Publications by authors named "Patrice Le Pape"

98 Publications

Genotypic, proteomic, and phenotypic approaches to decipher the response to caspofungin and calcineurin inhibitors in clinical isolates of echinocandin-resistant Candida glabrata.

J Antimicrob Chemother 2021 Dec 10. Epub 2021 Dec 10.

Unidad de Proteómica y Micosis Humanas, Grupo de Enfermedades Infecciosas Departamento de Microbiología, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá, D.C., Colombia.

Background: Echinocandin resistance represents a great concern, as these drugs are recommended as first-line therapy for invasive candidiasis. Echinocandin resistance is conferred by mutations in FKS genes. Nevertheless, pathways are crucial for enabling tolerance, evolution, and maintenance of resistance. Therefore, understanding the biological processes and proteins involved in the response to caspofungin may provide clues indicating new therapeutic targets.

Objectives: We determined the resistance mechanism and assessed the proteome response to caspofungin exposure. We then evaluated the phenotypic impact of calcineurin inhibition by FK506 and cephalosporine A (CsA) on caspofungin-resistant Candida glabrata isolates.

Methods: Twenty-five genes associated with caspofungin resistance were analysed by NGS, followed by studies of the quantitative proteomic response to caspofungin exposure. Then, susceptibility testing of caspofungin in presence of FK506 and CsA was performed. The effects of calcineurin inhibitor/caspofungin combinations on heat stress (40°C), oxidative stress (0.2 and 0.4 mM menadione) and on biofilm formation (polyurethane catheter) were analysed. Finally, a Galleria mellonella model using blastospores (1 × 109 cfu/mL) was developed to evaluate the impact of the combinations on larval survival.

Results: F659-del was found in the FKS2 gene of resistant strains. Proteomics data showed some up-regulated proteins are involved in cell-wall biosynthesis, response to stress and pathogenesis, some of them being members of calmodulin-calcineurin pathway. Therefore, the impact of calmodulin inhibition was explored. Calmodulin inhibition restored caspofungin susceptibility, decreased capacity to respond to stress conditions, and reduced biofilm formation and in vivo pathogenicity.

Conclusions: Our findings confirm that calmodulin-calcineurin-Crz1 could provide a relevant target in life-threatening invasive candidiasis.
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http://dx.doi.org/10.1093/jac/dkab454DOI Listing
December 2021

Fungal infections in mechanically ventilated patients with COVID-19 during the first wave: the French multicentre MYCOVID study.

Lancet Respir Med 2021 Nov 26. Epub 2021 Nov 26.

CHU Necker Enfants Malades-APHP, Paris, France.

Background: Patients with severe COVID-19 have emerged as a population at high risk of invasive fungal infections (IFIs). However, to our knowledge, the prevalence of IFIs has not yet been assessed in large populations of mechanically ventilated patients. We aimed to identify the prevalence, risk factors, and mortality associated with IFIs in mechanically ventilated patients with COVID-19 under intensive care.

Methods: We performed a national, multicentre, observational cohort study in 18 French intensive care units (ICUs). We retrospectively and prospectively enrolled adult patients (aged ≥18 years) with RT-PCR-confirmed SARS-CoV-2 infection and requiring mechanical ventilation for acute respiratory distress syndrome, with all demographic and clinical and biological follow-up data anonymised and collected from electronic case report forms. Patients were systematically screened for respiratory fungal microorganisms once or twice a week during the period of mechanical ventilation up to ICU discharge. The primary outcome was the prevalence of IFIs in all eligible participants with a minimum of three microbiological samples screened during ICU admission, with proven or probable (pr/pb) COVID-19-associated pulmonary aspergillosis (CAPA) classified according to the recent ECMM/ISHAM definitions. Secondary outcomes were risk factors of pr/pb CAPA, ICU mortality between the pr/pb CAPA and non-pr/pb CAPA groups, and associations of pr/pb CAPA and related variables with ICU mortality, identified by regression models. The MYCOVID study is registered with ClinicalTrials.gov, NCT04368221.

Findings: Between Feb 29 and July 9, 2020, we enrolled 565 mechanically ventilated patients with COVID-19. 509 patients with at least three screening samples were analysed (mean age 59·4 years [SD 12·5], 400 [79%] men). 128 (25%) patients had 138 episodes of pr/pb or possible IFIs. 76 (15%) patients fulfilled the criteria for pr/pb CAPA. According to multivariate analysis, age older than 62 years (odds ratio [OR] 2·34 [95% CI 1·39-3·92], p=0·0013), treatment with dexamethasone and anti-IL-6 (OR 2·71 [1·12-6·56], p=0·027), and long duration of mechanical ventilation (>14 days; OR 2·16 [1·14-4·09], p=0·019) were independently associated with pr/pb CAPA. 38 (7%) patients had one or more other pr/pb IFIs: 32 (6%) had candidaemia, six (1%) had invasive mucormycosis, and one (<1%) had invasive fusariosis. Multivariate analysis of associations with death, adjusted for candidaemia, for the 509 patients identified three significant factors: age older than 62 years (hazard ratio [HR] 1·71 [95% CI 1·26-2·32], p=0·0005), solid organ transplantation (HR 2·46 [1·53-3·95], p=0·0002), and pr/pb CAPA (HR 1·45 [95% CI 1·03-2·03], p=0·033). At time of ICU discharge, survival curves showed that overall ICU mortality was significantly higher in patients with pr/pb CAPA than in those without, at 61·8% (95% CI 50·0-72·8) versus 32·1% (27·7-36·7; p<0·0001).

Interpretation: This study shows the high prevalence of invasive pulmonary aspergillosis and candidaemia and high mortality associated with pr/pb CAPA in mechanically ventilated patients with COVID-19. These findings highlight the need for active surveillance of fungal pathogens in patients with severe COVID-19.

Funding: Pfizer.
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http://dx.doi.org/10.1016/S2213-2600(21)00442-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8626095PMC
November 2021

Collateral consequences of agricultural fungicides on pathogenic yeasts: A One Health perspective to tackle azole resistance.

Mycoses 2021 Nov 25. Epub 2021 Nov 25.

