Energy Metabolism and Survival of the Infective Juveniles of Steinernema carpocapsae under Oxygen-Deficient Conditions

Energy metabolism and its relation to survival of the infective juveniles (IJ) of S. carpocapsae under anaerobic and oxygen-deficient conditions were studied by monitoring changes in survival rate, levels of key energy reserve materials, oxygen consumption, and respiratory quotient (RQ). The effects of various factors on the survival of IJ under anaerobic conditions were also investigated. Under anaerobic conditions, the IJ were inactivated but could survive for several days in an immobile state, using the carbohydrate reserves glycogen and trehalose for energy supply. The survival time of IJ was mainly dependent on the availability of energy supply, which, in turn, was influenced by factors such as temperature and metabolic by-products. Surviving, anaerobically incubated IJ fully recovered upon return to aerobic conditions. Recovering IJ were characterized by regaining mobility and restoration of carbohydrate reserves consumed during the anaerobic period. Carbohydrate reserves were restored by conversion from lipid reserves and possibly from anaerobic metabolic by-products. The infectivity of IJ recovered from the anaerobic state was not affected. At 1% oxygen level, IJ were also immobile and mainly depended on carbohydrate reserves for energy supply and the RQ was greater than 1. However, some oxygen was consumed; the survival time of these IJ was shorter than those kept in natural air but longer than those under anaerobic conditions. When IJ were incubated at oxygen levels of 3% to 21%, the RQs were maintained at 0.7 to 0.8. Oxygen consumption rates and the reduction in both mean dry weight and lipid levels were proportional to oxygen levels while the survival time of IJ was inversely proportional to oxygen levels.     

Hydroxylamine reduction to ammonium by plant and cyanobacterial hemoglobins

Plants often face hypoxic stress as a result of flooding and waterlogged soils. During these periods, they must continue ATP production and nitrogen metabolism if they are to survive. The normal pathway of reductive nitrogen assimilation in non-legumes, nitrate, and nitrite reductase can be inhibited during low oxygen conditions that are associated with the buildup of toxic metabolites such as nitrite and nitric oxide, so the plant must also have a means of detoxifying these molecules. Compared to animal hemoglobins, plant and cyanobacterial hemoglobins are adept at reducing nitrite to nitric oxide under anaerobic conditions. Here we test their abilities to reduce hydroxylamine, a proposed intermediate of nitrite reductase, under anaerobic conditions. We find that class 1 rice nonsymbiotic hemoglobin (rice nsHb1) and the hemoglobin from the cyanobacterium Synechocystis (SynHb) catalyze the reduction of hydroxylamine to ammonium at rates 100-2500 times faster than animal hemoglobins including myoglobin, neuroglobin, cytoglobin, and blood cell hemoglobin. These results support the hypothesis that plant and cyanobacterial hemoglobins contribute to anaerobic nitrogen metabolism in support of anaerobic respiration and survival during hypoxia.     

Plant anaerobic stress as a novel trend in ecological physiology, biochemistry, and molecular biology: 2. Further development of the problem

This review is a logical development of a previous publication, which summarized the main results of the early period of the systematic and active studying of hypoxic and anoxic stresses in plants. These studies laid a foundation for a new scientific discipline in biology, the investigation relevant to plant anaerobic stress. This review considers a further development of this trend when the investigations embraced a wider set of topics and the discipline acquired an international recognition. The results obtained during last decades by physiologists, biochemists, and molecular biologists engaged in the problem of plant anaerobic stress confirmed the correctness of a concept of the two principal strategies of plant adaptation to hypoxia and anoxia conditions. They are “true” tolerance manifesting at the molecular level under conditions of oxygen deficiency or its absence and “apparent” tolerance, which is realized by avoidance of anaerobiosis due to the long-distance oxygen transport. Therefore, experimental material available now is considered and discussed in this review mainly in the light of these principal notions. Especial attention is paid to the role of stress proteins, which synthesis is induced under hypoxia and anoxia. The results of these experiments confirmed earlier conclusions about the key role of energy (glycolysis and alcoholic fermentation) and carbohydrate (mobilization and utilization of reserved carbohydrates) metabolism in plant adaptation to oxygen deficiency or its absence from the environment. The phenomenon of hypoxic acclimation and its role in plant adaptation to anoxia are also considered. Along with these topics, a further development of pH-stat theory is discussed. A special attention is paid to plant strategy realized by the formation of the net of air-filled spaces (aerenchyma) and long-distance oxygen transport from aerated plant parts to those located in anaerobic environment (apparent tolerance). Among other important aspects, we consider (1) post-anaerobic plant injury by free oxygen radicals; (2) the physiological role of alternative pathways of plant adaptation (nitrate reduction and lipid synthesis); (3) the phenomenon of the adaptation syndrome in plants and possible molecular mechanisms of its realization; and (4) some biotechnological advances in the field of genetic and cell engineering used for the creation of plants more tolerant to anaerobic stress.     

