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Biomedical subjects

B L Nielsen

Publications and source records attributed to B L Nielsen.

At least 19 recordsLinked to original sources

Why women want prenatal ultrasound in normal pregnancy.

OBJECTIVES: To investigate women's reasons for requesting prenatal ultrasound in the absence of clinical indications. METHODS: A postal questionnaire was completed by 370 pregnant women with no apparent obstetric risk factors, who had expressed a desire to have ultrasound scanning in their current pregnancy. The women were asked to indicate, from a list of 12 items, their three most important reasons for wanting scanning. Ninety per cent of the women were in the first trimester of pregnancy, and 10% in the second trimester. RESULTS: The items most frequently identified as important reasons for ultrasound were to check for fetal abnormalities (60% of women), to see that all was normal (55%) and for own reassurance (44%). Lower income was related to wanting to see the baby (P = 0.028) and wanting an ultrasound picture (P = 0.017); higher income was related to checking that all was normal (P = 0.003) and for own reassurance (P = 0.015). Women in their first pregnancy were more likely to want themselves and the father to see the baby (P = 0.001); women who had given birth previously were more likely to want reassurance (P = 0.002), as were women with a previous miscarriage or induced abortion. Women who believed that the presence of fetal trisomy justifies abortion or who would vote for free abortion were more likely to want to know about abnormalities (P < 0.001 and P < 0.004, respectively). Women in the second trimester were more likely to want to check for abnormalities (P = 0.041) and appropriate fetal growth (P = 0.047) than those in the first trimester. CONCLUSIONS: It would appear that women in normal pregnancy have specific reasons for wanting prenatal ultrasound that are influenced by sociodemographic, obstetric and attitudinal factors.

Adult↗

Foot pad dermatitis and hock burn in broiler chickens and degree of inheritance.

A total of 2,118 birds from 2 strains were allocated to 12 groups of 93 to 100 each in 2 time-separated replicates. The development of foot pad dermatitis (FPD) and hock burn (HB) were recorded weekly from d 8 to slaughter on a set sample of live animals (7 per group). In addition, feet and hocks of all birds were investigated at slaughter at either 4, 6 (fast-growing strain), 8, or 10 (slow-growing strain) wk of age. Lesions were scored for both the left and right foot and classified according to a scale from 1 (no lesion) to 9 (very severe lesions) for FPD and from 1 (no lesion) to 3 (very severe lesions) for HB. No FPD lesions and very few low-grade HB lesions were found in chickens from the slow-growing strain. In the fast-growing strain, the first signs of FPD and HB were seen in wk 2. The incidence of both types of lesions increased thereafter. Foot pad dermatitis was more frequent in females (49 vs. 36%, P < 0.05). Body weight did not affect FPD, but more HB were found at higher BW (P < 0.01). Egg weight influenced neither FPD nor HB. Variance and covariance components were analyzed using a multivariate animal model, in which scores for FPD and HB were transformed into logarithmic scale. The analyses were carried out using restricted maximum likelihood algorithm. Heritabilities were estimated to be 0.31 +/- 0.12 (SE) for FPD, 0.08 +/- 0.08 for HB, and 0.38 +/- 0.13 for BW. Genetic correlations among these traits were low and nonsignificant. Phenotypic correlation between BW and FPD was low and nonsignificant and between BW and HB was 0.17 +/- 0.05 (P < 0.01). The relative high heritability of FPD and the low genetic correlation to BW suggested that genetic selection against susceptibility to FPD should be possible without negative effects on BW gain.

Analysis of Variance↗

Breast blisters in groups of slow-growing broilers in relation to strain and the availability and use of perches.

