Development of a real-time PCR and a sandwich ELISA for detection of potentially allergenic trace amounts of peanut (Arachis hypogaea) in processed foods

Hidden allergens in food products are, especially for peanut-allergic consumers, a serious problem because even low amounts (approximately 200 microg) of peanut can elicit allergic reactions. Undeclared peanut traces can be found in processed food products, because contaminations with peanut during production processes are frequent. To minimize the risk of such cross-contaminations, it is necessary to develop sensitive analytical methods for the detection of hidden allergens in foods. For this approach we developed two peanut-specific assays based on the detection of peanut protein by specific antibodies (sandwich ELISA) and by the detection of peanut-specific DNA (part of the coding region of Ara h 2) by a real-time PCR. Both tests did not show any cross-reactivity with 22 common food ingredients (cereals, nuts, legumes), and the limit of detection is <10 ppm peanut in processed foods. Thirty-three random samples of food products were tested for the presence of peanut to compare both assay types with each other and to evaluate the percentage of foods on the German market that are contaminated with peanut traces. We found that four products (13.3%) without peanut in the list of ingredients contained peanut protein in a range from 1 to 74 ppm peanut protein and that the results of both tests correlated well. The real-time PCR was able to detect one more positive sample than the sandwich ELISA. In conclusion, both assays are sensitive and specific tools for the detection of hidden allergens in processed foods.     

Evaluation of oral fluids for surveillance of foodborne and zoonotic pathogens in pig farms

The use of oral fluid (OF) to detect zoonotic pathogens in pigs has been only scarcely assessed. We evaluated OF as a potential specimen for detection by culture of methicillin-resistant Staphylococcus aureus (MRSA) and Yersinia enterocolitica, and the detection of antibodies against Salmonella spp. and hepatitis E virus (HEV) using commercial ELISAs. Samples from 33 pig farms were collected at the beginning and end of the fattening period. Results of the OF samples were compared with the results of serum samples and nasal swabs from individual pigs and pen floor fecal samples, using the Cohen kappa (κ) and the McNemar test. For Salmonella spp. antibodies, OF samples were negative, although the corresponding serum samples were positive. The detection of HEV antibodies in sera and OF had agreement at the first sampling, and poor and significant agreement at the second sampling (κ = 0.185, McNemar p = 0.238; κ = 0.088, McNemar p < 0.001). At both sampling times, the detection of MRSA in nasal swabs and OF showed agreement (κ = 0.466, McNemar p = 0.077; κ = 0.603, McNemar p = 1); agreement was seen for the detection of Y. enterocolitica in fecal and OF samples (κ = 0.012, McNemar p = 0.868; κ = 0.082, McNemar p = 0.061, respectively). According to the McNemar test, the use of pen-based OFs is more feasible for the detection of MRSA and Y. enterocolitica by culture than is detection of antibodies by commercial ELISA.     

Territorial division: the least-time constraint behind the formation of subnational boundaries

Nations usually locate their smaller administrative subdivisions in regions of highest population density. This report derives a precise form of the size-density relationship from the general assumption that social structures evolve in such a way as to minimize the total time expended by society in their operation. The result is confirmed empirically.     

Common sequence polymorphisms shaping genetic diversity in Arabidopsis thaliana

The genomes of individuals from the same species vary in sequence as a result of different evolutionary processes. To examine the patterns of, and the forces shaping, sequence variation in Arabidopsis thaliana, we performed high-density array resequencing of 20 diverse strains (accessions). More than 1 million nonredundant single-nucleotide polymorphisms (SNPs) were identified at moderate false discovery rates (FDRs), and approximately 4% of the genome was identified as being highly dissimilar or deleted relative to the reference genome sequence. Patterns of polymorphism are highly nonrandom among gene families, with genes mediating interaction with the biotic environment having exceptional polymorphism levels. At the chromosomal scale, regional variation in polymorphism was readily apparent. A scan for recent selective sweeps revealed several candidate regions, including a notable example in which almost all variation was removed in a 500-kilobase window. Analyzing the polymorphisms we describe in larger sets of accessions will enable a detailed understanding of forces shaping population-wide sequence variation in A. thaliana.     

Whole-genome shotgun sequencing of mitochondria from ancient hair shafts

Although the application of sequencing-by-synthesis techniques to DNA extracted from bones has revolutionized the study of ancient DNA, it has been plagued by large fractions of contaminating environmental DNA. The genetic analyses of hair shafts could be a solution: We present 10 previously unexamined Siberian mammoth (Mammuthus primigenius) mitochondrial genomes, sequenced with up to 48-fold coverage. The observed levels of damage-derived sequencing errors were lower than those observed in previously published frozen bone samples, even though one of the specimens was >50,000 14C years old and another had been stored for 200 years at room temperature. The method therefore sets the stage for molecular-genetic analysis of museum collections.     