Environmental and functional limits to muscular exercise and body size in marine invertebrate athletes

Many similarities exist between the key characteristics of muscular metabolism in marine invertebrates and those found in vertebrate striated muscle, even though there are important phosphagens and glycolytic end products that differ between groups. Lifestyles and modes of locomotion also vary extremely among invertebrates thereby shaping the pattern of exercise metabolism. In accordance with the limited availability of integrated ecological and physiological information the present paper reports recent progress in the exercise physiology of cephalopods, which are characterized by high rates of aerobic and anaerobic energy turnover during high velocity hunts or escapes in their pelagic environment, and a sipunculid worm, which mostly uses anaerobic resources during extended marathon-like digging excursions in the hypoxic marine sediment. Particular attention is paid to how lifestyle and oxygen availability in various marine environments shapes the use and rates of aerobic and anaerobic metabolism and acidosis as they depend on activity levels and energy saving strategies. Whereas aerobic scope and, accordingly, use of ambient oxygen by blood oxygen transport and skin respiration is maximized in some squids, aerobic scope is very small in the worm and anaerobic metabolism readily used upon muscular activity. Until recently, it was widely accepted that the glycolytic end product octopine, produced in the musculature of these invertebrates, acted as a weak acid and so did not compromise acid-base balance. However, it has now been demonstrated that octopine does cause acidosis. Concomitant study of tissue energy and acid-base status allows to evaluate the contribution of glycolysis, pH and free ADP accumulation to the use of the phosphagen and to the delayed drop in the Gibb's free energy change of ATP hydrolysis. The analysis reveals species specific capacities of these mechanisms to support exercise beyond the anaerobic threshold. During high intensity anaerobic exercise observed in squid, the levels of ATP free energy change finally fall to critical minimum levels contributing to fatigue. Maintenance of sufficiently high energy levels is found at low but extended rates of anaerobic metabolism as observed in the long term digging sipunculid worm. The greatest aerobic and anaerobic performance levels are seen in squid inhabiting the open ocean and appear to be made possible by the uniform and stable physicochemical parameters (esp. high O(2) and low CO(2) levels) that characterize such an environment. It is suggested that at least some squid operate at their functional and environmental limits. In extremely different environments, both the worm and the squids display a tradeoff between oxygen availability, temperature, performance level and also, body size.     

The nature of anaerobic fungi and their polysaccharide degrading enzymes

Conclusion The discovery of anaerobic fungi has added a new member to the indigenous microorganisms that inhabit the rumen ecosystem. Anaerobic fungi do not appear essential for the survival of ruminants due to their presence in very low numbers, and sometimes absence, in ruminants fed low fiber diets, but their presence may likely be very important in the digestion of fibrous diets. The anaerobic fungi have adapted well to the rumen environment. They are able to ferment a large array of soluble carbohydrates and can synthesize cellular components in an anaerobic environment. The fungi posses hydrogenosomes for the removal of reducing equivalents in the form of molecular hydrogen and the removal of trace oxygen is a accomplished via removal by NADH oxidase. Their positive synergistic interaction with methanogenic bacteria eludes to their highly evolved role in the rumen environment. The fungi also produce resistant sporangia that allows for transfer of species to a new host in an oxygen environment. The anaerobic fungi posses a highly active array of polysaccharide degrading enzymes that may provide an advantage in the highly competitive rumen ecosystem. The production of specific enzymes that hydrolyze the lignocellulosic fraction of plant walls is unique in rumen microorganisms and allows for their attachment and growth on fibrous plant particles that are not available to the rumen bacteria.     

