Pluripotent stem cells--model of embryonic development,tool for gene targeting,and basis of cell therapy

Embryonic stem (ES) cells are pluripotent cell lines with the capacity of self-renewal and a broad differentiation plasticity. They are derived from pre-implantation embryos and can be propagated as a homogeneous, uncommitted cell population for an almost unlimited period of time without losing their pluripotency and their stable karyotype. Murine ES cells are able to reintegrate fully into embryogenesis when returned into an early embryo, even after extensive genetic manipulation. In the resulting chimeric offspring produced by blastocyst injection or morula aggregation, ES cell descendants are represented among all cell types, including functional gametes. Therefore, mouse ES cells represent an important tool for genetic engineering, in particular via homologous recombination, to introduce gene knock-outs and other precise genomic modifications into the mouse germ line. Because of these properties ES cell technology is of high interest for other model organisms and for livestock species like cattle and pigs. However, in spite of tremendous research activities, no proven ES cells colonizing the germ line have yet been established for vertebrate species other than the mouse (Evans and Kaufman, 1981; Martin, 1981) and chicken (Pain et al., 1996). The in vitro differentiation capacity of ES cells provides unique opportunities for experimental analysis of gene regulation and function during cell commitment and differentiation in early embryogenesis. Recently, pluripotent stem cells were established from human embryos (Thomson et al., 1998) and early fetuses (Shamblott et al., 1998), opening new scenarios both for research in human developmental biology and for medical applications, i.e. cell replacement strategies. At about the same time, research activities focused on characteristics and differentiation potential of somatic stem cells, unravelling an unexpected plasticity of these cell types. Somatic stem cells are found in differentiated tissues and can renew themselves in addition to generating the specialized cell types of the tissue from which they originate. Additional to discoveries of somatic stem cells in tissues that were previously not thought to contain these kinds of cells, they also appear to be capable of developing into cell types of other tissues, but have a reduced differentiation potential as compared to embryo-derived stem cells. Therefore, somatic stem cells are referred to as multipotent rather than pluripotent. This review summarizes characteristics of pluripotent stem cells in the mouse and in selected livestock species, explains their use for genetic engineering and basic research on embryonic development, and evaluates their potential for cell therapy as compared to somatic stem cells.     

DNA sequences coding for the F18 fimbriae and AIDA adhesin are localised on the same plasmid in Escherichia coli isolates from piglets

The adhesin-involved-in-diffuse-adherence (AIDA) afimbrial adhesin is produced by human, but not by animal, Escherichia coli, with the exception of German porcine verotoxigenic Escherichia coli (VTEC) [Clin. Diagn. Lab. Immunol. 8 (2001) 143]. Presence and localisation of DNA sequences (aidA) coding for and production of an AIDA adhesin were investigated in a collection of Belgian VTEC and non-VTEC E. coli isolated from piglets at weaning time. The 174 isolates were also studied by colony hybridisation for the presence of DNA sequences coding for the Stx2e verocytotoxin and the F18 fimbrial adhesin (fed): 71 were Stx2+F18+AIDA+, 26 were F18+AIDA+, 12 were AIDA+, two were Stx2+AIDA+, and one was Stx2+ only. Fifty-four of the 58 (F18+)AIDA+ isolates tested positive in a western blotting assay with an immune serum raised against the AIDA protein. Hybridisation with the AIDA gene probe on plasmid DNA profiles identified a probe-positive plasmid band in the 10 AIDA+ and in 24 of the 25 F18+AIDA+ isolates studied. Moreover in F18+AIDA+ isolates, only one plasmid band hybridised with both F18 and AIDA probes. These results confirm the presence of aidA-related genes in not only VTEC, but also non-VTEC, isolates from piglets and the production of an antigenically AIDA-related protein by the majority of probe-positive E. coli. Moreover the plasmid DNA hybridisation results suggest a localisation on the same plasmid of the aidA- and fed-related DNA sequences.     

