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A recent study carried out by Houdijk et al (2005), used a rodent model to assess whether a reduction in protein scarcity during lactation resulted in a reduced degree of parasitism. Feeding high protein foods resulted in a reduced worm burden, but was confounded with increased food intake per se. Therefore, effects observed on parasitism may not necessarily have been associated with an increased protein supply, but with changes in the gut environment due to the increased food intake. Before this model can be used to assess the underlying immune responses, further work is needed to verify that the effects observed are indeed related to changes in nutrient supply. This experiment aimed to provide further evidence on the nutritional control of parasitism during lactation by manipulating nutrient demand. It was expected that the latter would not be associated with changes in food intake per se and results could therefore be used to exclude the influence of non-immunological changes in the gut environment as a contributing factor of reduced parasitism.
Non-starch polysaccharide (NSP) content of barley is predominately in the form of β-glucan, whereas in wheat it is predominately pentosans (containing the sugars arabinose and xylose). It has generally been accepted therefore that a high level of β-glucanase is needed in broiler diets containing barley, whereas wheat based diets require a high level of endoxylanase. Trials have however previously suggested that including an endoxylanase based enzyme in broiler diets will perform at least as well as enzyme containing highlevels of β-glucanase in diets containing up to 30% barley. To confirm this finding, a trial was initiated at Roslin Institute, Edinburgh to compare the effectiveness of Natugrain Blend 66% (an endoxylanase based product) against Natugrain 33% (endoxylanase/β-glucanase product) in diets containing up to 30% barley.
The details of the formation of the first objects, stars and galaxies and their subsequent evolution remain a cosmological unknown. Few observational probes of these processes exist. The Cosmic Infrared Background (CIB) originates from this era and measurements of its anisotropy can provide information to test models of both galaxy evolution and the growth of primordial structure. Such measurements should provide a sensitive probe of the large-scale variation in protogalaxy density at redshifts, z ~ 0.5-3, while optical galaxy surveys provide complementary information at z < 0.5 and Lyman alpha absorption forest studies and Cosmic Microwave Background measurements add information at higher redshifts.
Diverse strain types of methicillin-resistant Staphylococcus aureus (MRSA) cause infections in community settings worldwide. To examine heterogeneity of spread within households and to identify common risk factors for household transmission across settings, primary data from studies conducted in New York (USA), Breda (The Netherlands), and Melbourne (Australia) were pooled. Following MRSA infection of the index patient, household members completed questionnaires and provided nasal swabs. Swabs positive for S. aureus were genotyped by spa sequencing. Poisson regression with robust error variance was used to estimate prevalence odds ratios for transmission of the clinical isolate to non-index household members. Great diversity of strain types existed across studies. Despite differences between studies, the index patient being colonized with the clinical isolate at the home visit (P < 0·01) and the percent of household members aged <18 years (P < 0·01) were independently associated with transmission. Targeted decolonization strategies could be used across geographical settings to limit household MRSA transmission.
We describe a technique for measuring localization of holes in mid-gap states in high quality a-Si:H devices. The localization of holes is determined by measuring quantum efficiency of a-Si:H devices as a function of reverse bias voltage and wavelength of light. It is shown that the QE of localized holes increases significantly upon application of high electric fields, whereas the QE of de-localized holes does not show such a behavior. The voltage-induced increase in QE is explained using a Frenkel-Poole tunneling model. It is also shown that the density of mid-gap states (states in which holes are localized) increases significantly upon light soaking, and that a major consequence of this increase in mid-gap density is a decrease in the electric field in the device. The decrease in electric field is experimentally estimated by fitting the increased current due to tunneling to the expression for Frenkel-Poole tunneling.
The interdiffusion mechanism in Hg(l-x)X(x)Te/CdTe superlattices where X is Cd, Mn, or Zn can be deduced from the magnitude of the interdiffusion activation energy. By comparing in-situ x-ray diffraction measurements (our work) with results from Tang and Stevenson (J. Vac. Sci. Technol. AS, 1987), it is found that anionic and cationic Frenkel pairs represent the most likely interdiffusion mechanism in Hg(l-x)X(x)Te/CdTe superlattices. This model mixes vacancies and interstitials, as well as maintaining the conduction type and the electronic mobility. It is further shown that interdiffusion sets in as soon as the growth starts.
The electronic and optical properties of device quality hydrogenated amorphous silicon (a-Si:H) films grown by electron cyclotron resonance (ECR) plasma deposition were studied together with in-situ plasma characteristics. Hydrogen and helium plasmas, excited by 50–250 watts of 2.45 GHz microwave power under ECR conditions, were used to decompose silane at 6 to 20 mtorr pressures during the deposition of a-Si:H films at a 297 C substrate temperature. Both the electron temperature and density, and ion flux are measured near the deposition surface using plane and cylindrical Langmuir probes. An attempt is made to correlate these plasma properties with the light and dark photoconductivity, optical gap, refractive index, and subband gap photoconductivity.
