Endocrine Profiles, Haematology and Pregnancy Outcomes of Late Pregnant Holstein Dairy Heifers Sired by Bulls Giving a High or Low Incidence of Stillbirth

The high incidence of stillbirth in Swedish Holstein heifers has increased continuously during the last 15 years to an average of 11% today. The pathological reasons behind the increased incidence of stillbirth are unknown. The present experiment was undertaken to investigate possible causes of stillbirth and to study possible physiological markers for predicting stillbirth. Twenty Swedish Holstein dairy heifers sired by bulls with breeding values for a high risk of stillbirth (n = 12) (experimental group) and a low risk of stillbirth (n = 8) (control group, group B) were selected based on information in the Swedish AI-data base. The experimental group consisted of 2 subgroups of heifers (groups A1 and A2) inseminated with 2 different bulls with 3.5% and 9% higher stillbirth rates than the average, and the control group consisted of heifers pregnant with 5 different bulls with 0%–6% lower stillbirth rates than the average. The bull used for group A1 had also calving difficulties due to large calves as compared to the bull in group A2 showing no calving difficulties. The heifers were supervised from 6–7 months of pregnancy up to birth, and the pregnancies and parturitions were compared between groups regarding hormonal levels, haematology, placental characteristics and calf viability. In group A1, 1 stillborn, 1 weak and 4 normal calves were recorded. In group A2, 2 stillborn and 4 normal calves were registered. All animals in the control group gave birth to a normal living calf without any assistance. The weak calf showed deviating profiles of body temperature, saturated oxygen and heart rates, compared with the normal living calves. No differences of the placentome thickness, measured in vivo by ultrasonography were seen between the groups. The number of leukocytes and differential cell counts in groups A1 and A2 followed the profiles found in the control group. In group A1, a slight decrease of oestrone sulphate (E1SO4) levels was found in the animal delivering a stillborn calf from the first 24-h blood sampling at 6 weeks to the second at 3 weeks prior to delivery, while the levels of E1SO4 at both periods in the animal delivering a weak calf followed the profile in animals delivering a normal living calf. During late pregnancy and at the time of parturition, the levels of E1SO4 and PAGs in animals delivering a stillborn or weak calf (from group A1) followed the normal profiles found in animals delivering a normal living calf. In group A2, low levels of E1SO4 and pregnancy associated glycoproteins (PAGs) over 24 h at both 3 and 6 weeks prior to parturition (<1.5 nmol/L) were recorded in animals delivering a stillborn calf. During late pregnancy and parturition, the levels of E1SO4 and PAGs were slightly lower during 30–50 days prior to delivery and increased with a lower magnitude at the time of parturition. In conclusion, our results indicate that the aetiology behind stillbirth varies depending on the AI-bulls used and is associated with dystocia or low viability of the calves. Deviating profiles of oestrone sulphate (E1SO4) and pregnancy associated glycoproteins (PAGs) in animals delivering a stillborn calf not caused by dystocia were observed, suggesting placental dysfunction as a possible factor. The finding suggests that the analyses of E1SO4 and PAGs could be used for monitoring foetal well-being in animals with a high risk of stillbirth at term.     

Staining of intracellular deposits of uranium in cultured murine macrophages

In our studies of the health effects of internalized depleted uranium, we developed a simple and rapid light microscopic method to stain specifically intracellular uranium deposits. Using J774 cells, a mouse macrophage line, treated with uranyl nitrate and the pyridylazo dye 2-(5-bromo-2- pyridylazo)-5-diethylaminophenol, uranium uptake by the cells was followed. Specificity of the stain for uranium was accomplished by using masking agents to prevent the interaction of the stain with other metals. Prestaining wash consisting of a mixture of sodium citrate and ethylenediaminetetraacetic acid eliminated staining of metals other than uranium. The staining solution consisted of the pyridylazo dye in borate buffer along with a quaternary ammonium salt, ethylhexadecyldimethylammonium bromide, and the aforementioned sodium citrate/ethylene-diaminetetraacetic acid mixture. The buffer was essential for maintaining the pH within the optimum range of 8 to 12, and the quaternary ammonium salt prevented precipitation of the dye. Staining was conducted at room temperature and was complete in 30 min. Staining intensity correlated with both uranyl nitrate concentration and incubation time. Our method provides a simple procedure for detecting intracellular uranium deposits in macrophages.     

