Mortality in Grey seal pups: incidence and causes

Pup production and mortality of beach breeding Grey seals was studied at two contrasting types of rookery, the cliff-bound island of Ramsey, Dyfed, and the low, grassy island of Auskerry, Orkney. Thirty-five per cent of pups died on Ramsey compared with 14 % on Auskerry. Almost half of the Ramsey mortality was accounted for by animals being lost from the beaches so that they were not available for analysis. The main proximate causes of death revealed by post-mortem examination were starvation and infections. The other conditions which accounted for a small proportion of deaths were drowning, trauma, non-viability, stillbirths, atelectasis, dystokia and heart abnormality. Certain pathogens associated with disease were common to both sites, while others were specific to Ramsey or Auskerry.
The most important ultimate cause of pup deaths appeared to be failure of the mother/pup bond. The differences in levels of mortality between the sites is thought to be due to differences in the topography of the beaches. Pup survival on the narrow cliff-bound beaches, or in caves, on Ramsey was reduced either directly through pups being washed off, or indirectly by overcrowding of animals at high tide.     

Direct comparison of binding equilibrium, thermodynamic, and rate constants determined by surface- and solution-based biophysical methods

The binding interactions of small molecules with carbonic anhydrase II were used as model systems to compare the reaction constants determined from surface- and solution-based biophysical methods. Interaction data were collected for two arylsulfonamide compounds, 4-carboxybenzenesulfonamide (CBS) and 5-dimethyl-amino-1-naphthalene-sulfonamide (DNSA), binding to the enzyme using surface plasmon resonance, isothermal titration calorimetry, and stopped-flow fluorescence. We demonstrate that when the surface plasmon resonance biosensor experiments are performed with care, the equilibrium, thermodynamic, and kinetic constants determined from this surface-based technique match those acquired in solution. These results validate the use of biosensor technology to collect reliable data on small molecules binding to immobilized macromolecular targets. Binding kinetics were shown to provide more detailed information about complex formation than equilibrium constants alone. For example, although carbonic anhydrase II bound DNSA with twofold higher affinity than CBS, kinetic analysis revealed that CBS had a fourfold slower dissociation rate. Analysis of the binding and transition state thermodynamics also revealed significant differences in the enthalpy and entropy of complex formation. The lack of labeling requirements, high information content, and high throughput of surface plasmon resonance biosensors will make this technology an important tool for characterizing the interactions of small molecules with enzymes and receptors.     

Identification of the Escherichia coli sohB gene, a multicopy suppressor of the HtrA (DegP) null phenotype.

We cloned and sequenced the sohB gene of Escherichia coli. The temperature-sensitive phenotype of bacteria that carry a Tn10 insertion in the htrA (degP) gene is relieved when the sohB gene is present in the cell on a multicopy plasmid (30 to 50 copies per cell). The htrA gene encodes a periplasmic protease required for bacterial viability only at high temperature, i.e., above 39 degrees C. The sohB gene maps to 28 min on the E. coli chromosome, precisely between the topA and btuR genes. The gene encodes a 39,000-Mr precursor protein which is processed to a 37,000-Mr mature form. Sequencing of a DNA fragment containing the gene revealed an open reading frame which could encode a protein of Mr 39,474 with a predicted signal sequence cleavage site between amino acids 22 and 23. Cleavage at this site would reduce the size of the processed protein to 37,474 Mr. The predicted protein encoded by the open reading frame has homology with the inner membrane enzyme protease IV of E. coli, which digests cleaved signal peptides. Therefore, it is possible that the sohB gene encodes a previously undiscovered periplasmic protease in E. coli that, when overexpressed, can partially compensate for the missing HtrA protein function.     

Safety zones for a Sclerotinia sclerotiorum-based mycoherbicide: Accounting for regional and yearly variation in climate

Variation in the width of 'safety zones' for sheep and dairy pasture treated with a Sclerotinia sclerotiorum (Lib.) de Bary-based mycoherbicide was quantified using 10 years of climate data from each of five regions in New Zealand. Acceptable and risk-averse zones were defined as the maximum distances in any direction from a treated pasture where 'added:natural' ratios of air-borne S. sclerotiorum spores are 1:1 and 1:10, respectively. The 10-year mean 1:1 safety zone had zero width for sheep pasture at all locations, and was at most, 50 m wide for dairy pasture. The width of the 10-year mean 1:10 zone varied regionally from 314 to 443 m for sheep and from 175 to 280 m for dairy pasture, and the 90th percentile 1:10 zones were up to 41% wider. Linear relationships between safety zone width and mean wind speed were evident and these could be used to derive region-specific safety zones.     

