Sensitive and Accurate Methodology for Measuring the Kinetics of Concentration-Dependent Hydrocarbon Metabolism Rates in Seawater by Microbial Communities

A method having sufficient sensitivity to resolve the kinetic constants for dissolved nonpolar substrate metabolism, together with the related rate constants in natural waters, is presented. The method is based on the rate of ¹⁴CO₂ recovery from radioactive dissolved substrate. Sensitivity is enhanced by using large seawater volumes, high-specific-activity isotopes, and by reducing background radioactivity. Before use, commercial isotopes are purified by mild alkaline hydrolysis followed by sublimation from base to remove ¹⁴CO₂ as well as interfering polar ¹⁴C-substrates. During sample analysis, chilled Tenax resin is used to remove volatile ¹⁴C-substrate from the nitrogen stream containing ¹⁴CO₂ recovered from substrate oxidation. Chromatographic evidence of purity, shown to be insufficient, is augmented by kinetic data from toluene utilization by mixed cultures and by rates in induced versus noninduced pure cultures. Accuracy is enhanced by using short (<10 h) incubation times and small hydrocarbon concentrations so that the metabolism rates in unamended natural water systems can be evaluated. Toluene metabolism rates in seawater as low as 1 pg/liter per h and at concentrations as low as 20 ng/liter have been determined.     

Dissolved Hydrocarbons and Related Microflora in a Fjordal Seaport: Sources, Sinks, Concentrations, and Kinetics

The continuous addition of toluene as a solute of treated ballast water from oil tankers into a well-defined estuary facilitated the study of the dynamics of dissolved hydrocarbon metabolism in seawater. Most rates of toluene oxidation were in the range of 1 to 30 pg/liter per h at 0.5 μg of toluene per liter. Near the ballast water injection point, a layer of warm ballast water, rich in bacteria, that was trapped below the less-dense fresh surface water was located. Toluene residence times were approximately 2 weeks in this layer, 2 years elsewhere in Port Valdez, and 2 decades in the surface water of a more oceanic receiving estuary adjacent. Mixing was adequate for a steady-state treatment which showed that 98% of the toluene was flushed from Port Valdez before metabolism and gave a steady-state concentration of 0.18 μg/liter. Total bacterial biomass from direct counts and organism size data was usually near 0.1 mg/liter, but ranged up to 0.8 mg/liter in the bacteria-rich layer. The origin of bacteria in this layer was traced to growth in oil tanker ballast during shipments. The biomass of toluene oxidizers in water samples was estimated from the average affinity of pure-culture isolates for toluene (28 liters per g of cells per h) and observed toluene oxidation kinetics. Values ranged from nearly all of the total bacterial biomass within the bacteria-rich layer down to 0.2% at points far removed. Because the population of toluene oxidizers was large with respect to the amount of toluene consumed and because water from a nearby nonpolluted estuary was equally active in facilitating toluene metabolism, we searched for an additional hydrocarbon source. It was found that terpenes could be washed from spruce trees by simulated rainfall, which suggested that riparian conifers provide an additional and significant hydrocarbon source to seawater.     

The intracellular distribution of adenylate kinase in the leaves of spinach,wheat and barley

Of the total adenylate-kinase activity in 10-d-old barley and wheat leaves, 40–50% is localised in the chloroplasts, while in mature spinach leaves 50–70% of the enzyme is chloroplastic. The extra-chloroplastic adenylate-kinase activity is associated with the mitochondria, very little, if any, is freely soluble in the cytoplasm. The adenylate pool of the cytoplasm could have access to adenylate-kinase activity in the intermitochondrial space because of the free permeation of adenylates across the outer mitochondrial membrane. Thus the adenylate pool of the cytoplasm could be subject to adenylate-kinase equilibrium. The mitochondrial adenylate kinase appeared to the localised exclusively in the intermembrane space.     

On describing microbial growth kinetics from continuous culture data: Some general considerations,observations, and concepts

Analysis of continuous culture methodology suggests that this potentially powerful tool for kinetic analysis can be improved by minimizing several inherent shortcomings. Medium background substrates — organic carbon, phosphate, and manganese — were shown to dominate kinetic observations at concentrations below chemical detection methods. Reactor wall growth, culture size distribution changes, sample removal-induced steady state perturbations, and limiting substrate leakage from organisms are treated in terms of kinetic measurement errors. Large variations in maximal growth rates and substrate uptake rates found are attributed to experimental protocol-induced transient states. Relationships are presented for correcting limiting substrate concentrations for lability during sampling, contamination with unreacted medium, and background substrate effects. Analytical procedures are discussed for improved measurement of limiting substrate kinetics involving enzymes, isotopes, and material balance manipulation. Relaxation methods as applied to continuous culture are introduced as a means for isolating separate rate constants describing net substrate transport and for evaluating cellular metabolite leakage. Low velocity growth, multiple substrate metabolism, and endogenous metabolism are discussed along with measurements showing that 1-month generation times for aquatic microorganisms can be quite normal and that the kinetics are compatible withμg/liter limiting substrate concentrations. The concept of regarding growth kinetics as the sum of several net accumulation processes is suggested.     

A global analysis of the determinants of alien geographical range size in birds

Aim Determining the causes of range size variation in the distributions of alien species is important for understanding the spread of invasive species. Factors influencing alien range size have been explored for some species at a regional level, but to date there has been no global analysis of an entire class. Here, we present such an analysis for birds, testing for the effects of introduction event, location and species‐level variables on alien range sizes. Location Global. Methods We used a novel dataset on the global distributions of alien bird species to test for relationships between alien range size and colonization pressure, residence time, extent of the global climatic niche, native range size, body mass and specialization, using a statistical approach based on phylogenetic generalized least squares models. We performed this analysis globally, and for separate biogeographical realms. Results Approximately half of the variation in alien bird range size is explained by colonization pressure in univariate analysis. We identified consistent effects of higher colonization pressure at global and realm levels, as well as support for effects of native range size and residence time. We found less support for effects of body mass, specialization or extent of the global climatic niche on alien range size. Main conclusions Alien bird range sizes are generally small relative to their native range sizes, and many are continuing to expand. Nevertheless, current variation is predictable, most strongly by the event‐level factor of colonization pressure. Whether a species is widespread is a better predictor of alien range size than whether a species could be widespread (estimated by global climatic niche extent), while we also find effects of residence time on alien range size. These relationships may help to identify those alien species that are more likely to spread and hence have greater environmental and economic impacts where they have been introduced.     

Insulin stimulation of GLUT4 exocytosis, but not its inhibition of endocytosis, is dependent on RabGAP AS160

Insulin maintains whole body blood glucose homeostasis, in part, by regulating the amount of the GLUT4 glucose transporter on the cell surface of fat and muscle cells. Insulin induces the redistribution of GLUT4 from intracellular compartments to the plasma membrane, by stimulating a large increase in exocytosis and a smaller inhibition of endocytosis. A considerable amount is known about the molecular events of insulin signaling and the complex itinerary of GLUT4 trafficking, but less is known about how insulin signaling is transmitted to GLUT4 trafficking. Here, we show that the AS160 RabGAP, a substrate of Akt, is required for insulin stimulation of GLUT4 exocytosis. A dominant-inhibitory mutant of AS160 blocks insulin stimulation of exocytosis at a step before the fusion of GLUT4-containing vesicles with the plasma membrane. This mutant, however, does not block insulin-induced inhibition of GLUT4 endocytosis. These data support a model in which insulin signaling to the exocytosis machinery (AS160 dependent) is distinct from its signaling to the internalization machinery (AS160 independent).