Development and validation of a qPCR-based method for quantifying Shiga toxin-encoding and other lambdoid bacteriophages

P>To address whether seasonal variability exists among Shiga toxin-encoding bacteriophage (Stx phage) numbers on a cattle farm, conventional plaque assay was performed on water samples collected over a 17 month period. Distinct seasonal variation in bacteriophage numbers was evident, peaking between June and August. Removal of cattle from the pasture precipitated a reduction in bacteriophage numbers, and during the winter months, no bacteriophage infecting Escherichia coli were detected, a surprising occurrence considering that 1031 tailed-bacteriophages are estimated to populate the globe. To address this discrepancy a culture-independent method based on quantitative PCR was developed. Primers targeting the Q gene and stx genes were designed that accurately and discriminately quantified artificial mixed lambdoid bacteriophage populations. Application of these primer sets to water samples possessing no detectable phages by plaque assay, demonstrated that the number of lambdoid bacteriophage ranged from 4.7 x 104 to 6.5 x 106 ml-1, with one in 103 free lambdoid bacteriophages carrying a Shiga toxin operon (stx). Specific molecular biological tools and discriminatory gene targets have enabled virus populations in the natural environment to be enumerated and similar strategies could replace existing propagation-dependent techniques, which grossly underestimate the abundance of viral entities.

Development and application of a method for the purification of free shigatoxigenic bacteriophage from environmental samples

Shiga toxin producing Escherichia coli (STEC) strains are foodborne pathogens whose ability to produce Shiga toxin (Stx) is due to the integration of Stx-encoding lambdoid bacteriophage (Stx phage). Circulating, infective Stx phages are very difficult to isolate, purify and propagate such that there is no information on their genetic composition and properties. Here we describe a novel approach that exploits the phage's ability to infect their host and form a lysogen, thus enabling purification of Stx phages by a series of sequential lysogen isolation and induction steps. A total of 15 Stx phages were rigorously purified from water samples in this way, classified by TEM and genotyped using a PCR-based multi-loci characterisation system. Each phage possessed only one variant of each target gene type, thus confirming its purity, with 9 of the 15 phages possessing a short tail-spike gene and identified by TEM as Podoviridae. The remaining 6 phages possessed long tails, four of which appeared to be contractile in nature (Myoviridae) and two of which were morphologically very similar to bacteriophage lambda (Siphoviridae).

Seed development and maturation in early spring-flowering Galanthus nivalis and Narcissus pseudonarcissus continues post-shedding with little evidence of maturation in planta

Background and Aims: Seeds of the moist temperate woodland species Galanthus nivalis and Narcissus pseudonarcissus, dispersed during spring or early summer, germinated poorly in laboratory tests. Seed development and maturation were studied to better understand the progression from developmental to germinable mode in order to improve seed collection and germination practices in these and similar species. Methods: Phenology, seed mass, moisture content, and ability to germinate and tolerate desiccation were monitored during seed development until shedding. Embryo elongation within seeds was investigated during seed development and at several temperature regimes after shedding. Key Results: Seeds were shed at high moisture content (> 59%) with little evidence that dry mass accumulation or embryo elongation were complete. Ability to germinate developed prior to the ability of some seeds to tolerate enforced desiccation. Germination was sporadic and slow. Embryo elongation occurred post-shedding in moist environments, most rapidly at 20C in G. nivalis and 15C in N. pseudonarcissus. The greatest germination also occurred in these regimes, 78 and 48%, respectively, after 700 d. Conclusions: Seeds of G. nivalis and N. pseudonarcissus seeds were comparatively immature at shedding and substantial embryo elongation occurred post-shedding. Seeds showed limited desiccation tolerance at dispersal.

