Population-Based Survey of Filamentous Fungi and Antifungal Resistance in Spain (FILPOP Study)

ABSTRACTA population-based survey was conducted to investigate the epidemiology of and antifungal resistance in Spanish clinical strains of filamentous fungi isolated from deep tissue samples, blood cultures, and respiratory samples. The study was conducted in two different periods (October 2010 and May 2011) to analyze seasonal variations. A total of 325 strains were isolated in 29 different hospitals. The average prevalence was 0.0016/1,000 inhabitants. Strains were identified by sequencing of DNA targets and susceptibility testing by the European Committee for Antimicrobial Susceptibility Testing reference procedure. The most frequently isolated genus wasAspergillus, accounting for 86.3% of the isolates, followed byScedosporiumat 4.7%; the order Mucorales at 2.5%;Penicilliumat 2.2%, andFusariumat 1.2%. The most frequent species wasAspergillus fumigatus(48.5%), followed byA. flavus(8.4%),A. terreus(8.1%),A. tubingensis(6.8%), andA. niger(6.5%). Cryptic/siblingAspergillusspecies accounted for 12% of the cases. Resistance to amphotericin B was found in 10.8% of the isolates tested, while extended-spectrum triazole resistance ranged from 10 to 12.7%, depending on the azole tested. Antifungal resistance was more common among emerging species such as those ofScedosporiumand Mucorales and also among cryptic species ofAspergillus, with 40% of these isolates showing resistance to all of the antifungal compounds tested. CrypticAspergillusspecies seem to be underestimated, and their correct classification could be clinically relevant. The performance of antifungal susceptibility testing of the strains implicated in deep infections and multicentric studies is recommended to evaluate the incidence of these cryptic species in other geographic areas.

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Antifungal Resistance to Fluconazole and Echinocandins Is Not Emerging in Yeast Isolates Causing Fungemia in a Spanish Tertiary Care Center

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02670-14 ◽

2014 ◽

Vol 58 (8) ◽

pp. 4565-4572 ◽

Cited By ~ 34

Author(s):

Laura Judith Marcos-Zambrano ◽

Pilar Escribano ◽

Carlos Sánchez ◽

Patricia Muñoz ◽

Emilio Bouza ◽

...

Keyword(s):

Susceptibility Testing ◽

Molecular Level ◽

Care Center ◽

Antifungal Susceptibility ◽

Tertiary Care ◽

Candida Guilliermondii ◽

Antifungal Resistance ◽

Its2 Region ◽

Content Type

ABSTRACTAccurate knowledge of fungemia epidemiology requires identification of strains to the molecular level. Various studies have shown that the rate of resistance to fluconazole ranges from 2.5% to 9% inCandidaspp. isolated from blood samples. However, trends in antifungal resistance have received little attention and have been studied only using CLSI M27-A3 methodology. We assessed the fungemia epidemiology in a large tertiary care institution in Madrid, Spain, by identifying isolates to the molecular level and performing antifungal susceptibility testing according to the updated breakpoints of European Committee for Antimicrobial Susceptibility Testing (EUCAST) definitive document (EDef) 7.2. We studied 613 isolates causing 598 episodes of fungemia in 544 patients admitted to our hospital (January 2007 to December 2013). Strains were identified after amplification and sequencing of the ITS1-5.8S-ITS2 region and further tested forin vitrosusceptibility to amphotericin B, fluconazole, posaconazole, voriconazole, micafungin, and anidulafungin. Resistance was defined using EUCAST species-specific breakpoints, and epidemiological cutoff values (ECOFFs) were applied as tentative breakpoints. Most episodes were caused byCandida albicans(46%),Candida parapsilosis(28.7%),Candida glabrata(9.8%), andCandida tropicalis(8%). Molecular identification enabled us to better detect cryptic species ofCandida guilliermondiiandC. parapsilosiscomplexes and episodes of polyfungal fungemia. The overall percentage of fluconazole-resistant isolates was 5%, although it was higher inC. glabrata(8.6%) and non-Candidayeast isolates (47.4%). The rate of resistance to echinocandins was 4.4% and was mainly due to the presence of intrinsically resistant non-Candidaspecies. Resistance mainly affected non-Candidayeasts. The rate of resistance to fluconazole and echinocandins did not change considerably during the study period.

