Cubicle Refusal and Rearing Accommodation as Possible Mastitis Risk Factors in Cubicle-Housed Dairy Heifers

Fifty-nine of the 65 dairy farms with cubicle sheds in the Norwegian county of Oppland in 1990 were included in a study of rearing accommodation, cubicle refusal and mastitis incidence. The farmers recorded the favoured resting location of the individual cows and heifers throughout the final week of pregnancy as well as during calving. The observations were matched with individual records of mastitis cases during the first 30 days after calving. Mastitis incidence in the heifers was analysed by logistic regression using rearing accommodation and cubicle refusal as independent variables, controlling for herd as a clustering factor. Cubicle refusal was found in 29% of the heifers, but in only 3% of older cows. The results of the analysis indicated a tendency for cubicle refusal to be associated with an increased mastitis incidence among the heifers (OR = 2.2, c.i._95%OR = 0.9–5.4, P = 0.08). Cubicle refusal accounted for 21% (0–32%) of the mastitis cases in the study population (PAF = 0.21).     

Use of a Fluorimetric Method to Assay the Binding and Transfer of Phospholipids by Lipid Transfer Proteins from Maize Seedlings and Arabidopsis Leaves

A fluorimetric method has been used to study the binding andtransfer of phospholipids mediated by a lipid transfer protein(LTP) from maize seedlings and by proteins extracted from Arabidopsisleaves. This method is based on the use of donor vesicles preparedby sonication or injection and consisting of either self quenchingvesicles of (1-palmitoyl 2-{12-{(7-nitrobenzoxadiazol-4-yl)amino}dodecanoyl}-Sn-glycero-3-phosphocholine(NBD-PC) or trinitrophenylphosphate phosphatidyl ethanolamine(TNP-PE) quenched vesicles of l-palmitoyl-2(l-pyredecanoyl)-Snglycero-3-phosphocholine(Pyr-PC). Acceptor vesicles consisted in a mixture (90/10; v/v)of dioleoylphosphatidylcholine and phosphatidyl inositol (DOPC-PI).The use of injected vesicles of Pyr-PC allowed to achieve sensitiveand qualitative tests of binding and transfer since 0.05 µMLTP were detected. By contrast, when sonicated vesicles of NBD-PCwere used, quantitative determinations of phospholipid-LTP complexas well as measurement of the initial rate of phospholipid transferon a large range of protein concentration (0.5 to 20 µM)were performed. This fluorimetric method has been successfullyused to study the activity of maize LTP during its purificationor the activity of LTPs present in Arabidopsis extracts. (Received December 5, 1993; Accepted December 7, 1993)     

通过接合作用质粒从大肠杆菌到棒状杆菌的转移

构建了可接合转移的穿梭质粒pXZ911、pBZ51和pBZ52。这些质粒中含有具转移功能的Mob片段和在棒状杆菌中复制的复制区。以大肠杆菌S17—1为供体菌,通过接合转移作用,可将这些质粒转移到谷氨酸棒杆菌ATCCl3032、谷氨酸棒杆菌ATCC21543、北京棒杆菌B3、北京棒杆菌1．299、裂氏棒杆菌B43、黄色短杆菌ATCC 14067等棒状杆菌苗株,接合转移频率分别为：9X10^-5,1X10^-4,8．5x10,2．3X10^-4×10^-5,2．9X10^-5。本文还探索了大肠杆菌和几种革兰氏阳性棒状杆菌问基因转移的方法、转移频率、影响转移频率的因素、宿主范围等问题。     

Density Transfer Studies of DNA Isolated from BACILLUS SUBTILIS after Exposure to Phenethyl Alcohol

