Juan-Carlos Argüelles

The World Health Organization WHO considers fungal infections as a significant global risk that necessitates the development of new therapies. The arsenal of antifungals is limited, and the eukaryotic organization of fungi makes it difficult to find selective antifungal targets. In the search for potential targets for the design of new antifungals, the Stress-Activated Protein Kinase (SAPKs) pathways, and specifically, the two-component system, could be a plausible option since this upstream signaling system is absent in metazoans. SAPK pathways are involved in the response and adaptation to different environmental conditions. In pathogenic fungi, these signaling pathways are crucial for virulence, and some of them become activated in response to certain antifungals. Although further experimental evidence is required on the role of SAPKs in antifungal signaling and resistance, the possibility of impairing SAPK signaling by tagging the two-component system can be considered a useful strategy for implementing future antifungal therapies. In particular, the beneficial value of SAPK modulators combined with antifungal drugs should be a preferred line of research. In this review, we focused on the connection between the SAPK pathways and antifungal signaling in the four fungal priority pathogens, Cryptococcus neoformans, Candidozyma (formerly Candida) auris, Aspergillus fumigatus, and Candida albicans, defined by the WHO.

The WHO considers fungal infections as a significant global risk that necessitates the development of new therapies. The arsenal of antifungals is limited, and the eukaryotic organization of fungi makes it difficult to find selective antifungal targets. In the search for potential targets for the design of new antifungals, the Stress-Activated Protein Kinase (SAPKs) pathways, and specifically, the two-component system, could be a plausible option since this upstream signaling system is absent in metazoans. SAPK pathways are involved in the response and adaptation to different environmental conditions. In pathogenic fungi, these signaling pathways are crucial for virulence, and some of them become activated in response to certain antifungals. Although further experimental evidence is required on the role of SAPKs in antifungal signaling and resistance, the possibility of impairing SAPK signaling by tagging the two-component system can be considered a useful strategy for implementing future antifungal therapies. In particular, the beneficial value of SAPK modulators combined with antifungal drugs should be a preferred line of research. In this review, we focused on the connection between the SAPK pathways and antifungal signaling in the four fungal priority pathogens defined by the WHO.

Since its discovery in the middle of the XX century, research into autophagy has undergone a spectacular expansion, particularly in the early 1990s. A number of physiological processes involving autophagy have been revealed and important human pathologies have been associated with perturbations in autophagy. In 2008 the “Guidelines for the use and interpretation of assays for monitoring autophagy” was launched with the purpose of collecting in a single document all the available information to monitor autophagy, which, it was thought, might be useful for established groups and any new scientists attracted by this field. The usefulness and success of this Guidelines has led to the subsequent publication of editions every 4 years, a task in which a growing number of authors have become involved and consequently included in the list of contributors. However, this worthy initiative and closely associated metric parameters has led to important scholarly repercussions in terms of perceived merits, grants and financial support obtained, professional careers and other areas concerning scientific activity. All these aspects are carefully examined in this contribution.

Fungi are eukaryotic organisms with relatively few pathogenic members dangerous for humans, usually acting as opportunistic infections. In the last decades, several life-threatening fungal infections have risen mostly associated with the worldwide extension of chronic diseases and immunosuppression. The available antifungal therapies cannot combat this challenge because the arsenal of compounds is scarce and displays low selective action, significant adverse effects, and increasing resistance. A growing isolation of outbreaks triggered by fungal species formerly considered innocuous is being recorded. From ancient times, natural substances harvested from plants have been applied to folk medicine and some of them recently emerged as promising antifungals. The most used are briefly revised herein. Combinations of chemotherapeutic drugs with natural products to obtain more efficient and gentle treatments are also revised. Nevertheless, considerable research work is still necessary before their clinical use can be generally accepted. Many natural products have a highly complex chemical composition, with the active principles still partially unknown. Here, we survey the field underlying lights and shadows of both groups. More studies involving clinical strains are necessary, but we illustrate this matter by discussing the potential clinical applications of combined carnosic acid plus propolis formulations.

