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OBJECTIVE: Candida auris has emerged as a health-care-associated and multidrug-resistant fungal pathogen of great clinical concern. As many as 50% of C. auris clinical isolates are reported to be resistant to amphotericin B, but no mechanisms contributing to this resistance have been identified. Here we describe a clinical case in which high-level amphotericin B resistance was acquired in vivo during therapy and undertake molecular and genetic studies to identify and characterize the genetic determinant of resistance. METHODS: Whole-genome sequencing was performed on four C. auris isolates obtained from a single patient case. Cas9-mediated genetic manipulations were then used to generate mutant strains harbouring mutations of interest, and these strains were subsequently subjected to amphotericin B susceptibility testing and comprehensive sterol profiling. RESULTS: A novel mutation in the C. auris sterol-methyltransferase gene ERG6 was found to be associated with amphotericin B resistance, and this mutation alone conferred a >32-fold increase in amphotericin B resistance. Comprehensive sterol profiling revealed an abrogation of ergosterol biosynthesis and a corresponding accumulation of cholesta-type sterols in isolates and strains harbouring the clinically derived ERG6 mutation. CONCLUSIONS: Together these findings definitively demonstrate mutations in C. auris ERG6 as the first identified mechanism of clinical amphotericin B resistance in C. auris and represent a significant step forward in the understanding of antifungal resistance in this emerging public health threat.

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.

Ines Matia-Garcia,1 Eduardo Vadillo,2 Rosana Pelayo,3 José F Muñoz-Valle,4 Mariel García-Chagollán,4 Jaqueline Loaeza-Loaeza,1 Amalia Vences-Velázquez,1 Lorenzo Salgado-Goytia,1 Samuel García-Arellano,4 Isela Parra-Rojas1 1Facultad de Ciencias Químico Biológicas, Universidad Autónoma de Guerrero, Chilpancingo, Guerrero, México; 2Unidad de Investigación Médica en Enfermedades Oncológicas, Hospital de Oncología, Centro Médico Nacional, Instituto Mexicano del Seguro Social, Ciudad de México, México; 3Centro de Investigación Biomédica de Oriente, Instituto Mexicano del Seguro Social, Atlixco, Puebla, México; 4Instituto de Investigación en Ciencias Biomédicas, Centro Universitario de Ciencias de la Salud, Universidad de Guadalajara, Guadalajara, Jalisco, MéxicoCorrespondence: Isela Parra-RojasLaboratorio de Investigación en Obesidad y Diabetes, Facultad de Ciencias Químico Biológicas, Universidad Autónoma de Guerrero, Avenida Lázaro Cárdenas S/N, Ciudad Universitaria, Chilpancingo, Guerrero, 39090, MéxicoTel +52 7474719310Email iprojas@yahoo.comPurpose: We aim to identify Th1 and Th2 cell clusters in young subjects, including their clinical and metabolic characteristics and the Th1/Th2 balance.Patients and Methods: A total of 100 participants were included. The frequencies of Th1 and Th2 cells in peripheral blood were determined by flow cytometry. Serum C-reactive protein was measured using a turbidimetric assay, and insulin levels were quantified with an enzyme-linked immunosorbent assay. Circulating cytokine levels were analyzed using a multiplex system.Results: A cluster analysis was performed to determine the Th1/Th2 balance in a group of young people, and 3 clusters were formed with the following characteristics: 1) subjects with a higher prevalence of hyperglycemia (38%), dyslipidemia (38– 75%), and insulin resistance (50%), as well as a higher percentage of Th1 cells and Th1/Th2 ratio, including elevated IFN-ɣ levels; 2) subjects with a lower prevalence of hyperglycemia (23%) and insulin resistance (15.4%), but a higher prevalence of dyslipidemia (8– 85%) with a predominance of Th2 cells, and lower Th1/Th2 ratio; 3) subjects with a lower prevalence of hyperglycemia (6%), insulin resistance (41%), and dyslipidemia (10– 63%), as well as a balance of Th1 and Th2 cells and lower Th1/Th2 ratio, including low IFN-ɣ levels. Positive correlations between Th1 cells with IFN-γ, IL-12, and IL-1β and between Th2 cells with IFN-γ, IL-2, and IL-4 were found (p < 0.05). A significant increase in Th1 cells was observed in the presence of hyperglycemia and high LDL-C levels, as well as increased Th2 cells in the absence of abdominal obesity and high blood pressure, including low HDL-C levels. The Th1/Th2 ratio was higher in the group with high cardiometabolic risk (p = 0.03).Conclusion: Th1/Th2 balance is related to metabolic abnormalities that may occur in young population, and thus the timely identification of different phenotypes may help predict an increased cardiometabolic risk.Keywords: Th1 cells, Th2 cells, hyperglycemia, dyslipidemias, IFN-ɣ levels