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Population-Based Survey of Filamentous Fungi and Antifungal Resistance in Spain (FILPOP Study)

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01287-13 ◽

2013 ◽

Vol 57 (9) ◽

pp. 4604-4604 ◽

Cited By ~ 7

Author(s):

A. Alastruey-Izquierdo ◽

E. Mellado ◽

T. Peláez ◽

J. Pemán ◽

S. Zapico ◽

...

Keyword(s):

Filamentous Fungi ◽

Population Based ◽

Antifungal Resistance ◽

Population Based Survey

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Molecular Identification and Susceptibility Testing of Molds Isolated in a Prospective Surveillance of Triazole Resistance in Spain (FILPOP2 Study)

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00358-18 ◽

2018 ◽

Vol 62 (9) ◽

Cited By ~ 16

Author(s):

Ana Alastruey-Izquierdo ◽

Laura Alcazar-Fuoli ◽

Olga Rivero-Menéndez ◽

Josefina Ayats ◽

Carmen Castro ◽

...

Keyword(s):

Molecular Identification ◽

Susceptibility Testing ◽

Antifungal Susceptibility ◽

Sensu Stricto ◽

Antifungal Resistance ◽

Azole Resistance ◽

Secondary Resistance ◽

Content Type ◽

Resistant Species ◽

Tertiary Hospitals

ABSTRACT Antifungal resistance is increasing by the emergence of intrinsically resistant species and by the development of secondary resistance in susceptible species. A previous study performed in Spain revealed levels of azole resistance in molds of between 10 and 12.7%, but secondary resistance in Aspergillus fumigatus was not detected. We used itraconazole (ITZ)-supplemented medium to select resistant strains. A total of 500 plates supplemented with 2 mg/liter of ITZ were sent to 10 Spanish tertiary hospitals, and molecular identification and antifungal susceptibility testing were performed. In addition, the cyp51A gene in those A. fumigatus strains showing azole resistance was sequenced. A total of 493 isolates were included in the study. Sixteen strains were isolated from patients with an infection classified as proven, 104 were isolated from patients with an infection classified as probable, and 373 were isolated from patients with an infection classified as colonization. Aspergillus was the most frequent genus isolated, at 80.3%, followed by Scedosporium-Lomentospora (7.9%), Penicillium-Talaromyces (4.5%), Fusarium (2.6%), and the order Mucorales (1%). Antifungal resistance was detected in Scedosporium-Lomentospora species, Fusarium, Talaromyces, and Mucorales. Three strains of A. fumigatus sensu stricto were resistant to azoles; two of them harbored the TR34+L98H mechanism of resistance, and the other one had no mutations in cyp51A. The level of azole resistance in A. fumigatus remains low, but cryptic species represent over 10% of the isolates and have a broader but overall higher range of antifungal resistance.

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Trends in Antifungal Drug Susceptibility of Cryptococcus neoformans Isolates Obtained through Population-Based Surveillance in South Africa in 2002-2003 and 2007-2008

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00048-11 ◽

2011 ◽

Vol 55 (6) ◽

pp. 2606-2611 ◽

Cited By ~ 43

Author(s):

Nelesh P. Govender ◽

Jaymati Patel ◽

Marelize van Wyk ◽

Tom M. Chiller ◽

Shawn R. Lockhart ◽

...

