Novel, synergistic antifungal combinations that target translation fidelity

There is an unmet need for new antifungal or fungicide treatments, as resistance to existing treatments grows. Combination treatments help to combat resistance. Here we develop a novel, effective target for combination antifungal therapy. Different aminoglycoside antibiotics combined with different sulphate-transport inhibitors produced strong, synergistic growth-inhibition of several fungi. Combinations decreased the respective MICs by ≥8 fold. Synergy was suppressed in yeast mutants resistant to effects of sulphate-mimetics (like chromate or molybdate) on sulphate transport. By different mechanisms, aminoglycosides and inhibition of sulphate transport cause errors in mRNA translation. The mistranslation rate was stimulated up to 10-fold when the agents were used in combination, consistent with this being the mode of synergistic action. A range of undesirable fungi were susceptible to synergistic inhibition by the combinations, including the human pathogens Candida albicans, C. glabrata and Cryptococcus neoformans, the food spoilage organism Zygosaccharomyces bailii and the phytopathogens Rhizoctonia solani and Zymoseptoria tritici. There was some specificity as certain fungi were unaffected. There was no synergy against bacterial or mammalian cells. The results indicate that translation fidelity is a promising new target for combinatorial treatment of undesirable fungi, the combinations requiring substantially decreased doses of active components compared to each agent alone

Insights into Candida tropicalis nosocomial infections and virulence factors

Candida tropicalis is considered the first or the second non-Candida albicans Candida (NCAC) species most frequently isolated from candidosis, mainly in patients admitted in intensive care units (ICUs), especially with cancer, requiring prolonged catheterization, or receiving broad-spectrum antibiotics. The proportion of candiduria and candidemia caused by C. tropicalis varies widely with geographical area and patient group. Actually, in certain countries, C. tropicalis is more prevalent, even compared with C. albicans or other NCAC species. Although prophylactic treatments with fluconazole cause a decrease in the frequency of candidosis caused by C. tropicalis, it is increasingly showing a moderate level of fluconazole resistance. The propensity of C. tropicalis for dissemination and the high mortality associated with its infections might be strongly related to the potential of virulence factors exhibited by this species, such as adhesion to different host surfaces, biofilm formation, infection and dissemination, and enzymes secretion. Therefore, the aim of this review is to outline the present knowledge on all the above-mentioned C. tropicalis virulence traits.The authors acknowledge Coordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES), Brazil, for supporting Melyssa Negri (BEX 4642/06-6) and Fundacao para a Ciencia e Tecnologia (FCT), Portugal, for supporting Sonia Silva (SFRH/BPD/71076/2010), and European Community fund FEDER, trough Program COMPETE under the Project FCOMP-01-0124-FEDER-007025 (PTDC/AMB/68393/2006) is gratefully acknowledged

Candida glabrata : a review of its features and resistance

Candida species belong to the normal microbiota of the oral cavity and gastrointestinal and vaginal tracts, and are responsible for several clinical manifestations, from mucocutaneous overgrowth to bloodstream infections. Once believed to be non-pathogenic, Candida glabrata was rapidly blamable for many human diseases. Year after year, these pathological circumstances are more recurrent and problematic to treat, especially when patients reveal any level of immunosuppression. These difficulties arise from the capacity of C. glabrata to form biofilms and also from its high resistance to traditional antifungal therapies. Thus, this review intends to present an excerpt of the biology, epidemiology, and pathology of C. glabrata, and detail an approach to its resistance mechanisms based on studies carried out up to the present.The authors are grateful to strategic project PTDC/SAU-MIC/119069/2010 for the financial support to the research center and for Celia F. Rodrigues' grant

Identification of Candida glabrata genes involved in pH modulation and modification of the phagosomal environment in macrophages

notes: PMCID: PMC4006850types: Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov'tCandida glabrata currently ranks as the second most frequent cause of invasive candidiasis. Our previous work has shown that C. glabrata is adapted to intracellular survival in macrophages and replicates within non-acidified late endosomal-stage phagosomes. In contrast, heat killed yeasts are found in acidified matured phagosomes. In the present study, we aimed at elucidating the processes leading to inhibition of phagosome acidification and maturation. We show that phagosomes containing viable C. glabrata cells do not fuse with pre-labeled lysosomes and possess low phagosomal hydrolase activity. Inhibition of acidification occurs independent of macrophage type (human/murine), differentiation (M1-/M2-type) or activation status (vitamin D3 stimulation). We observed no differential activation of macrophage MAPK or NFκB signaling cascades downstream of pattern recognition receptors after internalization of viable compared to heat killed yeasts, but Syk activation decayed faster in macrophages containing viable yeasts. Thus, delivery of viable yeasts to non-matured phagosomes is likely not triggered by initial recognition events via MAPK or NFκB signaling, but Syk activation may be involved. Although V-ATPase is abundant in C. glabrata phagosomes, the influence of this proton pump on intracellular survival is low since blocking V-ATPase activity with bafilomycin A1 has no influence on fungal viability. Active pH modulation is one possible fungal strategy to change phagosome pH. In fact, C. glabrata is able to alkalinize its extracellular environment, when growing on amino acids as the sole carbon source in vitro. By screening a C. glabrata mutant library we identified genes important for environmental alkalinization that were further tested for their impact on phagosome pH. We found that the lack of fungal mannosyltransferases resulted in severely reduced alkalinization in vitro and in the delivery of C. glabrata to acidified phagosomes. Therefore, protein mannosylation may play a key role in alterations of phagosomal properties caused by C. glabrata.Deutsche ForschungsgemeinschaftNational Institutes for HealthWellcome TrustBBSR

