods, the real-time PCR yielded accurate diagnosis with specific identification of the causative species in three hours compared to one or two weeks required for culture. The novel method reduces turnaround time as well as labor intensity and high costs associated with current reference methods.

Candida parapsilosis is the second most common cause of candidemia in some geographical areas and in children in particular. Yet, the proportion among children varies, for example, from 10.4% in Denmark to 24.7% in Tehran, Iran. As this species is also known to cause hospital outbreaks, we explored if the relatively high number of C. parapsilosis pediatric cases in Tehran could in part be explained by undiscovered clonal outbreaks. Among 56 C. parapsilosis complex isolates, 50 C. parapsilosis were genotyped by Amplified Fragment Length Polymorphism (AFLP) fingerprinting and microsatellite typing and analyzed for nucleotide polymorphisms by FKS1 and ERG11 sequencing. AFLP fingerprinting grouped Iranian isolates in two main clusters. Microsatellite typing separated the isolates into five clonal lineages, of which four were shared with Danish isolates, and with no correlation to the AFLP patterns. ERG11 and FKS1 sequencing revealed few polymorphisms in ERG11 leading to amino-acid substitutions (D133Y, Q250K, I302T, and R398I), with no influence on azole-susceptibilities. Collectively, this study demonstrated that there were no clonal outbreaks at the Iranian pediatric ward. Although possible transmission of a diverse C. parapsilosis community within the hospital cannot be ruled out, the study also emphasizes the necessity of applying appropriately discriminatory methods for outbreak investigation.

Abstract Background Candida albicans, Candida glabrata, and Candida parapsilosis are three prevalent causes of candidiasis, worldwide. These species are considered as nine medically important complex species. Limited knowledge about these newly recognized species prompted us to develop a one-step, multiplex PCR to detect and identify them in clinical settings. Methods Primers targeting Hyphal Wall Protein I gene for the C. albicans, C. dubliniensis, C. africana, Intergenic Spacer for the C. glabrata, C. nivariensis, C. bracarensis, and Intein and ITS rDNA for the C. parapsilosis, C. orthopsilosis, and C. metapsilosis were designed. Using 168 CBS reference strains and 280 clinical isolates, the specificity and reproducibility of the developed assay were evaluated. Results Our developed assay successfully identified and distinguished all the nine species. No cross-reaction with closely- and distantly-related yeast species, Aspergillus species and human DNA was observed, resulting in 100% specificity. The ambiguous results obtained by MALDI-TOF for C. albicans and C. africana were corrected by our 9-plex PCR assay. This assay identified all the cryptic complex species from two test sets from Iran and China, correctly. Conclusions Our developed multiplex assay is accurate, specific, cost/time-saving, and works without the tedious DNA extraction steps. It could be integrated into routine clinical laboratories and as a reliable identification tool and has the potential to be implemented into epidemiological studies to broaden the limited knowledge of cryptic species complexes.

Abstract Background Opportunistic infections due to Candida species occur frequently in cancer patients because of their inherent immunosuppression. The aim of the present study was to investigate the epidemiology of yeast species from the oral cavity of patients during treatment for oncological and haematological malignancies. Methods MALDI-TOF was performed to identify yeasts isolated from the oral cavity of 350 cancer patients. Moreover, antifungal susceptibility testing was performed in according to CLSI guidelines (M27-A3). Results Among 162 yeasts and yeast-like fungi isolated from the oral cavity of cancer patients, Candida albicans was the most common species (50.6%), followed by Candida glabrata (24.7%), Pichia kudriavzevii (Candida krusei (9.9%)), Candida tropicalis (4.3%), Candida dubliniensis (3.7%), Kluyveromyces marxianus (Candida kefyr (3.7%)) and Candida parapsilosis (1%). In addition, uncommon yeast species i.e., Saprochaete capitata, Saccharomyces cerevisiae, Clavispora lusitaniae (C. lusitaniae) and Pichia kluyveri (C. eremophila) were recovered from oral lesions. Oral colonization by C. albicans, non-albicans Candida species and uncommon yeasts were as follow; 55%, 44% and 1%, whereas oral infection due to C. albicans was 33.3%, non-albicans Candida species 60.6%, and uncommon yeasts 6.1%. Poor oral hygiene and xerostomia were identified as independent risk factors associated with oral yeast colonization. The overall resistance to fluconazole was 11.7% (19/162). Low MIC values were observed for anidulafungin for all Candida and uncommon yeast species. Conclusions This current study provides insight into the prevalence and susceptibility profiles of Candida species, including emerging Candida species and uncommon yeasts, isolated from the oral cavity of Iranian cancer patients. The incidence of oral candidiasis was higher amongst patients with hematological malignancies. The majority of oral infections were caused by non-albicans Candida species which were often more resistant to anti-fungal agents. Our findings suggest that anidulafungin should be used as antifungal of choice for prophylaxis in clinically high-risk patients with documented oral colonization or infection.

