Development of Trypanosoma cruzi in vitro assays to identify compounds suitable for progression in Chagas’ disease drug discovery

Chagas' disease is responsible for significant mortality and morbidity in Latin America. Current treatments display variable efficacy and have adverse side effects, hence more effective, better tolerated drugs are needed. However, recent efforts have proved unsuccessful with failure of the ergosterol biosynthesis inhibitor posaconazole in phase II clinical trials despite promising in vitro and in vivo studies. The lack of translation between laboratory experiments and clinical outcome is a major issue for further drug discovery efforts. Our goal was to identify cell-based assays that could differentiate current nitro-aromatic drugs nifurtimox and benznidazole from posaconazole. Using a panel of T. cruzi strains including the six major lineages (TcI-VI), we found that strain PAH179 (TcV) was markedly less susceptible to posaconazole in vitro. Determination of parasite doubling and cycling times as well as EdU labelling experiments all indicate that this lack of sensitivity is due to the slow doubling and cycling time of strain PAH179. This is in accordance with ergosterol biosynthesis inhibition by posaconazole leading to critically low ergosterol levels only after multiple rounds of division, and is further supported by the lack of effect of posaconazole on the non-replicative trypomastigote form. A washout experiment with prolonged posaconazole treatment showed that, even for more rapidly replicating strains, this compound cannot clear all parasites, indicative of a heterogeneous parasite population in vitro and potentially the presence of quiescent parasites. Benznidazole in contrast was able to kill all parasites. The work presented here shows clear differentiation between the nitro-aromatic drugs and posaconazole in several assays, and suggests that in vitro there may be clinically relevant heterogeneity in the parasite population that can be revealed in long-term washout experiments. Based on these findings we have adjusted our in vitro screening cascade so that only the most promising compounds are progressed to in vivo experiments

Cells of yeasts adhered in corn grains and the storage perspective for use as probiotic

Yeasts were applied to corn grains, containing 16 or 20% moisture, at concentrations of 1 and 2%. The cellular viability was assayed at 0, 15, 30, 90 and 110 days. The cellular viability did not differ statistically among the treatments up to 30 days of storage, with the median viability of 89.10%. The average viability found at 90 days (72.20%) was lower than in the first three storage periods. After 110 days, the average viability was 61.14%. In conclusion, since yeast cells were viable up to 110 days in storage on corn grains, these can be used as a vehicle for the application of yeast as a probiotic.<br>O comportamento da viabilidade celular de leveduras é essencial quando se almeja utilizá-la como um probiótico e o veículo que assegure esta propriedade até o momento do consumo é particularmente. Sobre grãos de milho, contendo 16 ou 20% de umidade, foram aplicadas leveduras desidratadas vivas, nas concentrações de 1 e de 2%. A viabilidade celular foi avaliada nos períodos de 0, 15, 30, 90 e 110 dias de armazenamento. A viabilidade celular nas leveduras não diferiu estatisticamente entre os tratamentos aos 30 dias em armazenamento, com a viabilidade média de 89,10%. A viabilidade média encontrada aos 90 dias de 72,20% foi menor do que aquelas nos três primeiros períodos de armazenamento. Após 110 dias de armazenamento foi de 61,14%. Conclue-se que leveduras apresentam viabilidade celular até 110 dias em armazenamento sobre milho e que estes podem ser utilizados como um veículo para a aplicação da levedura como um probiótico

Development and Prospects of Dedicated Tracers for the Molecular Imaging of Bacterial Infections

Bacterial infections have always been, and still are, a major global healthcare problem. For accurate treatment it is of utmost importance that the location(s), severity, type of bacteria, and therapeutic response can be accurately staged. Similar to the recent successes in oncology, tracers specific for molecular imaging of the disease may help advance the patient management. Chemical design and bacterial targeting mechanisms are the basis for the specificity of such tracers. The aim of this review is to provide a comprehensive overview of the molecular imaging tracers developed for optical and nuclear identification of bacteria and bacterial infections. Hereby we envision that such tracers can be used to diagnose infections and aid their clinical management. From these compounds we have set-out to identify promising targeting mechanisms and select the most promising candidates for further development.The Netherlands Organisation for Scientific Research (NWO; STW BGT 11272).http://pubs.acs.org/bchb201