Variability of Bacterial Community Composition on Leaves Between and Within Plant Species

The phyllosphere is one of the largest habitats for terrestrial microorganisms. To gain a better insight into the factors underlying the composition of bacterial communities inhabiting leaf surfaces we performed culture-dependent and independent (Denaturing Gradient Gel Electrophoresis) analyses on the bacteria associated with the leaves of three plant species: Amygdalus communis, Citrus paradisi, and Nicotiana glauca. We found that the culturable classes Bacilli and Actinobacteria were the predominant classes on the phyllosphere of all three plant species. In contrast to this consistency on the bacterial class level, we found a significant variation on the bacterial species-level based on the culturable methods. Although some variation was detected among individual plants within one plant species, the inter-specific variability exceeded the intra-specific variability. C. paradisi leaf surface had the highest predicted total species richness (Chao 2 and ICE) and the highest species diversity (βw) among the three plant species. Our findings demonstrate that environmental conditions, mainly the plant species within a site, govern the bacterial community composition on leaf surfaces.     

Evaluation of helmet and goggle designs by modeling non-penetrating projectile impacts

Despite the progress in developing personal combat-protective gear, eye and brain injuries are still widely common and carry fatal or long-term repercussions. The complex nature of the cranial tissues suggests that simple methods (e.g. crash-dummies) for testing the effectiveness of personal protective gear against non-penetrating impacts are both expensive and ineffective, and there are ethical issues in using animal or cadavers. The present work presents a versatile testing framework for quantitatively evaluating protective performances of head and eye combat-protective gear, against non-penetrating impacts. The biomimetic finite element (FE) head model that was developed provides realistic representation of cranial structure and tissue properties. Simulated crash impact results were validated against a former cadaveric study and by using a crash-phantom developed in our lab. The model was then fitted with various helmet and goggle designs onto which a non-penetrating ballistic impact was applied. Example data show that reduction of the elastic and shear moduli by 30% and 80% respectively of the helmet outer Kevlar-29 layer, lowered intracranial pressures by 20%. Our modeling suggests that the level of stresses that develop in brain tissues, which ultimately cause the brain damage, cannot be predicted solely by the properties of the helmet/goggle materials. We further found that a reduced contact area between goggles and face is a key factor in reducing the mechanical loads transmitted to the optic nerve and eye balls following an impact. Overall, this work demonstrates the simplicity, flexibility and usefulness for development, evaluation, and testing of combat-protective equipment using computational modeling.

• Highlights

• A finite element head model was developed for testing head gear.

• Reduced helmet’s outer layer elastic and shear moduli lowered intracranial stresses.

• Gear material properties could not fully predict impact-related stress in the brain.

• Reduced goggles-face contact lowered transmitted loads to the optic nerve and eyes.

     

Detection of light images by simple tissues as visualized by photosensitized magnetic resonance imaging

In this study, we show how light can be absorbed by the body of a living rat due to an injected pigment circulating in the blood stream. This process is then physiologically translated in the tissue into a chemical signature that can be perceived as an image by magnetic resonance imaging (MRI). We previously reported that illumination of an injected photosynthetic bacteriochlorophyll-derived pigment leads to a generation of reactive oxygen species, upon oxygen consumption in the blood stream. Consequently, paramagnetic deoxyhemoglobin accumulating in the illuminated area induces changes in image contrast, detectable by a Blood Oxygen Level Dependent (BOLD)-MRI protocol, termed photosensitized (ps)MRI. Here, we show that laser beam pulses synchronously trigger BOLD-contrast transients in the tissue, allowing representation of the luminous spatiotemporal profile, as a contrast map, on the MR monitor. Regions with enhanced BOLD-contrast (7-61 fold) were deduced as illuminated, and were found to overlap with the anatomical location of the incident light. Thus, we conclude that luminous information can be captured and translated by typical oxygen exchange processes in the blood of ordinary tissues, and made visible by psMRI (Fig. 1). This process represents a new channel for communicating environmental light into the body in certain analogy to light absorption by visual pigments in the retina where image perception takes place in the central nervous system. Potential applications of this finding may include: non-invasive intra-operative light guidance and follow-up of photodynamic interventions, determination of light diffusion in opaque tissues for optical imaging and possible assistance to the blind.     

The Potential Anti-Herbivory Role of Microorganisms on Plant Thorns

Thorns, spines and prickles are some of the anti-herbivore defenses that plants have evolved. They were recently found to be commonly aposematic (warning coloration). However, the physical anti-herbivore defense executed by these sharp structures seems to be only the tip of the iceberg. We show that thorns of various plant species commonly harbor an array of aerobic and anaerobic pathogenic bacteria including Clostridium perfringens the causative agent of the life-threatening gas gangrene, Bacillus anthracis, and Pantoea agglomerans. Septic inflammation caused by plant thorn injury can result not only from bacteria. Medical literature indicates that thorns, spines or prickles also introduce pathogenic fungi into animals or humans. Dermatophytes that cause subcutaneous mycoses are unable to penetrate the skin and must be introduced into the subcutaneous tissue by a puncture wound. The common microorganism-thorn combinations seem to have been an important contributor to the fact that so many plant thorns are aposematically colored, as a case of convergent evolution of aposematism in these organisms.Key Words: aposematism, herbivory, pathogen, spine, thorn, bacillus anthracis, clostridium perfringens, sporotrichosis, Mycetoma, subcutaneous mycotic disease     