The aim of this study was to determine whether there are specific weak points in the Bacillus subtilis chromosome and if so whether the replication point is the site of breakage. To answer these questions, B. subtilis chromosomes were partially labeled with 5-bromodeoxyuridine (5-BUdR). Sheared or unsheared preparations of partially labeled chromosomes which may or may not contain replication forks were analyzed for the distribution of genetic markers in a CsCl density gradient. Two sets of experiments based upon the density transfer experiments of Yoshikawa and Sueoka (1963) were performed: (1) experiments in which the origin of the chromosome was labeled and (2) experiments in which the terminus of the chromosome was labeled. In the first experiment, strain 23 (thy(-), his(-)) spores were germinated in the presence of 5-BUdR for various lengths of time and then transferred to fresh medium containing phenethyl alcohol (PEA) and thymidine (TdR). The DNA was isolated before and after transfer to PEA and TdR. In the second experiment strain 23 (thy(-), his(-)) spores were germinated in the presence of TdR and then PEA was added. After various lengths of time transfer was made to fresh medium containing PEA and 5-BUdR. The DNA was extracted by an extremely gentle technique to avoid breakage and centrifuged in a CsCl density gradient. PEA was added to the germinated spores to prevent dichotomous replication, but PEA did not prevent dichotomous replication in any of these experiments. This contradicts the conclusion of others that PEA prevents the chromosome from entering a new round of replication, but allows the chromosome to complete the round of replication already begun. The following observations offer support for the hypothesis that the replication point is a weak point in the chromosome: (1) when conditions were created to obtain partially labeled chromosomes with replication points: (a) labeled markers appeared at the hybrid density, (b) unlabeled markers appeared at the light density, (c) shearing of the DNA had little effect on the CsCl density gradient, except on a small proportion of labeled markers which had not appeared at the hybrid density prior to shearing; (2) when conditions were created to obtain partially labeled chromosomes with no replication points: (a) the majority of DNA molecules appeared at an intermediate density between the hybrid and the light densities, (b) the labeled and unlabeled markers appeared in the intermediate peak with approximately the same ratio as in the DNA preparations, (c) the labeled markers were found in the intermediate peak except where dichotomous replication had occurred, (d) after shearing, the labeled markers appeared at the hybrid density and the unlabeled markers appeared at the light density. Thus it is concluded that the replication point is a weak point in the B. subtilis chromosome where breakage easily occurs.     

通过不对称体细胞杂交向小麦转移青苗碱谷DNA

普通小麦"济南177"(Triticum aestivum L.cv.Jinan177)经继代培养和选择,形成了性质不同的愈伤组织,一种生长迅速,易于形成悬浮系,游离的原生质具有旺盛的分裂能力,但不能分化,称为Cha9;另一种具有一定能力,但游离原生质体分裂能力低,称为176.二者之一来源的原生质体与紫外线照射的青苗碱谷原生质体融合均不能获得再生植株.而将源于Cha9和176的两种原生质体混合,与经紫外线照射的青苗碱谷原生质体在PEG诱导下融合,融合产物再生了大量植株.再生的愈伤组织及植株经表现型、细胞学、有工酶、RAPD分析,证明了其杂种性质,用不麦叶绿体特异的简单得复序列(SSR)引物分析了再生杂种的叶绿体遗传组成情况.在不同的杂种克隆中,同时带有Cha9和176的遗传物质并含有供体核及胞质基因组的克隆4具有较高的分化能力,再生了大量生长旺盛的完整植株.     

通过不对称体细胞杂交向小麦转移青苗碱谷DNA(英文)

普通小麦“济南177”(Triticum aestzvum L.cv.Jinan 177)经继代培养和选择,形成了两种性质不同的愈伤组织,一种生长迅速,易于形成悬浮系,游离的原生质体具有旺盛的分裂能力,但不能分化,称为Cha 9;另一种具有一定分化能力,但游离的原生质体分裂能力低,称为176。二者之一来源的原生质体与紫外线照射的青苗碱谷原生质体融合均不能获得再生植株。而将来源于Cha 9和176的两种原生质体混合,与经紫外线照射的青苗碱谷原生质体在PEG诱导下融合,融合产物再生了大量植株。再生的愈伤组织及植株经表型、细胞学、同工酶、RAPD分析,证明了其杂种性质,用小麦叶绿体特异的简单重复序列(SSR)引物分析了再生杂种的叶绿体遗传组成情况。在不同的杂种克隆中,同时带有Cha 9和176的遗传物质并含有供体核及胞质基因组的克隆4具有较高的分化能力,再生了大量生长旺盛的完整植株。     

Isolating Contributions from Intersegmental Transfer to DNA Searching by Alkyladenine DNA Glycosylase

Large genomes pose a challenge to DNA repair pathways because rare sites of damage must be efficiently located from among a vast excess of undamaged sites. Human alkyladenine DNA glycosylase (AAG) employs nonspecific DNA binding interactions and facilitated diffusion to conduct a highly redundant search of adjacent sites. This ensures that every site is searched, but could be a detriment if the protein is trapped in a local segment of DNA. Intersegmental transfer between DNA segments that are transiently in close proximity provides an elegant solution that balances global and local searching processes. It has been difficult to detect intersegmental transfer experimentally; therefore, we developed biochemical assays that allowed us to observe and measure the rates of intersegmental transfer by AAG. AAG has a flexible amino terminus that tunes its affinity for nonspecific DNA, but we find that it is not required for intersegmental transfer. As AAG has only a single DNA binding site, this argues against the bridging model for intersegmental transfer. The rates of intersegmental transfer are strongly dependent on the salt concentration, supporting a jumping mechanism that involves microscopic dissociation and capture by a proximal DNA site. As many DNA-binding proteins have only a single binding site, jumping may be a common mechanism for intersegmental transfer.     