Background and Purpose Vulvovaginal candidiasis (VVC) is a common problem in women. The purpose of this study was to identify of Candida species isolated from vulvovaginitis woman suffering vulvovaginitis refered to Ghaem Hospital, Mashhad, Iran, by use of MALDI-TOF mass spectrometry. Materials and Methods The 65 clinical samples isolated from Vulvovaginitis women were collected in Ghaem Hospital. All specimens were identified using phenotypic techniques such as microscopy and culture on Sabouraud dextrose agar and corn meal agar medium,Then, All isolates were detected and were processed for MALDI TOF MS identification. Results Of the 65 isolates analyzed, 61 (93.8%) were recognised by MALDI-TOF mass spectrometry and for four isolates (6.1%) only not relabile identifications were achieved. In this study, the most frequently isolated species were Candida albicans (58.5%), followed by Candida tropicalis (16.9%), Candida glabrata (7.7%), Candida parapsilosis (7.7%) and Candida guillermondii (3.1%). Conclusion presented results demonstrate that the MALDI TOF mass spectrometry is a fast and reliable technique, and has the potential to replace conventional phenotypic identification of Candida species and other yeast strains routinely isolated in clinical microbiology laboratories.

Cystic fibrosis (CF) patients are colonized with a multitude of bacteria and fungi. From respiratory samples of two CF patients in our institute, a difficult to identify yeast was isolated repeatedly. This yeast was eventually identified as Cutaneotrichosporon (Cryptococcus) cyanovorans by internal transcribed spacer (ITS) and ribosomal large subunit (LSU) sequencing. C. cyanovorans is a basidiomycetous yeast originally reported as environmental isolate from South African soil and has not been described before as clinical isolate from CF patients. Keywords: Cryptococcus cyanovorans, Cutaneotrichosporon cyanovorans, Cystic fibrosis, MADLITOF-MS

Abstract Background Antifungal susceptibility testing is a subject of interest in the field of medical mycology. The aim of the present study were the distributions and antifungal susceptibility patterns of various Candida species isolated from colonized and infected immunocompromised patients admitted to ten university hospitals in Iran. Methods In totally, 846 Candida species were isolated from more than 4000 clinical samples and identified by the API 20 C AUX system. Antifungal susceptibility testing was performed by broth microdilution method according to CLSI. Results The most frequent Candida species isolated from all patients was Candida albicans (510/846). The epidemiological cutoff value and percentage of wild-type species for amphotericin B and fluconazole in Candida albicans, Candida tropicalis, Candida glabrata and Candida krusei were 0.5 μg/ml (95%) and 4 μg/ml (96%); 1 μg/ml (95%) and 8 μg/ml (95%); 0.5 μg/ml (99%) and 19 μg/ml (98%); and 4 μg/ml (95%) and 64 μg/ml (95%), respectively. The MIC90 and epidemiological cutoff values to posaconazole in Candida krusei were 0.5 μg/ml. There were significant differences between infecting and colonizing isolates of Candida tropicalis in MIC 90 values of amphotericin B, and isolates of Candida glabrata in values of amphotericin B, caspofungin, and voriconazole (P

