Characterization of extracellular oxygen consumption by the green alga Selenastrum minutum

Cells of the green alga Selenastrum minutum display a high capacity for extra-mitochondrial O₂ consumption in the presence of effectors such as salicylhydroxamic acid and/or NADH. We provide evidence that this O₂ consumption is mediated by extracellular peroxidase. Peroxidase capacity, measured as the potential for stimulation of O₂ consumption by a combination of salicylhydroxamic acid and NADH, changed over a 10-day time course. Maximal stimulation of O₂ consumption occurred at day three, at which point the capacity for peroxidase-mediated O₂ consumption was three-to four-fold higher than that of the control O₂ consumption rate. Peroxidase-mediated O₂ consumption was sensitive to inhibition by 50 m M ascorbate and by cyanide. Cyanide titration curves indicated that O₂ consumption by peroxidase was much more sensitive to inhibition by cyanide than was O₂ consumption by cytochrome oxidase (I₅₀ < 1.6 μ M and I₅₀= 18.3 μ M cyanide, respectively). By using evidence from a combination of cyanide titration curves and ascorbate inhibition, we concluded that despite a large capacity for peroxidase-mediated O₂ consumption, peroxidase did not measurably contribute to control rates of O₂ consumption. In the absence of effectors, O₂ consumption was mediated primarily by cytochrome oxidase.     

Storage Polysaccharide in Coccidial Sporozites after Excystation and Penetration of Cells

SYNOPSIS. Eimeria acervulina, E. necatrix , and E. meleagrimitis sporozoites were examined for carbohydrates by cytochemical methods during dormancy, after excystation, and after penetration of cells. The only carbohydrate found was amylopectin, a homogeneous polymer of glucose. It was distributed in 3 regions: (a) in front of the anterior refractile globule, (b) around the nucleus, and (c) behind the posterior refractile globule. The relative amounts decreased after excystation and penetration of cells until only small amounts remained around the nucleus. The quantity of amylopectin decreased following excystation from 30.0-36.7 to 9.4-13.3 μg glucose/10⁶ oocysts. Over a 6 yr period of storage at 4 C, there was a decrease in the quantity of amylopectin in dormant sporozoites of E. acervulina from 33.3 μg glucose/10⁶ oocysts at 3 mos to 1.5 μg at 6 years. Coincidentally, 3 month- and 1 year-old oocysts of E. acervulina produced patent infections in chicks with a dosage of 5 × 10⁴ oocysts, but only a few of the oocysts that had been stored for 2 years were infective; a dosage of 2 × 10⁶ oocysts was necessary to produce a patent infection. Oocysts which had been stored 6 years did not produce a patent infection.
It was concluded that amylopectin is the energy source for excystation and subsequent penetration of cells. Small amounts of amylopectin are used during dormancy and, when the content in the sporozoite falls below a certain level, the sporozoites lack sufficient energy to infect cells.     

Azide-stimulated oxygen consumption by the green alga Selenastrum minutum

Dark O₂ consumption by the green alga Selenastrum minutum was sensitive to inhibition by the cytochrome pathway respiration inhibitor cyanide in the absence of an alternative oxidase inhibitor, consistent with previous work that suggested that this alga lacks alternative oxidase capacity. In contrast, addition of low concentrations of the cytochrome pathway inhibitor azide (50–750 μ M ) resulted in a stimulation of dark O₂ consumption, while higher concentrations of azide (1–2 m M ) partially inhibited O₂ consumption. Measurements of changes in cellular levels of pyruvate, malate and pyridine nucleotides upon cyanide addition were consistent with the absence of alternative oxidase capacity, and suggested that cyanide inhibition of O₂ consumption was not due to nonspecific effects of cyanide. Addition of salicylhydroxamic acid (SHAM) also resulted in an increase in the rate of O₂ consumption. Both azide- and SHAM-stimulated O₂ consumption were sensitive to inhibition by 50 m M ascorbate or by cyanide. However, the ubiquinone analogs chloroquine and quinacrine specifically inhibited azide-stimulated O₂ consumption, with only minor effects on SHAM-stimulated O₂ consumption. These results suggest that azide-stimulated O₂ consumption was not mediated by the previously characterized SHAM-stimulated oxidase, and are consistent with the possibility that azide-stimulated O₂ consumption is mediated by a plasma membrane redox system.     

