A molecular comparison of T lymphocyte populations infiltrating the liver and circulating in the blood of patients with chronic hepatitis B: Evidence for antigen-driven selection of a public complementarity-determining region 3 (CDR3) motif

Despite a large number of T cells infiltrating the liver of patients with chronic hepatitis B, little is known about their complexity or specificity. To characterize the composition of these T cells involved with the pathogenesis of chronic hepatitis B (CHB), we have studied the clonality of V beta T cell receptor (TCR)-bearing populations in liver tissue by size spectratyping the complementarity-determining region (CDR3) lengths of TCR transcripts. We have also compared the CDR3 profiles of the lymphocytes infiltrating the liver with those circulating in the blood to see whether identical clonotypes may be detected that would indicate a virus-induced expansion in both compartments. Our studies show that in most of the patients examined, the T cell composition of liver infiltrating lymphocytes is highly restricted, with evidence of clonotypic expansions in 4 to 9 TCR V beta subfamilies. In contrast, the blood compartment contains an average of 1 to 3 expansions. This pattern is seen irrespective of the patient's viral load or degree of liver pathology. Although the TCR repertoire profiles between the 2 compartments are generally distinct, there is evidence of some T cell subsets being equally distributed between the blood and the liver. Finally, we provide evidence for a putative public binding motif within the CDR3 region with the sequence G-X-S, which may be involved with hepatitis B virus recognition.

Stimulus-driven selection of routes to imitation

Several models have proposed that an action can be imitated via one of two routes: a direct visuospatial route, which can in principle mediate imitation of both meaningful (MF) and meaningless (ML) actions, and an indirect semantic route, which can be used only for MF actions. The present study investigated whether selection between the direct and indirect routes is strategic or stimulus driven. Tessari and Rumiati (J Exp Psychol Hum Percept Perform 30:1107–1116, 2004) have previously shown, using accuracy measures, that imitation of MF actions is superior to imitation of ML actions when the two action types are presented in separate blocks, and that the advantage of MF over ML items is smaller or absent when they are presented in mixed blocks. We first replicated this finding using an automated reaction time (RT), as well as accuracy, measure. We then examined imitation of MF and ML actions in the mixed condition as a function of the action type presented in the previous trial and in relation to the number of previous test trials. These analyses showed that (1) for both action types, performance was worse immediately after ML than MF trials, and (2) even at the beginning of the mixed condition, responding to MF actions was no better than responding to ML items. These results suggest that the properties of the action stimulus play a substantial role in determining whether imitation is mediated by the direct or the indirect route, and that effects of block composition on imitation need not be generated through strategic switching between routes.

Hierarchical classification of G-protein-coupled receptors with data-driven selection of attributes and classifiers

We address the important bioinformatics problem of predicting protein function from a protein's primary sequence. We consider the functional classification of G-Protein-Coupled Receptors (GPCRs), whose functions are specified in a class hierarchy. We tackle this task using a novel top-down hierarchical classification system where, for each node in the class hierarchy, the predictor attributes to be used in that node and the classifier to be applied to the selected attributes are chosen in a data-driven manner. Compared with a previous hierarchical classification system selecting classifiers only, our new system significantly reduced processing time without significantly sacrificing predictive accuracy.

The role of parasite-driven selection in shaping landscape genomic structure in red grouse (Lagopus lagopus scotica)

This article is protected by copyright. All rights reserved. Acknowledgements This study was funded by a BBSRC studentship (MAW) and NERC grants NE/H00775X/1 and NE/D000602/1 (SBP). The authors are grateful to Mario Röder and Keliya Bai for fieldwork assistance, and all estate owners, factors and keepers for access to field sites, most particularly MJ Taylor and Mike Nisbet (Airlie), Neil Brown (Allargue), RR Gledson and David Scrimgeour (Delnadamph), Andrew Salvesen and John Hay (Dinnet), Stuart Young and Derek Calder (Edinglassie), Kirsty Donald and David Busfield (Glen Dye), Neil Hogbin and Ab Taylor (Glen Muick), Alistair Mitchell (Glenlivet), Simon Blackett, Jim Davidson and Liam Donald (Invercauld), Richard Cooke and Fred Taylor† (Invermark), Shaila Rao and Christopher Murphy (Mar Lodge), and Ralph Peters and Philip Astor (Tillypronie). Data accessibility • Genotype data (DataDryad: doi:10.5061/dryad.4t7jk) • Metadata (information on sampling sites, phenotypes and medication regimen) (DataDryad: doi:10.5061/dryad.4t7jk)

