Comparative pathogenicity of Scedosporium species in murine model of systemic scedosporiosis.

Systemic scedosporiosis is a devastating emerging fungal infection caused by several species of the genus Scedosporium in immunocompetent and immunocompromised individuals. In this study, we compared the virulence of different Scedosporium species in a murine model of systemic scedosporiosis by survival assays, fungal burden and histopathological analysis. We found that mice mortality was species-dependent, S. apiospermum, S. aurantiacum and S. dehoogii were the most virulent species. We also observed the dissemination and invasion of Scedosporium species to the brain, spleen and kidney by colony count and histopathological analysis at different times of infection. Particularly, the brain was the tissue most susceptible to invasion during systemic scedosporiosis. This study shows the virulence and pathophysiology of different Scedosporium species and will be useful in facilitating control and prevention strategies for systemic scedosporiosis.

Microwave-Assisted Synthesis of Aminophosphonic Derivatives and Their Antifungal Evaluation against Lomentospora prolificans.

Lomentospora prolificans is a pathogenic and multidrug-resistant fungus that can infect both immunocompetent and immunocompromised patients, with mortality rates up to 87%. The World Health Organization (WHO) included this fungal species in its first list of 19 priority fungal pathogens, which focused on fungal pathogens that can cause invasive acute and subacute systemic fungal infections. Therefore, there is a growing interest in finding new therapeutic alternatives. In this work, the synthesis of twelve α-aminophosphonates by the microwave-assisted Kabachnik-Fields reaction and twelve α-aminophosphonic acids by a monohydrolysis reaction is reported. All compounds were evaluated by the agar diffusion method as a preliminary screening in comparison with voriconazole, showing inhibition halos for compounds 7, 11, 13, 22 and 27. The five active compounds in the preliminary tests were evaluated against five strains of L. prolificans following protocol M38-A2 from CLSI. The results showed that these compounds exhibit antifungal activity in the concentration range of 900->900 µg/mL. Cytotoxicity against healthy COS-7 cells was also evaluated by the MTT assay, and it was shown that compound 22 was the least cytotoxic, with a viability of 67.91%, comparable to the viability exhibited by voriconazole (68.55%). Docking studies showed that the possible mechanism of action of the active compounds could be through the inhibition of the enzyme lanosterol-14-alpha-demethylase in an allosteric hydrophobic cavity.

Recent Antifungal Pipeline Developments against Candida auris: A Systematic Review.

The alarming spread and impact of multidrug-resistant Candida auris infections alongside the limited therapeutic options have prompted the development of new antifungals. These promising agents are currently in different stages of development, offering novel dosing regimens and mechanisms of action. A systematic search in MEDLINE, EMBASE, Web of Science, and Scopus up to 27 June 2022 was conducted to find relevant articles reporting data of in vitro activity and in vivo efficacy of investigational antifungals against C. auris. These included new additions to existing antifungal classes (rezafungin and opelconazole), first-in-class drugs such as ibrexafungerp, manogepix/fosmanogepix, olorofim and tetrazoles (quilseconazole, oteseconazole and VT-1598), as well as other innovative agents like ATI-2307, MGCD290 and VL-2397. From 592 articles retrieved in the primary search, 27 met the eligibility criteria. The most studied agent was manogepix/fosmanogepix (overall MIC90: 0.03 mg/L), followed by ibrexafungerp (overall MIC90: 1 mg/L) and rezafungin (overall MIC mode: 0.25 mg/L), while VT-1598 and ATI-2307 were the least explored drugs against C. auris. All these compounds demonstrated significant improvements in survival and reduction in tissue fungal burden on neutropenic animal models of candidemia due to C. auris. Continual efforts towards the discovery of new treatments against this multidrug-resistant fungus are essential.

Novel α-Aminophosphonates and α-Aminophosphonic Acids: Synthesis, Molecular Docking and Evaluation of Antifungal Activity against Scedosporium Species.

