Changes in race-specific virulence in pseudomonas syringae pv. phaseolicola are Associated with a chimeric transposable element and rare deletion events in a plasmid-borne pathogenicity island

Virulence for bean and soybean is determined by effector genes in a plasmid-borne pathogenicity island (PAI) in race 7 strain 1449B of Pseudomonas syringae pv. phaseolicola. One of the effector genes, avrPphF, confers either pathogenicity, virulence, or avirulence depending on the plant host and is absent from races 2, 3, 4, 6, and 8 of this pathogen. Analysis of cosmid clones and comparison of DNA sequences showed that the absence of avrPphF from strain 1448A is due to deletion of a continuous 9.5-kb fragment. The remainder of the PAI is well conserved in strains 1448A and 1449B. The left junction of the deleted region consists of a chimeric transposable element generated from the fusion of homologs of IS1492 from Pseudomonas putida and IS1090 from Ralstonia eutropha. The borders of the deletion were conserved in 66 P. syringae pv. phaseolicola strains isolated in different countries and representing the five races lacking avrPphF. However, six strains isolated in Spain had a 10.5-kb deletion that extended 1 kb further from the right junction. The perfect conservation of the 28-nucleotide right repeat of the IS1090 homolog in the two deletion types and in the other 47 insertions of the IS1090 homolog in the 1448A genome strongly suggests that the avrPphF deletions were mediated by the activity of the chimeric mobile element. Our data strongly support a clonal origin for the races of P. syringae pv. phaseolicola lacking avrPphF.

The influence of the accessory genome on bacterial pathogen evolution

Bacterial pathogens exhibit significant variation in their genomic content of virulence factors. This reflects the abundance of strategies pathogens evolved to infect host organisms by suppressing host immunity. Molecular arms-races have been a strong driving force for the evolution of pathogenicity, with pathogens often encoding overlapping or redundant functions, such as type III protein secretion effectors and hosts encoding ever more sophisticated immune systems. The pathogens’ frequent exposure to other microbes, either in their host or in the environment, provides opportunities for the acquisition or interchange of mobile genetic elements. These DNA elements accessorise the core genome and can play major roles in shaping genome structure and altering the complement of virulence factors. Here, we review the different mobile genetic elements focusing on the more recent discoveries and highlighting their role in shaping bacterial pathogen evolution.

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.

L1-regularisation for ill-posed problems in variational data assimilation

We consider four-dimensional variational data assimilation (4DVar) and show that it can be interpreted as Tikhonov or L2-regularisation, a widely used method for solving ill-posed inverse problems. It is known from image restoration and geophysical problems that an alternative regularisation, namely L1-norm regularisation, recovers sharp edges better than L2-norm regularisation. We apply this idea to 4DVar for problems where shocks and model error are present and give two examples which show that L1-norm regularisation performs much better than the standard L2-norm regularisation in 4DVar.

Evaluating raw ensembles with the continuous ranked probability score

The continuous ranked probability score (CRPS) is a frequently used scoring rule. In contrast with many other scoring rules, the CRPS evaluates cumulative distribution functions. An ensemble of forecasts can easily be converted into a piecewise constant cumulative distribution function with steps at the ensemble members. This renders the CRPS a convenient scoring rule for the evaluation of ‘raw’ ensembles, obviating the need for sophisticated ensemble model output statistics or dressing methods prior to evaluation. In this article, a relation between the CRPS score and the quantile score is established. The evaluation of ‘raw’ ensembles using the CRPS is discussed in this light. It is shown that latent in this evaluation is an interpretation of the ensemble as quantiles but with non-uniform levels. This needs to be taken into account if the ensemble is evaluated further, for example with rank histograms.

SANS/WANS time-resolving neutron scattering studies of polymer phase transitions using NIMROD

We use new neutron scattering instrumentation to follow in a single quantitative time-resolving experiment, the three key scales of structural development which accompany the crystallisation of synthetic polymers. These length scales span 3 orders of magnitude of the scattering vector. The study of polymer crystallisation dates back to the pioneering experiments of Keller and others who discovered the chain-folded nature of the thin lamellae crystals which are normally found in synthetic polymers. The inherent connectivity of polymers makes their crystallisation a multiscale transformation. Much understanding has developed over the intervening fifty years but the process has remained something of a mystery. There are three key length scales. The chain folded lamellar thickness is ~ 10nm, the crystal unit cell is ~ 1nm and the detail of the chain conformation is ~ 0.1nm. In previous work these length scales have been addressed using different instrumention or were coupled using compromised geometries. More recently researchers have attempted to exploit coupled time-resolved small-angle and wide-angle x-ray experiments. These turned out to be challenging experiments much related to the challenge of placing the scattering intensity on an absolute scale. However, they did stimulate the possibility of new phenomena in the very early stages of crystallisation. Although there is now considerable doubt on such experiments, they drew attention to the basic question as to the process of crystallisation in long chain molecules. We have used NIMROD on the second target station at ISIS to follow all three length scales in a time-resolving manner for poly(e-caprolactone). The technique can provide a single set of data from 0.01 to 100Å-1 on the same vertical scale. We present the results using a multiple scale model of the crystallisation process in polymers to analyse the results.

