Passive Acoustic Emissions Monitoring of Fluidized Bed Pellet Coating

Passive acoustic emissions were assessed for their potential as a non-invasive monitoring tool for the coating of pellets in a fluidized bed. Pharmaceutical pellets are small spherical particles that contain an active ingredient. They are film coated for the purpose of modified drug release and packed into capsules as a multiple unit dosage form. A more reliable monitoring and control method is desired to ensure the appropriate drug release profile is achieved by minimizing variations within and between coated pellets. Microphones attached to the exterior of a conical top spray fluidized bed measured acoustic emissions produced from the coating process. Statistical analysis of the signals was shown to provide information on fluidization quality and nozzle performance, while the amplitude of the acoustic emissions was shown to correspond to an increase in pellet film coat thickness. Overall, passive acoustic emissions reflected changes in process dynamics and particle interactions, indicating the ability to monitor fluidized bed pellet coating and potentially for the determination of a desired coating end-point

Engineering cell-free systems for synthetic biologists

Synthetic biology (synbio) has emerged as a transformative scientific field with immense potential to address a wide-range of global problems. A specific sub-field of synbio utilizes cellular biomolecular machinery outside of a living cell. In theory, these “cell-free” systems offer a simpler approach and unique features compared to cell-based systems for biotechnology development. However, in practice limited accessibility and poor protein synthesis capacity hinder the overall scope and application of cell-free synbio. To address these challenges, it was our goal to create new engineering tools that will help expand the overall utility of cell-free expression systems. Data presented here provides: 1) detailed methods for the in-house preparation of a cost-effective in vitro reconstituted cell-free system, 2) an in-depth proteomic analysis of the system building blocks as a tool to characterize the composition and inform optimization, and 3) an improvement to protein synthesis capacity by modifying the ribosome composition. Furthermore, a critical assessment of the regulatory landscape is provided, promoting the safe and responsible use of cell-free synbio

Filovirus receptor NPC1 contributes to species-specific patterns of ebolavirus susceptibility in bats

Biological factors that influence the host range and spillover of Ebola virus (EBOV) and other filoviruses remain enigmatic. While filoviruses infect diverse mammalian cell lines, we report that cells from African straw-colored fruit bats (Eidolon helvum) are refractory to EBOV infection. This could be explained by a single amino acid change in the filovirus receptor, NPC1, which greatly reduces the affinity of EBOV-NPC1 interaction. We found signatures of positive selection in bat NPC1 concentrated at the virus-receptor interface, with the strongest signal at the same residue that controls EBOV infection in Eidolon helvum cells. Our work identifies NPC1 as a genetic determinant of filovirus susceptibility in bats, and suggests that some NPC1 variations reflect host adaptations to reduce filovirus replication and virulence. A single viral mutation afforded escape from receptor control, revealing a pathway for compensatory viral evolution and a potential avenue for expansion of filovirus host range in nature

An Information Model For Exchanging Hydrological Rating Tables

Many hydrological data systems provide Internet access to observational and processed data in various forms, from websites to web services. This data is generally described with basic metadata, such as units, names of measured variables, spatial coordinates, and so on. This metadata is largely suitable for further analysis or ingestion into hydrological models. However, when the data has been processed through many – potentially complex – steps, more information is required to give users details of implicit assumptions, inaccuracies, or uncertainties that may have been introduced. A common example of this within hydrology is the use of ratings tables to derive variables such as river discharge. Rating tables are generally developed through field observations using a wide range of methods and are subject to constant revision to adapt to changes in the physical world. Extracting details of rating conversions used in any of the hydrological data repositories found on the Internet is currently either not possible or quite difficult. A contributing factor to this is the lack of standard representations for rating tables and their related concepts. This paper describes work by members of the joint World Meteorology Organisation/Open Geospatial Consortium’s Hydrology Domain Working Group on development of an internationally harmonized information model to describe rating tables, called WaterML2.0 part 2. The paper covers the core aspects of the information model, its implementation within web services, and a visualization client for web-based analysis of rating tables. An international data exchange experiment has been setup to further test the information model in a number of exchange scenarios. The results will be used to refine and progress WaterML2.0 part 2 towards an open standard for data exchange. The standard will lead to increased transparency for data derived using ratings, resulting in improved integration with models and other analytical processes

The role of phosphodiesterase 12 (PDE12) as a negative regulator of the innate immune response and the discovery of antiviral inhibitors

2',5'-Oligoadenylate synthetase (OAS) enzymes and RNase-L constitute a major effector arm of interferon (IFN)-mediated antiviral defense. OAS produces a unique oligonucleotide second messenger, 2',5'-oligoadenylate (2-5A), that binds and activates RNase-L. This pathway is down-regulated by virus- and host-encoded enzymes that degrade 2-5A. Phosphodiesterase 12 (PDE12) was the first cellular 2-5A- degrading enzyme to be purified and described at a molecular level. Inhibition of PDE12 may up-regulate the OAS/RNase-L pathway in response to viral infection resulting in increased resistance to a variety of viral pathogens. We generated a PDE12-null cell line, HeLaΔPDE12, using transcription activator-like effector nuclease-mediated gene inactivation. This cell line has increased 2-5A levels in response to IFN and poly(I-C), a double-stranded RNA mimic compared with the parental cell line. Moreover, HeLaΔPDE12 cells were resistant to viral pathogens, including encephalomyocarditis virus, human rhinovirus, and respiratory syncytial virus. Based on these results, we used DNA-encoded chemical library screening to identify starting points for inhibitor lead optimization. Compounds derived from this effort raise 2-5A levels and exhibit antiviral activity comparable with the effects observed with PDE12 gene inactivation. The crystal structure of PDE12 complexed with an inhibitor was solved providing insights into the structure-activity relationships of inhibitor potency and selectivity