EXTRACELLULAR PEROXIDASE-MEDIATED OXYGEN CONSUMPTION IN CHLAMYDOMONAS REINHARDTII (CHLOROPHYTA)1

The unicellular green alga Chlamydomonas reinhardtii Dang. displays a high capacity for salicylhydroxamic acid (SHAM)—stimulated O₂ consumption, mediated by extracellular peroxidaie. Addition of exogenous NADH also resulted in stimulation of O₂ consumption. The SHAM-and NADH-stimulated peroxidase activity was partially sensitive to inhibition by exogenous superoxide dismutase, ascorbate, and gentisic acid. These compounds did not inhibit O₂ consumption in the absence of effectors. SHAM-and NADH-stimulated peroxidase activity also was sensitive to inhibition by cyanide, and cyanide titration curves indicated that O₂ consumption by peroxidase was more cyanide-sensitive than O₂ consumption by cytochrome oxidase. The differential sensitivity to cyanide was used to estimate partitioning of O₂ consumption between mitochondrial respiration and extracellular peroxidase. We suggest that, despite a large capacity for peroxidase-me-diated O₂ consumption, peroxidase did not consume O₂ at detectable rates in the absence of effectors. Therefore, in the absence of effectors, measured rates of O₂ consumption represented the rate of mitochondrial respiration .     

Scaling of oxygen consumption of Lake Magadi tilapia, a fish living at 37°C

Rates of oxygen consumption were measured in the geothermal, hot spring fish, Oreochromis alcalicus grahami by stopped flow respirometry. At 37° C, routine oxygen consumption followed the allometric relationship: V o₂=0.738 M ^0.75, where V o₂ is ml O₂ h ⁻¹ and M is body mass (g). This represents a routine metabolic rate for a 10 g fish at 37° C of 0.415 ml O₂ g⁻¹ h ⁻¹ (16.4 μmol O₂ g ⁻¹ h ⁻¹). Acutely increasing the temperature from 37 to 42° C significantly elevated the rate of O₂ consumption from 0.739 to 0.970 ml O₂ g ⁻¹ h ⁻¹ ( Q ₁₀=l.72). In the field, O. a. grahami was observed to be 'gulping' air from the surface of the water especially in hot springs that exceeded 40° C. O. a. grahami may utilize aerial respiration when O₂ requirements are high.     

Hydrogen peroxide release from bacterial spores during germination

The release of free H₂O₂ from spores of Clostridium perfringens and Bacillus megaterium during germination has been demonstrated using the scopoletin fluorescence assay. Scopoletin oxidation was markedly inhibited when exogenous catalase was added, and was also influenced by the concentration of spores. H₂O₂ release into the germination medium was observed to parallel the O₂ consumption during germination, suggesting that the H₂O₂ may arise from certain O₂-dependent metabolism associated with initiation of spore germination.     

OXYGEN CONSUMPTION ASSOCIATED WITH FERRIC REDUCTASE ACTIVITY AND IRON UPTAKE BY IRON-LIMITED CELLS OF CHLORELLA KESSLERI (CHLOROPHYCEAE)

Plasma membrane ferric reductase activity was enhanced 5-fold under iron limitation in the unicellular green alga Chlorella kessleri Fott et Nováková. Furthermore, ferric reductase activity in iron-limited cells was approximately 50% higher in the light than in the dark. In contrast, iron uptake rates of iron-limited cells were unaffected by light versus dark treatments. Rates of iron uptake were much lower than rates of ferric reduction, averaging approximately 2% of the dark ferric reduction rate. Ferric reduction was associated with an increased rate of O₂ consumption in both light and dark, the increase in the light being approximately 1.5 times as large as in the dark. The increased rate of O₂ consumption could be decreased by half by the addition of catalase, indicating that H₂O₂ is the product of the O₂ consumption and that the increased O₂ consumption is nonrespiratory. The stimulation of O₂ consumption was almost completely abolished by the addition of bathophenanthroline disulfonate, a strong chelator of Fe^{2 +} . Anaerobic conditions or the presence of exogenous superoxide dismutase affected neither ferric reduction nor iron uptake. We suggest that the O₂ consumption associated with ferric reductase activity resulted from superoxide formation from the aerobic oxidation of Fe^{2 +} , which is the product of ferric reductase activity. At saturating concentrations of Fe^{3 +} chelates, ferric reductase activity is much greater than the iron uptake rate, leading to rapid oxidation of Fe^{2 +} and superoxide generation. Therefore, O₂ consumption is not an integral part of the iron assimilation process.     