Nantes University Hospital and EA1155 IICiMed, Nantes University, Nantes, France.

Candida and Cryptococcus affect millions of people yearly, being responsible for a wide array of clinical presentations, including life-threatening diseases. Interestingly, most human pathogenic yeasts are not restricted to the clinical setting, as they are also ubiquitous in the environment. Recent studies raise concern regarding the potential impact of agricultural use of azoles on resistance to medical antifungals in yeasts, as previously outlined with Aspergillus fumigatus. Thus, we undertook a narrative review of the literature and provide lines of evidence suggesting that an alternative, environmental route of azole resistance, may develop in pathogenic yeasts, in addition to patient route. However, it warrants sound evidence to support that pathogenic yeasts cross border between plants, animals and humans and that environmental reservoirs may contribute to azole resistance in Candida or other yeasts for humans. As these possibilities could concern public health, we propose a road map for future studies under the One Health perspective.
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http://dx.doi.org/10.1111/myc.13404DOI Listing
November 2021

Highly Conserved Gene of Pneumocystis jirovecii in Patients with or without Prior Exposure to Echinocandins.

Antimicrob Agents Chemother 2022 Jan 1;66(1):e0156321. Epub 2021 Nov 1.

Laboratoire de Parasitologie et Mycologie, Hôpital de La Cavale Blanche, CHU de Brest, Brest, France.

Echinocandins are noncompetitive inhibitors of the GSC1 subunit of the enzymatic complex involved in synthesis of 1,3-beta-d-glucan, a cell wall component of most fungi, including Pneumocystis spp. Echinocandins are widely used for treating systemic candidiasis and rarely used for treating Pneumocystis pneumonia. Consequently, data on gene diversity are still scarce compared to that for the homologous gene of spp. In this study, we analyzed gene diversity and the putative selection pressure of echinocandins on gene sequences of specimens from two patient groups were compared. One group of 27 patients had prior exposure to echinocandins, whereas the second group of 24 patients did not, at the time of infection diagnoses. Two portions of the gene, HS1 and HS2, homologous to hot spots described in spp., were sequenced. Three single-nucleotide polymorphisms (SNPs) at positions 2204, 2243, and 2303 close to the HS1 region and another SNP at position 4540 more distant from the HS2 region were identified. These SNPs represent synonymous mutations. Three HS1 alleles, A, B, and C, and two HS2 alleles, a and b, and four haplotypes, Ca, Cb, Aa, and Ba, were defined, without significant difference in haplotype distribution in both patient groups (0.57). Considering the identical diversity of gene and the detection of synonymous mutations in both patient groups, no selection pressure of echinocandins among microorganisms can be pointed out so far.
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http://dx.doi.org/10.1128/AAC.01563-21DOI Listing
January 2022

antifungal effect of a plant-based product, CIN-102, on antifungal resistant filamentous fungi and their biofilms.

J Med Microbiol 2021 Sep;70(9)

Université de Lorraine, SIMPA, F-54000 Nancy, France.

The increase of invasive fungal infections (IFIs) and associated treatment failure in populations at risk is driving us to look for new treatments. The CIN-102 compound, derived from cinnamon essential oil, could be a new antifungal class with an activity, in particular, on strains resistant to current antifungals but also on biofilms, a factor of virulence and resistance of fungi. The aim of this study is to show the activity of CIN-102 on various strains resistant to current antifungals, on the biofilm and to determine the possibility of resistance induced with this compound. We studied the MIC of CIN-102 and of current antifungals (voriconazole and amphotericin B) using CLSI techniques against eight different strains of three genera of filamentous fungi involved in IFIs and having resistance phenotypes to current antifungals. We also determined their effects on biofilm formation, and the induced resistance by voriconazole (VRC) and CIN-102. MIC values determined for CIN-102 were between 62.5 and 250 µg ml. We demonstrated the antifungal effect of CIN-102 on biofilm, and more particularly on its formation, with 100 % inhibition achieved for most of the strains. CIN-102 at a sub-inhibitory concentration in the medium did not induce resistance in our strains, even after 30 generations. In this study we show that CIN-102 is effective against resistant filamentous fungi and against biofilm formation. In addition, our strains did not acquire a resistance phenotype against CIN-102 over time, unlike with VRC. CIN-102 is therefore an interesting candidate for the treatment of IFIs, including in cases of therapeutic failure linked to resistance, although further studies on its efficacy, safety and mechanism of action are needed.
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http://dx.doi.org/10.1099/jmm.0.001399DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8697507PMC
September 2021

Confocal spectral microscopy, a non-destructive approach to follow contamination and biofilm formation of mCherry Staphylococcus aureus on solid surfaces.

Sci Rep 2021 08 2;11(1):15574. Epub 2021 Aug 2.

UMR703 PAnTher APEX, INRAE/ONIRIS - La Chantrerie, 101 Route de Gachet, 44307, Nantes, France.

Methods to test the safety of wood material for hygienically sensitive places are indirect, destructive and limited to incomplete microbial recovery via swabbing, brushing and elution-based techniques. Therefore, we chose mCherry Staphylococcus aureus as a model bacterium for solid and porous surface contamination. Confocal spectral laser microscope (CSLM) was employed to characterize and use the autofluorescence of Sessile oak (Quercus petraea), Douglas fir (Pseudotsuga menziesii) and poplar (Populus euramericana alba L.) wood discs cut into transversal (RT) and tangential (LT) planes. The red fluorescent area occupied by bacteria was differentiated from that of wood, which represented the bacterial quantification, survival and bio-distribution on surfaces from one hour to one week after inoculation. More bacteria were present near the surface on LT face wood as compared to RT and they persisted throughout the study period. Furthermore, this innovative methodology identified that S. aureus formed a dense biofilm on melamine but not on oak wood in similar inoculation and growth conditions. Conclusively, the endogenous fluorescence of materials and the model bacterium permitted direct quantification of surface contamination by using CSLM and it is a promising tool for hygienic safety evaluation.
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http://dx.doi.org/10.1038/s41598-021-94939-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8329050PMC
August 2021

Anti-protozoal and anti-fungal evaluation of 3,5-disubstituted 1,2-dioxolanes.

