Comparative Efficiency of the Fenwick Can and Schuiling Centrifuge in Extracting Nematode Cysts from Different Soil Types

The Fenwick can and Schuiling centrifuge are widely used to extract nematode cysts from soil samples. The comparative efficiencies of these two methods during cyst extraction have not been determined for different soil types under different cyst densities. Such information is vital for statutory laboratories that must choose a method for routine, high-throughput soil monitoring. In this study, samples of different soil types seeded with varying densities of potato cyst nematode (Globodera rostochiensis) cysts were processed using both methods. In one experiment, with 200 ml samples, recovery was similar between methods. In a second experiment with 500 ml samples, cyst recovery was higher using the Schuiling centrifuge. For each method and soil type, cyst extraction efficiency was similar across all densities tested. Extraction was efficient from pure sand (Fenwick 72%, Schuiling 84%) and naturally sandy soils (Fenwick 62%, Schuiling 73%), but was significantly less efficient from clay-soil (Fenwick 42%, Schuiling 44%) and peat-soil with high organic matter content (Fenwick 35%, Schuiling 33%). Residual moisture (<10% w/w) in samples prior to analyses reduced extraction efficiency, particularly for sand and sandy soils. For each soil type and method, there were significant linear relationships between the number of cysts extracted and the numbers of cysts in the samples. We discuss the advantages and disadvantages of each extraction method for cyst extraction in statutory soil laboratories.     

Effects of bund crops and insecticide treatments on arthropod diversity and herbivore regulation in tropical rice fields

Ecological engineering using vegetable or flower strips is promoted as a potential pest management strategy in irrigated rice. Farmers in the Philippines often plant rice levees (bunds) with vegetables, particularly string beans (Vigna unguiculata [L.] Walpers) to supplement income, but without considering the potential for pest management. This study examines the effects of planted bunds on rice herbivores and their natural enemies. We compared arthropods in (a) rice fields that had string beans planted on bunds, (b) fields without string beans and without any insecticide applications and (c) fields without string beans but with insecticide treatments (standard practice). Rice yield was similar across all treatments; however, the vegetation strips produced an extra 3.6 kg of fresh string bean pods per metre of bund. There were no apparent increases in major natural enemy groups in fields with string beans compared to fields with conventional bunds. Fields with insecticide treatments had higher damage from leaffolders (Lepidoptera: Pyralidae). The sprayed fields also had lower parasitism of planthopper eggs and fewer predatory dragonflies and damselflies (Odonata). Furthermore, the mortality of planthopper (Delphacidae: Hemiptera) and stemborer (Pyralidae) eggs by parasitoids and predators was density dependent only in the unsprayed fields (with and without string beans). Our results demonstrate that planting string beans on rice bunds improves the productivity of rice farms, but our ecological engineering system did not appreciably affect natural enemy or herbivore abundance; however, chemical insecticides adversely affected pest regulatory ecosystem functions leading to higher pest damage.     

Dung beetle assemblages in forests and pastures of El Salvador: a functional comparison

This study examines the functional attributes of Neotropical dung beetles, and, based on the analysis, evaluates the role of dung beetles in pastures and tropical dry forest in El Salvador. Dung beetle diversity was lower in pastures than in forests. However, the total biomass of beetles at similarly-sized dung baits was frequently higher in pastures. Diversity loss followed structured patterns: (1) carrion and fruit feeding beetles were well represented (species number) in the forests but were largely absent from pastures; (2) large ball rollers and small fast tunnellers were present in forests but were absent from pastures; (3) large fast tunnellers and small slow tunnellers were poorly represented in pastures compared to forests but the few species that survived in pastures attained extremely high populations; and (4) each functional group in the species-poor pasture assemblages was divided almost equally into day and night active species. Substrate and habitat generalists that were present in both the forests and pastures were attracted to fruit and carrion in forested habitat but not in open pastures. In open habitats, flies and other insects may be the principal decomposers of decaying fruit and carrion.     

