Mechanism of endocarp-imposed constraints of germination of Lannea microcarpa seeds

Lannea microcarpa, a multipurpose tree species from the dry African savanna, sheds seeds that often display inhibition of germination. The underlying mechanism was investigated using seeds processed from fully matured fruits collected from natural stands in Burkina Faso. Germination of fresh seeds was variable (16¿28%), while they did not germinate after drying and rehydration. Mechanical scarification of the endocarp at the proximal end of the seeds increased germination to 83¿94%. Scarification on the distal end led to delayed radicle emergence through the produced hole in c. 40% of the seeds. The endocarp was permeable to water and respiratory gases. Increased water content in scarified seeds was associated with radicle extension during germination. Intact and scarified non-germinated seeds displayed a moderate rate of respiration with respiratory quotient (RQ) values of c. 1. Respiration increased and RQ decreased to c. 0.7 with radicle emergence. Ethylene evolution peaked in both intact and scarified seeds at the beginning of incubation and then decreased to low values. Inhibition of ethylene production by 1¿5 mM 2-amino-ethoxyvinylglycine (AVG) caused only a partial decrease of germination of the scarified seeds. Intact non-germinated seeds gradually lost viability during incubation at 30°C, but could be rescued by delayed scarification before day 15 of incubation. It is concluded that radicle emergence in dry L. microcarpa seeds is inhibited only mechanically. The mechanical properties of the endocarp are attributed to irreversible structural changes of the lignin¿hemicellulose complex, which occur during drying

Subtidal water level variation controlled by river flow and tides

Subtidal water level dynamics in the Berau river, East Kalimantan, Indonesia, feature a pronounced fortnightly variation. The daily mean water levels at a station about 60 km from the sea are 0.2–0.6 m higher during spring tide than during neap tide. To explain the underlying mechanisms, a local subtidal momentum balance is set up from field data, using continuous discharge estimates inferred from measurements taken with a horizontal acoustic Doppler current profiler. It is demonstrated that terms accounting for friction and variation in the water surface gradient are dominant in the subtidal momentum balance. To further investigate the sources of subtidal water level variation, a generic method of analysis is proposed to decompose the subtidal friction term into contributions caused by river flow, by interaction between tidal motions and river flow, and by the tidal motions alone. At the station under study, mainly the river-tide interaction term is responsible for generating fortnightly variation of the subtidal water level. The contribution from interaction between diurnal, semidiurnal, and quarterdiurnal tides to subtidal friction is significantly smaller. Provided that the reduction of tidal velocity amplitudes with increasing discharges can be predicted from a regression model, the results presented herein can be used to predict changes in subtidal water levels as a result of increased river discharges

Vitality and metabolic properties of binucleate and trinucleate pollen species upon dehiscence

