The disruption of GDP-fucose de novo biosynthesis suggests the presence of a novel fucose-containing glycoconjugate in <i>Plasmodium</i> asexual blood stages

Glycosylation is an important posttranslational protein modification in all eukaryotes. Besides glycosylphosphatidylinositol (GPI) anchors and N-glycosylation, O-fucosylation has been recently reported in key sporozoite proteins of the malaria parasite. Previous analyses showed the presence of GDP-fucose (GDP-Fuc), the precursor for all fucosylation reactions, in the blood stages of Plasmodium falciparum. The GDP-Fuc de novo pathway, which requires the action of GDP-mannose 4,6-dehydratase (GMD) and GDP-L-fucose synthase (FS), is conserved in the parasite genome, but the importance of fucose metabolism for the parasite is unknown. To functionally characterize the pathway we generated a PfGMD mutant and analyzed its phenotype. Although the labelling by the fucose-binding Ulex europaeus agglutinin I (UEA-I) was completely abrogated, GDP-Fuc was still detected in the mutant. This unexpected result suggests the presence of an alternative mechanism for maintaining GDP-Fuc in the parasite. Furthermore, PfGMD null mutant exhibited normal growth and invasion rates, revealing that the GDP-Fuc de novo metabolic pathway is not essential for the development in culture of the malaria parasite during the asexual blood stages. Nonetheless, the function of this metabolic route and the GDP-Fuc pool that is generated during this stage may be important for gametocytogenesis and sporogonic development in the mosquito

Linking full-text grey literature to underlying research and post-publication data: An Enhanced Publications Project 2011-2012

1. The Project This project seeks to circumvent the data vs. documents camp in the grey literature community by way of a middle ground provided through enhanced publications. Enhanced publications allow for a fuller understanding of the process in which data and information are used and applied in the generation of knowledge. The enhanced publication of grey literature precludes the idea of a random selection of data and information, and instead focuses on the human intervention in data-rich environments. The definition of an enhanced publication is borrowed from the DRIVER-II project, “a publication that is enhanced with three categories of information: research data, extra materials, and post-publication data”. Enhanced publications combine textual resources i.e. documents intended to be read by human beings, which contain an interpretation or analysis of primary data. Enhanced publications inherently contribute to the review process of grey literature as well as the replication of research and improved visibility of research results in the scholarly communication chain. 2. Design of the Questionnaire and Author Survey The population of the survey was selected from among the 286 authors and co-authors in the International Conference Series on Grey Literature. It was decided that only first authors would receive the questionnaire, which narrowed the potential population of the survey to 162 authors of which only 95 were actually sent the online questionnaire. The reason the other 67 first authors were not included in the final survey population was due to a number of factors such as no current email address, retired, deceased, etc. The 95 authors were sent a personalized email with a standardized text inviting them to participate in the survey by completing the online questionnaire. The survey was carried out using the freeware ‘Survey Monkey’ and the questionnaire contained 10 items, three of which were open-ended. Subheadings were also inserted in the questionnaire set off by quotation marks. These subheadings preceded each odd numbered question and were deemed relevant in achieving informed responses. The final results are based on the response of 50 of the 95 survey recipients, which amounts to roughly a 53% response rate

Volumetric 3D printing for rapid production of medicines

3D printing (3DP), or additive manufacturing, has been actively investigated as one of the enabling technologies for the impending era of personalized medicines. However, existing 3DP technologies do not afford the speeds required for on-demand production of medicines in fast-paced clinical settings. Volumetric printing is a novel 3DP technology that offers rapid printing speed and overcomes the geometric and surface quality limitations of layer-based vat photopolymerization techniques. Unlike previous vat photopolymerization 3DP technologies, volumetric printing cures the entire desired 3D geometry simultaneously by exploiting the threshold behavior in the photopolymerization process that arise due to oxygen-induced polymerization inhibition. In this work, for the first time, a volumetric printer was used to fabricate drug-loaded 3D printed tablets (Printlets™) within seconds. Six resin formulations were evaluated using this printer, each composed of poly(ethylene glycol) diacrylate (PEGDA) as the crosslinking monomer, lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) as the photoinitiator, and paracetamol as the model drug. Water or PEG300 were included as diluents in varying concentrations to facilitate drug release. Paracetamol-loaded Printlets were successfully fabricated within 17 s. Drug release rates could be tuned by altering the monomer-to-diluent ratio of the photosensitive resin, with a lower ratio releasing drug faster. The present work confirms the suitability of volumetric 3DP for printing drug products in a matter of seconds. Upon further optimization, this novel technology can enable rapid, on-demand fabrication of medicines and medical devices

