Differential elution of histones from gel-trapped nuclei

A method has been developed for characterizing at, submicrogram levels the heterogeneity of histones from purified nuclei. The histones are eluted with a smooth concentration gradient from nuclei trapped in polyacrylamide–gel threads and are collected in a micro fraction collector suitable for volumes in the 10–100 μl range. The gradient and fraction collection systems are governed by cam driven syringes. Samples obtained are subjected to electrophoresis in a starch-gel system and the gels are stained with a highly sensitive stain specific for guanidinium groups. Seven major and a similar number of minor components are demonstrated in the histones. The method of differential elution of trapped macromolecules is suitable for use with systems other than nuclei and histones.     

The comparative efficiency of algorithms for the construction of support-vector machines for regression reconstruction tasks

Methods for support-vector machine construction do not require additional a priori information and allow for the processing of large datasets; therefore, they are of extreme importance for a range of computational biology tasks. The principal algorithms for support-vector machine construction are reviewed and a comparative analysis of algorithm efficiency is presented. The most efficient algorithms are identified using critical analysis of the results. The algorithms that were recommended are described in sufficient detail to enable practical implementation.     

Surface fine structure of the globiferous pedicellariae of the regular echinoid,Psammechinus miliaris gmelin

The globiferous pedicellariae of Psammechinus miliaris are described. Two fixation methods giving minimal distortion and rapid tissue hardening were adapted for soft tissue preparation for scanning electron microscopy. The pedicellarial valves are covered by a microvillous epithelium. The outer valve epithelial microvilli overlying red spherulocytes in the epidermis are characterized by a filament matrix radiating out from each microvillus. These microvilli may function in epidermal absorption of organic solutes. The inner valve microvilli are more densely packed and the filament matrix is absent. Ciliation is confined to the inner valve surface where the cilia are concentrated to form a distal sensory pad and sensory hillock. Behavioural evidence suggests a chemo- and mechanosensory role for the inner valve surface.     

Optimum walking techniques for idealized animals

The vertical component of the force exerted by a foot on the ground, in the course of a step, may rise to a single maximum and decline again (as in human running) or may show two distinct maxima (as in human walking). A foot may remain on the ground for a large or small fraction of the duration of a stride. Mathematical models are used to investigate the effects of these differences of technique on the energy cost of locomotion. The optimum technique for a biped at a given speed is different from the optimum for a hypothetical many-legged animal. The optima for quadrupedal walking are likely to lie between these extremes.
The walking techniques adopted by men at different speeds are close to the optima indicated by the bipedal model. The two maxima of the force exerted by a foot are higher, and have a lower minimum between them, at higher speeds of walking. The techniques adopted by a sheep are close to the optima indicated by the many-legged model but dogs use techniques rather closer to the optima for bipeds.
The limitations of the models are discussed.     

Large scale genotype-phenotype correlation analysis based on phylogenetic trees

We provide two methods for identifying changes in genotype that are correlated with changes in a phenotype implied by phylogenetic trees. The first method, VENN, works when the number of branches over which the change occurred are modest. VENN looks for genetic changes that are completely penetrant with phenotype changes on a tree. The second method, CCTSWEEP, allows for a partial matching between changes in phenotypes and genotypes and provides a score for each change using Maddison's concentrated changes test. The mutations that are highly correlated with phenotypic change can be ranked by score. We use these methods to find SNPs correlated with resistance to Bacillus anthracis in inbred mouse strains. Our findings are consistent with the current biological literature, and also suggest potential novel candidate genes.     

Uniform Terminology for the Protozoan Subphylum Apicomplexa*

SYNOPSIS. Uniform names for the stages, processes and structures of apicomplexan protozoa are proposed and defined, and names that should be superseded are listed. The same names are used for the same stages of all members of the group. Gregarines are designated as either septate or aseptate rather than as polycystid or monocystid. The gregarine stage often called a sporadin is recognized for what it is, a gamont. The cyst formed around 2 gregarine gamonts in which zygotes are formed is a gametocyst; it contains oocysts which in turn contain sporozoites. The term “spore” is inappropriate for these oocysts. The apical complex includes the polar ring, conoid, rhoptries, micronemes and subpellicular tubules. The gregarine “pseudocyst” is actually a gametocyst residuum. The term micropore is preferred to cytostome for the apicomplexan structure, since it is visible only with the electron microscope.     