Étude du polymorphisme du gène majeur d’histocompatibilité de classe IIb (MHIIb) chez l’omble de fontaine (Salvelinus fontinalis)

Les molécules classiques du complexe majeur d’histocompatibilité de classe II (CMHII) sont des glycoprotéines de surface spécialisées dans la présentation de peptides, principalement dérivés de pathogènes extracellulaires, aux récepteurs des lymphocytes T CD4+ afin d’initier la réponse immunitaire adaptative. Elles sont encodées, avec celles du CMH de classe I, par les gènes les plus polymorphiques identifiés jusqu’à maintenant, avec plusieurs loci et une grande diversité allélique à chacun d’eux. De plus, le polymorphisme des gènes du CMHII n’est pas limité qu’aux séquences codantes. Il est également observé dans les promoteurs où on a démontré ses effets sur le niveau d’expression des gènes. La variation de la régulation d’un gène est considérée comme un facteur important et pour laquelle des modifications morphologiques, physiologiques et comportementales sont observées chez tous les organismes. Des séquences d’ADN répétées impliquées dans cette régulation ont été identifiées dans les régions non-codantes des génomes. D’un autre côté, la sélection par les pathogènes permettrait l’évolution et le maintien du polymorphisme des gènes du CMH chez les vertébrés. À ce sujet, plusieurs études ont montré l’implication de différents allèles du CMH dans la résistance ou la susceptibilité aux maladies. Cette étude avait pour objectifs de caractériser le polymorphisme du gène MHIIb chez l’omble de fontaine (Salvelinus fontinalis) et de documenter ses effets au niveau de la survie conférée par des allèles et/ou génotypes particuliers lors d’une infection, ainsi que sur la variation du niveau d’expression du gène dans différentes conditions. Dans une première partie, nous avons identifié un total de 6 allèles du gène MHIIb, désignés Safo-DAB*0101 à Safo-DAB*0601, qui montrent une grande similarité avec les séquences codantes provenant de poissons téléostéens et de l’humain. L’analyse des séquences du domaine b1 a permis de détecter l’effet d’une pression sélective positive pour maintenir le polymorphisme dans cette région de la molécule. Quatre de ces allèles ont été testés lors d’une expérience d’infection avec le pathogène Aeromonas salmonicida afin d’évaluer l’effet qu’ils pouvaient avoir sur la survie des poissons. Nous avons trouvé que l’allèle DAB*0101 était significativement associé à la résistance à la furonculose. En plus d’avoir été identifié chez les individus homozygotes pour cet allèle, l’effet a également été remarqué au niveau de la survie les poissons de génotype DAB*0101/*0201. À l’opposé, les facteurs de risque élevé obtenus pour les génotypes DAB*0201/*0301 et DAB*0301/*0401 suggèrent plutôt une association à la susceptibilité. Étant donné la faible fréquence à laquelle l’allèle DAB*0101 a été retrouvé dans la population, le modèle de la sélection dépendante de la fréquence pourrait expliquer l’avantage conféré par ce dernier et souligne l’importance de ce mécanisme pour le maintien du polymorphisme du gène MHIIb chez l’omble de fontaine. Dans une seconde partie, nous avons rapporté la présence d’un minisatellite polymorphique formé d’un motif de 32 nucléotides dans le second intron du gène MHIIb, et pour lequel un nombre exclusif de répétitions du motif a été associé à chaque allèle (69, 27, 20, 40, 19 et 25 répétitions pour les allèles DAB*0101 à DAB*0601 respectivement). L’expression relative de quatre allèles a été évaluée dans des poissons hétérozygotes aux températures de 6 ºC et 18 ºC. Les résultats indiquent que les allèles possédant un long minisatellite montrent une réduction de l’expression du gène d’un facteur 1,67 à 2,56 par rapport aux allèles qui en contiennent un court. De même, des allèles qui incluent des minisatellites de tailles similaires n’affichent pas de différence significative au niveau de l’abondance du transcrit aux deux températures. De plus, l’effet répressif associé aux longs minisatellites est amplifié à la température de 18 ºC dans des poissons de trois génotypes différents. Nous avons finalement observé une augmentation significative par un facteur 2,08 de l’expression totale du gène MHIIb à la température de 6 ºC. Ces résultats appuient l’implication des séquences d’ADN répétées dans la régulation de l’activité transcriptionnelle d’un gène et suggèrent qu’un minisatellite sensible aux différences de températures pourrait être soumis aux forces sélectives et jouer un rôle important dans l’expression de gènes et l’évolution des organismes poïkilothermes.