Cases of tuberculosis/cryptococcosis co-infection are rapidly increasing in China. However, most studies addressing this co-infection have been published in Chinese journals, and this publication strategy has obscured this disease trend for scientists in other parts of the world. Our investigation found that 62.9% of all co-infection cases worldwide were reported in the Chinese population (n=197) between 1965 and 2016, and 56.3% of these Chinese cases were reported after 2010. Nearly all cases originated from the warm and wet monsoon regions of China. HIV-positive subjects tended to correlate with more severe manifestations of a tuberculosis/cryptococcosis co-infection than those without HIV. Notablely, dual tubercular/cryptococcal meningitis was the most frequent (54.0%) and most easily misdiagnosed (95.2%, n=40/42) co-infection. We also found that the combined use of cerebrospinal fluid pressure and concentrations of glucose, protein and chlorine might be an inexpensive and effective indicator to differentiate tubercular/cryptococcal co-infection meningitis from tubercular meningitis and cryptococcal meningitis.Emerging Microbes & Infections (2017) 6, e73; doi:10.1038/emi.2017.61; published online 23 August 2017

Mucormycosis is a rare fungal infection caused by Mucor indicus. Phylogenetic analysis of many M. indicus isolates, mainly sampled from different clinical and environmental specimens collected worldwide, revealed two genotypes, I and II, based on ITS and D1/D2 LSU rDNA sequences. A retrospective review of the literature revealed 13 cases. Eight (76.9%) patients had disseminated infections, and the overall mortality rate was 30.7%. A pulmonary infection caused by M. indicus genotype I in a liver transplant recipient was disseminated to include the skin and was successfully treated with liposomal amphotericin B and aggressive surgery. M. indicus can infect a wide variety of patients with no real preference for the site of infection. We concluded that M. indicus has emerged as a significant cause of invasive mycosis in severely immunocompromised patients worldwide. Early diagnosis and initiation of appropriate therapy could enhance survival in these immunocompromised patient populations.Emerging Microbes & Infections (2017) 6, e63; doi:10.1038/emi.2017.51; published online 12 July 2017

Abstract The PREDICTS project—Projecting Responses of Ecological Diversity In Changing Terrestrial Systems (https://www.nhm.ac.uk/our-science/our-work/biodiversity/predicts.html)—has collated from published studies a large, reasonably representative database of comparable samples of biodiversity from multiple sites that differ in the nature or intensity of human impacts relating to land use. We have used this evidence base to develop global and regional statistical models of how local biodiversity responds to these measures. We describe and make freely available this 2016 release of the database, containing more than 3.2 million records sampled at over 26,000 locations and representing over 47,000 species. We outline how the database can help in answering a range of questions in ecology and conservation biology. To our knowledge, this is the largest and most geographically and taxonomically representative database of spatial comparisons of biodiversity that has been collated to date; it will be useful to researchers and international efforts wishing to model and understand the global status of biodiversity.

In 2006 several yeast-like fungi were isolated from apples that showed a postharvest disorder named “white haze.” These strains were morphologically and molecularly assigned to the genus Tilletiopsis. Following the recent reclassification of yeasts in Ustilaginomycotina and the genus Tilletiopsis in particular, species that caused “white haze” disorder were re-identified based on the phylogenetic analysis of five DNA-loci (ITS, LSU, SSU, RPB2, and TEF1) and analysis of D1/D2 domains of the 26S/28S rRNA (LSU). Six novel species belonging to three orders in the Exobasidiomycetes, namely Entyloma belangeri (holotype: CBS 111600; ex-type: DSM 29114) MB 823155, Entyloma davenportii (holotype: CBS 111604; ex-type: DSM 100135) MB 823154, Entyloma elstari (holotype: CBS 111593; ex-type: DSM 29113) MB 823153, Entyloma randwijkense (holotype: CBS 111606; ex-type: DSM 100136) MB 823156, Jamesdicksonia mali (holotype: CBS 111625; ex-type: DSM 29121) MB 823151 and Golubevia heteromorpha (holotype: CBS 111610; ex-type: DSM 100176) MB 823152 are proposed to accommodate these strains. In addition, sequences representing phylogenetically related but yet undescribed fungi were obtained from GenBank in order to show the diversity of Tilletiopsis-like yeast states in Exobasidiomycetes.

