Hydrolysis of the amyloid prion protein and nonpathogenic meat and bone meal by anaerobic thermophilic prokaryotes and streptomyces subspecies

Transmissible spongiform encephalopathies are caused by accumulation of highly resistant misfolded amyloid prion protein PrPres and can be initiated by penetration of such pathogen molecules from infected tissue to intact organism. Decontamination of animal meal containing amyloid prion protein is proposed thanks to the use of proteolytic enzymes secreted by thermophilic bacteria Thermoanaerobacter, Thermosipho, and Thermococcus subsp. and mesophilic soil bacteria Streptomyces subsp. Keratins alpha and beta, which resemble amyloid structures, were used as the substrates for the screening for microorganisms able to grow on keratins and producing efficient proteases specific for hydrolysis of beta-sheeted proteic structures, hence amyloids. Secretion of keratin-degrading proteases was evidenced by a zymogram method. Enzymes from thermophilic strains VC13, VC15, and S290 and Streptomyces subsp. S6 were strongly active against amyloid recombinant ovine prion protein and animal meal proteins. The studied proteases displayed broad primary specificities hydrolyzing low molecular mass peptide model substrates. Strong amyloidolytic activity of detected proteases was confirmed by experiments of hydrolysis of PrPres in SAFs produced from brain homogenates of mice infected with the 6PB1 BSE strain. The proteases from Thermoanaerobacter subsp. S290 and Streptomyces subsp. S6 are the best candidates for neutralization/elimination of amyloids in meat and bone meal and other protein-containing substances and materials.     

Small urban woodlands as biodiversity conservation hot-spot: a multi-taxon approach

To evaluate the importance of urban woodlands to serve as potential sites for biodiversity conservation, we analysed bird, carabid beetle and small mammal community responses to urbanisation at different spatial scales. We analysed the relationships between the variations of the structure (species richness S, diversity H′ and dominance D) of animal communities of woodlands distributed along a rural–urban gradient, and the variations along this same gradient of (1) the vegetation within woodlands, (2) the landscape at 100 m and (3) 600 m around the woodlands. We identified the spatial scales whose variations along the gradient most affected each animal community structure, and characterised community responses to these variations. Our results showed that urbanisation affected taxa differently according to their dispersal ability. Carabid beetles, less mobile, seem to be sensitive to increasing fragmentation and built surfaces from periurban to town centre which could make their movement within the urban landscape difficult. Birds, mobile species, seem to be more sensitive to variations of the vegetation structure within woodlands from periurban to town centre that could affect their capacity to maintain in habitat patches. Although our study did not allow relating the small mammal community structure to urbanisation, it suggests that this taxa is sensitive to urban local disturbances. A relevant management scale of woodlands can be specified for each taxa conservation. Urban woodlands accommodate over 50% of the species present in periurban woodlands, and effective management could enhance this number. Woodlands seem to be a good choice for promoting biodiversity conservation in towns.     

La laccase acide des champignons supérieurs. Relation entre la présence de cet enzyme et les conditions de développement chez Agaricus campestris

Résumé Il a été montré que les champignons supérieurs produisent une oxydase qui catalyse l'oxydation des polyphénols, de la p. phénylènediamine et de l'acide ascorbique en milieu acide. D'après la nature des subtrats oxydés cet enzyme s'apparente à la laccase. Les différentes valeurs du pH optimum des enzymes provenant d'espèces ces diverses et qui s'étagent entre 3,5 et 7,2 pourraient s'interpréter par la production, dans chaque espèce étudiée, d'un mélange en proportions variées de deux enzymes très voisins: une laccase normale et une laccase acide qui ne se sépareraient pas au cours du traitement de purification mis en oeuvre. Le pH optimum de la laccase produite par le mycélium d'A. campestris cultivé dans les conditions habituelles est 7,0, mais, lorsque le mycélium est cultivé sur certains milieux synthétiques, le pH optimum de la laccase synthétisée est 5,6.Ainsi, la production de laccase acide n'apparaît pas comme une propriété spécifique, elle est en rapport avec la composition du milieu de culture.

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Summary It is shown, that in higher fungi an oxydase is produced which catalyses the oxydation of polyphenols, of p-phenylene-diamine and ascorbic acid in an acid medium.From the nature of substrates this enzyme is closely related to laccase. Different species of fungi show different values of the pH-optimum, between 3,5 and 7,2; this could be interpreted by assuming that every species contains a mixture of two related enzymes: a normal laccase and an acid laccase which could not be separated during the purification.The pH optimum of the laccase ofAgaricus campestris, cultivated in the usual medium, is 7,0; after cultivation in certain synthetic medium the pH optimum of the formed laccase is 5,6. Therefore the formation of an acid laccase is not a specific property but appears to be related with the composition of the culture medium.

