Nutrient limitation of algae and macrophytes in streams: Integrating laboratory bioassays, field experiments, and field data.

Successful eutrophication control strategies need to address the limiting nutrient. We conducted a battery of laboratory and in situ nutrient-limitation tests with waters collected from 9 streams in an agricultural region of the upper Snake River basin, Idaho, USA. Laboratory tests used the green alga Raphidocelis subcapitata, the macrophyte Lemna minor (duckweed) with native epiphytes, and in situ nutrient-limitation tests of periphyton were conducted with nutrient-diffusing substrates (NDS). In the duckweed/epiphyte test, P saturation occurred when concentrations reached about 100 µg/L. Chlorophyll a in epiphytic periphyton was stimulated at low P additions and by about 100 µg/L P, epiphytic periphyton chlorophyll a appeared to be P saturated. Both duckweed and epiphyte response patterns with total N were weaker but suggested a growth stimulation threshold for duckweed when total N concentrations exceeded about 300 µg/L and approached saturation at the highest N concentration tested, 1300 µg/L. Nutrient uptake by epiphytes and macrophytes removed up to 70 and 90% of the N and P, respectively. The green algae and the NDS nutrient-limitation test results were mostly congruent; N and P co-limitation was the most frequent result for both test series. Across all tests, when N:P molar ratios >30 (mass ratios >14), algae or macrophyte growth was P limited; N limitation was observed at N:P molar ratios up to 23 (mass ratios up to 10). A comparison of ambient periphyton chlorophyll a concentrations with chlorophyll a accrued on control artificial substrates in N-limited streams, suggests that total N concentrations associated with a periphyton chlorophyll a benchmark for desirable or undesirable conditions for recreation would be about 600 to 1000 µg/L total N, respectively. For P-limited streams, the corresponding benchmark concentrations were about 50 to 90 µg/L total P, respectively. Our approach of integrating controlled experiments and matched biomonitoring field surveys was cost effective and more informative than either approach alone.

The accumulation, transformation, and effects of quinestrol in duckweed (Spirodela polyrhiza L.).

Potential risk of endocrine disrupting compounds on non-target organisms has received extensive attentions in recent years. The present work aimed to investigate the behavior and effect of a synthetic steroid estrogen quinestrol in duckweed Spirodela polyrhiza L. Experimental results showed that quinestrol could be uptaken, accumulated, and biotransformed into 17 α-ethynylestradiol in S. polyrhiza L. The accumulation of quinestrol had a positive relation to the exposure concentration. The bioaccumulation rate was higher when the duckweed was exposed to quinestrol solutions at low concentrations than at high concentration. While the transformation of quinestrol showed no concentration-dependent manner. Quinestrol reduced the biomass and pigment content and increased superoxide dismutase and catalase activities and malondialdehyde contents in the duckweed. The results demonstrated that quinestrol could be accumulated and biotransformed in aquatic plant S. polyrhiza L. This work would provide supplemental data on the behavior of this steroid estrogen compound in aquatic system.

Diversity of pollination ecology in the Schismatoglottis Calyptrata Complex Clade (Araceae).

Field studies integrating pollination investigations with an assessment of floral scent composition and thermogenesis in tropical aroids are rather few. Thus, this study aimed to investigate the pollination biology of nine species belonging to Schismatoglottis Calyptrata Complex Clade. The flowering mechanism, visiting insect activities, reproductive system, thermogenesis and floral scent composition were examined. Anthesis for all species started at dawn and lasted 25-29 h. Colocasiomyia (Diptera, Drosophilidae) are considered the main pollinators for all the investigated species. Cycreon (Coleoptera, Hydrophilidae) are considered secondary pollinators as they are only present in seven of the nine host plants, despite the fact that they are the most effective pollen carrier, carrying up to 15 times more pollen grains than Colocasiomyia flies. However, the number of Colocasiomyia individuals was six times higher than Cycreon beetles. Chaloenus (Chrysomelidae, Galeuricinae) appeared to be an inadvertent pollinator. Atheta (Coleoptera, Staphylinidae) is considered a floral visitor in most investigated species of the Calyptrata Complex Clade in Sarawak, but a possible pollinator in S. muluensis. Chironomidae midges and pteromalid wasps are considered visitors in S. calyptrata. Thermogenesis in a biphasic pattern was observed in inflorescences of S. adducta, S. calyptrata, S. giamensis, S. pseudoniahensis and S. roh. The first peak occurred during pistillate anthesis; the second peak during staminate anthesis. Inflorescences of all investigated species of Calyptrata Complex Clade emitted four types of ester compound, with methyl ester-3-methyl-3-butenoic acid as a single major VOC (volatile organic compound). The appendix, pistillate zone, staminate zone and spathe emitted all these compounds. A mixed fly-beetle pollination system is considered an ancestral trait in the Calyptrata Complex Clade, persisting in Sarawak taxa, whereas the marked reduction of interpistillar staminodes in taxa from Peninsular Malaysia and especially, Ambon, Indonesia, is probably linked to a shift in these taxa to a fly-pollinated system.

