RESUMO
OBJECTIVES: The investigation of Candida auris outbreaks is needed to provide insights into its population structure and transmission dynamics. We genotypically and phenotypically characterised a C. auris nosocomial outbreak occurred in Consorcio Hospital General Universitario de Valencia (CHGUV), Spain. METHODS: Data and isolates were collected from CHGUV from September 2017 (first case) until September 2021. Thirty-five isolates, including one from an environmental source, were randomly selected for whole genome sequencing (WGS), and the genomes were analysed along with a database with 335 publicly available genomes, assigning them to one of the five major clades. In order to identify polymorphisms associated with drug resistance, we used the fully susceptible GCA_003014415.1 strain as reference sequence. Known mutations in genes ERG11 and FKS1 conferring resistance to fluconazole and echinocandins, respectively, were investigated. Isolates were classified into aggregating or non-aggregating. RESULTS: All isolates belonged to clade III and were from an outbreak with a single origin. They clustered close to three publicly available genomes from a hospital from where the first patient was transferred, being the probable origin. The mutation VF125AL in the ERG11 gene, conferring resistance to fluconazole, was present in all the isolates and one isolate also carried the mutation S639Y in the FKS1 gene. All the isolates had a non-aggregating phenotype (potentially more virulent). CONCLUSIONS: Isolates are genotypically related and phenotypically identical but one with resistance to echinocandins, which seems to indicate that they all belong to an outbreak originated from a single isolate, remaining largely invariable over the years. This result stresses the importance of implementing infection control practices as soon as the first case is detected or when a patient is transferred from a setting with known cases.
Assuntos
Antifúngicos , Candida auris , Candidíase , Infecção Hospitalar , Surtos de Doenças , Farmacorresistência Fúngica , Genótipo , Fenótipo , Sequenciamento Completo do Genoma , Humanos , Espanha/epidemiologia , Infecção Hospitalar/microbiologia , Infecção Hospitalar/epidemiologia , Candidíase/microbiologia , Candidíase/epidemiologia , Antifúngicos/farmacologia , Candida auris/genética , Candida auris/efeitos dos fármacos , Farmacorresistência Fúngica/genética , Testes de Sensibilidade Microbiana , Mutação , Masculino , Fluconazol/farmacologia , Feminino , Equinocandinas/farmacologia , Pessoa de Meia-Idade , Candida/genética , Candida/efeitos dos fármacos , Candida/classificação , Candida/isolamento & purificaçãoRESUMO
Despite the severe impact of uncommon yeast fungal infections and the pressing need for more research on the topic, there are still few studies available on the identification, epidemiology, and susceptibility profile of those pathogens. The aims of the current study were to define the profile of uncommon yeast species at Fattouma Bourguiba University Hospital using phenotypic, molecular, and proteomic methods and to study their antifungal susceptibility profile. Pre-identified uncommon yeast species were collected from 2018 to 2021. These isolates were further identified using phenotypic methods (ID32C® system and Vitek2® YST), matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), and sequencing. The antifungal susceptibility profile was studied using the reference CLSI broth microdilution method. In total, 30 strains were collected during the study period. Referring to the sequencing, the most isolated uncommon species were Saprochaete capitata, Candida lusitaniae, Candida kefyr, Candida inconspicua, and Candida guilliermondii. A total of 90% of isolates were correctly identified by MALDI-TOF MS compared to 76.7% and 63.3% by ID32® C and VITEK® 2 YST, respectively. The isolated species showed variable responses to antifungals. Candida guilliermondii showed increased azole minimum inhibitory concentrations. Misidentification of uncommon yeast species was common using commercial phenotypic methods. The high percentage of concordance of MALDI-TOF results with sequencing highlights its high performance and usefulness as a routine diagnosis tool.
There is still little information on the epidemiology of uncommon emergent yeasts, although their implication in severe diseases and mainly invasive infections. Thus, the importance of an accurate identification and antifungal susceptibility testing for a better monitoring of related infections.
