Bacillus thuringiensis A1 improve phenol tolerance and phytoextraction by Acorus calamus L.

Phenol, as a very toxic pollutant, exists widely in rivers in China. To explore the effect of bacterial augmentation on phytoremediation of phenol by Acorus calamus L., some plant growth and physiological parameters and percent removal of phenol were determined in hydroponics containing phenol with addition of Bacillus thuringiensis A1. The A. calamus L. and B. thuringiensis A1 consortium increased the growth rate of plant height, chlorophyll content, the activity of superoxide dismutase (SOD) and peroxidase (POD) in A. calamus L. 10.00-36.54%, 0.62 - 22.15%, 3.94 - 11.25% and 1.37-10.50% respectively compared with single plant treatments at same phenol concentrations. However, the addition of B. thuringiensis A1 decreased the content of malondialdehyde (MDA) and relative electrical conductivity (REC) in A. calamus L. 12.99-23.66% and 8.38-29.98% respectively compared with single plant treatments. The removal efficiency of phenol (increased from 1.56% to 13.78%) by the A. calamus L. and B. thuringiensis A1 consortium was higher than the removal efficiency of phenol of the independent A. calamus L. system. In conclusion, the addition of B. thuringiensis A1 alleviated phenol stress to A. calamus L and enhanced phenol removal due to phenol removal by bacterial augmentation.Novelty statementThe addition of B. thuringiensis A1 alleviated phenol stress to A. calamus L. and enhanced phenol removal due to phenol removal by bacterial augmentation.

Growth responses of eight wetland species to water level fluctuation with different ranges and frequencies.

Fluctuation range and frequency are two important components of water level fluctuation, but their effects on wetland plants have not been evaluated separately. We subjected eight wetland species to a control treatment with static water level and fluctuation treatments with different ranges or frequencies to examine their effects on plant growth. Acorus calamus, Butomus umbellatus and Iris wilsonii showed high survival rates in all treatments with various fluctuation ranges and frequencies. Their survival rates were higher at the medium fluctuation frequency than at the low and high frequencies, suggesting beneficial effects of the medium frequency. In the experiment comparing the fluctuation ranges, A. calamus and I. wilsonii could maintain the capacity for asexual propagation and accumulate higher biomass compared with the control plants, while biomass of the other six species dramatically decreased. In the experiment comparing fluctuation frequency, species with relatively high survival rates (≥ 50%) maintained or increased the capacity of asexual propagation, and A. calamus and I. wilsonii allocated relatively more biomass to roots, which may enhance plant growth and survival. In contrast, these species did not show increased biomass allocation to shoots in response to both fluctuation range and frequency, presumably because shoots are prone to mechanical damage caused by streaming floodwater. Taken together, biomass accumulation in roots rather than in shoots and the ability to asexually propagate are important for the survival of these species during water fluctuation.

[Influences of Biochar Application on Root Aerenchyma and Radial Oxygen Loss of Acorus calamus in Relation to Subsurface Flow in a Constructed Wetland].

In the subsurface flow of a constructed wetland (CW) used for treating wastewater, low oxygen diffusion results in long-term anoxic or anaerobic surroundings, which cannot meet the needs of plant respiration and poses a threat to the survival of macrophytes. Although sweet sedge (Acorus calamus L.) has a significant ability to resist hypoxia, membrane lipid oxidation would still occur in the plant due to the long-term hypoxia in the CW. According to reports in the existing literature, activation of the antioxidative response system could be promoted by adding biochar, thereby significantly decreasing the malonic dialdehyde in the plants. However, the specific reasons why biochar alleviates the stress from anoxia are still not clear. Thus, the responses of macrophyte roots to biochar application were studied in five different CWs built in a greenhouse, using plant ecology analyses combined with root aerenchyma, root porosity, and radial oxygen loss (ROL). The results showed that adding biochar to CW was beneficial for sweet sedge to form root aerenchyma and to increase root porosity. Moreover, there was a significant positive correlation between root porosity and the amount of biochar applied. Photosynthetic metabolism could be indirectly promoted by biochar application by increasing oxygen partial pressure in the blades, helping to transport O2 to underground parts through aerenchyma, and spreading O2 to the rhizosphere in the form of ROL. The reduction environment could be improved by applying biochar in CWs, which was also beneficial for ROL. Compared with other light conditions, 3000 µmol·(m2·s)-1 was more suitable for the growth of A. calamus in CWs with biochar, where the ability of the plants to secrete oxygen would be stimulated and enhanced. However, the effect of the biochar application ratio on ROL was not significant.