Proteins induced by anaerobiosis in Escherichia coli

The contribution of protein induction and repression to the adaptation of cells to changes in oxygen supply is only poorly understood. We assessed this contribution by measuring the levels of 170 individual polypeptides produced by Escherichia coli K-12 in cells growing aerobically or anaerobically with and without nitrate. Eighteen reached their highest levels during anaerobic growth. These 18 polypeptides include at least 4 glycolytic enzymes and pyruvate formate-lyase (beta-subunit). Most of these proteins were found at significant levels during aerobic growth and appeared to undergo metabolic regulation by stimuli other than anaerobiosis. Anaerobic induction ratios ranged from 1.8- to 11-fold, and nitrate antagonized the anaerobic induction of all of the proteins except one. The time course of synthesis of the proteins after shifts in oxygen supply revealed at least three distinct temporal patterns. These results are discussed in light of known physiological alterations associated with changes in oxygen availability.     

A Ratiometric Sensor Based on Plant N-Terminal Degrons Able to Report Oxygen Dynamics in Saccharomyces cerevisiae

The ability to perceive oxygen levels is crucial to many organisms because it allows discerning environments compatible with aerobic or anaerobic metabolism, as well as enabling rapid switch between these two energy strategies. Organisms from different taxa dedicate distinct mechanisms to associate oxygen fluctuations with biological responses. Following from this observation, we speculated that orthogonal oxygen sensing devices can be created by transfer of essential modules from one species to another in which they are not conserved. We expressed plant cysteine oxidase (PCOs) enzymes in Saccharomyces cerevisiae, to confer oxygen-conditional degradability to a bioluminescent protein tagged with the Cys-exposing N-degron typical of plant ERF-VII factors. Co-translation of a second luciferase protein, not subjected to oxygen-dependent proteolysis, made the resulting Double Luciferase Oxygen Reporter (DLOR) ratiometric. We show that DLOR acts as a proxy for oxygen dynamics in yeast cultures. Moreover, since DLOR activity was enabled by the PCO sensors, we employed this device to disclose some of their properties, such as the dispensability of nitric oxide for N-terminal cysteine oxidation and the individual performance of Arabidopsis PCO isoforms in vivo. In the future, we propose the synthetic DLOR device as a convenient, eukaryotic cell-based tool to easily screen substrates and inhibitors of cysteine oxidase enzymes in vivo. Replacement of the luminescent proteins with fluorescent proteins will further turn our system into a visual reporter for oxygen dynamics in living cells.     

Respiratory Development in Saccharomyces cerevisiae Grown at Controlled Oxygen Tension

Saccharomyces cerevisiae was grown in batch culture over a wide range of oxygen concentrations, varying from the anaerobic condition to a maximal dissolved oxygen concentration of 3.5 muM. The development of cells was assayed by measuring amounts of the aerobic cytochromes aa(3), b, c, and c(1), the cellular content of unsaturated fatty acids and ergosterol, and the activity of respiratory enzyme complexes. The half-maximal levels of membrane-bound cytochromes aa(3), b, and c(1), were reached in cells grown in O(2) concentrations around 0.1 muM; this was similar to the oxygen concentration required for half-maximal levels of unsaturated fatty acid and sterol. However, the synthesis of ubiquinone and cytochrome c and the increase in fumarase activity were essentially linear functions of the dissolved oxygen concentration up to 3.5 muM oxygen. The synthesis of the succinate dehydrogenase, succinate cytochrome c reductase, and cytochrome c oxidase complexes showed different responses to changes in O(2) concentration in the growth medium. Cyanide-insensitive respiration and P(450) cytochrome content were maximal at 0.25 muM oxygen and declined in both more anaerobic and aerobic conditions. Cytochrome c peroxidase and catalase activities in cell-free homogenates were high in all but the most strictly anaerobic cells.     

Utilization of aldehydes and alcohols by soybean bacteroids

Aldehydes, alcohols and acids were tested for their ability to support acetylene reduction and oxygen consumption by Rhizobium japonicum bacteroids isolated from soybean nodules. Several alcohols and aldehydes increased acetylene reduction and oxygen uptake. This is consistent with the concept that the plant nodule cytosol can metabolize carbohydrate via anaerobic fermentative pathways.     

