Identification and characterisation of five novel miniature inverted-repeat transposable elements (MITEs) in amphioxus (Branchiostoma floridae)

As the sister group to vertebrates, amphioxus is consistently used as a model of genome evolution for understanding the invertebrate/vertebrate transition. The amphioxus genome has not undergone massive duplications like those in the vertebrates or disruptive rearrangements like in the genome of Ciona, a urochordate, making it an ideal evolutionary model. Transposable elements have been linked to many genomic evolutionary changes including increased genome size, modified gene expression, massive gene rearrangements, and possibly intron evolution. Despite their importance in genome evolution, few previous examples of transposable elements have been identified in amphioxus. We report five novel Miniature Inverted-repeat Transposable Elements (MITEs) identified by an analysis of amphioxus DNA sequence, which we have named LanceleTn-1, LanceleTn-2, LanceleTn-3a, LanceleTn-3b and LanceleTn-4. Several of the LanceleTn elements were identified in the amphioxus ParaHox cluster, and we suggest these have had important implications for the evolution of this highly conserved gene cluster. The estimated high copy numbers of these elements implies that MITEs are probably the most abundant type of mobile element in amphioxus, and are thus likely to have been of fundamental importance in shaping the evolution of the amphioxus genome.

Miniature transposable sequences are frequently mobilized in the bacterial plant pathogen Pseudomonas syringae pv. phaseolicola

Mobile genetic elements are widespread in Pseudomonas syringae, and often associate with virulence genes. Genome reannotation of the model bean pathogen P. syringae pv. phaseolicola 1448A identified seventeen types of insertion sequences and two miniature inverted-repeat transposable elements (MITEs) with a biased distribution, representing 2.8% of the chromosome, 25.8% of the 132-kb virulence plasmid and 2.7% of the 52-kb plasmid. Employing an entrapment vector containing sacB, we estimated that transposition frequency oscillated between 2.661025 and 1.161026, depending on the clone, although it was stable for each clone after consecutive transfers in culture media. Transposition frequency was similar for bacteria grown in rich or minimal media, and from cells recovered from compatible and incompatible plant hosts, indicating that growth conditions do not influence transposition in strain 1448A. Most of the entrapped insertions contained a full-length IS801 element, with the remaining insertions corresponding to sequences smaller than any transposable element identified in strain 1448A, and collectively identified as miniature sequences. From these, fragments of 229, 360 and 679-nt of the right end of IS801 ended in a consensus tetranucleotide and likely resulted from one-ended transposition of IS801. An average 0.7% of the insertions analyzed consisted of IS801 carrying a fragment of variable size from gene PSPPH_0008/PSPPH_0017, showing that IS801 can mobilize DNA in vivo. Retrospective analysis of complete plasmids and genomes of P. syringae suggests, however, that most fragments of IS801 are likely the result of reorganizations rather than one-ended transpositions, and that this element might preferentially contribute to genome flexibility by generating homologous regions of recombination. A further miniature sequence previously found to affect host range specificity and virulence, designated MITEPsy1 (100-nt), represented an average 2.4% of the total number of insertions entrapped in sacB, demonstrating for the first time the mobilization of a MITE in bacteria.

Where theory meets practice: a critical comparison of research into identifying letters and craft knowledge of type design

This paper introduces scientific research findings and accounts of skilled design judgement to: (i) develop an interdisciplinary account of what affects our identification of letters when reading; (ii) analyse the relationship between the approaches of psychologists and designers to explaining how we identify letters; (iii) propose ways in which collaboration may work to make psychological research more relevant to typographic practice. The topics reviewed are addressed within each discipline and cover the contribution of letters and words to reading; letter features; essential or structural forms; uniformity within font design; and letter spacing. Analysis of the literature identifies possible means of reconciling different perspectives, points out some anomalies in interpretation of findings, and proposes how designers may contribute to research planning and dissemination.