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Zusammenfassung Es wurde gezeigt, dass höhere Pilze Oxydase erzeugen, die die Oxydation der Polyphenole, des p-Phenylendiamins und der Ascorbinsäure in saurer Lösung katalysieren. Die Natur der Substrate zeigt an, dass das Enzym mit der Laccasse verwandt ist.Das pH-Optimum des Enzyms ist je nach der Pilzart verschieden, und liegt zwischen den Werten von pH 3,5 und 7,3; es wäre möglich die Verschiedenheit dieser Werte dahin zu deuten, dass es sich je nach der Pilzart um ein Gemisch von zwei sich sehr nahe stehenden Enzymen handelt: einer normalen und einer sauren Laccase, die durch Reinigung nicht trennbar sind.Die Laccase, die — durch das Myzel vonAgaricus campestris gezüchtet — auf den üblichen Nährlösungen gebildet wird, besitzt eine optimale Aktivität bei pH 7; diese optimale Aktivität liegt bei pH 5,6 wenn das Myzel auf synthetischen Nährlösungen wächst.Die Bildung der sauren Laccase ist also in diesem Fall keine spezifische Eigenschaft der Pilzart, sondern ist durch die Zusammensetzung der Nährlösung bestimmt.

     

Expert judgement in a risk assessment model for Salmonella spp. in pork: The performance of different weighting schemes

A structured expert judgement study was carried out in order to obtain input parameters for a quantitative microbial risk assessment (QMRA) model. This model aimed to estimate the risk of human Salmonella infections associated with the consumption of minced pork meat. Judgements of 11 experts were used to derive subjective probability density functions (PDFs) to quantify the uncertainty on the model input parameters.The performance of experts as probability assessors was measured by the experts’ ability to correctly and precisely provide estimates for a set of seed variables (=variables from the experts’ area of expertise for which the true values were known to the analyst). Subsequently different weighting schemes or “decision makers” (DMs) were applied using Cooke's classical model in order to obtain combined PDFs as a weighted linear combination of the expert's individual PDFs.The aim of this study was to compare the performance of four DMs namely the equal weight DM (each expert's opinion received equal weight), the user weight DM (weights are determined by the expert's self-perceived level of expertise) and two performance-based DMs: the global weight DM and the item weight DM. Weights in the performance-based DMs were calculated based on the expert's calibration and information performance as measured on the set of seed variables.The item weight DM obtained the highest performance with a calibration score of 0.62 and an information score of 0.52, as compared to the other DMs. The weights of the performance-based DMs outperformed those of the best expert in the panel. The correlation between the scores for self-rating of expertise and the weights based on the experts’ performance on the calibration variables was low and not significant (r = 0.37, p = 0.13).The applied classical model provided a rational basis to use the combined distributions obtained by the item weight DM as input in the QMRA model since this DM yielded generally more informative distributions for the variables of interest than those obtained by the equal weight and user weight DM. Attention should be paid to find adequate and relevant seed variables, since this is important for the validation of the results of the weighting scheme.     

Meat retail conditions within the establishments of Kigali city (Rwanda): bacteriological quality and risk factors for <Emphasis Type="Italic">Salmonella</Emphasis> occurrence

Meat constitutes one of the major vehicles for human foodborne infections. This study aimed to assess the retail conditions and to determine the microbiological quality and safety of meat retailed within the establishments of Kigali (Rwanda). A questionnaire survey was carried out in 150 retail outlets to characterise meat retail conditions. Additionally, 270 retail meat samples were analysed for the enumeration of hygiene indicator bacteria (total mesophilic bacteria and Escherichia coli) and for the qualitative detection of Salmonella, using conventional culture methods. The results revealed that beef was the predominant meat sold within the retail premises of Kigali city, while meat from non-bovine animal species was mainly sold in large establishments. Salmonella was detected in 19.6% of all the retailed meat samples evaluated, whereas the mean loads for total mesophilic bacteria and E. coli were 7.3 and 3.5 log cfu/g, respectively. Three factors, namely the temperature conditions of the meat under retail, the cleanability of the used meat cutting boards, and the training of personnel in hygienic meat handling practices, were found to be significantly (p ≤ 0.05) associated with the risk of Salmonella occurrence in the retailed meat. The findings from this study highlight the need for improvements in hygienic meat handling practices, particularly, in small and medium meat retail establishments in Kigali.     

Effects of a quantitative trait locus for muscle hypertrophy from Belgian Texel sheep on carcass conformation and muscularity

A QTL for muscle hypertrophy has been identified in the Belgian Texel breed. A population of F2 and backcross lambs created from crosses of Belgian Texel rams with Romanov ewes was studied. Effects on carcass traits and muscle development of the Belgian Texel breed polygenes and Belgian Texel single QTL were compared. In both cases, carcass conformation and muscularity were improved. The Texel polygenic environment improved conformation mainly through changes in skeletal frame shape. Segments were shorter and bone weight lower. Muscles were more compact, shorter, and thicker. The single QTL affected muscle development. Thickness and weight of muscles were increased. Composition in myosin changed toward an increase of fast contractile type. The relative contribution of hind limb joint to carcass weight was increased. Differences in skeletal frame morphology among the three genotypes of the single QTL were small. Conformation scoring was mainly influenced by leg muscularity. Back and shoulder muscle development, which largely contributed to variability of muscularity, were less involved in the conformation scoring. Lastly, the QTL explains a small part of differences between these Belgian Texel and Romanov breeds for conformation or muscle development. A large part of genetic variability remains to be explored.     

