Concentrated patches of recent trembling aspen (Populus tremuloides) mortality covered 56,091 ha of Colorado forests in 2006. Mortality has progressed rapidly. Area affected increased 58% between 2005 and 2006 on the Mancos-Dolores Ranger District, San Juan National Forest, where it equaled nearly 10% of the aspen cover type. In four stands that were measured twice, incidence of mortality increased from 7-9% in 2002/2003 to 31-60% in 2006. Mortality generally decreased with increasing elevation over the primary elevation range of aspen and occurred on less steep slopes than healthy aspen. Slope-weighted mean aspects of aspen cover type were northern at low elevations and generally southern at high elevations. Relative frequency of mortality was generally highest on southern to western aspects. In 31 stands measured in detail, mortality ranged from 0 to 100% (mean 32%) and was negatively correlated with stand density (P = 0.033). Size of trees affected was strongly correlated with amount of current mortality (P < 0.001), and current mortality was skewed toward larger diameter classes. Density of regeneration was in a low range typical of undisturbed stands and did not increase with overstory mortality.Agents that typically kill mature trees in aspen stands were unimportant in this mortality. Instead, a group of interchangeable, usually secondary agents was most commonly associated with mortality, including Cytospora canker (usually caused by Valsa sordida), aspen bark beetles (Trypophloeus populi and Procryphalus mucronatus), poplar borer (Saperda calcarata), and bronze poplar borer (Agrilus liragus). The rapidity of mortality, mortality agents involved, and probably other causal factors distinguish this phenomenon from the long-term loss of aspen cover usually attributed to successional processes operating in an altered disturbance regime (and often exacerbated by ungulate browsing). Our data are consistent with a hypothesis that (a) predisposing factors include stand maturation, low density, southern aspects and low elevations; (b) a major inciting factor was the recent, acute drought accompanied by high temperatures, and; (c) contributing factors and proximate agents of mortality are the common biotic agents observed. On sites with poor regeneration and weak root systems, clones may die, resulting in the long-term loss of aspen forest cover. Published by Elsevier B.V.

Elucidating how antigen exposure and selection shape the human antibody repertoire is fundamental to our understanding of B-cell immunity. We sequenced the paired heavy-and light-chain variable regions (VH and VL, respectively) from large populations of single B cells combined with computational modeling of antibody structures to evaluate sequence and structural features of human antibody repertoires at unprecedented depth. Analysis of a dataset comprising 55,000 antibody clusters from CD19− IgM-naive B cells, >120,000 antibody clusters from CD19+ antigenexperienced B cells, and >2,000 RosettaAntibody-predicted structural models across three healthy donors led to a number of key findings: (i) VH and VL gene sequences pair in a combinatorial fashion without detectable pairing restrictions at the population level; (ii) certain VH:VL gene pairs were significantly enriched or depleted in the antigen-experienced repertoire relative to the naive repertoire; (iii) antigen selection increased antibody paratope net charge and solvent-accessible surface area; and (iv) public heavy-chain third complementarity-determining region (CDR-H3) antibodies in the antigen-experienced repertoire showed signs of convergent paired light-chain genetic signatures, including shared light-chain third complementarity-determining region (CDR-L3) amino acid sequences and/or Vκ,λ-Jκ,λ genes. The data reported here address several longstanding questions regarding antibody repertoire selection and development and provide a benchmark for future repertoire-scale analyses of antibody responses to vaccination and disease.antibody | B cell | immunology | high-throughput sequencing | computational modeling E ffective antigen recognition by the humoral immune system is predicated on the somatic generation of a large antibody repertoire that encompasses the sequence and conformational diversity to respond to a highly diversified set of antigens (1-3). Upon antigen challenge, naive B cells (NBCs) expressing unmutated antibodies capable of binding antigen with an affinity sufficient to initiate B-cell receptor (BCR) signaling may be stimulated to undergo somatic hypermutation (SHM) of the antibody genes. B cells expressing higher-affinity BCRs are better equipped to compete for antigen and thus receive signals that enable their preferential proliferation and further antibody sequence diversification in additional rounds of SHM. This process generates a repertoire of somatically mutated antibodies that, at the structural level, generally display decreased conformational flexibility (4, 5), slower antigen dissociation rates, and increased binding selectivity relative to the germline repertoire.Understanding the salient features of the human antibody repertoire is critical for immunology research (6, 7). Specifically, additional information is needed regarding how a history of pathogen and environmental exposure modulates the sequence and conformational properties of naive antibodies to yield a mature antibody repertoire that confers effective protection. Hi...

The Clostridium difficile toxins A and B are primarily responsible for symptoms of C. difficile associated disease and are prime targets for vaccine development. We describe a plasmid-based system for the production of genetically modified toxins in a non-sporulating strain of C. difficile that lacks the toxin genes tcdA and tcdB. TcdA and TcdB mutations targeting established glucosyltransferase cytotoxicity determinants were introduced into recombinant plasmids and episomally expressed toxin mutants purified from C. difficile transformants. TcdA and TcdB mutants lacking glucosyltransferase and autoproteolytic processing activities were ~10 000-fold less toxic to cultured human IMR-90 cells than corresponding recombinant or native toxins. However, both mutants retained residual cytotoxicity that could be prevented by preincubating the antigens with specific antibodies or by formalin treatment. Such non-toxic formalin-treated mutant antigens were immunogenic and protective in a hamster model of infection. The remaining toxicity of untreated TcdA and TcdB mutant antigens was associated with cellular swelling, a phenotype consistent with pore-induced membrane leakage. TcdB substitution mutations previously shown to block vesicular pore formation and toxin translocation substantially reduced residual toxicity. We discuss the implications of these results for the development of a C. difficile toxoid vaccine.

This paper considers distributed clustering of high-dimensional heterogeneous data using a distributed principal component analysis (PCA) technique called the collective PCA. It presents the collective PCA technique, which can be used independent of the clustering application. It shows a way to integrate the Collective PCA with a given offthe-shelf clustering algorithm in order to develop a distributed clustering technique. It also presents experimental results using different test data sets including an application for web mining.

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