Combined use of empirical data and mathematical modelling to better estimate the microbial turnover of isotopically labelled carbon substrates in soil

The flow of carbon (C) through soil is inherently complex due to the many thousands of different chemical transformations occurring simultaneously within the soil microbial community. The accurate modelling of this C flow therefore represents a major challenge. In response to this, isotopic tracers (e.g. 13C, 14C) are commonly used to experimentally parameterise models describing the fate and residence time of individual C compounds within soil. In this study, we critically evaluated the combined use of experimental 14C labelling and mathematical modelling to estimate C turnover times in soil. We applied 14C-labelled alanine and glucose to an agricultural soil and simultaneously measured their loss from soil solution alongside the rate of microbial C immobilization and mineralization. Our results revealed that chloroform fumigation-extraction (CFE) cannot be used to reliably quantify the amount of isotopically labelled 13C/14C immobilised by the microbial biomass. This is due to uncertainty in the extraction efficiency values (kec) within the CFE methodology which are both substrate and incubation time dependent. Further, the traditional mineralization approach (i.e. measuring 14/13CO2 evolution) provided a poor estimate of substrate loss from soil solution and mainly reflected rates of internal microbial C metabolism after substrate uptake from the soil. Therefore, while isotope addition provides a simple mechanism for labelling the microbial biomass it provides limited information on the behaviour of the substrate itself. We used our experimental data to construct a new empirical model to describe the simultaneous flow of substrate-C between key C pools in soil. This model provided a superior estimate of microbial substrate use and microbial respiration flux in comparison to traditional first order kinetic modelling approaches. We also identify a range of fundamental problems associated with the modelling of isotopic-C in soil, including issues with variation in C partitioning within the community, model pool connectivity and variation in isotopic pool dilution, which make interpretation of any C isotopic flux data difficult. We conclude that while convenient, the use of isotopic data (13C, 14C, 15N) has many potential pitfalls necessitating a critical evaluation of both past and future studies

Response of the Gypsy Moth, Lymantria dispar to Transgenic Poplar, Populus simonii x P. nigra, Expressing Fusion Protein Gene of the Spider Insecticidal Peptide and Bt-toxin C-peptide

The response of the Asian gypsy moth Lymantria dispar (L.) (Lepidoptera: Lymantriidae) to a fusion gene consisting of the spider, Atrax robustus Simon (Araneae: Hexanthelidae) ω?-ACTX-Ar1 sequence coding for an ω?-atracotoxin and a sequence coding for the Bt-toxin C-peptide, expressed in transgenic poplar Populus simonii x P. nigra L. (Malphigiales: Salicaceae) was investigated. Individual performance, feeding selection, midgut proteinase activity and nutrition utilization were monitored. The growth and development of L. dispar were significantly affected by continually feeding on the transgenic poplar, with the larval instars displaying significantly shorter developmental times than those fed on nontransgenic poplar, but pupation was delayed. Mortality was higher in populations fed transgenic poplar leaves, than for larvae fed nontransgenic poplar leaves. The cumulative mortality during all stages of larvae fed transgenic leaves was 92% compared to 16.7% of larvae on nontransgenic leaves. The highest mortality observed was 71.7% in the last larval instar stage. A two-choice test showed that fifth-instar larvae preferred to feed on nontransgenic leaves at a ratio of 1:1.4. Feeding on transgenic leaves had highly significant negative effects on relative growth of larvae, and the efficiency of conversion of ingested and digested food. Activity of major midgut proteinases was measured using substrates TAME and BTEE showed significant increases in tryptase and chymotrypsinlike activity (9.2- and 9.0-fold, respectively) in fifth-instar larvae fed on transgenic leaves over control. These results suggest transgenic poplar is resistant to L. dispar, and the mature L. dispar may be weakened by the transgenic plants due to Bt protoxins activated by elevated major midgut proteinase activity. The new transgenic poplar expressing fusion protein genes of Bt and a new spider insecticidal peptide are good candidates for managing gypsy moth

In vitro template-change PCR to create single crossover libraries: a case study with B. thuringiensis Cry2A toxins

During evolution the creation of single crossover chimeras between duplicated paralogous genes is a known process for increasing diversity. Comparing the properties of homologously recombined chimeras with one or two crossovers is also an efficient strategy for analyzing relationships between sequence variation and function. However, no well-developed in vitro method has been established to create single-crossover libraries. Here we present an in vitro template-change polymerase change reaction that has been developed to enable the production of such libraries. We applied the method to two closely related toxin genes from B. thuringiensis and created chimeras with differing properties that can help us understand how these toxins are able to differentiate between insect species

The Influence of Computational Mesh on the Prediction of Vortex Interactions about a Generic Missile Airframe

