Global Analysis of the Evolution and Mechanism of Echinocandin Resistance in Candida glabrata

The evolution of drug resistance has a profound impact on human health. Candida glabrata is a leading human fungal pathogen that can rapidly evolve resistance to echinocandins, which target cell wall biosynthesis and are front-line therapeutics for Candida infections. Here, we provide the first global analysis of mutations accompanying the evolution of fungal drug resistance in a human host utilizing a series of C. glabrata isolates that evolved echinocandin resistance in a patient treated with the echinocandin caspofungin for recurring bloodstream candidemia. Whole genome sequencing identified a mutation in the drug target, FKS2, accompanying a major resistance increase, and 8 additional non-synonymous mutations. The FKS2-T1987C mutation was sufficient for echinocandin resistance, and associated with a fitness cost that was mitigated with further evolution, observed in vitro and in a murine model of systemic candidemia. A CDC6-A511G(K171E) mutation acquired before FKS2-T1987C(S663P), conferred a small resistance increase. Elevated dosage of CDC55, which acquired a C463T(P155S) mutation after FKS2-T1987C(S663P), ameliorated fitness. To discover strategies to abrogate echinocandin resistance, we focused on the molecular chaperone Hsp90 and downstream effector calcineurin. Genetic or pharmacological compromise of Hsp90 or calcineurin function reduced basal tolerance and resistance. Hsp90 and calcineurin were required for caspofungin-dependent FKS2 induction, providing a mechanism governing echinocandin resistance. A mitochondrial respiration-defective petite mutant in the series revealed that the petite phenotype does not confer echinocandin resistance, but renders strains refractory to synergy between echinocandins and Hsp90 or calcineurin inhibitors. The kidneys of mice infected with the petite mutant were sterile, while those infected with the HSP90-repressible strain had reduced fungal burden. We provide the first global view of mutations accompanying the evolution of fungal drug resistance in a human host, implicate the premier compensatory mutation mitigating the cost of echinocandin resistance, and suggest a new mechanism of echinocandin resistance with broad therapeutic potential

Who uses emergency departments inappropriately and when - a national cross-sectional study using a monitoring data system

Contains fulltext : 126219.pdf (publisher's version ) (Open Access)BACKGROUND: Increasing pressures on emergency departments (ED) are straining services and creating inefficiencies in service delivery worldwide. A potentially avoidable pressure is inappropriate attendances (IA); typically low urgency, self-referred patients better managed by other services. This study examines demographics and temporal trends associated with IA to help inform measures to address them. METHODS: Using a national ED dataset, a cross-sectional examination of ED attendances in England from April 2011 to March 2012 (n = 15,056,095) was conducted. IA were defined as patients who were self-referred; were not attending a follow-up; received no investigation and either no treatment or 'guidance/advice only'; and were discharged with either no follow-up or follow-up with primary care. Small, nationally representative areas were used to assign each attendance to a residential measure of deprivation. Multivariate analysis was used to predict relationships between IA, demographics (age, gender, deprivation) and temporal factors (day, month, hour, bank holiday, Christmas period). RESULTS: Overall, 11.7% of attendances were categorized as inappropriate. IA peaked in early childhood (adjusted odds ratio (AOR) = 1.53 for both one and two year olds), and was elevated throughout late-teens and young adulthood, with odds reducing steadily from age 27 (reference category, age 40). Both IA and appropriate attendances (AA) were most frequent in the most deprived populations. However, relative to AA, those living in the least deprived areas had the highest odds of IA (AOR = 0.89 in most deprived quintile). Odds of IA were also higher for males (AOR = 0.95 in females). Both AA and IA were highest on Mondays, whilst weekends, bank holidays and the period between 8 am and 4 pm saw more IA relative to AA. CONCLUSIONS: Prevention of IA would be best targeted at parents of young children and at older youths/young adults, and during weekends and bank holidays. Service provision focusing on access to primary care and EDs serving the most deprived communities would have the most benefit. Improvements in coverage and data quality of the national ED dataset, and the addition of an appropriateness field, would make this dataset an effective monitoring tool to evaluate interventions addressing this issue

Discovery of a new azole resistance mechanism in Aspergillus fumigatus through whole genome sequencing and sexual crossing.

Contains fulltext : 110932.pdf (publisher's version ) (Open Access

In vitro and in vivo antifungal activities and mechanism of heteropolytungstates against Candida species

Abstract The antifungal activities of heteropolytungstates, α-1,2,3-K6H[SiW9V3O40] (SiW-3), K13[Ce(SiW11O39)2]·17H2O (SiW-5), K13[Eu(SiW11O39)2]·25H2O (SiW-10), K6PV3W9O40 (PW-6), α-K4PVW11O40 (PW-8), were screened in 29 Candida albicans, 8 Candida glabrata, 3 Candida krusei, 2 Candida parapsilosis, 1 Candida tropicalis, and 1 Cryptococcus neoformans strains using the CLSI M27-A3 method. SiW-5 had the highest efficacy with a minimum inhibitory concentration (MIC) values of <0.2–10.2 μM in vitro. The antifungal mechanism, acute toxicity and in vivo antifungal activity of SiW-5 were then evaluated in C. albicans. The results showed that SiW-5 damaged the fungal cell membrane, reduce the ergosterol content and its main mode of action was through inhibition of ergosterol biosynthesis. Real-time PCR showed that ERG1, ERG7, ERG11 and ERG28 were all significantly upregulated by SiW-5. An acute toxicity study showed the 50% lethal dose (LD50) of SiW-5 for ICR mice was 1651.5 mg/kg. And in vivo antifungal studies demonstrated that SiW-5 reduced both the morbidity and fungal burden of mice infected with C. albicans. This study demonstrates that SiW-5 is a potential antifungal candidate against the Candida species