Multilocus microsatellite analysis of European and African Candida glabrata isolates

This study aimed to elucidate the genetic relatedness and epidemiology of 127 clinical and environmental Candida glabrata isolates from Europe and Africa using multilocus microsatellite analysis. Each isolate was first identified using phenotypic and molecular methods and subsequently, six unlinked microsatellite loci were analyzed using automated fluorescent genotyping. Genetic relationships were estimated using the minimum-spanning tree (MStree) method. Microsatellite analyses revealed the existence of 47 different genotypes. The fungal population showed an irregular distribution owing to the over-representation of genetically different infectious haplotypes. The most common genotype was MG-9, which was frequently found in both European and African isolates. In conclusion, the data reported here emphasize the role of specific C. glabrata genotypes in human infections for at least some decades and highlight the widespread distribution of some isolates, which seem to be more able to cause disease than others.This research was supported in part by the EU Mare Nostrum (EUMN-III Call) program of the European Union, grant agreement number 2011-4050/001-EMA2. Dr Sanae Rharmitt was the recipient of a scholarship (10 months) signed within the EUMN program for PhD students (F.S. 1.04.11.01 UORI) under the supervision of Prof Orazio Romeo.info:eu-repo/semantics/publishedVersio

Method-dependent epidemiological cutoff values (ECVs) for detection of triazole resistance in Candida and Aspergillus species for the SYO colorimetric broth and Etest agar diffusion methods

Although the Sensitrite Yeast-One (SYO) and Etest methods are widely utilized, interpretive criteria are not available for triazole susceptibility testing of Candida or Aspergillus species. We collected fluconazole, itraconazole, posaconazole and voriconazole SYO and Etest MICs from 39 laboratories representing all continents for (method-agent-dependent): 11,171 Candida albicans, 215 C. dubliniensis, 4,418 C. glabrata species complex (SC), 157 C. (Meyerozyma) guilliermondii, 676 C. krusei (Pichia kudriavzevii), 298 C (Clavispora) lusitaniae, 911 and 3,691 C. parapsilosissensu stricto (SS) and C. parapsilosisSC, respectively, 36 C. metapsilosis, 110 C. orthopsilosis, 1,854 C. tropicalis, 244 Saccharomyces cerevisiae, 1,409 Aspergillus fumigatus, 389 A. flavus, 130 A. nidulans, 233 A. niger, and 302 A. terreus complexes. SYO/Etest MICs for 282 confirmed non-WT isolates were included: ERG11 (C. albicans), ERG11 and MRR1 (C. parapsilosis), cyp51A (A. fumigatus), and CDR2, CDR1 overexpression (C. albicans and C. glabrata, respectively). Interlaboratory modal agreement was superior by SYO for yeast spp., and by the Etest for Aspergillus spp. Distributions fulfilling CLSI criteria for ECV definition were pooled and we proposed SYO ECVs for S. cerevisiae, 9 yeast and 3 Aspergillus species, and Etest ECVs for 5 yeast and 4 Aspergillus species. The posaconazole SYO ECV of 0.06 \ub5g/ml for C. albicans and the Etest itraconazole ECV of 2 \ub5g/ml for A. fumigatus were the best predictors of non-WT isolates. These findings support the need for method-dependent ECVs, as overall, the SYO appears to perform better for susceptibility testing of yeast spp. and the Etest for Aspergillus spp. Further evaluations should be conducted with more Candida mutants

Phylogenomic analysis of a 55.1 kb 19-gene dataset resolves a monophyletic Fusarium that includes the Fusarium solani Species Complex

Scientific communication is facilitated by a data-driven, scientifically sound taxonomy that considers the end-user¿s needs and established successful practice. In 2013, the Fusarium community voiced near unanimous support for a concept of Fusarium that represented a clade comprising all agriculturally and clinically important Fusarium species, including the F. solani species complex (FSSC). Subsequently, this concept was challenged in 2015 by one research group who proposed dividing the genus Fusarium into seven genera, including the FSSC described as members of the genus Neocosmospora, with subsequent justification in 2018 based on claims that the 2013 concept of Fusarium is polyphyletic. Here, we test this claim and provide a phylogeny based on exonic nucleotide sequences of 19 orthologous protein-coding genes that strongly support the monophyly of Fusarium including the FSSC. We reassert the practical and scientific argument in support of a genus Fusarium that includes the FSSC and several other basal lineages, consistent with the longstanding use of this name among plant pathologists, medical mycologists, quarantine officials, regulatory agencies, students, and researchers with a stake in its taxonomy. In recognition of this monophyly, 40 species described as genus Neocosmospora were recombined in genus Fusarium, and nine others were renamed Fusarium. Here the global Fusarium community voices strong support for the inclusion of the FSSC in Fusarium, as it remains the best scientific, nomenclatural, and practical taxonomic option availabl