Effects of selenium on biological and physiological properties of the duckweed Landoltia punctata.

Duckweed can be used for bioremediation of selenium (Se) polluted water because of its capability of absorbing minerals from growing media. However, the presence of Se in the media may affect the growth of the duckweed. Landoltia punctata 7449 has been studied for its changes in chemical and biological properties with the presence of Se in the media. The duckweed was cultivated over a 12-day period at different initial concentrations of selenite (Na2 SeO3 ) from 0 to 80 µmol·l(-1) . The growth rate, the organic and total Se contents, the activity of antioxidant enzymes, the photosynthetic pigment contents, the chlorophyll a fluorescence OJIP transient, and the ultrastructure of the duckweed were monitored during the experiment. The results have shown that Se at low concentrations of ≤20 µmol·l(-1) promoted the growth of the L. punctata and inhibited lipid peroxidation. Substantial increases in duckweed growth rate and organic Se content in the duckweed were observed at low Se concentrations. The anti-oxidative effect occurred likely with the increases in guaiacol peroxidase, catalase and superoxide dismutase activities as well as the amount of photosynthetic pigments. However, negative impact to the duckweed was observed when the L. punctata was exposed to high Se concentrations (≥40 µmol·l(-1) ), in which the duckweed growth was inhibited by the selenium. The results indicate that L. punctata 7449 can be used for bioremediation of selenium (Se) polluted water when the Se concentration is ≤20 µmol·l(-1) .

Impact of exogenous GABA treatments on endogenous GABA metabolism in anthurium cut flowers in response to postharvest chilling temperature.

Anthurium flowers are susceptible to chilling injury, and the optimum storage temperature is 12.5-20 °C. The γ-aminobutyric acid (GABA) shunt pathway may alleviate chilling stress in horticultural commodities by providing energy (ATP), reducing molecules (NADH), and minimizing accumulation of reactive oxygen species (ROS). In this experiment, the impact of a preharvest spray treatment with 1 mM GABA and postharvest treatment of 5 mM GABA stem-end dipping on GABA shunt pathway activity of anthurium cut flowers (cv. Sirion) in response to cold storage (4 °C for 21 days) was investigated. GABA treatments resulted in lower glutamate decarboxylase (GAD) and higher GABA transaminase (GABA-T) activities in flowers during cold storage, which was associated with lower GABA content and coincided with higher ATP content. GABA treatments also enhanced accumulation of endogenous glycine betaine (GB) in flowers during cold storage, as well as higher spathe relative water content (RWC). These findings suggest that GABA treatments may alleviate chilling injury of anthurium cut flowers by enhancing GABA shunt pathway activity leading to provide sufficient ATP and promoting endogenous GB accumulation.

Floral diversity and pollination strategies of three rheophytic Schismatoglottideae (Araceae).

Homoplastic evolution of 'unique' morphological characteristics in the Schismatoglottideae - many previously used to define genera - prompted this study to compare morphology and function in connection with pollination biology for Aridarum nicolsonii, Phymatarum borneense and Schottarum sarikeense. Aridarum nicolsonii and P. borneense extrude pollen through a pair of horned thecae while S. sarikeense sheds pollen through a pair of pores on the thecae. Floral traits of spathe constriction, presence and movement of sterile structures on the spadix, the comparable role of horned thecae and thecae pores, the presence of stamen-associated calcium oxalate packages, and the timing of odour emission are discussed in the context of their roles in pollinator management. Pollinators for all investigated species were determined to be species of Colocasiomyia (Diptera: Drosophilidae).

