Immunogenicity of recombinant fiber-chimeric adenovirus serotype 35 vector-based vaccines in mice and rhesus monkeys

Preexisting immunity to adenovirus serotype 5 (Ad5) has been shown to suppress the immunogenicity of recombinant Ad5 (rAd5) vector-based vaccines for human immunodeficiency virus type 1 (HIV-1) in both preclinical studies and clinical trials. A potential solution to this problem is to utilize rAd vectors derived from rare Ad serotypes, such as Ad35. However, rAd35 vectors have appeared less immunogenic than rAd5 vectors in preclinical studies to date. In this study, we explore the hypothesis that the differences in immunogenicity between rAd5 and rAd35 vectors may be due in part to differences between the fiber proteins of these viruses. We constructed capsid chimeric rAd35 vectors containing the Ad5 fiber knob (rAd35k5) and compared the immunogenicities of rAd5, rAd35k5, and rAd35 vectors expressing simian immunodeficiency virus Gag and HIV-1 Env in mice and rhesus monkeys. In vitro studies demonstrated that rAd35k5 vectors utilized the Ad5 receptor CAR rather than the Ad35 receptor CD46. In vivo studies showed that rAd35k5 vectors were more immunogenic than rAd35 vectors in both mice and rhesus monkeys. These data suggest that the Ad5 fiber knob contributes substantially to the immunogenicity of rAd vectors. Moreover, these studies demonstrate that capsid chimeric rAd vectors can be constructed to combine beneficial immunologic and serologic properties of different Ad serotypes.     

An experimental test of the evolution of increased competitive ability hypothesis in goldenrod, <Emphasis Type="Italic">Solidago gigantea</Emphasis>

The mechanisms that allow introduced plants to become invasive are poorly understood. Here, we present a test of the evolution of increased competitive ability hypothesis, which holds that because specialized natural enemies may be absent from the introduced range, exotic plants may evolve to invest less in anti-herbivore defenses and thereby gain a competitive advantage over native plants. We grew Solidago gigantea plants derived from both the native range (North America) and the invasive range (Europe) in a common garden in the native range for 2 years. Half the plants were treated with insecticide to protect them from insect herbivores and the other half were exposed to insects that colonized the garden from nearby fields. Insect herbivore biomass was significantly higher on European plants than US plants in the first year but not the second. European plants were more heavily attacked by pathogens in both years of the study. When exposed to insect herbivores, US plants produced more seed than European plants, but when plants were protected from herbivores, seed production was equivalent between US plants and European plants. The presence of insect herbivores suppressed seed production of European plants much more than that of US plants, even though the level of herbivory experienced by European and US plants was similar in the second year, suggesting that the ability to tolerate herbivory was diminished in European plants. These results partially support the EICA hypothesis: plants from the introduced range were more susceptible to some natural enemies and benefited more from insect removal than plants from the native range. The prediction that European plants would perform better than US plants in the absence of insect herbivores was not supported. Electronic Supplementary Material Supplementary material is available for this article at     

Pathogen survival in the external environment and the evolution of virulence

Recent studies have provided evolutionary explanations for much of the variation in mortality among human infectious diseases. One gap in this knowledge concerns respiratory tract pathogens transmitted from person to person by direct contact or through environmental contamination. The sit-and-wait hypothesis predicts that virulence should be positively correlated with durability in the external environment because high durability reduces the dependence of transmission on host mobility. Reviewing the epidemiological and medical literature, we confirm this prediction for respiratory tract pathogens of humans. Our results clearly distinguish a high-virulence high-survival group of variola (smallpox) virus, Mycobacterium tuberculosis, Cornynebacterium diphtheriae, Bordetella pertussis, Streptococcus pneumoniae, and influenza virus (where all pathogens have a mean percent mortality > or = 0.01% and mean survival time >10 days) from a low-virulence low-survival group containing ten other pathogens. The correlation between virulence and durability explains three to four times of magnitude of difference in mean percent mortality and mean survival time, using both across-species and phylogenetically controlled analyses. Our findings bear on several areas of active research and public health policy: (1) many pathogens used in the biological control of insects are potential sit-and-wait pathogens as they combine three attributes that are advantageous for pest control: high virulence, long durability after application, and host specificity; (2) emerging pathogens such as the 'hospital superbug' methicillin-resistant Staphylococcus aureus (MRSA) and potential bioweapons pathogens such as smallpox virus and anthrax that are particularly dangerous can be discerned by quantifying their durability; (3) hospital settings and the AIDS pandemic may provide footholds for emerging sit-and-wait pathogens; and (4) studies on food-borne and insect pathogens point to future research considering the potential evolutionary trade-offs and genetic linkages between virulence and durability.     

PRACTICAL STEREOLOGICAL METHODS FOR MORPHOMETRIC CYTOLOGY

Stereological principles provide efficient and reliable tools for the determination of quantitative parameters of tissue structure on sections. Some principles which allow the estimation of volumetric ratios, surface areas, surface-to-volume ratios, thicknesses of tissue or cell sheets, and the number of structures are reviewed and presented in general form; means for their practical application in electron microscopy are outlined. The systematic and statistical errors involved in such measurements are discussed.     