Multi-year droughts in Europe: analysis of development and causes

Whilst hydrological systems can show resilience to short-term streamflow deficiencies during within-year droughts, prolonged deficits during multi-year droughts are a significant threat to water resources security in Europe. This study uses a threshold-based objective classification of regional hydrological drought to qualitatively examine the characteristics, spatio-temporal evolution and synoptic climatic drivers of multi-year drought events in 1962–64, 1975–76 and 1995–97, on a European scale but with particular focus on the UK. Whilst all three events are multi-year, pan-European phenomena, their development and causes can be contrasted. The critical factor in explaining the unprecedented severity of the 1975–76 event is the consecutive occurrence of winter and summer drought. In contrast, 1962–64 was a succession of dry winters, mitigated by quiescent summers, whilst 1995–97 lacked spatial coherence and was interrupted by wet interludes. Synoptic climatic conditions vary within and between multi-year droughts, suggesting that regional factors modulate the climate signal in streamflow drought occurrence. Despite being underpinned by qualitatively similar climatic conditions and commonalities in evolution and characteristics, each of the three droughts has a unique spatio-temporal signature. An improved understanding of the spatio-temporal evolution and characteristics of multi-year droughts has much to contribute to monitoring and forecasting capability, and to improved mitigation strategies.

A SEPALLATA gene is involved in the development and ripening of strawberry (Fragaria X ananassa Duch.) fruit, a non-climacteric tissue

Climacteric and non-climacteric fruits have traditionally been viewed as representing two distinct programmes of ripening associated with differential respiration and ethylene hormone effects. In climacteric fruits, such as tomato and banana, the ripening process is marked by increased respiration and is induced and co-ordinated by ethylene, while in non-climacteric fruits, such as strawberry and grape, it is controlled by an ethylene-independent process with little change in respiration rate. The two contrasting mechanisms, however, both lead to texture, colour, and flavour changes that probably reflect some common programmes of regulatory control. It has been shown that a SEPALLATA(SEP)4-like gene is necessary for normal ripening in tomato. It has been demonstrated here that silencing a fruit-related SEP1/2-like (FaMADS9) gene in strawberry leads to the inhibition of normal development and ripening in the petal, achene, and receptacle tissues. In addition, analysis of transcriptome profiles reveals pleiotropic effects of FaMADS9 on fruit development and ripening-related gene expression. It is concluded that SEP genes play a central role in the developmental regulation of ripening in both climacteric and non-climacteric fruits. These findings provide important information to extend the molecular control of ripening in a non-climacteric fruit beyond the limited genetic and cultural options currently available.

The influence of vegetation and building morphology on shadow patterns and mean radiant temperatures in urban areas: model development and evaluation

The solar and longwave environmental irradiance geometry (SOLWEIG) model simulates spatial variations of 3-D radiation fluxes and mean radiant temperature (T mrt) as well as shadow patterns in complex urban settings. In this paper, a new vegetation scheme is included in SOLWEIG and evaluated. The new shadow casting algorithm for complex vegetation structures makes it possible to obtain continuous images of shadow patterns and sky view factors taking both buildings and vegetation into account. For the calculation of 3-D radiation fluxes and T mrt, SOLWEIG only requires a limited number of inputs, such as global shortwave radiation, air temperature, relative humidity, geographical information (latitude, longitude and elevation) and urban geometry represented by high-resolution ground and building digital elevation models (DEM). Trees and bushes are represented by separate DEMs. The model is evaluated using 5 days of integral radiation measurements at two sites within a square surrounded by low-rise buildings and vegetation in Göteborg, Sweden (57°N). There is good agreement between modelled and observed values of T mrt, with an overall correspondence of R 2 = 0.91 (p < 0.01, RMSE = 3.1 K). A small overestimation of T mrt is found at locations shadowed by vegetation. Given this good performance a number of suggestions for future development are identified for applications which include for human comfort, building design, planning and evaluation of instrument exposure.

Development and evaluation of an Earth-System model – HadGEM2

We describe here the development and evaluation of an Earth system model suitable for centennial-scale climate prediction. The principal new components added to the physical climate model are the terrestrial and ocean ecosystems and gas-phase tropospheric chemistry, along with their coupled interactions. The individual Earth system components are described briefly and the relevant interactions between the components are explained. Because the multiple interactions could lead to unstable feedbacks, we go through a careful process of model spin up to ensure that all components are stable and the interactions balanced. This spun-up configuration is evaluated against observed data for the Earth system components and is generally found to perform very satisfactorily. The reason for the evaluation phase is that the model is to be used for the core climate simulations carried out by the Met Office Hadley Centre for the Coupled Model Intercomparison Project (CMIP5), so it is essential that addition of the extra complexity does not detract substantially from its climate performance. Localised changes in some specific meteorological variables can be identified, but the impacts on the overall simulation of present day climate are slight. This model is proving valuable both for climate predictions, and for investigating the strengths of biogeochemical feedbacks.

