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BACKGROUND: Paracoccidioidomycosis is an endemic mycosis caused by members of the Paracoccidioides genus. Brazil remains the focus area and, to a lesser extent, the disease has been reported from Argentina, Colombia and Venezuela. AIMS: A Venezuelan Paracoccidioides brasiliensis strain, isolated from a patient diagnosed with chronic multifocal paracoccidioidomycosis, was subjected to whole genome sequencing to provide more insight about Paracoccidioides outside the endemic focus area. METHODS: P. brasiliensis strain CBS 118890 was whole genome sequenced using nanopore; library preparation with the 'native barcoding genomic DNA kit' was followed by sequencing on Flongle and MinION flowcells. Batches of strain CBS 118890 were re-identified by sequencing the internal transcribed spacer (ITS) region, and final identification was made based on phylogenetic analysis. RESULTS: Surprisingly, the Venezuelan P. brasiliensis strain CBS 118890 turned out to be a Nannizziopsis species. The batches of this strain were ITS sequenced followed by phylogenetic analysis and resulted in the final identification of Nannizziopsis arthrosporioides. CONCLUSIONS: Nannizziopsis infections are commonly seen in a wide variety of reptiles, but are particularly rare in human infections. This case underlines the need for molecular characterization of cases that clinically mimic paracoccidioidomycosis but that are serologically negative for Paracoccidioides.

The greater wax moth, Galleria mellonella (L.) (Lepidoptera: Pyralidae), has gathered widespread interest in the biomedical research community as an alternative non-mammalian model to study bacterial and fungal infections in humans. Nevertheless, most laboratories rely on commercial breeding companies that grow these larvae in bulk. Variation in rearing conditions between these companies and shipping of the larvae affects the response of G. mellonella to infection, both decreasing the reproducibility and comparability of infection experiments across laboratories. To unlock the full potential of G. mellonella as an infection model, standardized rearing is essential to produce healthy larvae of uniform quality. To this end, we developed an optimized protocol using a high-nutritional artificial diet and cost-effective housing solutions to ensure optimal conditions during each stage of the G. mellonella life cycle. This resulted in the rapid generation of larvae with a short larval phase, large size, and increased weight compared to other protocols and commercial larvae. A survival analysis comparing the standardized larvae with commercially obtained larvae showed significantly higher survival of standardized reared larvae after infection with Candida albicans (CP Robin) Berkhout (Saccharomycetales), proving the higher quality of standardized reared larvae.

Sporotrichosis is a global occurring implantation (subcutaneous) mycosis, caused by Sporothrix species, usually affecting the skin and the lymphatic vessels, from where it can disseminate. Nowadays, the vast majority of sporotrichosis infections in Brazil are caused by zoonotic transmission of S. brasiliensis which is the main etiologic agent. We report a cutaneous disseminated case of this disease, observed in an immunocompetent farmer living in southern Brazil, the Sporothrix schenckii-infection was successfully treated with itraconazole.

Classic paracoccidioidomycosis (PCM) is a potentially deadly neglected tropical systemic mycosis caused by members of the Paracoccidioides brasiliensis complex (P. brasiliensis s. str., P. americana, P. restrepiensis, and P. venezuelensis) and P. lutzii. The laboratorial diagnosis of PCM relies on observing pathognomonic structures such as the “steering wheel” or “Mickey Mouse” shape in the direct mycological examination, fresh biopsied tissue in 10% KOH, histopathological analysis, and/or the isolation of the fungus in culture. However, these procedures are time-consuming and do not allow for the speciation of Paracoccidioides due to overlapping morphologies. Here, we propose a new one-tube multiplex probe-based qPCR assay to detect and recognize agents of the P. brasiliensis complex and P. lutzii. Primers (Paracoco-F and Paracoco-R) and TaqMan probes (PbraCx-Fam, Plu-Ned, and Paracoco-Vic) were developed to target the rDNA (ITS2/28S) in the Paracoccidioides genome. A panel of 77 Paracoccidioides isolates revealed a 100% specificity (AUC = 1.0, 95% CI 0.964–1.000, p < 0.0001) without cross-reacting with other medically relevant fungi or human and murine DNA. The lower limit of detection was 10 fg of gDNA and three copies of the partial rDNA amplicon. Speciation using qPCR was in perfect agreement with AFLP and TUB1-RFLP markers (kappa = 1.0). As a proof of concept, we assessed a panel of 16 formalin-fixed and paraffin-embedded specimens from histopathologically confirmed PCM patients to reveal a significant sensitivity of 81.25% and specificity of 100% (AUC = 0.906 ± 0.05, 95% CI = 0.756–0.979, p < 0.0001, Youden index J = 0.8125). Our assay achieved maximum sensitivity (100%) and specificity (100%) using fresh clinical samples (n = 9) such as sputum, bronchoalveolar lavage, and tissue fragments from PCM patients (AUC = 1.0, 95% CI 0.872–1.000, p < 0.0001, Youden index J = 1.0). Overall, our qPCR assay simplifies the molecular diagnosis of PCM and can be easily implemented in any routine laboratory, decreasing a critical bottleneck for the early treatment of PCM patients across a vast area of the Americas.

ObjectivesTo evaluate the in vitro activity of isavuconazole on 154 clinical and reference strains of Trichosporon asahii, Trichosporon asteroides, Trichosporon coremiiforme, Trichosporon faecale and Trichosporon inkin by using the EUCAST broth microdilution method (BMD) and Liofilchem MIC Test Strips (MTS).MethodsAntifungal susceptibility testing for isavuconazole, fluconazole, voriconazole and posaconazole was assessed by EUCAST E.DEF 7.3.2. MIC values of isavuconazole obtained by BMD after 48 h of incubation were compared with MTS MICs after 24 and 48 h of incubation.ResultsT. asahii and T. asteroides showed the highest isavuconazole MIC90 values (0.5 mg/L). In clinical isolates, T. asahii exhibited the highest MIC90 values (0.5 mg/L) compared with non-T. asahii (0.06–0.25 mg/L). The five non-WT T. asahii isolates for fluconazole, voriconazole and posaconazole also exhibited high MICs of isavuconazole (≥0.5 mg/L). A better correlation between MTS and BMD MICs was observed after 24 h incubation for all species tested. MTS measurements performed at 48 h increased by at least 122% the number of isolates with >2 dilutions compared with the standard method.ConclusionsIsavuconazole exhibited variable in vitro activity among the Trichosporon species tested, showing higher or equal MICs than the other azoles. The five non-WT T. asahii clinical isolates tested also exhibited high isavuconazole MICs, suggesting the occurrence of triazole cross-resistance. Our MTS data indicate that there is no advantage in extended reading time for MTS from 24 to 48 h for Trichosporon yeasts.

