Transposon-mediated Chromsomal Integration of Transgenes in the Parasitic Nematode STrongyloids ratti and Establsihment of Stable Transgenic Lines

Genetic transformation is a potential tool for analyzing gene function and thereby identifying new drug and vaccine targets in parasitic nematodes, which adversely affect more than one billion people. We have previously developed a robust system for transgenesis in Strongyloides spp. using gonadal microinjection for gene transfer. In this system, transgenes are expressed in promoter-regulated fashion in the F1 but are silenced in subsequent generations, presumably because of their location in repetitive episomal arrays. To counteract this silencing, we explored transposon-mediated chromosomal integration of transgenes in S. ratti. To this end, we constructed a donor vector encoding green fluorescent protein (GFP) under the control of the Ss-act-2 promoter with flanking inverted tandem repeats specific for the piggyBac transposon. In three experiments, free-living Strongyloides ratti females were transformed with this donor vector and a helper plasmid encoding the piggyBac transposase. A mean of 7.9% of F1 larvae were GFP-positive. We inoculated rats with GFP-positive F1 infective larvae, and 0.5% of 6014 F2 individuals resulting from this host passage were GFP-positive. We cultured GFP-positive F2 individuals to produce GFP-positive F3 L3i for additional rounds of host and culture passage. Mean GFP expression frequencies in subsequent generations were 15.6% in the F3, 99.0% in the F4, 82.4% in the F5 and 98.7% in the F6. The resulting transgenic lines now have virtually uniform GFP expression among all progeny after at least 10 generations of passage. Chromosomal integration of the reporter transgenes was confirmed by Southern blotting and splinkerette PCR, which revealed the transgene flanked by S. ratti genomic sequences corresponding to five discrete integration sites. BLAST searches of flanking sequences against the S. ratti genome revealed integrations in five contigs. This result provides the basis for two powerful functional genomic tools in S. ratti: heritable transgenesis and insertional mutagenesis

Geometric Exponents, SLE and Logarithmic Minimal Models

In statistical mechanics, observables are usually related to local degrees of freedom such as the Q < 4 distinct states of the Q-state Potts models or the heights of the restricted solid-on-solid models. In the continuum scaling limit, these models are described by rational conformal field theories, namely the minimal models M(p,p') for suitable p, p'. More generally, as in stochastic Loewner evolution (SLE_kappa), one can consider observables related to nonlocal degrees of freedom such as paths or boundaries of clusters. This leads to fractal dimensions or geometric exponents related to values of conformal dimensions not found among the finite sets of values allowed by the rational minimal models. Working in the context of a loop gas with loop fugacity beta = -2 cos(4 pi/kappa), we use Monte Carlo simulations to measure the fractal dimensions of various geometric objects such as paths and the generalizations of cluster mass, cluster hull, external perimeter and red bonds. Specializing to the case where the SLE parameter kappa = 4p'/p is rational with p < p', we argue that the geometric exponents are related to conformal dimensions found in the infinitely extended Kac tables of the logarithmic minimal models LM(p,p'). These theories describe lattice systems with nonlocal degrees of freedom. We present results for critical dense polymers LM(1,2), critical percolation LM(2,3), the logarithmic Ising model LM(3,4), the logarithmic tricritical Ising model LM(4,5) as well as LM(3,5). Our results are compared with rigourous results from SLE_kappa, with predictions from theoretical physics and with other numerical experiments. Throughout, we emphasize the relationships between SLE_kappa, geometric exponents and the conformal dimensions of the underlying CFTs.Comment: Added reference

Fusion algebra of critical percolation

We present an explicit conjecture for the chiral fusion algebra of critical percolation considering Virasoro representations with no enlarged or extended symmetry algebra. The representations we take to generate fusion are countably infinite in number. The ensuing fusion rules are quasi-rational in the sense that the fusion of a finite number of these representations decomposes into a finite direct sum of these representations. The fusion rules are commutative, associative and exhibit an sl(2) structure. They involve representations which we call Kac representations of which some are reducible yet indecomposable representations of rank 1. In particular, the identity of the fusion algebra is a reducible yet indecomposable Kac representation of rank 1. We make detailed comparisons of our fusion rules with the recent results of Eberle-Flohr and Read-Saleur. Notably, in agreement with Eberle-Flohr, we find the appearance of indecomposable representations of rank 3. Our fusion rules are supported by extensive numerical studies of an integrable lattice model of critical percolation. Details of our lattice findings and numerical results will be presented elsewhere.Comment: 12 pages, v2: comments and references adde

A longitudinal, observational study examining the relationships of patient satisfaction with services and mental well-being to their clinical course in young people with Type 1 diabetes mellitus during transition from child to adult health services