1. The relationship between perching behaviour, availability of perches and the incidence and severity of breast blisters in broilers was investigated together with a comparison between two slow-growing broiler strains. 2. Sixteen single strain groups (n = 60) of Labresse and i657 broilers were subjected throughout the experiment to one of three perch availabilities: 15 cm per bird (Labresse and i657), 7.5 cm per bird (i657 only), and 0 cm per bird (i657 only) with 4 replicates per treatment. The birds were housed indoors from one day old, and at 43 d of age 52 birds from each group were moved to outdoor housing facilities with access to grass-covered outdoor areas until slaughter at 84 d of age. 3. The use of perches was monitored via video recordings throughout the experimental period. The severity of breast blisters was recorded on a scale from 0 to 2 at slaughter. 4. Groups of i657 with 15 cm perch per bird used these more than groups with 7.5 cm perch per bird (19% vs 8% of birds perching at midnight). A positive association between access to perches and severe breast blisters (score 2) was found in the groups of i657, with odds ratios of 3.1 and 3.4 for 7.5 and 15.0 cm per bird, respectively, relative to the no-perch treatment. 5. Labresse were more likely to develop breast blisters than i657 (odds ratio 3.5), but used the perches less (0.1 vs 9.5 birds perching at midnight) and weighed less (2011 g vs 2246 g) than i657. 6. Males had a higher incidence of breast blisters than females (odds ratio 12.2), and this was most prominent in the Labresse strain (odds ratio 40.0). 7. In some broiler strains access to perches may be associated with an increase in the occurrence of severe breast blisters, but strain and sex of broiler chickens appear to have a much larger influence than access to perches on the incidence of breast blisters.

Animals↗

An Arabidopsis homologue of bacterial RecA that complements an E. coli recA deletion is targeted to plant mitochondria.

Homologous recombination results in the exchange and rearrangement of DNA, and thus generates genetic variation in living organisms. RecA is known to function in all bacteria as the central enzyme catalyzing strand transfer and has functional homologues in eukaryotes. Most of our knowledge of homologous recombination in eukaryotes is limited to processes in the nucleus. The mitochondrial genomes of higher plants contain repeated sequences that are known to undergo frequent rearrangements and recombination events. However, very little is known about the proteins involved or the biochemical mechanisms of DNA recombination in plant mitochondria. We provide here the first report of an Arabidopsis thaliana homologue of Escherichia coli RecA that is targeted to mitochondria. The mt recA gene has a putative mitochondrial presequence identified from the A. thaliana genome database. This nuclear gene encodes a predicted product that shows highest sequence homology to chloroplast RecA and RecA proteins from proteobacteria. When fused to the GFP coding sequence, the predicted presequence was able to target the fusion protein to isolated mitochondria but not to chloroplasts. The mitochondrion-specific localization of the mt recA gene product was confirmed by Western analysis using polyclonal antibodies raised against a synthetic peptide from a unique region of the mature mtRecA. The Arabidopsis mt recA gene partially complemented a recA deletion in E. coli, enhancing survival after exposure to DNA-damaging agents. These results suggest a possible role for mt recA in homologous recombination and/or repair in Arabidopsis mitochondria.

Amino Acid Sequence↗

Feed and strain effects on the use of outdoor areas by broilers.

1. The use of outdoor areas by two broiler strains, known to have different growth rates and with access to either moderate (M) or low (L) energy feeds, was examined to assess the consequences for production, meat quality, health and welfare. 2. Sixteen groups of either Ross 208 (ROSS) or a Labresse cross (LAB) were fed either feed M or L. They were raised indoors until 42 d of age when groups of 102 birds were moved to outdoor houses with free access to an outdoor area. 3. The number of chickens standing and lying, respectively, in each of 5 zones in the outdoor area were recorded at intervals throughout the outdoor growth period until slaughter at 84 d of age. Production variables were measured, and gait, feather condition, litter quality, and dermal lesions on foot-pads and hocks were scored together with objective meat quality measures. 4. Both strains showed a diurnal rhythm with most birds observed outside around sunrise and before sunset. Feed and strain both affected the distribution of the birds on the outdoor area, with more birds on feed M than L observed outside, and with more LAB birds outside and using more of the outdoor area than ROSS. This was reflected in some of the carcase and meat quality measures. Feather pecking and cannibalism only occurred in LAB. 5. ROSS was found to have a faster growth rate, poorer litter quality, more dermal lesions and impaired mobility, reflected in low usage of outdoor area and poor gait score compared with LAB. 6. This together with the presence of pectoral myopathies make ROSS unsuitable for 12 week growth in free range production systems. The occurrence of feather pecking and cannibalism in LAB make this particular Labresse cross questionable for meat-type poultry production.

Aggression↗

Staying good while playing god--the ethics of breeding farm animals.