Co-immunoprecipitation of the Ly-5 molecule and an endogenous protease: a proteolytic system requiring a reducing agent and Ca2+1

Sodium [3H]borohydride- and [35S]methionine-labeled Ly-5 molecules from mouse thymocytes and T lymphoma cells were isolated with specific antibody and Staphylococcus aureus Cowan I (SaCI) strain; after extensive washing of the complexes, elution with Laemmli's reducing buffer (0.05 M Tris [pH 6.8 or 6.0], 4% sodium dodecyl sulfate [SDS], and 2% 2-mercaptoethanol [2-ME]) resulted in partial breakdown of the isolated Ly-5 molecules from a Mr = 175,000 to 150,000. Other proteins present during the elution step showed no evidence of proteolysis. 2-ME and SDS were required for proteolysis; although addition of exogenous Ca2+ during elution was not necessary, both EDTA and EGTA inhibited breakdown of the molecule that could be overcome by excess Ca2+. Of a variety of protease inhibitors and thiol-reactive agents tested, only TAME and oxidized glutathione blocked proteolysis almost completely. SaCI, serum, and contaminating antibodies were ruled out as the source of the proteolytic activity. More stringent preclearing and washing conditions did not eliminate endogenous proteolysis of the Ly-5 molecule. The endogenous proteolytic fragment had a Mr distinct from the tryptic fragment of the Ly-5 molecule. We conclude that the Ly-5 molecule may be autolytic or tightly associated with a distinct cellular protease.     

Accuracy and Reliability of a Novel Method for Fusion of Digital Dental Casts and Cone Beam Computed Tomography Scans

Several methods have been proposed to integrate digital models into Cone Beam Computed Tomography scans. Since all these methods have some drawbacks such as radiation exposure, soft tissue deformation and time-consuming digital handling processes, we propose a new method to integrate digital dental casts into Cone Beam Computed Tomography scans. Plaster casts of 10 patients were randomly selected and 5 titanium markers were glued to the upper and lower plaster cast. The plaster models were scanned, impressions were taken from the plaster models and the impressions were also scanned. Linear measurements were performed on all three models, to assess accuracy and reproducibility. Besides that, matching of the scanned plaster models and scanned impressions was done, to assess the accuracy of the matching procedure. Results show that all measurement errors are smaller than 0.2 mm, and that 81% is smaller than 0.1 mm. Matching of the scanned plaster casts and scanned impressions show a mean error between the two surfaces of the upper arch of 0.14 mm and for the lower arch of 0.18 mm. The time needed for reconstructing the CBCT scans to a digital patient, where the impressions are integrated into the CBCT scan of the patient takes about 15 minutes, with little variance between patients. In conclusion, we can state that this new method is a reliable method to integrate digital dental casts into CBCT scans. As far as radiation exposure, soft tissue deformation and digital handling processes are concerned, it is a significant improvement compared to the previously published methods.     

Variants of a Thermus aquaticus DNA Polymerase with Increased Selectivity for Applications in Allele- and Methylation-Specific Amplification

The selectivity of DNA polymerases is crucial for many applications. For example, high discrimination between the extension of matched versus mismatched primer termini is desired for the detection of a single nucleotide variation at a particular locus within the genome. Here we describe the generation of thermostable mutants of the large fragment of Thermus aquaticus DNA polymerase (KlenTaq) with increased mismatch extension selectivity. In contrast to previously reported much less active KlenTaq mutants with mismatch discrimination abilities, many of the herein discovered mutants show conserved wild-type-like high activities. We demonstrate for one mutant containing the single amino acid exchange R660V the suitability for application in allele-specific amplifications directly from whole blood without prior sample purification. Also the suitability of the mutant for methylation specific amplification in the diagnostics of 5-methyl cytosines is demonstrated. Furthermore, the identified mutant supersedes other commercially available enzymes in human leukocyte antigen (HLA) analysis by sequence-specific primed polymerase chain reactions (PCRs).     

The alien flora of Europe: a taxonomic and biogeographic review

Abstract. A geographic and taxonomic overview of the non-indigenous plant species of Europe, based on the ‘Flora Europaea’ is given. The flora of Europe includes 1568 species which have either expanded their ranges within Europe under human influence (naturalized European species) or are of non-European origin (exotic species). The latter group consists of 580 species (37%) which form a diverse group in terms of their taxonomic composition and geographic origin. The exotics are represented by 113 families, of which the Compositae, Rosaceae and Gramineae are most important. The ratio of species to families is low. Most exotic species in Europe originate from the Americas and Asia. Countries of southern Europe have a higher relative number of exotics in their non-native flora than northern ones. The species-range size distribution differs between naturalized European and exotic species; the latter are on average more widespread than the naturalized.