Effects of ethidium bromide on DNA loop organisation in human lymphocytes measured by anomalous viscosity time dependence and single cell gel electrophoresis.

The effects of ethidium bromide (EtBr) on human lymphocytes were studied by the method of anomalous viscosity time dependence (AVTD) and by the comet assay. EtBr at low concentrations increased the maximum viscosity and time of radial migration as measured with AVTD at neutral conditions of lysis. A pronounced relaxation of DNA loops was observed with the neutral comet assay. The maximal comet length corresponded to 2 Mb DNA loops. At high concentrations of EtBr, 2 mg/ml, significant reduction in AVTD below control level was seen that suggested hypercondensation of chromatin. The hypercondensation was directly observed with the neutral comet assay. EtBr did not induce DNA strand breaks as measured by the alkaline comet assay. The hypercondensed nuclei could be decondensed by irradiation with gamma-rays or exposure to light. The data provide evidence that EtBr at high concentrations resulted in hypercondensation of chromatin below control level. The comet assay confirmed that the increase in AVTD peaks deals with relaxation of loops and AVTD decrease is caused by chromatin condensation. The prediction of the AVTD theory for a correlation between time of radial migration and condensation of chromatin was verified. Further, the data show that the comet assay at neutral conditions of lysis is rather sensitive to DNA loop relaxation in the absence of DNA damage. Finally, donor specificity was found for the hypercondensation.     

An improved negative staining technique using a thin quartz membrane as sample support

A negative staining technique is presented based on the use of 40-60 nm quartz membrane supported by a silicon grid. The quartz membrane is fabricated by thermal growth of silicon dioxide on a silicon substrate followed by an anisotropic silicon etching step giving rectangular holes in the silicon substrate. The hydrophilic membrane is shown to be ideally suited for negative staining due to its spreading characteristics, homogeneity, heat resistance and mechanical stability. Micrographs of phage lambda are presented showing the detailed structure of the tail. A simple method of calculating the number of adsorbed particles based on diffusion limited association is also presented.     

DNA-bound proteins contribute much more than soluble intracellular compounds to the intrinsic protection against radiation-induced DNA strand breaks in human cells

To assess the role of soluble intracellular compounds and DNA-bound proteins in the intrinsic protection against radiation-induced DNA strand breaks, the alkaline unwinding technique was applied to cellular, nuclear, and nucleoid monolayers. It was found that, when the soluble intracellular compounds were removed from human fibroblasts by permeabilization (nuclear monolayers) and irradiated in a phosphate buffer containing 150 mM monovalent cations (Na+ and K+) and 0.8 mM MgCl2, the frequency of radiation-induced DNA strand breaks increased twofold. Removal of both soluble intracellular compounds and DNA-bound proteins from the cells by a pretreatment with 2 M NaCl (nucleoid monolayers) resulted in a 100-fold increase in the frequency of strand-break induction by gamma radiation. Expressed as percentage of total intrinsic protection against radiation-induced DNA strand breaks, DNA-bound protein contributed 99% compared to 1% by soluble intracellular compounds. Using a different experimental approach it was found that the radioprotective capacity of soluble intracellular compounds was equivalent to about 5 mM dimethyl sulfoxide (DMSO) and DNA-bound proteins to about 70 mM DMSO. It is concluded that DNA-bound proteins play a much greater role than soluble intracellular compounds in the intrinsic protection against radiation-induced DNA strand breaks in cultured human cells.     

A molecular phylogeny of the north‐east Atlantic species of the genus Idotea (Isopoda) with focus on the Baltic Sea

Today, the marine isopods of the genus Idotea Fabricius, 1798 consist of 26 accepted species. Most species can be found in the Northern Hemisphere. While some species have a cosmopolitan distribution, others are endemic to a few coastlines. In the Baltic Sea, Idotea species have a key role as important meso‐grazers. Only three species can be found in this brackish environment, I. balthica, I. chelipes and I. granulosa, while nine species are described from the whole European coast. The goal of the present study was to reconstruct the phylogeny of the genus Idotea and to test whether the Baltic low‐salinity tolerant species represent a single lineage within the genus. The phylogeny of north‐east Atlantic Idotea species was investigated based on a fragment of the nuclear gene 28S and the mitochondrial gene COI for eight Idotea species. The phylogenetic reconstructions showed that the genus Idotea might not be monophyletic. Neither I. linearis nor I. urotoma did belong to the separated and well‐supported Idotea clade of I. balthica, I. chelipes, I. emarginata, I. granulosa, I. metallica and I. pelagica. The three Idotea species found in the Baltic belonged to different lineages, with estimated COI‐based divergence time older than 7 Myr. This suggests that the tolerance to low salinity has likely evolved in the genus Idotea more than once.     