The transforming growth factor-β (TGF-β) gene family regulates critical processes in animal development, and plays a crucial role in regulating the mammalian immune response. We aimed to identify TGF-β homologues from 2 laboratory model nematodes (Heligmosomoides polygyrus and Nippostrongylus brasiliensis) and 2 major parasites of ruminant livestock (Haemonchus contortus and Teladorsagia circumcincta). Parasite cDNA was used as a template for gene-specific PCR and RACE. Homologues of the TGH-2 subfamily were isolated, and found to differ in length (301, 152, 349 and 305 amino acids respectively), with variably truncated N-terminal pre-proteins. All contained conserved C-terminal active domains (>85% identical over 115 amino acids) containing 9 cysteine residues, as in C. elegans DAF-7, Brugia malayi TGH-2 and mammalian TGF-β. Surprisingly, only the H. contortus homologue retained a conventional signal sequence, absent from shorter proteins of other species. RT-PCR assays of transcription showed that in H. contortus and N. brasiliensis expression was maximal in the infective larval stage, and very low in adult worms. In contrast, in H. polygyrus and T. circumcincta, tgh-2 transcription is higher in adults than infective larvae. The molecular evolution of this gene family in parasitic nematodes has diversified the pre-protein and life-cycle expression patterns of TGF-β homologues while conserving the structure of the active domain.
The sheep scab mite, Psoroptes ovis, induces an intensely pruritic exudative dermatitis which is responsible for restlessness, loss of appetite and weight loss. Within the first 24 h of infection, there is a rapid inflammatory influx of eosinophils and apoptosis of the keratinocytes at the site of infection. The former cell type is capable of a sustained respiratory burst, toxic products of which may directly damage the mite and also contribute to lesion formation. Analysis of a P. ovis expressed sequence tag (EST) database identified a number of antioxidant enzyme-encoding sequences, including peroxiredoxin (thioredoxin peroxidase EC 184.108.40.206), all of which may help the mite endure the potentially toxic skin environment. A full length sequence encoding Po-TPx, a protein of 206 amino acids which showed high homology to a peroxiredoxin from the salivary gland of the tick Ixodes scapularis, was amplified from P. ovis cDNA. Recombinant Po-TPx was expressed in bacteria and antiserum to this protein was used to localize native Po-TPx in mite sections. Peroxiredoxin was localized, amongst other sites, to a subpharyngeal region in mite sections. The recombinant protein was recognized by sera from sheep infested with the mite suggesting that it may be secreted or excreted by the mite and interact with the host immune response.
Negative consumer opinion poses a potential barrier to the application of nutrigenomic intervention. The present study has aimed to determine attitudes toward genetic testing and personalised nutrition among the European public. An omnibus opinion survey of a representative sample aged 14–55+ years (n 5967) took place in France, Italy, Great Britain, Portugal, Poland and Germany during June 2005 as part of the Lipgene project. A majority of respondents (66 %) reported that they would be willing to undergo genetic testing and 27 % to follow a personalised diet. Individuals who indicated a willingness to have a genetic test for the personalising of their diets were more likely to report a history of high blood cholesterol levels, central obesity and/or high levels of stress than those who would have a test only for general interest. Those who indicated that they would not have a genetic test were more likely to be male and less likely to report having central obesity. Individuals with a history of high blood cholesterol were less likely than those who did not to worry if intervention foods contained GM ingredients. Individuals who were aware that they had health problems associated with the metabolic syndrome appeared particularly favourable toward nutrigenomic intervention. These findings are encouraging for the future application of personalised nutrition provided that policies are put in place to address public concern about how genetic information is used and held.
Molecular markers have been used to study genetic diversity within a set of Lablabpurpureus accessions collected from the southern states of India. Amplified fragment length polymorphism (AFLP) molecular marker studies using a total of 78 L. purpureus accessions with nine primer combinations showed there was very little genetic diversity within the L. purpureus accessions from the southern Indian germplasm collection as compared to a set of 15 accessions from other international germplasm collections that included African accessions. The set of 15 were selected from a random amplified length polymorphism (RAPD) marker study and chosen on the basis of widest genetic distance. Further molecular analysis with polymerase chain reaction (PCR) markers from 97 expressed sequence tag (EST) and gene-specific primer pairs, designed from a range of legume sequences, concurred with the AFLP analyses. Both of these approaches provide a wealth of markers for diversity and mapping studies. The 97 sequence-specific primer pairs tested in L. purpureus resulted in 70% amplification success, with 44% of primer pairs amplifying single bands and 10% double bands. Markers generated from these EST and genomic sequences provide useful cross-reference to comparative legume genomics that will potentially have long-term benefit to legume plant breeding.