Amperometric enzyme electrodes

Three different types of amperometric enzyme electrode are described. The first type uses a conducting organic-salt electrode to oxidize NADH. Results for sensors for ethanol and for bile acids are presented. In the second type of sensor, flavoenzymes are directly oxidized on the surface of the conducting organic-salt electrode. Results for five different enzymes are described. The mechanism of the enzyme oxidation is discussed and the reaction is shown to take place by heterogeneous redox catalysis and not by homogeneous mediation. The enzymes are strongly adsorbed on the electrode; microelectrodes for in vivo studies can be constructed without a membrane. Results for in vivo studies of changing glucose levels in the brain of a freely moving rat are presented. The third type of sensor is designed to measure low levels of toxic gases such as H2S and HCN. This is done by monitoring the inhibition by the toxic gas of the activity of the respiratory enzyme cytochrome oxidase.     

The mechanism for proton pumping in cytochrome c oxidase from an electrostatic and quantum chemical perspective

The mechanism for proton pumping in cytochrome c oxidase in the respiratory chain, has for decades been one of the main unsolved problems in biochemistry. However, even though several different suggested mechanisms exist, many of the steps in these mechanisms are quite similar and constitute a general consensus framework for discussing proton pumping. When these steps are analyzed, at least three critical gating situations are found, and these points are where the suggested mechanisms in general differ. The requirements for gating are reviewed and analyzed in detail, and a mechanism is suggested, where solutions for all the gating situations are formulated. This mechanism is based on an electrostatic analysis of a kinetic experiment fior the O to E transition. The key component of the mechanism is a positively charged transition state. An electron on heme a opens the gate for proton transfer from the N-side to a pump loading site (PLS). When the negative charge of the electron is compensated by a chemical proton, the positive transition state prevents backflow from the PLS to the N-side at the most critical stage of the pumping process. The mechanism has now been tested by large model DFT calculations, and these calculations give strong support for the suggested mechanism.     

The statistical analysis of truncated data: application to the Sverdlovsk anthrax outbreak

An outbreak of anthrax occurred in the city of Sverdlovsk in Russia in the spring of 1979. The outbreak was due to the inhalation of spores that were accidentally released from a military microbiology facility. In response to the outbreak a public health intervention was mounted that included distribution of antibiotics and vaccine. The objective of this paper is to develop and apply statistical methodology to analyse the Sverdlovsk outbreak, and in particular to estimate the incubation period of inhalational anthrax and the number of deaths that may have been prevented by the public health intervention. The data available for analysis from this common source epidemic are the incubation periods of reported deaths. The statistical problem is that incubation periods are truncated because some individuals may have had their deaths prevented by the public health interventions and thus are not included in the data. However, it is not known how many persons received the intervention or how efficacious was the intervention. A likelihood function is formulated that accounts for the effects of truncation. The likelihood is decomposed into a binomial likelihood with unknown sample size and a conditional likelihood for the incubation periods. The methods are extended to allow for a phase-in of the intervention over time. Assuming a lognormal model for the incubation period distribution, the median and mean incubation periods were estimated to be 11.0 and 14.2 days respectively. These estimates are longer than have been previously reported in the literature. The death toll from the Sverdlovsk anthrax outbreak could have been about 14% larger had there not been a public health intervention; however, the confidence intervals are wide (95% CI 0-61%). The sensitivity of the results to model assumptions and the parametric model for the incubation period distribution are investigated. The results are useful for determining how long antibiotic therapy should be continued in suspected anthrax cases and also for estimating the ultimate number of deaths in a new outbreak in the absence of any public health interventions.     

Sex-structured HIV/AIDS model to analyse the effects of condom use with application to Zimbabwe

We present a sex-structured model for heterosexual transmission of HIV/AIDS in a community. The model is formulated using integro-differential equations, which are shown to be equivalent to delay differential equations with a time delay due to incubation period. The sex-structured HIV/AIDS model divides the population into a two sex-structure consisting of females and males. The threshold and equilibria for the model are determined and stabilities are examined. We extend the model to focus on the effects of condom use as a single-strategy approach in HIV prevention in the absence of any treatment. Initially we model the use of male condoms and further extend the model to incorporate the use of both female and male condoms. The model includes two primary factors in condom use to control HIV that are condom efficacy and compliance. The exposure risk of infection after each intervention is obtained. Basic reproductive numbers for these models are computed and compared to assess the effectiveness of male and female condom use in a community. The models are numerically analysed to assess the effectiveness of condom use on the transmission dynamics of HIV/AIDS using demographic and epidemiological parameters for Zimbabwe. The study demonstrates the use of sex-structured HIV/AIDS models in assessing the effectiveness of female and male condom use as a preventative strategy in a heterosexually active population. Z. Mukandavire would like to acknowledge financial support given by the National University of Science and Technology through a Staff Development Scholarship. The authors are grateful to Eagle Insurance Company of Zimbabwe for financial support.     

