Distribution of legionellae in a hospital water system: prevalence of immunologically and genetically related Legionella pneumophila serogroup 6 isolates

A hospital warm water system was monitored for the presence and distribution of legionellae. Subtyping of ten selected Legionella pneumophila isolates, originating from four different sites in the system by using serogroup specific antisera in an indirect immunofluorescence test, revealed that nine of the ten isolates belong to serogroup 6, while the remaining one was serogroup 10. Two monoclonal antibodies (mAbs) specific for a subgroup of serogroup 6 strains were further used for characterization. None of the strains reacted with these mAbs. Genome analysis by elaborating NotI profiles using the pulsed field gel electrophoresis (PFGE) technique revealed that nearly all serogroup 6 isolates derived from different sites, including a new building connected by a ring pipe, were identical according to restriction fragment patterns. The patterns were distinguishable from those of the two L. pneumophila serogroup 6 reference strains, and from that of the L. pneumophila serogroup 10 isolate. These data argue for a relatively homogeneous L. pneumophila serogroup 6 population in the entire water system.     

Chromosomal assignment of two putative canine keratin gene clusters

Mutations in keratin genes account for a number of inherited keratodermas in humans. The groups of basic and acidic keratin genes are clustered on human chromosomes 12 and 17, respectively. The present authors have assigned the two putative keratin gene clusters to canine chromosomes using canine cosmid clones. Successful fluorescence in situ hybridization mapped the putative cluster of canine acidic genes to dog chromosome 20 and the putative cluster of basic keratin genes to a small autosome not yet included in the partial canine standard karyotype.     

Sensitivity of the woodland herb Anemone hepatica to changing environmental conditions

Abstract. The decline of deciduous woodland populations of Anemone hepatica L. in southern Sweden is documented and possible causes are discussed. The study was based on (1) re‐investigations of 6.25 km² grid‐squares first studied in 1938–1970, (2) distribution of A. hepatica in woodland sites with well‐known soil chemical properties and (3) a detailed study over 12 consecutive years into the relationships between biological characteristics of the species (number of individuals, vegetative development, flowering frequency) and environmental variables (temperature, precipitation), soil chemistry and time. There was a close relationship between soil acidity (pH, solubility of Al³⁺) and both distribution and biological characteristics. The biological variables declined significantly over time but were not related to climatic variability. Increasing soil acidity and Al³⁺ solubility are concluded to be the main factors responsible for the decline of A. hepatica in S. Swedish deciduous woodlands.     

Rapid changes in specificity within single clones of cytolytic effector cells

We find rapid changes in the specificity of the cytolytic effector cells in a mixed lymphocyte culture. The lysis patterns produced by cytolytic effector cells generated near limiting dilution in murine mixed lymphocyte reactions of three types, F₁ anti-parent (F₁(A × B) anti-A), allogeneic (C anti-F₁(A × B)), and F₁ anti-modified parent (F₁(A × B) anti-A-TNP), were investigated. Cultures were characterized by their ability or inability to lyse a panel of target cells (e.g., A, B, F₁). When individual cultures were tested at two different times, changes in lytic pattern were routinely seen, with some patterns reproducibly increasing in frequency and others reproducibly decreasing (e.g., patterns involving lysis of F₁ decreased in an F₁ anti-A response but increased in a C anti-F₁ response). X-linked isoenzyme analysis showed that changes can occur within a single clone of effector cells. These results imply that the T cell specificity repertoire continues to evolve during an ongoing immune response, a conclusion incompatible with clonal selection theory.     

Biophysical Characterization of the Tandem FHA Domain Regulatory Module from the Mycobacterium tuberculosis ABC Transporter Rv1747

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<Emphasis Type="Italic">PLP1</Emphasis> Gene Variation Modulates Leftward and Rightward Functional Hemispheric Asymmetries

Molecular neurobiological factors determining corpus callosum physiology and anatomy have been suggested to be one of the major factors determining functional hemispheric asymmetries. Recently, it was shown that allelic variations in two myelin-related genes, the proteolipid protein 1 gene PLP1 and the contactin 1 gene CNTN1, are associated with differences in interhemispheric integration. Here, we investigated whether three single nucleotide polymorphisms that were associated with interhemispheric integration via the corpus callosum in a previous study also are relevant for functional hemispheric asymmetries. To this end, we tested more than 900 healthy adults with the forced attention dichotic listening task, a paradigm to assess language lateralization and its modulation by cognitive control processes. Moreover, we used the line bisection task, a paradigm to assess functional hemispheric asymmetries in spatial attention. We found that a polymorphism in PLP1, but not CNTN1, was associated with performance differences in both tasks. Both functional hemispheric asymmetries and their modulation by cognitive control processes were affected. These findings suggest that both left and right hemisphere dominant cognitive functions can be modulated by allelic variation in genes affecting corpus callosum structure. Moreover, higher order cognitive processes may be relevant parameters when investigating the molecular basis of hemispheric asymmetries.     