Structural elements of the osteopontin SVVYGLR motif important for the interaction with alpha(4) integrins

The osteopontin SVVYGLR motif binds the integrins alpha(4)beta(1) and alpha(9)beta(1). We show that alpha(4)beta(7) also interacts with this motif and that an SVVYGLR-OH peptide antagonises the alpha(4)beta(7) MAdCAM interaction. The important elements of this motif required to bind alpha(4)beta(1) and alpha(4)beta(7) were probed using a series of mutated peptides based around SVVYGLR. Leu167 is important for the interaction with alpha(4) integrins, as is the C-terminal carboxylic acid of Arg168 exposed by thrombin cleavage. The importance of the acidic group means that SVVYGLR has structural elements in common with other alpha(4) integrin-binding motifs and suggests why thrombin cleavage activates this motif.     

Degeneration of the intervertebral disc

The intervertebral disc is a cartilaginous structure that resembles articular cartilage in its biochemistry, but morphologically it is clearly different. It shows degenerative and ageing changes earlier than does any other connective tissue in the body. It is believed to be important clinically because there is an association of disc degeneration with back pain. Current treatments are predominantly conservative or, less commonly, surgical; in many cases there is no clear diagnosis and therapy is considered inadequate. New developments, such as genetic and biological approaches, may allow better diagnosis and treatments in the future.     

Analysis of the alpha4beta1 integrin-osteopontin interaction

The integrin alpha4beta1 is involved in mediating exfiltration of leukocytes from the vasculature. It interacts with a number of proteins up-regulated during the inflammatory response including VCAM-1 and the CS-1 alternatively spliced region of fibronectin. In addition it binds the multifunctional protein osteopontin (OPN), which can act as both a cytokine and an extracellular matrix molecule. Here we map the region of human OPN that supports cell adhesion via alpha4beta1 using GST fusion proteins. We show that alpha4beta1 expressed in J6 cells interacts with intact OPN when the integrin is in a high activation state, and by deletion mapping that the alpha4beta1 binding region in OPN lies between amino acid residues 125 and 168 (aa125-168). This region contains the central RGD motif of OPN, which also interacts with integrins alphavbeta3, alphavbeta5, alphavbeta1, alpha8beta1, and alpha5beta1. Mutating the RGD motif to RAD had no effect on the interaction with alpha4beta1. To define the binding site the region incorporating aa125-168 was divided into 5 overlapping peptides expressed as GST fusion proteins. Two peptides supported adhesion via alpha4beta1, aa132-146, and aa153-168; of these only a synthetic peptide, SVVYGLR (aa162-168), derived from aa153-168 was able to inhibit alpha4beta1 binding to CS-1. These data identify the motif SVVYGLR as a novel peptide inhibitor of alpha4beta1, and the primary alpha4beta1 binding site within OPN.     

Neuronal calcium activates a Rap1 and B-Raf signaling pathway via the cyclic adenosine monophosphate-dependent protein kinase

Activity-dependent regulation of neuronal events such as cell survival and synaptic plasticity is controlled by increases in neuronal calcium levels. These actions often involve stimulation of intracellular kinase signaling pathways. For example, the mitogen-activated protein kinase, or extracellular signal-regulated kinase (ERK), signaling cascade has increasingly been shown to be important for the induction of gene expression and long term potentiation. However, the mechanisms leading to ERK activation by neuronal calcium are still unclear. In the present study, we describe a protein kinase A (PKA)-dependent signaling pathway that may link neuronal calcium influx to ERKs via the small G-protein, Rap1, and the neuronal Raf isoform, B-Raf. Thus, in PC12 cells, depolarization-mediated calcium influx led to the activation of B-Raf, but not Raf-1, via PKA. Furthermore, depolarization also induced the PKA-dependent stimulation of Rap1 and led to the formation of a Rap1/B-Raf signaling complex. In contrast, depolarization did not lead to the association of Ras with B-Raf. The major action of PKA-dependent Rap1/B-Raf signaling in neuronal cells is the activation of ERKs. Thus, we further show that, in both PC12 cells and hippocampal neurons, depolarization-induced calcium influx stimulates ERK activity in a PKA-dependent manner. Given the fact that both Rap1 and B-Raf are highly expressed in the central nervous system, we suggest that this signaling pathway may regulate a number of activity-dependent neuronal functions.     