Tracing carbon uptake from a natural CO2 spring into tree rings: an isotope approach

We analyzed 14C, 13C and 18O isotope variations over a 50-year period in tree rings of Quercus ilex L. trees growing at a natural CO2 spring in a Mediterranean ecosystem. We compared trees from two sites, one with high and one with low exposure to CO2 from the spring. The spring CO2 is free of 14C. Thus, this carbon can be traced in the wood, and the amount originating from the spring calculated. The amount decreased over time, from about 40% in 1950 to 15% at present for the site near the spring, indicating a potential difficulty in the use of natural CO2 springs for elevated CO2 research. The reason for the decrease may be decreasing emission from the spring or changes in stand structure, e.g., growth of the canopy into regions with lower concentrations. We used the 14C-calculated CO2 concentration in the canopy to determine the 13C discrimination of the plants growing under elevated CO2 by calculating the effective canopy air 13C/12C isotopic composition. The trees near the spring showed a 2.5 per thousand larger 13C discrimination than the more distant trees at the beginning of the investigated period, i.e., for the young trees, but this difference gradually disappeared. Higher discrimination under elevated CO2 indicated reduced photosynthetic capacity or increased stomatal conductance. The latter assumption is unlikely as inferred from the 18O data, which were insensitive to CO2 concentration. In conclusion, we found evidence for a downward adjustment of photosynthesis under elevated CO2 in Q. ilex in this dry, nutrient-poor environment.     

Soil carbon and nitrogen dynamics linked to Piliostigma species in ferugino-tropical soils in the Sudano-Sahelian zone of Burkina Faso, West Africa

In the Sudano-Sahelian zone of Burkina Faso, Piliostigma reticulatum (DC) Hochst and Piliostigma thonningii (Schumach) are precursor species of fallow land colonization and they are used by rural villagers. The present study aimed to assess the contribution of Piliostigma species to soil quality improvement. We quantified organic carbon, total nitrogen, soil microbial biomass, soil basal respiration and metabolic quotient from soil samples taken under and outside Piliostigma canopies. We used one-way ANOVA to test for differences in the above parameters between locations (beneath and outside Piliostigma canopies). We recorded increased total organic carbon under Piliostigma from 31%-105% and in total nitrogen from 23%-66%. Microbial biomass was13%-266% higher beneath canopies as compared to outside canopies. Basal respiration was also higher beneath canopies. The chemical elements varied by class of soil texture. Metabolic quotient (qCO2 ) was significantly correlated to clay (r=0.80) and silt (r=0.79) content. Piliostigma stands produced abundant litter due to their leaf biomass. Thus, they contribute to improved total organic carbon and total nitrogen content in the different phytogeographic zones and improve soil fertility     

Spatial pattern of beech (<Emphasis Type="Italic">Fagus sylvatica</Emphasis> L.) and oak (<Emphasis Type="Italic">Quercus pubescens</Emphasis> Mill.) seedlings in natural pine (<Emphasis Type="Italic">Pinus sylvestris</Emphasis> L.) woodlands

Spatial pattern of recruitment is an important factor influencing population dynamics of plant communities. The spatial pattern is determined by seed dispersal and by the spatial variability of germination and initial survival. In the process of forest expansion following farmland abandonment, mid- and late-successional species are often dispersed in pioneer forests by birds. This could result in an aggregated spatial pattern of seeds that could influence the dynamics of these species in mixed pioneer forests. In the sub-Mediterranean area, mid- and late-successional species such as Quercus pubescens (downy oak) and Fagus sylvatica (European beech) are expected to replace pioneer Pinus species. Using a point sampling method we demonstrated that beech and oak seedlings (height <2 m) have a clumped distribution in the understorey of pine. This could result from an aggregated dispersal by jays (dispersal effect) or from preferential recruitment in particular habitats (habitat effect). To test these hypotheses we proposed a statistical analysis of spatial patterns of regeneration of beech and oak. Ground cover variables (i.e. cover by rock outcrops, herbs, box shrubs, mosses or pine) did not differ significantly around seedlings as compared with random sample plots. Likewise, clumped seedlings had growth similar to isolated seedlings, thus refuting the hypothesis of preferential location in the most favourable microsites. Aggregated dispersal seems to be involved in the process of regeneration. Since beech and oak seedlings have contrasting ecological demands, we discuss the implication of this pattern for the replacement dynamics of pine by these species.