Interactions between respiration and inorganic phosphate uptake in phosphate-limited cells of Chlamydomonas reinhardtii

Phosphate addition to P-limited cells of Chlamydomonas reinhardtii resulted in an immediate increase in the rate of respiratory O₂ consumption. The respiration rate continued to increase for several minutes after the addition of P₁. Similar patterns of P₁ stimulation of respiratory O₂ consumption were observed in the presence of cyanide (cytochrome oxidase inhibitor) and propyl gallate (alternative oxidase inhibitor). Stimulation of O₂ consumption was accompanied by rapid changes in levels of glycolytic intermediates. These changes were consistent with activation of ATP-dependent phosphofructokinase and pyruvate kinase. The adenylate pool exhibited only minor perturbations, P₁, uptake resulted in extracellular acidification, which continued for several minutes after the exhaustion of added P₁, whereas exhaustion of extracellular P₁ resulted in a rapid decline in the O₂ consumption rate. These results are consistent with control of respiration in P-limited cells occurring largely at the level of glycolysis.     

Seasonal differences in routine oxygen consumption rates of the bonnethead shark

Routine oxygen consumption rates of bonnethead sharks, Sphyrna tiburo , increased from 141·3±29·7 mg O₂ kg⁻¹ h⁻¹ during autumn to 218·6±64·2 mg O₂ kg⁻¹ h⁻¹ during spring, and 329·7±38·3 mg O₂ kg⁻¹ h⁻¹ during summer. The rate of routine oxygen consumption increased over the entire seasonal temperature range (20–30° C) at a Q ₁₀=2·34.     

Oxygen uptake in developing eggs and larvae of the cod, Gadus morhua L.

Unfertilised cod eggs showed a mean oxygen uptake rate at 5°C of 0.089 μl O₂, dry wt.⁻¹ h⁻¹; this gradually rose to 0.768 μl O₂ mg dry wt.⁻¹ h⁻¹ in eggs about to hatch. From hatching to complete yolk absorption larvae respired at 1.6 μl O₂, mg dry wt.⁻¹ h⁻¹. During starvation following yolk absorption, uptake fell significantly to 1.1 μl O₂, mg dry ⁻¹ h⁻¹. Much of this decrease in oxygen consumption was shown to be caused by reduction in activity. Loss of weight during the embryo and larval phases could not easily be reconciled with total oxygen consumption; it is suggested that cod embryos and larvae may not rely solely upon endogenous energy reserves during development.     

Oxygen consumption by eggs and larvae of striped mullet, Mugil cephalus, in relation to development, salinity and temperature

Oxygen consumption rates during embryonic and the first 38 days of larval development of the striped mullet were measured at 24° C by differential respirometry. Measurements were obtained at the blastula, gastrula and four embryonic stages, and at the yolk-sac, preflexion, flexion and post-flexion larval stages.
Oxygen uptake rates of eggs increased linearly from 0.024 μl O₂ per egg h^-1 (0·323 μl O₂ mg^-1 dry wt h^-1) by blastulae to 0·177 μlO₂ per egg h^-1 (2·516 μlO₂mg ¹dry wth^-1) by embryos prior to hatching. Respiration rates did not vary significantly among four salinities (20,25, 30, 35%₀).
Larval oxygen consumption increased in a curvilinear manner from 0·243 μl O₂ per larva h^-1 shortly after hatching to 18·880 μl O₂ per larva h^-1 on day 38. Oxygen consumption varied in direct proportion to dry weight. Mass-specific oxygen consumption rates of preflexion, flexion, and postflexion larvae did not change with age (10·838 μl O₂ mg ¹dry wt h^-1).
Larval oxygen consumption rates did not vary significantly among salinities 10–35%. Acute temperature increases elicited significant increases in oxygen consumption, these being relatively greater in yolk-sac larvae ( Q₁₀ = 2·75) than in postflexion larvae ( Q₁₀ = 1·40).     