Assuntos
Antifúngicos , Hospitais Universitários , Testes de Sensibilidade Microbiana , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Leveduras , Humanos , Antifúngicos/farmacologia , Tunísia , Leveduras/efeitos dos fármacos , Leveduras/isolamento & purificação , Leveduras/classificação , Leveduras/genética , Micoses/microbiologia , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Criança , Candida/efeitos dos fármacos , Candida/classificação , Candida/isolamento & purificação , Candida/genética , Pré-Escolar , Adolescente , Adulto Jovem , Idoso , Farmacorresistência FúngicaRESUMO
Meyerozyma guilliermondii (Candida guilliermondii) is one of the Candida species associated with invasive candidiasis. With the potential for expressing industrially important enzymes, M. guilliermondii strain SO possessed 99 % proteome similarity with the clinical ATCC 6260 isolate and showed pathogenicity towards zebrafish embryos. Recently, three secreted aspartyl proteinases (SAPs) were computationally identified as potential virulence factors in this strain without in vitro verification of SAP activity. The quantification of Candida SAPs activity in liquid broth were also scarcely reported. Thus, this study aimed to characterize M. guilliermondii strain SO's ability to produce SAPs (MgSAPs) in different conditions (morphology and medium) besides analyzing its growth profile. MgSAPs' capability to cleave bovine serum albumin (BSA) was also determined to propose that MgSAPs as the potential virulence factors compared to the avirulent Saccharomyces cerevisiae. M. guilliermondii strain SO produced more SAPs (higher activity) in yeast nitrogen base-BSA-dextrose broth compared to yeast extract-BSA-dextrose broth despite insignificantly different SAP activity in both planktonic and biofilm cells. FeCl3 supplementation significantly increased the specific protein activity (â¼40 %). The BSA cleavage by MgSAPs at an acidic pH was proven through semi-quantitative SDS-PAGE, sharing similar profile with HIV-1 retropepsin. The presented work highlighted the MgSAPs on fungal cell wall and extracellular milieu during host infection could be corroborated to the quantitative production in different growth modes presented herein besides shedding lights on the potential usage of retropepsin's inhibitors in treating candidiasis. Molecular and expression analyses of MgSAPs and their deletion should be further explored to attribute their respective virulence effects.
Assuntos
Ácido Aspártico Proteases , Biofilmes , Candidíase , Soroalbumina Bovina , Fatores de Virulência , Fatores de Virulência/metabolismo , Fatores de Virulência/genética , Ácido Aspártico Proteases/metabolismo , Ácido Aspártico Proteases/genética , Candidíase/microbiologia , Soroalbumina Bovina/metabolismo , Biofilmes/crescimento & desenvolvimento , Animais , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Meios de Cultura/química , Candida/patogenicidade , Candida/metabolismo , Candida/genética , Saccharomycetales/metabolismo , Saccharomycetales/patogenicidade , Saccharomycetales/genética , VirulênciaRESUMO
BACKGROUND: Candida vulturna is an emerging pathogen belonging to the Metshnikowiaceae family together with Candida auris and Candida haemulonii species complex. Some strains of this species were reported to be resistant to several antifungal agents. OBJECTIVES: This study aims to address identification difficulties, evaluate antiungal susceptibilities and explore the molecular mechanisms of azole resistance of Candida vulturna. METHODS: We studied five C. vulturna clinical strains isolated in three Colombian cities. Identification was performed by phenotypical, proteomic and molecular methods. Antifungal susceptibility testing was performed following CLSI protocol. Its ERG11 genes were sequenced and a substitution was encountered in azole resistant isolates. To confirm the role of this substitution in the resistance phenotype, Saccharomyces cerevisiae strains with a chimeric ERG11 gene were created. RESULTS: Discrepancies in identification methods are highlighted. Sequencing confirmed the identification as C. vulturna. Antifungal susceptibility varied among strains, with four strains exhibiting reduced susceptibility to azoles and amphotericin B. ERG11 sequencing showed a point mutation (producing a P135S substitution) that was associated with the azole-resistant phenotype. CONCLUSIONS: This study contributes to the understanding of C. vulturna's identification challenges, its susceptibility patterns, and sheds light on its molecular mechanisms of azole resistance.
Assuntos
Antifúngicos , Azóis , Candida , Candidíase , Testes de Sensibilidade Microbiana , Antifúngicos/farmacologia , Azóis/farmacologia , Candida/efeitos dos fármacos , Candida/genética , Candida/classificação , Candida/isolamento & purificação , Candidíase/microbiologia , Humanos , Farmacorresistência Fúngica Múltipla/genética , Colômbia , Anfotericina B/farmacologia , Farmacorresistência Fúngica/genética , Mutação Puntual , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/efeitos dos fármacos , Sistema Enzimático do Citocromo P-450/genética , Proteínas Fúngicas/genética , Análise de Sequência de DNA , Proteínas de Saccharomyces cerevisiaeRESUMO
Lignocellulose (dry plant biomass) is an abundant cheap inedible residue of agriculture and wood industry with great potential as a feedstock for biotechnological processes. Lignocellulosic substrates can serve as valuable resources in fermentation processes, allowing the production of a wide array of chemicals, fuels, and food additives. The main obstacle for cost-effective conversion of lignocellulosic hydrolysates to target products is poor metabolism of the major pentoses, xylose and L-arabinose, which are the second and third most abundant sugars of lignocellulose after glucose. We study the oversynthesis of riboflavin in the flavinogenic yeast Candida famata and found that all major lignocellulosic sugars, including xylose and L-arabinose, support robust growth and riboflavin synthesis in the available strains of C. famata. To further increase riboflavin production from xylose and lignocellulose hydrolysate, genes XYL1 and XYL2 coding for xylose reductase and xylitol dehydrogenase were overexpressed. The resulting strains exhibited increased riboflavin production in both shake flasks and bioreactors using diluted hydrolysate, reaching 1.5 g L-1.