Trends in candidaemia in one Italian region, Lombardia

Objectives: To perform a 1-year survey on candidaemia in one Italian region, Lombardia, to verify the epidemiological changes in comparison with the results of a previous survey conducted ten years ago (Tortorano et al. J. Hosp. Infect. 2002; 51: 297). Methods: The survey was carried out prospectively during 2009 involving 30 Microbiology Laboratories of Lombardia that notified candidaemia episodes defined by at least one blood culture positive for Candida. Results: A total of 354 episodes of candidaemia occurred in 344 patients accounting for an incidence of 1.19 per 1000 admissions (range 0.19\u20132.3) and 1.20 per 10000 patient days (range 0.2\u20132.2). These rates were higher compared to those obtained in the previous '90s survey (0.38 per 1000 admissions, range 0.03\u20131.45, and 0.44 per 10000 patient days, range 0.04\u20131.64). In the present survey, candidaemia occurred more frequently in patients aged ?80 years (15.4 vs 8.1% of the cases) and remained associated mainly, even if at a reduced rate, to surgery (45% of the cases vs 56% in the previous study) and intensive care treatments (35% vs 45%). A shift of the species causing fungaemia was demonstrated by the comparison of the two periods: while the proportion due to C. albicans decreased from 58 to 52%, an increase of C. glabrata (from 13 to 20%) and C. tropicalis (from 6 to 8%) was noted. The proportion of C. parapsilosis remained unchanged (14.5%). A decreased of the crude mortality at day 30 from 35 to 32% was observed. The highest mortality rate was detected in patients with C. tropicalis (41%) and C. albicans (33%) bloodstream infections. Conclusion: The present study revealed an increasing incidence of candidaemia, mainly in aged subjects, and an increasing proportion of isolates with decreased susceptibility to fluconazole. Powered b

Studio multicentrico prospettico sulla resistenza agli azoli in ceppi di aspergillus isolati da tamponi di sorveglianza effettuati in pazienti ematologici e/o sottoposti a trapianto di cellule staminali ematopoietiche (Studio Arte)

L\u2019aspergillosi invasiva (AI) \ue8 una complicanza severa in pazienti ematologici ed in pazienti sottoposti a trapianto di cellule staminali ematopoietiche (HSCT). In questi pazienti la diagnosi si basa principalmente su metodi indiretti (galattomannano) per la difficolt\ue0 di ottenere campioni biologici con metodiche invasive, ma questo comporta la mancanza del ceppo fungino responsabile dell\u2019infezione. La colonizzazione delle vie aeree superiori rappresenta il primo passo nello sviluppo dell'infezione polmonare ed \ue8 stato dimostrato che le colture di sorveglianza delle secrezioni nasali ed orofaringee hanno un elevato valore predittivo positivo. La multi-azolo resistenza di Aspergillus fumigatus \ue8 stata riscontrata anche in Italia sia in ceppi isolati dall'ambiente che in ceppi clinici da pazienti in terapia antifungina e na\uefve. Alla base della resistenza vi sono mutazioni puntiformi di CYP51A, gene che codifica per l'enzima bersaglio di azoli. Lo studio multicentrico prospettico ARTE ha avuto come obiettivi: 1) raccogliere, dai diversi centri partecipanti, i ceppi di Aspergillus isolati da tamponi di sorveglianza di pazienti ematologici o sottoposti a HSCT; 2) analizzare la sensibilit\ue0 in vitro agli azoli (itraconazolo, voriconazolo e posaconazolo); 3) indagare, in presenza di resistenza, il meccanismo molecolare; 4) correlare la resistenza alle variabili demografiche e comportamentali ed agli eventuali trattamenti antifungini. Ventidue centri di ematologia di 11 regioni italiane hanno aderito al progetto ARTE. Tra il 2015 ed il 2016 sono stati raccolti 292 ceppi da 228 pazienti (et\ue0 media 55 anni; maschi 60%; il 41.6% viveva in campagna). Le specie pi\uf9 frequentemente isolate sono state: A. fumigatus (46.9%), A. niger (21.9%), A. flavus (16.1%), A. terreus (4.4%), A. nidulans (1.7%). Tre isolati sono risultati resistenti agli azoli: un ceppo di A. fumigatus (itraconazolo MIC 4 mg/L e posaconazolo MIC 0.5 mg/L), un ceppo di A. niger (itraconazolo MIC >16 mg/L) ed uno di A. lentulus (itraconazole MIC >16 mg/L). A. lentulus e A. niger sono noti in letteratura per avere valori di sensibilit\ue0 elevati agli azoli (in particolare ad itraconazolo). Il ceppo di A. fumigatus resistente \ue8 stato isolato da un paziente affetto da mieloma che aveva subito un autotrapianto di cellule staminali ematopoietiche, con antigene galattomannano positivo su lavaggio broncoalveolare con un quadro radiologico compatibile e che, inoltre, viveva in campagna ed era un fumatore. Tale ceppo aveva la mutazione TR34/L98H nel gene CYP51A e, dall\u2019analisi dei microsatelliti (MLST), \ue8 risultato differente dai ceppi ambientali ed altri ceppi clinici della stessa area geografica con i quali \ue8 stato confrontato. Il presente studio ha fatto emergere alcune problematiche gi\ue0 riscontrate in questi tipi di sorveglianze. La mancanza di protocolli comuni per le colture di sorveglianza nelle cliniche ematologiche e nei laboratori ha portato ad avere un numero molto diverso di campioni tra i centri partecipanti. Inoltre la difficolt\ue0 di ottenere, nei pazienti ematologici, le colture fungine, che rappresentano il principale strumento diagnostico, porta sicuramente ad una sottostima sia della colonizzazione/infezione sia della resistenza. Dato che pazienti con AI causata da un ceppo resistente hanno un tasso di mortalit\ue0 dell'88% vs il 30-50% dei pazienti con ceppo sensibile, il poter isolare precocemente il fungo responsabile potrebbe essere determinante per l\u2019outcome del paziente