Modern genetics has given us some very efficient tools with which to alter the characteristics of animals. To date, farm animal breeders have mainly used these tools to increase productivity. Thus, each new generation of farm animals matures faster, yields more milk, or produces more meat or eggs, than the previous one. Despite these apparent benefits, modern farm animal breeding has had severe negative consequences, including effects on the quality of the animals' lives and biodiversity. The aim of this paper is to discuss the goals and consequences of farm animal breeding within an ethical context. First, a description of what has happened to broilers and dairy cattle as breeders have pursued the goal of ever more efficient production is given. Second, the ethical values that ought to underlie future breeding schemes are discussed. It is suggested that there are in fact two very different ethical approaches: the 'quality of life-based' approach and the 'preservationist' approach. A view combining elements from both approaches is advanced. Finally, an example is given of how it is possible, in practice, to pursue an ethically defensible breeding goal without compromising production efficiency.

Animal Welfare↗

In vitro replication of mitochondrial plasmid mp1 from the higher plant Chenopodium album (L.): a remnant of bacterial rolling circle and conjugative plasmids?

According to the endosymbiotic theory, mitochondrial genomes evolved from the chromosome of an alpha-proteobacterium-like ancestor and developed during evolution an extraordinary variation in size, structure and replication. We studied in vitro DNA replication of the mitochondrial circular plasmid mp1 (1309 bp) from the higher plant Chenopodium album (L.) as a model system that replicates in a manner reminiscent of bacterial rolling circle plasmids. Several mp1 subclones were tested for their ability to support DNA replication using a newly developed in vitro system. Neutral/neutral two-dimensional gel electrophoresis of the in vitro products revealed typical simple Y patterns of intermediates consistent with a rolling circle type of replication. Replication activity was very high for a BamHI-restricted total plasmid DNA clone, a 464 bp BamHI/KpnI fragment and a 363 bp BamHI/SmaI fragment. Further subcloning of a 148 bp BamHI/EcoRI fragment resulted in the strongest in vitro DNA replication activity, while a 1161 bp-template outside of this region resulted in a substantial loss of activity. Electron microscopic studies of in vitro DNA replication products from the highly active clones also revealed sigma-shaped molecules. These results support our in vivo data for the presence of a predominant replication origin between positions 628 and 776 on the plasmid map. This sequence shares homology with double-stranded rolling circle origin (dso) or transfer origin (oriT) nicking motifs from bacterial plasmids. mp1 is the first described rolling circle plasmid in eukaryotes.

Base Sequence↗

Leucine-based receptor sorting motifs are dependent on the spacing relative to the plasma membrane.

Many integral membrane proteins contain leucine-based motifs within their cytoplasmic domains that mediate internalization and intracellular sorting. Two types of leucine-based motifs have been identified. One type is dependent on phosphorylation, whereas the other type, which includes an acidic amino acid, is constitutively active. In this study, we have investigated how the spacing relative to the plasma membrane affects the function of both types of leucine-based motifs. For phosphorylation-dependent leucine-based motifs, a minimal spacing of 7 residues between the plasma membrane and the phospho-acceptor was required for phosphorylation and thereby activation of the motifs. For constitutively active leucine-based motifs, a minimal spacing of 6 residues between the plasma membrane and the acidic residue was required for optimal activity of the motifs. In addition, we found that the acidic residue of leucine-based motifs must be located amino-terminal to the dileucine sequence for proper function of the motifs and that residues surrounding the motifs affect the activity of the motifs. Thus, our observations suggest that the position, the exact sequence, and surrounding residues are major determinants of the function of leucine-based receptor sorting motifs.

Amino Acid Sequence↗

Investigation of the relationship between farrowing environment, sex steroid concentrations and maternal aggression in gilts.