Uranium and cadmium provoke different oxidative stress responses in Lemna minor L.

Common duckweed (Lemna minor L.) is ideally suited to test the impact of metals on freshwater vascular plants. Literature on cadmium (Cd) and uranium (U) oxidative responses in L. minor are sparse or, for U, non-existent. It was hypothesised that both metals impose concentration-dependent oxidative stress and growth retardation on L. minor. Using a standardised 7-day growth inhibition test, the adverse impact of these metals on L. minor growth was confirmed, with EC50 values for Cd and U of 24.1 ± 2.8 and 29.5 ± 1.9 µm, respectively, and EC10 values of 1.5 ± 0.2 and 6.5 ± 0.9 µm, respectively. The metal-induced oxidative stress response was compared through assessing the activity of different antioxidative enzymes [catalase, glutathione reductase, superoxide dismutase (SOD), ascorbate peroxidase (APOD), guaiacol peroxidase (GPOD) and syringaldizyne peroxidase (SPOD)]. Significant changes in almost all antioxidative enzymes indicated their importance in counteracting the U- and Cd-imposed oxidative burden. However, some striking differences were also observed. For activity of APODs and SODs, a biphasic but opposite response at low Cd compared to U concentrations was found. In addition, Cd (0.5-20 µm) strongly enhanced plant GPOD activity, whereas U inhibited it. Finally, in contrast to Cd, U up to 10 µm increased the level of chlorophyll a and b and carotenoids. In conclusion, although U and Cd induce similar growth arrest in L. minor, the U-induced oxidative stress responses, studied here for the first time, differ greatly from those of Cd.

Utilization of two invasive free-floating aquatic plants (Pistia stratiotes and Eichhornia crassipes) as sorbents for oil removal.

Free-floating aquatic plants Pistia stratiotes and Eichhornia crassipes are well-known invasive species in the tropics and subtropics. The aim of this study was to utilize the plants as cost-effective and environmentally friendly oil sorbents. Multilevel wrinkle structure of P. stratiotes leaf (PL), rough surface of E. crassipes leaf (EL), and box structure of E. crassipes stalk (ES) were observed using the scanning electron microscope. The natural hydrophobic structures and capillary rise tests supported the idea to use P. stratiotes and E. crassipes as oil sorbents. Experiments indicated that the oil sorption by the plants was a fast process. The maximum sorption capacities for different oils reached 5.1-7.6, 3.1-4.8, and 10.6-11.7 g of oil per gram of sorbent for PL, EL, and ES, respectively. In the range of 5-35 °C, the sorption capacities of the plants were not significantly different. These results suggest that the plants can be used as efficient oil sorbents.

Cloning and functional analysis of a novel ascorbate peroxidase (APX) gene from Anthurium andraeanum.

An 888-bp full-length ascorbate peroxidase (APX) complementary DNA (cDNA) gene was cloned from Anthurium andraeanum, and designated as AnAPX. It contains a 110-bp 5'-noncoding region, a 28-bp 3'-noncoding region, and a 750-bp open reading frame (ORF). This protein is hydrophilic with an aliphatic index of 81.64 and its structure consisting of α-helixes, ß-turns, and random coils. The AnAPX protein showed 93%, 87%, 87%, 87%, and 86% similarities to the APX homologs from Zantedeschia aethiopica, Vitis pseudoreticulata, Gossypium hirsutum, Elaeis guineensis, and Zea mays, respectively. AnAPX gene transcript was measured non-significantly in roots, stems, leaves, spathes, and spadices by real-time polymerase chain reaction (RT-PCR) analysis. Interestingly, this gene expression was remarkably up-regulated in response to a cold stress under 6 °C, implying that AnAPX might play an important role in A. andraeanum tolerance to cold stress. To confirm this function we overexpressed AnAPX in tobacco plants by transformation with an AnAPX expression construct driven by CaMV 35S promoter. The transformed tobacco seedlings under 4 °C showed less electrolyte leakage (EL) and malondialdehyde (MDA) content than the control. The content of MDA was correlated with chilling tolerance in these transgenic plants. These results show that AnAPX can prevent the chilling challenged plant from cell membrane damage and ultimately enhance the plant cold tolerance.