Keyword(s):

South Africa ◽

Cryptococcus Neoformans ◽

Amphotericin B ◽

Fungal Infections ◽

Antifungal Susceptibility ◽

Drug Susceptibility ◽

Population Based ◽

Content Type ◽

Microdilution Method ◽

Broth Microdilution Method

ABSTRACTCryptococcus neoformansis the most common cause of meningitis among adult South Africans with HIV infection/AIDS. Widespread use of fluconazole for treatment of cryptococcal meningitis and other HIV-associated opportunistic fungal infections in South Africa may lead to the emergence of isolates with reduced fluconazole susceptibility. MIC testing using a reference broth microdilution method was used to determine if isolates with reduced susceptibility to fluconazole or amphotericin B had emerged among cases of incident disease. Incident isolates were tested from two surveillance periods (2002-2003 and 2007-2008) when population-based surveillance was conducted in Gauteng Province, South Africa. These isolates were also tested for susceptibility to flucytosine, itraconazole, voriconazole, and posaconazole. Serially collected isolate pairs from cases at several large South African hospitals were also tested for susceptibility to fluconazole. Of the 487 incident isolates tested, only 3 (0.6%) demonstrated a fluconazole MIC of ≥16 μg/ml; all of these isolates were from 2002-2003. All incident isolates were inhibited by very low concentrations of amphotericin B and exhibited very low MICs to voriconazole and posaconazole. Of 67 cases with serially collected isolate pairs, only 1 case was detected where the isolate collected more than 30 days later had a fluconazole MIC value significantly higher than the MIC of the corresponding incident isolate. Although routine antifungal susceptibility testing of incident isolates is not currently recommended in clinical settings, it is still clearly important for public health to periodically monitor for the emergence of resistance.

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Susceptibility of Filamentous Fungi to Voriconazole in Malaysia Tested by Sensititre YeastOne and CLSI Microdilution Methods

10.21203/rs.3.rs-199013/v1 ◽

2021 ◽

Author(s):

Xue Ting Tan ◽

Stephanie Jane Ginsapu ◽

Fairuz binti Amran ◽

Salina binti Mohamed Sukur ◽

Surianti binti Shukor

Keyword(s):

Filamentous Fungi ◽

Susceptibility Testing ◽

Rhizopus Oryzae ◽

Antifungal Susceptibility ◽

Sporothrix Schenckii ◽

Rank Test ◽

Broth Microdilution ◽

Syncephalastrum Racemosum ◽

Microdilution Method ◽

Broth Microdilution Method

Abstract Background: Voriconazole is a trizaole antifungal to treat fungal infection. In this study, the susceptibility pattern of voriconazole against filamentous fungi was studied using Sensititre® YeastOne and Clinical & Laboratory Standards Institute (CLSI) M38 broth microdilution method. Methods: The suspected cultures of Aspergillus niger, A. flavus, A. fumigatus, A. versicolor, A. sydowii, A. calidoutus, A. creber, A. ochraceopetaliformis, A. tamarii, Fusarium solani, F. longipes, F. falciferus, F. keratoplasticum, Rhizopus oryzae, R. delemar, R. arrhizus, Mucor sp., Poitrasia circinans, Syncephalastrum racemosum and Sporothrix schenckii were received from hospitals. Their identification had been confirmed in our lab and susceptibility tests were performed using Sensititre® YeastOne and CLSI M38 broth microdilution method. The significant differences between two methods were calculated using Wilcoxon Sign Rank test.Results: Mean of the minimum inhibitory concentrations (MIC) for Aspergillus spp. and Fusarium were within 0.25 μg/mL-2.00 μg/mL by two methods except A. calidoutus, F. solani and F. keratoplasticum. Moreover, mean of MIC for S. schenkii were around 3.00 μg/mL by two methods. In contrast, mean of MIC for Rhizopus spp., Mucor sp., P. circinans and S. racemosum were ≥6.00 μg/mL by two methods. Generally, the MIC obtained by Sensititre YeastOne was one two-fold increase or decrease compared with the results obtained by CLSI method. The overall agreement between Sensititre YeastOne and CLSI methods to test susceptibility testing of voricaonazole was more than 70% except A. sydowii. The significant differences between two methods were significant when tested on A. niger, A. flavus, A. fumigatus, A. versicolor, A. sydowii, F. solani and S. schenkii. Conclusions: In conclusion, Sensititre YeastOne method appears to be an alternative procedure for antifungal susceptibility testing for some Malaysian moulds.

