Biofilm Formation as a Pathogenicity Factor of Medically Important Fungi

To cause disease, the infectious agent makes use of both invasiveness factors—the pathogen virulence factors—and the ability to resist and evade the host immune system. The success of the infection process is the result of a complex equation involving pathogen interaction with the host, wherein the expression of several virulence factors (and not just one or the other) will favor the establishment of the pathogen in the host. Fungal pathogens are frequently associated with biofilm formation

Candida albicans quorum-sensing molecule farnesol modulates staphyloxanthin production and activates the thiol-based oxidative-stress response in Staphylococcus aureus

Microbial species utilize secreted-signaling molecules to coordinate their behavior. Our previous investigations demonstrated a key role for the Candida albicans-secreted quorum-sensing molecule farnesol in modulating Staphylococcus aureus response to antimicrobials in mixed biofilms. In this study, we aimed to provide mechanistic insights into the impact of farnesol on S. aureus within the context of inter-species interactions. To mimic biofilm dynamics, farnesol-sensitized S. aureus cells were generated via sequential farnesol exposure. The sensitized phenotype exhibited dramatic loss of the typical pigment, which we identified as staphyloxanthin, an important virulence factor synthesized by the Crt operon in S. aureus. Additionally, farnesol exposure exerted oxidative-stress as indicated by transcriptional analysis demonstrating alterations in redox-sensors and major virulence regulators. Paradoxically, the activated stress-response conferred S. aureus with enhanced tolerance to H2O2 and phagocytic killing. Since expression of enzymes in the staphyloxanthin biosynthesis pathway was not impacted by farnesol, we generated a theoretical-binding model which indicated that farnesol may block staphyloxanthin biosynthesis via competitive-binding to the CrtM enzyme crucial for staphyloxanthin synthesis, due to high structural similarity to the CrtM substrate. Finally, mixed growth with C. albicans was found to similarly induce S. aureus depigmentation, but not during growth with a farnesol-deficient C. albicans strain. Collectively, the findings demonstrate that a fungal molecule acts as a redox-cycler eliciting a bacterial stress response via activation of the thiol-based redox system under the control of global regulators. Therefore, farnesol-induced transcriptional modulations of key regulatory networks in S. aureus may modulate the pathogenesis of C. albicans- S. aureus co-infections

The role of hydrophobicity and surface receptors at hyphae of <i>Lyophyllum</i> sp. strain Karsten in the interaction with <i>Burkholderia terrae</i> BS001:Implications for interactions in soil

The soil bacterium Burkholderia terrae strain BS001 can interact with varying soil fungi, using mechanisms that range from the utilization of carbon/energy sources such as glycerol to the ability to reach novel territories in soil via co-migration with growing fungal mycelia. Here, we investigate the intrinsic properties of the B. terrae BS001 interaction with the basidiomycetous soil fungus Lyophyllum sp. strain Karsten. In some experiments, the ascomycetous Trichoderma asperellum 302 was also used. The hyphae of Lyophyllum sp. strain Karsten were largely hydrophilic on water-containing media versus hydrophobic when aerial, as evidenced by contact angle analyses (CA). Co-migration of B. terrae strain BS001 cells with the hyphae of the two fungi occurred preferentially along the - presumably hydrophilic - soil-dwelling hyphae, whereas aerial hyphae did not allow efficient migration, due to reduced thickness of their surrounding mucous films. Moreover, the cell numbers over the length of the hyphae in soil showed an uneven distribution, i.e. the CFU numbers increased from minima at the inoculation point to maximal numbers in the middle of the extended hyphae, then decreasing towards the terminal side. Microscopic analyses of the strain BS001 associations with the Lyophyllum sp. strain Karsten hyphae in the microcosms confirmed the presence of B. terrae BS001 cells on the mucous matter that was present at the hyphal surfaces of the fungi used. Cell agglomerates were found to accumulate at defined sites on the hyphal surfaces, which were coined ‘fungal-interactive’ hot spots. Evidence was further obtained for the contention that receptors for a physical bacterium-fungus interaction occur at the Lyophyllum sp. strain Karsten hyphal surface, in which the specific glycosphingolipid ceramide mono hexoside (CMH) plays an important role. Thus, bacterial adherence may be mediated by heterogeneously-distributed fungal-specific receptors, implying the CMH moieties. This study sheds light on the physical aspects of the B. terrae BS001 – L. sp strain Karsten interaction, highlighting heterogeneity along the hyphae with respect to hydrophobicity and the presence of potential anchoring sites

Proanthocyanidins polymeric tannin from Stryphnodendron adstringens are active against Candida albicans biofilms