Background Blastomycosis has been reported from countries in Africa and the Middle East, but a decades-long debate has persisted regarding whether this is the same disease known in North America and caused by Blastomyces dermatitidis and Blastomyces gilchristii. Methods We reviewed published cases of human and veterinary blastomycosis from Africa and the Middle East. We abstracted epidemiological and clinical features of cases, including sites of disease, diagnosis, management, outcomes, and, where available, genetic and antigenic typing of case isolates. In addition, we sequenced nucleic acids from 9 clinical isolates from Africa deposited in global collections as B. dermatitidis; for 5, we sequenced the internal transcribed spacer regions, and for the other 4 we sequenced the whole genomes. Results We identified 172 unique human patients with blastomycosis, including 159 patients from 25 African countries and 12 patients from 5 Middle Eastern countries, and also identified 7 reports of veterinary blastomycosis. In humans, cutaneous disease predominated (n = 100/137, 73%), followed by pulmonary (n = 73/129, 57%) and osteoarticular involvement (n = 61/128, 48%). Unusual direct microscopy/histopathological presentations included short hyphal fragments in tissues (n = 23/129, 18%). There were 34 genotyped case isolates that comprised 4 species: Blastomyces percursus (n = 22, 65%), from 8 countries throughout all regions; Blastomyces emzantsi (n = 9, 26%), from South Africa; B. dermatitidis (n = 1, 3%), from the Democratic Republic of Congo; and B. gilchristii (n = 2, 6%), from South Africa and Zimbabwe. Conclusions Blastomycosis occurs throughout Africa and the Middle East and is caused predominantly by B. percursus and, at least in South Africa, B. emzantsi, resulting in distinct clinical and pathological patterns of disease., Blastomycosis occurs throughout Africa and Middle East and is clinically and microbiologically distinct from the disease in North America. Cutaneous and osteoarticular involvement are common, and causative pathogens are Blastomyces percursus, Blastomyces emzantsi, and, rarely, Blastomyces dermatitidis and Blastomyces gilchristii.

Skin cancer is one of the main types of cancer worldwide, and non-melanoma skin cancer (NMSC) is the most frequent within this group. Basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) are the most common types. Multifactorial features are well-known for cancer development, and new hallmarks are gaining relevance. Genetics and epigenetic regulation play an essential role in cancer susceptibility and progression, as well as the variety of cells and molecules that interact in the tumor microenvironment. In this review, we provide an update on the genetic features of NMSC, candidate genes, and new therapies, considering diverse perspectives of skin carcinogenesis. The global health situation and the pandemic have been challenging for health care systems, especially in the diagnosis and treatment of patients with cancer. We provide innovative approaches to overcome the difficulties in the current clinical dynamics.

A western diet and increased intestinal permeability may contribute to systemic inflammation and the development of cardio-metabolic alterations. The aim of this study was to assess the relationship between diet, biomarkers of intestinal permeability, and chronic low-grade inflammation on the cardiometabolic profile. A cross-sectional study was carried out in 238 young subjects aged 18–29 years, divided into two groups: with

Candida auris has emerged as a healthcare-associated and multidrug-resistant fungal pathogen of great clinical concern. While as much as 50% of C. auris clinical isolates are reported to be resistant to amphotericin B, to date, no mechanisms contributing to this resistance have been identified. We report here mutations in the C. auris sterol-methyltransferase gene, ERG6, as the first identified mechanism of amphotericin B resistance in this emerging pathogen and describe the clinical case in which this high-level amphotericin B resistance was acquired in vivo during therapy. Whole genome sequencing revealed the four C. auris isolates obtained from this single patient case to be genetically related and identified a mutation in ERG6 as being associated with amphotericin B resistance. Cas9-mediated genetic manipulations confirmed this mutation alone to confer a >32-fold increase in amphotericin B resistance, and comprehensive sterol profiling revealed an abrogation of ergosterol biosynthesis and a corresponding accumulation of cholesta-type sterols in isolates and strains harboring the clinically derived ERG6 mutation. Together these findings represent the first significant step forward in the understanding of clinical amphotericin B resistance in C. auris.

Candida auris is a multidrug-resistant fungal pathogen that is endemic in South African hospitals. We tested bloodstream C. auris isolates that were submitted to a reference laboratory for national laboratory-based surveillance for candidemia in 2016 and 2017. We confirmed the species identification by phenotypic/molecular methods. We tested susceptibility to amphotericin B, anidulafungin, caspofungin, micafungin, itraconazole, posaconazole, voriconazole, fluconazole, and flucytosine using broth microdilution and Etest methods. We interpreted MICs using tentative breakpoints. We sequenced the genomes of a subset of isolates and compared them to the C. auris B8441 reference strain. Of 400 C. auris isolates, 361 (90%) were resistant to at least one antifungal agent, 339 (94%) to fluconazole alone (MICs of ≥32 µg/ml), 19 (6%) to fluconazole and amphotericin B (MICs of ≥2 µg/ml), and 1 (0.3%) to amphotericin B alone. Two (0.5%) isolates from a single patient were pan-resistant (resistant to fluconazole, amphotericin B, and echinocandins). Of 92 isolates selected for whole-genome sequencing, 77 clustered in clade III, including the pan-resistant isolates, 13 in clade I, and 2 in clade IV. Eighty-four of the isolates (91%) were resistant to at least one antifungal agent; both resistant and susceptible isolates had mutations. The common substitutions identified across the different clades were VF125AL, Y132F, K177R, N335S, and E343D in ERG11; N647T in MRR1; A651P, A657V, and S195G in TAC1b; S639P in FKS1HP1; and S58T in ERG3. Most South African C. auris isolates were resistant to azoles, although resistance to polyenes and echinocandins was less common. We observed mutations in resistance genes even in phenotypically susceptible isolates.