Genomic analyses are widely applied to epidemiological, population genetic and experimental studies of pathogenic fungi. A wide range of methods are employed to carry out these analyses, typically without including controls that gauge the accuracy of variant prediction. The importance of tracking outbreaks at a global scale has raised the urgency of establishing high-accuracy pipelines that generate consistent results between research groups. To evaluate currently employed methods for whole-genome variant detection and elaborate best practices for fungal pathogens, we compared how 14 independent variant calling pipelines performed across 35 Candida auris isolates from 4 distinct clades and evaluated the performance of variant calling, single-nucleotide polymorphism (SNP) counts and phylogenetic inference results. Although these pipelines used different variant callers and filtering criteria, we found high overall agreement of SNPs from each pipeline. This concordance correlated with site quality, as SNPs discovered by a few pipelines tended to show lower mapping quality scores and depth of coverage than those recovered by all pipelines. We observed that the major differences between pipelines were due to variation in read trimming strategies, SNP calling methods and parameters, and downstream filtration criteria. We calculated specificity and sensitivity for each pipeline by aligning three isolates with chromosomal level assemblies and found that the GATK-based pipelines were well balanced between these metrics. Selection of trimming methods had a greater impact on SAMtools-based pipelines than those using GATK. Phylogenetic trees inferred by each pipeline showed high consistency at the clade level, but there was more variability between isolates from a single outbreak, with pipelines that used more stringent cutoffs having lower resolution. This project generated two truth datasets useful for routine benchmarking of C. auris variant calling, a consensus VCF of genotypes discovered by 10 or more pipelines across these 35 diverse isolates and variants for 2 samples identified from whole-genome alignments. This study provides a foundation for evaluating SNP calling pipelines and developing best practices for future fungal genomic studies.

Invasive fungal infections caused by non-albicans Candidaspecies are increasingly reported. Recent advances in diagnostic and molecular tools enabled better identification and detection of these emerging pathogenic yeasts. Several of these emerging species belong to theCandida haemuloniispecies complex, which attracted much attention due to the rapid global emergence of its multi-drug resistant memberCandida auris. Here, we describe a new clinically relevant yeast isolated from geographically distinct regions, representing the proposed novel speciesCandida khanbhai, member of theC. haemuloniispecies complex. Moreover, several members of theC. haemuloniispecies complex were observed to be invalidly described, includingCandida aurisandCandida vulturna. Hence, the opportunity was taken to correct this here, formally validating the names of various yeast species.

This report is the first investigation of yeast biodiversity from the oligotrophic hypersaline coastal waters of the Arabian Gulf surrounding Qatar. Yeasts and yeast-like fungi, were cultured from seawater sampled at 13 coastal areas surrounding Qatar over a period of 2 years (December 2013-September 2015). Eight hundred and forty-two isolates belonging to 82 species representing two phyla viz., Ascomycota (23 genera) and Basidiomycota (16 genera) were identified by molecular sequencing. The results indicated that the coastal waters of the Qatari oligotrophic marine environment harbor a diverse pool of yeast species, most of which have been reported from terrestrial, clinical and aquatic sources in various parts of the world. Five species, i.e., Candida albicans, C. parapsilosis, C. tropicalis, Pichia kudriavzevii and Meyerozyma guilliermondii (n = 252/842; 30% isolates) are known as major opportunistic human pathogens. Fifteen species belonging to nine genera (n = 498/842; 59%) and 12 species belonging to seven genera (n = 459/842; 55%) are hydrocarbon degrading yeast and pollution indicator yeast species, respectively. Ascomycetous yeasts were predominant (66.38%; 559/842) as compared to their basidiomycetous counterparts (33.6%; 283/842). The most isolated yeast genera were Candida (28%; 236/842) (e.g., C. aaseri, C. boidinii, C. glabrata, C. intermedia, C. oleophila, C. orthopsilosis, C. palmioleophila, C. parapsilosis, C. pseudointermedia, C. rugopelliculosa, C. sake, C. tropicalis and C. zeylanoides), Rhodotorula (12.7%; 107/842), Naganishia (8.4%; 71/842), Aureobasidium (7.4%; 62/842), Pichia (7.3%; 62/842), and Debaryomyces (6.4%; 54/842). A total of eleven yeast species ( n = 38) isolated in this study are reported for the first time from the marine environment. Chemical testing demonstrated that seven out of the 13 sites had levels of total petroleum hydrocarbons (TPH) ranging from 200 to 900 µg/L, whereas 6 sites showed higher TPH levels (> 1000-21000 µg/L). The results suggest that the yeast community structure and density are impacted by various physico-chemical factors, namely total organic carbon, dissolved organic carbon and sulphur.

We report a severe case of rhinothecal cryptococcoma in a 13-year-old female African Grey Parrot (Psittacus erithacus). The bird was born and bred in captivity in Portugal. The beak was deformed and showed several round soft masses, and microscopic examination revealed the presence of cells suggestive for Cryptococcus. Phenotypic and molecular analyses identified the obtained yeast culture as C. bacillisporus (AFLP5/VGIII). By phylogenetic analysis the parrot strain clustered with clinical C. bacillisporus strains from Mexico.