Looking for Candida nivariensis and C. bracarensis among a large Italian collection of C. glabrata isolates: results of the FIMUA working group

Two recently described pathogenic Candida species, C. nivariensis and C. bracarensis, share many phenotypic characteristics with C. glabrata and are easily misidentified as such. The aim of this study was to determine the occurrence of these cryptic species in Italy. One thousand yeast isolates collected in 14 Italian regions and identified as C. glabrata by phenotypic and biochemical methods were included in this study: 928 were screened on CHROMagar and 72 were analysed by a multiplex PCR. None of these cryptic species was identified despite the nationwide distribution and the variety of biological origin of the isolates

European Confederation of Medical Mycology (ECMM) epidemiological survey on invasive infections due to Fusarium species in Europe

In order to better understand the epidemiology of fusariosis in Europe, a survey collecting information on the clinical characteristics of the patients infected by Fusarium as well as on the infecting isolates was launched. A total of 76 cases of invasive fusariosis occurring from January 2007 to June 2012 were collected and Fusarium isolates were identified by sequencing the translation elongation factor 1α (TEF) gene. Also, antifungal susceptibility was tested by broth microdilution according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the Etest. Disseminated disease was considered proven in 46 cases and probable in 17 cases. Localised infection was seen in 13 cases. Gibberella fujikuroi species complex (SC), including Fusarium verticillioides and F. proliferatum, and F. solani SC were the most frequent aetiology of disseminated and localised infections, respectively. The crude mortality rate was 46 %, the highest associated with F. solani SC (67 %) and F. proliferatum (62.5 %). A wide range of antifungal susceptibilities was observed. Amphotericin B was the most potent antifungal in vitro, and itraconazole the least effective. The azoles exhibited lower minimum inhibitory concentrations (MICs) against F. verticillioides strains, with posaconazole having a slightly better performance, while F. solani SC isolates were resistant to all three azoles tested. The essential agreement between the Etest and the EUCAST method was 100 % for itraconazole and voriconazole, and 96 % for amphotericin B and posaconazole. In conclusion, we confirm that fusariosis is a rare but severe event in Europe, that G. fujikuroi SC is the predominant cause of deep infections and that different species have different antifungal in vitro susceptibility patterns. © 2014 Springer-Verlag

European Confederation of Medical Mycology (ECMM) epidemiological survey on invasive infections due to Fusarium species in Europe