Decreased Mitochondrial DNA Mutagenesis in Human Colorectal Cancer

Genome instability is regarded as a hallmark of cancer. Human tumors frequently carry clonally expanded mutations in their mitochondrial DNA (mtDNA), some of which may drive cancer progression and metastasis. The high prevalence of clonal mutations in tumor mtDNA has commonly led to the assumption that the mitochondrial genome in cancer is genetically unstable, yet this hypothesis has not been experimentally tested. In this study, we directly measured the frequency of non-clonal (random) de novo single base substitutions in the mtDNA of human colorectal cancers. Remarkably, tumor tissue exhibited a decreased prevalence of these mutations relative to adjacent non-tumor tissue. The difference in mutation burden was attributable to a reduction in C∶G to T∶A transitions, which are associated with oxidative damage. We demonstrate that the lower random mutation frequency in tumor tissue was also coupled with a shift in glucose metabolism from oxidative phosphorylation to anaerobic glycolysis, as compared to non-neoplastic colon. Together these findings raise the intriguing possibility that fidelity of mitochondrial genome is, in fact, increased in cancer as a result of a decrease in reactive oxygen species-mediated mtDNA damage

Isolation and Characterization of Cytotoxic, Aggregative Citrobacter freundii

Citrobacter freundii is an infrequent but established cause of diarrhea in humans. However, little is known of its genetic diversity and potential for virulence. We analyzed 26 isolates, including 12 from human diarrheal patients, 2 from human fecal samples of unknown diarrheal status, and 12 from animals, insects, and other sources. Pulsed field gel electrophoresis using XbaI allowed us to divide the 26 isolates into 20 pulse types, while multi-locus sequence typing using 7 housekeeping genes allowed us to divide the 26 isolates into 6 sequence types (STs) with the majority belonging to 4 STs. We analyzed adhesion and cytotoxicity to HEp-2 cells in these 26 strains. All were found to adhere to HEp-2 cells. One strain, CF74, which had been isolated from a goat, showed the strongest aggregative adhesion pattern. Lactate dehydrogenase (LDH) released from HEp-2 cells was evaluated as a measure of cytotoxicity, averaging 7.46%. Strain CF74 induced the highest level of LDH, 24.3%, and caused >50% cell rounding, detachment, and death. We named strain CF74 “cytotoxic and aggregative C. freundii.” Genome sequencing of CF74 revealed that it had acquired 7 genomic islands, including 2 fimbriae islands and a type VI secretion system island, all of which are potential virulence factors. Our results show that aggregative adherence and cytotoxicity play an important role in the pathogenesis of C. freundii

Staphylococcal Enterotoxins

Staphylococcus aureus (S. aureus) is a Gram positive bacterium that is carried by about one third of the general population and is responsible for common and serious diseases. These diseases include food poisoning and toxic shock syndrome, which are caused by exotoxins produced by S. aureus. Of the more than 20 Staphylococcal enterotoxins, SEA and SEB are the best characterized and are also regarded as superantigens because of their ability to bind to class II MHC molecules on antigen presenting cells and stimulate large populations of T cells that share variable regions on the β chain of the T cell receptor. The result of this massive T cell activation is a cytokine bolus leading to an acute toxic shock. These proteins are highly resistant to denaturation, which allows them to remain intact in contaminated food and trigger disease outbreaks. A recognized problem is the emergence of multi-drug resistant strains of S. aureus and these are a concern in the clinical setting as they are a common cause of antibiotic-associated diarrhea in hospitalized patients. In this review, we provide an overview of the current understanding of these proteins

Fruit-Surface Flavonoid Accumulation in Tomato Is Controlled by a SlMYB12-Regulated Transcriptional Network

The cuticle covering plants' aerial surfaces is a unique structure that plays a key role in organ development and protection against diverse stress conditions. A detailed analysis of the tomato colorless-peel y mutant was carried out in the framework of studying the outer surface of reproductive organs. The y mutant peel lacks the yellow flavonoid pigment naringenin chalcone, which has been suggested to influence the characteristics and function of the cuticular layer. Large-scale metabolic and transcript profiling revealed broad effects on both primary and secondary metabolism, related mostly to the biosynthesis of phenylpropanoids, particularly flavonoids. These were not restricted to the fruit or to a specific stage of its development and indicated that the y mutant phenotype is due to a mutation in a regulatory gene. Indeed, expression analyses specified three R2R3-MYB–type transcription factors that were significantly down-regulated in the y mutant fruit peel. One of these, SlMYB12, was mapped to the genomic region on tomato chromosome 1 previously shown to harbor the y mutation. Identification of an additional mutant allele that co-segregates with the colorless-peel trait, specific down-regulation of SlMYB12 and rescue of the y phenotype by overexpression of SlMYB12 on the mutant background, confirmed that a lesion in this regulator underlies the y phenotype. Hence, this work provides novel insight to the study of fleshy fruit cuticular structure and paves the way for the elucidation of the regulatory network that controls flavonoid accumulation in tomato fruit cuticle