Bioorg Med Chem Lett 2021 09 8;47:128196. Epub 2021 Jun 8.

BioCIS, CNRS, Université Paris-Saclay, Châtenay-Malabry 92290, France. Electronic address:

Endoperoxides are a class of compounds, which is well-known for their antimalarial properties, but few reports exist about 3,5-disubstituted 1,2-dioxolanes. After having designed a new synthetic route for the preparation of these substances, they were evaluated against 4 different agents of infectious diseases, protozoa (Plasmodium and Leishmania) and Fungi (Candida and Aspergillus). Whereas moderate antifungal activity was found for our products, potent antimalarial and antileishmanial activities were observed for a few compounds. The nature of the substituents linked to the endoperoxide ring seems to play an important role in the bioactivities.
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http://dx.doi.org/10.1016/j.bmcl.2021.128196DOI Listing
September 2021

Characterisation of the antifungal effects of a plant-based compound, CIN-102, on the main septal filamentous fungi involved in human pathology.

J Glob Antimicrob Resist 2021 06 31;25:171-180. Epub 2021 Mar 31.

Université de Lorraine, SIMPA, F-54000 Nancy, France; Université de Lorraine, CHRU-Nancy, Laboratoire de Microbiologie, F-54000 Nancy, France.

Objectives: Today, the increase of invasive fungal infections and the emergence of resistant strains are observed in medical practice. New antifungals are expected, and the plant world offers a panel of potentially active molecules. CIN-102 is a mixture of seven different compounds of plant origin developed from the formulation of cinnamon essential oil.

Methods: The in vitro activity of CIN-102 was characterised against Aspergillus spp., Fusarium spp. and Scedosporium spp. by studying the minimum inhibitory concentration (MIC), inoculum effect, germination inhibition, fungal growth, post-antifungal effect (PAFE) and synergy.

Results: MICs determined for the three genera followed a unimodal distribution and their mean values ranged from 62-250 μg/mL. CIN-102 demonstrated an inoculum effect similar to voriconazole and amphotericin B, 100% inhibition of spore germination and a PAFE.

Conclusion: CIN-102 has significant activity against filamentous fungi involved in human pathologies and should be further explored as a potential new treatment. Other studies regarding its mechanisms of action as well as animal investigations are awaited.
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http://dx.doi.org/10.1016/j.jgar.2021.03.017DOI Listing
June 2021

The Extent of Aspergillosis in Critically Ill Patients With Severe Influenza Pneumonia: A Multicenter Cohort Study.

Crit Care Med 2021 06;49(6):934-942

Medical Intensive Care Unit, Centre Hospitalier et Universitaire de Brest, Université de Bretagne Occidentale, Brest, France.

Objectives: To determine the frequency and prognosis of invasive pulmonary aspergillosis in critically ill patients with severe influenza pneumonia.

Design: Retrospective multicenter cohort study.

Setting: Five French ICUs.

Patients: Patients with influenza admitted to ICU between 2009 and 2018.

Measurements And Main Results: Of the 524 patients admitted for severe influenza diagnosed with a positive airway reverse-transcriptase polymerase chain reaction test, 450 (86%) required mechanical ventilation. A lower respiratory tract sample yielded with Aspergillus (Asp+) in 28 patients (5.3%). Ten patients (1.9%) were diagnosed with putative or proven invasive pulmonary aspergillosis, based on the validated AspICU algorithm. A multivariate model was built to identify independent risk factors for Aspergillus-positive pulmonary culture. Factors independently associated with Aspergillus-positive culture were liver cirrhosis (odds ratio = 6.7 [2.1-19.4]; p < 0.01), hematologic malignancy (odds ratio = 3.3 [1.2-8.5]; p = 0.02), Influenza A(H1N1)pdm09 subtype (odds ratio = 3.9 [1.6-9.1]; p < 0.01), and vasopressor requirement (odds ratio = 4.1 [1.6-12.7]; p < 0.01). In-hospital mortality of Asp+ patients was 36% versus 21% in patients without Aspergillus-positive pulmonary culture (p = 0.09).

Conclusions: In this large retrospective multicenter cohort of critically ill patients, putative invasive pulmonary aspergillosis according to AspICU algorithm was a relatively rare complication of influenza. Patients at higher risk of Aspergillus pulmonary colonization included those with liver cirrhosis, hematologic malignancy, H1N1pdm09 influenza A virus, and requiring vasopressors. Our results provide additional data on the controversial association between severe influenza and invasive pulmonary aspergillosis. Reaching a consensual definition of invasive pulmonary aspergillosis becomes mandatory and confers further prospective research.
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http://dx.doi.org/10.1097/CCM.0000000000004861DOI Listing
June 2021

Multicenter Evaluation of an ELISA for the Detection of spp. Antigen in Clinical Human Stool Samples.

Microorganisms 2021 01 20;9(2). Epub 2021 Jan 20.

EA ESCAPE 7510, University of Medicine Pharmacy Rouen, 76000 Rouen, France.

Human cryptosporidiosis remains underdiagnosed, and rapid/accurate diagnosis is of clinical importance. Diagnosis of the oocyst in stool samples by conventional microscopy is labor-intensive, time-consuming, and requires skillful experience. Thus, we aimed to evaluate the usefulness of a coproantigen enzyme-linked immunosorbent assay (ELISA) test in detecting spp. from fecal specimens. For this aim, we evaluated the performances of a commercial ELISA (CoproELISA kit, Savyon Diagnostics, Israel) for the detection of spp. in random clinical stool samples through a multicenter study. The sensitivity and specificity for coproantigen ELISA were 98.86% and 94.32%, respectively. The coproantigen ELISA results indicate that the simple, rapid, reliable, and standardized immunoassay test is sensitive and specific for routine diagnosis, and may be useful for large-scale epidemiological studies of cryptosporidiosis.
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http://dx.doi.org/10.3390/microorganisms9020209DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7909390PMC
January 2021

Survival of Bacterial Strains on Wood () Compared to Polycarbonate, Aluminum and Stainless Steel.