Histoplasma capsulatum, a dimorphic fungal pathogen, is the most common cause of fungal respiratory infections in immunocompetent hosts. Histoplasma is endemic in the Ohio and Mississippi River Valleys in the United States and also distributed worldwide. Previous studies revealed at least eight clades, each specific to a geographic location: North American classes 1 and 2 (NAm 1 and NAm 2), Latin American groups A and B (LAm A and LAm B), Eurasian, Netherlands, Australian and African, and an additional distinct lineage (H81) comprised of Panamanian isolates. Previously assembled Histoplasma genomes are highly fragmented, with the highly repetitive G217B (NAm 2) strain, which has been used for most whole genome-scale transcriptome studies, assembled into over 250 contigs. In this study, we set out to fully assemble the repeat regions and characterize the large-scale genome architecture of Histoplasma species. We re-sequenced five Histoplasma strains (WU24 (NAm 1), G217B (NAm 2), H88 (African), G186AR (Panama), and G184AR (Panama)) using Oxford Nanopore Technologies long-read sequencing technology. Here we report chromosomal-level assemblies for all five strains, which exhibit extensive synteny among the geographically distant Histoplasma isolates. The new assemblies revealed that RYP2, a major regulator of morphology and virulence, is duplicated in G186AR. In addition, we mapped previously generated transcriptome datasets onto the newly assembled chromosomes. Our analyses revealed that the expression of transposons and transposon-embedded genes are upregulated in yeast phase compared to mycelial phase in G217B and H88 strains. This study provides an important resource for fungal researchers and further highlights the importance of chromosomal-level assemblies in analyzing high-throughput datasets.ImportanceHistoplasma species are dimorphic fungi causing significant morbidity and mortality worldwide. These fungi grow as mold in the soil and as budding yeast within the human host. Histoplasma can be isolated from soil in diverse regions, including North America, South America, Africa and Europe. Phylogenetically distinct species of Histoplasma have been isolated and sequenced. However, for the commonly used strains, genome assemblies have been fragmented, leading to underutilization of genome-scale data. This study provides chromosome-level assemblies of the commonly used Histoplasma strains using long-read sequencing technology. Comparative analysis of these genomes shows largely conserved gene order within the chromosomes. Mapping existing transcriptome data on these new assemblies reveals clustering of transcriptionally co-regulated genes. Results of this study highlight the importance of obtaining chromosome-level assemblies in understanding the biology of human fungal pathogens.

Candida auris is an emerging fungal pathogen of rising concern due to global spread, the ability to cause healthcare-associated outbreaks, and antifungal resistance. Genomic analyses revealed that early contemporaneously detected cases of C. auris were geographically stratified into four major clades. While Clades I, III, and IV are responsible for ongoing outbreaks of invasive and multidrug-resistant infections, Clade II, also termed the East Asian clade, consists primarily of cases of ear infection, is often susceptible to all antifungal drugs, and has not been associated with outbreaks. Here, we generate chromosome-level assemblies of twelve isolates representing the phylogenetic breadth of these four clades and the only isolate described to date from Clade V. This Clade V genome is highly syntenic with those of Clades I, III, and IV, although the sequence is highly divergent from the other clades. Clade II genomes appear highly rearranged, with translocations occurring near GC-poor regions, and large subtelomeric deletions in most chromosomes, resulting in a substantially different karyotype. Rearrangements and deletion lengths vary across Clade II isolates, including two from a single patient, supporting ongoing genome instability. Deleted subtelomeric regions are enriched in Hyr/Iff-like cell-surface proteins, novel candidate cell wall proteins, and an ALS-like adhesin. Cell wall proteins from these families and other drug-related genes show clade-specific signatures of selection in Clades I, III, and IV. Subtelomeric dynamics and the conservation of cell surface proteins in the clades responsible for global outbreaks causing invasive infections suggest an explanation for the different phenotypes observed between clades.

Water resources management requires specialized computer tools that allow explicit integration of surface and groundwater fluxes, which generally have domains with different spatial discretization. On one hand, a surface hydrological domain, D1, is typically segmented in sub-basins, elevation contour bands or hydrological response units. These elements usually are represented by grids, triangles, or simple irregular polygons. In D1, the elements are connected to each other and incorporated into a drainage network that defines a surface topology, t1. On the other hand, an aquifer domain, D2, is organized in hydrogeological units, which can be represented by geometrical elements such as grids, triangulations, Voronoi or Quadratree diagrams. In D2, a regular connection is typically associated to structured meshes that defines a groundwater topology, t2. We present a new tool called GeoLinkage (v.geolinkage) that creates an ESRI-format linkage shapefile of the new surface-groundwater topology, t1-2. This python-based open-source tool has a graphical user interface (GUI) as an add-on for GRASS-GIS, which was constructed using Pygrass and Flopy libraries. It was developed to be used in WEAP-MODFLOV models, but it can also be used with other water resources management models. GeoLinkage allows processing models with reasonable computation times, which facilitates scenario analysis. It calculates the locations of the surface element geometries (nodes and arcs) using the GRASS platform and connects them to each element of a structured mesh in MODFLOW models. GeoLinkage was applied to obtain groundwater levels and coverage of water demand in Azapa Valley, a hyper-arid zone in the desert of Chile, where a grid of 70.305 cells and six fields with detailed geometry were processed in only 12 min.

While emerging fungi threaten global biodiversity, the paucity of fungal genome assemblies impedes thoroughly characterizing epidemics and developing effective mitigation strategies. Here, we generate de novo genomic assemblies for six outbreaks of the emerging pathogen Batrachochytrium salamandrivorans (Bsal). We reveal the European epidemic currently damaging amphibian populations to comprise multiple, highly divergent lineages demonstrating isolate-specific adaptations and metabolic capacities. In particular, we show extensive gene family expansions and acquisitions, through a variety of evolutionary mechanisms, and an isolate-specific saprotrophic lifecycle. This finding both explains the chytrid’s ability to divorce transmission from host density, producing Bsal’s enigmatic host population declines, and is a key consideration in developing successful mitigation measures., M.K. was supported by FWO aspirant grant number 1111119N and would like to thank D. Gullick and JJADMS for their support in computational and practicalities.