Chapter 1Effects of various components upon germination in vitro were studied in order to develop an optimal germination medium for Compositae pollen. Equilibration of pollen in humid air, preceding germination, improved the reliability of results considerably.Irregular germination ability of pollen samples, originating from different collections, was studied by exposing flowering plants to different climatic conditions. High relative humidity and temperature at dehiscence cause a rapid decrease in pollen vitality. Data for an optimal germination medium and for acquisition of good pollen quality are presented.Chapter 2The respiration and vitality of ungerminated bi- and trinucleate pollen were studied in order to determine the influence of relative humidity and temperature on metabolic activity. The gas exchange, gerniination capacity and staining with tetrazolium bromide were followed under standardized conditions.A constant respiration rate occurred under conditions of high relative humidity (97 %). Per mg pollen, the trinucleate grains of Compositae and Gramineae respired 2 to 3 times as intense as 6 species of binucleate grains. Per unit of pollen protein the differences were even larger. In contrast to binucleate pollen, the longevity of trinucleate pollen was very short and the ability to germinate was lost twice as fast as the respiration capacity. This limits the use of tetrazolium bromide as an indicator of viability.At reduced relative humidities respiration was strongly restricted, but the longevity of bi- and trinucleate pollen considerably increased.Pollen of Gramineae, however, was very sensitive to changes in relative humidity; short exposure to low relative humidity decreased both the vitality and the capacity to respire.Chapter 3Bi- and trinucleate pollen generally differ in the extent of their mitochondrial development at anther dehiscence and in the rate of their attainment of maximum-phosphorylative capacity during germination in vitro, as judged from experiments with representatives of both groups.The typically trinucleate pollen of Aster tripolium L. immediately respired at a high rate, maintaining a high energy charge. Mitochondria attained maximum electron-transducing capacity within 2 min of incubation, while tube growth started within 3 min. In contrast, the binucleate pollen of Typha latifolia L. only gradually reached a relatively low rate of respiration, concomitant with a temporary decrease in energy charge, upon immersion in the germination medium. Development of the mitochondrial, electron-transducing system occurred in about 75 min, after which the first pollen tubes emerged. Starting from a poor differentiation, mitochondria became increasingly normal in appearance as germination proceeded.The binucleate pollen of Nicotiana alata Link et Otto and Tradescantia paludosa Anders. et Woods. showed intermediate characteristics: Nicotiana resembled Typha but mitochondria developed at a higher rate; Tradescantia germinated more rapidly and resembled the trinucleate pollen of Aster.Inhibitors of mitochondrial or cytoplasmic protein synthesis failed to affect the development of the mitochondrial, respiratory capacities during pollen germination. It is concluded that the duration of the lag period is determined by the level and rate of mitochondrial development and not by the division of the generative cell.Chapter 4The equal rates of water vapour absorption by both bi- and trinucleate pollen indicate that their widely-differing rates of respiration have an intrinsic, biochemical basis. This was investigated with various metabolic inhibitors that were previously introduced into dry pollen via anhydrous acetoneThe uncoupler, CCCP, inhibited the O 2 uptake of rapidly respiring pollen and stimulated that of slowly respiring types to similar absolute values, that probably reflect the rates of substrate transport across the mitochondrial membranes.The extent of inhibition of the O 2 uptake by oligomycin, DCCD, antimycin A, and SHAM, alone and in combinations, indicates that hardly any oxidative phosphorylation and anabolic activities occur in slowly respiring, binucleate pollen species, having low-developed mitochondria and high EC values. The presence of the alternative pathway was insignificant.In other binucleate pollen species, characterized by recognizable mitochondria and low EC values, a limited ATP synthesis was established. The low EC values point to imbalance between phosphorylative and anabolic activities.In rapidly respiring, trinucleate pollen, containing well- developed mitochondria, a significant activity of the alternative oxidase was found. The EC values were high notwithstanding the large demand for ATP, mounting to 1.7 μmol h -1mg pollen -1.In some pollen species, oligomycin highly stimulated the flow of electrons through the cytochrome pathway, which made an estimation of the ATP synthesis impossible.Chapter 5Under humid conditions both bi- and trinucleate pollen species incorporate very low amounts of leucine, 0.4 pmol min -1mg pollen -1on an average. During germination in vitro , however, the two types of pollen greatly differ in their capacity for protein synthesis.Binucleate pollen species such as Typha , which are characterized by slow respiration in humid air and prolonged lag periods during germination in vitro , contain large amounts of monoribosomes at dehiscence. Polyribosomes are formed soon after the pollen is wetted in the germination medium and a considerable incorporation of leucine is initiated after 10-15 min.More rapidly respiring, binucleate pollen, such as Tradescantia , showing a short lag period, may contain many polysomes at dehiscence already and incorporates leucine within 2 min of incubation.On the contrary, rapidly respiring, trinucleate Compositae pollen contains very limited amounts of ribosomal material and never attains any substantial level of incorporation.Cycloheximide completely inhibited both protein synthesis and tube emergence and growth in the slowly respiring binucleate pollen species. The more rapidly respiring types are less dependent on protein synthesis, while germination of the phylogenetically advanced, trinucleate Compositae pollen proceeds completely independently.It is concluded that the level of phylogenetic advancement of the male gametophyte is characterized by its overall state of metabolic development at dehiscence rather than by the number of its generative cells.</TT

Ageing increases the sensitivity of neem (Azadirachta indica) seeds to imbibitional stress

