An atomic model of the outer layer of the bluetongue virus core derived from X-ray crystallography and electron cryomicroscopy

BACKGROUND: Bluetongue virus (BTV), which belongs to the Reoviridae family and orbivirus genus, is a non-enveloped, icosahedral, double-stranded RNA virus. Several protein layers enclose its genome; upon cell entry the outer layer is stripped away leaving a core, the surface of which is composed of VP7. The structure of the trimeric VP7 molecule has previously been determined using X-ray crystallography. The articulated VP7 subunit consists of two domains, one which is largely alpha-helical and the other, smaller domain, is a beta barrel with jelly-roll topology. The relative orientations of these two domains vary in different crystal forms. The structure of VP7 and the organizations of 780 subunits of this molecule in the core of virus is central to the assembly and function of BTV. RESULTS: A 23 A resolution map of the core, determined using electron cryomicroscopy (cryoEM) data, reveals that the 260 trimers of VP7 are organized on a rather precise T = 13 laevo icosahedral lattice, in accordance with the theory of quasi-equivalence. The VP7 layer occupies a shell that is between 260 A and 345 A from the centre of the core. Below this radius (230-260 A) lies the T = 1 layer of 120 molecules of VP3. By fitting the X-ray structure of an individual VP7 trimer onto the cryoEM BTV core structure, we have generated an atomic model of the VP7 layer of BTV. This demonstrates that one of the molecular structures seen in crystals of the isolated VP7 corresponds to the in vivo conformation of the molecule in the core. CONCLUSIONS: The beta-barrel domains of VP7 are external to the core and interact with protein in the outer layer of the mature virion. The lower, alpha-helical domains of VP7 interact with VP3 molecules which form the inner layer of the BTV core. Adjacent VP7 trimer-trimer interactions in the T = 13 layer are mediated principally through well-defined regions in the broader lower domains, to form a structure that conforms well with that expected from the theory of quasi-equivalence with no significant conformational changes within the individual trimers. The VP3 layer determines the particle size and forms a rather smooth surface upon which the two-dimensional lattice of VP7 trimers is laid down.     

Head injury mechanisms and the concept of preventive management: a review and critical synthesis

Recent advances in head injury research have produced a plethora of useful data coupled with a paucity of conceptual integration across the four ways in which this research is pursued. These research orientations are the epidemiological, biomechanical, basic neuroscientific, and clinicopathologic/therapeutic (including rehabilitation). This overview of the history and current state of the art assumes that biomechanics is the basic science of causation in head injury research and when fully integrated with its counterparts, physiology and pathology, it can serve to overcome our conceptual handicaps. A paradigm integrating biomechanics; into the sequence of preventive, protective, acute therapeutic, and rehabilitative interventions will be described as the concept of preventive management. From this we derive the hypothesized claim that the exact biomechanics and the physiopathologic response at the time of injury (at the macroscopic and microscopic levels) determine the sequence of so-called secondary effects that are conceived as the inexorable delayed manifestations of the primary events and concomitant boundary conditions. Knowledge of these events will enable accurate predictions of the natural history and outcome of head injuries from observations carried out in the early acute phase. Examples to test this claim will be given with particular reference to the two types of traumatic brain injury (TBI) phenomenologically associated with disturbances of consciousness, the onset of which can be either immediate or delayed. The current economics and availability of computational power provide a significant opportunity for the development of selected experimental, physical, and simulated models of head injury on the basis of which the complex neurovascular and nonneural cellular and fluid elements of the nervous system may be accurately modeled. This approach will significantly improve the efficiency and quality of the essential biological and clinical observations and model experiments required to validate the theoretical methods and their predictions.     

Comparison of molecular changes in lung cancers in HIV-positive and HIV-indeterminate subjects

CONTEXT: Human immunodeficiency virus (HIV) infection has been associated with an increasing incidence of malignancy, and HIV-infected persons have an increased incidence of primary lung carcinoma compared with the general population. OBJECTIVE: To investigate the molecular changes present in HIV-associated lung tumors and compare them with those present in lung carcinomas arising in HIV-indeterminate subjects ("sporadic tumors"). DESIGN: Convenience sample. SUBJECTS: Archival tissues from 11 HIV-positive persons and from 35 persons of indeterminate HIV status. SETTING: University-based medical centers and affiliated hospitals. MAIN OUTCOME MEASURES: Analysis of frequency of loss of heterozygosity (LOH) and microsatellite alteration (MA) using polymerase chain reaction and 16 polymorphic microsatellite markers at 8 chromosomal regions frequently deleted in lung cancer. Presence of HIV and human papillomavirus (HPV) sequences. RESULTS: The overall frequency of LOH at all chromosomal regions tested and the frequencies at most of the individual regions were similar in the 2 groups. Frequency of MA present in the HIV-associated tumors (0.18) was 6-fold higher than in sporadic tumors (0.03) (P<.001). At least 1 MA was present in 10 (91%) of 11 HIV-associated tumors vs 17 (48%) of 35 sporadic tumors (P=.02). Molecular changes were independent of tumor stage and gender. HIV and HPV sequences were not detected in the HIV-associated lung carcinomas. CONCLUSIONS: Microsatellite alterations, which reflect widespread genomic instability, occur at greatly increased frequency in HIV-associated lung carcinomas. Although the mechanism underlying the development of increased MAs is unknown, it may play a crucial role in the development of many HIV-associated tumors.     

