Anthropometric specification of child crash dummy pelves through statistical analysis of skeletal geometry

The pelves of the child dummies of the widely used Hybrid-III family are based on minimal data from children. Because an accurate pelvis design is critical for realistic assessments of belt restraint interactions, an analysis of medical imaging data was conducted to develop guidance for improved pelvis design. Three-dimensional polygonal meshes of the bony pelvis were generated from computed tomography (CT) data from 81 children from ages 5 to 11. After aligning to a uniform anatomical coordinate system, the meshes were resampled to create a quadrilateral mesh with 12,960 vertices for each pelvis. A principal components analysis was conducted with the mesh vertex coordinates and the locations of 31 landmarks. Over 99% of the variance in size and shape was accounted for by the first 40 components. A three-dimensional model representing the target for a new dummy pelvis was developed using bispinous breadth as the predictor variable. To obtain the appropriate geometry for the six-year-old (6YO) and 10YO Hybrid-III dummies, a regression analysis was conducted using a large sample of child anthropometry data from a previous study to obtain a target dimension for bispinous breadth, using the design stature for each dummy as input. A separate regression analysis was conducted to predict principal component scores from bispinous breadth. Reconstructing a pelvis model from the principal components scores predicted for the target bispinous breadth values yielded a three-dimensional design target for the each dummy. The new pelvis target is similar in overall size to the current pelvis components, but the location of the anterior-superior iliac spine is markedly lower, which has important implications for belt interaction.     

Chlamydia pneumoniae Hides inside Apoptotic Neutrophils to Silently Infect and Propagate in Macrophages

Background

Intracellular pathogens have developed elaborate strategies for silent infection of preferred host cells. Chlamydia pneumoniae is a common pathogen in acute infections of the respiratory tract (e.g. pneumonia) and associated with chronic lung sequelae in adults and children. Within the lung, alveolar macrophages and polymorph nuclear neutrophils (PMN) are the first line of defense against bacteria, but also preferred host phagocytes of chlamydiae.

Methodology/Principal Findings

We could show that C. pneumoniae easily infect and hide inside neutrophil granulocytes until these cells become apoptotic and are subsequently taken up by macrophages. C. pneumoniae infection of macrophages via apoptotic PMN results in enhanced replicative activity of chlamydiae when compared to direct infection of macrophages, which results in persistence of the pathogen. Inhibition of the apoptotic recognition of C. pneumoniae infected PMN using PS- masking Annexin A5 significantly lowered the transmission of chlamydial infection to macrophages. Transfer of apoptotic C. pneumoniae infected PMN to macrophages resulted in an increased TGF-ß production, whereas direct infection of macrophages with chlamydiae was characterized by an enhanced TNF-α response.

Conclusions/Significance

Taken together, our data suggest that C. pneumoniae uses neutrophil granulocytes to be silently taken up by long-lived macrophages, which allows for efficient propagation and immune protection within the human host.     

Tissue distribution of L-fucokinase in rodents

L-fucose (fucose) is a monosaccharide normally present in mammals and is unique in being the only levorotatory sugar that can be synthesized and utilized by mammals. The metabolism of fucose is incompletely understood, but fucose can be synthesized de novo or salvaged. The utilization of fucose in the salvage pathway begins with phosphorylation by fucokinase. As part of an investigation of fucose metabolism in normal and disease states, we began an investigation of this enzyme. In this report, we present the tissue distribution of the enzyme in rat and mouse. The highest amount of activity was present in brain of both species. Some activity was found in all tissues examined (liver, kidney, heart, lung, spleen, brain, muscle, thymus, white adipose, testes, eye, aorta, small intestine, and submaxillary gland). Very low levels were found in small intestine. Varying levels in the tissues seems most likely to be the result of varying amounts of fucokinase protein as no difference in the Km values of crude enzyme could be shown. Protein-bound fucose levels were determined using the L-cysteine-phenol-sulfuric acid (CPS) assay. There is not a good correlation between fucokinase activity and protein-bound fucose, suggesting some tissues are more active in synthesis of fucose than others.     