Despite the increasing occurrence of Candida orthopsilosis and Candida metapsilosis in clinical settings, little is known about their microbiological and clinical properties. Herein, we conducted a national retrospective study (2014–2019) from multiple centers in Iran. Among the 1,770 Candida isolates collected, we identified 600 Candida parapsilosis species complex isolates. Isolate identification was performed by 9-plex PCR, matrix-assisted laser desorption-time of flight mass spectrometry (MALDI-TOF MS), and rDNA sequencing, and antifungal susceptibility testing (AFST) followed CLSI M27-A3/S4; genotyping was performed by amplified fragment length polymorphism (AFLP) analysis; and clinical information was mined. Thirty-one isolates of C. orthopsilosis from various clinical sources, one mixed sample (blood) concurrently containing C. orthopsilosis and C. parapsilosis and one isolate of C. metapsilosis from a nail sample were identified. Although both 9-plex PCR and MALDI-TOF successfully identified all isolates, only 9-plex PCR could identify the agents in a mixed sample. For the C. orthopsilosis isolates, resistance (non-wild type) was noted only for itraconazole (n = 4; 12.5%). Anidulafungin and fluconazole showed the highest and voriconazole had the lowest geometric mean values. AFLP analysis showed three main and four minor genotypes. Interestingly, 90% of nail isolates clustered with 80% of the blood isolates within two clusters, and four blood isolates recovered from four patients admitted to a hospital clustered into two genotypes and showed a high degree of similarity (>99.2%), which suggests that C. orthopsilosis disseminates horizontally. Supported by our data and published case studies, C. orthopsilosis and C. metapsilosis can be linked to challenging clinical failures, and successful outcomes are not always mirrored by in vitro susceptibility. Accordingly, conducting nationwide studies may provide more comprehensive data, which is required for a better prognosis and clinical management of patients.

Aspergillus are ubiquitous mold species that infect immunocompetent and immunocompromised patients. The symptoms are diverse and range from allergic reactions, bronchopulmonary infection, and bronchitis, to invasive aspergillosis. The aim of this study was to characterize 70 Aspergillus isolates recovered from clinical specimens of patients with various clinical conditions presented at Hamad general hospital in Doha, Qatar, by using molecular methods and to determine their in vitro antifungal susceptibility patterns using the Clinical and Laboratory Standards Institute (CLSI) M38-A2 reference method. Fourteen Aspergillus species were identified by sequencing β-tubulin and calmodulin genes, including 10 rare and cryptic species not commonly recovered from human clinical specimens. Aspergillus welwitschiae is reported in this study for the first time in patients with fungal rhinosinusitis (n = 6) and one patient with a lower respiratory infection. Moreover, Aspergillus pseudonomius is reported in a patient with fungal rhinosinusitis which is considered as the first report ever from clinical specimens. In addition, Aspergillus sublatus is reported for the first time in a patient with cystic fibrosis. In general, our Aspergillus strains exhibited low MIC values for most of the antifungal drugs tested. One strain of Aspergillus fumigatus showed high MECs for echinocandins and low MICs for the rest of the drugs tested. Another strain of A. fumigatus exhibited high MIC for itraconazole and categorized as non-wild type. These findings require further analysis of their molecular basis of resistance. In conclusion, reliable identification of Aspergillus species is achieved by using molecular sequencing, especially for the emerging rare and cryptic species. They are mostly indistinguishable by conventional methods and might exhibit variable antifungal susceptibility profiles. Moreover, investigation of the antifungal susceptibility patterns is necessary for improved antifungal therapy against aspergillosis.

Fungal infections such as those caused by Candida species are increasingly common complications in immunocompromised patients. The list of causative agents of candidiasis is growing and comprises a set of emerging species whose relative global incidence is rare but recurrent. This is the case of Candida inconspicua, which prevalence has increased 10-fold over the last years. To gain novel insights into the emergence of this opportunistic pathogen and its genetic diversity, we performed whole genome sequencing of the type strain (CBS180), and of 10 other clinical isolates. Our results revealed high levels of genetic heterozygosity structured in non-homogeneous patterns, which are indicative of a hybrid genome shaped by events of loss of heterozygosity (LOH). All analyzed strains were hybrids and could be clustered into two distinct clades. We found large variability across strains in terms of ploidy, patterns of LOH, and mitochondrial genome heterogeneity that suggest potential admixture between hybrids. Altogether, our results identify a new hybrid species with virulence potential toward humans and underscore the potential role of hybridization in the emergence of novel pathogenic lineages.