Imbibitional stress was imposed on neem (Azadirachta indica) seeds by letting them soak for 1 h in water at unfavourable, low temperatures before further incubation at 30degreesC. Sensitivity to low imbibition temperatures increased with a decrease in seed moisture content (MC). To investigate a possible involvement of seed age in the extent of imbibitional damage, initially high-quality seed lots that differed in storage history (10 weeks versus 10 months) were examined at 4 and 7% (fresh weight basis). After 10 months of storage, the 7% MC seeds had become sensitive to imbibitional stress. Further drying (1 week) to 4% MC affected aged seeds more than non-aged seeds. Barrier properties of cellular membranes in axes excised after 1 d of rehydration were estimated using a spin-probe technique. The proportion of cells with intact membranes increased with increasing imbibition temperature. For each temperature tested, there were more cells with leaky membranes after 10 months than after 10 weeks of dry storage. Localization of embryo cells displaying loss of turgor and abnormal cellular structure was accomplished using cryo-planing, followed by cryo-scanning electron microscopy. Inspection of the cryo-planed surfaces confirmed that imbibitional damage was temperature dependent, occurring at the periphery. Ageing increased the number of imbibitionally damaged, peripheral cell layers. Germination was estimated to fail when less than 70% of axis cells were alive. We conclude that ageing increases the sensitivity to imbibitional stress. Both the fast ageing and the sensitivity to imbibitional stress might explain the apparent controversies about neem seed desiccation tolerance and storage behaviour

Heterosis in the freezing tolerance, and sugar and flavonoid contents of crosses between Arabidopsis thaliana accessions of widely varying freezing tolerance

Heterosis is defined as the increased vigour of hybrids in comparison to their parents. We investigated 24 F1 hybrid lines of Arabidopsis thaliana generated by reciprocally crossing either C24 or Col with six other parental accessions (Can, Co, Cvi, Ler, Rsch, Te) that differ widely in their freezing tolerance. The crosses differed in the degree of heterosis for freezing tolerance, both in the non-acclimated state and after a 14 d cold acclimation period. Crosses with C24 showed more heterosis than crosses with Col, and heterosis was stronger in acclimated than in non-acclimated plants. Leaf content of soluble sugars and proline showed more deviation from mid-parent values in crosses involving C24 than in those involving Col, and deviations were larger in acclimated than in non-acclimated plants. There were significant correlations between the content of different sugars and leaf freezing tolerance, as well as between heterosis effects in freezing tolerance and sugar content. Flavonoid content and composition varied between accessions, and between non-acclimated and acclimated plants. In the crosses, large deviations from the mid-parent values in the contents of different flavonols occurred, and there were strikingly strong correlations between both flavonol content and freezing tolerance, and between heterosis effects in freezing tolerance and flavonol content

Glass-formingproperty of hydroxyectoine is the cause of its superior function as a desiccation protectant

We were able to demonstrate that hydroxyectoine, in contrast to ectoine, is a good glass-forming compound. Fourier transform infrared and spin label electron spin resonance studies of dry ectoine and hydroxyectoine have shown that the superior glass-forming properties of hydroxyectoine result from stronger intermolecular H-bonds with the OH group of hydroxyectoine. Spin probe experiments have also shown that better molecular immobilization in dry hydroxyectoine provides better redox stability of the molecules embedded in this dry matrix. With a glass transition temperature of 87°C (vs. 47°C for ectoine) hydroxyectoine displays remarkable desiccation protection properties, on a par with sucrose and trehalose. This explains its accumulation in response to increased salinity and elevated temperature by halophiles such as Halomonas elongata and its successful application in “anhydrobiotic engineering” of both enzymes and whole cells

Is oxidative stress involved in the loss of neem (Azadirachta indica) seed viability?