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Molecular Identification and Antifungal Susceptibility of Yeast Isolates Causing Fungemia Collected in a Population-Based Study in Spain in 2010 and 2011

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02155-13 ◽

2013 ◽

Vol 58 (3) ◽

pp. 1529-1537 ◽

Cited By ~ 82

Author(s):

Jesús Guinea ◽

Óscar Zaragoza ◽

Pilar Escribano ◽

Estrella Martín-Mazuelos ◽

Javier Pemán ◽

...

Keyword(s):

Hot Spot ◽

Antifungal Susceptibility ◽

Point Mutations ◽

Population Based ◽

Candida Guilliermondii ◽

Population Based Study ◽

Content Type ◽

Candida Lusitaniae ◽

Species Specific

ABSTRACTWe report the molecular identifications and antifungal susceptibilities of the isolates causing fungemia collected in the CANDIPOP population-based study conducted in 29 Spanish hospitals. A total of 781 isolates (from 767 patients, 14 of them having mixed fungemia) were collected. The species found most frequently wereCandida albicans(44.6%),Candida parapsilosis(24.5%),Candida glabrata(13.2%),Candida tropicalis(7.6%),Candida krusei(1.9%),Candida guilliermondii(1.7%), andCandida lusitaniae(1.3%). OtherCandidaand non-Candidaspecies accounted for approximately 5% of the isolates. The presence of cryptic species was low. Compared to findings of previous studies conducted in Spain, the frequency ofC. glabratahas increased. Antifungal susceptibility testing was performed by using EUCAST and CLSI M27-A3 reference procedures; the two methods were comparable. The rate of fluconazole-susceptible isolates was 80%, which appears to be a decrease compared to findings of previous studies, explained mainly by the higher frequency ofC. glabrata. Using the species-specific breakpoints and epidemiological cutoff values, the rate of voriconazole and posaconazolein vitroresistance was low (<2%). In the case ofC. tropicalis, using the EUCAST procedure, the rate of azole resistance was around 20%. There was a correlation between the previous use of azoles and the presence of fluconazole-resistant isolates. Resistance to echinocandins was very rare (2%), and resistance to amphotericin B also was very uncommon. The sequencing of the hot spot (HS) regions fromFKS1orFKS2genes in echinocandin-resistant isolates revealed previously described point mutations. The decrease in the susceptibility to fluconazole in Spanish isolates should be closely monitored in future studies.

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Antifungal Susceptibility Testing of Malassezia spp. with an Optimized Colorimetric Broth Microdilution Method

Journal of Clinical Microbiology ◽

10.1128/jcm.00338-17 ◽

2017 ◽

Vol 55 (6) ◽

pp. 1883-1893 ◽

Cited By ~ 22

Author(s):

Cheryl Leong ◽

Antonino Buttafuoco ◽

Martin Glatz ◽

Philipp P. Bosshard

Keyword(s):