Total sleep deprivation does not significantly degrade semantic encoding

ABSTRACT

Sleep deprivation impairs performance on cognitive tasks, but it is unclear which cognitive processes it degrades. We administered a semantic matching task with variable stimulus onset asynchrony (SOA) and both speeded and self-paced trial blocks. The task was administered at the baseline and 24 hours later after 30.8 hours of total sleep deprivation (TSD) or matching well-rested control. After sleep deprivation, the 20% slowest response times (RTs) were significantly increased. However, the semantic encoding time component of the RTs remained at baseline level. Thus, the performance impairment induced by sleep deprivation on this task occurred in cognitive processes downstream of semantic encoding.     

Effect of temperature on seed production in the invasive grass Glyceria maxima (Hartm.) Holmb. (Poaceae) in South Africa

Temperature is one of the main factors that determine sexual reproduction in terrestrial and emergent aquatic plant species. The effect of temperature on sexual reproduction and seed production of Glyceria maxima (Hartm.) Holmb. in the southern hemisphere is unknown. Glyceria maxima collections in February 2010 at three isolated infestations in KwaZulu-Natal failed to yield a single seed, only empty panicles. Laboratory experiments showed that vernalisation had no consistent effect on seed production. Field- and laboratory-grown plants produced seeds in the 2010/2011 season, because of having sufficient time at optimum temperatures required for seed production (1 491 and 1 585 hours, respectively), compared to a shorter period (1 352 hours) of suitable temperatures during the 2009/2010 growing season. An inadequate period of optimum temperatures (15–25°C) during seed production resulted in the lack of seeds in the field in the 2009/2010 growing season. This study showed that temperature and duration of exposure thereto during the seed-production period play vital roles in G. maxima sexual reproduction.     

Evidence for diet partitioning among three coexisting native freshwater fishes in South Africa's Cape Fold Ecoregion

The partitioning of limited resources commonly explains how different species can coexist within the same ecological community. In this 2010 study, the diets of three coexisting freshwater fishes (Cape galaxias Galaxias zebratus, n = 27; Cape kurper Sandelia capensis, n = 60; Breede River redfin Pseudobarbus burchelli, n = 77) were characterised and compared in three headwater streams in South Africa's Cape Fold Ecoregion using gut contents and stable isotope analyses. These data were analysed to ascertain whether the three species exploit distinct trophic niches. Both approaches provided evidence that these species occupy different trophic niches, though with some overlap. However, dietary differences among sites were not consistent and were probably influenced by site-specific factors like resource availability. Pseudobarbus burchelli had a broader niche breadth at Tierkloof Stream than the other two species, but not at Waaihoek or Tierstel Streams. Our results also suggest that P. burchelli consumed a more omnivorous diet than do the other two species, whereas S. capensis occupied a higher trophic position than the other two species and consumed vertebrates. Our findings suggest that these species occupy non-equivalent feeding niches in Cape Fold Ecoregion headwater streams, and that diet partitioning might facilitate their coexistence in these systems.     

Lactate dehydrogenase (LDH) gene duplication during chordate evolution: the cDNA sequence of the LDH of the tunicate Styela plicata

L-Lactate dehydrogenase (L-LDH, E.C. 1.1.1.27) is encoded by two or three loci in all vertebrates examined, with the exception of lampreys, which have a single LDH locus. Biochemical characterizations of LDH proteins have suggested that a gene duplication early in vertebrate evolution gave rise to Ldh-A and Ldh-B and that an additional locus, Ldh-C arose in a number of lineages more recently. Although some phylogenetic studies of LDH protein sequences have supported this pattern of gene duplication, others have contradicted it. In particular, a number of studies have suggested that Ldh-C represents the earliest divergence among vertebrate LDHs and that it may have diverged from the other loci well before the origin of vertebrates. Such hypotheses make explicit statements about the relationship of vertebrate and invertebrate LDHs, but to date, no closely related invertebrate LDH sequences have been available for comparison. We have attempted to provide further data on the timing of gene duplications leading to multiple vertebrate LDHs by determining the cDNA sequence of the LDH of the tunicate Styela plicata. Phylogenetic analyses of this and other LDH sequences provide strong support for the duplications giving rise to multiple vertebrate LDHs having occurred after vertebrates diverged from tunicates. The timing of these LDH duplications is consistent with data from a number of other gene families suggesting widespread gene duplication near the origin of vertebrates. With respect to the relationships among vertebrate LDHs, our data are not consistent with previous claims that Ldh-C represented the earliest divergence. However, the precise relationships among some of the main lineages of vertebrate LDHs were not resolved in our analyses.