Millimetre scale aeration of the rhizosphere and drilosphere

Soil aeration is a critical factor for oxygen-limited subsoil processes, as transport by diffusion and advection is restricted by the long distance to the free atmosphere. Oxygen transport into the soil matrix is highly dependent on its connectivity to larger pore channels like earthworm and root colonised biopores. Here we hypothesize that the soil matrix around biopores represents different connectivity depending on biopore genesis and actual coloniser. We analysed the soil pore system of undisturbed soil core samples around biopores generated or colonised by roots and earthworms and compared them with the pore system of soil, not in the immediacy of a biopore. Oxygen partial pressure profiles and gas relative diffusion was measured in the rhizosphere and drilosphere from the biopore wall into the bulk soil with microelectrodes. The measurements were linked with structural features such as porosity and connectivity obtained from X-ray tomography and image analysis. Aeration was enhanced in the soil matrix surrounding biopores in comparison to the bulk soil, shown by higher oxygen concentrations and higher relative diffusion coefficients. Biopores colonised by roots presented more connected lateral pores than earthworm colonised ones, which resulted in enhanced aeration of the rhizosphere compared to the drilosphere. This has influenced biotic processes (microbial turnover/mineralization or root respiration) at biopore interfaces and highlights the importance of microstructural features for soil processes and their dependency on the biopore's coloniser.     

Stereoselective formation of the varietal aroma compound rose oxide during alcoholic fermentation

The potent aroma compound rose oxide was quantified in several white wines by a headspace solid-phase microextration stable isotope dilution assay (HS-SPME-SIDA) and the enantiomeric ratios of the cis diastereomers were determined by enantioselective capillary GC. The most odor-active stereoisomer (23)-cis-rose oxide was detectable in all investigated white wines ranging from 0.2 to 12 microg/L. However, its contribution to the overall aroma in some white wine varieties can be neglected as indicated by a low odor activity value (OAV). The highest concentrations were found in Gewürztraminer wines, confirming the importance of rose oxide as a varietal aroma compound in this variety. Surprisingly, the enantiomeric ratio of cis-rose oxide in all investigated wines was substantially lower than in nonfermented musts and in some wines almost racemic cis-rose oxide was detected. Fermentation studies with a model must that contained deuterated water revealed that yeast is capable of reducing the precursor 3,7-dimethyl octa-2,5-dien-1,7-diol (geranyl diol I) yielding 3,7-dimethyl-5-octen-1,7-diol (citronellyl diol I) that gives rise to cis- and trans-rose oxide after acid catalyzed cylization. The deuterium labeling pattern of the resulting rose oxide stereoisomers and a clearly detectable kinetic isotope effect indicate that at least two different reductive pathways in yeast exist that yield cis-rose oxide with different enantiomeric ratios altering the genuine enantiomeric ratio in grape musts. The presence of (+)-cis-rose oxides in wines can therefore be attributed to the reductive yeast metabolism during fermentation. This observation corroborates recent findings that the modification of terpene derived varietal aroma is an integral part of yeast metabolism and not only a simple hydrolytical process.     

Soil-plant hydrology of indigenous and exotic trees in an Ethiopian montane forest

Fast-growing exotic trees are widely planted in the tropics to counteract deforestation; however, their patterns of water use could be detrimental to overall ecosystem productivity through their impact on ecosystem water budget. In a comparative field study on seasonal soil-plant water dynamics of two exotic species (Cupressus lusitanica Mill. and Eucalyptus globulus Labill.) and the indigenous Podocarpus falcatus (Thunb.) Mirb. in south Ethiopia, we combined a 2.5-year record for climate and soil water availability, natural-abundance oxygen isotope ratios (delta(18)O) of soil and xylem water, destructive root sampling and transpiration measurements. Soil was generally driest under C. lusitanica with its dense canopy and shallow root system, particularly following a relatively low-rainfall wet season, with the wettest soil under E. globulus. Wet season transpiration of C. lusitanica was twice that of the other species. In the dry season, P. falcatus and C. lusitanica reduced transpiration by a factor of six and two, respectively, whereas E. globulus showed a fivefold increase. In all species, there was a shift in water uptake to deeper soil layers as the dry season progressed, accompanied by relocation of live fine root biomass (LFR) of C. lusitanica and P. falcatus to deeper layers. Under P. falcatus, variability in soil matric potential, narrow delta(18)O depth gradients and high LFR indicated fast water redistribution. Subsoil water uptake was important only for E. globulus, which had low topsoil LFR and tap roots exploiting deep water. Although P. falcatus appeared better adapted to varying soil water availability than the exotic species, both conifers decreased growth substantially during dry weather. Growth of E. globulus was largely independent of topsoil water content, giving it the potential to cause substantial dry-season groundwater depletion.