Stomatal Conductance Measurement for Toxicity Assessment in Zero-Effluent Constructed Wetlands: Effects of Landfill Leachate on Hydrophytes.

In this research, we explore for the first time the use of leaf stomatal conductance (gs) for phytotoxicity assessment. Plants respond to stress by regulating transpiration. Transpiration can be correlated with stomatal conductance when the water vapor pressure gradient for transpiration is constant. Thus, our working hypothesis was that the gs measurement could be a useful indicator of the effect of toxic compounds on plants. This lab-scale study aimed to test the measurement of gs as a phytotoxicity indicator. Our model plants were two common hydrophytes used in zero-effluent constructed wetlands for treating landfill leachate. The toxic influence of two types of leachate from old landfills (L1, L2) on common reed (Phragmites australis (Cav.) Trin. ex Steud.) and sweet flag (Acorus calamus L.) was tested. The gs measurements correlated well with plant response to treatments with six solutions (0 to 100%) of landfill leachate. Sweet flag showed higher tolerance to leachate solutions compared to common reed. The estimated lowest effective concentration (LOEC) causing the toxic effect values for these leachates were 3.94% of L1 and 5.76% of L2 in the case of reed, and 8.51% of L1 and 10.44% of L2 in the case of sweet flag. Leachate L1 was more toxic than L2. The leaf stomatal conductance measurement can be conducted in vivo and in the field. The proposed approach provides a useful parameter for indicating plant responses to the presence of toxic factors in the environment.

Effect of plant species compositions on performance of lab-scale constructed wetland through investigating photosynthesis and microbial communities.

This study focused on the effects of plant compositions on removal rates of pollutants in microcosms through investigating rhizosphere microbial populations, photosynthetic efficiency and growth characteristics. Mixed-culture groups improved the removal efficiency of TN and TP significantly but exhibited lower COD removal rates. Total plant biomasses were improved as the species richness increased, but the N/P content in the plants was mainly affected by the type of species. The mixed-culture groups showed lower photosynthesis rates and oxygen supply generated from roots under high irradiation. Microbial communities of the cultured groups in the rhizosphere exhibited significant differences. According to principal component analysis (PCA), the fungi were the typical microbes of SPA, SPAB, and SPABC, resulted in improvement in nutrient accumulation. These results demonstrated that a mixed culture strategy can represent the overyielding of biomass, promote the photo-protection mechanism, and will further increase the removal rates of pollutants in a constructed wetland.

Comparative studies on growth and physiological responses of unicellular and colonial Microcystis aeruginosa to Acorus calamus.

In order to explore the growth inhibition and physiological responses of unicellular and colonial Microcystis aeruginosa during coexistence with Acorus calamus, algal densities, chlorophyll a contents, exopolysaccharide (EPS) concentrations, malondialdehyde (MDA) contents, catalase (CAT) activities, and peroxidase (POD) activities of the two algae strains were analyzed. Although the unicellular and colonial strains of M. aeruginosa were both inhibited by A. calamus, unicellular algae were more sensitive than the colonial algae. The measurement results for EPS, MDA, CAT, and POD showed that unicellular M. aeruginosa had higher levels of stress related damage than colonial strains when they were exposed to the same density of A. calamus, and the cellular defense system of colonial M. aeruginosa was stronger than that of unicellular M. aeruginosa. Natural blooms of Microcystis are typically composed of colonial forms of M. aeruginosa, therefore future efforts to control such blooms, possibly through the development of new algicides, should focus on the unique characteristics of colonial M. aeruginosa strains.

Spatio-temporal patterns in rhizosphere oxygen profiles in the emergent plant species Acorus calamus.