Morgawr: an experimental Bronze Age-type sewn-plank craft based on the Ferriby boats

This paper reports on the construction of a full-scale Bronze Age-type sewn-plank boat based on the Ferriby boats. The boat, which was named Morgawr, was constructed in the National Maritime Museum Cornwall in Falmouth, England, during 2012 and the first months of 2013, as part of a larger exhibition in the museum. This paper provides the background and context of the project, describes the process of building the craft, and reflects in particular on differences between Morgawr and the ‘hypothetical reconstruction of a complete sewn-plank boat’ published in 1990 by Ted Wright and John Coates which formed the basis for this project.

The phytocannabinoid Delta(9)-tetrahydrocannabivarin modulates inhibitory neurotransmission in the cerebellum

Background and purposeThe phytocannabinoid Delta(9)-tetrahydrocannabivarin (Delta(9)-THCV) has been reported to exhibit a diverse pharmacology; here, we investigate functional effects of Delta(9)-THCV, extracted from Cannabis sativa, using electrophysiological techniques to define its mechanism of action in the CNS.Experimental approachEffects of Delta(9)-THCV and synthetic cannabinoid agents on inhibitory neurotransmission at interneurone-Purkinje cell (IN-PC) synapses were correlated with effects on spontaneous PC output using single-cell and multi-electrode array (MEA) electrophysiological recordings respectively, in mouse cerebellar brain slices in vitro.Key resultsThe cannabinoid receptor agonist WIN 55,212-2 (WIN55) decreased miniature inhibitory postsynaptic current (mIPSC) frequency at IN-PC synapses. WIN55-induced inhibition was reversed by Delta(9)-THCV, and also by the CB(1) receptor antagonist AM251; Delta(9)-THCV or AM251 acted to increase mIPSC frequency beyond basal values. When applied alone, Delta(9)-THCV, AM251 or rimonabant increased mIPSC frequency. Pre-incubation with Delta(9)-THCV blocked WIN55-induced inhibition. In MEA recordings, WIN55 increased PC spike firing rate; Delta(9)-THCV and AM251 acted in the opposite direction to decrease spike firing. The effects of Delta(9)-THCV and WIN55 were attenuated by the GABA(A) receptor antagonist bicuculline methiodide.Conclusions and implicationsWe show for the first time that Delta(9)-THCV acts as a functional CB(1) receptor antagonist in the CNS to modulate inhibitory neurotransmission at IN-PC synapses and spontaneous PC output. Delta(9)-THCV- and AM251-induced increases in mIPSC frequency beyond basal levels were consistent with basal CB(1) receptor activity. WIN55-induced increases in PC spike firing rate were consistent with synaptic disinhibition; whilst Delta(9)-THCV- and AM251-induced decreases in spike firing suggest a mechanism of PC inhibition.British Journal of Pharmacology advance online publication, 3 March 2008; doi:10.1038/bjp.2008.57.

Investigation of the effects of the novel anticonvulsant compound carisbamate(RWJ-333369) on rat piriform cortical neurones in vitro