Immunizations for foreign travel.

One of the most important aspects of preparing travelers for destinations throughout the world is providing them with immunizations. Before administering any vaccines, however, a careful health and immunization history and travel itinerary should be obtained in order to determine vaccine indications and contraindications. There are three categories of immunizations for foreign travel. The first category includes immunizations which are routinely recommended whether or not the individual is traveling. Many travelers are due for primary vaccination or boosting against tetanus-diphtheria, measles-mumps-rubella, pneumococcal pneumonia, and influenza, for example, and the pre-travel visit is an ideal time to administer these. The second category are immunizations which might be required by a country as a condition for entry; these are yellow fever and cholera. The final category contains immunizations which are recommended because there is a risk of acquiring a particular disease during travel. Typhoid fever, meningococcal disease, rabies, and hepatitis are some examples. Travelers who are pregnant or who are infected with the human immunodeficiency virus require special consideration. Provision of appropriate immunizations for foreign travel is an important aspect of preventing illness in travelers.     

Constraints and missing reactions in the urea cycle.

The stoichiometric relations in a series of biochemical reactions are summarized by a stoichiometric number matrix (with a column for each reaction) and a conservation matrix (with a row for each constraint). These two matrices for a series or cycle of biochemical reactions are related because the columns of the stoichiometric number matrix are in the null space of the conservation matrix, and the rows of the transpose of the conservation matrix are in the null space of the transpose of the stoichiometric number matrix. The conservation matrix for a system of biochemical reactions is of interest because it shows the nature of the constraints in addition to the conservation of atoms and groups. Constraints beyond those for the conservation of atoms and groups indicate "missing reactions" that do not occur because the enzymes involved couple reactions that could occur and still conserve atoms and groups. The interpretation of conservation matrices and stoichiometric matrices for a reaction system is complicated by the fact that they are not unique. However, their row-reduced forms are unique, as are their dimensions, which represent the number of reactants and number of independent reactions. Two matrices that look different contain the same information if they have the same row-reduced form. The urea cycle, which involves five enzyme-catalyzed reactions, and its net reaction are discussed in terms of the linear constraints produced by enzyme catalysis. A procedure to obtain a set of conservation equations that will yield the correct net reaction is described.     

Segregating Sites in a Gene Conversion Model with Mutation

Formulae for the expectation and variance of the number of segregating and homogeneous sites in a sample of two chromosomes are found. The model includes gene conversion and infinitely-many-alleles mutation in a coalescent framework. The corresponding infinitely-many-sites model limits are also found. The formulae for the expectation are extended to any sample size. Comparisons are drawn between the pure mutation model and the model where gene conversion has been added.     

An ultrahigh-throughput screening assay for estrogen receptor ligands.

Members of the steroid and thyroid hormone receptor superfamily (nuclear receptors) play diverse roles in mammalian physiology, in both normal and pathological states. For this reason, and because nuclear receptors are natural receptors for lipophilic small molecules, they are important therapeutic targets for the pharmaceutical industry. Here we describe a method for screening for ligands for one of these receptors, the estrogen receptor. The method is rapid, robust, and reliable, and has been used in an ultrahigh-throughput robotic screen. The receptor is crosslinked to a scintillant-containing solid support (FlashPlate) via a receptor-specific antibody. Test compounds are assayed for competition with a radiolabeled estrogen for binding to the immobilized receptor. Receptor-ligand complexes are allowed to form and receptor-bound radioactivity is detected in a scintillation counter. The assay has been designed for both isoforms of the estrogen receptor, alpha and beta, using separate antibodies for each. Given a radioactive tracer and an appropriate antibody, many of which are now commercially available, the assay could be established for any nuclear receptor.     

A statistical method for the comparison of a discrete diagnostic test with several continuous diagnostic tests.

