Simple agarose gel electrophoretic method for the identification and characterization of plasmid deoxyribonucleic acid.

Agarose gel electrophoresis may be employed effectively for the detection and preliminary characterization of plasmid deoxyribonucleic acid (DNA) present in clinical isolates and laboratory strains of gram-negative microorganisms. The method is sensitive and does not require radioisotopes or ultracentrifugation. The estimation of plasmid mass from the extent of DNA migration in gels compares favorably with results obtained by electron microscopy of plasmid DNA purified by equilibrium density centrifugation. The method has proved to be a useful tool for survey work and the epidemiological investigation of plasmid dissemination, as well as an important adjunct to the genetic analysis of plasmids.     

The enterotoxin gene (cpe) of Clostridium perfringens can be chromosomal or plasmid-borne

The location of the cpe gene, encoding the enterotoxin responsible for food poisoning in humans, has been studied in a series of enterotoxigenic Ciostridium perfringens strains by means of pulsed field gel electrophoresis of genomic DNA. The cpe gene was found at the same chromosomal locus in strains associated with food poisoning in humans and was shown to be linked to a repetitive sequence, the Hin dlll repeat, and an open reading frame, ORF3, that may be part of an insertion sequence. In contrast, when the strains originated from domesticated livestock cpe was located on a large episome where it was often close to a copy of the transposable element IS 1151. In these cases, the Hin dlll repeat was not linked to the cpe gene although this was generally preceded by ORF3.     

Ants impact the composition of the aquatic macroinvertebrate communities of a myrmecophytic tank bromeliad

In an inundated Mexican forest, 89 out of 92 myrmecophytic tank bromeliads (Aechmea bracteata) housed an associated ant colony: 13 sheltered Azteca serica, 43 Dolichoderus bispinosus, and 33 Neoponera villosa. Ant presence has a positive impact on the diversity of the aquatic macroinvertebrate communities (n = 30 bromeliads studied). A Principal Component Analysis (PCA) showed that the presence and the species of ant are not correlated to bromeliad size, quantity of water, number of wells, filtered organic matter or incident radiation. The PCA and a generalized linear model showed that the presence of Azteca serica differed from the presence of the other two ant species or no ants in its effects on the aquatic invertebrate community (more predators). Therefore, both ant presence and species of ant affect the composition of the aquatic macroinvertebrate communities in the tanks of A. bracteata, likely due to ant deposition of feces and other waste in these tanks.     

Physiological girdling of pine trees via phloem chilling: proof of concept

Quantifying below-ground carbon (C) allocation is particularly difficult as methods usually disturb the root-mycorrhizal-soil continuum. We reduced C allocation below ground of loblolly pine trees by: (1) physically girdling trees and (2) physiologically girdling pine trees by chilling the phloem. Chilling reduced cambium temperatures by approximately 18 degrees C. Both methods rapidly reduced soil CO2 efflux, and after approximately 10 days decreased net photosynthesis (P(n)), the latter indicating feedback inhibition. Chilling decreased soil-soluble C, indicating that decreased soil CO2 efflux may have been mediated by a decrease in root C exudation that was rapidly respired by microbes. These effects were only observed in late summer/early autumn when above-ground growth was minimal, and not in the spring when above-ground growth was rapid. All of the effects were rapidly reversed when chilling was ceased. In fertilized plots, both chilling and physical girdling methods reduced soil CO2 efflux by approximately 8%. Physical girdling reduced soil CO2 efflux by 26% in non-fertilized plots. This work demonstrates that phloem chilling provides a non-destructive alternative to reducing the movement of recent photosynthate below the point of chilling to estimate C allocation below ground on large trees.     