Candida aurisis globally recognized as an opportunistic fungal pathogen of high concern, due to its extensive multidrug-resistance (MDR). Still, molecular mechanisms of MDR are largely unexplored. This is the first account of genome wide evolution of MDR inC. aurisobtained through serialin vitroexposure to azoles, polyenes and echinocandins. We show the stepwise accumulation of multiple novel mutations in genes known and unknown in antifungal drug resistance, albeit almost all new forC. auris. Echinocandin resistance evolved through a codon deletion inFKS1accompanied by a substitution inFKS1hot spot 3. Mutations inERG3andCIS2further increased the echinocandin MIC. Decreased azole susceptibility was acquired through a gain of function mutation in transcription factorTAC1byielding overexpression of the drug efflux pump Cdr1; a segmental duplication of chromosome 1 containingERG11; and a whole chromosome 5 duplication, which containsTAC1b. The latter was associated with increased expression ofERG11, TAC1bandCDR2, but notCDR1. The simultaneous emergence of nonsense mutations inERG3andERG11, presumably leading to the abrogation of ergosterol synthesis, was shown to decrease amphotericin B susceptibility, accompanied with fluconazole cross resistance. A mutation inMEC3, a gene mainly known for its role in DNA damage homeostasis, further increased the polyene MIC. Overall, this study shows the alarming potential and diversity for MDR development inC. auris, even in a clade until now not associated with MDR (clade II), hereby stressing its clinical importance and the urge for future research.ImportanceC. aurisis a recently discovered human fungal pathogens and has shown an alarming potential for multi- and pan-resistance towards all classes of antifungals most commonly used in the clinic. Currently,C. aurishas been globally recognized as a nosocomial pathogen of high concern due to this evolutionary potential. So far, this is the first study in which the stepwise progression of MDR inC. aurisis monitoredin vitro. Multiple novel mutations in known ‘resistance genes’ and genes previously not or vaguely associated with drug resistance reveal rapid MDR evolution in aC. aurisclade II isolate. Additionally, this study shows thatin vitroexperimental evolution can be a powerful tool to discover new drug resistance mechanisms, although it has its limitations.

Candida auris is a newly emerging fungal pathogen of humans and has attracted considerable attention from both the clinical and basic research communities. Clinical isolates of C. auris are often resistant to one or more antifungal agents. To explore how antifungal resistance develops, we performed experimental evolution assays using a fluconazole-susceptible isolate of C. auris (BJCA001). After a series of passages through medium containing increasing concentrations of fluconazole, fungal cells acquired resistance. By sequencing and comparing the genomes of the parental fluconazole-susceptible strain and 26 experimentally evolved strains of C. auris, we found that a portion of fluconazole-resistant strains carried one extra copy of chromosome V. In the absence of fluconazole, C. auris cells rapidly became susceptible and lost the extra copy of chromosome V. Genomic and transcriptome sequencing (RNA-Seq) analyses indicate that this chromosome carries a number of drug resistance-related genes, which were transcriptionally upregulated in the resistant, aneuploid strains. Moreover, missense mutations were identified in the genes TAC1B, RRP6, and SFT2 in all experimentally evolved strains. Our findings suggest that the gain of an extra copy of chromosome V is associated with the rapid acquisition of fluconazole resistance and may represent an important evolutionary mechanism of antifungal resistance in C. auris.

Paracoccidioidomycosis (PCM) is a life-threatening systemic mycosis widely reported in the Gran Chaco ecosystem. The disease is caused by different species from the genus Paracoccidioides, which are all endemic to South and Central America. Here, we sequenced and analyzed 31 isolates of Paracoccidioides across South America, with particular focus on isolates from Argentina and Paraguay. The de novo sequenced isolates were compared with publicly available genomes. Phylogenetics and population genomics revealed that PCM in Argentina and Paraguay is caused by three distinct Paracoccidioides genotypes, P. brasiliensis (S1a and S1b) and P. restrepiensis (PS3). P. brasiliensis S1a isolates from Argentina are frequently associated with chronic forms of the disease. Our results suggest the existence of extensive molecular polymorphism among Paracoccidioides species, and provide a framework to begin to dissect the connection between genotypic differences in the pathogen and the clinical outcomes of the disease. Fil: Teixeira, Marcus de Melo. Universidade do Brasília; Brasil Fil: Cattana, Maria Emilia. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste; Argentina Fil: Matute, Daniel R.. University of North Carolina; Estados Unidos Fil: Muñoz, José F.. Broad Institute Of Mit And Harvard; Estados Unidos Fil: Arechavala, Alicia. Hospital Francisco J Muñiz; Argentina Fil: Isbell, Kristin. University of North Carolina; Estados Unidos Fil: Schipper, Rafael. Universidade do Brasília; Brasil Fil: Santiso, Gabriela Maria. Gobierno de la Ciudad de Buenos Aires. Hospital de Infecciosas "Dr. Francisco Javier Muñiz"; Argentina Fil: Tracogna, Fernanda. Gobierno de la Provincia de Chaco. Hospital Julio Cecilio Perrando.; Argentina Fil: Sosa, María de los Ángeles. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina Fil: Cech, Norma. Hospital 4 de Junio; Argentina Fil: Alvarado, Primavera. Instituto de Biomedicina Dr. Jacinto Convit; Venezuela Fil: Barreto, Laura. Instituto Superior de Formación Docente Salome Ureña; República Dominicana Fil: Chacón, Yone. Provincia de Salta. Ministerio de Salud Pública. Hospital del Milagro; Argentina Fil: Ortellado, Juana. Universidad Nacional de Asunción; Paraguay Fil: Lima, Cleoni Mendes de. Universidade Federal de Rondonia; Brasil Fil: Chang, Marilene Rodrigues. Universidade Federal do Mato Grosso do Sul; Brasil Fil: Niño Vega, Gustavo. Universidad de Guanajuato; México Fil: Yasuda, Maria Aparecida Shikanai. Universidade de Sao Paulo; Brasil Fil: Felipe, Maria Sueli Soares. Universidade Catolica de Brasilia; Brasil Fil: Negroni, Ricardo. Gobierno de la Ciudad de Buenos Aires. Hospital de Infecciosas "Dr. Francisco Javier Muñiz"; Argentina Fil: Cuomo, Christina A.. Broad Institute of MIT And Harvard; Estados Unidos Fil: Barker, Bridget. Tgen Northern Arizona University; Estados Unidos Fil: Giusiano, Gustavo Emilio. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina. Universidad Nacional del Nordeste. Facultad de Medicina; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste; Argentina