Antibiotics (Basel) 2020 Nov 13;9(11). Epub 2020 Nov 13.

Laboratoire de Bactériologie-Hygiène, Centre Hospitalier Universitaire, 4 rue Larrey, 49933 Angers, France.

Healthcare-associated infections (HAI) remain a burden in healthcare facilities, environmental surfaces being a potential reservoir for healthcare-associated pathogens. In this context, exploration of materials with potential antimicrobial activities represents a way forward for the future. Here, we explored the survival of four bacterial species commonly involved in HAI (, , , ), on oak versus three other materials (aluminum, polycarbonate, stainless steel). Twenty microliters of each bacterial suspension (approximatively 10 bacteria) were deposited on each material. Bacterial counts were measured by grinding and culturing on day 0, 1, 2, 6, 7 and 15. Analyses were performed in triplicate for each material and each time evaluated. It appeared that the bacteria viable count decreased rapidly on transversal and tangential oak compared with the other materials for all bacterial species. Furthermore, no difference was noticed between transversal and tangential oak. These results underline the potential for use of oak materials in healthcare facilities, a consideration that should be supported by further investigations.
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http://dx.doi.org/10.3390/antibiotics9110804DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7698295PMC
November 2020

Comparison of MultiLocus Sequence Typing (MLST) and Microsatellite Length Polymorphism (MLP) for genotyping.

Comput Struct Biotechnol J 2020 15;18:2890-2896. Epub 2020 Oct 15.

Laboratoire de Parasitologie-Mycologie; AP-HP, Groupe Hospitalier Saint-Louis-Lariboisière-Fernand-Widal, Paris, France.

is an atypical fungus responsible for severe respiratory infections, often reported as local outbreaks in immunocompromised patients. Epidemiology of this infection, and transmission risk emphasises the need for developing genotyping techniques. Currently, two methods have emerged: Multilocus Sequence typing (MLST) and microsatellite length polymorphism (MLP). Here we compare an MLST strategy, including 2 nuclear loci and 2 mitochondrial loci, with an MLP strategy including 6 nuclear markers using 37 clinical PCR-positive respiratory samples from two French hospitals. MLST and MLP provided 30 and 35 different genotypes respectively. A higher number of mixed infections was detected using MLP (48.6% vs. 13.5% respectively;  = 0.002). Only one MLP marker (STR279) was statistically associated with the geographical origin of samples. Haplotype network inferred using the available genotypes yielded expanded network for MLP, characterized by more mutational steps as compared to MLST, suggesting that the MLP approach is more resolutive to separate genotypes. The correlation between genetic distances calculated based on MLST and MLP was modest with a value = 0.32 ( < 0.001). Finally, both genotyping methods fulfilled important criteria: (i) a discriminatory power from 97.5% to 99.5% and (ii) being quick and convenient genotyping tools. While MLP appeared highly resolutive regarding genotypes mixture within samples, using one genotyping method rather than the other may also depend on the context (i.e., MLST for investigation of suspected clonal outbreaks versus MLP for population structure study) as well as local facilities.
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http://dx.doi.org/10.1016/j.csbj.2020.10.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7593342PMC
October 2020

Wood materials for limiting the bacterial reservoir on surfaces in hospitals: would it be worthwhile to go further?

Future Microbiol 2020 10 6;15:1431-1437. Epub 2020 Nov 6.

Laboratoire de Bactériologie-Hygiène, Centre Hospitalier Universitaire, 4 Rue Larrey 49933 Angers Cedex, France.

To assess the activity of (oak) on five bacterial species/genus frequently involved in hospital-acquired infections for evaluating the interest of going further in exploring the possibilities of using untreated wood as a material in the hospital setting. We studied the activity of by the disk diffusion method. was active on and complex, two bacterial species particularly resistant in the hospital environment, independently from their resistance to antibiotics, and was slightly active on . Concurrently, was not active on Enterococci and . Overall, untreated wood material presented antimicrobial properties that could have an impact on the cross-transmission of certain bacterial species in healthcare settings.
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http://dx.doi.org/10.2217/fmb-2019-0339DOI Listing
October 2020

In vitro identification of imidazo[1,2-a]pyrazine-based antileishmanial agents and evaluation of L. major casein kinase 1 inhibition.

Eur J Med Chem 2021 Jan 23;210:112956. Epub 2020 Oct 23.

Université de Nantes, Cibles et Médicaments des Infections et du Cancer, IICiMed, EA 1155, F-44000, Nantes, France. Electronic address:

Leishmaniasis constitutes a severe public health problem, with an estimated prevalence of 12 million cases. This potentially fatal disease has a worldwide distribution and in 2012, the fatal Visceral Leishmaniasis (VL) was declared as new emerging disease in Europe, mainly due to global warming, with expected important public health impact. The available treatments are toxic, costly or lead to parasite resistance, thus there is an urgent need for new drugs with new mechanism of action. Previously, we reported the discovery of CTN1122, a potent imidazo[1,2-a]pyrazine-based antileishmanial hit compound targeting L-CK1.2 at low micromolar ranges. Here, we described structurally related, safe and selective compounds endowed with antiparasitic properties, better than miltefosine, the reference therapy by oral route. L-CK1.2 homology model gave the first structural explanations of the role of 4-pyridyl (CTN1122) and 2-aminopyrimidin-4-yl (compound 21) moieties, at the position 3 of the central core, in the low micromolar to nanomolar L-CK1.2 inhibition, whereas N-methylpyrazole derivative 11 remained inactive against the parasite kinase.
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http://dx.doi.org/10.1016/j.ejmech.2020.112956DOI Listing
January 2021

A One Health Perspective to Recognize as Important in Clinical Practice.

J Fungi (Basel) 2020 Oct 20;6(4). Epub 2020 Oct 20.

Grupo de Investigación Celular y Molecular de Microorganismos Patógenos (CeMoP), Departamento de Ciencias Biológicas, Universidad de Los Andes, Bogotá 111711, Colombia.

