Oxygen Limitation Research Articles

Highly efficient fermentation of 5-keto-d-fructose with Gluconobacter oxydans at different scales

The global market for sweeteners is increasing, and the food industry is constantly looking for new low-caloric sweeteners. The natural sweetener 5-keto-D-fructose is one such candidate. 5-Keto-D-fructose has a similar sweet taste quality as fructose. Developing a highly efficient 5-keto-D-fructose production process is key to being competitive with established sweeteners. Hence, the 5-keto-D-fructose production process was optimised regarding titre, yield, and productivity. For production of 5-keto-D-fructose with G. oxydans 621H ΔhsdR pBBR1-p264-fdhSCL-ST an extended-batch fermentation was conducted. During fructose feeding, a decreasing respiratory activity occurred, despite sufficient carbon supply. Oxygen and second substrate limitation could be excluded as reasons for the decreasing respiration. It was demonstrated that a short period of oxygen limitation has no significant influence on 5-keto-D-fructose production, showing the robustness of this process. Increasing the medium concentration increased initial biomass formation. Applying a fructose feeding solution with a concentration of approx. 1200g/L, a titre of 545g/L 5-keto-D-fructose was reached. The yield was with 0.98g5-keto-d-fructose/gfructose close to the theoretical maximum. A 1200g/L fructose solution has a viscosity of 450mPa∙s at a temperature of 55°C. Hence, the solution itself and the whole peripheral feeding system need to be heated, to apply such a highly concentrated feeding solution. Thermal treatment of highly concentrated fructose solutions led to the formation of 5-hydroxymethylfurfural, which inhibited the 5-keto-D-fructose production. Therefore, fructose solutions were only heated to about 100°C for approx. 10min. An alternative feeding strategy was investigated using solid fructose cubes, reaching the highest productivities above 10 g5-keto-d-fructose/L/h during feeding. Moreover, the scale-up of the 5-keto-D-fructose production to a 150L pressurised fermenter was successfully demonstrated using liquid fructose solutions (745g/L). We optimised the 5-keto-D-fructose production process and successfully increased titre, yield and productivity. By using solid fructose, we presented a second feeding strategy, which can be of great interest for further scale-up experiments. A first scale-up of this process was performed, showing the possibility for an industrial production of 5-keto-D-fructose.

Increased fire activity under high atmospheric oxygen concentrations is compatible with the presence of forests

Throughout Earth’s history, the abundance of oxygen in our atmosphere has varied, but by how much remains debated. Previously, an upper limit for atmospheric oxygen has been bounded by assumptions made regarding the fire window: atmospheric oxygen concentrations higher than 30–40% would threaten the regeneration of forests in the present world. Here we have tested these assumptions by adapting a Dynamic Global Vegetation Model to run over high atmospheric oxygen concentrations. Our results show that whilst global tree cover is significantly reduced under high O2 concentrations, forests persist in the wettest parts of the low and high latitudes and fire is more dependent on fuel moisture than O2 levels. This implies that the effect of fire on suppressing global vegetation under high O2 may be lower than previously assumed and questions our understanding of the mechanisms involved in regulating the abundance of oxygen in our atmosphere, with moisture as a potentially important factor.

Oxygen and irradiance constraints on visual habitat in a changing ocean: The luminoxyscape

Oxygen and irradiance constraints on visual habitat in a changing ocean: The luminoxyscape

Modeling of poly-β-hydroxybutyrate production by Bacillus subtilis and its use for feed-forward bioreactor studies.