Neem (Azadirachta indica) is a valuable multipurpose tree of tropical arid and semi-arid regions. The use of its seeds is hindered by their short storage longevity. The possible causes of rapid loss of viability were investigated on different seed lots during exposure to 32% and 75% RH at 20°C. Within 6 months the seeds almost lost germinability at 75% RH, whereas at 32% RH viability decreased only slightly. On rehydration, the axis cells from nongerminable seeds had lost turgor, whereas those from viable seeds were turgescent as visualized by low temperature scanning electron microscopy images of fractured axes. Glutathione oxidation status was used to estimate oxidative stress during storage. Oxidative stress was much higher at 75% RH storage than at 32% RH, mainly caused by the rapid loss of reduced glutathione at 75% RH. Oligosaccharides and phospholipids decreased, and free fatty acids increased during storage at the high RH but remained at a constant level at the low RH. However, the degree of fatty acid unsaturation between viable and nonviable seed lots was similar. During the (slow) dehydration of fresh seeds, total glutathione, oligosaccharides and phospholipids accumulated, particularly in the initially more hydrated seeds. We interpret this accumulation as a post-maturation process associated with acquisition of the capability for long-term survival in the dry state. The mass ratio of oligosaccharides to sucrose was 0.19 on average in dehydrated neem seeds. The data suggest that the storage behaviour of neem seeds has features that characterize it as orthodox

Suspended sediment fluxes in an Indonesian river draining a rainforested basin subject to land cover change

Forest clearing for reasons of timber production, open pit mining and the establishment of oil palm plantations generally results in excessively high sediment loads in the tropics. The increasing sediment fluxes pose a threat to coastal marine ecosystems such as coral reefs. This study presents observations of suspended sediment fluxes in the Berau river (Indonesia), which debouches into a coastal ocean that can be considered the preeminent center of coral diversity. The Berau is an example of a small river draining a mountainous, relatively pristine basin that receives abundant rainfall. Flow velocity was measured over a large part of the river width at a station under the influence of tides, using a Horizontal Acoustic Doppler Current Profiler (HADCP). Surrogate measurements of suspended sediment concentration were taken with an Optical Backscatter Sensor (OBS). Tidally averaged suspended sediment concentration increases with river discharge, implying that the tidally averaged suspended sediment flux increases non-linearly with river discharge. Averaged over the 6.5 weeks observations covered by the benchmark survey, the tidally averaged suspended sediment flux was estimated at 2 Mt y-1. Considering the wet conditions during the observation period, this figure may be considered as an upper limit of the yearly averaged flux. This flux is significantly smaller than what could have been expected from the characteristics of the catchment. The consequences of ongoing clearing of rainforest were explored using a plot scale erosion model. When rainforest, which still covered 50–60 % of the basin in 2007, is converted to production land, soil loss is expected to increase with a factor between 10 and 100. If this soil loss is transported seaward as suspended sediment, the increase in suspended sediment flux in the Berau river would impose a severe sediment stress on the global hotspot of coral reef diversity. The impact of land cover changes will largely depend on the degree in which the Berau estuary acts as a sediment tra

Suspended sediment load in the tidal zone of an Indonesian river

Forest clearing for reasons of timber production, open pit mining and the establishment of oil palm plantations generally results in excessively high sediment loads in tropical rivers. The increasing sediment loads pose a threat to coastal marine ecosystems, such as coral reefs. This study presents observations of suspended sediment loads in the Berau River (Kalimantan, Indonesia), which debouches into a coastal ocean that is a preeminent center of coral diversity. The Berau River is relatively small and drains a mountainous, still relatively pristine basin that receives abundant rainfall. In the tidal zone of the Berau River, flow velocity was measured over a large part of the river width using a horizontal acoustic Doppler current profiler (HADCP). Surrogate measurements of suspended sediment concentration were taken with an optical backscatter sensor (OBS). Averaged over the 6.5 weeks covered by the benchmark survey period, the suspended sediment load was estimated at 2 Mt yr-1. Based on rainfall-runoff modeling though, the river discharge peak during the survey was supposed to be moderate and the yearly averaged suspended sediment load is most likely somewhat higher than 2 Mt yr-1. The consequences of ongoing clearing of rainforest were explored using a plot-scale erosion model. When rainforest, which still covered 50–60% of the basin in 2007, is converted to production land, soil loss is expected to increase with a factor between 10 and 100. If this soil loss is transported seaward as suspended sediment, the increase in suspended sediment load in the Berau River would impose a severe stress on this global hotspot of coral reef diversity