Determination of early pregnancy in ewes utilizing transrectal ultrasonography

Transrectal ultrasonography was used in ewes to determine the earliest day at which pregnancy could be detected, the number of embryos present, and the pattern of growth of the embryos. Twenty-one ewes were placed with 2 fertile rams and 20 ewes with 2 vasectomized rams. All ewes were treated to synchronize estrus and were observed for estrus twice daily. The 36 ewes that showed synchronized estrus were separated from the rams following mating. Transrectal ultrasonography was performed daily from estrus (Day 0) to Day 25 for all ewes and on Days 30, 35 and 40 post breeding for the 20 ewes mated to fertile rams. A 7.5 MHz transducer (human prostate, linear array) was utilized, with the ewes in dorsal recumbency in a tilting squeeze chute. Extraembryonic fluid and membranes were observed in the uterine horns ipsilateral to corpora lutea by Day 15 post breeding in all 17 ewes subsequently diagnosed as pregnant. Rhythmic pulsations (heartbeat) within the embryonic vesicles were first detected on Day 18 or 19. At least 1 embryo was detected by Day 20 in all the pregnant ewes, but not all the embryos were counted accurately until Day 25 (main effect of day; P < 0.05). Two ewes each had an embryo which died (absence of previously observed heartbeat) by Day 25 or Day 40, respectively, but each maintained the remaining embryos to term. The pattern of embryonic growth, as determined by crown-rump lengths on Days 20, 25, 30, 35 and 40, did not differ with the number of embryos carried (n = 1 to 4). In conclusion, transrectal ultrasonography was found to provide a rapid, accurate means for the early detection of pregnancy in ewes.     

Molecular and Phenotypic Characterization of a New Mouse Insertional Mutation That Causes a Defect in the Distal Vertebrae of the Spine

We have identified and characterized the phenotype of a new insertional mutation in one line of transgenic mice. Mice carrying this mutation, which we have designated TgN(Imusd)370Rpw, display undulations of the vertebrae giving rise to a novel kinky-tail phenotype. Molecular characterization of the insertion site indicates that the transgene integration has occurred without any substantial alterations in the structure of the host sequences. Using probes that flank the insertion site, we have mapped the mutation to chromosome 5 near the semidominant mutation, thick tail (Tht). Thick tail does not complement the TgN(Imusd)370Rpw mutation; compound mutants containing one copy of each mutation display a more severe phenotype than either mutation individually.     

Endocrine profiles of dairy cows following experimentally induced clinical mastitis during early lactation

Concentrations of LH, cortisol, estradiol-17beta (E(2)), prolactin and 13,14-dihydro-15-keto-prostaglandin F(2alpha) (PGFM) were determined in cows with experimentally induced clinical mastitis during early lactation. Cows free of intramammary infection (IMI) and in the luteal phase of the estrous cycle were balanced by lactation number and days in milk and assigned to either control (n=5) or treatment (n=5) groups. Treated cows were infected experimentally (day 0), in two mammary quarters, with Streptococcus uberis and developed clinical mastitis within 60 h after inoculation as evidenced by increased mastitis scores, elevated rectal temperatures, mammary swelling and isolation of S. uberis pathogen. Four days following bacterial challenge, blood samples were collected every 20 min for 8 h for determination of PGFM and LH following administration of oxytocin and GnRH, respectively. Blood samples were also collected on days 0, 4 and 7 of the experiment to determine concentrations of E(2), prolactin and cortisol. Four days after bacterial challenge, concentrations of cortisol were higher (P=0.04) in experimentally infected cows than controls. Experimentally challenged cows had increased (P=0.02) concentrations of cortisol on days 4 and 7 compared with day 0. Control cows had no significant increase in blood cortisol during the experimental period. Baseline concentrations of PGFM did not differ between groups; however, peak concentrations of PGFM following oxytocin challenge were elevated (P=0.006) in cows with clinical mastitis compared with control animals. Prolactin, E(2) and LH did not differ between cows with clinical mastitis or controls. Experimentally induced mastitis during early lactation elevated concentrations of cortisol during the luteal phase of the estrous cycle. Furthermore, mastitic cows demonstrated an increased PGFM response following oxytocin administration. Altered reproductive efficiency in cows with clinical mastitis caused by Gram-positive pathogens may be the result of increased uterine sensitivity to prostaglandin F(2alpha) (PGF(2alpha)).     