Nonparametric correction for covariate measurement error in a stratified Cox model

Stratified Cox regression models with large number of strata and small stratum size are useful in many settings, including matched case-control family studies. In the presence of measurement error in covariates and a large number of strata, we show that extensions of existing methods fail either to reduce the bias or to correct the bias under nonsymmetric distributions of the true covariate or the error term. We propose a nonparametric correction method for the estimation of regression coefficients, and show that the estimators are asymptotically consistent for the true parameters. Small sample properties are evaluated in a simulation study. The method is illustrated with an analysis of Framingham data.     

Direct phenotypic analysis of human MHC class I antigen presentation: visualization,quantitation, and in situ detection of human viral epitopes using peptide-specific,MHC-restricted human recombinant antibodies

The advent in recent years of the application of tetrameric arrays of class I peptide-MHC complexes now enables us to detect and study rare populations of Ag-specific CD8(+) T cells. However, available methods cannot visualize or determine the number and distribution of these TCR ligands on individual cells nor detect APCs in tissues. In this study, we describe for the first time studies of human class I peptide-MHC ligand presentation. These studies were facilitated by applying novel tools in the form of peptide-specific, HLA-A2-restricted human recombinant Abs directed toward a viral epitope derived from human T cell lymphotropic virus type I. Using a large human Ab phage display library, we isolated a large panel of recombinant Fab Abs that are specific for a particular peptide-MHC class I complex in a peptide-dependent, MHC-restricted manner. We used these Abs to visualize the specific complex on APCs and virus-infected cells by flow cytometry, to quantify the number of, and visualize in situ, a particular complex on the surface of APCs bearing complexes formed by naturally occurring active intracellular processing of the cognate viral Ag. These findings demonstrate our ability to transform the unique fine specificity, but low intrinsic affinity of TCRs into high affinity soluble Ab molecules endowed with a TCR-like specificity toward human viral epitopes. These molecules may prove to be crucial useful tools for studying MHC class I Ag presentation in health and disease as well as for therapeutic purposes in cancer, infectious diseases, and autoimmune disorders.     

Estimating hypermutation rates from clonal tree data

To understand the mechanisms underlying the varying patterns of mutations that occur during immune and autoimmune responses, estimates of the somatic hypermutation rate are critical. However, despite its significance, precise estimates of the mutation rate do not currently exist. Microdissection studies of mutating B cell clones provide an opportunity to measure this rate more accurately than previously possible. Each microdissection provides a number of clonally related sequences that, through the analysis of shared mutations, can be genealogically related to each other. The shape of these clonal trees is influenced by many processes, including the hypermutation rate. We have developed two different methods to estimate the mutation rate based on these data. These methods are applied to two sets of experimental data, one from an autoimmune response and one from the antihapten response to (4-hydroxy-3-nitrophenyl)acetyl (NP). Comparable mutation rates are estimated for both responses, 0.7-0.9 x 10(-3) and 0.9-1.1 x 10(-3) bp(-1) division(-1) for the autoimmune and NP responses, respectively. In addition to comparing the results of the two procedures, we investigate the effect on our estimate of assumptions, such as the fraction of lethal mutations.     

The erythropoietin receptor transmembrane domain mediates complex formation with viral anemic and polycythemic gp55 proteins

Erythropoietin receptor (EpoR) activation is crucial for mature red blood cell production. The murine EpoR can also be activated by the envelope protein of the polycythemic (P) spleen focus forming virus (SFFV), gp55-P. Due to differences in the TM sequence, gp55 of the anemic (A) strain SFFV, gp55-A, cannot efficiently activate the EpoR. Using antibody-mediated immunofluorescence co-patching, we show that the majority of EpoR forms hetero-oligomers at the cell surface with gp55-P and, surprisingly, with gp55-A. The EpoR TM domain is targeted by gp55-P and -A, as only chimeric receptors containing EpoR TM sequences oligomerized with gp55 proteins. Both gp55-P and gp55-A are homodimers on the cell surface, as shown by co-patching. However, when the homomeric interactions of the isolated TM domains were assayed by TOXCAT bacterial reporter system, only the TM sequence of gp55-P was dimerized. Thus, homo-oligomerization of gp55 proteins is insufficient for full EpoR activation, and a correct conformation of the dimer in the TM region is required. This is supported by the failure of gp55-A-->P, a mutant protein whose TM domain can homo-oligomerize, to fully activate EpoR. As unliganded EpoR forms TM-dependent but inactive homodimers, we propose that the EpoR can be activated to different extents by homodimeric gp55 proteins, depending on the conformation of the gp55 protein dimer in the TM region.