Assessment of risk of insect-resistant transgenic crops to nontarget arthropods

An international initiative is developing a scientifically rigorous approach to evaluate the potential risks to nontarget arthropods (NTAs) posed by insect-resistant, genetically modified (IRGM) crops. It adapts the tiered approach to risk assessment that is used internationally within regulatory toxicology and environmental sciences. The approach focuses on the formulation and testing of clearly stated risk hypotheses, making maximum use of available data and using formal decision guidelines to progress between testing stages (or tiers). It is intended to provide guidance to regulatory agencies that are currently developing their own NTA risk assessment guidelines for IRGM crops and to help harmonize regulatory requirements between different countries and different regions of the world.     

Properties of the periplasmic nitrate reductases from Paracoccus pantotrophus and Escherichia coli after growth in tungsten-supplemented media

Paracoccus pantotrophus grown anaerobically under denitrifying conditions expressed similar levels of the periplasmic nitrate reductase (NAP) when cultured in molybdate- or tungstate-containing media. A native PAGE gel stained for nitrate reductase activity revealed that only NapA from molybdate-grown cells displayed readily detectable nitrate reductase activity. Further kinetic analysis showed that the periplasmic fraction from cells grown on molybdate (3 microM) reduced nitrate at a rate of V(max)=3.41+/-0.16 micromol [NO(3)(-)] min(-1) mg(-1) with an affinity for nitrate of K(m)=0.24+/-0.05 mM and was heat-stable up to 50 degrees C. In contrast, the periplasmic fraction obtained from cells cultured in media supplemented with tungstate (100 microM) reduced nitrate at a much slower rate, with much lower affinity (V(max)=0.05+/-0.002 micromol [NO(3)(-)] min(-1) mg(-1) and K(m)=3.91+/-0.45 mM) and was labile during prolonged incubation at >20 degrees C. Nitrate-dependent growth of Escherichia coli strains expressing only nitrate reductase A was inhibited by sub-mM concentrations of tungstate in the medium. In contrast, a strain expressing only NAP was only partially inhibited by 10 mM tungstate. However, none of the above experimental approaches revealed evidence that tungsten could replace molybdenum at the active site of E. coli NapA. The combined data show that tungsten can function at the active site of some, but not all, molybdoenzymes from mesophilic bacteria.     

Crystal structure of human MMP9 in complex with a reverse hydroxamate inhibitor

Matrix metalloproteinases (MMPs) and their inhibitors are important in connective tissue re-modelling in diseases of the cardiovascular system, such as atherosclerosis. Various members of the MMP family have been shown to be expressed in atherosclerotic lesions, but MMP9 is consistently seen in inflammatory atherosclerotic lesions. MMP9 over-expression is implicated in the vascular re-modelling events preceding plaque rupture (the most common cause of acute myocardial infarction). Reduced MMP9 activity, either by genetic manipulation or through pharmacological intervention, has an impact on ventricular re-modelling following infarction. MMP9 activity may therefore represent a key mechanism in the pathogenesis of heart failure. We have determined the crystal structure, at 2.3 A resolution, of the catalytic domain of human MMP9 bound to a peptidic reverse hydroxamate inhibitor as well as the complex of the same inhibitor bound to an active-site mutant (E402Q) at 2.1 A resolution. MMP9 adopts the typical MMP fold. The catalytic centre is composed of the active-site zinc ion, co-ordinated by three histidine residues (401, 405 and 411) and the essential glutamic acid residue (402). The main differences between the catalytic domains of various MMPs occur in the S1' subsite or selectivity pocket. The S1' specificity site in MMP9 is perhaps best described as a tunnel leading toward solvent, as in MMP2 and MMP13, as opposed to the smaller pocket found in fibroblast collagenase and matrilysin. The present structure enables us to aid the design of potent and specific inhibitors for this important cardiovascular disease target.     

