Dennis Konnerup

Linking oxygen availability with membrane potential maintenance and K+ retention of barley roots: implications for waterlogging stress tolerance

Oxygen deprivation is a key determinant of root growth and functioning under waterlogging. In this work, changes in net K(+) flux and membrane potential (MP) of root cells were measured from elongation and mature zones of two barley varieties under hypoxia and anoxia conditions in the medium, and as influenced by ability to transport O2 from the shoot. We show that O2 deprivation results in an immediate K(+) loss from roots, in a tissue- and time-specific manner, affecting root K(+) homeostasis. Both anoxia and hypoxia induced transient membrane depolarization; the extent of this depolarization varied depending on severity of O2 stress and was less pronounced in a waterlogging-tolerant variety. Intact roots of barley were capable of maintaining H(+) -pumping activity under hypoxic conditions while disrupting O2 transport from shoot to root resulted in more pronounced membrane depolarization under O2 -limited conditions and in anoxia a rapid loss of the cell viability. It is concluded that the ability of root cells to maintain MP and cytosolic K(+) homeostasis is central to plant performance under waterlogging, and efficient O2 transport from the shoot may enable operation of the plasma membrane H(+) -ATPase in roots even under conditions of severe O2 limitation in the soil solution.

Spatio-temporal relief from hypoxia and production of reactive oxygen species during bud burst in grapevine (Vitis vinifera)

Background and Aims Plants regulate cellular oxygen partial pressures (pO2), together with reduction/oxidation (redox) state in order to manage rapid developmental transitions such as bud burst after a period of quiescence. However, our understanding of pO2 regulation in complex meristematic organs such as buds is incomplete and, in particular, lacks spatial resolution. Methods The gradients in pO2 from the outer scales to the primary meristem complex were measured in grapevine (Vitis vinifera) buds, together with respiratory CO2 production rates and the accumulation of superoxide and hydrogen peroxide, from ecodormancy through the first 72 h preceding bud burst, triggered by the transition from low to ambient temperatures. Key Results Steep internal pO2 gradients were measured in dormant buds with values as low as 2·5 kPa found in the core of the bud prior to bud burst. Respiratory CO2 production rates increased soon after the transition from low to ambient temperatures and the bud tissues gradually became oxygenated in a patterned process. Within 3 h of the transition to ambient temperatures, superoxide accumulation was observed in the cambial meristem, co-localizing with lignified cellulose associated with pro-vascular tissues. Thereafter, superoxide accumulated in other areas subtending the apical meristem complex, in the absence of significant hydrogen peroxide accumulation, except in the cambial meristem. By 72 h, the internal pO2 gradient showed a biphasic profile, where the minimum pO2 was external to the core of the bud complex. Conclusions Spatial and temporal control of the tissue oxygen environment occurs within quiescent buds, and the transition from quiescence to bud burst is accompanied by a regulated relaxation of the hypoxic state and accumulation of reactive oxygen species within the developing cambium and vascular tissues of the heterotrophic grapevine buds.

Responses of rice to Fe2+ in aerated and stagnant conditions: growth, root porosity and radial oxygen loss barrier

Lowland rice (Oryza sativa L.) encounters flooded soils that are anaerobic and chemically reduced. Exposure of the roots to high soil Fe2+ concentrations can result in toxicity. Internal aeration delivering O2 to submerged roots via the aerenchyma is well understood, but the effect of Fe2+ on O2 transport in roots is less studied. We aimed to evaluate the effects of Fe2+ on growth and root aeration. O. sativa var. Amaroo was grown in aerobic and deoxygenated solutions with 0mM, 0.18mM, 0.36mM, 0.54mM or 0.72mM Fe2+ using FeSO4.7H2O and a control with 0.05mM Fe-EDTA. The treatments were imposed on 14-day-old plants (28-30 days old when harvested). Dry mass, shoot Fe concentration, root porosity and patterns of radial O2 loss (ROL) along roots were determined. In the aerobic solution, where Fe2+ was oxidised in the bulk medium, root dry mass increased with higher Fe2+; this was not the case in stagnant solutions, which had no significant root growth response, although Fe oxidation near the root surface was visible as a precipitate. In the highest Fe2+ treatment, shoot Fe concentrations in aerobic (667mgkg-1) and stagnant (433mgkg-1) solutions were below the level for toxicity (700mgkg-1). Rice responded to high Fe2+ in aerobic conditions by increasing root porosity and inducing strong barriers to ROL. In stagnant conditions, root porosity was already high and the ROL barrier induced, so these root aeration traits were not further influenced by the Fe2+ concentrations applied.