AIM: We hypothesized that participant well-being and satisfaction with services would be positively associated with a satisfactory clinical course during transition from child to adult health care. METHODS: Some 150 young people with Type 1 diabetes mellitus from five diabetes units in England were recruited to a longitudinal study of transition. Each young person was visited at home four times by a research assistant; each visit was 1 year apart. Satisfaction with services (Mind the Gap; MTG) and mental well-being (Warwick-Edinburgh Mental Well-being Scale; WEMWBS) were captured. Change in HbA1c , episodes of ketoacidosis, clinic and retinal screening attendance were used to assess clinical course. In total, 108 of 150 (72%) young people had sufficient data for analysis at visit 4. RESULTS: Mean age at entry was 16 years. By visit 4, 81.5% had left paediatric healthcare services. Median HbA1c increased significantly (P = 0.01) from 69 mmol/mol (8.5%) at baseline to 75 mmol/mol (9.0%) at visit 4. WEMWBS scores were comparable with those in the general population at baseline and were stable over the study period. MTG scores were also stable. By visit 4, some 32 individuals had a 'satisfactory' and 76 a 'suboptimal' clinical course. There were no significant differences in average WEMWBS and MTG scores between the clinical course groups (P = 0.96, 0.52 respectively); nor was there a significant difference in transfer status between the clinical course groups. CONCLUSIONS: The well-being of young people with diabetes and their satisfaction with transition services are not closely related to their clinical course. Investigating whether innovative psycho-educational interventions can improve the clinical course is a research priority

Gauge-invariant tree-level photoproduction amplitudes with form factors

We show how the gauge-invariance formulation given by Haberzettl is implemented in practice for photoproduction amplitudes at the tree level with form factors describing composite nucleons. We demonstrate that, in contrast to Ohta's gauge-invariance prescription, this formalism allows electric current contributions to be multiplied by a form factor, i.e., it does not require that they be treated like bare currents. While different in detail, this nevertheless lends support to previous ad hoc approaches which multiply the Born amplitudes by an overall form factor. Numerical results for kaon photoproduction off the nucleon are given. They show that the gauge procedure by Haberzettl leads to much improved $\chi^2$ values as compared to Ohta's prescription.Comment: 5 pages, RevTeX, two eps figure

Peculiar Velocities of Galaxy Clusters

We investigate the peculiar velocities predicted for galaxy clusters by theories in the cold dark matter family. A widely used hypothesis identifies rich clusters with high peaks of a suitably smoothed version of the linear density fluctuation field. Their peculiar velocities are then obtained by extrapolating the similarly smoothed linear peculiar velocities at the positions of these peaks. We test these ideas using large high resolution N-body simulations carried out within the Virgo supercomputing consortium. We find that at early times the barycentre of the material which ends up in a rich cluster is generally very close to a high peak of the initial density field. Furthermore the mean peculiar velocity of this material agrees well with the linear value at the peak. The late-time growth of peculiar velocities is, however, systematically underestimated by linear theory. At the time clusters are identified we find their rms peculiar velocity to be about 40% larger than predicted. Nonlinear effects are particularly important in superclusters. These systematics must be borne in mind when using cluster peculiar velocities to estimate the parameter combination $\sigma_8\Omega^{0.6}$.Comment: 8 pages, 4 figures; submitted to MNRA

Using legume-based mixtures to enhance the nitrogen use efficiency and economic viability of cropping systems - Final report (LK09106/HGCA3447)

As costs for mineral fertilisers rise, legume-based leys are recognised as a potential alternative nitrogen source for crops. Here we demonstrate that including species-rich legume-based leys in rotations helps to maximise synergies between agricultural productivity and other ecosystem services. By using functionally diverse plant species mixtures, these services can be optimised and fine-tuned to regional and farm-specific needs. Replicated field experiments were conducted over three years at multiple locations, testing the performance of 12 legume species and 4 grass species sown in monocultures, as well as in a mixture of 10 of the legumes and all 4 grasses (called the All Species Mix, ASM). In addition, we compared this complex mixture to farmer-chosen ley mixtures on 34 sites across the UK. The trials showed that there is a large degree of functional complementarity among the legume species. No single species scored high on all evaluation criteria. In particular, the currently most frequently used species, white clover, is outscored by other legume species on a number of parameters such as early development and resistance to decomposition. Further complementarity emerged from the different responses of legume species to environmental variables, with soil pH and grazing or cutting regime being among the more important factors. For example, while large birdsfoot trefoil showed better performance on more acidic soils, the opposite was true for sainfoin, lucerne and black medic. In comparison with the monocultures, the ASM showed increased ground cover, increased above-ground biomass and reduced weed biomass. Benefits of mixing species with regard to productivity increased over time. In addition, the stability of biomass production across sites was greater in the ASM than in the legume monocultures. Within the on-farm trials, we further found that on soils low in organic matter the biomass advantage of the ASM over the Control ley was more marked than on the soils with higher organic matter content. Ecological modelling revealed that the three best multifunctional mixtures all contained black medic, lucerne and red clover. Within the long term New Farming Systems (NFS) rotational study, the use of a clover bi-crop showed improvement to soil characteristics compared to current practice (e.g. bulk density and water infiltration rate). Improvements in wheat yield were also noted with respect to the inclusion of a clover bi-crop in 2010, but there was evidence of a decline in response as the N dose was increased. Cumulatively, over both the wheat crop and the spring oilseed rape crop, the clover bi-crop improved margin over N. The highest average yield response (~9%) was associated with the ASM legume species mix cover cropping approach