The Scedosporium genus is an emerging pathogen with worldwide prevalence and high mortality rates that gives multidrug resistance to antifungals; therefore, pharmacological alternatives must be sought for the treatment of diseases caused by this fungus. In the present project, six new α-aminophosphates were synthesized by the Kabachnik-Fields multicomponent reaction by vortex agitation, and six new monohydrolyzed α-aminophosphonic acids were synthesized by an alkaline hydrolysis reaction. Antifungal activity was evaluated using the agar diffusion method as an initial screening to determine the most active compound compared to voriconazole; then it was evaluated against 23 strains of the genus Scedosporium following the M38-A2 protocol from CLSI (activity range: 648.76-700 µg/mL). Results showed that compound 5f exhibited the highest antifungal activity according to the agar diffusion method (≤1 mg/mL). Cytotoxicity against healthy COS-7 cells was also evaluated by the MTT assay and it was shown that compound 5f exhibits a lower toxicity in comparison to voriconazole at the same concentration (1000 µM). A docking study was conducted afterwards, showing that the possible mechanism of action of the compound is through the inhibition of allosteric 14-α-demethylase. Taking these results as a basis, 5f is presented as a compound with attractive properties for further studies.

Managing raw materials of vegetable origin increases fungal indoor concentration in food companies.

Fungi in indoor environments is a known cause of disease and food spoilage. However, there is currently no legislation or normativity stablishing limits for fungal densities in correlation with these. Moreover, there is little knowledge of the diversity of fungi in indoor environments for industrial areas and in food-related companies in particular, a study has never been performed to evaluate the concentration and diversity of fungi in this type of places. We evaluated the fungal density of 20 food companies. We sampled 100 L of air onto rose bengal-malt extract-agar plates, using an Air Test Omega® sampler. After incubation, CFUs were counted and identified. Penicillium, Cladosporium and Aspergillus were the most commonly isolated genus, with Penicillium being the only genus to be present in every area sampled. Neither the companies' location nor their room temperature influenced the fungal densities significantly, however, companies using vegetable raw materials had a significantly greater concentration of fungi than the rest of the companies. While all concentrations were within previously suggested levels from a health-related point of view, more information is needed that correlates fungal concentration with food spoilage in order to suggest a range of concentrations focused for food companies' product preservation.

Phenotypical characterization and molecular identification of clinical isolates of Candida tropicalis / Caracterización fenotípica e identificación molecular de aislamientos clínicos de Candida tropicalis

Background. Candida tropicalis is an increasingly important human pathogen which usually affects neutropenic oncology patients with common hematogenous seeding to peripheral organs and high mortality rates. Candida pathogenicity is facilitated by several virulence attributes, including secretion of hydrolytic enzymes; however, little is known regarding the C. tropicalis ability to secrete them and their role in the disease. Aims. To confirm by molecular means the identification of 187 clinical isolates (127 from blood, 52 from urine, and 8 from diverse clinical origins) phenotypically identified as C. tropicalis, and to investigate their in vitro aspartyl proteinase, phospholipase, esterase, hemolysin, DNase and coagulase activities. Methods. The molecular confirmation was performed by ITS sequencing, and the enzymatic determinations were conducted using plate assays with specific substrates, with the exception of coagulase, which was determined by the classical tube test. Results. The majority of the strains exhibited a very strong or strong activity of aspartyl proteinase, phospholipase and esterase. A 4.7% of the bloodstream isolates were hemolysin producers, and all were negative for the coagulase and DNase assays. Conclusions. Very strong activities of aspartyl proteinase, phospholipase and esterase profiles were detected, and a statistical association between phospholipase production and blood and urine isolates was found (AU)