Immune Evasion,Stress Resistance,and Efficient Nutrient Acquisition Are Crucial for Intracellular Survival of Candida glabrata within Macrophages

Candida glabrata is both a human fungal commensal and an opportunistic pathogen which can withstand activities of the immune system. For example, C. glabrata can survive phagocytosis and replicates within macrophages. However, the mechanisms underlying intracellular survival remain unclear. In this work, we used a functional genomic approach to identify C. glabrata determinants necessary for survival within human monocyte-derived macrophages by screening a set of 433 deletion mutants. We identified 23 genes which are required to resist killing by macrophages. Based on homologies to Saccharomyces cerevisiae orthologs, these genes are putatively involved in cell wall biosynthesis, calcium homeostasis, nutritional and stress response, protein glycosylation, or iron homeostasis. Mutants were further characterized using a series of in vitro assays to elucidate the genes'' functions in survival. We investigated different parameters of C. glabrata-phagocyte interactions: uptake by macrophages, replication within macrophages, phagosomal pH, and recognition of mutant cells by macrophages as indicated by production of reactive oxygen species and tumor necrosis factor alpha (TNF-α). We further studied the cell surface integrity of mutant cells, their ability to grow under nutrient-limited conditions, and their susceptibility to stress conditions mirroring the harsh environment inside a phagosome. Additionally, resistance to killing by neutrophils was analyzed. Our data support the view that immune evasion is a key aspect of C. glabrata virulence and that increased immune recognition causes increased antifungal activities by macrophages. Furthermore, stress resistance and efficient nutrient acquisition, in particular, iron uptake, are crucial for intraphagosomal survival of C. glabrata.     

Systematic Phenotyping of a Large-Scale Candida glabrata Deletion Collection Reveals Novel Antifungal Tolerance Genes

The opportunistic fungal pathogen Candida glabrata is a frequent cause of candidiasis, causing infections ranging from superficial to life-threatening disseminated disease. The inherent tolerance of C. glabrata to azole drugs makes this pathogen a serious clinical threat. To identify novel genes implicated in antifungal drug tolerance, we have constructed a large-scale C. glabrata deletion library consisting of 619 unique, individually bar-coded mutant strains, each lacking one specific gene, all together representing almost 12% of the genome. Functional analysis of this library in a series of phenotypic and fitness assays identified numerous genes required for growth of C. glabrata under normal or specific stress conditions, as well as a number of novel genes involved in tolerance to clinically important antifungal drugs such as azoles and echinocandins. We identified 38 deletion strains displaying strongly increased susceptibility to caspofungin, 28 of which encoding proteins that have not previously been linked to echinocandin tolerance. Our results demonstrate the potential of the C. glabrata mutant collection as a valuable resource in functional genomics studies of this important fungal pathogen of humans, and to facilitate the identification of putative novel antifungal drug target and virulence genes.     

Design and Characterization of a 52K SNP Chip for Goats

The success of Genome Wide Association Studies in the discovery of sequence variation linked to complex traits in humans has increased interest in high throughput SNP genotyping assays in livestock species. Primary goals are QTL detection and genomic selection. The purpose here was design of a 50–60,000 SNP chip for goats. The success of a moderate density SNP assay depends on reliable bioinformatic SNP detection procedures, the technological success rate of the SNP design, even spacing of SNPs on the genome and selection of Minor Allele Frequencies (MAF) suitable to use in diverse breeds. Through the federation of three SNP discovery projects consolidated as the International Goat Genome Consortium, we have identified approximately twelve million high quality SNP variants in the goat genome stored in a database together with their biological and technical characteristics. These SNPs were identified within and between six breeds (meat, milk and mixed): Alpine, Boer, Creole, Katjang, Saanen and Savanna, comprising a total of 97 animals. Whole genome and Reduced Representation Library sequences were aligned on >10 kb scaffolds of the de novo goat genome assembly. The 60,000 selected SNPs, evenly spaced on the goat genome, were submitted for oligo manufacturing (Illumina, Inc) and published in dbSNP along with flanking sequences and map position on goat assemblies (i.e. scaffolds and pseudo-chromosomes), sheep genome V2 and cattle UMD3.1 assembly. Ten breeds were then used to validate the SNP content and 52,295 loci could be successfully genotyped and used to generate a final cluster file. The combined strategy of using mainly whole genome Next Generation Sequencing and mapping on a contig genome assembly, complemented with Illumina design tools proved to be efficient in producing this GoatSNP50 chip. Advances in use of molecular markers are expected to accelerate goat genomic studies in coming years.     