Maternal oestrogen and progesterone have been shown to be important in the initiation of maternal behaviour. Thirty-three Large White x Landrace gilts, housed in groups during pregnancy, were observed and aggressive interactions recorded. Individuals had jugular catheters implanted 14.5 (s.e. 0.34) days before their expected parturition date (EPD). Five days before EPD gilts were randomly allocated and moved to either a conventional farrowing crate (C; without straw, 16 gilts) or a pen (P; 2.1 x 3.1 m2; with straw bedding, 17 gilts). Blood samples were taken at frequencies determined by the proximity to farrowing onset. Piglets were removed at birth and returned 2 h after placental expulsion. The reaction of each gilt to her piglets was monitored. Gilts savaging piglets were sedated with azaperone (n = 8). There was no overall effect of farrowing environment on oestradiol and progesterone concentrations. The pre-farrowing ratio of progesterone to oestradiol was higher for (P) gilts (0.45 vs. 0.25, (P) vs. (C); S.E.D. 0.085, P < 0.05) as was their overall maximum oestradiol level (3.39 vs. 2.29 ng/ml, (P) vs. (C); S.E.D. 0.39, P < 0.01). In contrast to progesterone, oestradiol patterns varied considerably between individuals. Dominance rank value during pregnancy, but not levels of aggression, correlated positively to pre-farrowing oestradiol concentrations. Treatment with azaperone was not related to farrowing environment, piglet weight or litter size. Azaperone treated gilts showed a higher pre-farrowing oestradiol to progesterone ratio (0.55 vs. 0.29, +/- azaperone; S.E.D. 0.10, P < 0.05), significantly higher levels of oestradiol post-partum (0.7 vs. 0.19 ng/ml, +/- azaperone; S.E.D. 0.20, P < 0.001) and significantly lower levels of aggression during pregnancy (1.68 vs. 2.23 aggressive interactions/h, +/- azaperone; S.E.D. 0.15, P < 0.001). The results indicate that there are no major effects of farrowing environment on sex steroid concentrations. Maternal aggression under these conditions appears to be negatively related to aggression during pregnancy, but this is not reflected in plasma concentrations of sex steroids around parturition.

Aggression↗

Effects of feed composition and stage of lactation on the short-term feeding behavior of dairy cows.

Twenty Holstein-Friesian cows were assigned to one of four feeding groups throughout lactation in a full change-over experiment using two total mixed diets. The low concentrate total mixed diet contained 100 g of concentrate/kg of fresh matter, and the high concentrate total mixed diet contained 300 g of concentrate/kg of fresh matter. The remainder of the total mixed diet was grass silage. The two changeover groups switched total mixed diets at 153 d of lactation; the other two treatment groups remained on their assigned diets throughout lactation. For analysis of short-term feeding behavior, four periods of 3 wk each were identified. The midpoints of these periods were -102, -18, 18, and 102 d from the changeover. The concentrate content of the total mixed diet significantly affected dry matter intake and all short-term feeding behavior variables. Cows that consumed the high concentrate total mixed diet had fewer but longer visits to the feeders and ate more feed per visit than did cows consuming the low concentrate total mixed diet. With one exception, no significant effect of stage of lactation was detected for any of the short-term feeding behavior variables. Despite a highly significant decline in dry matter intake as lactation progressed for cows consuming the high concentrate total mixed diet, there were no interactions between total mixed diet and stage of lactation for any of the short-term feeding behavior variables. Large differences in feeding behavior were detected between cows consuming the same total mixed diet. These last two findings suggest that the use of short-term feeding behavior variables to predict daily intake is unlikely to be successful.

Animal Feed↗

Fine mapping of replication origins (ori A and ori B) in Nicotiana tabacum chloroplast DNA.

Using a partially purified replication complex from tobacco chloroplasts, replication origins have been localized to minimal sequences of 82 (pKN8, positions 137 683-137 764) and 243 bp (pKN3, positions 130 513-130 755) for ori A and ori B respectively. Analysis of in vitro replication products by two-dimensional agarose gel electrophoresis showed simple Y patterns for single ori sequence-containing clones, indicative of rolling circle replication. Double Y patterns were observed when a chloroplast DNA template containing both ori s (pKN9) was tested. Dpn I analysis and control assays with Escherichia coli DNA polymerase provide a clear method to distinguish between true replication and DNA repair synthesis. These controls also support the reliability of this in vitro chloroplast DNA replication system. EM analysis of in vitro replicated products showed rolling circle replication intermediates for single ori clones (ori A or ori B), whereas D loops were observed for a clone (pKN9) containing both ori s. The minimal ori regions contain sequences which are capable of forming stem-loop structures with relatively high free energy and other sequences which interact with specific protein(s) from the chloroplast replication fraction. Apparently the minimal ori sequences reported here contain all the necessary elements for support of chloroplast DNA replication in vitro.