Abstract\ud \ud Background\ud Biofilm formation is important in Candida albicans pathogenesis and constitutes a mechanism of antifungal resistance. Thus, we evaluated the effect of proanthocyanidins polymer-rich fractions from Stryphnodendron adstringens (fraction F2 and subfraction F2.4) against C. albicans biofilms.\ud \ud \ud Methods\ud Firstly, the antifungal activity of F2 and F2.4 against planktonic cells of Candida albicans (ATCC 10231) was determined using broth microdilution method. Anti-biofilm effect of F2 and F2.4 was evaluated during biofilm formation or on mature biofilm of C. albicans and compared with standard antifungals amphotericin B and fluconazole. Metabolic activity of sessile and dispersion cells from biofilms after antifungal treatments were measured using a tetrazolium reduction assay and the biofilm total biomass was quantified by crystal violet-based assay. Morphological alterations after treatments were observed using scanning electron microscopy.\ud \ud \ud Results\ud The anti-biofilm effect of F2 and F2.4 were comparable to standard antifungals (amphotericin B and fluconazole). F2 and F2.4 treatments reduced biofilm metabolic activity (in sessile and in dispersion cells) during biofilm formation, and in mature biofilms, unlike fluconazole, which only prevents the biofilm formation. Treatments with F2, F2.4 or fluconazole reduced biofilm biomass during biofilm formation, but not in mature biofilm. Amphotericin B presented higher inhibitory effect on biofilm formation and on mature biofilm of C. albicans. F2 and F2.4 treatments led to the appearance of dumbbell-shaped blastoconidia and of blastoconidia clusters in biofilms.\ud \ud \ud Conclusion\ud Proanthocyanidins polymer-rich fractions from S. adstringens successfully inhibited C. albicans planktonic growth and biofilm development, and they represent a potential new agent for the treatment of biofilm-associated candidiasis.This work was supported by Fundação de Amparo à Pesquisa\ud do Estado de São Paulo (FAPESP - Processo no. 2013/11232-0), Fundação\ud Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro\ud (FAPERJ), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior\ud (CAPES), and Conselho Nacional de Desenvolvimento e Pesquisa (CNPq).\ud RLFL and TVMV are fellows of the CNPq

Identification of two potential inhibitors of Sporothrix brasiliensis and Sporothrix schenckii in the Pathogen Box collection.

Sporotrichosis is a neglected endemic mycosis with a high incidence in Latin America, mainly in Brazil. Sporothrix schenckii is the most frequent species in Latin America, whereas Sporothrix brasiliensis is the predominant species observed in Brazil and is associated with both human and animal sporotrichosis. Sporotrichosis treatment remains restricted to a few options, itraconazole being the first choice for human and animal therapy. In this work, we screened the molecular library Pathogen Box (Medicines for Malaria Venture [MMV], Switzerland) in search of compounds with anti-Sporothrix activity. Our initial screen of the 400 compounds identified five compounds that inhibited more than 80% of S. brasiliensis and S. schenkii growth. Among those, three compounds (MMV675968, MMV102872, and MMV002817 (known as iodoquinol)) not previously described as antifungals or agrochemicals, were selected for further evaluation. MMV102872 and iodoquinol showed the most promising combination of antifungal activity (lower inhibitory concentration) and fungal selectivity (lower cytotoxicity in LLC-MK2 cells). Scanning electron microscopy and flow cytometry analyses revealed that MMV102872 and iodoquinol induced changes in cell morphology, membrane integrity, and the presence of neutral lipids, impairing fungal survival. Our results indicate that MMV102872 and iodoquinol are promising molecules for use as scaffolds for the development of new antifungal agents

Candida albicans quorum-sensing molecule farnesol modulates staphyloxanthin production and activates the thiol-based oxidative-stress response in Staphylococcus aureus

Microbial species utilize secreted-signaling molecules to coordinate their behavior. Our previous investigations demonstrated a key role for the Candida albicans-secreted quorum-sensing molecule farnesol in modulating Staphylococcus aureus response to antimicrobials in mixed biofilms. In this study, we aimed to provide mechanistic insights into the impact of farnesol on S. aureus within the context of inter-species interactions. To mimic biofilm dynamics, farnesol-sensitized S. aureus cells were generated via sequential farnesol exposure. The sensitized phenotype exhibited dramatic loss of the typical pigment, which we identified as staphyloxanthin, an important virulence factor synthesized by the Crt operon in S. aureus. Additionally, farnesol exposure exerted oxidative-stress as indicated by transcriptional analysis demonstrating alterations in redox-sensors and major virulence regulators. Paradoxically, the activated stress-response conferred S. aureus with enhanced tolerance to H2O2 and phagocytic killing. Since expression of enzymes in the staphyloxanthin biosynthesis pathway was not impacted by farnesol, we generated a theoretical-binding model which indicated that farnesol may block staphyloxanthin biosynthesis via competitive-binding to the CrtM enzyme crucial for staphyloxanthin synthesis, due to high structural similarity to the CrtM substrate. Finally, mixed growth with C. albicans was found to similarly induce S. aureus depigmentation, but not during growth with a farnesol-deficient C. albicans strain. Collectively, the findings demonstrate that a fungal molecule acts as a redox-cycler eliciting a bacterial stress response via activation of the thiol-based redox system under the control of global regulators. Therefore, farnesol-induced transcriptional modulations of key regulatory networks in S. aureus may modulate the pathogenesis of C. albicans- S. aureus co-infections