Sporothrix schenckii and allied species are thermodimorphic fungi widely distributed in nature which causes human and animal sporotrichosis, the most common subcutaneous mycosis globally. Sporotrichosis is acquired after a traumatic inoculation of soil or plant material contaminated with Sporothrix propagules or through bites and scratches from diseased cats. In Ascomycota, the master regulators of sex are MAT genes that lie in a single mating-type locus, in Sporothrix these are determined by two nonhomologous alleles, MAT1-1 and MAT1-2. We assessed the whole-genome sequences of medically relevant Sporothrix to develop a single-tube duplex PCR assay to screen S. brasiliensis, S. schenckii, S. globosa, and S. luriei idiomorphs (MAT1-1 or MAT1-2) and understand the distribution and incidence of mating-type strains from natural populations. Using our duplex PCR assay, a 673 bp amplicon (α-box protein) was consistently amplified from all MAT1-1 isolates, while a 291 bp fragment was only amplified from the isolates harboring MAT1-2 (HMG box). Molecular evidence suggests heterothallism (self-sterility) as the unique mating strategy among the species evaluated. The mating-type identity of 93 isolates revealed a nearly equal distribution (1:1 ratio) of mating type alleles within species but deviating between different outbreak areas. Remarkably, for S. brasiliensis in Rio de Janeiro, we report an overwhelming occurrence of MAT1-2 (1:13 ratio; χ2 = 10.286, P = 0.0013) opposing the high prevalence MAT1-1 in the Rio Grande do Sul (10:1 ratio; χ2 = 7.364, P = 0.0067). Therefore, the population structure of Sporothrix species refers from paucity to regular cycles of sexual recombination in most of the studied regions. Our PCR-based mating-type diagnostic assay is proposed here as an important marker to track the geographical expansion during the long-lasting outbreak of cat-transmitted sporotrichosis driven by S. brasiliensis.

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.

Trichosporon yeasts are classical agents of superficial mycoses, and they are ranked as the first to second predominant basidiomycetous yeast able to cause invasive infections. The clinical presentation of Trichosporon infections varies with the affected anatomical site, with fungaemia present in the majority of invasive trichosporonosis cases. Only a limited number of antifungal compounds can be used to treat Trichosporon infections. Azoles are the first choice due to their intrinsic resistance to echinocandins. Better laboratory methods and up-to-date databases of commercial platforms are required to improve identification, susceptibility testing and surveillance of this potentially threating infection.

INTRODUCTION: Chromoblastomycosis is a disease caused by melanized fungi, primarily belonging to the genera Fonsecaea and Cladophialophora, mainly affecting individuals who are occupationally exposed to soil and plant products. This research aimed to determine the clinical, epidemiological and laboratory characteristics of chromoblastomycosis in the state of Mato Grosso, Brazil. MATERIALS AND METHODS: Patients diagnosed with chromoblastomycosis treated at the Júlio Müller University Hospital, Cuiabá, Brazil, from January 2015 to December 2020, whose isolates were preserved in the Research Laboratory of the Faculty of Medicine of the Federal University of Mato Grosso. Isolates were identified by partly sequencing the Internal Transcribed Spacer (ITS) and β-tubulin (BT2) loci. AFLP fingerprinting was used to explore the genetic diversity. Susceptibility to itraconazole, voriconazole, 5-fluorocytosine, terbinafine and amphotericin B was determined by the broth microdilution technique. RESULTS: Ten patients were included, nine were male (mean age=64.1 years). Mean disease duration was 68 years. Lesions were mainly observed in the lower limbs. Predominant clinical forms were verrucous and scarring. Systemic arterial hypertension and type II diabetes mellitus were the predominant comorbidities. Leprosy was the main concomitant infectious disease. Fonsecaea pedrosoi was the unique etiological agent identified with moderate genetic diversity (H=0.3934-0.4527; PIC=0.3160-0.3502). Antifungal agents with the highest activity were terbinafine, voriconazole and itraconazole. CONCLUSION: Chromoblastomycosis is affecting the poor population in rural and urban areas, mainly related to agricultural activities, with F. pedrosoi being the dominant etiologic agent. All isolates had low MICs for itraconazole, voriconazole and terbinafine, confirming their importance as therapeutic alternatives for chromoblastomycosis.

OBJECTIVES: Diutina (Candida) catenulata is an ascomycetous yeast isolated from environmental sources and animals, occasionally infecting humans. The aim of this study is to shed light on the in vitro antifungal susceptibility and the genetic diversity of this opportunistic yeast. METHODS: Forty-five D. catenulata strains isolated from various sources (including human and environmental) and originating from nine countries, were included. Species identification was obtained by MALDI-TOF mass spectrometry and confirmed by ITS rDNA barcoding. In vitro antifungal susceptibility was determined for seven systemic antifungals by gradient strip method after 48h incubation at 35°C by Etest® (Biomérieux) or Liofilchem® strips. Isolates exhibiting fluconazole MICs ≥8 μg/mL were investigated for mutations in the ERG11 gene. A novel microsatellite genotyping scheme consisting of four markers was developed to assess the genetic diversity. RESULTS: MICs ranges for amphotericin B, caspofungin, micafungin, isavuconazole and posaconazole were [0.19-1], [0.094-0.5], [0.012-0.064], [0.003-0.047] and [0.006-0.032] μg/mL, respectively. By comparison, a broad range of MICs was noted for fluconazole (0.75 to >256 μg/mL), and voriconazole (0.012 to 0.38 mg/L), the higher values being observed among clinical strains. The Y132F amino acid substitution, associated with azole resistance in various Candida species (C. albicans, C. tropicalis, C. parapsilosis, C. orthopsilosis), was the main substitution identified. Although microsatellite typing showed extensive genetic diversity, most strains with high fluconazole MICs clustered together, suggesting human to human transmission or a common source of contamination. CONCLUSIONS: The high rate of acquired fluconazole resistance among clinical isolates of D. catenulata is of concern. We highlight a link between the genetic diversity of D. catenulata and its antifungal resistance patterns, suggesting possible clonal transmission of resistant isolates.

Members of the Cryptococcus gattii species complex are notorious causes of cryptococcosis as they often cause severe, life-threatening infections. Here we describe a case of a severe disseminated C. deuterogattii infection in a previously healthy patient who was initially treated with amphotericin B, 5-fluorocytosine and fluconazole, which led to a good neurological response, but the infection in the lungs remained unaltered and was not completely resolved until switching the antifungal therapy to isavuconazole. The infection was likely acquired during a one-month stay at the Azores Islands, Portugal. Environmental sampling did not yield any cryptococcal isolate; therefore, the source of this apparent autochthonous case could not be determined. Molecular typing showed that the cultured C. deuterogattii isolates were closely related to the Vancouver Island outbreak-genotype.