Successful scale-up of Bacillus subtilis culture for poly-β-hydroxybutyrate (PHB) production was performed in 5-l stirred-tank reactor using batch, fed-batch, and two-stage culture strategies. The kinetics of biomass production, substrate consumption, and PHB production were established in the stirred tank bioreactor in all the studies. A mathematical model was developed to investigate the role of limiting substrate on overall culture metabolism. A fed-batch strategy was predicted on the basis of computer simulations, for maximum PHB production. This was performed by extrapolation of batch model for predicting the feeding rate and suitable time of feeding. Substrate inhibition was studied and the substrate inhibition terms were incorporated in the model. The maximum cell biomass concentration in batch culture (24h) and fed-batch culture (30h) was 1.79 ± 0.03g/l on dry cell weight (DCW) basis and 1.66 ± 0.050g/l on DCW basis and the corresponding PHB content was 68.71% and 85.54% of DCW, respectively. Glucose was found to be the major limiting nutrient during the bioreactor culture. A two-stage culture, where cells were first grown in stage I in LBG media containing excess carbon and thereafter in stage II in OM media, showed biomass production of 1.95 ± 0.045g/l at 4h and PHB production of 93.33% of DCW at 16h. A 9% increase in growth and 25% increase in PHB yield were obtained using two-stage culture with computer-simulated feeding strategy in the 5l reactor. Oxygen limitation was overcome in modified two-stage culture to obtain a PHB production of 98% at 30h. KEY POINTS: • Polyhydroxybutyrate production was studied in a 5-l stirred-tank bioreactor using HPLC • Mathematical model-assisted fed-batch strategy was implemented in bioreactor • Two-stage fed-batch cultivation was implemented and PHB production was 93% of dry weight in Gram-positive bacteria.

Galactitol Transport Factor GatA Relieves ATP Supply Restriction to Enhance Acid Tolerance of Escherichia coli in the Two-Stage Fermentation Production of D-Lactate

Escherichia coli is a major contributor to the industrial production of organic acids, but its production capacity and cost are limited by its acid sensitivity. Enhancing acid resistance in E. coli is essential for improving cell performance and production value. Here, we propose a feasible strategy for improving cellular acid tolerance by reducing ATP supply restriction. Transcriptome assays of acid-tolerant evolved strains revealed that the galactitol phosphotransferase system transporter protein GatA is an acid-tolerance factor that assists E. coli in improving its resistance to a variety of organic acids. Enhanced GatA expression increased cell survival under conditions of lethal stress due to D-lactic acid, itaconic acid and succinic acid by 101.8-fold, 29.4-fold and 41.6-fold, respectively. In addition, fermentation patterns for aerobic growth and oxygen-limited production of D-lactic acid were identified, and suitable transition and induction stages were evaluated. GatA effectively compensated for the lack of cellular energy during oxygen limitation and enabled the D-lactic acid producing strain to exhibit more sustainable productivity in acidic fermentation environments with a 55.7% increase in D-lactic acid titer from 9.5 g·L−1 to 14.8 g·L−1 and reduced generation of by-product. Thus, this study developed a method to improve the acid resistance of E. coli cells by compensating for the energy gap without affecting normal cell metabolism while reducing the cost of organic acid production.

Enhancement of fatty acid degradation pathway promoted glucoamylase synthesis in Aspergillus niger

BackgroundOur recent multi-omics analyses of glucoamylase biosynthesis in Aspergillus niger (A. niger) suggested that lipid catabolism was significantly up-regulated during high-yield period under oxygen limitation. Since the catabolism of fatty acids can provide energy compounds such as ATP and important precursors such as acetyl-CoA, we speculated that enhancement of this pathway might be beneficial to glucoamylase overproduction.ResultsBased on previous transcriptome data, we selected and individually overexpressed five candidate genes involved in fatty acid degradation under the control of the Tet-on gene switch in A. niger. Overexpression of the fadE, fadA and cyp genes increased the final specific enzyme activity and total secreted protein on shake flask by 21.3 ~ 31.3% and 16.0 ~ 24.2%, respectively. And a better inducible effect by doxycycline was obtained from early logarithmic growth phase (18 h) than stationary phase (42 h). Similar with flask-level results, the glucoamylase content and total extracellular protein in engineered strains OE-fadE (overexpressing fadE) and OE-fadA (overexpressing fadA) on maltose-limited chemostat cultivation were improved by 31.2 ~ 34.1% and 35.1 ~ 38.8% compared to parental strain B36. Meanwhile, intracellular free fatty acids were correspondingly decreased by 41.6 ~ 44.6%. The metabolomic analysis demonstrated intracellular amino acids pools increased 24.86% and 18.49% in two engineered strains OE-fadE and OE-fadA compared to B36. Flux simulation revealed that increased ATP, acetyl-CoA and NADH was supplied into TCA cycle to improve amino acids synthesis for glucoamylase overproduction.ConclusionThis study suggested for the first time that glucoamylase production was significantly improved in A. niger by overexpression of genes fadE and fadA involved in fatty acids degradation pathway. Harnessing the intracellular fatty acids could be a strategy to improve enzyme production in Aspergillus niger cell factory.