Susceptibility Testing ◽

Skin Diseases ◽

Antifungal Susceptibility ◽

Drug Susceptibility ◽

Drug Susceptibility Testing ◽

Broth Microdilution ◽

Content Type ◽

Microdilution Method ◽

Broth Microdilution Method ◽

Colorimetric Indicator

ABSTRACTMalasseziais a genus of lipid-dependent yeasts. It is associated with common skin diseases such as pityriasis versicolor and atopic dermatitis and can cause systemic infections in immunocompromised individuals. Owing to the slow growth and lipid requirements of these fastidious yeasts, convenient and reliable antifungal drug susceptibility testing assays forMalasseziaspp. are not widely available. Therefore, we optimized a broth microdilution assay for the testing ofMalasseziathat is based on the CLSI and EUCAST assays forCandidaand other yeasts. The addition of ingredients such as lipids and esculin provided a broth medium formulation that enabled the growth of allMalasseziaspp. and could be read, with the colorimetric indicator resazurin, by visual and fluorescence readings. We tested the susceptibility of 52 strains of 13Malasseziaspecies to 11 commonly used antifungals. MIC values determined by visual readings were in good agreement with MIC values determined by fluorescence readings. The lowest MICs were found for the azoles itraconazole, posaconazole, and voriconazole, with MIC90values of 0.03 to 1.0 μg/ml, 0.06 to 0.5 μg/ml, and 0.03 to 2.0 μg/ml, respectively. AllMalasseziaspp. were resistant to echinocandins and griseofulvin. SomeMalasseziaspp. also showed high MIC values for ketoconazole, which is the most widely recommended topical antifungal to treatMalasseziaskin infections. In summary, our assay enables the fast and reliable susceptibility testing ofMalasseziaspp. with a large panel of different antifungals.

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Evaluation of ID Fungi Plates Medium for Identification of Molds by MALDI Biotyper

Journal of Clinical Microbiology ◽

10.1128/jcm.01687-19 ◽

2020 ◽

Vol 58 (5) ◽

Cited By ~ 2

Author(s):

Marie Gladys Robert ◽

Charlotte Romero ◽

Céline Dard ◽

Cécile Garnaud ◽

Odile Cognet ◽

...

Keyword(s):

Filamentous Fungi ◽

Susceptibility Testing ◽

Antifungal Susceptibility ◽

Antifungal Susceptibility Testing ◽

Maldi Tof Ms ◽

Extraction Protocol ◽

Maldi Tof ◽

Morphological Aspects ◽

Routine Identification ◽

Tof Ms

ABSTRACT MALDI-TOF mass spectrometry (MS) identification of pathogenic filamentous fungi is often impaired by difficulties in harvesting hyphae embedded in the medium and long extraction protocols. The ID Fungi Plate (IDFP) is a novel culture method developed to address such difficulties and improve the identification of filamentous fungi by MALDI-TOF MS. We cultured 64 strains and 11 clinical samples on IDFP, Sabouraud agar-chloramphenicol (SAB), and ChromID Candida agar (CAN2). We then compared the three media for growth, ease of harvest, amount of material picked, and MALDI-TOF identification scores after either rapid direct transfer (DT) or a long ethanol-acetonitrile (EA) extraction protocol. Antifungal susceptibility testing and microscopic morphology after subculture on SAB and IDFP were also compared for ten molds. Growth rates and morphological aspects were similar for the three media. With IDFP, harvesting of fungal material for the extraction procedure was rapid and easy in 92.4% of cases, whereas it was tedious on SAB or CAN2 in 65.2% and 80.3% of cases, respectively. The proportion of scores above 1.7 (defined as acceptable identification) were comparable for both extraction protocols using IDFP (P = 0.256). Moreover, rates of acceptable identification after DT performed on IDFP (93.9%) were significantly higher than those obtained after EA extraction with SAB (69.7%) or CAN2 (71.2%) (P = <0.001 and P = 0.001, respectively). Morphological aspects and antifungal susceptibility testing were similar between IDFP and SAB. IDFP is a culture plate that facilitates and improves the identification of filamentous fungi, allowing accurate routine identification of molds with MALDI-TOF-MS using a rapid-extraction protocol.

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Molecular Identification, Antifungal Susceptibility Testing, and Mechanisms of Azole Resistance in Aspergillus Species Received within a Surveillance Program on Antifungal Resistance in Spain

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00865-19 ◽

2019 ◽

Vol 63 (9) ◽

Cited By ~ 10

Author(s):

Olga Rivero-Menendez ◽

Juan Carlos Soto-Debran ◽

Narda Medina ◽

Jose Lucio ◽

Emilia Mellado ◽

...