Homology and functional similarity of an hrp-linked pathogenicity locus, dspEF, of Erwinia amylovora and the avirulence locus avrE of Pseudomonas syringae pathovar tomato

The "disease-specific" (dsp) region next to the hrp gene cluster of Erwinia amylovora is required for pathogenicity but not for elicitation of the hypersensitive reaction. A 6.6-kb apparent operon, dspEF, was found responsible for this phenotype. The operon contains genes dspE and dspF and is positively regulated by hrpL. A BLAST search revealed similarity in the dspE gene to a partial sequence of the avrE locus of Pseudomonas syringae pathovar tomato. The entire avrE locus was sequenced. Homologs of dspE and dspF were found in juxtaposed operons and were designated avrE and avrF. Introduced on a plasmid, the dspEF locus rendered P. syringae pv. glycinea race 4 avirulent on soybean. An E. amylovora dspE mutant, however, elicited a hypersensitive reaction in soybean. The avrE locus in trans restored pathogenicity to dspE strains of E. amylovora, although restored strains were low in virulence. DspE and AvrE are large (198 kDa and 195 kDa) and hydrophilic. DspF and AvrF are small (16 kDa and 14 kDa) and acidic with predicted amphipathic alpha helices in their C termini; they resemble chaperones for virulence factors secreted by type III secretion systems of animal pathogens.     

Medical challenges of internal conflicts

The most prevalent menace since the end of the cold war is the occurrence of civil wars and local and regional conflicts. The term "low intensity conflict" describes the new threat environment and covers a multitude of phenomena, such as civil wars, guerrilla warfare, terrorism and counterinsurgency operations occurring between routine, peaceful inter- or intrastate competition, and a sustained conventional conflict. There is a great challenge to alert the physicians in general, and the surgical community of the world in particular, to the new threat environment and the medical challenges involved in treating casualties of low intensity conflicts. Specifically, a new international body of surgeons might be required to coordinate the recruitment, training, and creditation for surgeons with special expertise in the management of victims of such conflicts and to facilitate research and general knowledge of the medical challenges of modern conflicts.     

Spatiotemporal imaging of human brain activity using functional MRI constrained magnetoencephalography data: Monte Carlo simulations

The goal of our research is to develop an experimental and analytical framework for spatiotemporal imaging of human brain function. Preliminary studies suggest that noninvasive spatiotemporal maps of cerebral activity can be produced by combining the high spatial resolution (millimeters) of functional MRI (fMRI) with the high temporal resolution (milliseconds) of electroencephalography (EEG) and magnetoencephalography (MEG). Although MEG and EEG are sensitive to millisecond changes in mental activity, the ability to resolve source localization and timing is limited by the ill-posed "inverse" problem. We conducted Monte Carlo simulations to evaluate the use of MRI constraints in a linear estimation inverse procedure, where fMRI weighting, cortical location and orientation, and sensor noise statistics were realistically incorporated. An error metric was computed to quantify the effects of fMRI invisible ("missing") sources, "extra" fMRI sources, and cortical orientation errors. Our simulation results demonstrate that prior anatomical and functional information from MRI can be used to regularize the EEG/MEG inverse problem, giving an improved solution with high spatial and temporal resolution. An fMRI weighting of approximately 90% was determined to provide the best compromise between separation of activity from correctly localized sources and minimization of error caused by missing sources. The accuracy of the estimate was relatively independent of the number and extent of the sources, allowing for incorporation of physiologically realistic multiple distributed sources. This linear estimation method provides an operator-independent approach for combining information from fMRI, MEG, and EEG and represents a significant advance over traditional dipole modeling.     

Ultrarapid metabolizers of debrisoquine: characterization and PCR-based detection of alleles with duplication of the CYP2D6 gene

During a six-month period 2,221 haemocultures obtained from patients hospitalized in the Faculty Hospital Olomouc were examined. In all 304 isolated bacteria the sensitivity was assessed by the standard dilution micromethod and moreover all positive haemocultures were examined the "direct" disc method. Agreement between the results of the two methods was proved in 84% of pairs of tests and within a range from 67 to 100%, depending on the type of antimicrobial preparation. Based on these findings it may be stated that assessment of the sensitivity by the "direct" method agrees significantly with assessment of the sensitivity according to minimal inhibitory concentrations (MIC). In patients with septicaemia this procedure makes it possible to change empirical antibiotherapy by 24 hours sooner to aimed therapy.     

Random walks for selected boolean implication and equivalence problems

This paper is concerned with the design and analysis of a random walk algorithm for the 2CNF implication problem (2CNFI). In 2CNFI, we are given two 2CNF formulas f₁{\phi_{1}} and f₂{\phi_{2}} and the goal is to determine whether every assignment that satisfies f₁{\phi_{1}} , also satisfies f₂{\phi_{2}} . The implication problem is clearly coNP-complete for instances of kCNF, k ≥ 3; however, it can be solved in polynomial time, when k ≤ 2. The goal of this paper is to provide a Monte Carlo algorithm for 2CNFI with a bounded probability of error. The technique developed for 2CNFI is then extended to derive a randomized, polynomial time algorithm for the problem of checking whether a given 2CNF formula Nae-implies another 2CNF formula.