Contrasting submergence tolerance in two species of stem-succulent halophytes is not determined by differences in stem internal oxygen dynamics

BACKGROUND AND AIMS Many stem-succulent halophytes experience regular or episodic flooding events, which may compromise gas exchange and reduce survival rates. This study assesses submergence tolerance, gas exchange and tissue oxygen (O2) status of two stem-succulent halophytes with different stem diameters and from different elevations of an inland marsh. METHODS Responses to complete submergence in terms of stem internal O2 dynamics, photosynthesis and respiration were studied for the two halophytic stem-succulents Tecticornia auriculata and T. medusa. Plants were submerged in a glasshouse experiment for 3, 6 and 12 d and O2 levels within stems were measured with microelectrodes. Photosynthesis by stems in air after de-submergence was also measured. KEY RESULTS Tecticornia medusa showed 100 % survival in all submergence durations whereas T. auriculata did not survive longer than 6 d of submergence. O2 profiles and time traces showed that when submerged in water at air-equilibrium, the thicker stems of T. medusa were severely hypoxic (close to anoxic) when in darkness, whereas the smaller diameter stems of T. auriculata were moderately hypoxic. During light periods, underwater photosynthesis increased the internal O2 concentrations in the succulent stems of both species. Stems of T. auriculata temporally retained a gas film when first submerged, whereas T. medusa did not. The lower O2 in T. medusa than in T. auriculata when submerged in darkness was largely attributed to a less permeable epidermis. The submergence sensitivity of T. auriculata was associated with swelling and rupturing of the succulent stem tissues, which did not occur in T. medusa. CONCLUSIONS The higher submergence tolerance of T. medusa was not associated with better internal aeration of stems. Rather, this species has poor internal aeration of the succulent stems due to its less permeable epidermis; the low epidermal permeability might be related to resistance to swelling of succulent stem tissues when submerged.

Leaf gas film retention during submergence of 14 cultivars of wheat (Triticum aestivum)

Flooding of fields after sudden rainfall events can result in crops being completely submerged. Some terrestrial plants, including wheat (Triticum aestivum L.), possess superhydrophobic leaf surfaces that retain a thin gas film when submerged, and the gas films enhance gas exchange with the floodwater. However, the leaves lose their hydrophobicity during submergence, and the gas films subsequently disappear. We tested gas film retention time of 14 different wheat cultivars and found that wheat could retain the gas films for a minimum of 2 days, whereas the wild wetland grass Glyceria fluitans (L.) R.Br. had thicker gas films and could retain its gas films for a minimum of 4 days. Scanning electron microscopy showed that the wheat cultivars and G. fluitans possessed high densities of epicuticular wax platelets, which could explain their superhydrophobicity. However, G. fluitans also had papillae that contributed to higher hydrophobicity during the initial submergence and could explain why G. fluitans retained gas films for a longer period of time. The loss of gas films was associated with the leaves being covered by an unidentified substance. We suggest that leaf gas film is a relevant trait to use as a selection criterion to improve the flood tolerance of crops that become temporarily submerged.

Leaf gas films contribute to rice (Oryza sativa) submergence tolerance during saline floods

Floods and salinization of agricultural land adversely impact global rice production. We investigated whether gas films on leaves of submerged rice delay salt entry during saline submergence. Two-week-old plants with leaf gas films (+GF) or with gas films experimentally removed (-GF) were submerged in artificial floodwater with 0 or 50 mm NaCl for up to 16 d. Gas films were present >9 d on GF plants after which gas films were diminished. Tissue ion analysis (Na+ , Cl- and K+ ) showed that gas films caused some delay of Na+ entry, as leaf Na+ concentration was 36-42% higher in -GF leaves than +GF leaves on days 1-5. However, significant net uptakes of Na+ and Cl- , and K+ net loss, occurred despite the presence of gas films, indicating the likely presence of some leaf-to-floodwater contact, so that the gas layer must not have completely separated the leaf surfaces from the water. Natural loss and removal of gas films resulted in severe declines in growth, underwater photosynthesis, chlorophylla and tissue porosity. Submergence was more detrimental to leaf PN and growth than the additional effect of 50 mm NaCl, as salt did not significantly affect underwater PN at 200 μm CO2 nor growth.

Flood tolerance of wheat – the importance of leaf gas films during complete submergence

Submergence invokes a range of stressors to plants with impeded gas exchange between tissues and floodwater being the greatest challenge. Many terrestrial plants including wheat (Triticum aestivum L.), possess superhydrophobic leaf cuticles that retain a thin gas film when submerged, and the gas films enhance gas exchange with the floodwater. However, leaf hydrophobicity is lost during submergence and the gas films disappear accordingly. Here, we completely submerged wheat (with or without gas films) for up to 14 days and found that plants with gas films survived significantly longer (13 days) than plants without (10 days). Plants with gas films also had less dead tissue following a period of recovery. However, this study also revealed that reflections by gas films resulted in a higher light compensation point for underwater net photosynthesis for leaves with gas films compared with leaves without (IC=52 vs 35µmol photons m-2 s-1 with or without gas films, respectively). Still, already at ~5% of full sunlight the beneficial effect of gas films overcame the negative under ecologically relevant CO2 concentrations. Our study showed that dryland crops also benefit from leaf gas films during submergence and that this trait should be incorporated to improve flood tolerance of wheat.