In order to better understand the epidemiology of fusariosis in Europe, a survey collecting information on the clinical characteristics of the patients infected by Fusarium as well as on the infecting isolates was launched. A total of 76 cases of invasive fusariosis occurring from January 2007 to June 2012 were collected and Fusarium isolates were identified by sequencing the translation elongation factor 1\u3b1 (TEF) gene. Also, antifungal susceptibility was tested by broth microdilution according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the Etest. Disseminated disease was considered proven in 46 cases and probable in 17 cases. Localised infection was seen in 13 cases. Gibberella fujikuroi species complex (SC), including Fusarium verticillioides and F. proliferatum, and F. solani SC were the most frequent aetiology of disseminated and localised infections, respectively. The crude mortality rate was 46 %, the highest associated with F. solani SC (67 %) and F. proliferatum (62.5 %). A wide range of antifungal susceptibilities was observed. Amphotericin B was the most potent antifungal in vitro, and itraconazole the least effective. The azoles exhibited lower minimum inhibitory concentrations (MICs) against F. verticillioides strains, with posaconazole having a slightly better performance, while F. solani SC isolates were resistant to all three azoles tested. The essential agreement between the Etest and the EUCAST method was 100 % for itraconazole and voriconazole, and 96 % for amphotericin B and posaconazole. In conclusion, we confirm that fusariosis is a rare but severe event in Europe, that G. fujikuroi SC is the predominant cause of deep infections and that different species have different antifungal in vitro susceptibility patterns

Prospective multicentre study on azole resistance in Aspergillus isolates from surveillance cultures in haematological patients in Italy

Objectives: This study was conducted to assess the prevalence of azole resistance in Aspergillus isolates from patients with haematological malignancies or who were undergoing haematopoietic stem cell transplantation and to identify the molecular mechanism of resistance. Methods: In this 28-month prospective study involving 18 Italian centres, Aspergillus isolates from surveillance cultures were collected and screened for azole resistance, and mutations in the cyp51A gene were identified. Resistant isolates were genotyped by microsatellite analysis, and the allelic profiles were compared with those of resistant environmental and clinical isolates from the same geographical area that had been previously genotyped. Results: There were 292 Aspergillus isolates collected from 228 patients. The isolates belonged mainly to the section Fumigati (45.9%), Nigri (20.9%), Flavi (16.8%) and Terrei (4.8%). Three isolates showed itraconazole resistance: Aspergillus fumigatus sensu stricto, Aspergillus lentulus (section Fumigati) and Aspergillus awamori (section Nigri). The itraconazole resistance rates were 1% and 1.48% considering all Aspergillus spp. isolates and the Aspergillus section Fumigati, respectively. The prevalence of azole resistance among all the patients was 1.3%. Among patients harbouring A. fumigatus sensu stricto isolates, the resistance rate was 0.79%. The A. fumigatus isolate, with the TR34/L98H mutation, was genotypically distant from the environmental and clinical strains previously genotyped. Conclusions: In this study, the Aspergillus azole resistance rate was 1% (3/292). In addition to A. fumigatus sensu stricto, A. lentulus and A. awamori azole-resistant isolates were identified. Therefore, it is important have a correct identification at the species level to address a rapid therapy better, quickly understand the shift towards cryptic species and have an updated knowledge of the local epidemiology

Prospective multicentre study on azole resistance in Aspergillus isolates from surveillance cultures in haematological patients in Italy

Objectives: This study was conducted to assess the prevalence of azole resistance in Aspergillus isolates from patients with haematological malignancies or who were undergoing haematopoietic stem cell transplantation and to identify the molecular mechanism of resistance. Methods: In this 28-month prospective study involving 18 Italian centres, Aspergillus isolates from surveillance cultures were collected and screened for azole resistance, and mutations in the cyp51A gene were identified. Resistant isolates were genotyped by microsatellite analysis, and the allelic profiles were compared with those of resistant environmental and clinical isolates from the same geographical area that had been previously genotyped. Results: There were 292 Aspergillus isolates collected from 228 patients. The isolates belonged mainly to the section Fumigati (45.9%), Nigri (20.9%), Flavi (16.8%) and Terrei (4.8%). Three isolates showed itraconazole resistance: Aspergillus fumigatus sensu stricto, Aspergillus lentulus (section Fumigati) and Aspergillus awamori (section Nigri). The itraconazole resistance rates were 1% and 1.48% considering all Aspergillus spp. isolates and the Aspergillus section Fumigati, respectively. The prevalence of azole resistance among all the patients was 1.3%. Among patients harbouring A. fumigatus sensu stricto isolates, the resistance rate was 0.79%. The A. fumigatus isolate, with the TR34/L98H mutation, was genotypically distant from the environmental and clinical strains previously genotyped. Conclusions: In this study, the Aspergillus azole resistance rate was 1% (3/292). In addition to A. fumigatus sensu stricto, A. lentulus and A. awamori azole-resistant isolates were identified. Therefore, it is important have a correct identification at the species level to address a rapid therapy better, quickly understand the shift towards cryptic species and have an updated knowledge of the local epidemiology