Interfacial structure of wild-type and mutant forms of puroindoline-b bound to DPPG monolayers

The interaction of wild-type puroindoline-b (Pin-b+) and two mutant forms having single residue substitutions (G46S or W44R) with L-alpha-dipalmitoylphosphatidyl-dl-glycerol (DPPG) as a Langmuir monolayer at the air/water interface was investigated by neutron reflectivity (NR) and Brewster angle microscopy (BAM). NR profiles were fitted using a three-layer model to enable differences in penetration of protein between the lipid headgroup and acyl regions to be determined. The data showed similar surface excesses for each of the three proteins at the interface; however, it was revealed that the depth of penetration of protein into the lipid region differed for each protein with Pin-b+ penetrating further into the acyl region of the lipid compared to the mutant forms of the protein that interacted with the headgroup region only. BAM images revealed that the domain structure of the DPPG monolayers was disrupted when Pin-b+ adsorption had reached equilibrium, suggesting protein penetration had led to compression of the lipid region. In contrast, the domain structure was unaffected by the W44R mutant, suggesting no change in compression of the lipid region and hence little or no penetration of protein into the lipid layer.

In-street wind direction variability in the vicinity of a busy intersection in central London

We present results from fast-response wind measurements within and above a busy intersection between two street canyons (Marylebone Road and Gloucester Place) in Westminster, London taken as part of the DAPPLE (Dispersion of Air Pollution and Penetration into the Local Environment; www.dapple.org.uk) 2007 field campaign. The data reported here were collected using ultrasonic anemometers on the roof-top of a building adjacent to the intersection and at two heights on a pair of lamp-posts on opposite sides of the intersection. Site characteristics, data analysis and the variation of intersection flow with the above-roof wind direction (θref) are discussed. Evidence of both flow channelling and recirculation was identified within the canyon, only a few metres from the intersection for along-street and across-street roof-top winds respectively. Results also indicate that for oblique rooftop flows, the intersection flow is a complex combination of bifurcated channelled flows, recirculation and corner vortices. Asymmetries in local building geometry around the intersection and small changes in the background wind direction (changes in 15-min mean θref of 5–10 degrees) were also observed to have profound influences on the behaviour of intersection flow patterns. Consequently, short time-scale variability in the background flow direction can lead to highly scattered in-street mean flow angles masking the true multi-modal features of the flow and thus further complicating modelling challenges.

Puroindoline-b mutations control the lipid binding interactions in mixed puroindoline-a: puroindoline-b systems

The interactions have been investigated of puroindoline-a (Pin-a) and mixed protein systems of Pin-a and wild-type puroindoline-b (Pin-b+) or puroindoline-b mutants (G46S mutation (Pin bH) or W44R mutation (Pin-bS)) with condensed phase monolayers of an anionic phospholipid (L-α-dipalmitoylphosphatidyl-dl-glycerol (DPPG)) at the air/water interface. The interactions of the mixed systems were studied at three different concentration ratios of Pin-a:Pin-b, namely 3:1, 1:1 and 1:3 in order to establish any synergism in relation to lipid binding properties. Surface pressure measurements revealed that Pin-a interaction with DPPG monolayers led to an equilibrium surface pressure increase of 8.7 ± 0.6 mN m-1. This was less than was measured for Pin-a:Pin-b+ (9.6 to 13.4 mN m-1), but was significantly more than was measured for Pin-a:Pin-bH (4.0 to 6.2 mN m-1) or Pin-a:Pin-bS (3.8 to 6.3 mN m-1) over the complete range of concentration ratio. Consequently, surface pressure increases were shown to correlate to endosperm hardness phenotype, with puroindolines present in hard-textured wheat varieties yielding lower equilibrium surface pressure changes. Integrated amide I peak areas from corresponding external reflectance Fourier-transform infrared (ER-FTIR) spectra, used to indicate levels of protein adsorption to the lipid monolayers, showed that differences in adsorbed amount were less significant. The data therefore suggest that Pin-b mutants having single residue substitutions within their tryptophan-rich loop that are expressed in some hard-textured wheat varieties influence the degree of penetration of Pin-a and Pin-b into anionic phospholipid films. These findings highlight the key role of the tryptophan-rich loop in puroindoline-lipid interactions.