Assuntos
Candida , Lignina , Engenharia Metabólica , Riboflavina , Xilose , Lignina/metabolismo , Riboflavina/metabolismo , Riboflavina/biossíntese , Candida/metabolismo , Candida/genética , Xilose/metabolismo , Aldeído Redutase/metabolismo , Aldeído Redutase/genética , Fermentação , Reatores Biológicos/microbiologia , D-Xilulose Redutase/metabolismo , D-Xilulose Redutase/genética , Arabinose/metabolismoRESUMO
Candida albicans causes millions of mucosal infections in humans annually. Hyphal overgrowth on mucosal surfaces is frequently associated with tissue damage caused by candidalysin, a secreted peptide toxin that destabilizes the plasma membrane of host cells thereby promoting disease and immunopathology. Candidalysin was first identified in C. albicans strain SC5314, but recent investigations have revealed candidalysin "variants" of differing amino acid sequence in isolates of C. albicans, and the related species C. dubliniensis, and C tropicalis, suggesting that sequence variation among candidalysins may be widespread in natural populations of these Candida species. Here, we analyzed ECE1 gene sequences from 182 C. albicans isolates, 10 C. dubliniensis isolates, and 78 C. tropicalis isolates and identified 10, 3, and 2 candidalysin variants in these species, respectively. Application of candidalysin variants to epithelial cells revealed differences in the ability to cause cellular damage, changes in metabolic activity, calcium influx, MAPK signalling, and cytokine secretion, while biophysical analyses indicated that variants exhibited differences in their ability to interact with and permeabilize a membrane. This study identifies candidalysin variants with differences in biological activity that are present in medically relevant Candida species. IMPORTANCE: Fungal infections are a significant burden to health. Candidalysin is a toxin produced by Candida albicans that damages host tissues, facilitating infection. Previously, we demonstrated that candidalysins exist in the related species C. dubliniensis and C. tropicalis, thereby identifying these molecules as a toxin family. Recent genomic analyses have highlighted the presence of a small number of candidalysin "variant" toxins, which have different amino acid sequences to those originally identified. Here, we screened genome sequences of isolates of C. albicans, C. dubliniensis, and C. tropicalis and identified candidalysin variants in all three species. When applied to epithelial cells, candidalysin variants differed in their ability to cause damage, activate intracellular signaling pathways, and induce innate immune responses, while biophysical analysis revealed differences in the ability of candidalysin variants to interact with lipid bilayers. These findings suggest that intraspecies variation in candidalysin amino acid sequence may influence fungal pathogenicity.
Assuntos
Candida albicans , Células Epiteliais , Proteínas Fúngicas , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/química , Humanos , Candida albicans/genética , Candida albicans/efeitos dos fármacos , Células Epiteliais/microbiologia , Candidíase/microbiologia , Candidíase/imunologia , Sequência de Aminoácidos , Variação Genética , Candida/genética , Candida/patogenicidade , Candida tropicalis/genética , Candida tropicalis/metabolismoRESUMO
The use of recreational waters is a widespread activity worldwide, and one of the risks associated with this practice is the exposure of bathers to microorganisms that may arise due to pollution caused by inadequate infrastructure and sanitation. In the present work, we isolated Candida spp. (n = 24) from five recreational beaches in Rio de Janeiro, Brazil, in order to evaluate their susceptibility to antifungals, the production of virulence attributes and the in vivo virulence using Tenebrio molitor larvae as a model. The ITS1-5.8S-ITS2 gene sequencing identified thirteen isolates (54.1 %) as C. tropicalis, seven (29.1 %) as C. krusei (Pichia kudriavzevii), one (4.2 %) as C. rugosa (Diutina rugosa), one (4.2 %) as C. mesorugosa (Diutina mesorugosa), one (4.2 %) as C. utilis (Cyberlindnera jadinii) and one (4.2 %) as C. parapsilosis. C. tropicalis isolates showed resistance to azoles and susceptibility to amphotericin B, flucytosine and caspofungin. C. krusei isolates were resistant to fluconazole, caspofungin and itraconazole, with 42.8 % resistance to flucytosine, besides susceptibility to voriconazole and amphotericin B. The remaining species were susceptible to all tested antifungals. All Candida isolates adhered to abiotic surfaces and formed biofilm on polystyrene, albeit to varying degrees, and produced aspartic protease and hemolytic activity, which are considered fungal virulence attributes. C. tropicalis, C. krusei and C. utilis isolates produced phytase, while the only esterase producer was C. tropicalis. Regarding resistance to osmotic stress, all isolates of C. tropicalis, C. parapsilosis and C. mesorugosa grew up to 7.5 % NaCl; the remaining isolates grew up to 1.87-3.75 % NaCl. The mortality caused by fungal challenges in T. molitor larvae was variable, with C. tropicalis, C. utilis and C. parapsilosis being more virulent than C. krusei and C. rugosa complex. Collectively, the presence of these yeasts, particularly the virulent and resistant isolates, in recreational waters can pose a significant health risk to bathers.