Rhizosphere oxygen profiles are the key to understanding the role of wetland plants in ecological remediation. Though in situ determination of the rhizosphere oxygen profiles has been performed occasionally at certain growing stages within days, comprehensive study on individual roots during weeks is still missing. Seedlings of Acorus calamus, a wetland monocot, were cultivated in silty sediment and the rhizosphere oxygen profiles were characterized at regular intervals, using micro-optodes to examine the same root at four positions along the root axis. The rhizosphere oxygen saturation culminated at 42.9% around the middle part of the root and was at its lowest level, 3.3%, at the basal part of the root near the aboveground portion. As the plant grew, the oxygen saturation at the four positions remained nearly constant until shoot height reached 15 cm. When shoot height reached 60 cm, oxygen saturation was greatest at the point halfway along the root, followed by the point three-quarters of the way down the root, the tip of the root, and the point one-quarter of the way down. Both the internal and rhizosphere oxygen saturation steadily increased, as did the thickness of stably oxidized microzones, which ranged from 20 µm in younger seedlings to a maximum of 320 µm in older seedlings. The spatial patterns of rhizosphere oxygen profiles in sediment contrast with those from previous studies on radial oxygen loss in A. calamus that used conventional approaches. Rhizosphere oxygen saturation peaked around the middle part of roots and the thickness of stably oxidized zones increased as the roots grew.

[Influence of perennial flooding and drought on growth restoration of Acorus calamus in water-level-fluctuation zone of the Three Gorges Reservoir].

Acorus calamus L. is a common kind of wetland plant species in the Three Gorges Reservoir. In this study, we investigated the influence of perennial flooding on growth restoration of A. calamus in the lightless conditions and the drought stress on this plant species' growth after flooding. Our research provided the scientific basis for the selection of candidate species for vegetations restoration in water-level-fluctuation zone of the Three Gorges Reservoir. A. calamus plants were exposed to waters in the lightless conditions in September 2009 and September 2010 respectively and taken away from the waters and grew in natural conditions in the following March, April and May (named as S1, S2, S3). All plants in the control, S1 and S2 groups were challenged with drought stress in May for 20 days. During the experiment, the plant number and leaf number were recorded regularly, as well as leaf length and leaf width. The results showed that flooding restrained the germination of the plants with much less plant in flooding groups than the control, and the plant germination rate had inverse relation to the flooding time. Flooding promoted formation and elongation of the leaves in S1 and S2 groups, which showed higher leaf growth parameters, such as leaf length, leaf number, total leaf length of one plant and total leaf length of all plants than the control. However, all of these growth parameters in S3 group had significantly lower values compared to the control. The survival rate of the plants after flooding decreased significantly with longer flooding time. Besides, the leaf length and leaf width in S1 and S2 groups increased significantly but with decreased leaf number. Additionally, all growth parameters (leaf length, leaf width, leaf number, total leaf number, total leaf length of one plant, total leaf length of all plants) in S3 group decreased remarkably. Furthermore, drought decreased the values of all growth parameters and the plant number in the control, S1 and S2 groups notably. When drought stress was removed for 25 days, the leaf number in the control, S1 and S2 groups increased by 67.0% (P < 0.05), 66.7% (P < 0.05) and 36.2% (P < 0.05), respectively, and the total leaf length of one plant, total leaf length of all plants and total leaf number in S1 and S2 groups increased by 48.2%, 18.1%, 66.7%, 35.0%, 75.0% and 64.3%, respectively (P < 0.05). Therefore, A. calamus exhibited not only strong adaption and tolerance to flooding,but also robust growth restoration ability after flooding, as well as good restoration ability to the drought stress. In summary, A. calamus could be used as one kind of restoration or reconstruction species in water-level-fluctuation zone (especially not exposed to flooding in March or April) of the Three Gorges Reservoir.

Diurnal fluctuations in oxygen release from roots of Acorus calamus Linn in a modeled constructed wetland.

Oxygen is known to be released from plant roots, but has seldom been quantified for wetland plants. Our study aims to quantify oxygen release from the roots of one wetland species in China, and use this knowledge as a basis for future modeling. We measured diurnal fluctuations in oxygen release from the roots of Acorus calamus Linn in a modeled constructed wetland (CW) using a titanium ([image omitted]) citrate buffer. Oxygen release was monitored every two hours. Maximum oxygen release was recorded in the range of 215.2-750.8 µmolg(-1)h(-1) and occurred around 15:00. The maximum value of photosynthetically active radiation (PAR) was in the range of 1281.8-1712.0 mmolm(-2)s(-1) and occurred around 13:00. Both the oxygen release rate and PAR were found to approach zero at night. Our results indicate that oxygen release depends largely on light intensity and exhibits a diurnal periodicity with release occurring only during daytime. Rate of root oxygen release varied during the daytime and this temporal variation was well described by the Gaussian function. While further validation is needed, we suggest that the Gaussian function may be used as the basis for modeling root oxygen release in natural and constructed wetlands.