Single amino acid substitutions in puroindoline-b mutants influence lipid binding properties

External reflectance Fourier transform infrared (ER-FTIR) spectroscopy and surface pressure measurements have been used to characterize the interaction of wild-type puroindoline-b (Pin-b) and two mutant forms featuring single residue substitutions-namely, Gly-46 to Ser-46 (Pin-bH) and Trp-44 to Arg-44 (Pin-bS)-with condensed-phase monolayers of zwitterionic (L-alpha-dipalmitoylphosphatidylcholine, DPPC) and anionic (L-alpha-dipalmitoylphosphatidyl-dl-glycerol, DPPG) phospholipids. The interaction with anionic DPPG monolayers, monitored by surface pressure isotherms, was influenced significantly by mutations in Pin-b (p < 0.05); wild-type Pin-b showed the highest surface pressure change of 10.6 +/- 1.0 mN m(-1), followed by Pin-bH (7.9 +/- 1.6 mN m(-1)) and Pin-bS (6.3 +/- 1.0 mN m(-1)), and the surface pressure isotherm kinetics were also different in each case. Integrated Amide I peak areas from corresponding ER-FTIR spectra confirmed the differences in adsorption kinetics, but also showed that differences in adsorbed amount were less significant, suggesting that mutations influence the degree of penetration into DPPG films. All Pin-b types showed evidence of interaction with DPPC films, detected as changes in surface pressure (5.6 +/- 1.1 mN m(-1)); however, no protein peaks were detected in the ER-FTIR spectra, which indicated that the interaction was via penetration with limited adsorption at the lipid/water interface. The expression of Pin-b mutants is linked to wheat endosperm hardness; therefore, the data presented here suggest that the lipid binding properties may be pivotal within the mechanism for this quality trait. In addition, the data suggest antimicrobial activities of Pin-b mutants would be lower than those of the wild-type Pin-b, because of decreased selectivity toward anionic phospholipids.

Optimal protection of stabilised dry live bacteria from bile toxicity in oral dosage forms by bile acid adsorbent resins

We previously found that dried live bacteria of a vaccine strain can be temporarily sensitive to bile acids and suggested that Bile Adsorbing Resins (BAR) can be used in oral vaccine tablets to protect dried bacteria from intestinal bile. Here, we report a quantitative analysis of the ability of BAR to exclude the dye bromophenol blue from penetrating into matrix tablets and also sections of hard capsule shells. Based on this quantitative analysis, we made a fully optimised formulation, comprising 25% w/w of cholestyramine in Vcaps™ HPMC capsules. This gave effectively 100% protection of viability from 4% bile, with 4200-fold more live bacteria recovered from this formulation compared to unprotected dry bacteria. From the image analysis, we found that the filler material or compaction force used had no measurable effect on dye exclusion but did affect the rate of tablet hydration. Increasing the mass fraction of BAR gave more exclusion of dye up to 25% w/w, after which a plateau was reached and no further dye exclusion was seen. More effective dye exclusion was seen with smaller particle sizes (i.e. cholestyramine) and when the BAR was thoroughly dried and disaggregated. Similar results were found when imaging dye penetration into capsule sections or tablets. The predictions of the dye penetration study were tested using capsules filled with dried attenuated Salmonella vaccine plus different BAR types, and the expected protection from bile was found, validating the imaging study. Surprisingly, depending on the capsule shell material, some protection was given by the capsule alone without adding BAR, with Vcaps™ HPMC capsules providing up to 174-fold protection against 1% bile; faster releasing Vcaps Plus™ HPMC capsules and Coni Snap™ gelatin capsules gave less protection.