Micronucleation by mitosis inhibitors in developing microspores of Spathiphyllum wallisii Regel.

KEY MESSAGE : We developed an efficient protocol for chromosome scattering in Spathiphyllum microspores. The effects of plant material, developmental age, genotype and antimicrotubular toxin type, exposure and concentration were evaluated. Asymmetric hybridization through microprotoplast-mediated chromosome transfer (MMCT) is a known method for overcoming sexual breeding barriers between distantly related plant species. To obtain microprotoplasts, it is necessary to induce mass micronucleation either in somatic or gametic cells. We have tested the efficiency for micronuclei induction of five mitosis inhibitors, amiprophos-methyl (APM), butamiphos (BUT), chlorpropham (CIPC), oryzalin (ORY) and propyzamide (PRO), on developing microspores of diploid Spathiphyllum wallisii Regel. Besides the used toxins, also the effect of their concentrations and incubation period as well as plant genotypes and material was tested. We observed micronuclei (MNi) in pollen mother cells, dyads and tetrads as well as other abnormalities such as ball metaphases and chromosome bridges. The flower position on the spadix and the type of starting material (dissected anthers vs. complete spadices) did not significantly influence micronucleation frequencies. The highest micronucleation index of 86 % was obtained in microspores treated with 10 µM ORY during 72 h. All six genotypes tested formed micronuclei after this particular treatment, although the efficiency varied between cultivars. Next to ORY, CIPC was also a very efficient MNi inducer. The average number of MNi found in micronucleated cells varied between 1.67-6.44 for CIPC and 0.83-5.50 for ORY. The maximal number of MNi observed was 12 for CIPC and 9 for ORY. Our results demonstrate that CIPC and ORY can be applied for mass micronucleation on developing microspores of S. wallisii as a first step of MMCT in aroid interspecific or intergeneric breeding.

Nutrient removal of effluent from quail farm through cultivation of Wolffia arrhiza.

The objective of this work was to study the nutrient removal using the Wolffiaarrhiza during the treatment of laying quails farm effluent. The relationship between W. arrhiza biomass and treatment time, the change in water qualities, and nitrogen-balance (N-balance) were evaluated. The results showed that a biomass of 12.0g of W. arrhiza per liter of effluent and a treatment period of 30 days were found to provide the best conditions for W. arrhiza's growth and the quality of the treated effluent in terms of biological oxygen demand, suspended solids, total phosphorus, nitrate, total ammonia nitrogen and total Kjeldahl nitrogen. The pH and salinity were similar for each level of biomass. The W. arrhiza biomasses of 4.00-12.0g/l of effluent were suitable for W. arrhiza survival over time. Since W. arrhiza can fix N in the atmosphere, it can grow very well in effluent containing a low level of N.

Real-time visualization of oxidative stress in a floating macrophyte Lemna minor L. exposed to cadmium, copper, menadione, and AAPH.

An ultra-sensitive digital imaging system was employed to visualize oxidative stress in intact L. minor plants exposed to Cd, Cu, menadione, AAPH, and ascorbate in real time. The increase of ROS production was assessed by measuring the rate of fluorescence intensity increases of the test medium supplemented with a fluorescing probe (dichlorofluorescein diacetate). The addition of 100 µM CdCl2 or 100 µM CuSO4 to the growth medium resulted in a significant increase of medium fluorescence. Additionally, CuSO4 caused a significantly higher fluorescence intensity than CdCl2 did. A strong positive correlation (R² = 0.99) between menadione concentration and fluorescence intensity was observed. The positive correlation between AAPH concentration and fluorescence intensity was not as strong as in the case of menadione (R² = 0.81). Menadione induced a stronger oxidative stress than similar concentration of AAPH. The addition of 100 µM ascorbate to L. minor treated with 50 µM menadione significantly reduced the fluorescence intensity increase. A linear trend of the fluorescence increase was observed in all treatments, indicating that chemical-induced oxidative stress is a gradual process and that the applied concentrations of the chemicals caused a constant increased production of ROS with different intensities, depending on the treatment. This is the combined result of a gradual diminishing of antioxidant reserves and accumulating oxidative damage. The observed rates of ROS production were slower than those in the studies using cell cultures.