High pressures and asymmetrical stresses in the scoliotic disc in the absence of muscle loading

Background

Loads acting on scoliotic spines are thought to be asymmetric and involved in progression of the scoliotic deformity; abnormal loading patterns lead to changes in bone and disc cell activity and hence to vertebral body and disc wedging. At present however there are no direct measurements of intradiscal stresses or pressures in scoliotic spines. The aim of this study was to obtain quantitative measurements of the intradiscal stress environment in scoliotic intervertebral discs and to determine if loads acting across the scoliotic spine are asymmetric. We performed in vivo measurements of stresses across the intervertebral disc in patients with scoliosis, both parallel (termed horizontal) and perpendicular (termed vertical) to the end plate, using a side mounted pressure transducer (stress profilometry)

Methods

Stress profilometry was used to measure horizontal and vertical stresses at 5 mm intervals across 25 intervertebral discs of 7 scoliotic patients during anterior reconstructive surgery. A state of hydrostatic pressure was defined by identical horizontal and vertical stresses for at least two consecutive readings. Results were compared with similar stress profiles measured during surgery across 10 discs of 4 spines with no lateral curvature and with data from the literature.

Results

Profiles across scoliotic discs were very different from those of normal, young, healthy discs of equivalent age previously presented in the literature. Hydrostatic pressure regions were only seen in 14/25 discs, extended only over a short distance. Non-scoliotic discs of equivalent age would be expected to show large centrally placed hydrostatic nuclear regions in all discs. Mean pressures were significantly greater (0.25 MPa) than those measured in other anaesthetised patients (<0.07 MPa). A stress peak was seen in the concave annulus in 13/25 discs. Stresses in the concave annulus were greater than in the convex annulus indicating asymmetric loading in these anaesthetised, recumbent patients.

Conclusion

Intradiscal pressures and stresses in scoliotic discs are abnormal, asymmetrical and high in magnitude even in the absence of significant applied muscle loading. The origin of these abnormal stresses is unclear.     

Population genetic structure of Bombus terrestris in Europe: Isolation and genetic differentiation of Irish and British populations

The genetic structure of the earth bumblebee (Bombus terrestris L.) was examined across 22 wild populations and two commercially reared populations using eight microsatellite loci and two mitochondrial genes. Our study included wild bumblebee samples from six populations in Ireland, one from the Isle of Man, four from Britain and 11 from mainland Europe. A further sample was acquired from New Zealand. Observed levels of genetic variability and heterozygosity were low in Ireland and the Isle of Man, but relatively high in continental Europe and among commercial populations. Estimates of F_st revealed significant genetic differentiation among populations. Bayesian cluster analysis indicated that Irish populations were highly differentiated from British and continental populations, the latter two showing higher levels of admixture. The data suggest that the Irish Sea and prevailing south westerly winds act as a considerable geographical barrier to gene flow between populations in Ireland and Britain; however, some immigration from the Isle of Man to Ireland was detected. The results are discussed in the context of the recent commercialization of bumblebees for the European horticultural industry.     

A fluorometric assay for the potassium-dependent phosphatase activity of the (Na+ + K+)-adenosine triphosphatase

A fluorometric assay for the K+-dependent phosphatase activity of the (Na+ + K+)-ATPase in both purified and membrane-bound forms is described. The assay utilizes 3-O-methylfluorescein phosphate as substrate and measures the fluorescence of the 3-O-methylfluorescein produced by hydrolysis of the substrate. The assay described is an order of magnitude more sensitive than the assay employing p-nitrophenylphosphate, the substrate most commonly used to measure this activity. The assay is also suitable for the specific measurement of (Na+ + K+)-ATPase activities in membranes which contain high levels of other ATPase activities.