In less than a decade, C. auris has emerged in health care settings worldwide; this species is capable of colonizing skin and causing outbreaks of invasive candidiasis. In contrast to other Candida species, C. auris is unique in its ability to spread via nosocomial transmission and its high rates of drug resistance. As part of the public health response, whole-genome sequencing has played a major role in characterizing transmission dynamics and detecting new C. auris introductions. Through a global collaboration, we assessed genome evolution of isolates of C. auris from 19 countries. Here, we described estimated timing of the expansion of each C. auris clade and of fluconazole resistance, characterized discrete phylogeographic population structure of each clade, and compared genome data to sensitivity measurements to describe how antifungal resistance mechanisms vary across the population. These efforts are critical for a sustained, robust public health response that effectively utilizes molecular epidemiology., Candida auris has emerged globally as a multidrug-resistant yeast that can spread via nosocomial transmission. An initial phylogenetic study of isolates from Japan, India, Pakistan, South Africa, and Venezuela revealed four populations (clades I, II, III, and IV) corresponding to these geographic regions. Since this description, C. auris has been reported in more than 30 additional countries. To trace this global emergence, we compared the genomes of 304 C. auris isolates from 19 countries on six continents. We found that four predominant clades persist across wide geographic locations. We observed phylogeographic mixing in most clades; clade IV, with isolates mainly from South America, demonstrated the strongest phylogeographic substructure. C. auris isolates from two clades with opposite mating types were detected contemporaneously in a single health care facility in Kenya. We estimated a Bayesian molecular clock phylogeny and dated the origin of each clade within the last 360 years; outbreak-causing clusters from clades I, III, and IV originated 36 to 38 years ago. We observed high rates of antifungal resistance in clade I, including four isolates resistant to all three major classes of antifungals. Mutations that contribute to resistance varied between the clades, with Y132F in ERG11 as the most widespread mutation associated with azole resistance and S639P in FKS1 for echinocandin resistance. Copy number variants in ERG11 predominantly appeared in clade III and were associated with fluconazole resistance. These results provide a global context for the phylogeography, population structure, and mechanisms associated with antifungal resistance in C. auris.

Introduction: Strongyloidiasis is a prevailing helminth infection ubiquitous in tropical and subtropical areas, however, seroprevalence data are scarce in migrant populations, particularly for those coming for Asia. Methods: This study aims at evaluating the prevalence of S. stercoralis at the hospital level in migrant populations or long term travellers being attended in out-patient and in-patient units as part of a systematic screening implemented in six Spanish hospitals. A cross-sectional study was conducted and systematic screening for S. stercoralis infection using serological tests was offered to all eligible participants. Results: The overall seroprevalence of S. stercoralis was 9.04% (95%CI 7.76&ndash, 10.31). The seroprevalence of people with a risk of infection acquired in Africa and Latin America was 9.35% (95%CI 7.01&ndash, 11.69), 9.22% (7.5&ndash, 10.93), respectively. The number of individuals coming from Asian countries was significantly smaller and the overall prevalence in these countries was 2.9% (95%CI &minus, 0.3&ndash, 6.2). The seroprevalence in units attending potentially immunosuppressed patients was significantly lower (5.64%) compared with other units of the hospital (10.20%) or Tropical diseases units (13.33%) (p &lt, 0.001). Conclusions: We report a hospital-based strongyloidiasis seroprevalence of almost 10% in a mobile population coming from endemic areas suggesting the need of implementing strongyloidiasis screening in hospitalized patients coming from endemic areas, particularly if they are at risk of immunosuppression.