Selection Signatures in Worldwide Sheep Populations

The diversity of populations in domestic species offers great opportunities to study genome response to selection. The recently published Sheep HapMap dataset is a great example of characterization of the world wide genetic diversity in sheep. In this study, we re-analyzed the Sheep HapMap dataset to identify selection signatures in worldwide sheep populations. Compared to previous analyses, we made use of statistical methods that (i) take account of the hierarchical structure of sheep populations, (ii) make use of linkage disequilibrium information and (iii) focus specifically on either recent or older selection signatures. We show that this allows pinpointing several new selection signatures in the sheep genome and distinguishing those related to modern breeding objectives and to earlier post-domestication constraints. The newly identified regions, together with the ones previously identified, reveal the extensive genome response to selection on morphology, color and adaptation to new environments.     

Rethinking Human–Nonhuman Primate Contact and Pathogenic Disease Spillover

Zoonotic transmissions are a major global health risk, and human–animal contact is frequently raised as an important driver of transmission. A literature examining zooanthroponosis largely agrees that more human–animal contact leads to more risk. Yet the basis of this proposition, the term contact, has not been rigorously analyzed. To understand how contact is used to explain cross-species spillovers, we conducted a multi-disciplinary review of studies addressing human–nonhuman primate (NHP) engagements and pathogenic transmissions and employing the term contact. We find that although contact is frequently invoked, it is employed inconsistently and imprecisely across these studies, overlooking the range of pathogens and their transmission routes and directions. We also examine a related but more expansive approach focusing on human and NHP habitats and their spatial overlap, which can potentially facilitate pathogenic transmission. Contact and spatial overlap investigations cannot, however, explain the processes that bring together people, animals and pathogens. We therefore examine another approach that enhances our understanding of zoonotic spillovers: anthropological studies identifying such historical, social, environmental processes. Comparable to a One Health approach, our ongoing research in Cameroon draws contact, spatial overlap and anthropological–historical approaches into dialog to suggest where, when and how pathogenic transmissions between people and NHPs may occur. In conclusion, we call for zoonotic disease researchers to specify more precisely the human–animal contacts they investigate and to attend to how broader ecologies, societies and histories shape pathogen–human–animal interactions.     

Cell kinetic studies of in situ human brain tumors with bromodeoxyuridine

At the time of surgery, 18 patients with various brain tumors were given a 1-h i.v. infusion of bromodeoxyuridine (BrdUrd), 150-200 mg/m2. At an infusion rate of 200 mg/m2/h, serum BrdUrd levels of 8 microM were achieved. After the infusion, tumor tissue was obtained and divided into two portions. One portion was fixed in 70% ethanol, embedded in paraffin, and sectioned; the sections were deparaffinized, denatured with 2 N HCl, and reacted with monoclonal antibodies against BrdUrd (anti-BrdUrd MAb). BrdUrd-labeled nuclei were demonstrated satisfactorily by an indirect peroxidase method. The other portion was dissociated into single cells with a DNase enzyme cocktail and reacted with FITC-conjugated anti-BrdUrd MAb to determine the percentage of BrdUrd-labeled cells or with chromomycin A3 for DNA analysis. The single-cell suspensions were analyzed by flow cytometry. The fraction of S-phase cells in the tissue sections was similar to both the percentage of BrdUrd-labeled nuclei and the S-phase fraction determined by flow cytometric analysis. The results obtained with BrdUrd-labeled nuclei were similar to those obtained from previous autoradiographic studies of various brain tumors exposed to a pulse of 3H-thymidine. Since BrdUrd is not radioactive and is nontoxic at the dosage used, these techniques, together with the histopathological diagnosis, may help to predict the biological malignancy of individual tumors.     

The TGGCA protein binds to the MMTV-LTR, the adenovirus origin of replication, and the BK virus enhancer.

TGGCA-binding proteins are nuclear proteins with high affinity for double-stranded DNA homologous to the prototype recognition sequence 5'YTGGCANNNTGCCAR 3'. Their ubiquitous tissue distribution in higher vertebrates characterizes them as a class of highly conserved proteins which may exert a basic function. To obtain clues to this function, specific binding sites were mapped on three viral genomes. Recognition sites were identified in the enhancer region of the BK virus, in the LTR of the mouse mammary tumor virus, and in the origin of replication of adenovirus 12. The TGGCA-binding protein from HeLa cells appears to be identical to nuclear factor I described by others, which stimulates initiation of adenovirus DNA replication in vitro. However, data from MMTV, BKV, and from cellular genes suggest that this specific protein-DNA interaction may also be involved in the control of gene activity.