Clinical and pathological effects of the dermonecrotic toxin of Bordetella bronchiseptica and Pasteurella multocida in specific-pathogen-free piglets

The role of dermonecrotic toxin (DNT) of Bordetella bronchiseptica and Pasteurella multocida, purified by repeated chromatography in Sephacryl S-200 gel, in the pathogenesis of atrophic rhinitis (AR) of swine was studied bacteriologically, clinically and pathologically. Two-week-old specific pathogen-free (SPF) piglets were parenterally treated with 30 micrograms of DNT 3 times at 2-day interval and 7-week-old piglets were treated with 15 micrograms of DNT twice a week for 5 weeks. In 2- to 3-week-old piglets, both B. bronchiseptica DNT and P. multocida DNT produced nasal turbinate lesions with similar severity, characterized by damage of the cilia, epithelial metaplasia, intensive proliferation of osteoblasts, regressive changes, and diffuse osteocytic osteolysis. In 7- to 12-week-old piglets, treatment with B. bronchiseptica DNT failed to produce progressive changes in the nasal turbinates. Histopathological examination revealed osteogenic processes and osteoid synthesis besides the proliferation of osteoblasts and mild osteocytic osteolysis. Moreover, severe gross pathological lesions developed in the stomach, liver, kidneys, and lymphoid organs. The piglets' appetite and body weight gain gradually decreased during the DNT treatment and in the last week when the toxic signs appeared. Treatment of 7- to 12-week-old piglets with P. multocida DNT resulted in progressive AR. Histopathologically, diffuse osteocytic osteolysis was observed in the nasal turbinates. Neither clinical signs nor pathological lesions of the visceral organs developed in these piglets. The authors emphasize that the DNT of B. bronchiseptica basically differs from that of P. multocida in biological properties, though there are certain similarities between the DNTs.     

Flavonoids of zoysiagrass (Zoysia spp.) cultivars varying in fall armyworm (Spodoptera frugiperda) resistance

Flavonoid profiles of 12 zoysiagrass (Zoysia spp.) cultivars sampled six times in 1998 were correlated to fall armyworm (Spodoptera frugiperda J. E. Smith) larval weights and survival on replicated field-grown plant material and analyzed to determine genetic and seasonal variations of flavonoids among zoysiagrass cultivars. From multiple regression analyses and correlations, flavonoid peak 10 (luteolin-glucoside) had the greatest positive association with average fall armyworm weight; however, resistance appeared to be correlated with a number of other flavonoids. The flavonoid profiles of cultivars subjected to clustering procedures showed consistent genetic variability for five of six samplings and was used to genotype 23 cultivars. The dendrogram supported the results of the FASTCLUS procedure in clustering certain genotypes such as fall armyworm-resistant Cavalier and Zeon together, as well as J-36 and Meyer. Flavonoid evaluations measure genetic relatedness among cultivars and could be used for selective breeding of resistance to fall armyworm.     

Conventional analyses of data from dietary validation studies may misestimate reporting accuracy: illustration from a study of the effect of interview modality on children's reporting accuracy

OBJECTIVE: To compare two approaches to analysing energy- and nutrient-converted data from dietary validation (and relative validation) studies - conventional analyses, in which the accuracy of reported items is not ascertained, and reporting-error-sensitive analyses, in which reported items are classified as matches (items actually eaten) or intrusions (items not actually eaten), and reported amounts are classified as corresponding or overreported. DESIGN: Subjects were observed eating school breakfast and lunch, and interviewed that evening about that day's intake. For conventional analyses, reference and reported information were converted to energy and macronutrients; then t-tests, correlation coefficients and report rates (reported/reference) were calculated. For reporting error-sensitive analyses, reported items were classified as matches or intrusions, reported amounts were classified as corresponding or overreported, and correspondence rates (corresponding amount/reference amount) and inflation ratios (overreported amount/reference amount) were calculated. SUBJECTS: Sixty-nine fourth-grade children (35 girls) from 10 elementary schools in Georgia (USA). RESULTS: For energy and each macronutrient, conventional analyses found that reported amounts were significantly less than reference amounts (every P < 0.021; paired t-tests); correlations between reported and reference amounts exceeded 0.52 (every P < 0.001); and median report rates ranged from 76% to 95%. Analyses sensitive to reporting errors found median correspondence rates between 67% and 79%, and that median inflation ratios, which ranged from 7% to 17%, differed significantly from 0 (every P < 0.0001; sign tests). CONCLUSIONS: Conventional analyses of energy and nutrient data from dietary reporting validation (and relative validation) studies may overestimate accuracy and mask the complexity of dietary reporting error.     