Stocking may increase mitochondrial DNA diversity but fails to halt the decline of endangered Atlantic salmon populations

Over the last 50 years, Spanish Atlantic salmon (Salmo salar) populations have been in decline. In order to bolster these populations, rivers were stocked with fish of northern European origin during the period 1974-1996, probably also introducing the furunculosis-inducing pathogen, Aeromonas salmonicida. Here we assess the relative importance of processes influencing mitochondrial (mt)DNA variability in these populations from 1948 to 2002. Genetic material collected over this period from four rivers in northern Spain (Cantabria) was used to detect variability at the mtDNA ND1 gene. Before stocking, a single haplotype was found at high frequency (0.980). Following stocking, haplotype diversity (h) increased in all rivers (mean h before stocking was 0.041, and 0.245 afterwards). These increases were due principally to the dramatic increase in frequency of a previously very low frequency haplotype, reported at higher frequencies in northern European populations proximate to those used to stock Cantabrian rivers. Genetic structuring increased after stocking: among-river differentiation was low before stocking (1950s/1960s Phi(ST) = -0.00296-0.00284), increasing considerably at the height of stocking (1980s Phi(ST) = 0.18932) and decreasing post-stocking (1990s/2002 Phi(ST) = 0.04934-0.03852). Gene flow from stocked fish therefore seems to have had a substantial role in increasing mtDNA variability. Additionally, we found significant differentiation between individuals that had probably died from infectious disease and apparently healthy, angled fish, suggesting a possible role for pathogen-driven selection of mtDNA variation. Our results suggest that stocking with non-native fish may increase genetic diversity in the short term, but may not reverse population declines.

Variety matters: adaptive genetic diversity and parasite load in two mouse opossums from the Brazilian Atlantic forest

The adaptive potential of a species to a changing environment and in disease defence is primarily based on genetic variation. Immune genes, such as genes of the major histocompatibility complex (MHC), may thereby be of particular importance. In marsupials, however, there is very little knowledge about natural levels and functional importance of MHC polymorphism, despite their key role in the mammalian evolution. In a previous study, we discovered remarkable differences in the MHC class II diversity between two species of mouse opossums (Gracilinanus microtarsus, Marmosops incanus) from the Brazilian Atlantic forest, which is one of the most endangered hotspots for biodiversity conservation. Since the main forces in generating MHC diversity are assumed to be pathogens, we investigated in this study gastrointestinal parasite burden and functional associations between the individual MHC constitution and parasite load. We tested two contrasting scenarios, which might explain differences in MHC diversity between species. We predicted that a species with low MHC diversity would either be under relaxed selection pressure by low parasite diversity (`Evolutionary equilibrium` scenario), or there was a recent loss in MHC diversity leading to a lack of resistance alleles and increased parasite burden (`Unbalanced situation` scenario). In both species it became apparent that the MHC class II is functionally important in defence against gastrointestinal helminths, which was shown here for the first time in marsupials. On the population level, parasite diversity did not markedly differ between the two host species. However, we did observe considerable differences in the individual parasite load (parasite prevalence and infection intensity): while M. incanus revealed low MHC DAB diversity and high parasite load, G. microtarsus showed a tenfold higher population wide MHC DAB diversity and lower parasite burden. These results support the second scenario of an unbalanced situation.