Targeting of an enteropathogenic Escherichia coli (EPEC) effector protein to host mitochondria

Many Gram-negative pathogens use a type III secretion apparatus to deliver effector molecules into host cells to subvert cellular processes in favour of the pathogen. Enteropathogenic Escherichia coli (EPEC) uses such a system to deliver the Tir effector molecule into host cells. In this paper, we show that the gene upstream of tir , orf 19, encodes an additional type III secreted effector protein. Orf19 is delivered into host cells by a mechanism independent of endocytosis, but dependent on EspB. Orf19 is targeted to host mitochondria, where it appears to interfere with the ability to maintain membrane potential. Although the precise role of Orf19 remains to be elucidated, its interaction with mitochondria suggests a possible role in the subversion of key functions of these organelles, such as energy production or control of cell death. This is the first example of a type III secreted protein targeted to mitochondria; it is probable that homologues (present in EPEC and Shigella species) and other bacterial effectors will also target this organelle.     

Hoverfly oviposition response to aphids in broccoli fields

The oviposition response of predacious hoverflies (Diptera: Syrphidae) to Brevicoryne brassicae L. and Myzus persicae (Sulzer) (Homoptera: Aphididae) in commercial broccoli, Brassica oleracea var. italica L., Plenck (Brassicaceae), fields was investigated at two sites over the course of a growing season. The hoverfly oviposition responses to these aphid species on different parts of the broccoli plant canopy were also examined. There were no hoverfly eggs on broccoli plants without aphids, egg numbers were very low on plants with fewer than 50 aphids, and no peak in oviposition relative to aphid numbers was observed. Within individual plants that were colonized by aphids, there was some oviposition on individual leaves without aphids, and no hoverfly eggs were seen on leaves that had more than 400 aphids. Leaves in the broccoli plant canopy, and the datasets associated with them, were divided into three sections vertically, ‘upper’, ‘middle’, and ‘lower’. Brevicoryne brassicae was more abundant in the upper and middle canopy sections, while M. persicae was found mostly in the lower section. The rate of hoverfly oviposition per aphid was higher in the upper section than in the two other sections. Modeling of the oviposition response using logistic regression showed that the presence of hoverfly eggs was positively correlated with numbers of each aphid species and sampling date.     

A mass stranding of white-sided dolphins (Lagenorhynchus acutus) in Ireland: biological and pathological studies

We give details of the biological and pathological findings in a mass stranding of 19 white-sided dolphins on the west coast of Ireland. The pod consisted of adults and calves; no juveniles were present. The calf of a recently parturient female was not found. The presence of small foetuses suggested that mating took place in late August or early September. The largest and oldest male was severely diseased but the remainder of the pod were in good health although some minor lesions were found. The site where the animals came ashore was compatible with the geomagnetic theory of mass strandings.     

Temporal and spatial changes in aphid and parasitoid populations following applications of deltamethrin in winter wheat

The spatial and temporal effects of the pyrethroid insecticide, deltamethrin, on populations of cereal aphids and their primary parasitoids and hyperparasitoids were investigated in a large scale field experiment in winter wheat. Four hectare plots were treated at the recommended field concentration or a reduced concentration representing one twentieth of field rate. A control plot was left unsprayed. Invertebrate populations were sampled at sites within a grid layout covering the whole plots, enabling the use of geostatistical analysis. Hymenopteran populations were monitored using transparent sticky traps and suction sampling. Aphid populations were recorded by visual counts. Monitoring continued for 36 days after treatment. The full rate deltamethrin treatment resulted in initial reductions of aphid populations by 78%. Primary parasitoid and hyperparasitoid populations were reduced in suction samples by 90% and 47% respectively, when corrected for control fluctuations. The reduced deltamethrin concentration caused reductions of aphid, primary parasitoid and hyperparasitoid populations of 40, 60 and 54% respectively. Aphid population recovery over the full rate plot occurred slowly and in a patchy manner following treatment. No significant reinvasion gradients were detected. Aphid population density recovered more rapidly in the reduced rate treatment; with initial evidence for increased densities at the plot centre. Significant patterns of reinvasion were initially detected for both groups of Hymenoptera in the full rate treatment, suggesting that reinvasion of the sprayed area was taking place from untreated surrounding reservoirs. It was concluded that experiments that examine pesticide impacts within small plots will lead to underestimates of effects on dispersive groups including parasitic Hymenoptera.