Development of a pre-glycoengineered CHO-K1 host cell line for the expression of antibodies with enhanced Fc mediated effector function

Novel biotherapeutic glycoproteins, like recombinant monoclonal antibodies (mAbs) are widely used for the treatment of numerous diseases. The N-glycans attached to the constant region of an antibody have been demonstrated to be crucial for the biological efficacy. Even minor modifications of the N-glycan structure can dictate the potency of IgG effector functions such as the antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC).

Here, we present the development of a glycoengineered CHO-K1 host cell line (HCL), stably expressing β1,4-N-Acetylglucoseaminyltransferase III (GnT-III) and α-mannosidase II (Man-II), for the expression of a-fucosylated antibodies with enhanced Fc-mediated effector function. Glycoengineered HCLs were generated in a two-step strategy, starting with generating parental HCLs by stable transfection of CHO-K1 cells with GnT-III and Man-II. In a second step, parental HCLs were stably transfected a second time with these two transgenes to increase their copy number in the genetic background. Generated glycoengineered CHO-K1 cell lines expressing two different mAbs deliver antibody products with a content of more than 60% a-fucosylated glycans. In-depth analysis of the N-glycan structure revealed that the majority of the Fc-attached glycans of the obtained mAbs were of complex bisected type. Furthermore, we showed the efficient use of FcγRIIIa affinity chromatography as a novel method for the fast assessment of the mAbs a-fucosylation level. By testing different cultivation conditions for the pre-glycoengineered recombinant CHO-K1 clones, we identified key components essential for the production of a-fucosylated mAbs. The prevalent effect could be attributed to the trace element manganese, which leads to a strong increase of a-fucosylated complex- and hybrid-type glycans. In conclusion, the novel pre-glycoengineered CHO-K1 HCL can be used for the production of antibodies with high ratios of a-fucosylated Fc-attached N-glycans. Application of our newly developed FcγRIIIa affinity chromatography method during cell line development and use of optimized cultivation conditions can ultimately support the efficient development of a-fucosylated mAbs.     

EvoSketch: Simple simulations for learning random and probabilistic processes in evolution,and effects of instructional support on learners’ conceptual knowledge

Background

Students’ knowledge of scientific principles of evolution is often inadequate, despite its recognized importance for understanding biology. Moreover, difficulties associated with underlying abstract concepts such as randomness and probability can hinder successful learning of evolutionary concepts. Studies show that visualizations, particularly simulations together with appropriate instructional support, facilitate the learning of abstract concepts. Therefore, we have developed interactive, web-based simulation software called EvoSketch in efforts to help learners grasp the nature and importance of random and probabilistic processes in evolutionary contexts. We applied EvoSketch in an intervention study comparing four self-directed study conditions: learning with EvoSketch (1) alone, (2) combined with interpretative support, (3) combined with reflective support, and (4) using texts about randomness and probability instead of EvoSketch. All conditions received no support from any instructors. Knowledge about evolution as well as randomness and probability in the context of evolution, time-on-task, and perceived cognitive load were measured. A sample of 269 German secondary school students (M_age?=?15.6 years, SD?=?0.6 years) participated in the study.

Results

Learners using EvoSketch without additional support obtained higher follow-up test scores regarding their knowledge of randomness and probability than those using the text-based approach. However, use of the simulations together with given instructional support (interpretative or reflective) did not increase students’ performance, relative to the text-based approach. In addition, no significant between-intervention differences were found concerning the knowledge of evolution, while significant differences between the groups were detected concerning students’ perceived cognitive load and time-on-task.

Conclusions

From our findings, we conclude that EvoSketch seems to have a very small positive effect on students’ understanding of randomness and probability. Contrary to our expectations, additional self-directed instructional support did not improve students’ understanding, probably because it was not necessary to understand EvoSketch simulations. When using EvoSketch in the classroom, we recommend increasing the intervention timeframe to several sessions and a variety of evolutionary examples for which EvoSketch serves as an underlying framework.

     

Structure and function of molybdopterin containing enzymes

Molybdopterin containing enzymes are present in a wide range of living systems and have been known for several decades. However, only in the past two years have the first crystal structures been reported for this type of enzyme. This has represented a major breakthrough in this field. The enzymes share common structural features, but reveal different polypeptide folding topologies. In this review we give an account of the related spectroscopic information and the crystallographic results, with emphasis on structure-function studies.