Effect of salinity on natural community and production of Litopenaeus vannamei (Boone), within experimental zero-water exchange culture systems

Recent efforts have been made to culture marine shrimp in systems operating under low or zero‐water exchange and with decreased water salinity. The aim of this study was to investigate the impact of various salinity levels on qualitative and quantitative characteristics of the natural community and, more particularly, ciliated protozoa, and compare this information with shrimp growth and survival. Tanks with 9‰ salinity were characterized by a higher pH, but also by a significantly higher concentration of chlorophyll a (Chl a) per weight of suspended matter (1.93 ± 0.72 µg Chl a/mg TSS) than tanks with 18‰ (1.29 ± 0.68 µg Chl a/mg TSS) or 36‰ (1.37 ± 0.61 µg Chl a/mg TSS) salinity. Concentrations of ciliates (max 6000 cells mL^?1) showed considerable fluctuations over the sampling period, reflecting the impact of water salinity, dynamic interactions between ciliates and their diverse roles within the shrimp production system. There was no significant difference between survival rates of shrimp reared at 9‰, 18‰ or 36‰, but decreasing salinity from 36‰ to 9‰ led to a significant decrease in final shrimp body weight (from 13.40 ± 0.26 g to 10.23 ± 2.72 g). Future work should address the potential of ciliates as an indicator of aquaculture water quality, as is currently being done in the wastewater industry, and the contribution of ciliates as food sources.     

Biomass equations for seven different tree species growing in coppice-with-standards forests in Central Germany

With an increasing demand of sustainable raw materials for bioenergy use, coppicing as management approach to increase the biomass production of forests is becoming of greater importance. This study describes the parameterization of biomass equations for six tree species traditionally used in coppices forests, namely sycamore maple (Acer pseudoplatanus L.), field maple (Acer campestre L.), European ash (Fraxinus excelsior L.), European hornbeam (Carpinus betulus L.), downy birch (Betula pubescens Ehrh.), and common hazel (Corylus avellana L.) growing in coppice-with-standard systems in Lower Saxony, Germany. The parameterization was based on measurements of over 950 trees sampled from two forest sites. The sampled trees were felled and separated into three biomass compartments (stem, coarse branches, and fine brushwood) and weighed on site. The dry weight of sub samples from each compartment was measured. Equations were derived for total aboveground biomass, stem biomass, and crown biomass using regression analyses. We either used diameter at breast height as single independent explanatory variable or in combination with tree height. Biomass production of stump sprouts and generatively grown stems was compared for ash and sycamore maple. In the same age classes, it was found that ash stump sprouts had a slightly higher production than seed-grown stems. For sycamore maple, no difference was detected.     

Screening for low grain cadmium phenotypes in sunflower, durum wheat and flax

Cadmium (Cd) level in nonoilseed sunflower (Helianthus annuus L.), flax (Linum usitatissimum L.), and durum wheat (Triticum turgidum L. var. durum) grown on uncontaminated, alkaline soils has exceeded limits established in Northern Europe. Separate field experiments were conducted to investigate variability of grain Cd levels among sunflower, durum wheat and flax germplasm, and to seek an efficient screening method for future breeding. There were large variations in leaf Cd concentration among 200 sunflower lines. These lines performed more consistently for Cd uptake at the R5 stage than at the V8 stage across 4 locations with markedly differing soils. Cd concentration in V8 leaves was not related to Cd in grain. The positive correlation between R5 leaf Cd and kernel Cd level was obtained from nonoilseed hybrid (Sigco 954) (R²; = 0.74**), and 200 lines (R² = 0.44**) tested over 4 locations in 2 field trials, respectively. This indicates that an efficient and low cost screening method can be developed for genotype selection, but plants must be grown to the R5 stage. A preliminary evaluation of 30 durum wheat and 74 flax lines indicated large variations in grain Cd level of durum wheat and flax. Grain Cd concentration ranged from 0.11 to 0.34 mg Cd kg^-1 DW for durum wheat, and 0.14 to 1.37 mg Cd kg^-1 DW for flax, respectively. This variability indicates that breeding for low grain Cd in durum wheat and flax should be feasible. This revised version was published online in July 2006 with corrections to the Cover Date.