Photoinduced Carbene for Effective Photodynamic Therapy Against Hypoxic Cancer Cells

Photoinduced Carbene for Effective Photodynamic Therapy Against Hypoxic Cancer Cells

The effects of different temperature conditions on sludge characteristics and microbial communities of nitritation denitrification

The effects of different temperature conditions on nitritation denitrification are still unclear. In this study, two sequencing batch reactors (SBR) were used to explore the effects of stepwise cooling (28.2, 24.5, 20.6, 17.0 °C) and continuous low temperature (17.0 °C) on partial nitrification denitrification (PND). The PND performance, sludge characteristics, and microbial communities were evaluated. The results showed that PND completed start-up faster and achieved >80 % nitrite accumulation under stepwise cooling conditions, while a longer acclimation period was required under continuous low temperature conditions. Higher dissolved oxygen limits should be used for real-time control at low temperatures to achieve complete ammonia removal. Fitting using Arrhenius equation, it was found that PND was not significantly affected by temperature at 17–28 °C. However, the denitrification activity was significantly inhibited by temperatures below 17 °C. Bacteroidota phylum proliferated at low temperatures, while DGAOs proliferated only under stepwise cooling conditions, and they may contribute to efficient denitrification. Sludge bulking under sustained low temperature conditions eased with increasing PND activity, suggesting that sludge settling performance needs attention when activity is low.

Fast neutron induced gamma rays from (n,n’), (n,p) and (n,α) reactions on CaCO3

Emission of prompt gamma rays following (n,n’), (n,p) and (n,α) reactions induced by irradiation of a calcium carbonate (CaCO3) sample with a beam of fission neutrons was investigated with a modified version of the FaNGaS (Fast Neutron-induced Gamma-ray Spectrometry) instrument operated at the Heinz Maier-Leibnitz Zentrum (MLZ) in Garching. The neutron spectrum has an average energy of 2.30 MeV and at sample position the fast neutron flux was (1.13 ± 0.04) × 108 cm−2 s−1. The measurement was performed at an angle of 90° between neutron beam and detector. In total, we claim the identification of 38 prompt gamma lines, from which we have assigned 12 to the (n,n’) reaction in calcium (7 for 40Ca, 1 for 42Ca and 4 for 44Ca), 1 to the 12C(n,n’)12C reaction, and 2 to the (n,n’) reaction in oxygen (1 for 16O and 1 for 18O). The other observed gamma lines are attributed to the 40Ca(n,p)40K reaction (20 lines) and to the 40Ca(n,α)37Ar reaction (3 lines). Relative intensities and fast neutron spectrum averaged partial cross sections of the gamma lines are presented and compared with available literature data. Our results were found to be in good agreement with available literature data but also improved and complemented it, as we have pointed out possible errors and identified new lines. Additionally, for a counting time of 12 h the detection limits of calcium, carbon and oxygen were determined as 5, 13 and 64 mg, respectively.

Impacts of nighttime hypoxia on the physiological performance of Red Sea macroalgae under peak summer temperature

Eutrophication-induced hypoxic sites are increasingly reported in coastal regions. At the same time, ocean warming, water column stratification, and changing circulation lead to open-ocean deoxygenation. In coastal areas and reefs with dense vegetation, aquatic organisms can be exposed to oxygen limitation stress where oxygen concentration reaches extremely low levels, particularly during nighttime once photosynthetic O2 production has ceased. Despite scientists being aware of this for decades, little is known about the impact of deoxygenation on the physiology of marine primary producers, such as macroalgae. In the Red Sea, in particular, the physiological adaptations of macroalgae under future climate scenarios are nonexistent. Here, we investigate the impact of different oxygen levels (6.5, 2.5, and 1.3 mg O2 L-1) at night for three conspicuous Red Sea macroalgae species Halimeda opuntia and Padina boryana (calcareous) and the brown algae Sargassum latifolium (noncalcifying). We monitored algal physiological responses during a 12-hour nighttime (dark) period at 32°C by measuring photochemical efficiency (Fv/Fm), respiration rates, and cellular viability. No lethal thresholds were detected. However, both deoxygenation treatments decreased respiration rates and induced changes in cellular activity, and only under severe hypoxia was a decrease in photochemical efficiency observed in all species. We calculated sublethal O2 thresholds SLC(50) of 1.2 ± 0.1, 1.5 ± 0.1, and 1.7 ± 0.1 mg O2 L-1 for H. opuntia, P. boryana, and S. latifolium, respectively. Therefore, the effects of nighttime hypoxia are evident over short timescales and may impact ecosystems via reduced primary production. Future consequences of persistent hypoxia and subsequent performance in multifaceted stressor exposures will provide a fundamental understanding of hypoxia’s threat to biodiversity and ecosystems.