Background and purpose: Carisbamate is being developed for adjuvant treatment of partial onset epilepsy. Carisbamate produces anticonvulsant effects in primary generalized, complex partial and absence-type seizure models, and exhibits neuroprotective and antiepileptogenic properties in rodent epilepsy models. Phase IIb clinical trials of carisbamate demonstrated efficacy against partial onset seizures; however, its mechanisms of action remain unknown. Here, we report the effects of carisbamate on membrane properties, evoked and spontaneous synaptic transmission and induced epileptiform discharges in layer II-III neurones in piriform cortical brain slices. Experimental approach: Effects of carisbamate were investigated in rat piriform cortical neurones by using intracellular electrophysiological recordings. Key results: Carisbamate (50–400 mmol·L-1) reversibly decreased amplitude, duration and rise-time of evoked action potentials and inhibited repetitive firing, consistent with use-dependent Na+ channel block; 150–400 mmol·L-1 carisbamate reduced neuronal input resistance, without altering membrane potential. After microelectrode intracellular Cl- loading, carisbamate depolarized cells, an effect reversed by picrotoxin. Carisbamate (100–400 mmol·L-1) also selectively depressed lateral olfactory tract-afferent evoked excitatory synaptic transmission (opposed by picrotoxin), consistent with activation of a presynaptic Cl conductance. Lidocaine (40–320 mmol·L-1) mimicked carisbamate, implying similar modes of action. Carisbamate (300–600 mmol·L-1) had no effect on spontaneous GABAA miniature inhibitory postsynaptic currents and at lower concentrations (50–200 mmol·L-1) inhibited Mg2+-free or 4-aminopyridine-induced seizure-like discharges. Conclusions and implications: Carisbamate blocked evoked action potentials use-dependently, consistent with a primary action on Na+ channels and increased Cl- conductances presynaptically and, under certain conditions, postsynaptically to selectively depress excitatory neurotransmission in piriform cortical layer Ia-afferent terminals.

G-protein-coupled-receptor-mediated presynaptic inhibition in the cerebellum

Throughout the central nervous system a dominant form of inhibition of neurotransmitter release from presynaptic terminals is mediated by G-protein-coupled receptors (GPCRs). Neurotransmitter release is typically induced by action potentials (APs), but can also occur spontaneously. Presynaptic inhibition by GPCRs has been associated with modulation of voltage-dependent ion channels. However, electrophysiological recordings of spontaneous, AP-independent (so-called ‘miniature’) postsynaptic events reveal an additional, important form of GPCR-mediated presynaptic inhibition, distinct from effects on ionic conductances and consistent with a direct action on the vesicle release machinery. Recent studies suggest that such miniature events might be of physiological relevance not only in signalling but also in development. In the cerebellum, neurotransmitter release onto Purkinje cells occurs by AP-dependent and AP-independent pathways. Here, I focus on inhibitory synapses between interneurons and Purkinje cells, which are subject to strong, identifiable regulation by endogenous GPCR agonists, to consider mechanisms of GPCR-mediated presynaptic inhibition.

Genomic and genetic analyses of diversity and plant interactions of Pseudomonas fluorescens

Background: Pseudomonas fluorescens are common soil bacteria that can improve plant health through nutrient cycling, pathogen antagonism and induction of plant defenses. The genome sequences of strains SBW25 and Pf0-1 were determined and compared to each other and with P. fluorescens Pf-5. A functional genomic in vivo expression technology (IVET) screen provided insight into genes used by P. fluorescens in its natural environment and an improved understanding of the ecological significance of diversity within this species. Results: Comparisons of three P. fluorescens genomes (SBW25, Pf0-1, Pf-5) revealed considerable divergence: 61% of genes are shared, the majority located near the replication origin. Phylogenetic and average amino acid identity analyses showed a low overall relationship. A functional screen of SBW25 defined 125 plant-induced genes including a range of functions specific to the plant environment. Orthologues of 83 of these exist in Pf0-1 and Pf-5, with 73 shared by both strains. The P. fluorescens genomes carry numerous complex repetitive DNA sequences, some resembling Miniature Inverted-repeat Transposable Elements (MITEs). In SBW25, repeat density and distribution revealed 'repeat deserts' lacking repeats, covering approximately 40% of the genome. Conclusions: P. fluorescens genomes are highly diverse. Strain-specific regions around the replication terminus suggest genome compartmentalization. The genomic heterogeneity among the three strains is reminiscent of a species complex rather than a single species. That 42% of plant-inducible genes were not shared by all strains reinforces this conclusion and shows that ecological success requires specialized and core functions. The diversity also indicates the significant size of genetic information within the Pseudomonas pan genome.