Molecular characterization of four β-tubulin genes from dinitroaniline susceptible and resistant biotypes of Eleusine indica

Dinitroaniline herbicides are antimicrotubule drugs that bind to tubulins and inhibit polymerization. As a result of repeated application of dinitroaniline herbicides, resistant biotypes of goosegrass (Eleusine indica) developed in previously susceptible wild-type populations. We have previously reported that -tubulin missense mutations correlate with dinitroaniline response phenotypes (Drp) (Plant Cell 10: 297–308, 1998). In order to ascertain associations of other tubulins with dinitroaniline resistance, four -tubulin cDNA classes (designated TUB1, TUB2, TUB3, and TUB4) were isolated from dinitroaniline-susceptible and -resistant biotypes. Sequence analysis of the four -tubulin cDNA classes identified no missense mutations. Identified nucleotide substitutions did not result in amino acid replacements. These results suggest that the molecular basis of dinitroaniline resistance in goosegrass differs from those of colchicine/dinitroaniline cross-resistant Chlamydomonas reinhardtii and benzimidazole-resistant fungi and yeast. Expression of the four -tubulins was highest in inflorescences. This is in contrast to -tubulin TUA1 that is expressed predominantly in roots. Collectively, these results imply that -tubulin genes are not associated with dinitroaniline resistance in goosegrass. Phylogenetic analysis of the four -tubulins, together with three -tubulins, suggests that the resistant biotype developed independently in multiple locations rather than spreading from one location.     

Chemical modification of nucleic acids. Methylation of calf thymus DNA investigated by mass spectrometry and liquid chromatography

Mass spectrometry provides an extremely sensitive method for the identification and quantification of modified nucleosides and hence for determining chemical modifications of nucleic acids. When mass spectrometry is used in conjunction with a new high-performance liquid chromatographic system capable of separating 15 methylated and naturally occurring nucleosides, this allows the quantification of products of in vitro DNA methylation. With synthetic (2H3)methyl-labeled methylnucleosides as internal references, the distribution of methylated products formed when calf thymus DNA was reacted with N-methyl-N-nitrosourea(MeNU) was determined. Five modified products, 1-methyldeoxyadenosine(m1dA), 3-methyldeoxycytidine(m3dC), 7-methyldeoxyguanosine(m7dG), 3-methylthymidine(m3T) and O4-methylthymidine(m4T) were detected and the relative distributions were measured. The ability of mass spectrometry/mass spectrometry (tandem mass spectrometry) to increase specificity and sensitivity in this determination is demonstrated and its application to in vivo studies is suggested.     

Is prostaglandin F2 alpha involved in the increased myometrial contractility of primary dysmenorrhoea?

The concentrations of prostaglandin F2 alpha (PGF2 alpha) and E2 (PGE2) in menstrual fluid collected daily from 13 women with primary dysmenorrhoea and 11 matched controls, were compared with the pattern of uterine contractility during the hour following the menstrual fluid collection. The intra-uterine pressure (IUP) was measured using a micro-transducer catheter and the tracings analysed. On Day 2 the concentration of PGF2 alpha correlated with the peak area, but not with amplitude, duration or rate of contraction. These findings add additional support to the hypothesis that increased production of PGF2 alpha could contribute to the increased uterine contractility in primary dysmenorrhoea.     

An in vitro micro-volume procedure for rapid measurement of erythrocytic hexose monophosphate shunt activity

A radiometric micro-volume procedure for measurement of erythrocytic hexose monophosphate shunt (HMS) activity in intact cells in vitro is described. The procedure is rapid, allowing 200 individual HMS determinations in a single experiment of 5 hr duration. The procedure is reproducible, yielding HMS activity means insignificantly different (P greater than 0.05) between replicate experiments. A profile of sodium nitrite-induced HMS stimulation is reported: HMS was elevated 2-fold (P less than 0.001) between zero and 2.5 mM NaNO2; HMS elevation was more distinct (7-fold) between 2.5 and 5.0 mM NaNO2; maximum activity (22-fold) was observed between 10 and 20 mM NaNO2; greater than 20 mM NaNO2 caused significant (P less than 0.001) diminution of HMS; glucose carbon recycling through the HMS occurred only with greater than 2.5 mM NaNO2 where this process contributed less than or equal to 16% to total HMS activity.     

Microbial tropicalization driven by a strengthening western ocean boundary current

Western boundary currents (WBCs) redistribute heat and oligotrophic seawater from the tropics to temperate latitudes, with several displaying substantial climate change‐driven intensification over the last century. Strengthening WBCs have been implicated in the poleward range expansion of marine macroflora and fauna, however, the impacts on the structure and function of temperate microbial communities are largely unknown. Here we show that the major subtropical WBC of the South Pacific Ocean, the East Australian Current (EAC), transports microbial assemblages that maintain tropical and oligotrophic (k‐strategist) signatures, to seasonally displace more copiotrophic (r‐strategist) temperate microbial populations within temperate latitudes of the Tasman Sea. We identified specific characteristics of EAC microbial assemblages compared with non‐EAC assemblages, including strain transitions within the SAR11 clade, enrichment of Prochlorococcus, predicted smaller genome sizes and shifts in the importance of several functional genes, including those associated with cyanobacterial photosynthesis, secondary metabolism and fatty acid and lipid transport. At a temperate time‐series site in the Tasman Sea, we observed significant reductions in standing stocks of total carbon and chlorophyll a, and a shift towards smaller phytoplankton and carnivorous copepods, associated with the seasonal impact of the EAC microbial assemblage. In light of the substantial shifts in microbial assemblage structure and function associated with the EAC, we conclude that climate‐driven expansions of WBCs will expand the range of tropical oligotrophic microbes, and potentially profoundly impact the trophic status of temperate waters.