A method for long-term measurement of respiration in intertidal fishes during simulated intertidal conditions

Aquatic and aerial respiration of the amphibious fishes Lipophrys pholis and Periophthalmus barbarus were examined using a newly designed flow-through respirometer system. The system allowed long-term measurements of oxygen consumption and carbon dioxide release during periods of aquatic and aerial respiration. The M o ₂ of L. pholis , measured at 15° C, was 2·1 μmol O₂ g^–1 h^–1 during aquatic and 1·99 μmol O₂ g^–1 h^–1 during aerial exposure. The corresponding values of the M co₂ were 1.67 and 1.59 μmol O₂ g^–1 h^–1 respectively, giving an aquatic respiratory exchange ratio (RER) of 0·80 and an aerial RER of 0·79. The M o₂ of P. barbarus , measured at 28°C, was 4·05 μmol O₂ g^–1 h^–1 during aquatic and 3·44 μmol O₂ g^–1 h^–1 during aerial exposure. The corresponding values of the Mco₂ were 3·29 μmol CO₂ g^–1 h^–1 and 2·63 μmol CO₂ g^–1 h^–1 respectively, giving an aquatic RER of 0·81 and an aerial RER of 0·77. While exposed to air for at least 10 h, both species showed no decrease in metabolic rate or carbon dioxide release. The RER of these fishes equalled their respiratory quotient. After re-immersion an increased oxygen consumption, due to the payment of an oxygen debt, could not be detected.     

Reduced growth of Atlantic cod in non-lethal hypoxic conditions

Growth in length and mass, improvements in condition, as well as final condition of c. 700 g Atlantic cod Gadus morhua were significantly less at 45% and 56% O₂ saturation than at 65%, 75%, 84% and 93% O₂ saturation. Hypoxia decreased food consumption. In turn, food consumption explained 97% of the variation in growth. Conversion efficiency varied slightly, but significantly, with level of dissolved O₂, except that the group reared at 93% O₂ had a lower than expected conversion efficiency. Slow growth in low O₂ was not due to increased activity, because activity decreased in hypoxia. In the Gulf of St Lawrence, waters deeper than 200 m usually are <65% saturated in O₂, and thus should impact negatively on cod growth.     

The occurrence of aerial respiration in Rhinelepis strigosa during progressive hypoxia

Rhinelepis strigosa did not surface for air breathing in normoxic or moderate hypoxic water. This species initiated air breathing when the P _io₂ in the water reached 22 ± 1 mmHg. Once begun, the air-breathing frequency increased with decreasing P _io₂. Aquatic oxygen consumption was 21·0 ± 1·9ml O₂ kg⁻¹h⁻¹ in normoxic water, and was almost constant during progressive hypoxia until the P _io₂ reached 23·9 mmHg, considered the critical oxygen tension (P_co₂). Gill ventilation increased until close to the P _co₂ (7·9-fold) as a consequence of a greater increase in ventilatory volume than in breathing frequency. Gill oxygen extraction was 42 ± 5% and decreased with hypoxia, but under severe hypoxia returned to values characteristic of normoxic. The critical threshold for air breathing was coincident with the P_co₂ during aquatic respiration. This suggests that the air-breathing response is evoked by the aquatic oxygen tension at which the respiratory mechanisms fail to compensate for environmental hypoxia, and the gill O₂ uptake becomes insufficient to meet O₂ requirements.     

Salt stress induces a decrease in the oxygen uptake of soybean nodules and in their permeability to oxygen diffusion

The effects of short-term NaCl-salinity on nodules of soybean ( Glycine max L. cv. Kingsoy) were studied on hydroponically-grown plants. Both acetylene reducing activity (ARA) and nodule respiration (O₂ uptake and CO₂ evolution) were immediately inhibited, and the stimulation of them by rising the external partial pressure of O₂ (pO₂) was diminished by the application of 0.1 M NaCl in the nutrient solution. The permeability of the nodule to O₂ diffusion, estimated by O₂ consumption or CO₂ evolution, was significantly lower in the stressed nodules than in the cootrol ones. The respiratory quotient of intact nodules and the ethanol production of excised nodules were increased by low pO₂ and by salt stress. These data confirm that in salt-stressed soybean nodules, O₂ availability is reduced and fermentative pathways are stimulated.     

Physiological differences between strains of Tribolium castaneum selected for resistance to hypoxia and hypercarbia, and the unselected strain

Abstract. The metabolic rates, as expressed by oxygen (O₂) consumption, carbon dioxide (CO₂) production, and losses in wet and dry weights, were examined for adults of three strains of the red flour beetle Tribolium castaneum (Herbst), during exposure to two modified atmospheres (MAs). Exposure of a strain selected for resistance over twenty-one generations to an atmosphere of 65% CO₂, 20% O₂ and the balance nitrogen (N₂), termed a high carbon dioxide concentration atmosphere (HCC) and exposure of an unselected strain to HCC, showed considerable levels of aerobic metabolism during exposure. For the unselected strain water loss and mobilization of energy reserves were rapid and mortality was followed by rapid desiccation. For the HCC-resistant strain water balance was maintained and energy reserves were utilized more slowly over a prolonged period. Exposure of a strain selected for resistance over twenty-one generations to a low oxygen concentration atmosphere (LOC) of 0.5% O₂ in N₂, and an unselected strain to LOC, revealed that even at 0.5% O₂, metabolism was largely aerobic in both strains. Maintenance of water balance was not a major factor causing mortality of either strain during exposure to LOC. In air, metabolic rates of both the resistant strains were lower than that of the unselected strain.     