In this paper we study a statistic that is suitable for comparing a discrete diagnostic marker to one or more continuous diagnostic markers. Test procedures and confidence intervals are based on asymptotic normality. The statistic is applicable for correlated data in which all the markers are obtained for each subject. The statistic was studied for use in comparing two markers for rectal bleeding. Examples for this application and two more general applications are presented.     

Analyzing the normal mode dynamics of macromolecules by the component synthesis method: Residue clustering and multiple-component approach

A study of the component synthesis method (CSM) for analyzing the normal mode dynamics of macromolecules is reported. The procedure involves a reduction of the dimensions of the normal mode problems for large molecular systems and the accurate extraction of the low-frequency modes. A macromolecule is divided into small components based on a hierarchical clustering of the residues in the structure. Interactions between coupled components are treated by the method of static correlation. The normal modes of the components are obtained first, and a fraction of the low-frequency normal modes of the components under mutual correlations are then used as a reduced basis for solving for the normal modes of the whole molecule. Multiple components are introduced for large macromolecules so that the dimensions of the eigenvalue problems at the component level are small. The method is applied to the protein crambin. In test calculations in which the dimensions of the eigenvalue equations are reduced to 1/6 of their natural size, the errors in the normal mode frequencies calculated by the CSM procedure are only about 1–2% when compared with the exact values. The rms fluctuations of all atoms in crambin calculated by the CSM procedure are basically identical to the exact results. The CSM procedure is shown to be accurate for calculating the normal modes of large macromolecules with a significant reduction of the size of the problem. © 1994 John Wiley & Sons, Inc.     

Genetic analysis of Tourette syndrome suggesting major gene effect.

Data on Gilles de la Tourette syndrome are analyzed by multiple threshold models in inheritance that incorporate sex effect. The polygenic-multifactorial model is rejected. Single major locus inheritance can account for the data, although many of the occurrences of Tourette are due to nongenetic phenocopies. In both models, males and females share a common genetic environmental liability, but the less prevalent sex, that is, females, has a higher genetic loading for the disorder. The predicted population prevalences in the single major locus model are 2.3% for males and 0.8% for females. The implications for genetic and biological research in Tourette syndrome are discussed.     

Study of the surface barrier discharge generated by electrodes in the form of a series of parallel metal strips

A surface discharge in a system where metal electrodes in the form of a series of parallel strips are positioned on the dielectric surface is studied. Analytical formulas for calculating the spatial distribution of the potential and the electric field in a discharge cell are derived. It is shown that the geometry of the metal electrodes should be taken into account (along with physical and chemical characteristics of the dielectric, the voltage applied to the electrodes, and other parameters of the system) for generation of the electric field with optimal configuration in the discharge cell. The obtained results are also applicable for analysis of discharge cells with a coplanar barrier discharge where metal electrodes are positioned in the dielectric at small depths. The results are of interest since a barrier discharge is one of the efficient methods for generating non-equilibrium plasma at high pressures for a variety of technological applications.     

Biotope prioritisation in the Central Apennines (Italy): species rarity and cross-taxon congruence

The conservation status of invertebrates is usually lesser known than that of vertebrates, and strategies to identify biotopes to preserve invertebrate diversity are typically based on a single surrogate taxon, or even on the use of vertebrates as surrogates. Aim of this research is to illustrate a method for biotope prioritisation that can be easily adapted to different animal groups and geographical contexts. A two-step protocol for biotope prioritisation is proposed on the basis of a multidimensional characterisation of species vulnerability. Firstly, species vulnerability is estimated from rarity measures which include geographical range, abundance and biotope specialisation. Then, these values of vulnerability are used to rank biotopes. The method was applied here to the tenebrionid beetles, the butterflies, the birds and the mammals of the Central Apennines, a montane area of high conservation concern for South Europe. This study provides evidence for the importance of including insects in conservation decisions, because vertebrates are poor surrogates for insects. Conservation efforts in the reserves included in the study area are mostly focused on vertebrates, for which woodlands are considered particularly important. However high altitude open biotopes are crucial for both tenebrionids and butterflies, and preservation of such kind of biotopes would be beneficial also for vertebrates. The approach applied here demonstrates that (1) vertebrates are poor surrogates for insects, and (2) measures of species rarity, typically used in vertebrate conservation, can be obtained also for insects, for which a veritable amount of data are hidden in specialised literature and museum collections.