Terahertz spectroscopy and imaging at the nanoscale for biological and security applications

The chemical specificity of terahertz spectroscopy, when combined with techniques for sub-wavelength sensing, is giving new understanding of processes occurring at the nanometre scale in biological systems and offers the potential for single molecule detection of chemical and biological agents and explosives. In addition, terahertz techniques are enabling the exploration of the fundamental behaviour of light when it interacts with nanoscale optical structures, and are being used to measure ultrafast carrier dynamics, transport and localisation in nanostructures. This chapter will explain how terahertz scale modelling can be used to explore the fundamental physics of nano-optics, it will discuss the terahertz spectroscopy of nanomaterials, terahertz near-field microscopy and other sub-wavelength techniques, and summarise recent developments in the terahertz spectroscopy and imaging of biological systems at the nanoscale. The potential of using these techniques for security applications will be considered.

Chapter 4: Simulation of reaction injection moulding

Reaction Injection Moulding is a technology that enables the rapid production of complex plastic parts directly from a mixture of two reactive materials of low viscosity. The reactants are mixed in specific quantities and injected into a mould. This process allows large complex parts to be produced without the need for high clamping pressures. This chapter explores the simulation of the complex processes involved in reaction injection moulding. The reaction processes mean that the dynamics of the material in the mould are in constant evolution and an effective model which takes full account of these changing dynamics is introduced and incorporated in to finite element procedures, which are able to provide a complete simulation of the cycle of mould filling and subsequent curing.

Sensitivity of Bolivian seasonally-dry tropical forest to precipitation and temperature changes over glacial-interglacial timescales

We used fossil pollen to investigate the response of the eastern Chiquitano seasonally-dry tropical forest (SDTF), lowland Bolivia, to high-amplitude climate change associated with glacial–interglacial cycles. Changes in the structure, composition and diversity of the past vegetation are compared with palaeoclimate data previously reconstructed from the same record, and these results shed light on the biogeographic history of today’s highly disjunct blocks of SDTF across South America. We demonstrate that lower glacial temperatures limited tropical forest in the Chiquitanía region, and suggest that SDTF was absent or restricted at latitudes below 17°S, the proposed location of the majority of the hypothesized ‘Pleistocene dry forest arc’ (PDFA). At 19500 yrs b.p., warming supported the establishment of a floristically-distinct SDTF, which showed little change throughout the glacial–Holocene transition, despite a shift to significantly wetter conditions beginning ca. 12500–12200 yrs b.p. Anadenanthera colubrina, a key SDTF taxon, arrived at 10000 yrs b.p., which coincides with the onset of drought associated with an extended dry season. Lasting until 3000 yrs b.p., Holocene drought caused a floristic shift to more drought-tolerant taxa and a reduction in α-diversity (shown by declining palynological richness), but closed-canopy forest was maintained throughout. In contrast to the PDFA, the modern distribution of SDTF most likely represents the greatest spatial coverage of these forests in southern South America since glacial times. We find that temperature is a key climatic control upon the distribution of lowland South American SDTF over glacial-interglacial timescales, and seasonality of rainfall exerts a strong control on their floristic composition.

Geographical variation in total and inorganic arsenic content of polished (white) rice

An extensive data set of total arsenic analysis for 901 polished (white) grain samples, originating from 10 countries from 4 continents, was compiled. The samples represented the baseline (i.e., not specifically collected from arsenic contaminated areas), and all were for market sale in major conurbations. Median total arsenic contents of rice varied 7-fold, with Egypt (0.04 mg/kg) and India (0.07 mg/kg) having the lowest arsenic content while the U.S. (0.25 mg/kg) and France (0.28 mg/kg) had the highest content. Global distribution of total arsenic in rice was modeled by weighting each country’s arsenic distribution by that country’s contribution to global production. A subset of 63 samples from Bangladesh, China, India, Italy, and the U.S. was analyzed for arsenic species. The relationship between inorganic arsenic content versus total arsenic content significantly differed among countries, with Bangladesh and India having the steepest slope in linear regression, and the U.S. having the shallowest slope. Using country-specific rice consumption data, daily intake of inorganic arsenic was estimated and the associated internal cancer risk was calculated using the U.S. Environmental Protection Agency (EPA) cancer slope. Median excess internal cancer risks posed by inorganic arsenic ranged 30-fold for the 5 countries examined, being 0.7 per 10,000 for Italians to 22 per 10,000 for Bangladeshis, when a 60 kg person was considered.