Use of entomopathogenic bacterium Pseudomonas putida (Enterobacteriaceae) and it secretions against the Greater Wax Moth Galleria mellonella pupae

The bacterium from Pseudomonas putida from Steinernema abbasi and its metabolic secretions caused the mortality of the Galleria mellonella pupae. Experiments were conducted in sand and filter paper on time exposure, temperature, moisture, dose and time of penetration of bacterium in pupae and tested stored or dried toxic metabolites using G. mellonella pupae as a test target organism. Death of pupae was probably due to the toxic metabolites. Pseudomonas putida cells were recovered from the haemocoele when bacterial cells were applied to the G. mellonella pupae indicating that bacterial cells can enter the haemocoele in the absence of nematode vector. Penetration of bacterium was found rapidly after application on G. mellonella pupae. Pseudomonas putida or its toxic secretions can be used as a microbial control for insect control. The experimental results indicate that there is possibility of using P. putida and its toxic secretions as a biopesticide and can contribute in the development of new microbial and biological control against insect pests.

Pathogenicity of the bacterium Pseudomonas putida from the entomopathogenic nematode Steinernema abbasi and its secretion against Galleria mellonella larvae

The Entomopathogenic bacterium Pseudomonas putida from Steinernema abbasi and its metabolic secretions were lethal to the Galleria mellonella larvae. Different laboratory experiments on time interval, substrate, moisture, temperature, dose, penetration of cells, stored and dried metabolites were conducted in sand and filter paper bioassays. It was concluded that death was probably due to the toxic metabolites. This bacterium and its metabolites were found very effective at 30 degree C. Penetration of bacterium was rapid after application on G. mellonella larvae. P. putida cells were recovered from the haemocoele when suspensions containing bacterial cells were applied to the G. mellonella indicating that bacterial symbionts do have a free-living existence and can enter the haemocoele in the absence of nematode vector. Stored metabolite and dried metabolites were found persistent for long time. This bacterium or its toxic secretions can be used for insect control that can be important component of integrated pest management against different insect pests. P. putida and its secretions are suggested as the most appropriate suspension to apply against insect pest control program in tropical ecological regions.

The rapid rise of supermarkets?

A series of articles, many of them published in this journal, have charted the rapid spread of supermarkets in developing and middle-income countries and forecast its continuation. In this article, the level of supermarket penetration (share of the retail food market) is modelled quantitatively on a cross-section of 42 countries for which data could be obtained, representing all stages of development. GDP per capita, income distribution, urbanisation, female labour force participation and openness to inward foreign investment are all significant explanators. Projections to 2015 suggest significant but not explosive further penetration; increased openness and GDP growth are the most significant factors.

Plasmodiophora brassicae in its Environment

Plasmodiophora brassicae Wor. is viewed in this article from the standpoint of a highly evolved and successful organism, well fitted for the ecological niche that it occupies. Physical, chemical, and biological components of the soil environment are discussed in relation to their effects on the survival, growth, and reproduction of this microbe. It is evident that P. brassicae is well equipped by virtue of its robust resting spores for survival through many seasonal cycles. Germination is probably triggered as a result of signals initiated by root exudates. The resultant motile zoospore moves rapidly to the root hair surface and penetration and colonization follow. The short period between germination and penetration is one of greatest vulnerability for P. brassicae. In this phase survival is affected at the very least by soil texture and structure; its moisture; pH; calcium, boron, and nitrogen content; and the presence of active microbial antagonists. These factors influence the inoculum potential (sensu Garrett, 1956) and its viability and invasive capacity. There is evidence that these effects may also influence differentially the survival of some physiologic races of P. brassicae. Considering the interaction of P. brassicae with the soil environment from the perspective of its biological fitness is an unusual approach; most authors consider only the opportunities to destroy this organism. The approach adopted here is borne of several decades spent studying P. brassicae and the respect that has been engendered for it as a biological entity. This review stops at the point of penetration, although some of the implications of the environment for successful colonization are included because they form a continuum. Interactions with the molecular and biochemical cellular environment are considered in other sections in this special edition.