Quantitative thermal imaging analysis of aircraft materials: through skin sensing (Invited Paper)

Thermal non-destructive testing (NDT) is commonly used for assessing aircraft structures. This research work evaluates the potential of pulsed -- transient thermography for locating fixtures beneath aircraft skins in order to facilitate accurate automated assembly operations. Representative aluminium and carbon fibre aircraft skin-fixture assemblies were modelled using thermal modelling software. The assemblies were also experimentally investigated with an integrated pulsed thermographic evaluation system, as well as using a custom built system incorporating a miniature un-cooled camera. Modelling showed that the presence of an air gap between skin and fixture significantly reduced the thermal contrast developed, especially in aluminium. Experimental results show that fixtures can be located to accuracies of 0.5 mm.

Visualizing knowledge in project-based work

This article considers how visual practices are used to manage knowledge in project-based work. It compares project-based work in a capital goods manufacturer and an architectural firm. Visual representations are used extensively in both cases, but the nature of visual practice differs significantly between the two. The research explores the kinds of knowledge that are (and aren't) developed and made visible in strategizing and planning activities. For example, whereas the emphasis of project-based work in the former firm is on exploitation of knowledge and it visualizes its project context largely in commercial and processual terms, the emphasis in the latter is on exploration and it uses a wide range of visual materials to understand physical interdependencies across the project boundary. We contend particular kinds of visual tools can help project teams step between exploration and exploitation within a project, and articulate the types of representations, foci of attention and patterns of interaction involved. The findings suggest that business managers can make more deliberate choices about how knowledge is made visible, and can change visual practice to align the project with exploring and exploiting opportunities. It raises the question: What don't you see within your organization? The work contributes to academic debates about managing through projects, strategising and organizing, while the focus on visual representation disrupts the tacit-codified dichotomy in the broad debate on knowledge and learning, and highlights the craft skills central to strategizing and organizing.

Reducing fouling during UHT treatment of goat's milk

Whole fresh goat's milk was heat treated at 135 degrees C for 4 s using a miniature UHT plant. The temperature of the milk in the preheating and sterilizer sections, and the milk flow rate were monitored to evaluate the overall heat transfer coefficient (OHTC). The decrease in OHTC was used to estimate the extent of fouling. Goat's milk fouled very quickly and run times of the UHT plant were short. The use of sodium hexametaphosphate, trisodium citrate and cation exchange resins to reduce ionic calcium prior to UHT processing, increased the pH and alcohol stability of the milk and markedly increased the run time of the UHT plant.

In vitro fermentation by human fecal microflora of wheat arabinoxylans

The fermentation of three arabinoxylan (AX) fractions from wheat by the human fecal microflora was investigated in vitro. Three AX fractions, with average molecular masses of 354, 278, and 66 kDa, were incorporated into miniature-scale batch cultures (with inulin as a positive prebiotic control) with feces from three healthy donors, aged 23-29. Microflora changes were monitored by the culture-independent technique, fluorescent in situ hybridization, and short chain fatty acid (SCFA) and lactic acid production were measured by high-performance liquid chromatography. Total cell numbers increased significantly in all treated cultures, and the fermentation of AX was associated with a proliferation of the bifidobacteria, lactobacilli, and eubacteria groups. Smaller but statistically significant increases in bacteroides and clostridia groups were also observed. All AX fractions had comparable bifidogenic impacts on the microflora at 5 and 12 h, but the 66 kDa AX was particularly selective for lactobacilli. Eubacteria increased significantly on all AX fractions, particularly on 66 kDa AX. As previously reported, inulin gave a selective increase in bifidobacteria. All supplemented cultures showed significant rises in total SCFA production, with a particularly high proportion of butyric acid being produced from AX fermentation. The prebiotic effect, that is, the selectivity of AX for bifidobacteria and lactobacilli groups, increased as the molecular mass of the AX decreased. This suggests that molecular mass may influence the fermentation of AX in the colon.