Base Sequence↗

Regulation and function of the CD3gamma DxxxLL motif: a binding site for adaptor protein-1 and adaptor protein-2 in vitro.

Several receptors are downregulated by internalization after ligand binding. Regulation of T cell receptor (TCR) expression is an important step in T cell activation, desensitization, and tolerance induction. One way T cells regulate TCR expression is by phosphorylation/dephosphorylation of the TCR subunit clusters of differentiation (CD)3gamma. Thus, phosphorylation of CD3gamma serine 126 (S126) causes a downregulation of the TCR. In this study, we have analyzed the CD3gamma internalization motif in three different systems in parallel: in the context of the complete multimeric TCR; in monomeric CD4/CD3gamma chimeras; and in vitro by binding CD3gamma peptides to clathrin-coated vesicle adaptor proteins (APs). We find that the CD3gamma D127xxxLL131/132 sequence represents one united motif for binding of both AP-1 and AP-2, and that this motif functions as an active sorting motif in monomeric CD4/ CD3gamma molecules independently of S126. An acidic amino acid is required at position 127 and a leucine (L) is required at position 131, whereas the requirements for position 132 are more relaxed. The spacing between aspartic acid 127 (D127) and L131 is crucial for the function of the motif in vivo and for AP binding in vitro. Furthermore, we provide evidence indicating that phosphorylation of CD3gamma S126 in the context of the complete TCR induces a conformational change that exposes the DxxxLL sequence for AP binding. Exposure of the DxxxLL motif causes an increase in the TCR internalization rate and we demonstrate that this leads to an impairment of TCR signaling. On the basis of the present results, we propose the existence of at least three different types of L-based receptor sorting motifs.

Adaptor Protein Complex alpha Subunits↗

Analysis of the tobacco chloroplast DNA replication origin (oriB) downstream of the 23 S rRNA gene.

We have mapped the origin of DNA replication (oriB) downstream of the 23 S rRNA gene in each copy of the inverted repeat (IR) of tobacco chloroplast DNA between positions 130,502 and 131,924 (IR(A)) by a combination of approaches. In vivo chloroplast DNA replication intermediates were examined by two-dimensional agarose gel electrophoresis. Extended arc patterns suggestive of replication intermediates containing extended single-stranded regions were observed with the 4.29 kb SspI fragment and an overlapping EcoRI fragment from one end of the inverted repeat, while only simple Y patterns were observed with a 3.92 kb BamHI-KpnI fragment internal to the SspI fragment. Other restriction fragments of tobacco chloroplast DNA besides those at the oriA region also generated only simple Y patterns in two-dimensional agarose gels. Several chloroplast DNA clones from this region were tested for their ability to support in vitro DNA replication using a partially purified chloroplast protein fraction. Templates with a deletion of 154 bp from the SspI to the BamHI sites near the end of the inverted repeat resulted in a considerable loss of in vitro DNA replication activity. These results support the presence of a replication origin at the end of the inverted repeat. The 5' end of nascent DNA from the replication displacement loop was identified at position 130,697 for IR(A) (111,832 for IR(B)) by primer extension. A single major product insensitive to alkali and RNase treatment was observed and mapped to the base of a stem-loop structure which contains one of two neighboring BamHI sites near the end of each inverted repeat. This provides the first precise determination of the start site of DNA synthesis from oriB. Adjacent DNA fragments containing the stem-loop structure and the 5' region exhibit sequence-specific gel mobility shift activity when incubated with the replication protein fraction, suggesting the presence of multiple binding sites.

Base Sequence↗

A 7.5-kbp region of the maize (T cytoplasm) mitochondrial genome contains a chloroplast-like trnI (CAT) pseudo gene and many short segments homologous to chloroplast and other known genes.

