The passage of spermatozoa through the cervix of ovariectomized ewes treated with progesterone and oestrogen.

Thirty spayed ewes were used in a 2 x 3 split-plot factorial experiment (n = 5) in which oestrus was induced with 30 or 90 mug oestradiol benzoate (OB) following a period of progesterone priming. They were inseminated 36 hr after oestrogen and the numbers of spermatozoa in the caudal, mid- and cranial regions of the cervix (sub-plots) were estimated 1, 12 and 24 hr later. At each interval of time and for each region of the cervix, fewer spermatozoa were recovered from the ewes treated with 30 mug OB than from those treated with 90 mug (P less than 0-05).     

Induction of ovulation and cyclic activity in anestrous ewes with testosterone treated wethers and ewes

An experiment was conducted to evaluate the effectiveness of wethers and ewes treated with testosterone preparations to induce ovulation and breeding activity in anestrous ewes. The testosterone was administered three times at weekly intervals. Wethers and ewes treated with 105 mg testosterone propionate and wethers treated with 100 mg testosterone from testosterone cyclopentyl propionate were as successful as vasectomised rams in inducing ovulation and cyclic activity in ewes. Seven days after the last injection the concentrations of testosterone in peripheral bloods were not significantly different from that in the vasectomised rams. By day 28 the concentrations were significantly lower than in the rams. The testosterone preparations tested are suitable for the induction of male sexual behavior and are rapidly excreted by sheep.     

Non-operative removal of bile duct stones by duodenoscopic sphincterotomy in the elderly.

Between January 1975 and December 1979, 71 patients over the age of 70 underwent attempted duodenoscopic sphincterotomy for stones in the common bile duct. Fifteen patients still had gall bladders in situ. Sphincterotomy was possible in 69 of the patients and in 65 of these duct clearance was achieved, giving an overall success rate of 92%. Failure to achieve sphincterotomy in two cases was due to substantial peripapillary diverticula. Duct clearance failed in four patients, mostly due to the size of the retained stones. The largest stone extracted was 24 mm diameter. There were no deaths but complications occurred in nine patients (13%); these were haemorrhage in four (requiring surgery in one), cholangitis in four (two of whom required surgical extraction of stones), and pancreatitis in one. The average duration of hospital stay in successful cases was 11 days (range three to 30). Clinical follow-up of 55 patients one to five years after sphincterotomy showed no evidence of stones or of stenosis of the sphincter. Duodenoscopic sphincterotomy is a major advance in the management of elderly patients with stones in the common bile duct.     

The decolorization of betacyanin by polyamines

The pigment betacyanin, the efflux of which has been used as a measure of membrane integrity in red beet discs, has now been shown to react with di- and polyamines with consequent decolorization.     

Characterization of 13 newly isolated strains of anaerobic, cellulolytic, thermophilic bacteria

Characteristics of 13 newly isolated thermophilic, anaerobic, and cellulolytic strains were compared with previously described strains of Clostridium thermocellum: ATCC 27405 and JW20 (ATCC 31549). Colony morphology, antibiotic sensitivity, fermentation end-products, and cellulose degradation were documented. All 13 strains were sensitive to erythromycin (5 μg/ml) and chloramphenicol (25 μg/ml), and all strains but one were sensitive to kanamycin (20 μg/ml). Polymerase chain reaction (PCR) amplification using primers based on gene sequences from C. thermocellum ATCC 27405 was successful for all 13 strains in the case of the hydrogenase gene and 11 strains in the case of phosphotransacetylase/acetate kinase genes. Ten strains amplified a product of the expected size with primers developed to be specific for C. thermocellum 16SrRNA primers. Two of the 13 strains did not amplify any product with the PCR primers designed for the phosphotransacetylase/acetate kinase and 16SrRNA primers. A MboI-like GATC- recognizing restriction activity was present in all of the five strains examined. The results of this study have several positive implications with respect to future development of a transformation system for cellulolytic thermophiles. Journal of Industrial Microbiology & Biotechnology (2001) 27, 275–280. Received 12 September 2000/ Accepted in revised form 20 November 2000     

Interactions of SARS-CoV-2 envelope protein with amilorides correlate with antiviral activity

SARS-CoV-2 is the novel coronavirus that is the causative agent of COVID-19, a sometimes-lethal respiratory infection responsible for a world-wide pandemic. The envelope (E) protein, one of four structural proteins encoded in the viral genome, is a 75-residue integral membrane protein whose transmembrane domain exhibits ion channel activity and whose cytoplasmic domain participates in protein-protein interactions. These activities contribute to several aspects of the viral replication-cycle, including virion assembly, budding, release, and pathogenesis. Here, we describe the structure and dynamics of full-length SARS-CoV-2 E protein in hexadecylphosphocholine micelles by NMR spectroscopy. We also characterized its interactions with four putative ion channel inhibitors. The chemical shift index and dipolar wave plots establish that E protein consists of a long transmembrane helix (residues 8–43) and a short cytoplasmic helix (residues 53–60) connected by a complex linker that exhibits some internal mobility. The conformations of the N-terminal transmembrane domain and the C-terminal cytoplasmic domain are unaffected by truncation from the intact protein. The chemical shift perturbations of E protein spectra induced by the addition of the inhibitors demonstrate that the N-terminal region (residues 6–18) is the principal binding site. The binding affinity of the inhibitors to E protein in micelles correlates with their antiviral potency in Vero E6 cells: HMA ≈ EIPA > DMA >> Amiloride, suggesting that bulky hydrophobic groups in the 5’ position of the amiloride pyrazine ring play essential roles in binding to E protein and in antiviral activity. An N15A mutation increased the production of virus-like particles, induced significant chemical shift changes from residues in the inhibitor binding site, and abolished HMA binding, suggesting that Asn15 plays a key role in maintaining the protein conformation near the binding site. These studies provide the foundation for complete structure determination of E protein and for structure-based drug discovery targeting this protein.     