Introduction: Aspergillus exhibits a wide variation of susceptibility against antifungals according to genetic and environmental factors. Identification to the species level is necessary for appropriate treatment. Our objective was to determine the Aspergillus species involved in invasive pulmonary aspergillosis (IPA) among ICU patients in Jakarta, Indonesia. Methodology: The incidence of IPA in ICU patients at six hospitals in Jakarta from October 2012 – January 2015 was investigated. It involved a collection of endotracheal aspirates (ETA), nasal swabs and environmental samples around the hospitals, phenotypic screening, molecular characterization, and antifungal susceptibility testing. Results: Of the 405 patients investigated, 31 patients (7.7%) were diagnosed with putative IPA, from whom 45 Aspergillus isolates were collected. Aspergillus isolates were identified from pulmonary secretions in 24 patients, from nasal swabs in 7 patients and from both pulmonary secretions and nasal swabs in 7 patients. The phenotypic method showed 33 isolates of Aspergillus flavus (73.4%), nine Aspergillus fumigatus (20%), two Aspergillus niger (4.4%), and one Aspergillus nidulans (2.2%) isolate. Molecular identification showed 27 isolates of A. flavus (60.0%), eight isolates of A. fumigatus (17.8%), two isolates of A. niger (4.4%) and one isolate of A. nidulans (2.2%), while seven isolates (15.6%) were cryptic species or mixed isolates. Conclusions: Susceptibility testing showed all isolates were susceptible to amphotericin B, azoles and micafungin. Aspergillus flavus was the main causative organism in IPA cases in Jakarta, followed by A. fumigatus.

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.

BACKGROUND: Mucormycosis is classified as the third leading cause of invasive fungal disease in immunocompromised patients and is characterized by high morbidity and mortality (33%-56%). The aim of this study is to describe presentation, treatment and outcome of Dutch pediatric hemato-oncology patients recently diagnosed with mucormycosis and to review the literature to gain more insight specifically into contemporary outcome data. METHODS: Ten cases were diagnosed in the Princess Máxima Center for Pediatric Oncology from 2018 to 2021 and were retrospectively reviewed. In addition, 9 case series (n = 148) were included from literature. RESULTS: In our case series, 5 patients of 10 children (age 2-17 years) had disseminated invasive fungal disease. Four patients had localized pulmonary disease and 1 had a localized renal infection. One diagnosis was made postmortem. The underlying diseases were acute lymphoblastic leukemia (n = 6), acute myeloid leukemia (n = 2) and lymphoma (n=2). Seven patients received combination therapy comprising of a lipid amphotericin B formulation and a triazole, surgery was performed in 67%. All neutropenic patients received granulocyte transfusions and/or granulocyte colony-stimulating factor. Mucormycosis-related mortality was 20%. In the literature review, mucormycosis-related mortality was 36% for all patients and 66% for patients with disseminated disease. Survival rates were similar over the past 2 decades. The most common underlying disorder was acute lymphoblastic leukemia. Liposomal amphotericin B was the mainstay of treatment. Seventy percent of patients underwent surgery. CONCLUSIONS: Although survival of mucormycosis improved significantly overtime, it plateaued in the past decades. This series shows that with screening, early diagnostics and early antifungal and if possible surgical treatment, mortality is low and even disseminated disease is salvageable if approached aggressively with a combination of surgery and antifungal treatment. Further research focused on diagnostics, combination antifungal and adjunctive therapy is necessary to enhance the survival of mucormycosis in children.

Diutina catenulata is an ascomycetous yeast, that is regularly fluconazole-resistant and increasingly reported as the cause of invasive infection in humans. Here, we describe the de novo genome assembly of the clinical D. catenulata type-strain CBS565 and provide insights into the genome and compared it to an Illumina-sequenced environmental strain.

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.

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.

Sporothrix (order Ophiostomatales) comprises a genus with 53 species, of which S. brasiliensis, S. schenckii, S. globosa, and S. luriei cause skin infections in humans and other mammals. Remarkably, closely related Sporothrix can follow different strategies in epidemics. For example, during the cat-transmitted sporotrichosis, there is an increased prevalence of the highly virulent S. brasiliensis in South America, whereas S. schenckii and S. globosa are generally associated with a sapronotic route worldwide. Therefore, species-specific types of transmission may require distinct public health strategies to mitigate the advance of sporotrichosis, including early diagnosis, isolation of new animal cases, administration of adequate antifungal therapy, and population education on the main aspects of the disease. Here, we shed light on the system Sporothrix-sporotrichosis covering hot topics in the epidemiology and diagnosis of this important neglected disease.

Identification of Candida auris by conventional identification methodologies can be challenging. While whole genome sequencing is seen as the golden standard to genotype C. auris at an inter- and intraspecies level, it is costly and time-consuming. Sequencing the transcribed spacer (ITS) region and microsatellite typing provide simple, fast, and inexpensive alternatives for identification and genotyping of C. auris. Here we will describe both molecular approaches.

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.

Introduction: We report here the presence of Cryptococcus neoformans, and other potentially pathogenic yeasts and yeast-like fungi in environmental sources in Bonaire. Methods: Seventy environmental samples comprising 40 samples of old pigeon droppings, 18 of woody debris from hollows of living trees of Caesalpinia ('Divi Divi'), Ziziphus jujuba (Red Indian date), Tamarindus indica (Tamarind), Terminalia catappa (Tropical almond), Azadirachta indica (Neem) and 3 of other unidentified species of trees, 3 of latex from a rubber tree and 6 of coral dust were processed for isolation of pathogenic Cryptococcus spp. and other potentially pathogenic yeasts and yeast-like fungi. A variety of mycological media were employed. Identification of the isolates was done with conventional techniques and species identification was done by matrix-assisted laser desorption ionization-time of flight mass spectrometry. Results: Three of the 40 samples from old pigeon droppings yielded Cryptococcus neoformans, constituting the first record of environmental occurrence of this important pathogenic yeast in the Dutch Caribbean. Other potentially pathogenic yeasts and yeast-like fungi recovered from these environmental samples included 6 isolates each of Candida albicans, 8 of Candida parapsilosis, 4 each of Candida metapsilosis and Candida orthopsilosis, 2 each of Candida carpophila, Candida famata, Candida fabianii and Candida pelliculosa, 7 of Candida spp., 5 of Trichosporon spp. and 2 of Sporobolomyces spp. Conclusions: This study has demonstrated for the first time the occurrence of C. neoformans in a natural habitat in the Dutch Caribbean. The recovery of many species of potentially pathogenic yeast-like fungi and yeasts from environmental sources is remarkable.