Acid- and base-induced proteins during aerobic and anaerobic growth of Escherichia coli revealed by two-dimensional gel electrophoresis

Proteins induced by acid or base, during long-term aerobic or anaerobic growth in complex medium, were identified in Escherichia coli. Two-dimensional gel electrophoresis revealed pH-dependent induction of 18 proteins, nine of which were identified by N-terminal sequencing. At pH 9, tryptophan deaminase (TnaA) was induced to a high level, becoming one of the most abundant proteins observed. TnaA may reverse alkalinization by metabolizing amino acids to produce acidic products. Also induced at high pH, but only in anaerobiosis, was glutamate decarboxylase (GadA). The gad system (GadA/GadBC) neutralizes acidity and enhances survival in extreme acid; its induction during anaerobic growth may help protect alkaline-grown cells from the acidification resulting from anaerobic fermentation. To investigate possible responses to internal acidification, cultures were grown in propionate, a membrane-permeant weak acid which acidifies the cytoplasm. YfiD, a homologue of pyruvate formate lyase, was induced to high levels at pH 4.4 and induced twofold more by propionate at pH 6; both of these conditions cause internal acidification. At neutral or alkaline pH, YfiD was virtually absent. YfiD is therefore a strong candidate for response to internal acidification. Acid or propionate also increased the expression of alkyl hydroperoxide reductase (AhpC) but only during aerobic growth. At neutral or high pH, AhpC showed no significant difference between aerobic and anaerobic growth. The increase of AhpC in acid may help protect the cell from the greater concentrations of oxidizing intermediates at low pH. Isocitrate lyase (AceA) was induced by oxygen across the pH range but showed substantially greater induction in acid or in base than at pH 7. Additional responses observed included the induction of MalE at high pH and induction of several enzymes of sugar metabolism at low pH: the phosphotransferase system components ManX and PtsH and the galactitol fermentation enzyme GatY. Overall, our results indicate complex relationships between pH and oxygen and a novel permeant acid-inducible gene, YfiD.     

The ability of aquatic macrophytes to increase root porosity and radial oxygen loss determines their resistance to sediment anoxia

Radial oxygen loss (ROL) has been suggested to be a major process to protect plants exposed to root anaerobic stress. In the present study, we aimed to test the importance of root porosity and radial oxygen loss on the aquatic macrophyte resistance to sediment anoxia. We expected that species living in eutrophic environments characterized by anaerobic conditions in sediments exhibited higher root porosity and radial oxygen loss than species restrained to oligotrophic environments. In this way, we compared the responses to sediment anoxia of two hydrophyte species growing under meso-eutrophic conditions in the field (Myriophyllum spicatum L. and Vallisneria spiralis L.) and three species growing under oligotrophic conditions (Potamogeton coloratus Horne, Elodea canadensis Michx and Sparganium emersum Michx.). Under laboratory conditions, ROL, root porosity, plant metabolism (aerobic respiration, photosynthesis, root fermentative activity) and plant growth were analysed after 3?months of acclimation in anaerobic sediments and compared with control values obtained from aerobic sediments. The results showed that two meso-eutrophic species (M. spicatum and V. spiralis) survived in anaerobic sediments and maintained similar photosynthesis rates than those measured under aerobic conditions. In contrast, the three oligotrophic species (P. coloratus, E. canadensis and S. emersum) suffered net biomass loss and depressed their photosynthesis rates under anaerobic conditions. All variables associated with plant tolerance to anaerobic conditions (maintenance of photosynthesis, aerobic respiration and growth rate, and limitation of root fermentative activity) were positively linked to root porosity and ROL. According to our hypothesis, species that could survive to anaerobic conditions were the species able to increase their root porosity and ROL under these conditions. Thus, in ecological studies, it would be useful to use the root porosity and ROL plasticity as biological traits in order to model the distribution of macrophytes in river floodplains.     

Response of plant metabolism to too little oxygen

Oxygen can fall to low concentrations within plant tissues, either because of environmental factors that decrease the external oxygen concentration or because the movement of oxygen through the plant tissues cannot keep pace with the rate of oxygen consumption. Recent studies document that plants can decrease their oxygen consumption in response to low oxygen concentrations to avoid internal anoxia. This adaptive response involves a restriction of respiration and a concomitant decrease in ATP consumption that results from the inhibition of a wide range of biosynthetic processes. The inhibition of respiration is rapid and occurs at oxygen concentrations well above the K(m)(oxygen) of cytochrome oxidase, indicating that an oxygen-sensing system triggers a coordinated inhibition of ATP formation and consumption. In addition to this, low oxygen concentrations lead to the induction of a plant-specific and energy-conserving pathway of sucrose degradation, which decreases oxygen consumption and improves plant performance. Low oxygen concentrations also lead to long-term morphological adaptations, which allow respiration per volume tissue to be decreased and oxygen entry to be increased. Recently, advances have been made in elucidating possible oxygen-sensing systems and regulatory components that are involved in these responses.     