Computer modelling and estimation of recruitment patterns of non-branching coral colonies at three sites in Wakatobi Marine Park, S.E. Sulawesi, Indonesia; implications for coral reef conservation

We have studied growth and estimated recruitment of massive coral colonies at three sites, Kaledupa, Hoga and Sampela, separated by about 1.5 km in the Wakatobi Marine National Park, S.E. Sulawesi, Indonesia. There was significantly higher species richness (P<0.05), coral cover (P<0.05) and rugosity (P<0.01) at Kaledupa than at Sampela. A model for coral reef growth has been developed based on a rational polynomial function, where dx/dt is an index of coral growth with time; W is the variable (for example, coral weight, coral length or coral area), up to the power of n in the numerator and m in the denominator; a1……an and b1…bm are constants. The values for n and m represent the degree of the polynomial, and can relate to the morphology of the coral. The model was used to simulate typical coral growth curves, and tested using published data obtained by weighing coral colonies underwater in reefs on the south-west coast of Curaçao [‘Neth. J. Sea Res. 10 (1976) 285’]. The model proved an accurate fit to the data, and parameters were obtained for a number of coral species. Surface area data was obtained on over 1200 massive corals at three different sites in the Wakatobi Marine National Park, S.E. Sulawesi, Indonesia. The year of an individual's recruitment was calculated from knowledge of the growth rate modified by application of the rational polynomial model. The estimated pattern of recruitment was variable, with little numbers of massive corals settling and growing before 1950 at the heavily used site, Sampela, relative to the reef site with little or no human use, Kaledupa, and the intermediate site, Hoga. There was a significantly greater sedimentation rate at Sampela than at either Kaledupa (P<0.0001) or Hoga (P<0.0005). The relative mean abundance of fish families present at the reef crests at the three sites, determined using digital video photography, did not correlate with sedimentation rates, underwater visibility or lack of large non-branching coral colonies. Radial growth rates of three genera of non-branching corals were significantly lower at Sampela than at Kaledupa or at Hoga, and there was a high correlation (r=0.89) between radial growth rates and underwater visibility. Porites spp. was the most abundant coral over all the sites and at all depths followed by Favites (P<0.04) and Favia spp. (P<0.03). Colony ages of Porites corals were significantly lower at the 5 m reef flat on the Sampela reef than at the same depth on both other reefs (P<0.005). At Sampela, only 2.8% of corals on the 5 m reef crest are of a size to have survived from before 1950. The Scleractinian coral community of Sampela is severely impacted by depositing sediments which can lead to the suffocation of corals, whilst also decreasing light penetration resulting in decreased growth and calcification rates. The net loss of material from Sampela, if not checked, could result in the loss of this protective barrier which would be to the detriment of the sublittoral sand flats and hence the Sampela village.

Using a water-soluble melamine-formaldehyde resin to improve the hardness of Norway spruce wood

Samples of Norway spruce wood were impregnated with a water-soluble melamine formaldehyde resin by using short-term vacuum treatment and long-term immersion, respectively. By means of Fourier transform infrared (FTIR) spectroscopy and UV microspectrophotometry, it was shown that only diffusion during long-term immersion leads to sufficient penetration of melamine resin into the wood structure, the flow of liquids in Norway spruce wood during vacuum treatment being greatly hindered by aspirated pits. After an immersion in aqueous melamine resin solution for 3 days, the resin had penetrated to a depth > 4 mm, which, after polymerization of the resin, resulted in an improvement of hardness comparable to the hardwood beech. A finite element model describing the effect of increasing depth of modification on hardness demonstrated that under the test conditions chosen for this study, a minimum impregnation depth of 2 mm is necessary to achieve an optimum increase in hardness. (C) 2004 Wiley Periodicals, Inc.