Any strategy that proposes solutions to health-related problems recognizes that people, animals, and the environment are interconnected. is an example of this interaction because it is capable of infecting plants, animals, and humans. This review provides information on various aspects of these relations and proposes how to approach fusariosis with a One Health methodology (a multidisciplinary, and multisectoral approach that can address urgent, ongoing, or potential health threats to humans, animals, and the environment). Here, we give a framework to understand infection pathogenesis, through the epidemiological triad, and explain how the broad utilization of fungicides in agriculture may play a role in the treatment of human fusariosis. We assess how plumbing systems and hospital environments might play a role as a reservoir for animal and human infections. We explain the role of antifungal resistance mechanisms in both humans and agriculture. Our review emphasizes the importance of developing interdisciplinary research studies where aquatic animals, plants, and human disease interactions can be explored through coordination and collaborative actions.
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http://dx.doi.org/10.3390/jof6040235DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7711799PMC
October 2020

Synthesis, Optimization, Antifungal Activity, Selectivity, and CYP51 Binding of New 2-Aryl-3-azolyl-1-indolyl-propan-2-ols.

Pharmaceuticals (Basel) 2020 Aug 8;13(8). Epub 2020 Aug 8.

EA1155-IICiMed, Institut de Recherche en Santé 2, Département de Pharmacochimie, Nantes Atlantique Universités, Université de Nantes, F-44200 Nantes, France.

A series of 2-aryl-3-azolyl-1-indolyl-propan-2-ols was designed as new analogs of fluconazole (FLC) by replacing one of its two triazole moieties by an indole scaffold. Two different chemical approaches were then developed. The first one, in seven steps, involved the synthesis of the key intermediate 1-(1-benzotriazol-1-yl)methyl-1-indole and the final opening of oxiranes by imidazole or 1-1,2,4-triazole. The second route allowed access to the target compounds in only three steps, this time with the ring opening by indole and analogs. Twenty azole derivatives were tested against and other species. The enantiomers of the best anti- compound, 2-(2,4-dichlorophenyl)-3-(1-indol-1-yl)-1-(1-1,2,4-triazol-1-yl)-propan-2-ol (), were analyzed by X-ray diffraction to determine their absolute configuration. The (-)- enantiomer (Minimum inhibitory concentration (MIC) = IC = 0.000256 µg/mL on CA98001) was found with the -absolute configuration. In contrast the ()- enantiomer was found with the -absolute configuration (MIC = 0.023 µg/mL on CA98001). By comparison, the MIC value for FLC was determined as 0.020 µg/mL for the same clinical isolate. Additionally, molecular docking calculations and molecular dynamics simulations were carried out using a crystal structure of lanosterol 14α-demethylase (CaCYP51). The (-)-()- enantiomer aligned with the positioning of posaconazole within both the heme and access channel binding sites, which was consistent with its biological results. All target compounds have been also studied against human fetal lung fibroblast (MRC-5) cells. Finally, the selectivity of four compounds on a panel of human P450-dependent enzymes (CYP19, CYP17, CYP26A1, CYP11B1, and CYP11B2) was investigated.
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http://dx.doi.org/10.3390/ph13080186DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7464559PMC
August 2020

Impact of fecal microbiota transplantation on chronic recurrent pouchitis in ulcerative colitis with ileo-anal anastomosis: study protocol for a prospective, multicenter, double-blind, randomized, controlled trial.

Trials 2020 Jun 3;21(1):455. Epub 2020 Jun 3.

Gastroenterology Department, Institute of Digestive Diseases (Institut des Maladies de l'Appareil Digestif - IMAD), CHU Nantes and Nantes University, Nantes, France.

Background: Almost 15% of patients with ulcerative colitis (UC) will require a proctocolectomy with ileal pouch-anal anastomosis (IPAA) as a result of fulminant colitis, dysplasia, cancer, or medical refractory diseases. Around 50% will experience pouchitis, an idiopathic inflammatory condition involving the ileal reservoir, responsible for digestive symptoms, deterioration in quality of life, and disability. Though the majority of initial cases of pouchitis are easily managed with a short course of antibiotics, in about 10% of cases, inflammation of the pouch becomes chronic with very few treatments available. Previous studies have suggested that manipulating the composition of intestinal flora through antibiotics, probiotics, and prebiotics achieved significant results for treating acute episodes of UC-associated pouchitis. However, there is currently no established effective treatment for chronic antibiotic-dependent pouchitis. Fecal microbiota transplantation (FMT) is a novel therapy involving the transfer of normal intestinal flora from a healthy donor to a patient with a medical condition potentially caused by the disrupted homeostasis of intestinal microbiota or dysbiosis.

Methods: Our project aims to compare the delay of relapse of chronic recurrent pouchitis after FMT versus sham transplantation. Forty-two patients with active recurrent pouchitis after having undergone an IPAA for UC will be enrolled at 12 French centers. The patients who respond to antibiotherapy will be randomized at a ratio of 1:1 to receive either FMT or sham transplantation.

Discussion: On April 30, 2014, the World Health Organization published an alarming report on antibiotic resistance. Finding an alternative medical treatment to antibiotics in order to prevent relapses of pouchitis is therefore becoming increasingly important given the risk posed by multiresistant bacteria. Moreover, if the results of this study are conclusive, FMT, which is less expensive than biologics, could become a routine treatment in the future.

Trial Registration: ClinicalTrials.gov, NCT03524352. Registered on 14 May 2018.
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http://dx.doi.org/10.1186/s13063-020-04330-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7267479PMC
June 2020

Medicopsis romeroi nodular subcutaneous infection in a kidney transplant recipient.

Int J Infect Dis 2020 Jun 24;95:262-264. Epub 2020 Apr 24.