Background Cryptococcal meningitis is most commonly found in HIV-infected patients. In HIV-negative patients, its low incidence can lead to prolonged time to diagnosis. Detailed case reports of chronic cryptococcal meningitis are scarce, but could provide clues for earlier diagnosis in this patient category. Case presentation A 60-year old man presented June 2015 with intermittent headaches for several months without any fever. Initial work-up showed a leukocytosis, raised CSF opening pressure and raised leukocytes and protein in the CSF. An MRI revealed leptomeningeal contrast enhancement and cerebellar oedema. While malignancy and various infectious causes were excluded, the patient had a spontaneous clinical and radiological recovery. One year later, the patient returned with complaints of headaches. Also, cerebellar oedema and leptomeningeal contrast enhancement had recurred. Eventually in March 2017, the novel cryptococcal antigen lateral flow assay (CrAg LFA) was positive on CSF, and one colony of Cryptococcus neoformans was cultured from CSF. The patient was treated with the standard antifungal regimen which resulted in resolution of his headaches. In retrospect, the cryptococcal antigen test was already positive on a serum sample from June 2015. Interestingly, post-treatment immunological analysis revealed both a low mannose-binding lectin (MBL) concentration and low naïve CD4 counts. Conclusions We present a patient with cryptococcal meningitis in an HIV-negative patient with low MBL and low naïve CD4 count suffering a chronic relapsing meningo-encephalitis with relatively mild symptoms for around 2 years. In patients with an unexplained meningo-encephalitis such as this case, early performance of CrAg LFA on serum and/or CSF is an inexpensive and rapid method to reduce time-to diagnosis.

Cryptococcosis is an infection caused by encapsulated basidiomycetous yeasts belonging to the Cryptococcus neoformans/Cryptococcus gattii species complexes. It is acquired through inhalation of infectious propagules, often resulting in meningitis and meningoencephalitis. The ecological niche of these agents is a wide variety of trees species, as well as pigeon, parrot and passerine excreta. The objective of this study was to isolate Cryptococcus yeasts from excreta of commercially traded parrots and passerines. The 237 samples were collected between October 2018 and April 2019 and processed using conventional methodologies. Nineteen colonies with a dark brown phenotype, caused by phenol oxidase activity, were isolated, suggesting the presence of pathogenic Cryptococcus yeasts. All isolates tested positive for urease activity. URA5-RFLP fingerprinting identified 14 isolates (68.4%) as C. neoformans (genotype AFLP1/VNI) and 5 (26.3%) as C. deuterogattii (genotype AFLP6/VGII). Multi-locus sequence typing was applied to investigate the relatedness of the C. deuterogattii isolates with those collected globally, showing that those originating from bird-excreta were genetically indistinguishable from some clinical isolates collected during the past two decades.

Paracoccidioidomycosis (PCM) is a life-threatening systemic fungal infection acquired after inhalation of Paracoccidioides propagules from the environment. The main agents include members of the P. brasiliensis complex (phylogenetically-defined species S1, PS2, PS3, and PS4) and P. lutzii. DNA-sequencing of protein-coding loci (e.g., GP43, ARF, and TUB1) is the reference method for recognizing Paracoccidioides species due to a lack of robust phenotypic markers. Thus, developing new molecular markers that are informative and cost-effective is key to providing quality information to explore genetic diversity within Paracoccidioides. We report using new amplified fragment length polymorphism (AFLP) markers and mating-type analysis for genotyping Paracoccidioides species. The bioinformatic analysis generated 144 in silico AFLP profiles, highlighting two discriminatory primer pairs combinations (#1 EcoRI-AC/MseI-CT and #2 EcoRI-AT/MseI-CT). The combinations #1 and #2 were used in vitro to genotype 165 Paracoccidioides isolates recovered from across a vast area of South America. Considering the overall scored AFLP markers in vitro (67-87 fragments), the values of polymorphism information content (PIC = 0.3345-0.3456), marker index (MI = 0.0018), effective multiplex ratio (E = 44.6788-60.3818), resolving power (Rp = 22.3152-34.3152), discriminating power (D = 0.5183-0.5553), expected heterozygosity (H = 0.4247-0.4443), and mean heterozygosity (H avp = 0.00002-0.00004), demonstrated the utility of AFLP markers to speciate Paracoccidioides and to dissect both deep and fine-scale genetic structures. Analysis of molecular variance (AMOVA) revealed that the total genetic variance (65-66 %) was due to variability among P. brasiliensis complex and P. lutzii (PhiPT = 0.651-0.658, P

The conventional baker's yeast, Saccharomyces cerevisiae, is the indispensable baking yeast of all times. Its monopoly coupled to its major drawbacks, such as streamlined carbon substrate utilisation base and a poor ability to withstand a number of baking associated stresses, prompt the need to search for alternative yeasts to leaven bread in the era of increasingly complex consumer lifestyles. Our previous work identified the inefficient baking attributes of Wickerhamomyces subpelliculosus and Kazachstania gamospora as well as preliminarily observations of improving the fermentative capacity of these potential alternative baker's yeasts using evolutionary engineering. Here we report on the characterisation and improvement in baking traits in five out of six independently evolved lines incubated for longer time and passaged for at least 60 passages relative to their parental strains as well as the conventional baker's yeast. In addition, the evolved clones produced bread with a higher loaf volume when compared to bread baked with either the ancestral strain or the control conventional baker's yeast. Remarkably, our approach improved the yeasts' ability to withstand baking associated stresses, a key baking trait exhibited poorly in both the conventional baker's yeast and their ancestral strains. W. subpelliculosus evolved the best characteristics attractive for alternative baker's yeasts as compared to the evolved K. gamospora strains. These results demonstrate the robustness of evolutionary engineering in development of alternative baker's yeasts.