Restricted pollen flow of Dieffenbachia seguine populations in fragmented and continuous tropical forest.

Habitat fragmentation can change the ecological context of populations, rupturing genetic connectivity among them, changing genetic structure, and increasing the loss of genetic diversity. We analyzed mating system and pollen structure in two population fragments and two continuous forest populations of Dieffenbachia seguine (Araceae), an insect-pollinated understory herb in the tropical rain forest of Los Tuxtlas, México, using nine allozyme loci. Mating system analysis indicated almost complete outcrossing but some inbreeding among the adults. Pollen structure analysis indicated highly restricted pollen flow, both within and among populations. We showed that the effective pollination neighborhood was small in all populations, and slightly (though not significantly) smaller in fragments, partially as a consequence of an increase in density of reproductive individuals in those fragments. Using assignment analysis, we showed that all populations were strongly structured, suggesting that pollen and seed flow across the Los Tuxtlas landscape has been spatially restricted, though sufficient to maintain connectedness. Forest fragmentation at Los Tuxtlas has (so far) had limited impact on pollen dynamics, despite the changing ecological context, with reduced pollinator abundance being partially offset by increased flowering density in fragments. Continued outcrossing and limited pollen immigration, coupled with more extensive seed migration, should maintain genetic connectedness in D. seguine, if fragmentation is not further exacerbated by additional deforestation.

Effect of excess iron and copper on physiology of aquatic plant Spirodela polyrrhiza (L.) Schleid.

To elucidate effect of chemical reagents addition on growth of aquatic plants in restoration of aquatic ecosystem, Spirodela polyrrhiza (L.) Schleid was used to evaluate its physiological responses to excess iron (Fe(3+)) and copper (Cu(2+)) in the study. Results showed that accumulation of iron and copper both reached maximum at 100 mg L(-1) iron or copper after 24 h short-term stress, but excess iron and copper caused plants necrosis or death and colonies disintegration as well as roots abscission at excess metal concentrations except for 1 mg L(-1) iron. Significant differences in chlorophyll fluorescence (Fv/Fm) were observed at 1-100 mg L(-1) iron or copper. The synthesis of chlorophyll and protein as well as carbohydrate and the uptake of phosphate and nitrogen were inhibited seriously by excess iron and copper. Proline content decreased with increasing iron or copper concentration, however, MDA content increased with increasing iron or copper concentration.

Effects of floral thermogenesis on pollen function in Asian skunk cabbage Symplocarpus renifolius.

The effects of temperature on pollen germination and pollen tube growth rate were measured in vitro in thermogenic skunk cabbage, Symplocarpus renifolius Schott ex Tzvelev, and related to floral temperatures in the field. This species has physiologically thermoregulatory spadices that maintain temperatures near 23 degrees C, even in sub-freezing air. Tests at 8, 13, 18, 23, 28 and 33 degrees C showed sharp optima at 23 degrees C for both variables, and practically no development at 8 degrees C. Thermogenesis is therefore a requirement for fertilization in early spring. The narrow temperature tolerance is probably related to a long period of evolution in flowers that thermoregulate within a narrow range.

Caladium bicolor (Araceae) and Cyclocephala celata (Coleoptera, Dynastinae): a well-established pollination system in the Northern Atlantic rainforest of Pernambuco, Brazil.