Laboratoire de Parasitologie et Mycologie Médicale, CHU de Nantes, France; Département de Parasitologie et Mycologie Médicale, Université de Nantes, Nantes Atlantique Universités, EA1155 - IICiMed, Institut de Recherche en Santé 2, Nantes, France. Electronic address:

Phaeohyphomycosis is a set of fungal infections caused by various dematiaceous fungi such as coelomycetes. These infections can occur either in immunocompetent or immunocompromised patients like solid organ transplants. Here we describe a nodular lesion of the right hallux that occurred in a kidney transplant patient. Microscopic examination of the biopsy revealed fungal hyphae and culture was positive to a grey to black mould that lacked characteristic elements to be identified. Nucleic acid sequencing targeting the internal transcribed spacer of the ribosomal DNA identified this mould as Medicopsis romeroi. The patient benefited of an antifungal therapy with voriconazole associated with surgical excision of the lesion. No relapse of the lesion was observed during a six-month follow-up. In solid organ transplants, phaeohyphomycosis caused by Medicopsis romeroi are very rare with only 12 cases reported. The clinical history should be well assessed since the lesion can appear several years after a cutaneous trauma that happened in a tropical region. Therapy generally combines antifungals with surgical excision of the lesion in order to avoid any relapse or dissemination of the infection.
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http://dx.doi.org/10.1016/j.ijid.2020.04.028DOI Listing
June 2020

In vitro immune responses of human PBMCs against Candida albicans reveals fungal and leucocyte phenotypes associated with fungal persistence.

Sci Rep 2020 04 10;10(1):6211. Epub 2020 Apr 10.

Nantes Université, CHU de Nantes, Cibles et médicaments des infections et du cancer, IICiMed, EA 1155, F-44000, Nantes, France.

Although there is a growing understanding of immunity against Candida albicans, efforts need to be pursued in order to decipher the cellular mechanisms leading to an uncontrolled immune response that eventually oppose disease eradication. We describe here significant intra- and inter-subject variations in immune response patterns of major human leucocyte subsets following an in vitro challenge with C. albicans clinical isolates. We also observed that there are Candida isolate-dependent changes in leucocyte phenotypes. Through a combination of multiple fungal growth and flow cytometric measurements, coupled to the tSNE algorithm, we showed that significant proliferation differences exist among C. albicans isolates, leading to the calculation of a strain specific persistent index. Despite substantial inter-subject differences in T cells and stability of myeloid cells at baseline, our experimental approach highlights substantial immune cell composition changes and cytokine secretion profiles after C. albicans challenge. The significant secretion of IL-17 by CD66+ cells, IFN-γ and IL-10 by CD4+ T cells 2 days after C. albicans challenge was associated with fungal control. Fungal persistence was associated with delayed secretion of IFN-γ, IL-17, IL-4, TNF-α and IL-10 by myeloid cells and IL-4 and TNF-α secretion by CD4+ and CD8+ T cells. Overall, this experimental and analytical approach is available for the monitoring of such fungal and human immune responses.
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http://dx.doi.org/10.1038/s41598-020-63344-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7148345PMC
April 2020

Persistence of Clonal Azole-Resistant Isolates of from a Patient with Chronic Mucocutaneous Candidiasis in Colombia.

J Glob Infect Dis 2020 Jan-Mar;12(1):16-20. Epub 2020 Feb 19.

Unidad de Proteómica y Micosis Humanas, Grupo de Enfermedades Infecciosas Departamento de Microbiología, Facultad de Ciencias, Pontificia Universidad Javeriana, Bogotá, Colombia.

Purpose: The present article describes retrospectively a case of a patient with chronic mucocutaneous candidiasis (CMC) who presented recurrent infection since he was 6 months old. We obtained 16 isolates recovered during a 4-year period. Our purpose was to determinate the susceptibility, genotyping, and the pathogenicity profile in all the isolates.

Methods: Sixteen were isolated from a 25-year-old male with several recurrent fungal infections admitted to Hospital. The isolates were recovered during 4 years from a different anatomical origin. We typified them by multilocus sequence typing, also we evaluated susceptibility to fluconazole, itraconazole, voriconazole, posaconazole, isavuconazole, caspofungin, and amphotericin B by microdilution method and we also test the pathogenic capacity in the model.

Results: Genotyping of all clinical isolates showed the persistence of the same diploid sequence type (DST). Isolates changed their susceptibility profile over time, but there were no significant statistical differences in pathogenicity.

Conclusion: Herein, a persistent clonal isolates of (DST 918) in a patient with CMC, showed changes in its susceptibility profile after several antifungal treatments acquiring gradual resistance to the azole drugs, which did not affect their pathogenicity.
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http://dx.doi.org/10.4103/jgid.jgid_74_19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7045761PMC
February 2020

Mucorales DNA detection in serum specimens for early diagnosis of mucormycosis.

Diagn Microbiol Infect Dis 2020 Jun 29;97(2):115004. Epub 2020 Jan 29.

Laboratoire de Parasitologie et Mycologie, Hôpital de La Cavale Blanche, CHU de Brest, Brest, France; Groupe d'Etude des Interactions Hôte-Pathogène (GEIHP; EA 3142), Université de Bretagne Occidentale-Université d'Angers, Brest, France. Electronic address:

We report a case of pulmonary mucormycosis in a patient with T-cell acute lymphoblastic leukemia. The diagnosis of mucormycosis was initially based on mycological examination of a pulmonary specimen. However, we describe how it could have been made 2 months earlier using polymerase chain reaction assays targeting Mucorales species on serum specimens.
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http://dx.doi.org/10.1016/j.diagmicrobio.2020.115004DOI Listing
June 2020

Invasive Aspergillosis Due to Aspergillus Section Usti: A Multicenter Retrospective Study.

Clin Infect Dis 2021 04;72(8):1379-1385

Department of Infectious Diseases, Faculty of Medicine, Hacettepe University, Ankara, Turkey.

Background: Aspergillus spp. of section Usti (A. ustus) represent a rare cause of invasive aspergillosis (IA). This multicenter study describes the epidemiology and outcome of A. ustus infections.

Methods: Patients with A. ustus isolated from any clinical specimen were retrospectively identified in 22 hospitals from 8 countries. When available, isolates were sent for species identification (BenA/CaM sequencing) and antifungal susceptibility testing. Additional cases were identified by review of the literature. Cases were classified as proven/probable IA or no infection, according to standard international criteria.