Sporothrix schenckii and related species are the agents of human and animal sporotrichosis. Routine diagnoses using classical mycological approaches are unspecific due to overlapping phenotypes. As the frequency and prevalence of sporotrichosis increases worldwide, developing specific, sensitive and cost-effective diagnostic tools is essential to understand the distribution patterns, map-affected areas and promote specific public health strategies to mitigate future outbreaks. Polymorphisms among the β-tubulin gene were exploited to speciate S. brasiliensis, S. schenckii and S. globosa in a one-tube multiplex probe-based qPCR assay. A panel of 84 Sporothrix revealed 100% specificity (AUC = 1.000, 95% CI = 0.971-1.000, p

Candida auris has been reported worldwide, but only in December 2020, the first strain from a COVID-19 patient in Brazil was isolated. Here, we describe the genome sequence of this susceptible C. auris strain and performed variant analysis of the genetic relatedness with strains from other geographic localities.

Introduction Pigeon droppings are among the major environmental sources of Cryptococcus neoformans AFLP1/VNI, from where the organism infects susceptible humans and animals resulting in cryptococcosis. Until now, C. neoformans AFLP1B/VNII was the only molecular type reported in Nigeria. Effective clinical treatment of this infection has occasionally been stymied by the emergence of antifungal non-susceptible, and resistant strains of C. neoformans AFLP1/VNI. Hypothesis/Gap Statement Pigeon droppings harbour C. neoformans and HIV/AIDS patients are among the susceptible population to develop cryptococcal infection. Epidemiological data on cryptococcal prevalence is limited in Nigeria. Aim To investigate the environmental prevalence of C. neoformans in South-eastern Nigeria and compare the isolates with other lineages by using molecular and microbiological tools. Methodology A total of 500 pigeon droppings and 300 blood samples of HIV/AIDS patients were collected, respectively, from five market squares and three tertiary healthcare centres within the Nsukka area of South-eastern Nigeria. The antifungal susceptibility of the C. neoformans isolates to amphotericin B, fluconazole, 5-fluorocytosine, itraconazole, voriconazole, posaconazole, and isavuconazole was investigated based on the CLSI M27-A3 protocol. Yeasts were identified by MALDI-TOF MS, thereafter Cryptococcus MLST was performed according to the International Society for Human and Animal Mycology (ISHAM) consensus scheme. Results C. neoformans was recovered from 6 (1.2 %) pigeon droppings and 6 (2 %) blood cultures of HIV/AIDS patients. Molecular analyses indicated that all cryptococcal isolates belong to serotype A and the AFLP1/VNI molecular type with sequence type (ST)32. Infection with C. neoformans was independent of sex and age of the patients investigated. All C. neoformans isolates were susceptible to the seven antifungal agents. Conclusion This is the first report on the prevalence of C. neoformans AFLP1/VNI (ST32) in environmental and clinical samples from Nigeria. The antifungal susceptibility indicates that antifungal resistance by C. neoformans is yet a rare occurrence in Nigeria.

Chromoblastomycosis is a chronic disease caused by melanized fungi that mainly affect individuals performing soil-related labor. The objective of this study was to analyze the epidemiological and clinical characteristics of chromoblastomycosis in Latin America and the Caribbean by an extensive literature review. An integrative review was performed of English, French, Portuguese, and Spanish publications in LILACS, SciELO, PubMed, SCOPUS and Web of Science databases covering the period 1969–2019. A total of 1211 articles were identified, of which 132 were included in the review, covering 2081 patients, 80.3% were males, the mean age was 56.1 years. The mean duration of the disease was 10.8 years. The lesions were mainly described in the lower limbs (60%). The most frequent clinical forms were verrucous (46.4%) and tumorous (21.7%). Major disease symptoms and signs consisted of itching and pain. Bacterial infection and functional limitation were important complications. Immunosuppression post-kidney transplantation was the most frequent comorbidity while leprosy was the main concomitant infectious disease. Fonsecaea pedrosoi and Cladophialophora carrionii were the predominant etiological agents. Majority of the cured cases were treated with itraconazole as monotherapy or in combination with other antifungals, surgery or cryosurgery. Chromoblastomycosis affects hundreds of rural workers in Latin America and the Caribbean, causing disability and personal, family and economic losses. It is important to prioritize epidemiological surveillance and early diagnosis of this disease in order to reveal its real prevalence and direct resources to preventive actions, diagnosis and early treatment. Lay summary Chromoblastomycosis is a slowly progressing chronic disease caused by melanized fungi. We collected data from South America and the Caribbean covering 1969–2019, the 132 articles included 2081 patients, mean disease duration was 10.8 years. Fonsecaea pedrosoi and Cladophialophora carrionii predominated.

Sporothrix (Ophiostomatales) comprises species that are pathogenic to humans and other mammals as well as environmental fungi. Developments in molecular phylogeny have changed our perceptions about the epidemiology, host-association, and virulence of Sporothrix. The classical agent of sporotrichosis, Sporothrix schenckii, now comprises several species nested in a clinical clade with S. brasiliensis, S. globosa, and S. luriei. To gain a more precise view of outbreaks dynamics, structure, and origin of genetic variation within and among populations of Sporothrix, we applied three sets of discriminatory AFLP markers (#3 EcoRI-GA/MseI-TT, #5 EcoRI-GA/MseI-AG, and #6 EcoRI-TA/MseI-AA) and mating-type analysis to a large collection of human, animal and environmental isolates spanning the major endemic areas. A total of 451 polymorphic loci were amplified in vitro from 188 samples, and revealed high polymorphism information content (PIC = 0.1765–0.2253), marker index (MI = 0.0001–0.0002), effective multiplex ratio (E = 15.1720–23.5591), resolving power (Rp = 26.1075–40.2795), discriminating power (D = 0.9766–0.9879), expected heterozygosity (H = 0.1957–0.2588), and mean heterozygosity (Havp = 0.000007–0.000009), demonstrating the effectiveness of AFLP markers to speciate Sporothrix. Analysis using the program structure indicated three genetic clusters matching S. brasiliensis (population 1), S. schenckii (population 2), and S. globosa (population 3), with the presence of patterns of admixture amongst all populations. AMOVA revealed highly structured clusters (PhiPT = 0.458–0.484, P

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.