Dimorphic fungal pathogens cause a significant human disease burden and unlike most fungal pathogens affect immunocompetent hosts. Most dimorphic fungi are found in the family Ajellomycetaceae, including the genera Histoplasma, Blastomyces, Paracoccidioides, and the recently described Emergomyces. To examine the origin of virulence and host adaptation in these fungal pathogens, we compared the gene content of classic systemic, opportunistic, and non-pathogenic species, including new genomes for Emmonsia species and two closely non-pathogenic species, Helicocarpus griseus and Polytolypa hystricis. We examined differences in gene content between pathogens and environmental fungi, and found that gene families related to plant degradation, synthesis of secondary metabolites, and amino acid and lipid metabolism are retained in H. griseus and P. hystricis. Genes of central importance in dimorphic pathogenic fungi such as heat shock response proteins, morphological switch related proteins are conserved in non-pathogenic species. However, we found that changes in the copy number of proteases, phosphotransferases and transcription factors in systemic dimorphic relative to non-dimorphic species may have aided the evolution of specialized gene regulatory programs to rapidly adapt to higher temperatures and new nutritional environments. Notably, we found that both more basal non-pathogenic species appear homothallic, with both mating type locus idiomorphs fused at a single locus, whereas all related pathogenic species are heterothallic. These differences revealed that independent changes in nutrient acquisition capacity but similar host/substrate shift from plants to animals have occurred in the Onygenaceae and Ajellomycetaceae, and highlighted how the dimorphic pathogens from Ajellomycetaceae have adapted to the host immune response and decreased their capacity for life stages in environmental niches.

Dimorphic human pathogenic fungi interact with host effector cells resisting their microbicidal mechanisms. Yeast cells are able of surviving within the tough environment of the phagolysosome by expressing an antioxidant defense system that provides protection against host-derived reactive oxygen species (ROS). This includes the production of catalases (CATs). Here we identified and analyzed the role of CAT isoforms in Paracoccidioides, the etiological agent of paracoccidioidomycosis. Firstly, we found that one of these isoforms was absent in the closely related dimorphic pathogen Coccidioides and dermatophytes, but all of them were conserved in Paracoccidioides, Histoplasma and Blastomyces species. We probed the contribution of CATs in Paracoccidioides by determining the gene expression levels of each isoform through quantitative RT-qPCR, in both the yeast and mycelia phases, and during the morphological switch (transition and germination), as well as in response to oxidative agents and during interaction with neutrophils. PbCATP was preferentially expressed in the pathogenic yeast phase, and was associated to the response against exogenous H2O2. Therefore, we created and analyzed the virulence defects of a knockdown strain for this isoform, and found that CATP protects yeast cells from H2O2 generated in vitro and is relevant during lung infection. On the other hand, CATA and CATB seem to contribute to ROS homeostasis in Paracoccidioides cells, during endogenous oxidative stress. CAT isoforms in Paracoccidioides might be coordinately regulated during development and dimorphism, and differentially expressed in response to different stresses to control ROS homeostasis during the infectious process, contributing to the virulence of Paracoccidioides.

We reevaluated 20 cases of blastomycosis diagnosed in South Africa between 1967 and 2014, with Blastomyces dermatitidis considered to be the etiological agent, in light of newly described species and the use of more advanced technologies. In addition to histopathological and/or culture-based methods, all 20 isolates were phenotypically and genotypically characterized, including multilocus typing of five genes and whole-genome sequencing. Antifungal susceptibility testing was performed as outlined by Clinical and Laboratory Standards Institute documents M27-A3 and M38-A2. We merged laboratory and corresponding clinical case data, where available. Morphological characteristics and phylogenetic analyses of five-gene and whole-genome sequences revealed two groups, both of which were closely related to but distinct from B. dermatitidis, Blastomyces gilchristii, and Blastomyces parvus. The first group (n = 12) corresponded to the recently described species Blastomyces percursus, and the other (n = 8) is described here as Blastomyces emzantsi sp. nov. Both species exhibited incomplete conversion to the yeast phase at 37°C and were heterothallic for mating types. All eight B. emzantsi isolates belonged to the α mating type. Whole-genome sequencing confirmed distinct species identities as well as the absence of a full orthologue of the BAD-1 gene. Extrapulmonary (skin or bone) disease, probably resulting from hematogenous spread from a primary lung infection, was more common than pulmonary disease alone. Voriconazole, posaconazole, itraconazole, amphotericin B, and micafungin had the most potent in vitro activity. Over the 5 decades, South African cases of blastomycosis were caused by species that are distinct from B. dermatitidis. Increasing clinical awareness and access to simple rapid diagnostics may improve the diagnosis of blastomycosis in resource-limited countries.

The Propopis tamarugo Phil, also known as Tamarugo, is an endemic and protected tree that survives in the Atacama Desert&mdash, a hyper arid and highly saline environment. The Tamarugo is threatened because of groundwater overexploitation, and its preservation depends on the soil moisture in the vadose zone, as many of the tree roots do not reach the current water table levels. To improve the estimation of soil moisture available for the Tamarugo trees, we applied a hydrogeological model that couples the unsaturated and saturated zones. The model was used to represent different groundwater exploitation and recharge scenarios between February 2006 and September 2030 to predict simultaneously groundwater levels and soil moisture. The model results show that even at locations where water table depletion is relatively small (~1&ndash, 1.5 m), soil moisture can drastically decrease (0.25&ndash, 0.30 m3/m3). Therefore, Tamarugo survival can be better addressed, as the applied model provides a management tool to estimate response of Tamarugo trees to changing soil moisture. To further improve the model and its use to assess Tamarugo survival, more field data, such as soil hydrodynamic properties and soil moisture, should be collected. Additionally, relationships between the state of the Tamarugo trees and soil moisture should be further constructed. In this way, the developed model will be able to predict future conditions associated to the Tamarugo&rsquo, s health state.

