Antifungal Activity of Amphiphilic Perylene Bisimides

[EN] Perylene-based compounds, either naturally occurring or synthetic, have shown interesting biological activities. In this study, we report on the broad-spectrum antifungal properties of two lead amphiphilic perylene bisimides, compounds 4 and 5, which were synthesized from perylene-3,4,9,10-tetracarboxylic dianhydride by condensation with spermine and an ammonium salt formation. The antifungal activity was evaluated using a collection of fungal strains and clinical isolates from patients with onychomycosis or sporotrichosis. Both molecules displayed an interesting antifungal profile with MIC values in the range of 2-25 mu M, being as active as several reference drugs, even more potent in some particular strains. The ammonium trifluoroacetate salt 5 showed the highest activity with a MIC value of 2.1 mu M for all tested Candida spp., two Cryptococcus spp., two Fusarium spp., and one Neoscytalidium spp. strain. Therefore, these amphiphilic molecules with the perylene moiety and cationic ammonium side chains represent important structural features for the development of novel antifungals.This study was supported by grant 201680I008 (awarded to M.A.G.-C.) from the Spanish Government (Consejo Superior de Investigaciones Cientificas) and grant 3756 of the University of Antioquia.Roa-Linares, VC.; Mesa-Arango, AC.; Zaragoza, RJ.; González-Cardenete, MA. (2022). Antifungal Activity of Amphiphilic Perylene Bisimides. Molecules. 27(20):1-12. https://doi.org/10.3390/molecules27206890112272

Development and validation of an in-house library for filamentous fungi identification by MALDI-TOF MS in a clinical laboratory in Medellin (Colombia)

Identification of filamentous fungi by conventional phenotypic methods are time-consuming, and a correct identification at the species level is prone to errors. Therefore, a more accurate and faster time-to-results, and cost-effective technique, is required, such as the Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS). In this study, we describe the development of an in-house spectra library for the identification of filamentous fungi frequently isolated from patients with infections. An in-house spectra library was constructed using 14 reference strains grown in solid medium. Clinical isolates were identified either by the in-house spectra library or the Biotyper commercial library from Bruker Daltonics. Fungal identification was carried following the Biotyper’s established scores: ≤1.699: not reliably identified (NRI); 1.700–1.999: genus-level; ≥2.000: species-level. Clinical isolates were identified, with the in-house library, at species- and genus-level at 88.70% (55) and 3.22% (2), respectively. While 4.80% (3) was NRI and 3.22% (2) was discrepant concerning sequencing. On the contrary, identification up to species and genus-level with the commercial library was 44.44% (16) and 22.22% (8), respectively. NRI and the discrepancy was 30.55% (11) and 2.77% (1), respectively. For the reaming 26 isolates, 16 from Neoscytalidium dimidiatum and 10 from Sporothrix spp., respectively, the absence of spectrum and the specific spectra within the Sporothrix complex in the commercial library resulted in the inability to obtain an identification. In conclusion, the current results advocate the importance that each clinical microbiological laboratory needs to develop an ad hoc library associated with the MALDI-TOF MS fungal identification to overcome the limitations of the available commercial libraries.Comité para el Desarrollo de la Investigación, Universidad de Antioquia, Medellín, Colombia. Grant No 2604 and by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020-Programa Operacional Regional do Norteinfo:eu-repo/semantics/publishedVersio

Identificación de aislamientos clínicos del complejo Sporothrix schenckii por espectrometría de masas MALDI-TOF

[Extracto] Introducción: Sporothrix schenckii es un hongo dimórfico, que por mucho tiempo fue considerado la única especie responsable de la esporotricosis. Sin embargo, con el advenimiento de las técnicas de biología molecular, aplicadas a la investigación taxonómica de hongos, se demostró que S. schenckii es un complejo de especies crípticas conformado por S. brasiliensis, S. schenckii sensu stricto, S. globosa, S. mexicana y S. luriei. Estas especies difieren en sensibilidad antifúngica, distribución geográfica y virulencia. Tradicionalmente la identificación de Sporothrix spp., se ha llevado a cabo con métodos fenotípicos clásicos, sin embargo, estos no son suficientes para la clasificación correcta de las diferentes especies, por la similitud morfológica entre ellas. La espectrometría de masas (EM) MALDI-TOF es una técnica ampliamente utilizada para la identificación rápida de diferentes microorganismos. El objetivo del presente trabajo fue la creación y validación de una base de espectros de diferentes especies del complejo S. schenckii a partir de la forma micelial del hongo para identificar aislamientos clínicos por EM- MALDI-TOF. [...]info:eu-repo/semantics/publishedVersio

Impact of Resistance to Fluconazole on Virulence and Morphological Aspects of Cryptococcus neoformans and Cryptococcus gattii Isolates