Gene expression in abnormal ovarian structures of ewes homozygous for the inverdale prolificacy gene

Animals heterozygous (I+) for the Inverdale prolificacy gene (FecX(I)) have an increased ovulation rate whereas those homozygous (II) for FecX(I) are infertile with "streak" ovaries and follicular development arrested at the primary (type 2 follicle) stage. The streak ovaries also contain small oocyte-free nodules with granulosa-like cells and often tumor-like structures. It has been hypothesized that these abnormal structures are of granulosa cell origin, and the aim of this study was to determine whether genes normally expressed in granulosa cells are also expressed in the nodules and tumor-like structures. The mRNAs encoding c-kit and its ligand stem cell factor (SCF), FSH receptor (FSH-R), follistatin, alpha-inhibin subunit, and the beta(A)- and beta(B)-activin/inhibin subunits were localized in ovaries of ewes with 0 (++), 1 (I+), or 2 (II) copies of the FecX(I) gene (n = 4-9 animals per genotype per gene) using in situ hybridization. Ontogeny of expression of all mRNAs examined was similar between ++ and I+ ewes. Expression of c-kit mRNA was observed in the oocyte of all follicular types present in ++, I+, and II ewes. Moreover, granulosa cells of type 2 (II) and type 2 and larger follicles (++, I+) expressed SCF mRNA. The mRNAs encoding FSH-R, follistatin, alpha-inhibin subunit, and beta(B)-activin/inhibin subunit were identified in type 3 and larger follicles of ++ and I+ ewes but not in follicles of II ewes that were only at the type 1, 1a, or 2 stages of development. However, the cells within the oocyte-free nodules of II ewes expressed all of these genes. The mRNAs encoding c-kit and beta(A)-activin/inhibin subunit were not observed in granulosa cells until antrum formation (type 5 follicles) or in the nodules of II ewes. Tumors from 4 ewes were obtained and classified as cystic, semisolid, or solid structures containing granulosa-like cells or as solid structures containing predominately fibroblast- and luteal-like cells. Often, two tumors were present on the same ovary. Tumors containing granulosa-like cells (n = 3-4 per gene) expressed the mRNAs encoding alpha-inhibin subunit, beta(A)-, and beta(B)-activin/inhibin subunits, follistatin, and the FSH-R but did not contain detectable amounts of mRNA for c-kit or SCF. Tumors composed predominately of fibroblast- and luteal-like cells expressed very low levels of SCF mRNA; of the other mRNAs examined, none were detected. Also, none of the genes examined were found to be expressed by the surface epithelium, theca externa, fibroblast, or vascular cells within the ovary of animals of any genotype. These findings are consistent with the hypothesis that the somatic cells in oocyte-free nodules and tumor-like tissue in II ewes originate from the granulosa cells of the small follicles.     

Phylogenetic relationships within the Alcidae (Charadriiformes: Aves) inferred from total molecular evidence

The Alcidae is a unique assemblage of Northern Hemisphere seabirds that forage by "flying" underwater. Despite obvious affinities among the species, their evolutionary relationships are unclear. We analyzed nucleotide sequences of 1,045 base pairs of the mitochondrial cytochrome b gene and allelic profiles for 37 allozyme loci in all 22 extant species. Trees were constructed on independent and combined data sets using maximum parsimony and distance methods that correct for superimposed changes. Alternative methods of analysis produced only minor differences in relationships that were supported strongly by bootstrapping or standard error tests. Combining sequence and allozyme data into a single analysis provided the greatest number of relationships receiving strong support. Addition of published morphological and ecological data did not improve support for any additional relationship. All analyses grouped species into six distinct lineages: (1) the dovekie (Alle alle) and auks, (2) guillemots, (3) brachyramphine murrelets, (4) synthliboramphine murrelets, (5) true auklets, and (6) the rhinoceros auklet (Cerorhinca monocerata) and puffins. The two murres (genus Uria) were sister taxa, and the black guillemot (Cepphus grylle) was basal to the other guillemots. The Asian subspecies of the marbled murrelet (Brachyramphus marmoratus perdix) was the most divergent brachyramphine murrelet, and two distinct lineages occurred within the synthliboramphine murrelets. Cassin's auklet (Ptychoramphus aleuticus) and the rhinoceros auklet were basal to the other auklets and puffins, respectively, and the Atlantic (Fratercula arctica) and horned (Fratercula corniculata) puffins were sister taxa. Several relationships among tribes, among the dovekie and auks, and among the auklets could not be resolved but resembled "star" phylogenies indicative of adaptive radiations at different depths within the trees.      