[Purification efficiency of several wetland macrophytes on COD and nitrogen removal from domestic sewage].

In order to investigate the role of wetland macrophytes in waster water purification and to select appropriate native filter plants in constructed wetland, three vertical-flow constructed wetlands were built with river sands as the substrates of Acorus gramineus, Juncus effusus and Iris japonica, and one without plant as the control. Investigation on the removal of COD and total nitrogen (TN) from domestic sewage showed that within lower concentrations of COD (<200 mg x L(-1)) and TN (<30 mg x L(-1)), more than 90% of COD and 80% of TN were removed from domestic sewage in all constructed wetlands. When the concentration of COD and TN increased, the purification efficiency of all constructed wetlands decreased to some extent. The constructed wetlands with macrophytes had a higher efficiency than control. Among the three constructed wetlands with macrophytes, the one with Acorus gramineus had an average purification efficiency of 80.46% for COD and 77.77% for TN, that with Juncus effusus was 75.53% for COD and 71.17% for TN, and the one with Iris japonica was 70.50% for COD and 66.38% for TN. The constructed wetland without vegetation had an average purification efficiency of 61.39% for COD and 55.81% for TN. Acorus gramineus was more capable of removing COD and TN than Juncus effusus and Iris japonica. Vegetation biomass was the main factor affecting the removal rate of COD and nitrogen, because it significantly correlated with the ability of absorbing organic substance and nitrogen, and with the nitrification and denitrification around roots.

Allelopathic effect of Acorus tatarinowii upon algae.

Besides competing with algae for light and mineral nutrients (i.e. N, P, etc.), the root system of Acorus tatarinowii excretes some chemical substances, which injure and eliminate alga cells, to inhibit the growth of the algae. When the algae cells were treated in "A. tatarinowii water", some of the chlorophyll a were destroyed and the photosynthetic rate of algae decreased markedly and the ability of alga cells to deoxidize triphenyltetrazolium chloride (TTC) reduced greatly. Then alga cells turned from bright red to bluish green under fluorescence microscope. These showed that the allelopathic effects of A. tatarinowii on algae were obvious and planting A. tatarinowii can control some green algae. The experiment on the extractions of the secretions of the root system showed that the inhibitory effect had a concentration effect. If the concentration of the root secretion was below 30 microliters/disc, the inhibitory rate was negative; if it was over 45 microliters/disc, the inhibitory rate was positive. This proved that the influence of the root secretion on the same acceptor was a kind of concentration effect. When the concentration of the root secretion was low, it promoted the growth of algae; when the concentration reached a definite threshold value, it restrained the growth of algae. In present case, the threshold value was between 30 microliters/disc and 45 microliters/disc.

Long-term anoxia tolerance in leaves of Acorus calamus L. and Iris pseudacorus L.

Mature green leaves of Acorus calamus and Iris pseudacorus have been shown to survive at least 28 d of total anoxia in the dark during the growing season, increasing up to 75 d and 60 d in overwintering leaves in A. calamus and I. pseudacorus, respectively. During the period of anaerobic incubation the glycolytic rate is reduced, carbohydrate reserves are conserved and ethanol levels in the tissues reached an equilibrium. Prolonged anoxia significantly suppressed leaf capacity for respiration and photosynthesis. After 28 d of anoxia, respiratory capacity was reduced in A. calamus and I. pseudacorus by 80% and 90%, respectively. The photosynthetic capacity of leaves decreased by 83% in A. calamus and by 97% in I. pseudacorus after 28 d of anoxia. This reduction in photosynthetic capacity was accompanied by a modification of the chlorophyll fluorescence pattern indicating damage to the PSII reaction centre and subsequent electron transport. Chlorophyll content was only slightly reduced after 28 d under anoxia and darkness in A. calamus, whereas there was a 50% reduction in I. pseudacorus. On return to air A. calamus leaves that endured 28 d of anoxia recovered full photosynthetic activity within 7 d while those of I. pseudacorus had a lag phase of 3-10 d. This well-developed ability to endure prolonged periods of oxygen deprivation in both these species is associated with a down-regulation in metabolic activity in response to the imposition of anaerobiosis. It is suggested that when leaf damage eventually does take place in these species after protracted oxygen deprivation, it is anoxic rather than post-anoxic stress that is responsible.