Candida auris is an emerging fungal pathogen of rising concern due to its increasing incidence, its ability to cause healthcare-associated outbreaks and antifungal resistance. Genomic analysis revealed that early cases of C. auris that were detected contemporaneously were geographically stratified into four major clades. Clade II, also termed East Asian clade, consists of the initial isolates described from cases of ear infection, is less frequently resistant to antifungal drugs and to date, the isolates from this group have not been associated with outbreaks. Here, we generate nearly complete genomes (“telomere-to-telomere”) of an isolate of this clade and of the more widespread Clade IV. By comparing these to genome assemblies of the other two clades, we find that the Clade II genome appears highly rearranged, with 2 inversions and 9 translocations resulting in a substantially different karyotype. In addition, large subtelomeric regions have been lost from 10 of 14 chromosome ends in the Clade II genomes. We find that shorter telomeres and genome instability might be a consequence of a naturally occurring loss-of-function mutation in DCC1 exclusively found in Clade II isolates, resulting in a hypermutator phenotype. We also determine that deleted subtelomeric regions might be linked to clade-specific adaptation as these regions are enriched in Hyr/Iff-like cell surface proteins, novel candidate cell surface proteins, and an ALS-like adhesin. The presence of these cell surface proteins in the clades responsible for global outbreaks causing invasive infections suggests an explanation for the different phenotypes observed between clades.IMPORTANCECandida auris was unknown prior to 2009 and since then it has quickly spread around the world, causing outbreaks in healthcare facilities and representing a high fraction of candidemia cases in some regions. The emergence of C. auris is a major concern, since it is often multidrug-resistant, easily spread between patients, and causes invasive infections. While isolates from three global clades cause invasive infections, isolates from Clade II primarily cause ear infections and have not been implicated in outbreaks, though cases of Clade II infections have been reported on different continents. Here, we describe genetic differences between Clade II and Clades I, III and IV, including a loss-of-function mutation in a gene associated with telomere length maintenance and genome stability, and the loss of cell wall proteins involved in adhesion and biofilm formation, that may suggest an explanation for the lower virulence and potential for transmission of Clade II isolates.

Rheumatoid arthritis is a clinical autoimmune syndrome that causes joint damage. The positive or negative anti-cyclic citrullinated protein (CCP) antibodies serodiagnosis differentiates two subsets of the disease, each with different genetic background. Previous studies have identified associations between KIR genes and rheumatoid arthritis but not with anti-CCP serodiagnosis. Therefore, we investigated the proportion of patients seropositive and seronegative to anti-CCP and its possible association with KIR (killer cell immunoglobulin-like receptor) genes. We included 100 patients with rheumatoid arthritis from western Mexico, who were determined for anti-CCP serodiagnosis by ELISA, and 16 KIR genes were genotyped by PCR-SSP. The proportion of seropositive anti-CCP patients was 83%, and they presented a higher frequency of KIR2DL2 genes than the seronegative group (73.6% vs. 46.2%, p = 0.044) which, in turn, presented a higher KIR2DL2-/KIR2DL3+ genotype frequency than the first ones (46.2% vs. 17.2%, p = 0.043). These results suggest different KIR genetic backgrounds for each subset of the disease according to anti-CCP serodiagnosis.La artritis reumatoide es un síndrome clínico autoinmune que causa daño en las articulaciones. El serodiagnóstico positivo o negativo para anticuerpos proteicos anticíclicos citrulinados (CCP) diferencia dos subconjuntos de la enfermedad, cada uno con diferente fondo genético. Estudios previos han identificado asociaciones entre los genes killer cell immunoglobulin- like receptor (KIR) y la artritis reumatoide, pero no con el serodiagnóstico de anti-CCP. Por lo tanto, investigamos la proporción de seropositividad y seronegatividad anti-CCP y su posible asociación con genes KIR. Se incluyeron 100 pacientes con artritis reumatoide del occidente de México, a quienes se les determinó su serodiagnóstico anti-CCP por ELISA y también se les realizó genotipificación de 16 genes KIR por PCR-SSP. La proporción de pacientes seropositivos anti-CCP fue del 83% y presentaron una mayor frecuencia génica KIR2DL2 que el grupo seronegativo (73.6% vs. 46.2%, p = 0.044), estos últimos presentaron mayor frecuencia genotípica KIR2DL2-/KIR2DL3+ que los primeros (46.2% vs. 17.2%, p = 0.043). Los resultados sugieren diferente fondo genético KIR para cada subconjunto de la enfermedad, de acuerdo con el serodiagnóstico anti-CCP.