Evaluation of root porosity and radial oxygen loss of disomic addition lines of Hordeum marinum in wheat

Hordeum marinum Huds. is a waterlogging-tolerant wild relative of wheat (Triticum aestivum L.). Greater root porosity (gas volume per root volume) and formation of a barrier to reduce root radial O2 loss (ROL) contribute to the waterlogging tolerance of H. marinum and these traits are evident in some H. marinum-wheat amphiploids. We evaluated root porosity, ROL patterns and tolerance to hypoxic stagnant conditions for 10 various H. marinum (two accessions) disomic chromosome addition (DA) lines in wheat (two varieties), produced from two H. marinum-wheat amphiploids and their recurrent wheat parents. None of the DA lines had a barrier to ROL or higher root porosity than the wheat parents. Lack of a root ROL barrier in the six DA lines for H. marinum accession H21 in Chinese Spring (CS) wheat indicates that the gene(s) for this trait do not reside on one of these six chromosomes; unfortunately, chromosome 3 of H. marinum has not been isolated in CS. Unlike the H21-CS amphiploid, which formed a partial ROL barrier in roots, the H90-Westonia amphiploid and the four derived DA lines available did not. The unaltered root aeration traits in the available DA lines challenge the strategy of using H. marinum as a donor of these traits to wheat.

Contrasting oxygen dynamics in Limonium narbonense and Sarcocornia fruticosa during partial and complete submergence

Terrestrial saltmarsh plants inhabiting flood-prone habitats undergo recurrent and prolonged flooding driven by tidal regimes. In this study, the role of internal plant aeration in contrasting hypoxic/anoxic conditions during submergence was investigated in the two halophytes Limonium narbonense Mill. and Sarcocornia fruticosa (L.) A.J. Scott. Monitoring of tissue O2 dynamics was performed in shoots and roots using microelectrodes under drained conditions, waterlogging, partial and complete submergence, in light or darkness. For both species, submergence in darkness resulted in significant declines in tissue O2 status and when in light, in rapid O2 increases first in shoot tissues and subsequently in roots. During partial submergence, S. fruticosa benefitted from snorkelling and efficiently transported O2 to roots, whereas the O2 concentration in roots of L. narbonense declined by more than 90%. Significantly thinner leaves and articles were recorded under high degree of flooding stress and both species showed considerably high tissue porosity. The presence of aerenchyma seemed to support internal aeration in S. fruticosa whereas O2 diffusion in L. narbonense seemed impeded, despite the higher porosity (up to 50%). Thus, the results obtained for L. narbonense, being well adapted to flooding, suggests that processes other than internal aeration could be involved in better flooding tolerance e.g. fermentative processes, and that traits resulting in flooding tolerance in plants are not yet fully understood.

Waterlogging tolerance, tissue nitrogen and oxygen transport in the forage legume Melilotus siculus: a comparison of nodulated and nitrate-fed plants

Background and Aims Soil waterlogging adversely impacts most plants. Melilotus siculus is a waterlogging-tolerant annual forage legume, but data were lacking for the effects of root-zone hypoxia on nodulated plants reliant on N2 fixation. The aim was to compare the waterlogging tolerance and physiology of M. siculus reliant on N2 fixation or with access to NO3-. Methods A factorial experiment imposed treatments of water level (drained or waterlogged), rhizobia (nil or inoculated) and mineral N supply (nil or 11 mm NO3-) for 21 d on plants in pots of vermiculite in a glasshouse. Nodulation, shoot and root growth and tissue N were determined. Porosity (gas volume per unit tissue volume) and respiration rates of root tissues and nodules, and O2 microelectrode profiling across nodules, were measured in a second experiment. Key Results Plants inoculated with the appropriate rhizobia, Ensifer (syn. Sinorhizobium) medicae, formed nodules. Nodulated plants grew as well as plants fed NO3-, both in drained and waterlogged conditions. The growth and total N content of nodulated plants (without any NO3- supplied) indicated N2 fixation. Respiration rates (mass basis) were highest in nodules and root tips and lowest in basal root tissues. Secondary aerenchyma (phellem) formed along basal root parts and a thin layer of this porous tissue also covered nodules, which together enhanced gas-phase diffusion of O2 to the nodules; O2 was below detection within the infected zone of the nodule interior. Conclusions Melilotus siculus reliant on N2 fixation grew well both in drained and waterlogged conditions, and had similar tissue N concentrations. In waterlogged conditions the relatively high respiration rates of nodules must rely on O2 movement via the aerenchymatous phellem in hypocotyl, roots and the outer tissue layers of nodules.