Objectives: Historically, the Brazilian Central-West region has had high numbers of paracoccidioidomycosis (PCM) cases caused by the dimorphic fungus Paracoccidioides lutzii. Methods: This epidemiological, observational, analytical, cross-sectional study was performed to investigate the clinical and laboratory data of 44 PCM patients with a culture-proven P. lutzii infection. All patients were referred to the Systemic Mycosis Center, Júlio Muller University Hospital, Cuiabá, Brazil, during January 2017 to March 2020. The neutrophil to lymphocyte ratio (NLR) was calculated and dichotomized by its median value to include in the identification of factors associated with severity. Results: At admission, 13 (31.7%) patients showed the disseminated multifocal chronic form of PCM and 16 (36.4%) patients met the clinical severity criteria. Treatment prescribed on admission did not follow the recommendations of the Brazilian Guideline for the Clinical Management of Paracoccidioidomycosis in 26% of the severe PCM cases (prevalence ratio 0.26, 95% confidence interval 0.14–0.49; P < 0.0001). Patients with severe PCM had a higher NLR that was greater than the median (≥4.11). Conclusions: The NLR biomarker complements the criteria for PCM severity. Applying the low-cost NLR test can greatly increase the diagnostic sensitivity when screening patients for PCM and contribute to better control of the disease, management of complications, and therapeutic strategies.

BACKGROUND PCM is a neglected systemic mycosis endemic in Brazil. The middle-west region of Brazil has shown the highest number of PCM by Paracoccidioides lutzii (P lutzii) cases. Differentiating cases of severe PCM from non-severe ones should be a concern at the bedside. Diagnosis of severe PCM by P lutzii is based on the subjectivity of clinical manifestations, which can result in a delay in starting its treatment and, consequently evolution to severe sequelae. There is not laboratory biomarker available to support the early diagnosis of severe PCM that is feasible for all the realities that coexist in Brazil. OBJECTIVES The aim of this study was to investigate the usefulness of laboratory biomarkers as erythrocyte sedimentation rate (ESR), C-reactive protein (CRP) and neutrophil/lymphocyte ratio (NLR) in the diagnosis of severe PCM. PATIENTS/METHODS ESR, CRP and NLR were analysed for 44 patients with PCM by P lutzii and a Receiver Operation Characteristic (ROC) curve were generated to identify the NLR cut-off point and point out the presence of severe PCM. RESULTS Sixteen (36.4%) had severe PCM and 28 (63.6%) had non-severe PCM. The mean NLR was higher and statistically significant among patients with severe PCM than among those with non-severe PCM. The area under the ROC curve was 0.859 for the diagnosis of severe PCM. The cut-off point for NLR for the diagnosis of severe PCM was 3.318 (sensitivity of 100%, specificity of 77%). CONCLUSIONS According to results, it is plausible to conclude that NLR represents a potential biomarker for the diagnosis of severe PCM.

More than a decade ago, the first cryptococcal lateral flow assay (LFA) became available. This affordable point-of-care test has undeniably led to tremendous improvements in early detection of cryptococcosis ([1][1], [2][2]). However, from several studies, it must be concluded that the reliability

We report on a family of three in a rabbit farm, especially children with Majocchi granuloma as the main clinical feature of cross-infection of Trichophyton mentagrophytes, provides experience in reducing the rate of clinical misdiagnosis. Based on the pathological features of Majocchi's granuloma, taking into account that severe inflammatory reactions are also responsible for the damage of the body. In Majocchi's granuloma treatment strategy, we recommend the use of anti-inflammatory Chinese medicines that have a hormonal effect and have no side effects. Keywords: Child, Rabbits, Trichophyton, Granuloma, Anti-inflammatory agents

Autosomal recessive (AR) CARD9 (caspase recruitment domain-containing protein 9) deficiency underlies invasive infections by fungi of the ascomycete phylum in previously healthy individuals at almost any age. Although CARD9 is expressed mostly by myeloid cells, the cellular basis of fungal infections in patients with inherited CARD9 deficiency is unclear. Therapy for fungal infections is challenging, with at least 20% premature mortality. We report two unrelated patients from Brazil and Morocco with AR CARD9 deficiency, both successfully treated with hematopoietic stem cell transplantation (HSCT). From childhood onward, the patients had invasive dermatophytic disease, which persisted or recurred despite multiple courses of antifungal treatment. Sanger sequencing identified homozygous missense CARD9 variants at the same residue, c.302G>T (p.R101L) in the Brazilian patient and c.301C>T (p.R101C) in the Moroccan patient. At the ages of 25 and 44 years, respectively, they received a HSCT. The first patient received a HLA-matched HSCT from his CARD9-mutated heterozygous sister. There was 100% donor chimerism at D + 100. The other patient received a T cell-depleted haploidentical HSCT from his CARD9-mutated heterozygous brother. A second HSCT from the same donor was performed due to severe amegakaryocytic thrombocytopenia despite achieving full donor chimerism (100%). At last follow-up, more than 3 years after HSCT, both patients have achieved complete clinical remission and stopped antifungal therapy. HSCT might be a life-saving therapeutic option in patients with AR CARD9 deficiency. This observation strongly suggests that the pathogenesis of fungal infections in these patients is largely due to the disruption of leukocyte-mediated CARD9 immunity.