Evaluating cardiac oxygen limitation as a mechanism for female-biased mortality in coho salmon (Oncorhynchus kisutch)

Female-biased mortality has been consistently reported in Pacific salmon during their adult upriver migration. We collected coho salmon ( Oncorhynchus kisutch (Walbaum, 1792)) upon arrival at their spawning grounds to test whether females are more prone to cardiac oxygen limitations following exercise stress. We used a surgical approach to periodically sample arterial and venous blood over 48 h following recovery from a chase protocol to induce maximum metabolic rate. We found no significant differences in arterial or venous partial pressures of O2 between males and females. Female salmon had significantly elevated plasma cortisol levels but there were no effects of sex on either plasma lactate or K+. Our data show that female coho salmon do not suffer oxygen limitations to the spongy myocardium after a single exercise event at moderate temperatures (14 °C)—at least not when arriving to their spawning grounds. This study found no clear support for a cardiac oxygen limitation underlying elevated female mortality in Pacific salmon. Neither, however, does our study design nor specific findings allow us to rule out cardiac limitations in these fish. Future work should address whether potential oxygen limitations to the spongy myocardium at high temperatures or oxygen limitations to the compact myocardium via coronary blood flow contribute to female-biased mortality earlier on the migratory route.

Long term analysis of N2O emission from partial nitritation anammox process under oxygen limitation

Partial nitritation with anammox (PNA) process was achieved during 300 days to investigate the fate of N2O and NO. N2O emission factor increased from 0.4 ± 0.1 % up to 6.1 ± 0.3 % during intensification phase of 5 months and progressively stabilized at 2.0 ± 0.1 % after 10 months without any significant emission of NO. The emission of N2O correlated with the ammonium consumption rate and the oxygen transfer rate. Experiment carried out in anoxic condition suggested that heterotrophic denitrification poorly contributed. Measurements of δ15N, δ18O and SP of N2O demonstrated that its production resulted from the reduction of nitrite. The stimulating effect of nitrite on N2O production was demonstrated with specific tests and observed during transient nitrite accumulation periods. This study highlighted the importance of nitrite and oxygen transfer rate control for future control strategies aiming to mitigate N2O emission in PNA system.

Unique thermal sensitivity imposes a cold-water energetic barrier for vertical migrators

Alterations of marine species’ ranges with climate change are often attributed to oxygen limitation in warming oceans. Here we report unique metabolic temperature sensitivities for the myriad of vertically migrating oceanic species that daily cross depth-related gradients in temperature and oxygen. In these taxa, selection favours high metabolic activity for predator–prey interactions in warm shallow water and hypoxia tolerance in the cold at depth. These diverging selective pressures result in thermal insensitivity of oxygen supply capacity and enhanced thermal sensitivity of active metabolic rate. Aerobic scope is diminished in the cold, well beyond thermodynamic influences and regardless of ambient oxygen levels, explaining the native distributions of tropical migrators and their recent range expansions following warming events. Cold waters currently constitute an energetic barrier to latitudinal range expansion in vertical migrators. As warming due to climate change approaches, and eventually surpasses, temperatures seen during past warming events, this energetic barrier will be relieved.

Controversial Roles of Oxygen in Organismal Responses to Climate Warming.