Cryptococcus spp. are responsible for around one million cases of meningitis every year. Fluconazole (FLU) is commonly used in the treatment of cryptococcosis, mainly in immunocompromised patients and the resistance is usually reported after long periods of treatment. In this study, the morphological characterization and virulence profile of FLU-susceptible and FLU-resistant clinical and environmental isolates of C. neoformans and C. gattii were performed both in vitro and in vivo using the Galleria mellonella model. FLU-susceptible isolates from C. neoformans were significantly more virulent than the FLU-resistant isolates. FLU-susceptible C. gattii isolates showed a different virulence profile from C. neoformans isolates where only the environmental isolate, CL, was more virulent compared with the resistant isolates. Cell morphology and capsule size were analyzed and the FLU-resistant isolates did not change significantly compared with the most sensitive isolates. Growth at 37°C was also evaluated and in both species, the resistant isolates showed a reduced growth at this temperature, indicating that FLU resistance can affect their growth. Based on the results obtained is possible suggest that FLU resistance can influence the morphology of the isolates and consequently changed the virulence profiles. The most evident results were observed for C. neoformans showing that the adaptation of isolates to antifungal selective pressure influenced the loss of virulence

Polyphasic identification and typing Trichophytum rubrum strains of clinical origin

The dermatophyte Trychophytum rubrum is the most frequent aetiological agent of dermatophytosis in humans around the world representing a major public health problem, not just for European countries but also for tropical countries where climate conditions allow major propagation for this ascomycete. For instance, in Portugal, T. rubrum was the dermatophyte most frequently isolated (83.3%) in a toenail onychomycosis geriatric population survey [1]. The identification, pathogenicity, biology, and epidemiology of T. rubrum, is of interest for both dermatologists and medical mycologists [1,2]. Currently, in many countries and clinical laboratories, the T. rubrum strains isolated from lesions are primarily identified by conventional culture-based methods, including colony morphology and slide culture only. This approach does not provide evidence of intraspecific variations with a lack of information to track infections, determine common sources of infections and recurrence or reinfection after treatment, and analyse their virulence and drug resistance [3]. The aim of this work is to use a polyphasic approach to study T. rubrum from different geographic origins in order to identify intraspecific characteristics with clinical interest. Materials and Methods About 40 European and South American T. rubrum and reference strains were used. Macro and micro morphological techniques, urease assay, dermatophyte milk agar test and hair perforation test (HPT) where combined with molecular biology techniques, such as the analysis of internal transcribed spacer (ITS) region, Trubrum specific primers for species differentiation among close related species, mating type MAT1-1 -box characterisation and DNA fingerprinting (e.g., (GACA)4). Results and conclusions Culturally T. rubrum strains showed on the plates white and cottony colonies on the obverse and blood-red pigment on the reverse. T. rubrum strains were urease negative and inhibited in dermatophyte milk agar. In the HPT, which is useful to differentiate T. rubrum from T. interdigitale, any strain was able to perforate the hair despite normal growth being observed. The analysis of ITS region confirmed all the strains as a T. rubrum species as well as the Trubrum primers generate a typical amplicon of 200 bp. The DNA fingerprinting is now explored in order to find the best approach to differentiate intraspecific variations and/or geographic differences. In conclusion, there are several techniques that can be applied to identify and characterise T. rubrum from different origins depending of the technologies available in each clinical laboratory or country.info:eu-repo/semantics/publishedVersio

Identificación por secuenciación y por espectrometría de masas MALDI-TOF y evaluación de la susceptibilidad antifúngica in vitro de aislamientos clínicos de Neoscytalidium spp.

[Extracto] Introducción: Neoscytalidium spp. son hongos dematiáceos, queratinofílicos que habitan en suelo y plantas. La taxonomía de este género ha sido problemática y constantemente revisada. Actualmente se reconocen las especies N. novaehollandiae, N. orchidacearum y N. dimidiatum, ampliamente distribuidos en regiones tropicales y subtropicales. Aunque estas especies son generalmente fitopatógenas, estudios han demostrado la implicación de N. dimidiatum en infecciones de piel, uñas, tejido celular subcutáneo y extracutáneo. La identificación de estos hongos se ha basado en la observación de las características macroscópicas y microscópicas de los cultivos in vitro, lo que puede conducir a errores. Como alternativa se puede considerar la secuenciación y la espectrometría de masas (EM) MALDI-TOF; esta última se utiliza con éxito en la identificación de bacterias y levaduras debido a la rapidez y costo-efectividad. Sin embargo, en la identificación de hongos filamentosos aún presenta inconvenientes. Por otro lado, N. dimidiatum es poco sensible a diferentes antifúngicos de uso clínico. Considerando lo anterior, el objetivo de este estudio fue identificar aislamientos clínicos de Neoscytalidium spp. por secuenciación y por EM MALDI-TOF y evaluar la susceptibilidad in vitro a tres antifúngicos. [...]info:eu-repo/semantics/publishedVersio

Design and baseline characteristics of the finerenone in reducing cardiovascular mortality and morbidity in diabetic kidney disease trial