Fungal peritonitis is frequent on peritoneal dialysis, with rare cases by Exophiala dermatitidis. A 25-month-old female admitted to the pediatric ICU with acute renal failure was submitted to peritoneal dialysis. After 10 days patient presented fever. Peritoneal fluid culture showed yeast colonies molecularly identified as E. dermatitidis. Patient was treated with voriconazole and hemodialysis. The literature was reviewed. Disseminated infections are frequently fatal, but appropriate diagnose and therapeutic led to cure in this case. Keywords: Phaeohyphomycosis, Fungal infection, Dialysis, Therapeutic management, Molecular diagnosis

Necropsy examination of an adult Atlantic salmon (Salmo salar) from the Dalälven River in Sweden revealed numerous large, white nodules, with spherical cysts and granulomata in kidney and liver. Histopathology showed dark, septate, thin-walled hyphae. The aetiologic agent was found to be an Ochroconis species (Venturiales) that differed from known fish-associated species of the genus. Molecular phylogenetic studies of the culture (strain UIII09 = CBS 135766) demonstrated that Ochroconis globalis was concerned. The isolate proved to be susceptible to all investigated antifungals, as it is known for another Ochroconis species. The role of Ochroconis in opportunism of cold-blooded animals was discussed, and the diagnostic methods using DNA sequences for routine identification of the fungus were proposed.

A 3-year-old boy presented with multiple lesions of tinea corporis with dermatophytids, and subsequent inflammatory lesions with alopecia on the scalp. At the beginning, topical clobetasone butyrate was prescribed. The infection was diagnosed as dermatophytosis on the basis of positive direct microscopy and fungal culture. The etiological agent was isolated from all sampled sites and identified as Trichophyton verrucosum. Clonal nature of the infection was confirmed by random amplified polymorphic DNA (RAPD) analysis. The child lived in close vicinity of cattle. He was successfully treated with itraconazole.

Aspergillus fumigatus resistant to azole as first-line therapy has been reported in azole-naïve patients. This worldwide resistance phenomenon has been linked to fungicide-driven alterations in the cyp51A gene and its promoter region (such as TR34/L98H and TR46/Y121F/T289A). Azole-resistant A. fumigatus related to the use of triazole fungicides in flower fields was recently reported In Colombia. The purpose of this study was to investigate the presence of azole-resistant A. fumigatus in soil samples from vegetable crops such as carrots, potatoes, maize, strawberries, and pea, and from prepared farming land surrounding the city of Bogotá. Species identification was based on sequencing of the β-tubulin and calmodulin genes. All A. fumigatus strains were screened for azole resistance on agar supplemented with itraconazole or voriconazole. Among the 60 soil samples, 34 (56.6%) were positive for A. fumigatus and 15 samples exhibited strains (n = 18) that grew on agar supplemented with itraconazole or voriconazole. Triazole-resistant strains were isolated from soil samples associated with carrot, potato, maize, and pea crops. Sequencing of the cyp51A gene and its promoter region indicated polymorphism, mainly with the presence of TR46/Y121F/T289A (n = 8), TR34/L98H, and TR53. Eight resistant isolates exhibited cyp51A wild type without alterations in the promoter region. Our study showed evidence of dissemination of azole-resistant A. fumigatus, with high genetic diversity, in vegetable crops in Colombia. These data underline the need to determine the prevalence of azole resistance in A. fumigatus in clinical and environmental settings for other regions of Colombia as well as Latin America.

Background Progressive lung injury in Cystic Fibrosis (CF) patients can lead to chronic colonization with bacteria and fungi. Fungal colonization is obtained from the environment which necessitates locally performed epidemiology studies. We prospectively analyzed respiratory samples of CF patients during a 3-year period, using a uniform fungal culture protocol, focusing on filamentous fungi and azole resistance in Aspergillus fumigatus. Methods Over a 3-year period, all respiratory specimens collected from CF patients in 5 Dutch CF centers, were analyzed. Samples were inoculated onto the fungal culture media Sabouraud dextrose agar (SDA) and Medium B+. All fungal isolates were collected and identified in one centre, using Amplified Fragment Length Polymorphism (AFLP) fingerprinting, rDNA PCR and ITS, calmodulin and β-tubulin sequencing. Azole resistance was assessed for all A. fumigatus using a qPCR assay followed by phenotypic confirmation. Results Filamentous fungi were recovered from 699 patients from at least one respiratory sample, corresponding with 3787 cultured fungal species. A. fumigatus was cultured most often with a mean prevalence of 31.7%, followed by Penicillium species (12.6%), non-fumigatus Aspergillus species (5.6%), Scedosporium species (4.5%) and Exophiala dermatitidis and Cladosporium species (1.1% each). In total 107 different fungal species were identified, with 39 Penicillium species and 15 Aspergillus species. Azole resistance frequency in A. fumigatus was 7.1%, with TR34/L98H being the dominant resistance mechanism. Conclusion A vast diversity of filamentous fungi was demonstrated, dominated by Aspergillus and Penicillium species. We observed a mean azole resistance prevalence of 7.1% of A. fumigatus culture positive patients.

Objectives: To improve the quality of invasive pulmonary aspergillosis (IPA) management for intensive care unit (ICU) patients using a practical diagnostic scoring model. Methods: This nested case-control study aimed to determine the incidence of IPA in 405 ICU patients, between July 2012 and June 2014, at 6 hospitals in Jakarta, Indonesia. Phenotypic identifications and galactomannan (GM) tests of sera and lung excreta were performed in mycology laboratory, Parasitology Department, Faculty of Medicine, Universitas Indonesia in Jakarta, Indonesia. Results: The incidence of IPA in the ICUs was 7.7% (31 of 405 patients). A scoring model used for IPA diagnosis showed 4 variables as the most potential risk factors: lung excreta GM index (score 2), solid organ malignancy (score 2), pulmonary tuberculosis (score 2), and systemic corticosteroids (score 1). Patients were included in a high-risk group if their score was greater than 2, and in a low-risk group if their score was less than 2. Conclusion: This study provides a novel diagnosis scoring model to predict IPA in ICU patients. Using this model, a more rapid diagnosis and treatment of IPA may be possible. The application of the diagnosis scoring should be preceded by specified pre-requisites. Saudi Med J 2019; Vol. 40 (2): 140-146 doi: 10.15537/smj.2019.2.22940 How to cite this article: Rozaliyani A, Sedono R, Jusuf A, Rumende CM, Aniwidyaningsih W, Burhan E, et al. A novel diagnosis scoring model to predict invasive pulmonary aspergillosis in the intensive care unit. Saudi Med J . 2019 Feb;40(2):140-146. doi: 10.15537/smj.2019.2.22940.