Glucosylceramides are critical for cell‐type differentiation and organogenesis,but not for cell viability in Arabidopsis

Glucosylceramides (GlcCer), glucose‐conjugated sphingolipids, are major components of the endomembrane system and plasma membrane in most eukaryotic cells. Yet the quantitative significance and cellular functions of GlcCer are not well characterized in plants and other multi‐organ eukaryotes. To address this, we examined Arabidopsis lines that were lacking or deficient in GlcCer by insertional disruption or by RNA interference (RNAi) suppression of the single gene for GlcCer synthase (GCS, At2g19880), the enzyme that catalyzes GlcCer synthesis. Null mutants for GCS (designated ‘gcs‐1’) were viable as seedlings, albeit strongly reduced in size, and failed to develop beyond the seedling stage. Heterozygous plants harboring the insertion allele exhibited reduced transmission through the male gametophyte. Undifferentiated calli generated from gcs‐1 seedlings and lacking GlcCer proliferated in a manner similar to calli from wild‐type plants. However, gcs‐1 calli, in contrast to wild‐type calli, were unable to develop organs on differentiation media. Consistent with a role for GlcCer in organ‐specific cell differentiation, calli from gcs‐1 mutants formed roots and leaves on media supplemented with the glucosylated sphingosine glucopsychosine, which was readily converted to GlcCer independent of GCS. Underlying these phenotypes, gcs‐1 cells had altered Golgi morphology and fewer cisternae per Golgi apparatus relative to wild‐type cells, indicative of protein trafficking defects. Despite seedling lethality in the null mutant, GCS RNAi suppression lines with ≤2% of wild‐type GlcCer levels were viable and fertile. Collectively, these results indicate that GlcCer are essential for cell‐type differentiation and organogenesis, and plant cells produce amounts of GlcCer in excess of that required for normal development.     

Sequence analysis of the human hTg737 gene and its polymorphic sites in patients with autosomal recessive polycystic kidney disease

DNA sequence analysis of the human Tg737 gene was performed in 36 patients with the autosomal recessive form of polycystic kidney disease (ARPKD). Coding exons and their adjacent splice sites were screened for mutations. Pathogenic exon or splice region mutations were not identified although one exonic and two intronic polymorphic sites were discovered. These results are in agreement with another study that has recently reported linkage to Chromosome (Chr) 6p21-cen in a set of 16 ARPKD families. STS mapping has localized the gene to a YAC contig that includes D13S175 on chromosome 13q12.1. The polymorphisms found in the hTg737 gene will permit its future evaluation as a candidate gene for other recessive cystic renal diseases and as a modifier gene in human PKD.     

Relationship of pregnancy rate to peripheral concentrations of progesterone and estradiol in beef cows

The relationship between pregnancy rate and concentrations of progesterone (P(4)) and estradiol-17beta (E(2)) in serum was examined in inseminated beef cows. Jugular blood was collected twice daily on Days 4 through 7 and Days 14 through 17 after estrus to establish patterns of secretion of P(4) and E(2). Pregnancy rate was determined by palpation per rectum at 45 d. Mean concentrations of each hormone, ratio of E(2):P(4) and regressions of hormone on day were the variables measured for each of the 2 periods. Cows were classified into low (n=26), medium (n=50) and high (n=26) groups for each variable. The relationship of pregnancy rate to each variable was tested using Chi-square analyses. Pregnancy rates to the first service decreased linearly as relative mean concentrations of E(2) increased on Days 14 through 17 (P<0.05) but were not affected by any of the other hormonal variables studied during either period. Pregnancy rates to the second service were not related to concentrations of P(4) or E(2) during the luteal phase before mating (Days 14 through 17). The effects of pregnancy on concentrations of E(2) and P(4) also were tested. On Days 14 through 17, P(4) increased slightly in pregnant cows and declined slightly in nonpregnant cows (P<0.05), but pregnancy did not affect E(2) during either period or P(4) on Days 4 through 7. In summary, pregnancy rate to the first service decreased significantly as concentrations of E(2) increased on Days 14 through 17.