Novel high-resolution characterization of ancient DNA reveals C > U-type base modification events as the sole cause of post mortem miscoding lesions

Ancient DNA (aDNA) research has long depended on the power of PCR to amplify trace amounts of surviving genetic material from preserved specimens. While PCR permits specific loci to be targeted and amplified, in many ways it can be intrinsically unsuited to damaged and degraded aDNA templates. PCR amplification of aDNA can produce highly-skewed distributions with significant contributions from miscoding lesion damage and non-authentic sequence artefacts. As traditional PCR-based approaches have been unable to fully resolve the molecular nature of aDNA damage over many years, we have developed a novel single primer extension (SPEX)-based approach to generate more accurate sequence information. SPEX targets selected template strands at defined loci and can generate a quantifiable redundancy of coverage; providing new insights into the molecular nature of aDNA damage and fragmentation. SPEX sequence data reveals inherent limitations in both traditional and metagenomic PCR-based approaches to aDNA, which can make current damage analyses and correct genotyping of ancient specimens problematic. In contrast to previous aDNA studies, SPEX provides strong quantitative evidence that C > U-type base modifications are the sole cause of authentic endogenous damage-derived miscoding lesions. This new approach could allow ancient specimens to be genotyped with unprecedented accuracy.     

Structural studies on the acidic exopolysaccharide from Haloferax denitrificans ATCC 35960.

The structure of a linear, acidic exopolysaccharide isolated from the Archaeon Haloferax denitrificans ATCC 35960 has been determined using NMR spectroscopy. The sugar residues in the repeating unit of the polysaccharide were identified as Gal and GlcA2,3NAc after the assignment of the 1H and 13C resonances using COSY, HOHAHA, HMQC and HMQC-TOCSY experiments. The sequence of the residues in the polysaccharide was established from the inter-residue connectivities observed in the HMQC-NOESY plot. The only sugar released on acid hydrolysis was shown to be D-Gal by GLC analysis, while the absolute configuration of the acidic sugars was shown to be D by comparison of the carbon chemical shifts with those of model compounds. Partial acid hydrolysis yielded a tetrasaccharide, terminated by D-Gal at the reducing end, whose structure confirmed that of the repeating unit of the polysaccharide as-->4)-beta-D-GlcpA2,3NAc-(1-->4)-beta-D-GlcpA2, 3NAc-(1-->4)-alpha-D-GlcpA2,3NAc-(1-->3)-alpha-D-Galp- (1-->, where D-GlcpA2,3NAc is 2,3-diacetamido-2,3-dideoxy-D-glucopyranosiduronic acid.     

Cell motility measurements with an automated microscope system

The motility of 3T3 cells has been studied using a newly developed automated microscope system which is capable of recognizing live unstained cells growing in tissue culture. A large number of individual cells can be rapidly identified and characterized and their precise positions recorded. All cells can be revisited automatically every few minutes, and the new cell positions can be determined. Quantitative data from up to 1 000 cells can then be obtained, and cell movement parameters like cell speed, distance travelled, direction of movement, etc., can be measured for individual cells and for the whole cell population. In addition, for any number of chosen cells, high-resolution digitized images can be taken for further morphological studies, including acquisition of images of individual cells.     

Patterns of dispersion, density and dispersal in alpine populations of the land snail Arianta arbustorum (L.) (Helicidae)

In studying populations of the land snail Arianta arbustorum in the eastern Swiss Alps, juvenile and subadult snails were found to be aggregated all the time, while adults were mostly aggregated during summer, the main mating period. The mean size of aggregations was 25 cm × 25 cm but varied, according to the area a grass tuft covered, while the intensity of aggregations was influenced by weather conditions. Densities of 10–15 subadults and 3–6 adults m⁻² were found along ditches and streams, 2–25 subadults and 2–20 adults on uncultivated alpine meadows, while on scree-covered alpine grassland the density of subadults and adults was 0.3–0.5 m⁻². The extent of daily movements varied with microclimatic factors and season and depended on the structure of the habitat. The dispersal rate was greatest in June (2.6 m month⁻¹) and least in August (1.1 m month⁻¹). In summer and autumn, active uphill movements were noted, a process which compensates for the loss of altitude by passive downhill displacements. Roiling down on snowfields and being carried away by avalanches and streams were observed.