A research program has been underway for four years to study vortex interaction aerodynamics that are relevant to military air vehicle performance. The program has been conducted under the auspices of the NATO Science and Technology Organization (STO), Applied Vehicle Technology (AVT) panel by a Task Group with the identification of AVT-316. The Missile Facet of this group has concentrated their work on the vortical flow field around a generic missile airframe and its prediction via computational methods. This paper focuses on mesh-related effects and RANS simulations. Simulated vortex characteristics were found to depend strongly on the properties of the employed mesh, in terms of both resolution and topology. Predictions of missile aerodynamic coefficients show a great dependence on mesh properties as they are sensitive to computed vortex dynamics. Key suggestions about the desired mesh characteristics have been made. Based on these, a shared mesh was constructed to perform common analyses between the AVT-316 Missile Facet members. Mesh based uncertainties of the aerodynamic coefficient predictions were estimated via Richardson Extrapolation method

Constraints on the χ_(c1) versus χ_(c2) polarizations in proton-proton collisions at √s = 8 TeV

The polarizations of promptly produced χ_(c1) and χ_(c2) mesons are studied using data collected by the CMS experiment at the LHC, in proton-proton collisions at √s=8  TeV. The χ_c states are reconstructed via their radiative decays χ_c → J/ψγ, with the photons being measured through conversions to e⁺e⁻, which allows the two states to be well resolved. The polarizations are measured in the helicity frame, through the analysis of the χ_(c2) to χ_(c1) yield ratio as a function of the polar or azimuthal angle of the positive muon emitted in the J/ψ → μ⁺μ⁻ decay, in three bins of J/ψ transverse momentum. While no differences are seen between the two states in terms of azimuthal decay angle distributions, they are observed to have significantly different polar anisotropies. The measurement favors a scenario where at least one of the two states is strongly polarized along the helicity quantization axis, in agreement with nonrelativistic quantum chromodynamics predictions. This is the first measurement of significantly polarized quarkonia produced at high transverse momentum

Canada-U.S. Relations

This report provides a short overview of Canada's political scene, its economic conditions, and its recent security and foreign policy, focusing particularly on issues that may be relevant to U.S. policymakers. This brief country survey is followed by several summaries of current bilateral issues in the political, trade, and environmental arenas

Protist-Type Lysozymes of the Nematode Caenorhabditis elegans Contribute to Resistance against Pathogenic Bacillus thuringiensis

Pathogens represent a universal threat to other living organisms. Most organisms express antimicrobial proteins and peptides, such as lysozymes, as a protection against these challenges. The nematode Caenorhabditis elegans harbours 15 phylogenetically diverse lysozyme genes, belonging to two distinct types, the protist- or Entamoeba-type (lys genes) and the invertebrate-type (ilys genes) lysozymes. In the present study we characterized the role of several protist-type lysozyme genes in defence against a nematocidal strain of the Gram-positive bacterium Bacillus thuringiensis. Based on microarray and subsequent qRT-PCR gene expression analysis, we identified protist-type lysozyme genes as one of the differentially transcribed gene classes after infection. A functional genetic analysis was performed for three of these genes, each belonging to a distinct evolutionary lineage within the protist-type lysozymes (lys-2, lys-5, and lys-7). Their knock-out led to decreased pathogen resistance in all three cases, while an increase in resistance was observed when two out of three tested genes were overexpressed in transgenic lines (lys-5, lys-7, but not lys-2). We conclude that the lysozyme genes lys-5, lys-7, and possibly lys-2 contribute to resistance against B. thuringiensis, thus highlighting the particular role of lysozymes in the nematode's defence against pathogens

Genome Characteristics of a Novel Phage from Bacillus thuringiensis Showing High Similarity with Phage from Bacillus cereus

Bacillus thuringiensis is an important entomopathogenic bacterium belongs to the Bacillus cereus group, which also includes B. anthracis and B. cereus. Several genomes of phages originating from this group had been sequenced, but no genome of Siphoviridae phage from B. thuringiensis has been reported. We recently sequenced and analyzed the genome of a novel phage, BtCS33, from a B. thuringiensis strain, subsp. kurstaki CS33, and compared the gneome of this phage to other phages of the B. cereus group. BtCS33 was the first Siphoviridae phage among the sequenced B. thuringiensis phages. It produced small, turbid plaques on bacterial plates and had a narrow host range. BtCS33 possessed a linear, double-stranded DNA genome of 41,992 bp with 57 putative open reading frames (ORFs). It had a typical genome structure consisting of three modules: the “late” region, the “lysogeny-lysis” region and the “early” region. BtCS33 exhibited high similarity with several phages, B. cereus phage Wβ and some variants of Wβ, in genome organization and the amino acid sequences of structural proteins. There were two ORFs, ORF22 and ORF35, in the genome of BtCS33 that were also found in the genomes of B. cereus phage Wβ and may be involved in regulating sporulation of the host cell. Based on these observations and analysis of phylogenetic trees, we deduced that B. thuringiensis phage BtCS33 and B. cereus phage Wβ may have a common distant ancestor