AbstractDespite the global ecological importance of climate change, controversy surrounds how oxygen affects the fate of aquatic ectotherms under warming. Disagreements extend to the nature of oxygen bioavailability and whether oxygen usually limits growth under warming, explaining smaller adult size. These controversies affect two influential hypotheses: gill oxygen limitation and oxygen- and capacity-limited thermal tolerance. Here, we promote deeper integration of physiological and evolutionary mechanisms. We first clarify the nature of oxygen bioavailability in water, developing a new mass-transfer model that can be adapted to compare warming impacts on organisms with different respiratory systems and flow regimes. By distinguishing aerobic energy costs of moving oxygen from environment to tissues from costs of all other functions, we predict a decline in energy-dependent fitness during hypoxia despite approximately constant total metabolic rate before reaching critically low environmental oxygen. A new measure of oxygen bioavailability that keeps costs of generating water convection constant predicts a higher thermal sensitivity of oxygen uptake in an amphipod model than do previous oxygen supply indices. More importantly, by incorporating size- and temperature-dependent costs of generating water flow, we propose that oxygen limitation at different body sizes and temperatures can be modeled mechanistically. We then report little evidence for oxygen limitation of growth and adult size under benign warming. Yet occasional oxygen limitation, we argue, may, along with other selective pressures, help maintain adaptive plastic responses to warming. Finally, we discuss how to overcome flaws in a commonly used growth model that undermine predictions of warming impacts.

Integrative Approaches to Understanding Organismal Responses to Aquatic Deoxygenation.

AbstractOxygen bioavailability is declining in aquatic systems worldwide as a result of climate change and other anthropogenic stressors. For aquatic organisms, the consequences are poorly known but are likely to reflect both direct effects of declining oxygen bioavailability and interactions between oxygen and other stressors, including two-warming and acidification-that have received substantial attention in recent decades and that typically accompany oxygen changes. Drawing on the collected papers in this symposium volume ("An Oxygen Perspective on Climate Change"), we outline the causes and consequences of declining oxygen bioavailability. First, we discuss the scope of natural and predicted anthropogenic changes in aquatic oxygen levels. Although modern organisms are the result of long evolutionary histories during which they were exposed to natural oxygen regimes, anthropogenic change is now exposing them to more extreme conditions and novel combinations of low oxygen with other stressors. Second, we identify behavioral and physiological mechanisms that underlie the interactive effects of oxygen with other stressors, and we assess the range of potential organismal responses to oxygen limitation that occur across levels of biological organization and over multiple timescales. We argue that metabolism and energetics provide a powerful and unifying framework for understanding organism-oxygen interactions. Third, we conclude by outlining a set of approaches for maximizing the effectiveness of future work, including focusing on long-term experiments using biologically realistic variation in experimental factors and taking truly cross-disciplinary and integrative approaches to understanding and predicting future effects.

Compound Extreme Events Induce Rapid Mortality in a Tropical Sea Urchin.

AbstractThe frequency, magnitude, and duration of marine heatwaves and deoxygenation events are increasing globally. Recent research suggests that their co-occurrence is more common than previously thought and that their combination can have rapid, dire biological impacts. We used the sea urchin Echinometra lucunter to determine whether mortality occurs faster when deoxygenation events are combined with extreme heating (compound events), compared to deoxygenation events alone. We also tested whether prior exposure to local heatwave conditions accentuates the impacts of compound events. Animals were first exposed for five days to either ambient temperature (28 °C) or a warmer temperature that met the minimum criteria for a local heatwave (30.5 °C). Animals were then exposed to hypoxia, defined as oxygen levels 35% below their average critical oxygen limit, combined with ambient or extreme field temperatures (28 °C, 32 °C). Subsets of animals were removed from the hypoxic treatments every 3 hours for 24 hours to determine how long they could survive. Prior exposure to heatwave conditions did not help or hinder survival under hypoxic conditions, and animals exposed to hypoxia under ambient temperatures experienced little mortality. However, when hypoxia was coupled with extreme temperatures (32 °C), 55% of the animals died within 24 hours. On the reefs at our Panama study site, we found that extreme hypoxic conditions only ever occurred during marine heatwave events, with four compound events occurring in 2018. These results show that short durations (∼1 day) of compound events can be catastrophic and that increases in their duration will severely threaten sea urchin populations.

Chronic experimental hyperoxia elevates aerobic scope: a valid method to test for physiological oxygen limitations in fish.