Keyword(s):

Aspergillus Terreus ◽

Fungal Infections ◽

Novel Mutation ◽

Antifungal Susceptibility ◽

Resistance Mechanism ◽

Sensu Stricto ◽

Antifungal Resistance ◽

Surveillance Program ◽

Azole Resistance ◽

Content Type

ABSTRACT Antifungal resistance is one of the major causes of the increasing mortality rates for fungal infections, especially for those caused by Aspergillus spp. A surveillance program was established in 2014 in the Spanish National Center for Microbiology for tracking resistance in the most prevalent Aspergillus species. A total of 273 samples were included in the study and were initially classified as susceptible or resistant according to EUCAST breakpoints. Several Aspergillus cryptic species were found within the molecularly identified isolates. Cyp51 mutations were characterized for Aspergillus fumigatus, Aspergillus terreus, and Aspergillus flavus sensu stricto strains that were classified as resistant. Three A. fumigatus sensu stricto strains carried the TR34/L98H resistance mechanism, while two harbored G54R substitution and one harbored the TR46/Y121F/T289A mechanism. Seventeen strains had no mutations in cyp51A, with ten of them resistant only to isavuconazole. Three A. terreus sensu stricto strains harbored D344N substitution in cyp51A, one of them combined with M217I, and another carried an A249G novel mutation. Itraconazole-resistant A. flavus sensu stricto strains harbored P220L and H349R alterations in cyp51A and cyp51C, respectively, that need further investigation on their implication in azole resistance.

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Epidemiology and Antifungal Susceptibility Profile of Aspergillus Species: Comparison between Environmental and Clinical Isolates from Patients with Hematologic Malignancies

Journal of Clinical Microbiology ◽

10.1128/jcm.02023-18 ◽

2019 ◽

Vol 57 (7) ◽

Cited By ~ 6

Author(s):

Sung-Yeon Cho ◽

Dong-Gun Lee ◽

Won-Bok Kim ◽

Hye-Sun Chun ◽

Chulmin Park ◽

...

Keyword(s):

Invasive Aspergillosis ◽

Cryptic Species ◽

Hematologic Malignancy ◽

Antifungal Susceptibility ◽

Hematologic Malignancies ◽

Clinical Isolates ◽

Environmental Isolates ◽

Content Type ◽

Culture Positive ◽

Global Data

ABSTRACT Global data on the epidemiology and susceptibility of Aspergillus are crucial in the management of invasive aspergillosis. Here, we aimed to determine the characteristics of clinical and environmental Aspergillus isolates, focusing mainly on hematologic malignancy patients. We prospectively collected all consecutive cases and clinical isolates of culture-positive proven/probable invasive aspergillosis patients from January 2016 to April 2018 and sampled the air inside and outside the hospital. Cryptic species-level identification of Aspergillus, antifungal susceptibilities, and cyp51 gene sequencing were performed, and clinical data were analyzed. This study was conducted as part of the Catholic Hematology Hospital Fungi Epidemiology (CAFÉ) study. A total of 207 proven/probable invasive aspergillosis and 102 clinical and 129 environmental Aspergillus isolates were included in this analysis. The incidence of proven/probable invasive aspergillosis was 1.3 cases/1,000 patient-days during the study period. Cryptic Aspergillus species accounted for 33.8%, with no differences in proportions between the clinical and environmental isolates. Section Nigri presented a high proportion (70.5%) of cryptic species, mainly from A. tubingensis and A. awamori: the former being dominant in environmental samples, and the latter being more common in clinical isolates (P < 0.001). Of 91 A. fumigatus isolates, azole-resistant A. fumigatus was found in 5.3% of all A. fumigatus isolates. Three isolates presented the TR34/L98H mutation of the cyp51A gene. Patients with invasive aspergillosis caused by azole-resistant A. fumigatus showed 100% all-cause mortality at 100 days. This study demonstrates the significant portion of cryptic Aspergillus species and clinical implications of azole resistance and underscores the comparison between clinical and environmental isolates.

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