Assuntos
Antifúngicos , Candida , Farmacorresistência Fúngica , Brasil , Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Candida/patogenicidade , Candida/genética , Virulência , Testes de Sensibilidade Microbiana , Animais , PraiasRESUMO
A single strain of Candida anglica, isolated from cider, is available in international yeast collections. We present here seven new strains isolated from French PDO cheeses. For one of the cheese strains, we achieved a high-quality genome assembly of 13.7 Mb with eight near-complete telomere-to-telomere chromosomes. The genomes of two additional cheese strains and of the cider strain were also assembled and annotated, resulting in a core genome of 5966 coding sequences. Phylogenetic analysis showed that the seven cheese strains clustered together, away from the cider strain. Mating-type locus analysis revealed the presence of a MATa locus in the cider strain but a MATalpha locus in all cheese strains. The presence of LINE retrotransposons at identical genome position in the cheese strains, and two different karyotypic profiles resulting from chromosomal rearrangements were observed. Together, these findings are consistent with clonal propagation of the cheese strains. Phenotypic trait variations were observed within the cheese population under stress conditions whereas the cider strain was found to have a much greater capacity for growth in all conditions tested.
Assuntos
Candida , Queijo , Alimentos Fermentados , Genoma Fúngico , Filogenia , Queijo/microbiologia , Candida/genética , Candida/metabolismo , Candida/classificação , Candida/isolamento & purificação , Candida/crescimento & desenvolvimento , Alimentos Fermentados/microbiologia , Adaptação Fisiológica , Microbiologia de Alimentos , Fermentação , Genes Fúngicos Tipo AcasalamentoRESUMO
Candida auris, initially identified in 2009, has rapidly become a critical concern due to its antifungal resistance and significant mortality rates in healthcare-associated outbreaks. To date, whole-genome sequencing (WGS) has identified five unique clades of C. auris, with some strains displaying resistance to all primary antifungal drug classes. In this study, we presented the first WGS analysis of C. auris from Bangladesh, describing its origins, transmission dynamics, and antifungal susceptibility testing (AFST) profile. Ten C. auris isolates collected from hospital settings in Bangladesh were initially identified by CHROMagar Candida Plus, followed by VITEK2 system, and later sequenced using Illumina NextSeq 550 system. Reference-based phylogenetic analysis and variant calling pipelines were used to classify the isolates in different clades. All isolates aligned ~90% with the Clade I C. auris B11205 reference genome. Of the 10 isolates, 8 were clustered with Clade I isolates, highlighting a South Asian lineage prevalent in Bangladesh. Remarkably, the remaining two isolates formed a distinct cluster, exhibiting >42,447 single-nucleotide polymorphism differences compared to their closest Clade IV counterparts. This significant variation corroborates the emergence of a sixth clade (Clade VI) of C. auris in Bangladesh, with potential for international transmission. AFST results showed that 80% of the C. auris isolates were resistant to fluconazole and voriconazole, whereas Clade VI isolates were susceptible to azoles, echinocandins, and pyrimidine analogue. Genomic sequencing revealed ERG11_Y132F mutation conferring azole resistance while FCY1_S70R mutation found inconsequential in describing 5-flucytosine resistance. Our study underscores the pressing need for comprehensive genomic surveillance in Bangladesh to better understand the emergence, transmission dynamics, and resistance profiles of C. auris infections. Unveiling the discovery of a sixth clade (Clade VI) accentuates the indispensable role of advanced sequencing methodologies.IMPORTANCECandida auris is a nosocomial fungal pathogen that is commonly misidentified as other Candida species. Since its emergence in 2009, this multidrug-resistant fungus has become one of the five urgent antimicrobial threats by 2019. Whole-genome sequencing (WGS) has proven to be the most accurate identification technique of C. auris which also played a crucial role in the initial discovery of this pathogen. WGS analysis of C. auris has revealed five distinct clades where isolates of each clade differ among themselves based on pathogenicity, colonization, infection mechanism, as well as other phenotypic characteristics. In Bangladesh, C. auris was first reported in 2019 from clinical samples of a large hospital in Dhaka city. To understand the origin, transmission dynamics, and antifungal-resistance profile of C. auris isolates circulating in Bangladesh, we conducted a WGS-based surveillance study on two of the largest hospital settings in Dhaka, Bangladesh.
