Fractionation of DNA with Sephacryl S-1000

In this study the application of gel filtration for purification of heterogeneous DNA is described. The fractionation of partial restriction enzyme digests of bacterial chromosomal DNA on a Sephacryl S-1000 -column is easy and rapid. Simultaneously intact chromosomal DNA and low molecular weight substances are eliminated in the run. The method is also applicable to the purification of plasmid DNA, as has been previously reported (3). Thus we are able to get pure DNA with yields over 80%.     

Effect of boundary type and season on predatory arthropods associated with field margins on New Zealand farmland

Pitfall traps were used to monitor predatory arthropod numbers along two types of field boundary, a post and wire fence line and a Cupressus macrocarpa hedge, along the same paddock margin in Canterbury, New Zealand, over 24 months. The seven most abundant predator groups recorded were: Araneae > Phalangiidae > Staphylinidae > Coccinellidae > Chilopoda > Hemerobiidae > Carabidae. Araneae, Phalangiidae, Staphylinidae, Chilopoda and Hemerobiidae were found in larger numbers at the wire fence than at the hedge site, whereas the numbers of Carabidae and Coccinellidae adults exhibited no field margin preference. However, more species of Araneae and Staphylinidae were caught at the hedge site, whereas species richness of carabid beetles was greatest at the wire fence. Principal component analysis clearly separated the samples collected from the two habitats based on the assemblages of Araneae, Staphylinidae and Carabidae, and certain species in each of these taxonomic groups appeared to be particularly associated with one boundary type or the other. All the main taxonomic groups exhibited clear seasonal patterns, with distinct peaks in abundance occurring at certain times of the year. The results of the study reinforce the idea that management of field boundaries can be used to manipulate the type and abundance of particular groups of predatory arthropods, and that seasonal patterns should be taken into account in schemes of integrated pest management so that any adverse effects of biocide application on these beneficial species may be minimised.     

Ecological inference on bacterial succession using curve-based community fingerprint data analysis, demonstrated with rhizoremediation experiment

Nucleic acid-based community fingerprinting methods are valuable tools in microbial ecology, as they offer rapid and robust means to compare large series of replicates and references. To avoid the time-consuming and potentially subjective procedures of peak-based examination, we assessed the possibility to apply direct curve-based data analysis on community fingerprints produced with bacterial length heterogeneity PCR (LH-PCR). The dataset comprised 180 profiles from a 21-week rhizoremediation greenhouse experiment with three treatments and 10 sampling times. Curve-based analysis quantified the progressive effect of the plant (Galega orientalis) and the reversible effect of the contaminant (fuel oil) on bacterial succession. The major observed community shifts were assigned to changes in plant biomass and contamination level by canonical correlation analysis. A novel method to extract relative abundance data from the fingerprint curves for Shannon diversity index revealed contamination to reversibly decrease community complexity. By cloning and sequencing the fragment lengths, recognized to change in time in the averaged LH-PCR profiles, we identified Aquabacterium (Betaproteobacteria) as the putative r-strategic fuel oil degrader, and K-strategic Alphaproteobacteria growing in abundance later in succession. Curve-based community fingerprint analysis can be used for rapid data prescreening or as a robust alternative for the more heavily parameterized peak-based analysis.     

Side-to-side differences in bone strength in master jumpers and sprinters

Introduction: This study evaluated side-to-side difference in tibial bone structure, calf muscle cross-sectional area (CSA) and hopping force in master athletes as a result of training for sports with different magnitudes of inter-leg loading difference. Methods: Tibial bone parameters (at 4%, 14%, 38% and 66% tibial length proximal to distal end), muscle CSA (at 66% tibial length) and hopping forces of both legs of 51 master athletes (conditioned jumpers, conditioned triple jumpers, unconditioned jumpers, hurdlers and sprinters) were examined using pQCT. In epiphyseal 4% slice bone CSA (Ar.tot), total BMC (vBMC.tot), trabecular BMC (vBMC.tb) cortical BMC (vBMC.ct), and trabecular BMD (vBMD.tb) were measured. In diaphyseal slices, Ar.tot, vBMC.ct, cortical density (vBMD.ct), cross-sectional moment of inertia (CSMI) and calf muscle CSA (MuscA) were examined. Results: In conditioned jumpers, side-to-side differences in favour of take-off leg were found in 4% slice in vBMC.tb (+4.1%) (P<0.05). A side-to-side difference was found in 66% slice vBMC.ct and CSMI (both P<0.05), with conditioned jumper (+2.8% and 6.6%) and triple jumper (+2.7% and 7.2%) values higher than other groups. Conclusion: The results suggest that regular training in high-impact sports with uneven lower limb loading results in side-to-side differences in skeletal adaptation independent of age and gender, suggesting that high-impact exercise is effective in maintaining bone strength throughout human lifespan.     