The treatment and prevention of parasitism in both humans and livestock continues to rely almost exclusively on the use of antiparasitic drugs – an approach which has limitations, particularly as reinfection, which occurs rapidly in endemic regions, is not prevented. In addition, the widespread appearance of drug-resistant parasites of animals (Kaplan, 2004;) together with emerging evidence of resistance problems in human parasites (Fallon et al. 1995; Ismail et al. 1996; De Clerq et al. 1997; East African Network for Monitoring Antimalarial Treatment, 2003), emphasise the importance of developing alternative methods of control, with anti-parasite vaccines a prime target.
This paper summarises the progress towards vaccine development against the major blood-feeding nematodes of man and livestock, the hookworms and Haemonchus contortus, respectively. The impact of the diseases and the drivers for vaccine development are summarized as well as the anticipated impact of the host immune response on vaccine design. The performance requirements are discussed and progress towards these objectives using defined larval and adult antigens, many of these being shared between species. Specific examples include the Ancylostoma secreted proteins and homologues in Haemonchus as well as proteases used for digestion of the blood meal. This discussion shows that many of the major vaccine candidates are shared between these blood-feeding species, not only those from the blood-feeding stages but also those expressed by infective L3s in the early stages of infection. Challenges for the future include: exploiting the expanding genome information for antigen discovery, use of different recombinant protein expression systems, formulation with new adjuvants, and novel methods of field testing vaccine efficacy.
RNA interference (RNAi) on parasitic nematodes has been described as successful and useful for the identification of novel drug and vaccine candidates. In this study we have evaluated this technology on the cattle parasite Ostertagia ostertagi. Eight different genes were targeted in L1 and L3 O. ostertagi larvae, by electroporation and soaking in dsRNA respectively. Down-regulation of target transcript levels was evaluated by semi-quantitative reverse transcriptase (RT) PCR. In L3 larvae, variable decreases in mRNA levels were observed for 5 genes, ranging from a complete knock down (tropomyosin, β-tubulin) to a minor decrease (ATPsynthase, superoxide dismutase, polyprotein allergen). However, repeated experiments indicated that effects were sometimes difficult to reproduce. RNAi for ubiquitin, a transthyretin-like protein and a 17 kDa excretion secretion (ES) protein never resulted in a knock down of the transcript. The mRNA levels of 7 non-target genes showed no difference between larvae soaked in C. elegans control dsRNA versus O. ostertagi tropomyosin dsRNA, supporting that the observed reductions are specific for the target gene. Electroporation of L1 larvae proved to be less effective. Reductions in mRNA levels were only noticed for 2 genes and were not reproducible. In conclusion, the results indicate that the RNAi pathway is probably present in O. ostertagi but that the current RNAi techniques can not be used as a reliable screening method.
Asparaginyl proteinases (or legumains) are a recently identified, novel class of cysteine proteinase which specifically hydrolyse peptide bonds after asparagine residues. Legumains have been implicated in the activation of cysteine proteases, particularly cathepsin B-like proteinases which are thought to help degrade the bloodmeal in blood-feeding helminths such as schistosomes, hookworms and other nematode species. An EST sequence representing a full-length legumain was identified from the Haemonchus contortus datasetNucleotide sequence data described in this paper have been deposited in the EMBL database under the Accession number AM177177.. This encoded a protein with a predicted Mr of 49 kDa, the amino acid sequence of which showed good homology (34–40% identity) to legumains from Schistosoma mansoni, human and rat and contained a legumain-like active site. RT-PCR indicated that the legumain transcript was expressed from the L4 life-cycle stage onwards. The coding sequence was expressed in E. coli and antibodies to the resultant recombinant protein indicated that the enzyme was expressed in the microvillar surface of the intestinal cells. Legumain activity was detected in extracts of the adult parasite but not the host protective Thiol-Sepharose-binding fraction, although it was detectable in the latter by immunoblot. Activity was relatively insensitive to E64, an inhibitor of cysteine proteinases and completely inhibited by the alkylating agent, N-ethylmaleimide, consistent with inhibitor effects on previously characterized legumains.
RNA interference (RNAi) has become an invaluable tool for the functional analysis of genes in a wide variety of organisms including the free-living nematode Caenorhabditis elegans. Recently, attempts have been made to apply this technology to parasitic helminths of animals and plants with variable success. Gene knockdown has been reported for Schistosoma mansoni by soaking or electroporating different life-stages in dsRNA. Similar approaches have been tested on parasitic nematodes which clearly showed that, under certain conditions, it was possible to interfere with gene expression. However, despite these successes, the current utility of this technology in parasite research is questionable. First, problems have arisen with the specificity of RNAi. Treatment of the parasites with dsRNA resulted, in many cases, in non-specific effects. Second, the current RNAi methods have a limited efficiency and effects are sometimes difficult to reproduce. This was especially the case in strongylid parasites where only a small number of genes were susceptible to RNAi-mediated gene knockdown. The future application of RNAi in parasite functional genomics will greatly depend on how we can overcome these difficulties. Optimization of the dsRNA delivery methods and in vitro culture conditions will be the major challenges.