Assuntos
Antifúngicos , Candida auris , Candidíase , Testes de Sensibilidade Microbiana , Filogenia , Sequenciamento Completo do Genoma , Bangladesh/epidemiologia , Humanos , Antifúngicos/farmacologia , Candidíase/microbiologia , Candidíase/epidemiologia , Candida auris/genética , Candida auris/efeitos dos fármacos , Candida auris/isolamento & purificação , Farmacorresistência Fúngica , Genoma Fúngico , Polimorfismo de Nucleotídeo Único , Candida/genética , Candida/efeitos dos fármacos , Candida/classificação , Candida/isolamento & purificação , Fluconazol/farmacologia , FemininoRESUMO
Candida auris is an emerging fungal pathogen associated with multi-drug resistance rates and widespread outbreaks in hospitals and healthcare units worldwide. Sequencing studies have revealed that different clonal lineages of the fungus seem to be prevalent among distinct geographical sites. The first case of C. auris in Northern Greece was reported in Thessaloniki in October 2022, almost 2 years after the first isolation in Greece (Athens 2019). The Mycology Laboratory of the Medical School of Aristotle University of Thessaloniki stands as the reference laboratory for fungal diseases in Northern Greece and a meticulous search for the yeast, in plenty of suspicious samples, has been run since 2019 in the Lab as well as a retrospective analysis of all its yeasts' collection, back to 2008, with negative results for the presence of C. auris. Here, are presented the findings concerning the outbreak and surveillance of C. auris in Northern Greece, mainly the region of Thessaloniki and the broader area of Macedonia, from October 2022 until August 2023. The isolates from Northern Greece continue to fall in Clade I and present with an almost equal and stable sensitivity profile until now.
The study concerns the outbreak of Candida auris in Northern Greece since October 2022 and the effort for surveillance and epidemiological monitoring. All isolates continue to fall in Clade I and present with an almost equal and stable sensitivity profile till now.
Assuntos
Candida auris , Candidíase , Surtos de Doenças , Monitoramento Epidemiológico , Grécia/epidemiologia , Humanos , Candidíase/epidemiologia , Candidíase/microbiologia , Candida auris/genética , Candida auris/isolamento & purificação , Estudos Retrospectivos , Antifúngicos/farmacologia , Testes de Sensibilidade Microbiana , Masculino , Farmacorresistência Fúngica Múltipla , Candida/isolamento & purificação , Candida/classificação , Candida/genética , FemininoRESUMO
The rise of Candida auris, a multidrug-resistant fungal pathogen, across more than 40 countries, has signaled an alarming threat to global health due to its significant resistance to existing antifungal therapies. Characterized by its rapid spread and robust drug resistance, C. auris presents a critical challenge in managing infections, particularly in healthcare settings. With research on its biological traits and genetic basis of virulence and resistance still in the early stages, there is a pressing need for a concerted effort to understand and counteract this pathogen. This review synthesizes current knowledge on the epidemiology, biology, genetic manipulation, pathogenicity, diagnostics, and resistance mechanisms of C. auris, and discusses future directions in research and therapeutic development. By exploring the complexities surrounding C. auris, we aim to underscore the importance of advancing research to devise effective control and treatment strategies.
Assuntos
Antifúngicos , Candida auris , Candidíase , Farmacorresistência Fúngica Múltipla , Humanos , Antifúngicos/farmacologia , Antifúngicos/uso terapêutico , Farmacorresistência Fúngica Múltipla/genética , Candidíase/microbiologia , Candidíase/tratamento farmacológico , Candida auris/genética , Candida auris/efeitos dos fármacos , Virulência , Animais , Candida/efeitos dos fármacos , Candida/genética , Candida/patogenicidadeRESUMO
Fungal diseases are often linked to poverty, which is associated with poor hygiene and sanitation conditions that have been severely worsened by the COVID-19 pandemic. Moreover, COVID-19 patients are treated with Dexamethasone, a corticosteroid that promotes an immunosuppressive profile, making patients more susceptible to opportunistic fungal infections, such as those caused by Candida species. In this study, we analyzed the prevalence of Candida yeasts in wastewater samples collected to track viral genetic material during the COVID-19 pandemic and identified the yeasts using polyphasic taxonomy. Furthermore, we investigated the production of biofilm and hydrolytic enzymes, which are known virulence factors. Our findings revealed that all Candida species could form biofilms and exhibited moderate hydrolytic enzyme activity. We also proposed a workflow for monitoring wastewater using Colony PCR instead of conventional PCR, as this technique is fast, cost-effective, and reliable. This approach enhances the accurate taxonomic identification of yeasts in environmental samples, contributing to environmental monitoring as part of the One Health approach, which preconizes the monitoring of possible emergent pathogenic microorganisms, including fungi.