In Candida parapsilosis, homozygous disruption of the two genes encoding trehalase activity increased the susceptibility to Itraconazole compared with the isogenic parental strain. The fungicidal effect of this azole can largely be counteracted by preincubating growing cells with rotenone and the protonophore 2,4-Dinitrophenol. In turn, measurement of endogenous reactive oxygen species formation by flow cytometry confirmed that Itraconazole clearly induced an internal oxidative stress, which can be significantly abolished in rotenone-exposed cells. Analysis of the antioxidant enzymatic activities of catalase and superoxide dismutase pointed to a moderate decrease of catalase in trehalase-deficient mutant cells compared to the wild type, with an additional increase upon addition of rotenone. These enzymatic changes were imperceptible in the case of superoxide dismutase. Alternative assays with Voriconazole led to a similar profile in the results regarding cell growth and antioxidant activities. Collectively, our data suggest that the antifungal action of Itraconazole on C. parapsilosis is dependent on a functional mitochondrial activity. They also suggest that the central metabolic pathways in pathogenic fungi should be considered as preferential antifungal targets in new research.

Fungi have traditionally been considered opportunistic pathogens in primary infections caused by virulent bacteria, protozoan, or viruses. Consequently, antimycotic chemotherapy is clearly less developed in comparison to its bacterial counterpart. Currently, the three main families of antifungals (polyenes, echinocandins, and azoles) are not sufficient to control the enormous increase in life-threatening fungal infections recorded in recent decades. Natural substances harvested from plants have traditionally been utilized as a successful alternative. After a wide screening of natural agents, we have recently obtained promising results with distinct formulations of carnosic acid and propolis on the prevalent fungal pathogens Candida albicans and Cryptococcus neoformans. Here, we extended their use to the treatment against the emerging pathogenic yeast Candida glabrata, which displayed lower susceptibility in comparison to the fungi mentioned above. Taking into account the moderate antifungal activity of both natural agents, the antifungal value of these combinations has been improved through the obtention of the hydroethanolic fractions of propolis. In addition, we have demonstrated the potential clinical application of new therapeutical designs based on sequential pre-treatments with carnosic/propolis mixtures, followed by exposure to amphotericin B. This approach increased the toxic effect induced by this polyene.

Central metabolic pathways may play a major role in the virulence of pathogenic fungi. Here, we have investigated the susceptibility of a Candida parapsilosis mutant deficient in trehalase activity (atc1Δ/ntc1Δ strain) to the azolic compounds fluconazole and itraconazole. A time-course exposure to itraconazole but not fluconazole induced a significant degree of cell killing in mutant cells compared to the parental strain. Flow cytometry determinations indicated that itraconazole was able to induce a marked production of endogenous ROS together with a simultaneous increase in membrane potential, these effects being irrelevant after fluconazole addition. Furthermore, only itraconazole induced a significant synthesis of endogenous trehalose. The recorded impaired capacity of mutant cells to produce structured biofilms was further increased in the presence of both azoles, with itraconazole being more effective than fluconazole. Our results in the opportunistic pathogen yeast C. parapsilosis reinforce the study of trehalose metabolism as an attractive therapeutic target and allow extending the hypothesis that the generation of internal oxidative stress may be a component of the antifungal action exerted by the compounds currently available in medical practice.

Central metabolic pathways may play a major role in the virulence of pathogenic fungi. Here, we have investigated the susceptibility of a Candida parapsilosis mutant deficient in trehalase activity (atc1Δ/ntc1Δ strain) to the azolic compounds Fluconazole and Itraconazole. A time-course exposure to Itraconazole but not Fluconazole induced a significant degree of cell-killing in mutant cells compared to the parental strain. Flow cytometry determinations indicated that Itraconazole was able to induce a marked production of endogenous ROS together with a simultaneous increase in membrane potential, these effects being irrelevant after Fluconazole addition. Furthermore, only Itraconazole induced a significant synthesis of endogenous trehalose. The recorded impaired capacity of mutant cells to produce structured biofilms was further increased in the presence of both azoles, with Itraconazole being more effective than Fluconazole. Our results in the opportunistic pathogen yeast C. parapsilosis reinforce the study of trehalose metabolism as an attractive therapeutic target and allow extending the hypothesis that the generation of internal oxidative stress may be a component of the antifungal action exerted by the compounds currently available in medical practice.