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Phase-contrast illumination is simple and most commonly used microscopic method to observe nonstained living cells. Automatic cell segmentation and motion analysis provide tools to analyze single cell motility in large cell populations. However, the challenge is to find a sophisticated method that is sufficiently accurate to generate reliable results, robust to function under the wide range of illumination conditions encountered in phase-contrast microscopy, and also computationally light for efficient analysis of large number of cells and image frames. To develop better automatic tools for analysis of low magnification phase-contrast images in time-lapse cell migration movies, we investigated the performance of cell segmentation method that is based on the intrinsic properties of maximally stable extremal regions (MSER). MSER was found to be reliable and effective in a wide range of experimental conditions. When compared to the commonly used segmentation approaches, MSER required negligible preoptimization steps thus dramatically reducing the computation time. To analyze cell migration characteristics in time-lapse movies, the MSER-based automatic cell detection was accompanied by a Kalman filter multiobject tracker that efficiently tracked individual cells even in confluent cell populations. This allowed quantitative cell motion analysis resulting in accurate measurements of the migration magnitude and direction of individual cells, as well as characteristics of collective migration of cell groups. Our results demonstrate that MSER accompanied by temporal data association is a powerful tool for accurate and reliable analysis of the dynamic behaviour of cells in phase-contrast image sequences. These techniques tolerate varying and nonoptimal imaging conditions and due to their relatively light computational requirements they should help to resolve problems in computationally demanding and often time-consuming large-scale dynamical analysis of cultured cells. [ABSTRACT FROM AUTHOR]

We report the isolation and cross species amplification of 42 Drosophila virilis microsatellite loci. Nine loci were isolated from mapped P1 bacteriophage clones and 33 were obtained from genomic DNA or GenBank searches. Cross species amplification was tested for all members of the D. virilis group. The amplification success was high (varying from 45% to 100%) and most of the loci were polymorphic. This set of loci can be applied for several genetic studies such as mapping behavioural quantitative trait loci (QTL) and for studying population structure in a phylogeographical framework in D. virilis group species. [ABSTRACT FROM AUTHOR]

ABSTRACTThe effects of enhanced UV-B radiation on the needle anatomy of loblolly pine (Pinus taeda L.) and Scots pine (Pinus sylvestris L.) were studied in the field under supplemental UV-B radiation supplied by a modulated irradiation system. The supplemental UV-B levels were designed to simulate either a 16 or 25% loss of stratospheric ozone over College Park, Maryland. Enhanced UV-B radiation caused different responses in these two species. The needles of loblolly pine had larger amounts of tannin in the lumen of epidermal cells and more wall-bound phenolics in the outer epidermal walls of UV-B-treated needles, whereas the most pronounced effect on Scots pine needles was increased cutinization. In both species, the outer epidermal cell walls thickened and the needle cross-sectional and mesophyll areas decreased (statistically significantly only in Scots pine). This suggests that more carbon may have been allocated to the protection mechanisms at the expense of photosynthetic area. The difference in response between these species suggests that the response to UV-B radiation is not mediated by a single mechanism and that no generalization with regard to the effects of UV-B on conifers can be made. [ABSTRACT FROM AUTHOR]

The superoxide dismutase (SOD; EC 1.15.1.1.) activities in homogenates of Scots pine (Pinus sylvestris) and Norway spruce (Picea abies L. Karst.) needles from rural and urban forest sites were analysed. The cloned pines from northern (specific SOD mean 11.7 U/mg protein) and eastern Finland (specific SOD mean 24.1 U/mg protein) had significantly different SOD activity levels, which indicates different responses under pollution stress. The rural pines had a mean specific SOD activity of 24.9 U/mg protein. The corresponding value for rural spruces was 14.7 U/mg protein. The urban pines under SO2influence had a mean of 29.1 U/mg protein, while the urban spruces under SO2 pollution had a mean of 19.2 U/mg protein. The NOx polluted site revealed high total activity, but low specific activity. [ABSTRACT FROM AUTHOR]