Antecedentes. Candida tropicalis es un patógeno del ser humano cada vez más importante que afecta especialmente a pacientes oncológicos neutropénicos, en los cuales es frecuente la diseminación hematógena del microorganismo a órganos periféricos, lo que conlleva elevadas tasas de mortalidad. La patogenicidad de Candida es facilitada por diversos factores de virulencia, incluyendo la secreción de enzimas hidrolíticas; sin embargo, poco se sabe respecto a la habilidad de C. tropicalis para su secreción, así como el papel que desempeña en la enfermedad. Objetivos. Confirmar por un método molecular la identidad de 187 aislamientos clínicos (127 de sangre, 52 de orina y 8 de orígenes diversos) fenotípicamente identificados como C. tropicalis y estudiar la actividad in vitro de las enzimas proteinasa aspártica, fosfolipasa, esterasa, hemolisina, DNasa y coagulasa. Métodos. La confirmación molecular se llevó a cabo mediante secuenciación del ITS y las determinaciones enzimáticas se llevaron a cabo mediante ensayos en placa con sustratos específicos, a excepción de la coagulasa, que se determinó mediante la clásica prueba en tubo. Resultados. La mayoría de los aislamientos analizados mostraron un perfil de actividad muy fuerte o fuerte de proteinasa aspártica, fosfolipasa y esterasa. El 4,7% de las cepas sanguíneas fue productora de hemolisinas y todas fueron negativas para coagulasa y DNasa. Conclusiones. Se detectaron perfiles con una actividad proteinasa aspártica, fosfolipasa y esterasa muy fuerte entre los aislamientos clínicos analizados, así como también se encontró asociación estadística entre la producción de fosfolipasa y aquellos aislamientos obtenidos de sangre y orina (AU)

Biofilm formation and antifungal susceptibility of Trichosporon asahii isolates from Mexican patients / Formación de biopelículas y sensibilidad a los antifúngicos de aislamientos de Trichosporon asahii procedentes de pacientes mexicanos

Background. Trichosporon asahii is a yeast-like fungus that has recently gained importance as a cause of opportunistic systemic infections. The pathogenicity and virulence factors of T. asahii remain largely unknown. Because of the association between invasive infections and the use of catheters and related devices, the ability of the microorganism to adhere and form biofilms may play an important role in the pathogenicity during a trichosporonosis. Aims. The aim of this study is to identify an association between biofilm formation by T. asahii isolates and their genotype and/or clinical source. Methods. The biofilm production of 49 T. asahii strains isolated from Mexican patients was measured using the crystal violet stain method, and a comparison made with different adhesion phase incubation times. Antifungal susceptibility testing was performed using a modified CLSI protocol coupled with the quantification of the viable cells with the XTT reduction method. Results. All the T. asahii isolates assayed were able to produce biofilm in vitro, with an intraspecific variability being observed. Overall, increased biofilm production was found when extending the adhesion phase incubation time from 2 to 4h. No association could be established between the biofilm-producing phenotype and either the genotype or clinical source. Higher antifungal resistance to amphotericin B and fluconazole was linked to increased biofilm production by T. asahii. Conclusions. All clinical isolates tested were able to produce biofilm. No association could be established between biofilm formation and genotype or clinical source (AU)

Antecedentes. Trichosporon asahii es un hongo levaduriforme de gran importancia en los últimos años por causar infecciones sistémicas oportunistas. Existe escasa información sobre su patogenicidad y factores de virulencia. Debido a la asociación entre las infecciones invasivas y el uso de catéteres y otros dispositivos médicos, la capacidad de este microorganismo para adherirse y formar biopelículas puede incrementar su patogenicidad durante la tricosporonosis. Objetivos. En el presente trabajo se estudia la asociación entre la formación de biopelícula por diferentes aislamientos de T. asahii y su genotipo y/o lugar de aislamiento. Métodos. Se cuantificó la producción de biopelícula en 49 aislamientos de T. asahii aislados de pacientes mexicanos mediante el método de tinción con cristal violeta, y se compararon diferentes tiempos de incubación para la fase de adhesión. Además se realizaron pruebas de sensibilidad a los antifúngicos mediante el protocolo del CLSI, con modificaciones, acoplado a la cuantificación de células viables con el método de reducción del XTT. Resultados. Todos los aislamientos de T. asahii analizados produjeron biopelícula; se observó variabilidad intraespecifica en dicha producción. En general se observó un incremento en la producción de biopelícula al aumentar el tiempo de incubación de la fase de adhesión de 2 a 4h. No se logró establecer una asociación entre la producción de biopelícula y el genotipo u origen clínico de los diferentes aislamientos de T. asahii. El incremento en la resistencia a la anfotericina B y el fluconazol resultó proporcional al incremento en la producción de biopelícula. Conclusiones. Todos los aislamientos clínicos evaluados fueron capaces de producir biopelícula. No se logró establecer una asociación entre la formación de biopelícula y el genotipo u origen clínico del aislamiento (AU)

Biofilm formation and antifungal susceptibility of Trichosporon asahii isolates from Mexican patients.