The role of location in knowledge creation and diffusion: evidence of centripetal and centrifugal forces in the City of London financial services agglomeration

In this paper we report on a major empirical study of centripetal and centrifugal forces in the City of London financial services agglomeration. The study sheds light on (1) the manner and magnitude of firm interaction in the agglomeration; (2) the characteristics of the agglomeration that aid the competitiveness of incumbent firms; and (3) the problems associated with agglomeration. In addressing these issues, we use the data to (1) test emerging theory that explains the high productivity and innovation of agglomerations in terms of their ability to generate and diffuse knowledge; and (2) evaluate the ‘end of geography’ thesis.

Strong armies, slow adaptation: civil-military relations and diffusion of military power

Why are some states more willing to adopt military innovations than others? Why, for example, were the great powers of Europe able to successfully reform their military practices to better adapt to and participate in the so-called military revolution of the sixteenth and seventeenth centuries while their most important extra-European competitor, the Ottoman Empire, failed to do so? This puzzle is best explained by two factors: civil-military relations and historical timing. In the Ottoman Empire, the emergence of an institutionally strong and internally cohesive army during the early stages of state formation—in the late fourteenth century—equipped the military with substantial bargaining powers. In contrast, the great powers of Europe drew heavily on private providers of military power during the military revolution and developed similar armies only by the second half of the seventeenth century, limiting the bargaining leverage of European militaries over their rulers. In essence, the Ottoman standing army was able to block reform efforts that it believed challenged its parochial interests. Absent a similar institutional challenge, European rulers initiated military reforms and motivated officers and military entrepreneurs to participate in the ongoing military revolution.

Development and validation of a parent report measure for detection of cognitive delay in infancy

Aim To develop a brief, parent-completed instrument (‘ERIC’) for detection of cognitive delay in 10-24 month-olds born preterm, or with low birth weight, or with perinatal complications, and to establish its diagnostic properties. Method Scores were collected from parents of 317 children meeting ≥1 inclusion criteria (birth weight <1500g; gestational age <34 completed weeks; 5-minute Apgar <7; presence of hypoxic-ischemic encephalopathy) and meeting no exclusion criteria. Children were assessed for cognitive delay using a criterion score on the Bayley Scales of Infant and Toddler Development Cognitive Scale III1 <80. Items were retained according to their individual associations with delay. Sensitivity, specificity, Positive and Negative Predictive Values were estimated and a truncated ERIC was developed for use <14 months. Results ERIC detected 17 out of 18 delayed children in the sample, with 94.4% sensitivity (95% CI [confidence interval] 83.9-100%), 76.9% specificity (72.1-81.7%), 19.8% positive predictive value (11.4-28.2%); 99.6% negative predictive value (98.7-100%); 4.09 likelihood ratio positive; and 0.07 likelihood ratio negative; the associated Area under the Curve was .909 (.829-.960). Interpretation ERIC has potential value as a quickly-administered diagnostic instrument for the absence of early cognitive delay in preterm or premature infants of 10-24 months, and as a screen for cognitive delay. Further research may be needed before ERIC can be recommended for wide-scale use.

Development and exploitation of a controlled vocabulary in support of climate modelling

There are three key components for developing a metadata system: a container structure laying out the key semantic issues of interest and their relationships; an extensible controlled vocabulary providing possible content; and tools to create and manipulate that content. While metadata systems must allow users to enter their own information, the use of a controlled vocabulary both imposes consistency of definition and ensures comparability of the objects described. Here we describe the controlled vocabulary (CV) and metadata creation tool built by the METAFOR project for use in the context of describing the climate models, simulations and experiments of the fifth Coupled Model Intercomparison Project (CMIP5). The CV and resulting tool chain introduced here is designed for extensibility and reuse and should find applicability in many more projects.