Description of two biovars in the Rhizobium galegae species: biovar orientalis and biovar officinalis

Twenty-six Rhizobium galegae strains, representing the center of origin of the host plants Galega orientalis and G. officinalis as well as other geographic regions, were used in a polyphasic analysis of the relationships of R. galegae strains. Phage typing, lipopolysaccharide (LPS) profiling, pulsed field gel electrophoresis (PFGE) profiling and rep-PCR (use of repetitive sequences as PCR primers for genomic fingerprinting) with REP and ERIC primers investigated nonsymbiotic properties, whereas plasmid profiling and hybridisation with a nif gene probe, and with nodB, nodD, nod box and an IS sequence from the symbiotic region as probes, were used to reveal the relationships of symbiotic genes. The results were used in pairwise calculations of distances between the strains, and the distances were visualised as a dendrogram. Indexes of association were compared for all tests pooled, and for chromosomal tests and symbiotic markers separately, to display the input of the different categories of tests on the grouping of the strains. Our study shows that symbiosis related genetic traits in R. galegae divide strains belonging to the species into two groups, which correspond to strains forming an effective symbioses with G. orientalis and G. officinalis respectively. We therefore propose that Rhizobium galegae strains forming an effective symbiosis with Galega orientalis are called R. galegae bv. orientalis and strains forming an effective symbiosis with Galega officinalis are called R. galegae bv. officinalis.     

Oral health and reduction in respiratory capacity in a cohort of community-dwelling elderly people: a population-based 5-year follow-up study

Objectives: Several anaerobic bacteria originating in periodontal pockets have been isolated from infected lungs and pharyngeal microflora. Increased bacterial load in lungs is known to be a risk factor for decline in forced expiratory volume during the first second. The aim was to evaluate both cross‐sectionally and longitudinally the association between oral health status and forced expiratory volume during the first second (FEV₁) in older residents of the city of Jyväskylä, Finland. Design: Cross‐sectional and prospective cohort study over a 5‐year follow‐up. Setting: Research laboratory of the University of Jyväskylä. Participants: In 1990 dental status and FEV₁ were examined in 203 80‐year‐old people, of whom 88 survivors were retested 5 years later. Main outcome measures: Primary: dental status and FEV₁. Secondary: existence of pulmonary diseases, height, handgrip strength, smoking, and length of education. Results: Participants were regrouped into three categories according to their baseline oral health status. At baseline, men with complete prostheses had the lowest FEV₁. Five years later the greatest reduction in FEV₁ was seen in subjects with poor periodontal status or complete prostheses (?9.4%) while those with healthy periodontal status showed no reduction in FEV₁ values (+1.0%, p = 0.006). Conclusions: Periodontal infections and complete prostheses may be reservoirs for pathogens which may be harmful and partly explain the observed reduction in FEV₁ during ageing.     

Incorporation of cholesterol into serum high density lipoprotein apoprotein and recombinants

The uptake of cholesterol (CHL) by serum high density lipoprotein (HDL) delipidated apoproteins and phospholipid-apoprotein recombinants has been studied with two methods; by incubation with Celite-dispersed cholesterol or with cholesterol crystals. The apoproteins bind very small amounts of cholesterol with a maximum of about 6 micrograms/mg apoprotein. Recombinants with dimyristoyl phosphatidylcholine (DMPC) or egg phosphatidylcholine (EPC) as phospholipid component gave similar values for cholesterol uptake. The initial rate for uptake from Celite-cholesterol by recombinants was high (0.1 mol cholesterol/mol phospholipid/h) and somewhat higher than that for phospholipid vesicles. The maximal uptake was by gel filtration shown to depend on the size of the complexes with values about 0.95 mol cholesterol per phospholipid for vesicular complexes, 0.75 for discoidal complexes and between 0.5 and 0.2 for small 'protein-rich' complexes. During the incubation of recombinants with cholesterol there was considerable decomposition of discoidal complexes and formation of larger ones. The results show that phospholipid-apoprotein complexes are efficient acceptors for cholesterol but also that about 25% of the phospholipid in the discoidal complexes is excluded from interaction with cholesterol by interaction with apoprotein.     

Association between chloroplast and mitochondrial lineages in oaks

Patterns of chloroplast DNA (cpDNA) and mitochondrial DNA (mtDNA) variation were studied in 378 populations of oak trees sampled throughout the southern half of France. Six cpDNA haplotypes detected in a previous European survey and three new cpDNA haplotypes were found in this region. Two mitochondrial polymorphisms detected earlier by restriction analysis of PCR-amplified fragments alone, or in combination with single-strand conformation polymorphism (SSCP), were compared with the cpDNA data. Sequencing revealed the nature of the two mitochondrial mutations: a single-base substitution and a 4-bp inversion associated with a 22-bp hairpin secondary structure. The single-base substitution was then analyzed by allele-specific amplification. Results for the two cytoplasmic genomes were combined, which allowed the identification of 12 cpDNA-mtDNA haplotypes. The 4-bp mtDNA inversion has appeared independently in different cpDNA lineages. Given the peculiar nature of this mtDNA mutation, we suggest that intramolecular recombination leading to repeated inversions of the 4-bp sequence (rather than paternal leakage of one of the two genomes) is responsible for this pattern. Furthermore, the geographic locations of the unusual cpDNA-mtDNA associations (due to the inversion) usually do not match the zones of contact between divergent haplotypes. In addition, in southern France, the groupings of populations based on the mtDNA substitution were strictly congruent with those based on cpDNA. Because many populations that are polymorphic for both cpDNA and mtDNA have remained in contact since postglacial recolonization in this area without producing any new combination of cytoplasms involving the mitochondrial substitution, we conclude that paternal leakage is not a significant factor at this timescale. Such results confirm and expand our earlier conclusions based on controlled crosses.