BACKGROUND: Trichosporon asahii is a yeast-like fungus that has recently gained importance as a cause of opportunistic systemic infections. The pathogenicity and virulence factors of T. asahii remain largely unknown. Because of the association between invasive infections and the use of catheters and related devices, the ability of the microorganism to adhere and form biofilms may play an important role in the pathogenicity during a trichosporonosis. AIMS: The aim of this study is to identify an association between biofilm formation by T. asahii isolates and their genotype and/or clinical source. METHODS: The biofilm production of 49 T. asahii strains isolated from Mexican patients was measured using the crystal violet stain method, and a comparison made with different adhesion phase incubation times. Antifungal susceptibility testing was performed using a modified CLSI protocol coupled with the quantification of the viable cells with the XTT reduction method. RESULTS: All the T. asahii isolates assayed were able to produce biofilm in vitro, with an intraspecific variability being observed. Overall, increased biofilm production was found when extending the adhesion phase incubation time from 2 to 4h. No association could be established between the biofilm-producing phenotype and either the genotype or clinical source. Higher antifungal resistance to amphotericin B and fluconazole was linked to increased biofilm production by T. asahii. CONCLUSIONS: All clinical isolates tested were able to produce biofilm. No association could be established between biofilm formation and genotype or clinical source.

Phenotypical characterization and molecular identification of clinical isolates of Candida tropicalis.

BACKGROUND: Candida tropicalis is an increasingly important human pathogen which usually affects neutropenic oncology patients with common hematogenous seeding to peripheral organs and high mortality rates. Candida pathogenicity is facilitated by several virulence attributes, including secretion of hydrolytic enzymes; however, little is known regarding the C. tropicalis ability to secrete them and their role in the disease. AIMS: To confirm by molecular means the identification of 187 clinical isolates (127 from blood, 52 from urine, and 8 from diverse clinical origins) phenotypically identified as C. tropicalis, and to investigate their in vitro aspartyl proteinase, phospholipase, esterase, hemolysin, DNase and coagulase activities. METHODS: The molecular confirmation was performed by ITS sequencing, and the enzymatic determinations were conducted using plate assays with specific substrates, with the exception of coagulase, which was determined by the classical tube test. RESULTS: The majority of the strains exhibited a very strong or strong activity of aspartyl proteinase, phospholipase and esterase. A 4.7% of the bloodstream isolates were hemolysin producers, and all were negative for the coagulase and DNase assays. CONCLUSIONS: Very strong activities of aspartyl proteinase, phospholipase and esterase profiles were detected, and a statistical association between phospholipase production and blood and urine isolates was found.

Comparative Pathogenicity of Lomentospora prolificans (Scedosporium prolificans) Isolates from Mexican Patients.

We identified 11 Lomentospora prolificans isolates recovered from Mexican patients using phenotypic and molecular characteristics. The identification of isolates was assessed by internal transcribed spacer (ITS rDNA) sequencing. In vitro susceptibility to amphotericin B, fluconazole, voriconazole, posaconazole, caspofungin, anidulafungin and micafungin was determined according to Clinical and Laboratory Standards Institute (CLSI) procedures. Three isolates (07-2239, 11-2242 and 04-2673) were used to induce systemic infection in immunocompetent ICR mice. Survival and tissue burden studies were used as markers of pathogenicity. All of the strains were resistant to every antifungal tested with MIC's for AmB (8->8 µg/ml), VRC (16->16 µg/ml), PSC (16->16 µg/ml), FLC (64->64 µg/ml) and echinocandins with MICs ≥8 µg/ml. One hundred, ninety and sixty percent of the infected mice with the strains 07-2239, 11-2242 and 04-2673 died during the study, respectively. Regarding tissue burden, the highest fungal load of the infected mice was detected in brain followed by spleen and kidney, regardless of the strain.