Maternal and paternal contributions to pathogen resistance dependent on development stage in a whitefish (Salmonidae)

It is often assumed that maternal and paternal contributions to offspring phenotype change over the lifetime of an individual. However, studies on parental effects typically suffer from the problems that heritabilities and maternal environmental effects are difficult to separate, and that both may depend on environmental factors and developmental stage. In order to experimentally disentangle maternal from paternal contributions and the likely effects of developmental stage from ecological effects, we sampled a natural population of the whitefish Coregonus palaea, used gametes for full-factorial in vitro fertilizations, raised over 10000 of the resulting offspring singly at controlled conditions, and exposed them at different points during embryonic development to one of two strains of Pseudomonas fluorescens that differed in their virulence characteristics (only one caused mortality, while both delayed hatching and reduced growth). Vulnerability to infection increased markedly over embryo development. This change coincided with a distinct shift in the importance of maternal to additive genetic effects on survival. Timing of exposure also affected the variance components for hatching time and larval length, but in a less consistent direction than the variance components for mortality. No significant genetic variation was found for any reaction norms across time points of exposure, indicating a uniformity among genotypes in how susceptibility changed over development. Phenotypes were also typically correlated across time points, which could constrain the evolution of the reaction norms. Our experiment demonstrates that the relative maternal and paternal contributions to susceptibility to an infection, and hence the evolutionary potential to respond to pathogen-induced selection, depends not only on the kind of pathogenic stress but also on the timing of the challenge.

Attentional Selection in Object Recognition

A key problem in object recognition is selection, namely, the problem of identifying regions in an image within which to start the recognition process, ideally by isolating regions that are likely to come from a single object. Such a selection mechanism has been found to be crucial in reducing the combinatorial search involved in the matching stage of object recognition. Even though selection is of help in recognition, it has largely remained unsolved because of the difficulty in isolating regions belonging to objects under complex imaging conditions involving occlusions, changing illumination, and object appearances. This thesis presents a novel approach to the selection problem by proposing a computational model of visual attentional selection as a paradigm for selection in recognition. In particular, it proposes two modes of attentional selection, namely, attracted and pay attention modes as being appropriate for data and model-driven selection in recognition. An implementation of this model has led to new ways of extracting color, texture and line group information in images, and their subsequent use in isolating areas of the scene likely to contain the model object. Among the specific results in this thesis are: a method of specifying color by perceptual color categories for fast color region segmentation and color-based localization of objects, and a result showing that the recognition of texture patterns on model objects is possible under changes in orientation and occlusions without detailed segmentation. The thesis also presents an evaluation of the proposed model by integrating with a 3D from 2D object recognition system and recording the improvement in performance. These results indicate that attentional selection can significantly overcome the computational bottleneck in object recognition, both due to a reduction in the number of features, and due to a reduction in the number of matches during recognition using the information derived during selection. Finally, these studies have revealed a surprising use of selection, namely, in the partial solution of the pose of a 3D object.

MetaVaccinology: a new vaccine discovery tool

In the last decade, the reverse vaccinology approach shifted the paradigm of vaccine discovery from conventional culture-based methods to high-throughput genome-based approaches for the development of recombinant protein-based vaccines against pathogenic bacteria. Besides reaching its main goal of identifying new vaccine candidates, this new procedure produced also a huge amount of molecular knowledge related to them. In the present work, we explored this knowledge in a species-independent way and we performed a systematic in silico molecular analysis of more than 100 protective antigens, looking at their sequence similarity, domain composition and protein architecture in order to identify possible common molecular features. This meta-analysis revealed that, beside a low sequence similarity, most of the known bacterial protective antigens shared structural/functional Pfam domains as well as specific protein architectures. Based on this, we formulated the hypothesis that the occurrence of these molecular signatures can be predictive of possible protective properties of other proteins in different bacterial species. We tested this hypothesis in Streptococcus agalactiae and identified four new protective antigens. Moreover, in order to provide a second proof of the concept for our approach, we used Staphyloccus aureus as a second pathogen and identified five new protective antigens. This new knowledge-driven selection process, named MetaVaccinology, represents the first in silico vaccine discovery tool based on conserved and predictive molecular and structural features of bacterial protective antigens and not dependent upon the prediction of their sub-cellular localization.