Respiratory responses of Oreochromis niloticus (Pisces, Cichlidae) to environmental hypoxia under different thermal conditions

The oxygen uptake ( V O₂), breathing frequency ( f _R), breath volume ( V _S.R), gill ventilation ( V G) and oxygen extraction (%) from the ventilatory current of four groups of Oreochromis niloticus during graded hypoxia were measured under the following acclimation temperatures: 20. 25. 30 and 35°C. The critical oxygen tensions ( P O₂), determined from V O₂ v. P O₂ of inspired water at each experimental temperature were, respectively. 19±1±3±1. 18±0±4±9, 29±7± 4±1 and 30±2± 0.6 mmHg. The f _R remained nearly constant during the reductions of O₂ at all the temperatures studied. V G increased discretely from normoxic levels until the P O₂ was reached, below which it assumed extremely high values (17-fold higher or more). The increases observed in V G resulted, at all the acclimation temperatures, in an elevation in V _S.R rather than in f _R. The extraction of O₂ decreased gradually from normoxia until the P O₂ was reached, below which an abrupt reduction of extraction was recorded, except at 35°C when fish showed a gradual reduction in extraction just below the tension of 80 mmHg.     

Routine metabolism of juvenile spot, Leiostomus xanthums (Lacépède), as a function of temperature, salinity and weight

Routine oxygen consumption rates of juvenile spot, Leiostomus xanthums , were measured over a range of temperatures, salinities and fish weights. As predicted, Q O₂ increased with temperature and decreased with body weight. However, Q O₂ decreased with decreasing salinity and did not show the expected minimum at isosmotic concentrations. The data are best described by the relationship: log₁₀ Q O₂ (mg O₂ g⁻¹ h⁻¹) = 0.129 log_lo salinity (%0) + 1.604 log₁₀ temperature (°C)-0.1401og₁₀(g)-2.767.     

An Investigation of the Oxidation-reduction Potential and of the Effect of Oxygen on the Germination and Outgrowth of Clostridium butyricum Spores, using Platinum Electrodes

S ummary . The changes in electrode potential recorded by a platinum foil electrode, and in pH, during the incubation of Clostridium butyricum spores in a medium continuously sparged with a sterile gas mixture have been correlated with the biological events occurring simultaneously. The marked fall in E _h which occurs during incubation is recorded by this type of electrode as occurring during the period of rapid cell replication; this observation may be an artefact associated with foil electrodes and the authors feel that results obtained with such electrodes should be interpreted with caution. The stimulating effect of CO₂ and the increasingly inhibitory effect on germination of O₂, as the concentration of the latter in the gas mixture is increased, have been noted. The inhibitory effect is thought to be due to the O₂ itself and not to the high E _h produced in the medium by the O₂.     

Swimming performance, oxygen consumption and excess post-exercise oxygen consumption in adult transgenic and ocean-ranched coho salmon

Routine oxygen consumption ( M o ₂) was 35% higher in 1 day starved and 21% higher in 4 day starved adult transgenic coho salmon Oncorhynchus kisutch relative to end of migration ocean-ranched coho salmon. Critical swimming speed ( U _crit) and M o ₂ at U _crit ( M o _2max) were significantly lower in 4 day starved transgenic coho salmon (1·25 BL s⁻¹; 8·79 mg O₂ kg⁻¹ min⁻¹) compared to ocean-ranched coho salmon (1·60 BL s⁻¹; 9·87 mg O₂ kg⁻¹ min⁻¹). Transgenic fish swam energetically less efficiently than ocean-ranched fish, as indicated by a poorer swimming economy at U _crit ( M o _2max     ). Although M o _2max was lower in transgenic coho salmon, the excess post-exercise oxygen consumption (EPOC) measured during the first 20 min of recovery was significantly larger in transgenic coho salmon (44·1 mg O₂ kg⁻¹) compared with ocean-ranched coho salmon (34·2 mg O₂ kg⁻¹), which had a faster rate of recovery.