Results: Clinical report forms were obtained for 90 patients, of whom 27 had proven/probable IA. An additional 45 cases were identified from literature review for a total of 72 cases of proven/probable IA. Hematopoietic cell and solid-organ transplant recipients accounted for 47% and 33% cases, respectively. Only 8% patients were neutropenic at time of diagnosis. Ongoing antimold prophylaxis was present in 47% of cases. Pulmonary IA represented 67% of cases. Primary or secondary extrapulmonary sites of infection were observed in 46% of cases, with skin being affected in 28% of cases. Multiple antifungal drugs were used (consecutively or in combination) in 67% of cases. The 24-week mortality rate was 58%. A. calidoustus was the most frequent causal agent. Minimal inhibitory concentrations encompassing 90% isolates (MIC90) were 1, 8, >16, and 4 µg/mL for amphotericin B, voriconazole, posaconazole, and isavuconazole, respectively.

Conclusions: Aspergillus ustus IA mainly occurred in nonneutropenic transplant patients and was frequently associated with extrapulmonary sites of infection. Mortality rate was high and optimal antifungal therapy remains to be defined.
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http://dx.doi.org/10.1093/cid/ciaa230DOI Listing
April 2021

Impact of calmodulin inhibition by fluphenazine on susceptibility, biofilm formation and pathogenicity of caspofungin-resistant Candida glabrata.

J Antimicrob Chemother 2020 05;75(5):1187-1193

Department of Parasitology and Medical Mycology, University of Nantes, Nantes Atlantique Universities, Faculty of Pharmacy, Nantes, France.

Background: In recent decades, Candida glabrata has emerged as a frequent cause of life-threatening fungal infection. In C. glabrata, echinocandin resistance is associated with mutations in FKS1/FKS2 (β-1,3-glucan synthase). The calmodulin/calcineurin pathway is implicated in response to antifungal stress and calcineurin gene disruption specifically reverses Fks2-mediated resistance of clinical isolates.

Objectives: We evaluated the impact of calmodulin inhibition by fluphenazine in two caspofungin-resistant C. glabrata isolates.

Methods: C. glabrata isolates were identified by ITS1/ITS4 (where ITS stands for internal transcribed spacer) sequencing and the echinocandin target FKS1/FKS2 genes were sequenced. Susceptibility testing of caspofungin in the presence of fluphenazine was performed by a modified CLSI microbroth dilution method. The effect of the fluphenazine/caspofungin combination on heat stress (37°C or 40°C), oxidative stress (0.2 and 0.4 mM menadione) and biofilm formation (polyurethane catheter) was analysed. A Galleria mellonella model using blastospores (1 × 109 cfu/mL) was developed to evaluate the impact of this combination on larval survival.

Results: F659del was found in the FKS2 gene of both resistant strains. In these clinical isolates, fluphenazine increased susceptibility to caspofungin and reduced their thermotolerance. Furthermore, the fluphenazine/caspofungin combination significantly impaired biofilm formation in an in vitro polyurethane catheter model. All these features participated in the increasing survival of infected G. mellonella after combination treatment in comparison with caspofungin alone.

Conclusions: In a repurposing strategy, our findings confirm that calmodulin could provide a relevant target in life-threatening fungal infectious diseases.
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http://dx.doi.org/10.1093/jac/dkz565DOI Listing
May 2020

New azole antifungals with a fused triazinone scaffold.

Eur J Med Chem 2020 Mar 20;189:112082. Epub 2020 Jan 20.

Université de Nantes, Nantes Atlantique Universités, Département de Chimie Thérapeutique, Cibles et Médicaments des Infections et du Cancer, IICIMED- EA1155, Institut de Recherche en Santé 2, F-44200, Nantes, France. Electronic address:

We identified a new series of azole antifungal agents bearing a pyrrolotriazinone scaffold. These compounds exhibited a broad in vitro antifungal activity against pathogenic Candida spp. (fluconazole-susceptible and fluconazole-resistant) and were 10- to 100-fold more active than voriconazole against two Candida albicans isolates with known mechanisms of azole resistance (overexpression of efflux pumps and/or specific point substitutions in the Erg11p/CYP51 enzyme). Our lead compound 12 also displayed promising in vitro antifungal activity against some filamentous fungi such as Aspergillus fumigatus and the zygomycetes Rhizopus oryzae and Mucor circinelloides and an in vivo efficiency against two murine models of lethal systemic infections caused by Candida albicans.
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http://dx.doi.org/10.1016/j.ejmech.2020.112082DOI Listing
March 2020

Biological exploration of a novel 1,2,4-triazole-indole hybrid molecule as antifungal agent.

J Enzyme Inhib Med Chem 2020 Dec;35(1):398-403

Département de Parasitologie et Mycologie Médicale, Université de Nantes, Nantes Atlantique Universités, EA1155 - IICiMed, Institut de Recherche en Santé 2, Nantes, France.

(2-(2,4-Dichlorophenyl)-3-(1-indol-1-yl)-1-(1,2,4-1-triazol-1-yl)propan-2-ol (), a new 1,2,4-triazole-indole hybrid molecule, showed a broad-spectrum activity against particularly against low fluconazole-susceptible species. Its activity was higher than fluconazole and similar to voriconazole on (MIC = 0.25, 64 and 1 µg/mL, respectively), (MIC = 0.125, 64 and 0.125 µg/mL, respectively) and (MIC = 0.5, 8 and 0.25 µg/mL, respectively). The action mechanisms of were also identified as inhibition of ergosterol biosynthesis and phospholipase A2-like activity. At concentration as low as 4 ng/mL, 8g inhibited ergosterol production by 82% and induced production of 14a-methyl sterols, that is comparable to the results obtained with fluconazole at higher concentration. demonstrated moderate inhibitory effect on phospholipase A2-like activity being a putative virulence factor. Due to a low MRC5 cytotoxicity, this compound presents a high therapeutic index. These results pointed out that is a new lead antifungal candidate with potent ergosterol biosynthesis inhibition.
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http://dx.doi.org/10.1080/14756366.2019.1705292DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6968525PMC
December 2020

A Decade of Antifungal Leads from Natural Products: 2010-2019.

Pharmaceuticals (Basel) 2019 Dec 12;12(4). Epub 2019 Dec 12.