Assuntos
COVID-19 , Candida , Águas Residuárias , Fluxo de Trabalho , Águas Residuárias/microbiologia , Águas Residuárias/virologia , Brasil/epidemiologia , Candida/isolamento & purificação , Candida/genética , Candida/classificação , COVID-19/epidemiologia , COVID-19/virologia , Humanos , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Biofilmes , Monitoramento Ambiental/métodos , PandemiasRESUMO
Candida auris is a multidrug-resistant fungal pathogen with a propensity to colonize humans and persist on environmental surfaces. C. auris invasive fungal disease is being increasingly identified in acute and long-term care settings. We have developed a prototype cartridge-based C. auris surveillance assay (CaurisSurV cartridge; "research use only") that includes integrated sample processing and nucleic acid amplification to detect C. auris from surveillance skin swabs in the GeneXpert instrument and is designed for point-of-care use. The assay limit of detection (LoD) in the skin swab matrix was 10.5 and 14.8 CFU/mL for non-aggregative (AR0388) and aggregative (AR0382) strains of C. auris, respectively. All five known clades of C. auris were detected at 2-3-5× (31.5-52.5 CFU/mL) the LoD. The assay was validated using a total of 85 clinical swab samples banked at two different institutions (University of California Los Angeles, CA and Wadsworth Center, NY). Compared to culture, sensitivity was 96.8% (30/31) and 100% (10/10) in the UCLA and Wadsworth cohorts, respectively, providing a combined sensitivity of 97.5% (40/41), and compared to PCR, the combined sensitivity was 92% (46/50). Specificity was 100% with both clinical (C. auris negative matrix, N = 31) and analytical (non-C. auris strains, N = 32) samples. An additional blinded study with N = 60 samples from Wadsworth Center, NY yielded 97% (29/30) sensitivity and 100% (28/28) specificity. We have developed a completely integrated, sensitive, specific, and 58-min prototype test, which can be used for routine surveillance of C. auris and might help prevent colonization and outbreaks in acute and chronic healthcare settings. IMPORTANCE: This study has the potential to offer a better solution to healthcare providers at hospitals and long-term care facilities in their ongoing efforts for effective and timely control of Candida auris infection and hence quicker response for any potential future outbreaks.
Assuntos
Candida auris , Candidíase , Sensibilidade e Especificidade , Humanos , Candidíase/diagnóstico , Candidíase/microbiologia , Candida auris/genética , Controle de Infecções/métodos , Monitoramento Epidemiológico , Pele/microbiologia , Limite de Detecção , Sistemas Automatizados de Assistência Junto ao Leito , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Diagnóstico Molecular/métodos , Candida/isolamento & purificação , Candida/genética , Candida/classificaçãoRESUMO
Candida auris is a multidrug-resistant opportunistic fungal pathogen capable of causing serious infections and healthcare-associated outbreaks. Screening for colonization with C. auris has become routine and is recommended in many hospitals and healthcare facilities as an infection control and prevention strategy. Subsequently, and since there are currently no FDA-approved tests for this purpose, clinical microbiology laboratories have become responsible for developing protocols to detect C. auris using axial and inguinal screening swabs. In a College of American Pathologists-accredited large academic healthcare center setting, we implemented a laboratory-developed nucleic-acid amplification test for the detection of C. auris DNA. Our test validation evaluated the performance of the DiaSorin C. auris primer set used in a real-time qualitative PCR assay on the LIAISON MDX thermocycler with the Simplexa Universal Disc. The assay was highly sensitive and specific, with a limit of detection of 1-2 CFU/reaction, with no observed cross-reactivity with other Candida spp., bacterial skin commensal organisms or commonly encountered viruses. When run in parallel with a culture-based detection method, the PCR assay was 100% sensitive and specific. The assay was precise, with low variability between replicates within and between runs. Lastly, pre-analytical factors, including swab storage time, temperature, and transport media, were assessed and found to have no significant effect on the detection of C. auris at variable concentrations. Taken together, this study expands the available options for nucleic acid detection of C. auris and characterizes pre-analytical factors for implementation in both high- and low-volume laboratory settings. IMPORTANCE: This study overviews the validation and implementation of a molecular screening tool for the detection of Candida auris in a College of American Pathologist-accredited clinical laboratory. This molecular laboratory-developed test is both highly sensitive and specific and has significant health-system cost-savings associated with significantly reduced turn-around-time compared to traditional standard-of-care culture-based work up. This method and workflow is of interest to support clinical microbiology diagnostics and to help aid in hospital inpatient, and infection prevention control screening.