Characterization of the Core Rumen Microbiome in Cattle during Transition from Forage to Concentrate as Well as during and after an Acidotic Challenge

This study investigated the effect of diet and host on the rumen bacterial microbiome and the impact of an acidotic challenge on its composition. Using parallel pyrosequencing of the V3 hypervariable region of 16S rRNA gene, solid and liquid associated bacterial communities of 8 heifers were profiled. Heifers were exclusively fed forage, before being transitioned to a concentrate diet, subjected to an acidotic challenge and allowed to recover. Samples of rumen digesta were collected when heifers were fed forage, mixed forage, high grain, during challenge (4 h and 12 h) and recovery. A total of 560,994 high-quality bacterial sequences were obtained from the solid and liquid digesta. Using cluster analysis, prominent bacterial populations differed (P≤0.10) in solid and liquid fractions between forage and grain diets. Differences among hosts and diets were not revealed by DGGE, but real time qPCR showed that several bacteria taxon were impacted by changes in diet, with the exception of Streptococcus bovis. Analysis of the core rumen microbiome identified 32 OTU''s representing 10 distinct bacterial taxa including Bacteroidetes (32.8%), Firmicutes (43.2%) and Proteobacteria (14.3%). Diversity of OTUs was highest with forage with 38 unique OTUs identified as compared to only 11 with the high grain diet. Comparison of the microbial profiles of clincial vs. subclinical acidotic heifers found a increases in the relative abundances of Acetitomaculum, Lactobacillus, Prevotella, and Streptococcus. Increases in Streptococcus and Lactobacillus likely reflect the tolerance of these species to low pH and their ability to proliferate on surplus fermentable carbohydrate. The acetogen, Acetitomaculum may thereforeplay a role in the conversion of lactate to acetate in acidotic animals. Further profiling of the bacterial populations associated with subclinical and clinical acidosis could establish a microbial fingerprint for these disorders and provide insight into whether there are causative microbial populations that could potentially be therapeutically manipulated.     

Transfer of polygenic resistance to ryegrass mosaic virus from perennial to Italian ryegrass by backcrossing

Polygenic resistance to ryegrass mosaic virus (RMV) was transferred from perennial to Italian ryegrass cv. Tribune by repeated cycles of backcrossing, polycrossing and selection. The resulting synthetic variety (Bb 2113) was highly resistant to infection by RMV whereas the Italian ryegrass cultivars Tribune, RvP, Bartolini, Augusta, Wilo and Dalita all were highly susceptible. Leaf width and length of Bb 2113 was similar to that of cv. Tribune and, like Tribune, Bb 2113 had awned florets. In a field trial, Bb 2113 had a similar persistency rating and number of inflorescences in the regrowth to cv. Tribune but produced 11% less dry matter. The results emphasise the need for a more efficient means of transferring small numbers of genes from one species of Lolium to another than backcrossing provides.     

Bacterial Synthesis of Nucleosides by the Pentosyl Transfer Reaction from Nucleoside to Base

Synthesis of nucleosides by the pentosyl transfer reaction in Ps. trifolii (IAM-1555) was studied. The ribosyl transfer reaction between purine or pyrimidine bases and their nucleosides as an acceptor and a donor, respectively, was observed in the presence or absence of inorganic phosphate, and the participations of nucleoside phosphorylase in the former and nucleoside N-ribosyltransferase in the latter were suggested. This transribosylation to the base in the latter was observed to proceed stoichiometrically between pH 6 and 9.5, but the apparent optimal pH in the former was observed at around 10.5. Effects of cultural condition of bacterium, reaction temperature and metallic ions on this reaction and acceptor-and donor-specificities were studied in detail.     

Transfer of cytoplasm from newBeta CMS sources to sugar beet by asymmetric fusion

Summary For our program on the transfer of cytoplasmic male sterility (CMS) by cybridization inBeta vulgaris L. (sugar beet), we have developed a procedure for the isolation and culture of mesophyll protoplasts of sugar beet followed by shoot regeneration. A prerequisite proved to be the presence in the media of n-propylgallate (nPG), a lipoxygenase inhibitor. Sustained divisions were found in all accessions that were tested. Plating efficiencies and regeneration ability varied greatly from one experiment to the other and appeared to be accession-dependent. Shoots could be easily transferred to soil. A majority of the regenerants (72%) retained the diploid chromosome number. Somaclonar variation in phenotype was low (4.9%). Mitochondrial DNA probes, capable of discriminating different cytoplasms ofBeta spp. showed no rearrangements due to the protoplast and in vitro culture phase, indicating that these probes can be used to identify cybrids after asymmetric fusions. The data presented here open up possibilities for genetic engineering using protoplasts in one of the world's most important arable crops.     