School of Pharmacy, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK.

In this review, we discuss novel natural products discovered within the last decade that are reported to have antifungal activity against pathogenic species. Nearly a hundred natural products were identified that originate from bacteria, algae, fungi, sponges, and plants. Fungi were the most prolific source of antifungal compounds discovered during the period of review. The structural diversity of these antifungal leads encompasses all the major classes of natural products including polyketides, shikimate metabolites, terpenoids, alkaloids, and peptides.
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http://dx.doi.org/10.3390/ph12040182DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6958371PMC
December 2019

Double positive CD4+CD8+ T cells are part of the adaptive immune response against Candida albicans.

Hum Immunol 2019 Dec 24;80(12):999-1005. Epub 2019 Sep 24.

Nantes Université, CHU de Nantes, Cibles et médicaments des infections et du cancer, IICiMed, EA 1155, F-44000 Nantes, France. Electronic address:

Although multiple immune cells participate in the innate and adaptive immune response against Candida albicans, the elucidation of cellular and inflammation kinetics may be a promising strategy to decipher events propitious to infection eradication. We used an in vitro Candida-human leucocyte coculture approach to study the dynamics of rare CD4+CD8+ double positive T lymphocytes (DP T) produced in response to this fungus. Our results highlight the presence of two phenotypically distinct subsets of DP T cells: CD4hiCD8lo and CD4loCD8hi, and that the different ratio of these cells correlates with infection outcome. The ratio of CD4hiCD8lo over CD4loCD8hi by day 6 was significantly higher in controlled infections and decreased when infection persisted due to a significant increase in the proportion of CD4loCD8hi. When infection was controlled, CD4hiCD8lo T cells secreted IFNγ, TNFα, IL-4 and IL-10 cytokines two days after challenge. By day 2, under conditions of persistent infection, CD4hiCD8lo and CD4loCD8hi T cells secreted significant levels of IL-4 and IL-10, respectively, compared to uninfected cultures. Frequency kinetics and original cytokine profiles detailed in this work indicate that DP T cells could participate in the adaptive immune response to C. albicans.
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http://dx.doi.org/10.1016/j.humimm.2019.09.008DOI Listing
December 2019

Comparison between MALDI-TOF MS and MicroScan in the identification of emerging and multidrug resistant yeasts in a fourth-level hospital in Bogotá, Colombia.

BMC Microbiol 2019 05 23;19(1):106. Epub 2019 May 23.

Departamento de Microbiología, Facultad de Ciencias, Unidad de Proteomica y Micosis Humanas, Grupo de Enfermedades Infecciosas, Pontificia Universidad Javeriana, Bogotá, Colombia.

Background: The introduction of MALDI-TOF MS in the clinical microbiology laboratory has modified the approaches for the identification of fungi. Thanks to this tool, it is possible to identify cryptic species, which possess critical susceptibility patterns. Clinical strains were identified using the MicroScan and MALDI-TOF MS systems. Discrepant results from both methods were investigated using ITS rDNA barcoding. Finally, these isolates were also tested for in vitro susceptibility.

Results: The percentage of agreement between both methods to 498 yeast isolates was of 93.6% (32 discrepant isolates). The concordance of ITS sequencing with MALDI-TOF MS was higher (99%) than that of MicroScan (94%). Several of these discordant yeasts displayed high MICs for antifungal agents.

Conclusions: Our study highlights the need of the MS and molecular approaches such as MALDI-TOF MS and ITS rDNA barcoding for the correct identification of emerging or cryptic yeast species; besides, some of these could be multidrug resistant. This work was the first experience in the implementation of the MALDI-TOF MS technology in Colombia. We found the first uncommon yeasts including Candida auris and we could identify Trichosporon faecalis. Our work highlights a clear necessity of an accurate yeast identification as a much more pertinent technique than the susceptibility profiles, because the most unusual yeasts exhibit resistance profiles to the few available antifungals.
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http://dx.doi.org/10.1186/s12866-019-1482-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6533654PMC
May 2019

Precise genome editing using a CRISPR-Cas9 method highlights the role of CoERG11 amino acid substitutions in azole resistance in Candida orthopsilosis.

J Antimicrob Chemother 2019 08;74(8):2230-2238

Laboratoire de Parasitologie-Mycologie, CHU de Nantes, Nantes, France.

Background: Azoles are one of the main antifungal classes for the treatment of candidiasis. In the current context of emerging drug resistance, most studies have focused on Candida albicans, Candida glabrata or Candida auris but, so far, less is known about the underlying mechanisms of resistance in other species, including Candida orthopsilosis.

Objectives: We investigated azole resistance in a C. orthopsilosis clinical isolate recovered from a patient with haematological malignancy receiving fluconazole prophylaxis.

Methods: Antifungal susceptibility to fluconazole was determined in vitro (CLSI M27-A3) and in vivo (in a Galleria mellonella model of invasive candidiasis). The CoERG11 gene was then sequenced and amino acid substitutions identified were mapped on the predicted 3D structure of CoErg11p. A clustered regularly interspaced short palindromic repeat-Cas9 (CRISPR-Cas9) genome-editing strategy was used to introduce relevant mutations into a fluconazole-susceptible C. orthopsilosis isolate.

Results: Compared with unrelated C. orthopsilosis isolates, the clinical isolate exhibited both in vitro and in vivo fluconazole resistance. Sequencing of the CoERG11 gene identified several amino acid substitutions, including two possibly involved in fluconazole resistance (L376I and G458S). Both mutations mapped close to the active site of CoErg11p. Engineering these mutations in a different genetic background using CRISPR-Cas9 demonstrated that G458S, but not L376I, confers resistance to fluconazole and voriconazole.

Conclusions: Our data show that the G458S amino acid substitution in CoERG11p, but not L376I, contributes to azole resistance in C. orthopsilosis. In addition to highlighting the potential of CRISPR-Cas9 technology for precise genome editing in the field of antifungal resistance, we discuss some points that are critical to improving its efficiency.
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http://dx.doi.org/10.1093/jac/dkz204DOI Listing
August 2019
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