The DNA sequence of a 7.5-kbp region from the maize cmsT mt genome was determined. Nucleotide-sequence analysis of this region reveals the presence of a ct-like trnI (CAT) gene, orf31 and orf48 which are respectively homologous to the trnI (CAT) gene in the maize and rice chloroplast genomes, and to orf28 and orf64 in the rice chloroplast genome. Northern-blot analysis indicates that the ct-like trnI (CAT) gene is likely to be a pseudo gene in the maize cmsT mt genome because it is not transcribed. A nucleotide-sequence homology search of this 7.5-kbp region reveals several short segments homologous to portions of chloroplast (ct), mitochondrial (mt) and other known genes. These segments range from 17 bp to 187 bp in length with homology from 71 to 100%. These observations also suggest that the transfer of DNA fragments from the ct genome to the mt genome may have occurred at different times during the evolution of the maize mt genome, and that multiple recombination events and rearrangements in both mt and ct genomes have occurred after the transfer of DNA fragments from the ct to the mt genome. A few segments were identified by their homology to be portions of genes from sources other than the ct genome. While it is known that large fragments of ctDNA have been transferred to maize mt genomes, this is the first report of the presence of numerous short ctDNA, or other foreign DNA segments, in the maize cmsT mt genome. These segments may provide valuable information regarding the evolution of plant mt genomes.

Base Sequence↗

Molecular characterization of avian reoviruses using nested PCR and nucleotide sequence analysis.

A nested polymerase chain reaction (PCR) with subsequent nucleotide sequence analysis identified and differentiated avian reoviruses (ARVs). PCR products amplified from the S1 gene segment of ARV of USA isolates were 738 and 342 bp, respectively. PCR products were conformed by Southern and dot blot hybridizations. The amplified cDNA fragments were cloned into the pUC18 vector and subjected to DNA sequencing. The nucleotide and deduced amino acid sequences of four USA (S1133, 1733, 2408, and CO8) and two Australian isolates (RAM-1 and SOM-4) were compared. Results of paired difference analysis and a predicted dendrogram revealed that USA isolates were closely related, but different from, Australian isolates. The deduced amino acid sequences of the N-terminal region of ARV sigma C showed a heptapeptide repeat of hydrophobic residues in all ARV isolates.

Amino Acid Sequence↗

Characterization of replication origins flanking the 23S rRNA gene in tobacco chloroplast DNA.

Using 5' end-labeled nascent strands of tobacco chloroplast DNA (ctDNA) as a probe, replication displacement loop (D-loop) regions were identified. The strongest hybridization was observed with restriction fragments containing the rRNA genes from the inverted repeat region. Two-dimensional gel analysis of various digests of tobacco ctDNA suggested that a replication origin is located near each end of the 7.1 kb BamHI fragment containing part of the rRNA operon. Analysis of in vitro replication products indicated that templates from either of the origin regions supported replication, while the vector alone or ctDNA clones from other regions of the genome did not support in vitro replication. Sequences from both sides of the BamHI site in the rRNA spacer region were required for optimal in vitro DNA replication activity. Primer extension was used for the first time to identify the start site of DNA synthesis for the D-loop in the rRNA spacer region. The major 5' end of the D-loop was localized to the base of a stem-loop structure which contains the rRNA spacer BamHI site. Primer extension products were insensitive to both alkali and RNase treatment, suggesting that RNA primers had already been removed from the 5' end of nascent DNA. Location of an origin in the rRNA spacer region of ctDNA from tobacco, pea and Oenothera suggests that ctDNA replication origins may be conserved in higher plants.

Base Composition↗

Isolation and characterization of an in vitro DNA replication system from maize mitochondria.

An in vitro DNA replication system from maize mitochondria has been isolated and characterized. Maize mtDNA polymerase activity was purified about 1100-fold through DEAE cellulose and Heparin-Sepharose columns. In addition to the DNA polymerase activity, this in vitro replication system also contained topoisomerase I, DNA primase and RNA polymerase activities. Optimal conditions for enzyme activity, preferred templates and inhibitors were determined in order to further characterize this in vitro replication system; this system was devoid of any detectable extramitochondrial activity as determined by: a) the mt origin of the DNA polymerase activity as evidenced by studies using different templates and inhibitors, b) absence of chloroplast or nuclear DNA, glucose -6-P-dehydrogenase (known to be present only in the cytosol and chloroplasts) and photosynthetic pigments in the mitochondrial fraction and c) the ability of maize mt topoisomerase I to relax positively supercoiled DNA.

Chromatography, Affinity↗