Parametric study of diethyl phthalate biodegradation

Summary The effects of temperature, dissolved oxygen, and other environmental parameters under both aerobic and anaerobic conditions were investigated using one aerobic and one facultative strain isolated from wastewater treatment plant sludge. Among other results, we found that low dissolved oxygen levels and low temperatures decreased the rate of DEP degradation and the growth rate, and that the facultative strain was much less affected by the lower DO concentrations than the aerobic strain.     

Aerobic and two-stage anaerobic-aerobic sludge digestion with pure oxygen and air aeration

The degradability of excess activated sludge from a wastewater treatment plant was studied. The objective was establishing the degree of degradation using either air or pure oxygen at different temperatures. Sludge treated with pure oxygen was degraded at temperatures from 22 degrees C to 50 degrees C while samples treated with air were degraded between 32 degrees C and 65 degrees C. Using air, sludge is efficiently degraded at 37 degrees C and at 50-55 degrees C. With oxygen, sludge was most effectively degraded at 38 degrees C or at 25-30 degrees C. Two-stage anaerobic-aerobic processes were studied. The first anaerobic stage was always operated for 5 days HRT, and the second stage involved aeration with pure oxygen and an HRT between 5 and 10 days. Under these conditions, there is 53.5% VSS removal and 55.4% COD degradation at 15 days HRT - 5 days anaerobic, 10 days aerobic. Sludge digested with pure oxygen at 25 degrees C in a batch reactor converted 48% of sludge total Kjeldahl nitrogen to nitrate. Addition of an aerobic stage with pure oxygen aeration to the anaerobic digestion enhances ammonium nitrogen removal. In a two-stage anaerobic-aerobic sludge digestion process within 8 days HRT of the aerobic stage, the removal of ammonium nitrogen was 85%.     

Regulation of plant reactive oxygen species (ROS) in stress responses: learning from AtRBOHD

Reactive oxygen species (ROS) are constantly produced in plants, as the metabolic by-products or as the signaling components in stress responses. High levels of ROS are harmful to plants. In contrast, ROS play important roles in plant physiology, including abiotic and biotic tolerance, development, and cellular signaling. Therefore, ROS production needs to be tightly regulated to balance their function. Respiratory burst oxidase homologue (RBOH) proteins, also known as plant nicotinamide adenine dinucleotide phosphate oxidases, are well studied enzymatic ROS-generating systems in plants. The regulatory mechanisms of RBOH-dependent ROS production in stress responses have been intensively studied. This has greatly advanced our knowledge of the mechanisms that regulate plant ROS production. This review attempts to integrate the regulatory mechanisms of RBOHD-dependent ROS production by discussing the recent advance. AtRBOHD-dependent ROS production could provide a valuable reference for studying ROS production in plant stress responses.     

Inhibition of Programmed Cell Death in Tobacco Plants during a Pathogen-Induced Hypersensitive Response at Low Oxygen Pressure

The hypersensitive response (HR) of plants to invading pathogens is thought to involve a coordinated activation of plant defense mechanisms and programmed cell death (pcd). To date, little is known about the mechanism underlying death of plant cells during this response. In addition, it is not known whether suppression of pcd affects the induction of other defense mechanisms during the HR. Here, we report that death of tobacco cells (genotype NN) infected with tobacco mosaic virus (TMV) is inhibited at low oxygen pressure. In contrast, virus replication and activation of defense mechanisms, as measured by synthesis of the pathogenesis-related protein PR-1a, were not inhibited at low oxygen pressure. Bacterium-induced pcd was also inhibited at low oxygen pressure. However, pcd induced by TMV or bacteria was not inhibited in transgenic tobacco plants expressing the mammalian anti-pcd protein Bcl-XL. Our results suggest that ambient oxygen levels are required for efficient pcd induction during the HR of plants and that activation of defense responses can be uncoupled from cell death. Furthermore, pcd that occurs during the interaction of tobacco with TMV or bacteria may be distinct from some cases of pcd or apoptosis in animals that are insensitive to low oxygen or inhibited by the Bcl-XL protein.