Assuntos
Candida auris , Candidíase , Reação em Cadeia da Polimerase em Tempo Real , Sensibilidade e Especificidade , Humanos , Reação em Cadeia da Polimerase em Tempo Real/métodos , Candidíase/diagnóstico , Candidíase/microbiologia , Candida auris/genética , Programas de Rastreamento/métodos , Pacientes Internados , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/normas , Hospitais , Candida/genética , Candida/isolamento & purificação , DNA Fúngico/genéticaRESUMO
NAD is a redox coenzyme and is the center of energy metabolism. In metabolic engineering modifications, an insufficient NAD(H) supply often limits the accumulation of target products. In this study, Candida glycerinogenes was found to be able to supply NAD(H) in large fluxes, up to 7.6 times more than Saccharomyces cerevisiae in aerobic fermentation. Aerobic fermentation in a medium without amino nitrogen sources demonstrated that C. glycerinogenes NAD synthesis was not dependent on NAD precursors in the medium. Inhibition by antisense RNA and the detection of transcript levels indicated that the main NAD supply pathway is the de novo biosynthesis pathway. It was further demonstrated that NAD(H) supply was unaffected by changes in metabolic flow through C. glycerinogenes ΔGPD aerobic fermentation (80 g/L ethanol). In conclusion, the ability of C. glycerinogenes to supply NAD(H) in large fluxes provides a new approach to solving the NAD(H) supply problem in synthetic biology.
Assuntos
Candida , Fermentação , Engenharia Metabólica , NAD , NAD/metabolismo , Candida/metabolismo , Candida/genética , Aerobiose , Engenharia Metabólica/métodos , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Etanol/metabolismo , RNA Antissenso/genética , RNA Antissenso/metabolismoRESUMO
Candida auris represents one of the most urgent threats to public health, although its ecology remains largely unknown. Because amphibians and reptiles may present favorable conditions for C. auris colonization, cloacal and blood samples (n = 68), from several snake species, were cultured and molecularly screened for C. auris using molecular amplification of glycosylphosphatidylinositol protein-encoding genes and ribosomal internal transcribed spacer sequencing. Candida auris was isolated from the cloacal swab of one Egyptian cobra (Naja haje legionis) and molecularly identified in its cloaca and blood. The isolation of C. auris from wild animals is herein reported for the first time, thus suggesting the role that these animals could play as reservoirs of this emerging pathogen. The occurrence of C. auris in blood requires further investigation, although the presence of cationic antimicrobial peptides in the plasma of reptiles could play a role in reducing the vitality of the fungus.
Candida auris represents one of the most urgent threats to public health. In this study, we reported for the first time the isolation of C. auris from snake thus suggesting the role of these animals as reservoirs of this emerging pathogen.
Assuntos
Candida , Candidíase , DNA Espaçador Ribossômico , Reservatórios de Doenças , Animais , Candida/genética , Candida/classificação , Candida/isolamento & purificação , Candida/efeitos dos fármacos , Reservatórios de Doenças/microbiologia , Candidíase/microbiologia , Candidíase/veterinária , DNA Espaçador Ribossômico/genética , DNA Espaçador Ribossômico/química , Cloaca/microbiologia , Análise de Sequência de DNA , DNA Fúngico/genética , Sangue/microbiologia , Serpentes/microbiologia , Elapidae , Egito , FilogeniaRESUMO
Candida lusitaniae fungemia is a serious infection that is rarely reported in children. The aim of this study is to describe a case series of C. lusitaniae fungemia and review previous publications regarding this rare pathogen. This is a multicenter case series of children diagnosed with C. lusitaniae fungemia. A total of 18 cases that occurred over a 15-year period in five tertiary hospitals were included. Additionally, a review of the literature regarding C. lusitaniae fungemia in children was performed. A total of 18 cases were enrolled; 11/18 (61%) were males, with a mean age of 2.3 years. All patients had severe underlying diseases and risk factors for opportunistic infection, most commonly prematurity and malignancies. More than one-third of cases occurred during the last 2 years of the study period. All isolates were susceptible to all tested antifungals. The survival rate following the acute infection was 94%, whereas the survival rate of 14 previously published cases was 71%, with the most common underlying diseases being CGD and malignancies. Candida lusitaniae fungemia is not a common event in the pediatric population, occurring exclusively in children with severe underlying diseases and significant risk factors. This cohort revealed better clinical outcomes than previously reported. All tested isolates were susceptible to all antifungal agents; variability in susceptibility as previously reported was not found in this study. The allegedly higher rate of infection in recent years is in need of further investigation in larger prospective studies in order to conclude if a real trend is at play.