Patch- and reach-scale dynamics of a macrophyte-invertebrate system in a New Zealand lowland stream

Abundant growths of macrophytes are a common feature of streams in open lowland areas of New Zealand during summer, but the values of these to aquatic biota are poorly understood. We studied the temporal dynamics of, and associations amongst, elements of a macrophyte-invertebrate system to provide an improved information base for lowland stream management. The biomass of macrophytes increased significantly over the four quarterly sampling occasions from 43.8 g m^-2 in June to 370.8 g m^-2 in March; biomass was dominated by Egeria densa on all dates, except in December when Potamogeton crispus was dominant. We did not detect strong associations between epiphyton biomass and invertebrate abundance in our study, but this may reflect the fact that we sampled loosely-adhering epiphyton on young, surface-reaching shoots whereas invertebrates were collected from macrophytes growing through the water column. Density of some invertebrate species per gram dry weight of plant material varied by macrophyte type, with the chironomids Tanytarsus vespertinus and Naonella forsythi displaying positive correlations with Egeria and Potamogeton biomass, respectively. The shrimp Paratya curvirostris accounted for 50% of phytophilous invertebrate biomass, with Chironomidae the only other group to comprise more than 9%. Abundance of total phytophilous invertebrates displayed a positive linear relationship with macrophyte biomass in a sample (0.1 m²), and a humped relationship with species richness, such that highest numbers of taxa occurred at macrophyte biomass levels around 400 g dw m^-2. Our study suggests that intermediate macrophyte biomass levels are likely to enhance macroinvertebrate biodiversity in sandy-bottomed lowland streams. This revised version was published online in July 2006 with corrections to the Cover Date.     

Neutrophils require SHP1 to regulate IL-1β production and prevent inflammatory skin disease

The regulation of neutrophil recruitment, activation, and disposal is pivotal for circumscribed inflammation. SHP1(Y208N/Y208N) mutant mice develop severe cutaneous inflammatory disease that is IL-1R dependent. Genetic reduction in neutrophil numbers and neutrophilic responses to infection is sufficient to prevent the spontaneous initiation of this disease. Neutrophils from SHP1(Y208N/Y208N) mice display increased pro-IL-1β production due to altered responses to MyD88-dependent and MyD88-independent signals. The IL-1R-dependent inflammatory disease in SHP1(Y208N/Y208N) mice develops independently of caspase 1 and proteinase 3 and neutrophil elastase. In response to Fas ligand, a caspase 1-independent inducer of IL-1β production, neutrophils from SHP1(Y208N/Y208N) mice produce elevated levels of IL-1β but display reduced caspase 3 and caspase 7 activation. In neutrophils deficient in SHP1, IL-1β induces high levels of pro-IL-1β suggesting the presence of a paracrine IL-1β loop. These data indicate that the neutrophil- and IL-1-dependent disease in SHP1(Y208N/Y208N) mice is a consequence of loss of negative regulation of TLR and IL-1R signaling.     

Cyclin-dependent kinase inhibitor Cdkn2c regulates B cell homeostasis and function in the NZM2410-derived murine lupus susceptibility locus Sle2c1

Sle2c1 is an NZM2410- and NZB-derived lupus susceptibility locus that induces an expansion of the B1a cell compartment. B1a cells have a repertoire enriched for autoreactivity, and an expansion of this B cell subset occurs in several mouse models of lupus. A combination of genetic mapping and candidate gene analysis presents Cdkn2c, a gene encoding for cyclin-dependent kinase inhibitor p18(INK4c) (p18), as the top candidate gene for inducing the Slec2c1-associated expansion of B1a cells. A novel single nucleotide polymorphism in the NZB allele of the Cdkn2c promoter is associated with a significantly reduced Cdkn2c expression in the splenic B cells and peritoneal cavity B1a cells from Sle2c1-carrying mice, which leads to a defective G1 cell cycle arrest in splenic B cells and increased proliferation of peritoneal cavity B1a cells. As the cell cycle is differentially regulated in B1a and B2 cells, these results suggest that Cdkn2c plays a critical role in B1a cell self-renewal and that its impaired expression leads to an accumulation of these cells with high autoreactive potential.     

Potential for fecundin to influence the reproductive performance of merino ewes in Western Australia

Four experiments were conducted at different locations in Western Australia to evaluate the effectiveness of immunizing young (maiden, 1 1/2 year old) and adult Merino ewes with Fecundin to improve their reproductive performances. The ovulation rates of immunized maiden ewes was increased (0.06-0.30) above that of untreated ewes in both experiments 1 and 2. However, there were no significant improvements in the marking percentages for the immunized ewes with the differences between the untreated and immunized ewes ranging from -16.4 to 5.8%. In comparison with untreated ewes immunization 6 and 2 weeks before the start of joining depressed ewe fertility by 26.1% whereas immunization 8 and 4 weeks before the start of joining did not significantly affect fertility. The availability of oat grain ad libitum prior to parturition and during early lactation did not improve the survival of lambs born to immunized ewes. In experiments 3 and 4 immunization of adult Merino ewes increased their ovulation rates (0.41-0.63) above untreated controls and tended to increase the proportion of pregnant ewes which had multiple pregnancies (from -2.3 to 34.2%). The responses at the end of lambing were variable (from -19.8 to 37.5% lambs marked) with high lamb mortalities occurring in some experiments. There was no adverse effect on the reproductive performances following consecutive annual immunizations over 3 years and the absence of treatment for 1 year did not prevent a response in the following year.