Rheumatoid arthritis is a clinical autoimmune syndrome that causes joint damage. The positive or negative anti-cyclic citrullinated protein (CCP) antibodies serodiagnosis differentiates two subsets of the disease, each with different genetic background. Previous studies have identified associations between KIR genes and rheumatoid arthritis but not with anti-CCP serodiagnosis. Therefore, we investigated the proportion of patients seropositive and seronegative to anti-CCP and its possible association with KIR (killer cell immunoglobulin-like receptor) genes. We included 100 patients with rheumatoid arthritis from western Mexico, who were determined for anti-CCP serodiagnosis by ELISA, and 16 KIR genes were genotyped by PCR-SSP. The proportion of seropositive anti-CCP patients was 83%, and they presented a higher frequency of KIR2DL2 genes than the seronegative group (73.6% vs. 46.2%, p = 0.044) which, in turn, presented a higher KIR2DL2-/ KIR2DL3+ genotype frequency than the first ones (46.2% vs. 17.2%, p = 0.043). These results suggest different KIR genetic backgrounds for each subset of the disease according to anti-CCP serodiagnosis. La artritis reumatoide es un síndrome clínico autoinmune que causa daño en las articulaciones. El serodiagnóstico positivo o negativo para anticuerpos proteicos anti-cíclicos citrulinados (CCP) diferencia dos subconjuntos de la enfermedad, cada uno con diferente fondo genético. Estudios previos han identificado asociaciones entre los genes killer cell immunoglobulin- like receptor (KIR) y la artritis reumatoide, pero no con el serodiagnóstico de anti-CCP. Por lo tanto, investigamos la proporción de seropositividad y seronegatividad anti-CCP y su posible asociación con genes KIR. Se incluyeron 100 pacientes con artritis reumatoide del occidente de México, a quienes se les determinó su serodiagnóstico anti-CCP por ELISA y también se les realizó genotipificación de 16 genes KIR por PCR-SSP. La proporción de pacientes seropositivos anti-CCP fue del 83% y presentaron una mayor frecuencia génica KIR2DL2 que el grupo seronegativo (73.6% vs. 46.2%, p = 0.044), estos últimos presentaron mayor frecuencia genotípica KIR2DL2-/KIR2DL3+ que los primeros (46.2% vs. 17.2%, p = 0.043). Los resultados sugieren diferente fondo genético KIR para cada subconjunto de la enfermedad, de acuerdo con el serodiagnóstico anti-CCP.

Volume 10, no. 1, e02529-18, 2019, [https://doi.org/10.1128/mBio.02529-18][1]. [Figure 1A][2] spotting images were incorrectly duplicated for the Candida albicans and Candida auris plates. We confirmed the essentiality of HSP90 expression in C. albicans and C. auris with multiple biological

The outcome of fungal infections depends on interactions with innate immune cells. Within a population of macrophages encountering Candida albicans, there are distinct host-pathogen trajectories; however, little is known about the molecular heterogeneity that governs these fates. Here we developed an experimental system to separate interaction stages and single macrophage cells infected with C. albicans from uninfected cells and assessed transcriptional variability in the host and fungus. Macrophages displayed an initial up-regulation of pathways involved in phagocytosis and proinflammatory response after C. albicans exposure that declined during later time points. Phagocytosed C. albicans shifted expression programs to survive the nutrient poor phagosome and remodeled the cell wall. The transcriptomes of single infected macrophages and phagocytosed C. albicans displayed a tightly coordinated shift in gene expression co-stages and revealed expression bimodality and differential splicing that may drive infection outcome. This work establishes an approach for studying host-pathogen trajectories to resolve heterogeneity in dynamic populations., The outcomes of the interactions between individual host cells and pathogens are heterogeneous. Here, the authors assess transcriptional variability in both host and pathogen during infection of macrophages with the fungus Candida albicans, using sorted subpopulations and single macrophages.

In this work, powders of ZnO-CeO2 nanocomposites were synthesized and with these powders ceramic pieces were shaped using a colloidal method (slip casting). The effect of the pieces on gases emitted by a motorcycle was then determined. The nanocomposites were characterized using X-ray diffraction (XRD), infrared (IR) – UV–visible absorption – diffuse reflectance and photoluminescence spectroscopies (PL), and scanning and transmission electron microscopies (TEM). The pieces sintered at 700 °C had a porosity of ∼ 12% and a grain size of the nanometer order. On coming into contact with the exhaust fumes emitted by the motorcycle, the ceramic pieces reduced the amount of polluting gases emitted by between ∼ 10% and ∼ 20%, depending on the speed of the motorcycle. It was found that the most effective of the composites for most gases examined was that containing 5% ceria, with a gas emission reduction higher than 15%, while for CO the 1% ceria ZnO-CeO2 composite was most effective, this time with a ~ 13% reduction.

The need to understand and simulate hydrological phenomena and their interactions, and the impact of anthropogenic and climate changes on natural environments have promoted the study of evaporation from bare soils in arid climates. In closed Altiplano basins, such as those encountered in arid and hyper arid basins in northern Chile, evaporation from shallow groundwater is the main source of aquifer depletion, and thus, its study is crucial for water resources management. The objective of this work is to understand the mechanisms of evaporation in saline soils with shallow water tables, in order to better quantify evaporation fluxes and improve our understanding of the water balance in these regions. To achieve this objective, a model that couples fluid flow with heat transfer was developed and calibrated using column experiments with saline soils from the Huasco salt flat basin, Chile. The model enables determination of both liquid and water vapour fluxes, as well as the location of the evaporation front. Experimental results showed that salt transport inside the soil profile modified the water retention curve, highlighting the importance of including salt transport when modelling the evaporation processes in these soils. Indeed, model simulations only agreed with the experimental data when the effect of salt transport on water retention curves was taken into account. Model results also showed that the evaporation front is closer to the soil surface as the water table depth reduces. Therefore, the model allows determining the groundwater level depth that results in disconnection of liquid fluxes in the vadose zone. A sensitivity analysis allowed understanding the effect of water-flux enhancements mechanisms on soil evaporation. The results presented in this study are important as they allow quantifying the evaporation that occurs in bare soils from Altiplano basins, which is typically the main water discharge in these closed basins. Copyright © 2016 John Wiley & Sons, Ltd.