Gene Transfer to Lentil Protoplasts by Lipofection and Electroporation

Abstract

Cationic liposomes made of dipalmitoylphosphatidylcholine and stearylamine (9:1) were prepared by reverse-phase evaporation and were able to interact spontaneously with plasmid DNA. The loaded vesicles delivered a β-glucuronidase (GUS)-carrying plasmid to lentil Lens culinaris) protoplasts, leading to transient expression of the GUS reporter gene. The transfection efficiency achieved by using stearylamine-containing liposomes (lipofection) was comparable to the one obtained by electroporating the protoplasts at 500 μF and different field strengths. Furthermore, the combination of electroporation and lipofection, though reducing cell survival, increased the activity of the reporter enzyme detected in the cell lysates, yielding transient expression levels higher than those recorded after lipofection or electroporation alone.     

Transfer factor of131I from the fallout to human thyroid dose equivalent after the Chernobyl accident

Summary A similar pattern of variation with time in observed maxima of daily dose equivalent rates in human thyroids (TD - µSv·d^–1) and of daily fallout radioactivities (FR - kBq·m^–2) has been found after the Chernobyl accident. An estimate of the time-lag between the maxima in TD lines and the preceding FR peaks was made of about seven days for adult and nine days for juveniles. Applying this time-lag it was possible to estimate transfer factors from the fall-out to thyroid dose equivalent: the highest estimated values were 221 µSv/kBq·m^–2 for adult and 641 µSv/kBq·m^–2 for juvenile thyroids. These values differ from those published by UNSCEAR (United Nations 1988), which have been calculated for various regions of Czechoslovakia, from ingestion and inhalation intake estimates. A broad variation of transfer factor values could be expected to result from such transfer calculations using ingestion and inhalation estimates. The findings also support the concept of a need for prolonged iodine prophylaxy after emissions of radioiodine into the environment.Abbreviations TD dose equivalent rates in thyroids [µSv·d^–1] - FR fall-out radioactivity (-ies) [kBq·m^–2]     

Nitrogen Transfer from Four Nitrogen-Fixer Associations to Plants and Soils

Nitrogen (N) fixation is the main source of ‘new’ N for N-limited ecosystems like subarctic and arctic tundra. This crucial ecosystem function is performed by a wide range of N₂ fixer (diazotroph) associations that could differ fundamentally in their timing and amount of N release to the soil. To assess the importance of different associative N₂ fixers for ecosystem N cycling, we tracked ¹⁵N-N₂ into four N₂-fixer associations (with a legume, lichen, free-living, moss) and into soil, microbial biomass and non-diazotroph-associated plants 3 days and 5 weeks after in situ labelling. In addition, we tracked ¹³C from ¹³CO₂ labelling to assess if N and C fixation are linked. Three days after labelling, half of the fixed ¹⁵N was recovered in the legume soils, indicating a fast release of fixed N₂. Within 5 weeks, the free-living N₂ fixers released two-thirds of the fixed ¹⁵N into the soil, whereas the lichen and moss retained the fixed ¹⁵N. Carbon and N₂ fixation were linked in the lichen shortly after labelling, in free-living N₂ fixers 5 weeks after labelling, and in the moss at both sampling times. The four investigated N₂-fixer associations released fixed N₂ at different rates into the soil, and non-diazotroph-associated plants have no access to ‘new’ N within several weeks after N₂ fixation. Although legumes and free-living N₂ fixers are immediate sources of ‘new’ N for N-limited tundra ecosystems, lichens and especially mosses, do not contribute to increase the N pool via N₂ fixation in the short term.     

Failure to transmit scrapie infection by transferring preimplantation embryos from naturally infected donor sheep

The objective of the study was to examine whether or not the preimplantation embryo can act as a carrier of classic scrapie infection. The study was carried out on quarantined premises with sheep of highly susceptible scrapie genotypes. Uninfected embryos, collected from New Zealand–derived Suffolk ewes, were surgically transferred into recipient ewes that were also of New Zealand origin. Seventeen negative control lambs were born on the study premises from these embryo transfers. Thirty-nine experimental lambs were from embryos collected from naturally infected donor ewes. The experimental lambs were also born on the study premises after their surgical transfer into recipient ewes of New Zealand origin. These embryos had been collected from donor ewes in a scrapie-infected flock where the ewes were clinically sick with scrapie or developed clinical scrapie after embryo collection. All lambs were confirmed as scrapie susceptible of the ARQ/ARQ genotype. Twenty-eight experimental animals survived to the end point of the study at 5 yr of age with a mean survival of 1579 d. In the negative control group, 12 of 17 sheep survived to 5 yr of age with a mean survival of 1508 d. Postmortem examinations were carried out on all animals derived by embryo transfer, and in none was histologic or immunohistochemical evidence of scrapie found. In contrast, in the originating flock the majority of scrapie cases occurred in ARQ/ARQ genotyped animals where a 56% mortality from scrapie had been recorded in animals of this genotype. Thus, the study provides no evidence for transmission of scrapie and reinforces published evidence that vertical transmission of scrapie may be circumvented by embryo transfer procedures.