Molecular identification and in vitro antifungal susceptibility of Scedosporium complex isolates from high-human-activity sites in Mexico.

The genus Scedosporium is a complex of ubiquitous moulds associated with a wide spectrum of clinical entities, with high mortality principally in immunocompromised hosts. Ecology of these microorganisms has been studied performing isolations from environmental sources, showing a preference for human-impacted environments. This study aimed to evaluate the presence and antifungal susceptibility of Scedosporium complex species in soil samples collected in high-human-activity sites of Mexico. A total of 97 soil samples from 25 Mexican states were collected. Identifications were performed by microscopic morphology and confirmed by sequencing of the rDNA (internal transcribed spacer [ITS], D1/D2) and ß-tubulin partial loci. Antifungal susceptibility testing was performed according to the Clinical and Laboratory Standards Institute (CLSI) protocols. Soil samples of urban gardens and industrial parks constituted the best sources for isolation of Scedosporium complex species. S. apiospermum sensu stricto was the most prevalent species (69%), followed by S. boydii (16%). Voriconazole (minimal inhibitory concentration [MIC] geometric mean ≤2.08 µg/mL), followed by posaconazole (MIC geometric mean ≤2.64 µg/mL), exhibited excellent in vitro activity for most species. Amphotericin B and fluconazole demonstrated limited antifungal activity, and all of the strains were resistant to echinocandins. This is the first report in Mexico of environmental distribution and antifungal in vitro susceptibility of these emergent pathogens.

Efficacy of ravuconazole in a murine model of vaginitis by Candida albicans / Eficacia del ravuconazol en un modelo murino de vaginitis por Candida albicans

Background. The incidence of vulvovaginal candidiasis, a common infection among healthy women primarily caused by the yeast Candida albicans, has increased significantly in recent years. Aims. The purpose of this study was to compare the efficacy of ravuconazole (RVC) and fluconazole (FLC) in the treatment of experimental C. albicans vaginitis. Methods. Forty isolates of C. albicans were screened for their in vitro susceptibility to RVC and FLC. A strain of C. albicans that was resistant to FLC (minimum inhibitory concentration [MIC] of >64 μg/ml) was selected for the in vivo study. Treatment regimens for the murine vaginal infection model were (1) 1, 5, 10, and 20 mg/kg RVC once daily, (2) 20 mg/kg RVC twice daily, (3) 20 mg/kg FLC once daily, and (4) 20 mg/kg FLC twice daily. Results. The geometric means of the MIC values at 48 h for all isolates tested were 0.05 and 0.5 μg/ml for RVC and FLC, respectively. Regimens of either RVC or FLC at 20 mg/kg twice daily were more effective to reduce the load of FLC-resistant C. albicans than single dose administration. Conclusions. Complete eradication of C. albicans from the vagina was not observed with RVC or FLC treatment in the animal model, although RVC treatment showed a lower fungal concentration 14 days after drug administration (AU)

Antecedentes. En los últimos años, ha aumentado sustancialmente la incidencia de candidiasis vulvovaginal, una infección frecuente entre mujeres sanas, causada sobre todo por la levadura Candida albicans. Objetivos. El objetivo del presente estudio fue comparar la eficacia del ravuconazol (RVC) y del fluconazol (FLC) en el tratamiento de la vaginitis experimental inducida por C. albicans. Métodos. Se examinó la sensibilidad in vitro de 40 aislamientos de C. albicans frente a RVC y FLC. Para el estudio in vivo se seleccionó una cepa de C. albicans que fue resistente a FLC (concentración inhibitoria mínima [CIM] >64 μg/ml). Las pautas de tratamiento para el modelo murino de infección vaginal fueron 1) 1, 5, 10 y 20 mg/kg de RVC una vez al día, 2) 20 mg/kg de RVC dos veces al día, 3) 20 mg/de FLC una vez al día, y 4) 20 mg/kg de FLC dos veces al día. Resultados. Para todos los aislamientos las medias geométricas de los valores de la CIM a las 48 h fueron de 0,05 y 0,5 μg/ml para RVC y FLC, respectivamente. Las pautas de 20 mg/kg de RVC o FLC dos veces al día fueron más eficaces para reducir la carga infectiva de C. albicans resistente a FLC que las administradas una vez al día. Conclusiones. En el modelo animal no se eliminó completamente C. albicans del tracto vaginal estéril mediante tratamiento con RVC o FLC. Sin embargo, el tratamiento con RVC derivó en concentraciones fúngicas más bajas 14 días después de su administración (AU)