Linking full-text grey literature to underlying research and post-publication data: An Enhanced Publications Project 2011-2012

1. The Project This project seeks to circumvent the data vs. documents camp in the grey literature community by way of a middle ground provided through enhanced publications. Enhanced publications allow for a fuller understanding of the process in which data and information are used and applied in the generation of knowledge. The enhanced publication of grey literature precludes the idea of a random selection of data and information, and instead focuses on the human intervention in data-rich environments. The definition of an enhanced publication is borrowed from the DRIVER-II project, “a publication that is enhanced with three categories of information: research data, extra materials, and post-publication data”. Enhanced publications combine textual resources i.e. documents intended to be read by human beings, which contain an interpretation or analysis of primary data. Enhanced publications inherently contribute to the review process of grey literature as well as the replication of research and improved visibility of research results in the scholarly communication chain. 2. Design of the Questionnaire and Author Survey The population of the survey was selected from among the 286 authors and co-authors in the International Conference Series on Grey Literature. It was decided that only first authors would receive the questionnaire, which narrowed the potential population of the survey to 162 authors of which only 95 were actually sent the online questionnaire. The reason the other 67 first authors were not included in the final survey population was due to a number of factors such as no current email address, retired, deceased, etc. The 95 authors were sent a personalized email with a standardized text inviting them to participate in the survey by completing the online questionnaire. The survey was carried out using the freeware ‘Survey Monkey’ and the questionnaire contained 10 items, three of which were open-ended. Subheadings were also inserted in the questionnaire set off by quotation marks. These subheadings preceded each odd numbered question and were deemed relevant in achieving informed responses. The final results are based on the response of 50 of the 95 survey recipients, which amounts to roughly a 53% response rate

Inhibition of the master regulator of Listeria monocytogenes virulence enables bacterial clearance from spacious replication vacuoles in infected macrophages

A hallmark of Listeria (L.) monocytogenes pathogenesis is bacterial escape from maturing entry vacuoles, which is required for rapid bacterial replication in the host cell cytoplasm and cell-to-cell spread. The bacterial transcriptional activator PrfA controls expression of key virulence factors that enable exploitation of this intracellular niche. The transcriptional activity of PrfA within infected host cells is controlled by allosteric coactivation. Inhibitory occupation of the coactivator site has been shown to impair PrfA functions, but consequences of PrfA inhibition for L. monocytogenes infection and pathogenesis are unknown. Here we report the crystal structure of PrfA with a small molecule inhibitor occupying the coactivator site at 2.0 Å resolution. Using molecular imaging and infection studies in macrophages, we demonstrate that PrfA inhibition prevents the vacuolar escape of L. monocytogenes and enables extensive bacterial replication inside spacious vacuoles. In contrast to previously described spacious Listeria-containing vacuoles, which have been implicated in supporting chronic infection, PrfA inhibition facilitated progressive clearance of intracellular L. monocytogenes from spacious vacuoles through lysosomal degradation. Thus, inhibitory occupation of the PrfA coactivator site facilitates formation of a transient intravacuolar L. monocytogenes replication niche that licenses macrophages to effectively eliminate intracellular bacteria. Our findings encourage further exploration of PrfA as a potential target for antimicrobials and highlight that intra-vacuolar residence of L. monocytogenes in macrophages is not inevitably tied to bacterial persistence.Originally included in thesis in manuscript form. </p