Impact damage characterisation of thermoplastic matrix composites using transmission transient thermography

In this work, IR thermography is used as a non-destructive tool for impact damage characterisation on thermoplastic E-glass/polypropylene composites for automotive applications. The aim of this experimentation was to compare impact resistance and to characterise damage patterns of different laminates, in order to provide indications for their use in components. Two E-glass/polypropylene composites, commingled ®Twintex (with three different weave structures: directional, balanced and 3-D) and random reinforced GMT, were in particular characterised. Directional and balanced Twintex were also coupled in a number of hybrid configurations with GMT to evaluate the possible use of GMT/Twintex hybrids in high-energy absorption components. The laminates were impacted using a falling weight tower, with impact energies ranging from 15 J to penetration. Using IR thermography during cooling down following a long pulse (3 s), impact damaged areas were characterised and the influence of weave structure on damage patterns was studied. IR thermography offered good accuracy for laminates with thickness not exceeding 3.5 mm: this appears to be a limit for the direct use of this method on components, where more refined signal treatment would probably be needed for impact damage characterisation.

Predicting the thermal inactivation of bacteria in a solid matrix: simulation studies on the relative effects of microbial thermal resistance parameters and process conditions

A combined mathematical model for predicting heat penetration and microbial inactivation in a solid body heated by conduction was tested experimentally by inoculating agar cylinders with Salmonella typhimurium or Enterococcus faecium and heating in a water bath. Regions of growth where bacteria had survived after heating were measured by image analysis and compared with model predictions. Visualisation of the regions of growth was improved by incorporating chromogenic metabolic indicators into the agar. Preliminary tests established that the model performed satisfactorily with both test organisms and with cylinders of different diameter. The model was then used in simulation studies in which the parameters D, z, inoculum size, cylinder diameter and heating temperature were systematically varied. These simulations showed that the biological variables D, z and inoculum size had a relatively small effect on the time needed to eliminate bacteria at the cylinder axis in comparison with the physical variables heating temperature and cylinder diameter, which had a much greater relative effect. (c) 2005 Elsevier B.V All rights reserved.

Liposomes and skin: From drug delivery to model membranes

The early eighties saw the introduction of liposomes as skin drug delivery systems, initially promoted primarily for localised effects with minimal systemic delivery. Subsequently, a novel ultradeformable vesicular system (termed "Transfersomes" by the inventors) was reported for transdermal delivery with an efficiency similar to subcutaneous injection. Further research illustrated that the mechanisms of liposome action depended on the application regime and the vesicle composition and morphology. Ethical, health and supply problems with human skin have encouraged researchers to use skin models. 'IYaditional models involved polymer membranes and animal tissue, but whilst of value for release studies, such models are not always good mimics for the complex human skin barrier, particularly with respect to the stratum corneal intercellular lipid domains. These lipids have a multiply bilayered organization, a composition and organization somewhat similar to liposomes, Consequently researchers have used vesicles as skin model membranes. Early work first employed phospholipid liposomes and tested their interactions with skin penetration enhancers, typically using thermal analysis and spectroscopic analyses. Another approach probed how incorporation of compounds into liposomes led to the loss of entrapped markers, analogous to "fluidization" of stratum corneum lipids on treatment with a penetration enhancer. Subsequently scientists employed liposomes formulated with skin lipids in these types of studies. Following a brief description of the nature of the skin barrier to transdermal drug delivery and the use of liposomes in drug delivery through skin, this article critically reviews the relevance of using different types of vesicles as a model for human skin in permeation enhancement studies, concentrating primarily on liposomes after briefly surveying older models. The validity of different types of liposome is considered and traditional skin models are compared to vesicular model membranes for their precision and accuracy as skin membrane mimics. (c) 2008 Elsevier B.V. All rights reserved.