Experimental hyperoxia has been shown to enhance the maximum oxygen uptake capacity of fishes under acute conditions, potentially offering an avenue to test prominent physiological hypotheses attempting to explain impacts of climate warming on fish populations (e.g., gill-oxygen limitation driving declines in fish size). Such benefits of experimental hyperoxia must persist under chronic conditions if it is to provide a valid manipulation to test the relevant hypotheses, yet the long-term benefits of experimental hyperoxia to oxygen uptake capacity have not been examined. Here, the authors measured aerobic metabolic performance of Galaxias maculatus upon acute exposure to hyperoxia (150% air saturation) and after 5months of acclimation, at both 15°C and 20°C. Acute hyperoxia elevated aerobic scope by 74%-94% relative to normoxic controls, and an elevation of 58%-73% persisted after 5months of hyperoxia acclimation. When hyperoxia-acclimated fish were acutely transitioned back to normoxia, they maintained superior aerobic performance compared with normoxic controls, suggesting an acclimation of the underlying metabolic structures/processes. In demonstrating the long-term benefits of experimental hyperoxia on the aerobic performance of a fish, the authors encourage the use of such approaches to disentangle the role of oxygen in driving the responses of fish populations to climate warming.

The AAA+ ATPase RavA and its binding partner ViaA modulate E. coli aminoglycoside sensitivity through interaction with the inner membrane

Enteric bacteria have to adapt to environmental stresses in the human gastrointestinal tract such as acid and nutrient stress, oxygen limitation and exposure to antibiotics. Membrane lipid composition has recently emerged as a key factor for stress adaptation. The E. coli ravA-viaA operon is essential for aminoglycoside bactericidal activity under anaerobiosis but its mechanism of action is unclear. Here we characterise the VWA domain-protein ViaA and its interaction with the AAA+ ATPase RavA, and find that both proteins localise at the inner cell membrane. We demonstrate that RavA and ViaA target specific phospholipids and subsequently identify their lipid-binding sites. We further show that mutations abolishing interaction with lipids restore induced changes in cell membrane morphology and lipid composition. Finally we reveal that these mutations render E. coli gentamicin-resistant under fumarate respiration conditions. Our work thus uncovers a ravA-viaA-based pathway which is mobilised in response to aminoglycosides under anaerobiosis and engaged in cell membrane regulation.

Brain dysfunction during warming is linked to oxygen limitation in larval zebrafish

Understanding the physiological mechanisms that limit animal thermal tolerance is crucial in predicting how animals will respond to increasingly severe heat waves. Despite their importance for understanding climate change impacts, these mechanisms underlying the upper thermal tolerance limits of animals are largely unknown. It has been hypothesized that the upper thermal tolerance in fish is limited by the thermal tolerance of the brain and is ultimately caused by a global brain depolarization. In this study, we developed methods for measuring the upper thermal limit (CTmax) in larval zebrafish (Danio rerio) with simultaneous recordings of brain activity using GCaMP6s calcium imaging in both free-swimming and agar-embedded fish. We discovered that during warming, CTmax precedes, and is therefore not caused by, a global brain depolarization. Instead, the CTmax coincides with a decline in spontaneous neural activity and a loss of neural response to visual stimuli. By manipulating water oxygen levels both up and down, we found that oxygen availability during heating affects locomotor-related neural activity, the neural response to visual stimuli, and CTmax. Our results suggest that the mechanism limiting the upper thermal tolerance in zebrafish larvae is insufficient oxygen availability causing impaired brain function.

Effects of Ion-Transporting Proteins on the Digestive System Under Hypoxia.

Hypoxia refers to a state of oxygen limitation, which mainly mediates pathological processes in the human body and participates in the regulation of normal physiological processes. In the hypoxic environment, the main regulator of human body homeostasis is the hypoxia-inducible factor family (HIF). HIF can regulate the expression of many hypoxia-induced genes and then participate in various physiological and pathological processes of the human body. Ion-transporting proteins are extremely important types of proteins. Ion-transporting proteins are distributed on cell membranes or organelles and strictly control the inflow or outflow of ions in cells or organelles. Changes in ions in cells are often closely related to extensive physiological and pathological processes in the human body. Numerous studies have confirmed that hypoxia and its regulatory factors can regulate the transcription and expression of ion-transporting protein-related genes. Under hypoxic stress, the regulation and interaction of ion-transporting proteins by hypoxia often leads to diseases of various human systems and even tumors. Using ion-transporting proteins and hypoxia as targets to explore the mechanism of digestive system diseases and targeted therapy is expected to become a new breakthrough point.