Marilou G. Nicolas

Convergent Evolution of Calcineurin Pathway Roles in Thermotolerance and Virulence in Candida glabrata.

Candida glabrata is an emerging human fungal pathogen that is frequently drug tolerant, resulting in difficulties in treatment and a higher mortality in immunocompromised patients. The calcium-activated protein phosphatase calcineurin plays critical roles in controlling drug tolerance, hyphal growth, and virulence in diverse fungal pathogens via distinct mechanisms involving survival in serum or growth at host temperature (37° and higher). Here, we comprehensively studied the calcineurin signaling cascade in C. glabrata and found novel and uncharacterized functions of calcineurin and its downstream target Crz1 in governing thermotolerance, intracellular architecture, and pathogenesis in murine ocular, urinary tract, and systemic infections. This represents a second independent origin of a role for calcineurin in thermotolerant growth of a major human fungal pathogen, distinct from that which arose independently in Cryptococcus neoformans. Calcineurin also promotes survival of C. glabrata in serum via mechanisms distinct from C. albicans and thereby enables establishment of tissue colonization in a murine systemic infection model. To understand calcineurin signaling in detail, we performed global transcript profiling analysis and identified calcineurin- and Crz1-dependent genes in C. glabrata involved in cell wall biosynthesis, heat shock responses, and calcineurin function. Regulators of calcineurin (RCN) are a novel family of calcineurin modifiers, and two members of this family were identified in C. glabrata: Rcn1 and Rcn2. Our studies demonstrate that Rcn2 expression is controlled by calcineurin and Crz1 to function as a feedback inhibitor of calcineurin in a circuit required for calcium tolerance in C. glabrata. In contrast, the calcineurin regulator Rcn1 activates calcineurin signaling. Interestingly, neither Rcn1 nor Rcn2 is required for virulence in a murine systemic infection model. Taken together, our findings show that calcineurin signaling plays critical roles in thermotolerance and virulence, and that Rcn1 and Rcn2 have opposing functions in controlling calcineurin signaling in C. glabrata.

Calcineurin Is Required for Pseudohyphal Growth, Virulence, and Drug Resistance in Candida lusitaniae

Candida lusitaniae is an emerging fungal pathogen that infects immunocompromised patients including HIV/AIDS, cancer, and neonatal pediatric patients. Though less prevalent than other Candida species, C. lusitaniae is unique in its ability to develop resistance to amphotericin B. We investigated the role of the calcium-activated protein phosphatase calcineurin in several virulence attributes of C. lusitaniae including pseudohyphal growth, serum survival, and growth at 37°C. We found that calcineurin and Crz1, a C. albicans Crz1 homolog acting as a downstream target of calcineurin, are required for C. lusitaniae pseudohyphal growth, a process for which the underlying mechanism remains largely unknown in C. lusitaniae but hyphal growth is fundamental to C. albicans virulence. We demonstrate that calcineurin is required for cell wall integrity, ER stress response, optimal growth in serum, virulence in a murine systemic infection model, and antifungal drug tolerance in C. lusitaniae. To further examine the potential of targeting the calcineurin signaling cascade for antifungal drug development, we examined the activity of a calcineurin inhibitor FK506 in combination with caspofungin against echinocandin resistant C. lusitaniae clinical isolates. Broth microdilution and drug disk diffusion assays demonstrate that FK506 has synergistic fungicidal activity with caspofungin against echinocandin resistant isolates. Our findings reveal that pseudohyphal growth is controlled by the calcineurin signaling cascade, and highlight the potential use of calcineurin inhibitors and caspofungin for emerging drug-resistant C. lusitaniae infections.

Modelling the Coverage of Dipterocarp Trees in Central Visayas, Philippines

With the rapid decline of dipterocarp coverage in the Philippines, efforts in restoration would benefit from having a suitability map that would indicate the suitable areas where different dipterocarp species could thrive. Another use of these maps would be for assessing whether particular locations should be considered as Protected Areas in which exploitation would be limited, if not completely prohibited. We obtained data from the Department of Environment and Natural Resources of the Philippines, which consists of dipterocarp-related information gathered over localities in the countrys Central Visayas region. Six climate data parameters were then chosen and obtained from the Philippine Atmospheric Geophysical and Astronomical Services Administration. We employed a maximum entropy-based niche modelling approach to generate a suitability map. The model that we produced obtained an area under the curve (AUC) score of 0.955 with minimum temperature being the variable with the highest contribution. The map produced indicates that dipterocarp trees are more likely to appear in the lower regions of Central Visayas.

In vitro and #945;-glucosidase inhibition and antioxidant activities of partially purified Antidesma bunius fruit and Gynura nepalensis leaf extracts

Diabetes mellitus is a metabolic disorder characterized by increased blood glucose levels. Current treatments involve the use of sulfonylureas, α-glucosidase inhibitors, and other synthetic drugs. The study demonstrated the α-glucosidase inhibition and antioxidant properties of partially purified ethanolic extracts of Antidesma bunius fruits and Gynura nepalensis leaves, as possible herbal drug candidates. After ethanol extraction, the extracts were fractionated using normal phase liquid column chromatography, with elution solvents ethyl acetate, methanol and water. Fractionation resulted in five fractions for A. bunius (A1, A2, A3, A4 and A5) and seven fractions for G. nepalensis (G1, G2, G3, G4, G5, G6 and G7). Fraction G1 showed the highest α-glucosidase inhibition activity (90.61±8.05%) and possibly acted via a mixed mode of inhibition. For the antioxidant activities, Fraction A1 exhibited highest radical scavenging activity via DPPH assay (97.39±2.48%), Fraction G7 exhibited highest iron (II)-chelating activity (95.85±1.46%) and Fraction G6 exhibited highest ferric-ion reducing activity via FRAP assay (272.60 μg/mL FeSO4 equivalents). Phytochemical screening revealed that flavonoids and tannins were common among all fractions. The results demonstrated the potential of these plants as antidiabetic herbal treatments; however, further studies and to isolate the active component(s), structure elucidation and toxicity assays need be done first.