Efficacy of ravuconazole in a murine model of vaginitis by Candida albicans.

BACKGROUND: The incidence of vulvovaginal candidiasis, a common infection among healthy women primarily caused by the yeast Candida albicans, has increased significantly in recent years. AIMS: The purpose of this study was to compare the efficacy of ravuconazole (RVC) and fluconazole (FLC) in the treatment of experimental C. albicans vaginitis. METHODS: Forty isolates of C. albicans were screened for their in vitro susceptibility to RVC and FLC. A strain of C. albicans that was resistant to FLC (minimum inhibitory concentration [MIC] of >64 µg/ml) was selected for the in vivo study. Treatment regimens for the murine vaginal infection model were (1) 1, 5, 10, and 20 mg/kg RVC once daily, (2) 20 mg/kg RVC twice daily, (3) 20 mg/kg FLC once daily, and (4) 20 mg/kg FLC twice daily. RESULTS: The geometric means of the MIC values at 48 h for all isolates tested were 0.05 and 0.5 µg/ml for RVC and FLC, respectively. Regimens of either RVC or FLC at 20 mg/kg twice daily were more effective to reduce the load of FLC-resistant C. albicans than single dose administration. CONCLUSIONS: Complete eradication of C. albicans from the vagina was not observed with RVC or FLC treatment in the animal model, although RVC treatment showed a lower fungal concentration 14 days after drug administration.

Killer behavior within the Candida parapsilosis complex.

A group of 29 isolates of Candida parapsilosis sensu stricto, 29 of Candida orthopsilosis, and 4 of Candida metapsilosis were assayed for the presence of killer activity using Saccharomyces cerevisiae ATCC 26609 as a sensitive strain. All C. metapsilosis isolates showed killer activity at 25 °C while strains of C. parapsilosis sensu stricto or C. orthopsilosis did not exhibit this activity. Sensitivity to killer toxins was evaluated using a set of previously reported killer strains of clinical origin. Only 11 isolates of the C. parapsilosis complex were inhibited by at least one killer isolate without resulting in any clear pattern, except for C. parapsilosis sensu stricto ATCC 22019, which was inhibited by every killer strain with the exception of C. parapsilosis and Candida utilis. The lack of sensitivity to killer activity among isolates of the genus Candida suggests that their toxins belong to the same killer type. Differentiation of species within the C. parapsilosis complex using the killer system may be feasible if a more taxonomically diverse panel of killer strains is employed.

Susceptibility of Dermatophytes to Thiabendazole Using CLSI Broth Macrodilution.

Objective. To evaluate in vitro antifungal activity of thiabendazole against strains of dermatophytes using a reference method for filamentous fungi. Materials and Methods. Dermatophytes' susceptibility to thiabendazole (TBZ) and fluconazole (FCZ) was evaluated using macrodilution method of protocol M38-A2 of the Clinical and Laboratory Standards Institute (CLSI). Results. MIC ranges of TBZ for all strains were narrower and/or smaller than those of FCZ. TBZ showed a significantly greater potency than FCZ (P = 0.05) against all isolates. Discussion. Although there have been approaches to evaluate the antifungal activity of TBZ in human mycoses, no tests had been made with a standardized protocol. Susceptibility data resulted from this study shows that although TBZ is not a particularly strong inhibitor of dermatophytes, it displays a stable and constant effect against all isolates tested. Conclusion. Results show that TBZ is more effective against strains of dermatophytes than FCZ. We acknowledge the antifungal effect of TBZ against dermatophyte isolates.