Candida auris was first reported in an ear swab from Japan in 2009; it then promptly spread over five continents and turned into a global nosocomial problem. The main challenges faced by many researchers are the mis-identification by conventional methods in clinical laboratories and failure in treatment. About 90% of C. auris strains are intrinsically resistant to fluconazole (FLU), and it is developing resistance to multiple classes of available antifungals. Echinocandins are the most potent class of antifungals against C. auris; however, reduced susceptibility to one or many echinocandin drugs has been recently observed. Thus, the main issues addressed in this paper are the fast and accurate identification of C. auris derived from Sabouraud dextrose agar and blood culture bottles as well as the rapid antifungal susceptibility test by MALDI-TOF MS. This study successfully identified all isolates of C. auris (n = 50) by MALDI-TOF MS, with an average log score of ≥ 2. An accuracy of 100% was found on both agar plate and blood culture bottles. MALDI Biotyper antibiotic susceptibility test-rapid assay (MBT ASTRA) was used for rapid antifungal susceptibility testing (AFST). A comparison between MBT ASTRA and the Clinical and Laboratory Standards Institute guidelines (CLSI) detected a sensitivity and specificity of 100% and 98% for anidulafungin, and 100% and 95.5% for micafungin, respectively. A categorical agreement of 98% and 96% was calculated for the two methods. For caspofungin, sensitivity and specificity of 100 and 73% were found, respectively, with a categorical agreement of 82%. MBT ASTRA has the great potential to detect C. auris isolates non-susceptible against echinocandin antifungals within 6 h, which makes it a promising candidate for AFST in clinical laboratories in the future.

Occurrence of non-Candida albicans Candida (NCAC) species that are associated with elevated MIC values and therapeutic failures are increasing. As a result, timely and accurate means of identification to the species level is becoming an essential part of diagnostic practices in clinical settings. In this study, 301 clinically isolated yeast strains recovered from various anatomical sites [Blood (n = 145), other sites (n = 156)] were used to assess the accuracy and practicality of API 20C AUX and 21-plex PCR compared to MALDI-TOF MS and large subunit rDNA (LSU rDNA). MALDI-TOF MS correctly identified 98.33% of yeast isolates, 100% of top five Candida species, 95.7% of rare yeast species, while 1.3% of isolates were misidentified. API 20C AUX correctly identified 83.7% of yeast isolates, 97.2% of top five Candida species, 61.8% of rare yeast species, while 16.2% of yeast isolates were misidentified. The 21-plex PCR, accurately identified 87.3% of yeast isolates, 100% of top five Candida species, 72% of rare yeast species, but it misidentified 1.3% of rare yeast species while 9.9% of whole yeast isolates were not identified. The combination of rapidity of 21-plex PCR and comprehensiveness of API 20C AUX, led to correct identification of 92% of included yeast isolates. Due to expensiveness of MALDI-TOF MS and sequencing, this combination strategy could be the most accurate and inexpensive alternative identification strategy for developing countries. Moreover, by the advent and development of cost-effective, reliable, and rapid PCR machines that cost 130 US dollars, 21-plex could be integrated in routine laboratories of developing and resource-limited countries to specifically identify 95% causative agents of yeast-related infections in human. Databases of MALDI-TOF MS, API 20C AUX, and the number of target species identified by 21-plex require further improvement to keep up with the diverse spectrum of yeast species.