Genome-wide scans to uncover loci underlying bovine reproductive biology

Pós-graduação em Medicina Veterinária - FCAV

Comparative morpho-physiology of the metapleural glands of two Atta leaf-cutting ant queens nesting in clayish and organic soils

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Rapid evolution in plant chitinases: Molecular targets of selection in plant-pathogen coevolution

Many pathogen recognition genes, such as plant R-genes, undergo rapid adaptive evolution, providing evidence that these genes play a critical role in plant-pathogen coevolution. Surprisingly, whether rapid adaptive evolution also occurs in genes encoding other kinds of plant defense proteins is unknown. Unlike recognition proteins, plant chitinases attack pathogens directly, conferring disease resistance by degrading chitin, a component of fungal cell walls. Here, we show that nonsynonymous substitution rates in plant class I chitinase often exceed synonymous rates in the plant genus Arabis (Cruciferae) and in other dicots, indicating a succession of adaptively driven amino acid replacements. We identify individual residues that are likely subject to positive selection by using codon substitution models and determine the location of these residues on the three-dimensional structure of class I chitinase. In contrast to primate lysozymes and plant class III chitinases, structural and functional relatives of class I chitinase, the adaptive replacements of class I chitinase occur disproportionately in the active site cleft. This highly unusual pattern of replacements suggests that fungi directly defend against chitinolytic activity through enzymatic inhibition or other forms of chemical resistance and identifies target residues for manipulating chitinolytic activity. These data also provide empirical evidence that plant defense proteins not involved in pathogen recognition also evolve in a manner consistent with rapid coevolutionary interactions.

Partitioning The Net Effect Of Host Diversity On An Emerging Amphibian Pathogen.

The 'dilution effect' (DE) hypothesis predicts that diverse host communities will show reduced disease. The underlying causes of pathogen dilution are complex, because they involve non-additive (driven by host interactions and differential habitat use) and additive (controlled by host species composition) mechanisms. Here, we used measures of complementarity and selection traditionally employed in the field of biodiversity-ecosystem function (BEF) to quantify the net effect of host diversity on disease dynamics of the amphibian-killing fungus Batrachochytrium dendrobatidis (Bd). Complementarity occurs when average infection load in diverse host assemblages departs from that of each component species in uniform populations. Selection measures the disproportionate impact of a particular species in diverse assemblages compared with its performance in uniform populations, and therefore has strong additive and non-additive properties. We experimentally infected tropical amphibian species of varying life histories, in single- and multi-host treatments, and measured individual Bd infection loads. Host diversity reduced Bd infection in amphibians through a mechanism analogous to complementarity (sensu BEF), potentially by reducing shared habitat use and transmission among hosts. Additionally, the selection component indicated that one particular terrestrial species showed reduced infection loads in diverse assemblages at the expense of neighbouring aquatic hosts becoming heavily infected. By partitioning components of diversity, our findings underscore the importance of additive and non-additive mechanisms underlying the DE.

Mitochondrial targeting adaptation of the hominoid-specific glutamate dehydrogenase driven by positive Darwinian selection.

Many new gene copies emerged by gene duplication in hominoids, but little is known with respect to their functional evolution. Glutamate dehydrogenase (GLUD) is an enzyme central to the glutamate and energy metabolism of the cell. In addition to the single, GLUD-encoding gene present in all mammals (GLUD1), humans and apes acquired a second GLUD gene (GLUD2) through retroduplication of GLUD1, which codes for an enzyme with unique, potentially brain-adapted properties. Here we show that whereas the GLUD1 parental protein localizes to mitochondria and the cytoplasm, GLUD2 is specifically targeted to mitochondria. Using evolutionary analysis and resurrected ancestral protein variants, we demonstrate that the enhanced mitochondrial targeting specificity of GLUD2 is due to a single positively selected glutamic acid-to-lysine substitution, which was fixed in the N-terminal mitochondrial targeting sequence (MTS) of GLUD2 soon after the duplication event in the hominoid ancestor approximately 18-25 million years ago. This MTS substitution arose in parallel with two crucial adaptive amino acid changes in the enzyme and likely contributed to the functional adaptation of GLUD2 to the glutamate metabolism of the hominoid brain and other tissues. We suggest that rapid, selectively driven subcellular adaptation, as exemplified by GLUD2, represents a common route underlying the emergence of new gene functions.