Candida lusitaniae fungemia is a serious infection rarely reported in children. This cohort revealed better clinical outcomes than previously reported. All tested isolates were susceptible to all antifungal agents. The higher rate of infection in recent years is in need of further investigation.
Assuntos
Antifúngicos , Candida , Pré-Escolar , Feminino , Humanos , Masculino , Antifúngicos/uso terapêutico , Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Candida/genética , Candida/isolamento & purificação , Candida/patogenicidade , Candidemia/microbiologia , Candidemia/epidemiologia , Fungemia/microbiologia , Fungemia/mortalidade , Testes de Sensibilidade Microbiana , Estudos Retrospectivos , Fatores de Risco , Centros de Atenção Terciária/estatística & dados numéricosAssuntos
Candida auris , Candidíase , Humanos , Candidíase/microbiologia , Candida auris/genética , Candida auris/fisiologia , Candida auris/metabolismo , Candida auris/efeitos dos fármacos , Candida auris/patogenicidade , Antifúngicos/farmacologia , Farmacorresistência Fúngica , Candida/patogenicidade , Candida/fisiologia , Candida/genética , Candida/efeitos dos fármacosRESUMO
Fungal infections in neonatal intensive care units (NICU) are mainly related to Candida species, with high mortality rates. They are predominantly of endogenous origin, however, cross-infection transmitted by healthcare professionals' hands has occurred. The aim of this study was to identify Candida species isolated from the hands of healthcare professionals in a NICU before and after hygiene with 70% ethanol-based gel and evaluate virulence factors DNase, phospholipase, proteinase, hemolysin, biofilm biomass production, and metabolic activity. In vitro antifungal susceptibility testing and similarity by random amplified polymorphic DNA (RAPD) were also performed. C. parapsilosis complex was the most frequent species (57.1%); all isolates presented at least one virulence factor; three isolates (Candida parapsilosis complex) were resistant to amphotericin B, two (Candida famata [currently Debaryomyces hansenii] and Candida guilliermondii [currently Meyerozyma guilliermondii]) was resistant to micafungin, and six (Candida parapsilosis complex, Candida guilliermondii [=Meyerozyma guilliermondii], Candida viswanathi, Candida catenulata [currently Diutina catenulata] and Candida lusitaniae [currently Clavispora lusitaniae]) were resistant to fluconazole. Molecular analysis by RAPD revealed two clusters of identical strains that were in the hands of distinct professionals. Candida spp. were isolated even after hygiene with 70% ethanol-based gel, highlighting the importance of stricter basic measures for hospital infection control to prevent nosocomial transmission.
Assuntos
Antifúngicos , Candida , Infecção Hospitalar , Etanol , Mãos , Testes de Sensibilidade Microbiana , Fatores de Virulência , Humanos , Mãos/microbiologia , Antifúngicos/farmacologia , Fatores de Virulência/genética , Candida/efeitos dos fármacos , Candida/isolamento & purificação , Candida/genética , Candida/patogenicidade , Etanol/farmacologia , Infecção Hospitalar/microbiologia , Infecção Hospitalar/prevenção & controle , Candidíase/microbiologia , Pessoal de Saúde , Técnica de Amplificação ao Acaso de DNA Polimórfico , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Unidades de Terapia Intensiva Neonatal , Farmacorresistência Fúngica , Géis , Desinfecção das MãosRESUMO
Cystic fibrosis (CF) is an inherited disease that results from mutations in the gene responsible for the cystic fibrosis transmembrane conductance regulator (CFTR). The airways become clogged with thick, viscous mucus that traps microbes in respiratory tracts, facilitating colonization, inflammation and infection. CF is recognized as a biofilm-associated disease, it is commonly polymicrobial and can develop in biofilms. This review discusses Candida spp. and both Gram-positive and Gram-negative bacterial biofilms that affect the airways and cause pulmonary infections in the CF context, with a particular focus on mixed-species biofilms. In addition, the review explores the intricate interactions between fungal and bacterial species within these biofilms and elucidates the underlying molecular mechanisms that govern their dynamics. Moreover, the review addresses the multifaceted issue of antimicrobial resistance in the context of CF-associated biofilms. By synthesizing current knowledge and research findings, this review aims to provide insights into the pathogenesis of CF-related infections and identify potential therapeutic approaches to manage and combat these complex biofilm-mediated infections.