Flowering, pollination ecology, and floral thermogenesis of Caladium bicolor were studied in the Atlantic Rainforest of Pernambuco, NE Brazil. Inflorescences of this species are adapted to the characteristic pollination syndrome performed by Cyclocephalini beetles. They bear nutritious rewards inside well-developed floral chambers and exhibit a thermogenic cycle which is synchronized to the activity period of visiting beetles. Heating intervals of the spadix were observed during consecutive evenings corresponding to the beginning of the female and male phases of anthesis. Highest temperatures were recorded during the longer-lasting female phase. An intense sweet odour was volatized on both evenings. Beetles of a single species, Cyclocephala celata, were attracted to odoriferous inflorescences of C. bicolor and are reported for the first time as Araceae visitors. All the inflorescences visited by C. celata developed into infructescences, whereas unvisited inflorescences showed no fruit development. Findings of previous studies in the Amazon basin of Surinam indicated that Cyclocephala rustica is a likely pollinator of C. bicolor. This leads to the assumption that locally abundant Cyclocephalini species are involved in the pollination of this species.

Conventional and novel modes of exine patterning in members of the Araceae--the consequence of ecological paradigm shifts?

In the family Araceae, the members of all subfamilies except Aroideae follow the conventional mode of exine formation pattern, which conforms with the textbook view of sporoderm stratification and chemistry (sporopollenin ektexine formed before the endexine). Only members of the subfamily Aroideae show a quite uncommon mode of exine formation pattern, with an endexine formed prior to the nonsporopollenin, polysaccharidic outer exine layer. The intine is formed simultaneously with this non-sporopollenin layer. From the differing timetable and especially from the different origin it is concluded that this outer exine layer is not homologous to the angiosperm ektexine. The fundamental question, why members of the Aroideae lack an elaborated sporopollenin ektexine, is discussed in terms of functionality of the nonsporopollenin outer exine layer. It seems that a major change in aroid evolution took place at the point when the family phylogenetically and ecologically shifted from bisexual (most subfamilies) to unisexual flowers (Aroideae only). The hypothesis is that ephemeral spathes and the absence of sporopollenin are the consequence of an adaptive syndrome for a short pollination time window in many members of the Aroideae, with short-lived pollen, an energetically not costly pollen wall, rapid germination of pollen tube, and brief receptivity of stigma.

Causes of low oxygen in a lowland, regulated eutrophic river in Eastern England.

In the River Brett, Eastern England, over the period 1955-1998 there was a significant long-term decline in dissolved oxygen (DO), as well as increases in TON (total oxidised nitrogen) and SRP (soluble reactive phosphorus). Flow decreased from 1963 to 1998. Field studies in 1998-2000 showed increased pH and a gradient of DO beneath the filamentous alga Cladophora glomerata. DO decreased through the summer. Macrophytes accounted for 45% of community respiration at the study site, while sediment accounted for 36%. In container studies, muddy sediments had the highest maximum sediment oxygen demand (SOD), but canopies of C. glomerata and Lemna minor together increased the SOD by up to 90% over control samples. During periods of high temperature, abundant growths of C. glomerata and/or L. minor would increase the SOD of organic mud in river areas with shallow, ponded water, eventually leading to anoxic conditions and the release of nutrients from the sediment. If a river had large areas of mud, these processes could dramatically affect the river's oxygen budget, and hence its ecology.

The capacity of duckweed to treat wastewater: ecological considerations for a sound design.

Duckweed species are promising macrophytes for use in sustainable wastewater treatment due to their rapid growth, ease of harvest, and feed potential as a protein source. This paper reviews growth rates of different duckweed species on wastewater and ammonia toxicity to duckweed and summarizes insights into the mechanism of organic matter and nutrient removal. Results were gained from laboratory experiments in small, shallow, duckweed-covered semicontinuous batch systems. Growth rates on different types of wastewater vary considerably among different species. Ammonia is toxic for duckweed in both the ionized and un-ionized forms. Duckweed, however, can be used to treat wastewater containing very high total ammonia concentrations as long as certain pH levels are not exceeded. The degradation of organic material is enhanced by duckweed through both additional oxygen supply and additional surface for bacterial growth. The duckweed mat with attached bacteria and algae is, independent of the loading rates, responsible for three-quarters of the total nitrogen (N) and phosphorus (P) loss in very shallow systems. Based on our results we suggest that full-scale pilot plants with duckweed should be shallower than the range encountered in the literature. A harvesting schedule that allows doubling times of 2 to 3.5 d, maintenance of a full coverage, and plug flow conditions are recommended.