The response of epicuticular wax of Scots pine (Pinus sylvestris L.) needles to dry- and wet- deposited sulphur and heavy metals was investigated at six sites located 10-110 km from the Monchegorsk Cu-Ni smelter on the Kola Peninsula, North-West Russia, and in a long-term irrigation experiment where pines were exposed over four growing seasons (1991-1994) to either acid rain treatment at pH 3.1 (H2SO4), metal treatment at pH 5.7 (Cu and Ni) or a combination of these at pH 3.1. Needle wettability exhibited a closer relationship with epistomatal wax tube distribution (WTD) than with chemical composition of epicuticular wax. Water droplet contact angles (DCA) decreased towards the smelter, and significant differences due to site were noted for 26-month-old and 38-month-old needles. Significant differences due to site were determined for secondary alcohols, dehydroabietic acid and hydroxy fatty acids, the proportions of which ranged from 22.5 to 48.9%, 6.2 to 22.4 % and 0.6 to 2.6% respectively, depending on site and needle age class. The proportion of dehydroabietic acid increased towards the smelter, but no gradient was observed in the proportion of secondary alcohols or hydroxy fatty acids. No major effect of experimentally applied pollutants on the chemical composition or structure of the epicuticular wax was observed. The effect of treatment on DCA was significant in 1993 and 1994 due to a 6.5-13.2 degree greater wettability of the 37-49-month-old acid-treated needles relative to the irrigated or dry controls. Sulphuric acid at pH 3.1 did not increase needle wettability when combined with copper and nickel sulphate in similar concentrations. These data indicate that S deposition, especially H2SO4, plays a more important role in needle surface deterioration than Cu and Ni. Pollutant-induced changes in epicuticular wax structure and needle wettability mimic natural wax ageing, but at an accelerated rate. Changes in wax chemical composition might also be caused by pollutant-induced metabolic changes in elongating needles. [ABSTRACT FROM AUTHOR]

The article reports on the effects of excess nitrogen fertilization on the structure of Scots-pine needles. The thickness and width of the needle and thickness of the tissues were measured, and the conditions of various cell types were classified at the light microscopy level. The natural xeromorphic structure of the conifer needles became mesomorphic, possibly influencing the protection afforded against environmental stresses. A high level of nitrogen fertilization caused changes in the central cylinder, especially in the vascular bundle and the sclerenchyma.

The growth and morphology of Scots pine needles were studied in a long-term acid rain experiment in the far north of Finnish Lapland. Pine trees 5 m tall of age 50-70 years were exposed, by spraying the foliage and soil from a height of 2 m, to either clean water (IC) or acidified water over the period 1985-1992, the acidification site being divided into sub-areas in which the precipitation contained two levels of either sulphuric (Sm, Sh) or nitric (Nm, Nh) acid, or both (SNm, SNh). The treatments with medium and high sulphate-S over eight consecutive years yielded a total sulphur deposition of 3.4 and 17.1 g m-2, respectively, and those with medium and high nitrate-N a total nitrogen deposition of 11 and 59 g m-2. Needles were collected for light and electron microscopy, growth measurements and morphometry. Growth in branch height had decreased by about 40% after 6 years of SNm or SNh treatment, and needle growth by 15% in the SNh trees as compared with the irrigated control trees (IC), although decreases were statistically significant only with respect to the non-irrigated control trees (DC). Growth of branches and needles was slightly better in the Nh treatment than in the IC group. The areas of the whole needle, the mesophyll and the phloem decreased in response to SNh treatment as compared with IC or DC, and a statistically significant decrease of about 30-40% was seen in the area of the xylem in comparison with DC. Cellular damage was observed following the acid treatments, especially with a high acid load. The damage was manifested in collapse of the cellular compartments, increases in lipid accumulations and swelling or disorganization of the protoplast. Increased vacuolization of the cytoplasm, plasmalemma irregularities and chilling-type damage to the mitochondria were also observed. [ABSTRACT FROM AUTHOR]