Background: Among people with diabetes, those with kidney disease have exceptionally high rates of cardiovascular (CV) morbidity and mortality and progression of their underlying kidney disease. Finerenone is a novel, nonsteroidal, selective mineralocorticoid receptor antagonist that has shown to reduce albuminuria in type 2 diabetes (T2D) patients with chronic kidney disease (CKD) while revealing only a low risk of hyperkalemia. However, the effect of finerenone on CV and renal outcomes has not yet been investigated in long-term trials. Patients and Methods: The Finerenone in Reducing CV Mortality and Morbidity in Diabetic Kidney Disease (FIGARO-DKD) trial aims to assess the efficacy and safety of finerenone compared to placebo at reducing clinically important CV and renal outcomes in T2D patients with CKD. FIGARO-DKD is a randomized, double-blind, placebo-controlled, parallel-group, event-driven trial running in 47 countries with an expected duration of approximately 6 years. FIGARO-DKD randomized 7,437 patients with an estimated glomerular filtration rate >= 25 mL/min/1.73 m(2) and albuminuria (urinary albumin-to-creatinine ratio >= 30 to <= 5,000 mg/g). The study has at least 90% power to detect a 20% reduction in the risk of the primary outcome (overall two-sided significance level alpha = 0.05), the composite of time to first occurrence of CV death, nonfatal myocardial infarction, nonfatal stroke, or hospitalization for heart failure. Conclusions: FIGARO-DKD will determine whether an optimally treated cohort of T2D patients with CKD at high risk of CV and renal events will experience cardiorenal benefits with the addition of finerenone to their treatment regimen. Trial Registration: EudraCT number: 2015-000950-39; ClinicalTrials.gov identifier: NCT02545049

It only takes one to do many jobs: Amphotericin B as antifungal and immunomodulatory drug

Amphotericin B acts through pore formation at the cell membrane after binding to ergosterol is an accepted dogma about the action mechanism of this antifungal, and this sentence is widely found in the literature. But after 60 years of investigation, the action mechanism of Amphotericin B is not fully elucidated. Amphotericin B is a polyene substance that is one of the most effective drugs for the treatment of fungal and parasite infections. As stated above, the first mechanism of action described was pore formation after binding to the ergosterol present in the membrane. But it has also been demonstrated that AmB induces oxidative damage in the cells. Moreover, amphotericin B modulates the immune system, and this activity has been related to the protective effect of the molecule, but also to its toxicity in the host. This review tries to provide a general overview of the main aspects of this molecule, and highlight the multiple effects that this molecule has on both the fungal and host cells. © 2012 Mesa-Arango, Scorzoni and Zaragoza

Establish in Medellin (Colombia) an in-house library to identify clinically important filamentous fungi by MALDI-TOF MS

[Excerpt] Epidemiology of fungal infections has changed in an important manner in the last decades, however timely diagnosis and treatment are still a current challenge. Accurate and swift identification of the agents that cause fungal infections are paramount, since sources of infection, differences between therapeutic regimes, and in-vitro susceptibility profiles may vary amongst species and strains. The application of Matrix Assisted Laser Desorption/Ionization Time-Of-Flight (MALDI-TOF) mass spectrometry (MS) for the identification of fungal samples is currently wellestablished based on the remarkable reproducibility for the measurement of constantly expressed and highly abundant proteins, such as ribosomal proteins, that are used to generate a fingerprint profile. However, the use of this tool for filamentous fungi identification in routine clinical laboratories is still limited due to several reasons, such as, 1) the biological variations among fungal clinical isolates or 2) the lack of a spectrum of reference in commercial libraries for the identification of emerging, endemic and prevalent fungi in tropical regions. Consequently, the establishment of an in-house library that will contain spectra for prevalent fungi found in routine mycological diagnosis at a local level, and of those not included in commercial libraries, is a demand. The aim of this research was to establish a library of spectra for the identification of clinically important filamentous fungi through MALDI-TOF MS in a mycological diagnosis laboratory. [...]info:eu-repo/semantics/publishedVersio

In Vitro Activity of Essential Oils Distilled from Colombian Plants against Candidaauris and Other Candida Species with Different Antifungal Susceptibility Profiles

Multi-drug resistant species such as Candida auris are a global health threat. This scenario has highlighted the need to search for antifungal alternatives. Essential oils (EOs), or some of their major compounds, could be a source of new antifungal molecules. The aim of this study was to evaluate the in vitro activity of EOs and some terpenes against C. auris and other Candida spp. The eleven EOs evaluated were obtained by hydro-distillation from different Colombian plants and the terpenes were purchased. EO chemical compositions were obtained by gas chromatography/mass spectrometry (GC/MS). Antifungal activity was evaluated following the CLSI standard M27, 4th Edition. Cytotoxicity was tested on the HaCaT cell line and fungal growth kinetics were tested by time&ndash;kill assays. Candida spp. showed different susceptibility to antifungals and the activity of EOs and terpenes was strain-dependent. The Lippia origanoides (thymol + p-cymene) chemotype EO, thymol, carvacrol, and limonene were the most active, mainly against drug-resistant strains. The most active EOs and terpenes were also slightly cytotoxic on the HaCaT cells. The findings of this study suggest that some EOs and commercial terpenes can be a source for the development of new anti-Candida products and aid the identification of new antifungal targets or action mechanisms