The Mitochondrial Genome in Embryo Technologies

The mammalian mitochondrial genome encodes for 37 genes which are involved in a broad range of cellular functions. The mitochondrial DNA (mtDNA) molecule is commonly assumed to be inherited through oocyte cytoplasm in a clonal manner, and apparently species‐specific mechanisms have evolved to eliminate the contribution of sperm mitochondria after natural fertilization. However, recent evidence for paternal mtDNA inheritance in embryos and offspring questions the general validity of this model, particularly in the context of assisted reproduction and embryo biotechnology. In addition to normal mt DNA haplotype variation, oocytes and spermatozoa show remarkable differences in mtDNA content and may be affected by inherited or acquired mtDNA aberrations. All these parameters have been correlated with gamete quality and reproductive success rates. Nuclear transfer (NT) technology provides experimental models for studying interactions between nuclear and mitochondrial genomes. Recent studies demonstrated (i) a significant effect of mtDNA haplotype or other maternal cytoplasmic factors on the efficiency of NT; (ii) phenotypic differences between transmitochondrial clones pointing to functionally relevant nuclear–cytoplasmic interactions; and (iii) neutral or non‐neutral selection of mtDNA haplotypes in heteroplasmic conditions. Mitochondria form a dynamic reticulum, enabling complementation of mitochondrial components and possibly mixing of different mtDNA populations in heteroplasmic individuals. Future directions of research on mtDNA in the context of reproductive biotechnology range from the elimination of adverse effects of artificial heteroplasmy, e.g. created by ooplasm transfer, to engineering of optimized constellations of nuclear and cytoplasmic genes for the production of superior livestock.     

Impact of RYR1 genotype of Piétrain boars on litter traits of Czech Large White × Czech Landrace crossbred sows

Summary Piétrain boars of different ryanodine receptor (RYR1) genotypes (NN, Nn and nn, three boars each) were mated with approximately 10 Czech Large White × Czech Landrace sows (genotype NN) each to produce one litter per sow. The progeny of nn boars had a significantly higher individual weaning weight (7.31 kg versus 6.86 kg) and average daily gain from birth to weaning (252 g/day versus 240 g/day) than the progeny of NN boars. Furthermore, piglets from nn boars differed statistically significant also from piglets of boars with the Nn genotype for these two traits. The litters of the NN boars were greater by 0.5 piglets than the litters of the nn boars for the total number of piglets born, piglets born alive and piglets weaned, but the differences were not significant. The Nn genotype showed for all litter size traits and the number of stillborn piglets the best values. The difference between the genotypes Nn and nn was statistically significant at the 0.05 level for the number of piglets born alive and the number of piglets weaned.     

High-dose-rate brachytherapy for intranasal tumours in dogs: results of a pilot study

This prospective study describes the feasibility and toxicity of ¹⁹²Iridium high‐dose‐rate (HDR) brachytherapy as an alternative strategy for the treatment of canine intranasal tumours. Fifteen dogs with malignant intranasal tumours were treated twice weekly using a hypofractionated protocol with eight fractions, 5 Gy per fraction, resulting in a total dose of 40 Gy. Acute and chronic adverse side‐effects appeared to be rare. Only 7% of the acute side‐effects and 5% of the chronic were classified as severe (grade 3). Eight dogs showed clinical complete remission, and five dogs had partial remission, with a resolution of tumour‐related symptoms. Magnetic resonance imaging showed a reduced tumour mass in 12 cases. Median survival time was 17 months (range 4–48 months), with four dogs (three without disease) still alive. Median time to recurrence of these dogs was 14 months. In nine dogs, progression or recurrence of the tumour was the cause of death. This study suggests that HDR brachytherapy is feasible and well tolerated.     

A longitudinal study of Salmonella enterica infections in high-and low-seroprevalence finishing swine herds in The Netherlands

The purpose of this investigation was to study the incidence and course of Salmonella infections in finishing pig herds in order to asses the stability of a given Salmonella herd status. Five low- and 7 high-seroprevalence herds were followed for seven sampling rounds. Each round, blood and faecal samples were tested in an indirect ELISA and by bacteriological culturing, respectively. In high-seroprevalence herds a positive Salmonella status was an indication of a long-term problem and the status was relatively stable over time. The herds experiencing clinical salmonellosis were not necessarily the herds with the highest seroprevalence. It is possible to deliver sero-negative finishers to the slaughterhouse, even though these pigs were seropositive as growers. In three out of five low-prevalence herds, major infection incidents occurred, indicating that changes in the Salmonella status should be anticipated. Low-prevalence herds can remain negative over a longer period of time as a result feeding a complete liquid feed containing fermented by-products.     