Voriconazole efficacy against Candida glabrata and Candida krusei: preclinical data using a validated in vitro pharmacokinetic/pharmacodynamic model

Background: Voriconazole exhibits in vitro activity against Candida glabrata and Candida krusei (EUCAST/CLSI epidemiological cut-off values 1/0.25 and 1/0.5 mg/L, respectively). Yet, EUCAST found insufficient evidence to set breakpoints for these species. We explored voriconazole pharmacodynamics (PD) in an in vitro dynamic model simulating human pharmacokinetics (PK). Methods: Four C. glabrata and three C. krusei isolates (voriconazole EUCAST and CLSI MICs of 0.03–2 mg/L) were tested in the PK/PD model simulating voriconazole exposures (t1=2 6 h q12h dosing for 3 days). PK/PD breakpoints were determined calculating the PTA for exposure indices fAUC0–24/MIC associated with half-maximal activity (EI50) using Monte Carlo simulation analysis. Results: Fungal load increased from 3.60±0.35 to 8.41±0.24 log10 cfu/mL in the drug-free control, with a maximum effect of 1 log10 kill of C. glabrata and C. krusei isolates with MICs of 0.06 and 0.25 mg/L, respectively, at high drug exposures. The 72 h log10 cfu/mL change versus fAUC0–24/MIC relationship followed a sigmoid curve for C. glabrata (R2 =0.85–0.87) and C. krusei (R2 =0.56–0.76) with EI50 of 49 (32–76) and 52 (33–78) fAUC/MIC for EUCAST and 55 (31–96) and 80 (42–152) fAUC/MIC for CLSI, respectively. The PTAs for C. glabrata and C. kr

A multicentre study to optimize echinocandin susceptibility testing of Aspergillus species with the EUCAST methodology and a broth microdilution colorimetric method

BACKGROUND: The determination of the minimal effective concentration (MEC) of echinocandins against Aspergillus species is subjective, time consuming and has been associated with very major errors. METHODS: The MECs/MICs of 40 WT [10 each of Aspergillus fumigatus species complex (SC), Aspergillus flavus SC, Aspergillus terreus SC and Aspergillus niger SC] and 4 non-WT A. fumigatus isolates were determined with EUCAST E.Def 9.3.1 read microscopically, macroscopically, spectrophotometrically and colorimetrically in three centres. The optimal conditions for spectrophotometric (single- versus multi-point readings) and colorimetric (XTT/menadione concentration and stability, incubation time) methods were evaluated in preliminary studies using different cut-offs for the determination of macroscopic, spectrophotometric and colorimetric MIC endpoints compared with the microscopically determined MEC. Inter-centre and inter-method essential (within one 2-fold dilution) agreement (EA) and categorical agreement (CA) were determined. RESULTS: Both macroscopic and spectr

Synthesis and Structure-Activity Relationships of Novel Non-Steroidal CYP17A1 Inhibitors as Potential Prostate Cancer Agents.

Twenty new compounds, targeting CYP17A1, were synthesized, based on our previous work on a benzimidazole scaffold, and their biological activity evaluated. Inhibition of CYP17A1 is an important modality in the treatment of prostate cancer, which remains the most abundant cancer type in men. The biological assessment included CYP17A1 hydroxylase and lyase inhibition, CYP3A4 and P450 oxidoreductase (POR) inhibition, as well as antiproliferative activity in PC3 prostate cancer cells. The most potent compounds were selected for further analyses including in silico modeling. This combined effort resulted in a compound (comp 2, IC50 1.2 µM, in CYP17A1) with a potency comparable to abiraterone and selectivity towards the other targets tested. In addition, the data provided an understanding of the structure-activity relationship of this novel non-steroidal compound class

Invasive pulmonary aspergillosis 10 years post bone marrow transplantation: a case report

Abstract Introduction Invasive pulmonary aspergillosis is a leading cause of mortality and morbidity in bone marrow transplant recipients. Establishing the diagnosis remains a challenge for clinicians working in acute care setting. However, prompt diagnosis and treatment can lead to favourable outcomes Case presentation We report a case of invasive aspergillosis occurring in a 39-year-old Caucasian female 10 years after an allogeneic haematopoietic bone marrow transplant, and 5 years after stopping all immunosuppression. Possible risk factors include bronchiolitis obliterans and exposure to building dust (for example, handling her husband's dusty overalls). There are no similar case reports in the literature at this time. Conclusion High clinical suspicion, especially in the setting of failure to respond to broad-spectrum antibiotics, should alert clinicians to the possibility of invasive pulmonary aspergillosis, which, in this case, responded to antifungal therapy.</p