Evidence for para Dechlorination of Polychlorobiphenyls by Methanogenic Bacteria

When microorganisms eluted from upper Hudson River sediment were cultured without any substrate except polychlorobiphenyl (PCB)-free Hudson River sediment, methane formation was the terminal step of the anaerobic food chain. In sediments containing Aroclor 1242, addition of eubacterium-inhibiting antibiotics, which should have directly inhibited fermentative bacteria and thereby should have indirectly inhibited methanogens, resulted in no dechlorination activity or methane production. However, when substrates for methanogenic bacteria were provided along with the antibiotics (to free the methanogens from dependence on eubacteria), concomitant methane production and dechlorination of PCBs were observed. The dechlorination of Aroclor 1242 was from the para positions, a pattern distinctly different from, and more limited than, the pattern observed with untreated or pasteurized inocula. Both methane production and dechlorination in cultures amended with antibiotics plus methanogenic substrates were inhibited by 2-bromoethanesulfonic acid. These results suggest that the methanogenic bacteria are among the physiological groups capable of anaerobic dechlorination of PCBs, but that the dechlorination observed with methanogenic bacteria is less extensive than the dechlorination observed with more complex anaerobic consortia.     

Effects of oestradiol, zeranol or trenbolone acetate implants on puberty, reproduction and fertility in heifers

The effects of anabolic agents on reproduction in beef heifers were determined by using 300 mg trenbolone acetate (TBA), 36 mg zeranol and 19 mg oestradiol-17 beta in a biodegradable pellet (1E: American Cyanamid, USA), or two such pellets (2E). On Day 1 of experiment, 81 Hereford x Friesian heifers (mean age = 84 +/- 1.2 days) were allocated at random to the following treatments: (1) controls (N = 15); (2) TBA (N = 15); (3) 1E (N = 12); (4) 2E (N = 15); (5) zeranol (N = 13); (6) TBA + 2E (N = 11). The 1 (1E), or 2 (2E) oestradiol implants were administered on Day 1 of the experiment only. Heifers assigned to receive TBA and zeranol were implanted on Days 1, 84, 168 and 252. Blood progesterone concentrations and oestrous activity were monitored from Days 137 and 200 respectively. Mean age (days) and weight (kg) at puberty (first ovulation), for heifers that reached puberty in Groups 1-6 respectively were 352 and 308, 419 and 356, 373 and 325, 381 and 331, 400 and 353, 423 and 383 [residual standard deviation (r.s.d.) = 43.8 and 39.4 for age and weight respectively]. Heifers in Group 4 were older (P less than 0.05), but not heavier (P greater than 0.05), while those in Groups 2 and 5 were both older (P less than 0.005) and heavier (P less than 0.005) than the controls at puberty. Age and weight at puberty were not different in heifers assigned to Groups 3 and 4, or to Groups 2 and 6. The proportion of heifers showing oestrus before puberty (prepubertal oestrus) were 3/15, 12/15, 6/12, 7/15, 10/13 and 11/11 in Groups 1-6 respectively. Heifers in Groups 2 and 5 had higher incidences of prepubertal oestrus than controls, while those in other treatment groups were not different. There was no treatment effect on the incidence of silent ovulations, but the incidence of non-ovulatory oestrus, after puberty, was increased from 4/48 in Group 1 to 26/40 (P less than 0.001), 15/56 (P less than 0.05) and 34/57 (P less than 0.001) in Groups 2, 4 and 5, respectively. Heifers in Group 6 had a higher incidence of non-ovulatory oestrus (P less than 0.05), but not of prepubertal oestrus, than did those in Group 2.(ABSTRACT TRUNCATED AT 400 WORDS)     

Glycosylation sites and site-specific glycosylation in human Tamm- Horsfall glycoprotein

The N-glycosylation sites of human Tamm-Horsfall glycoprotein from one healthy male donor have been characterized, based on an approach using endoproteinase Glu-C (V-8 protease, Staphylococcus aureus ) digestion and a combination of chromatographic techniques, automated Edman sequencing, and fast atom bombardment mass spectrometry. Seven out of the eight potential N-glycosylation sites, namely, Asn52, Asn56, Asn208, Asn251, Asn298, Asn372, and Asn489, turned out to be glycosylated, and the potential glycosylation site at Asn14, being close to the N-terminus, is not used. The carbohydrate microheterogeneity on three of the glycosylation sites was studied in more detail by high-pH anion-exchange chromatographic profiling and 500 MHz1H-NMR spectroscopy. Glycosylation site Asn489 contains mainly di- and tri-charged oligosaccharides which comprise, among others, the GalNAc4 S (beta1-4)GlcNAc terminal sequence. Only glycosylation site Asn251 bears oligomannose-type carbohydrate chains ranging from Man5GlcNAc2to Man8GlcNAc2, in addition to a small amount of complex- type structures. Profiling of the carbohydrate moieties of Asn208 indicates a large heterogeneity, similar to that established for native human Tamm-Horsfall glycoprotein, namely, multiply charged complex-type carbohydrate structures, terminated by sulfate groups, sialic acid residues, and/or the Sda-determinant.