Sodium transport across the isolated epithelium of sheep omasum is influenced by luminal ammonia

Ammonia is a physiological fermentation product in the forestomachs and is absorbed from the rumen and omasum. Cellular uptake of ammonia affects the intracellular pH of polar and non-polar cells. The effect of the uptake on the pH of the cytosol depends on the predominant form of ammonia. NH(3) uptake and its intracellular protonation tend to alkalinize the cytoplasm, whereas the uptake of NH(4)(+) acidifies the cytoplasm by reversing this reaction. Consequently, the absorption of ammonia across the omasal epithelium could cause a change of the intracellular pH and pH-dependent transport mechanisms like Na/H exchange. Because no information is available about the form of ammonia absorbed in the omasum and, hence, possible modulation of Na transport by ammonia, the effect of increasing luminal ammonia concentrations (0, 5, 15 and 30 mmol/l) on Na transport were studied. In epithelia of hay-fed animals, ammonia linearly inhibited Na transport in a dose-dependent manner, at a luminal pH of 7.40, but not at a pH of 6.40. Ammonia did not influence Na transport in epithelia of concentrate-fed animals. Because luminal ammonia did not consistently change the short circuit current or tissue conductance absorption of ammonia as NH(4)(+) appears to be unlikely. The predominant form of ammonia absorbed in the omasum is probably NH(3), which is protonated in the cytosol. The reduced availability of protons may be the cause of inhibition of Na transport via Na/H exchange.     

Practice, Policy, and Pedagogy in a Mandatory Environmental Law Course

All students at the University of Richmond School of Law are required to complete a first-year course in Environmental Law, as an introduction to the strategies and technicalities of statutory and regulatory regimes. This requirement poses special pedagogical challenges for the instructors. How, for example, is the class to address pollution questions regarding diverse media? Which of the many federal and nonfederal statutory provisions deserve the most (and least attention)? How does the instructor hold the attention of, and gain the most effective work product from, students whose major interests lie outside the area of Environmental Law? What is the proper mix of statutory, regulatory, and judge-made (pre- and post-regulatory) law? Our solution has been to craft a unique combination of traditional lectures and readings regarding specific statutes, regulations, and case law; weekly discussion questions that immerse the students in the intricacies of statutory and regulatory provisions; a series of small-group simulations designed to highlight substantive legal points and to place the students in a wide range of legal and extralegal settings (client interviews, appellate argument, advocacy before administrative bodies, negotiation, lobbying public officials, legislative drafting); and periodic written assignments that focus the students' attention on technical legal and public policy issues.     

Rapid Trichloroacetic Acid Extraction and Liquid Chromatography Method for Determination of Nicotinamide in Commercial Cereals

Determination of niacin in fortified infant and dairy products has been accomplished using a variety of analytical liquid chromatography (LC) methods. Applications of these LC techniques to other food matrices suffer due to the presence of endogenous absorbing peaks at 260 nm that co‐chromatograph with the nicotinic acid and nicotinamide vitamers. We have successfully adapted the LC method of Woollard and Indyk for the determination of nicotinamide in reference and commercial cereal products. Unbound nicotinamide in fortified cereal was extracted with 0.6M tri‐chloroacetic acid and chromatographed on a C₁₈ reversed‐phase column using a mobile phase of 75% methanol and water (pH 2.8, with formic acid) with sodium dioctylsulphosuccinate (0.1%) as the ion‐pairing agent. Using Spectral Analysis ChromQuest software, a three‐dimensional view showed only nicotinamide under the LC peak. Similarity index spectral matches of nicotinamide standard and the LC peak were ≈100%, indicating the absence of interferences. Nicotinamide recoveries for the reference cereals of VMA195 and VMA 399 (from AACC International, St. Paul, MN) and GM 19B (from General Mills, Medallion Laboratories, Minneapolis, MN) were 90–103% of assigned value. Experimental values for oat, corn, rice, and bran cereals showed that actual niacin content in commercial cereals may be significantly above (111–170%) declared label values. Because manufacturers may fortify at a level higher than the declared label level to ensure shelf life compliance, these data do have significant implications when making precise estimates of niacin intake based on label claims.