Background: Oral candidiasis (OC) has a profound effect on the life quality of immunocompromised patients, such as those undergoing chemotherapy. Objective: Systematic investigation of clinical outcome and microbiological features of yeast isolates recovered from the oral cavity of 150 Iranian patients with hematological malignancies. Design: MALDI-TOF MS, 21-plex PCR, and rDNA sequencing were used for identification. Antifungal susceptibility testing (broth microdilution, CLSI M27-A3/S4) and genotypic diversity of yeast isolates (amplified fragment length polymorphism) were assessed. Results: Nystatin treatment resulted in 70% therapeutic failure and administration of 150 mg fluconazole (FLZ) + nystatin for patients with OC relapse showed 70% clinical failure. Previous history of OC was significantly correlated with FLZ treatment requirement and nystatin failure (P = 0.005, α < 0.05). Candida albicans (80.3%) and Kluyveromyces marxianus (C. kefyr) (12.7%) were the two most prevalent yeast species isolated. FLZ and AMB exhibited the highest geometric mean values. 21-PCR showed 98.9% agreement with MALDI-TOF MS. K. marxianus isolates had the same genotype, while C. albicans isolates grouped in 15 genotypes. Conclusions: Marked rate of therapeutic failure of nystatin necessitated OC treatment with systemic antifungals. K. marxianus was the second most prevalent yeast and 21-plex PCR could be considered as an inexpensive identification tool.

Cryptococcal meningitis is an important fungal infection among systemic lupus erythematosus patients. We conducted a pooled analysis and systematic review to describe the epidemiological and clinical profile of cryptococcal meningitis in systemic lupus erythematosus patients. From two hospitals in China and nine literature databases, cases and prevalence data were collected for pooled analysis and meta-analysis, respectively. Categorical variables of cases were compared using a χ2-test on the statistical program of SAS. A multiple regression analysis was performed to ascertain independent predictors significantly correlated with prognosis. Meta-analysis was conducted by the statistical program of R. The prevalence of cryptococcal meningitis in systemic lupus erythematosus patients was 0.5%. Patients were predominantly females and adults. A prednisone equivalent of more than 30 mg/day before infection was associated with higher mortality (odds ratio (OR)=9.69 (1.54, 60.73)). In all, 36.8–38.9% patients showed low lupus activity when they developed the crytococcal infection. Moreover, 38.2% of the patients were misdiagnosed. The estimated case-fatality rate was 23.6%. Our results suggest that more emphasis should be placed to further understand lupus-related cryptococcal meningitis and to develop better prophylaxis and management strategies to combat this condition.

Human fungal pathogens are attributable to a significant economic burden and mortality worldwide. Antifungal treatments, although limited in number, play a pivotal role in decreasing mortality and morbidities posed by invasive fungal infections (IFIs). However, the recent emergence of multidrug-resistant Candida auris and Candida glabrata and acquiring invasive infections due to azole-resistant C. parapsilosis, C. tropicalis, and Aspergillus spp. in azole-naïve patients pose a serious health threat considering the limited number of systemic antifungals available to treat IFIs. Although advancing for major fungal pathogens, the understanding of fungal attributes contributing to antifungal resistance is just emerging for several clinically important MDR fungal pathogens. Further complicating the matter are the distinct differences in antifungal resistance mechanisms among various fungal species in which one or more mechanisms may contribute to the resistance phenotype. In this review, we attempt to summarize the burden of antifungal resistance for selected non-albicansCandida and clinically important Aspergillus species together with their phylogenetic placement on the tree of life. Moreover, we highlight the different molecular mechanisms between antifungal tolerance and resistance, and comprehensively discuss the molecular mechanisms of antifungal resistance in a species level.

Candida auris is an emerging opportunistic yeast species causing nosocomial outbreaks at a global scale. A few studies have focused on the C. auris genotypic structure. Here, we compared five epidemiological typing tools using a set of 96 C. auris isolates from 14 geographical areas. Isolates were analyzed by microsatellite typing, ITS sequencing, amplified fragment length polymorphism (AFLP) fingerprint analysis, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS), and Fourier-transform infrared (FTIR) spectroscopy methods. Microsatellite typing grouped the isolates into four main clusters, corresponding to the four known clades in concordance with whole genome sequencing studies. The other investigated typing tools showed poor performance compared with microsatellite typing. A comparison between the five methods showed the highest agreement between microsatellite typing and ITS sequencing with 45% similarity, followed by microsatellite typing and the FTIR method with 33% similarity. The lowest agreement was observed between FTIR spectroscopy, MALDI-TOF MS, and ITS sequencing. This study indicates that microsatellite typing is the tool of choice for C. auris outbreak investigations. Additionally, FTIR spectroscopy requires further optimization and evaluation before it can be used as an epidemiological typing method, comparable with microsatellite typing, as a rapid method for tracing nosocomial fungal outbreaks.