The Chara Genome: Secondary Complexity and Implications for Plant Terrestrialization.

Land plants evolved from charophytic algae, among which Charophyceae possess the most complex body plans. We present the genome of Chara braunii; comparison of the genome to those of land plants identified evolutionary novelties for plant terrestrialization and land plant heritage genes. C. braunii employs unique xylan synthases for cell wall biosynthesis, a phragmoplast (cell separation) mechanism similar to that of land plants, and many phytohormones. C. braunii plastids are controlled via land-plant-like retrograde signaling, and transcriptional regulation is more elaborate than in other algae. The morphological complexity of this organism may result from expanded gene families, with three cases of particular note: genes effecting tolerance to reactive oxygen species (ROS), LysM receptor-like kinases, and transcription factors (TFs). Transcriptomic analysis of sexual reproductive structures reveals intricate control by TFs, activity of the ROS gene network, and the ancestral use of plant-like storage and stress protection proteins in the zygote.

A shady phytoplankton paradox: when phytoplankton increases under low light.

Light is a fundamental driver of ecosystem dynamics, affecting the rate of photosynthesis and primary production. In spite of its importance, less is known about its community-scale effects on aquatic ecosystems compared with those of nutrient loading. Understanding light limitation is also important for ecosystem management, as human activities have been rapidly altering light availability to aquatic ecosystems. Here we show that decreasing light can paradoxically increase phytoplankton abundance in shallow lakes. Our results, based on field manipulation experiments, field observations and models, suggest that, under competition for light and nutrients between phytoplankton and submersed macrophytes, alternative stable states are possible under high-light supply. In a macrophyte-dominated state, as light decreases phytoplankton density increases, because macrophytes (which effectively compete for nutrients released from the sediment) are more severely affected by light reduction. Our results demonstrate how species interactions with spatial heterogeneity can cause an unexpected outcome in complex ecosystems. An implication of our findings is that partial surface shading for controlling harmful algal bloom may, counterintuitively, increase phytoplankton abundance by decreasing macrophytes. Therefore, to predict how shallow lake ecosystems respond to environmental perturbations, it is essential to consider effects of light on the interactions between pelagic and benthic producers.

The seasonal effects on the encrustation of charophytes in two hard-water lakes.

Encrustation and element content of six charophyte species from two hard-water lakes were investigated monthly for a period of 1 year. Seasonal patterns were analyzed for the interaction of water chemistry. Encrustation followed a seasonal pattern for Chara contraria, Chara subspinosa, and Nitellopsis obtusa in Lake Krüselin and for Chara globularis and Chara tomentosa in Lake Lützlow. However, no seasonality in the precipitated CaCO3 was observed for C. subspinosa in Lake Lützlow and for C. tomentosa in Lake Krüselin, indicating a lake-specific dependency. Species-specific encrustation was found. Chara contraria and N. obtusa encrusted the most in June and August, whereas C. subspinosa and Nitella flexilis/opaca exhibited lowest encrustation in March and April. The precipitated CaCO3 of charophytes correlated negatively to the concentration of total inorganic carbon in both lakes. Element content of plant dry weight was species-specific for Ca and K, and lake-specific for Mg. No specific pattern was found for the TP and Fe contents. The results showed seasonal, species, and lake-specific influences on the encrustation of charophytes.

Influence of the residue from an iron mining dam in the growth of two macrophyte species.

On November 5th, 2015 the worst environmental disaster in Brazil spilled 60 million m3 of iron mining residue into Gualaxo do Norte River (Minas Gerais State), an affluent of the highest River Basin of the Brazilian Southeast (Doce River Basin), reaching the Atlantic Ocean. To assess the impact of the iron residue on the aquatic plant metabolism, we performed macrophyte growth experiments under controlled light and temperature conditions using two species (Egeria densa and Chara sp.). The plants' growth data were fitted in a kinetic model to obtain the biomass yields (K) and growth rates (µ). Turbidity and electrical conductivity of the water were measured over time. Both plants showed the highest growth rates in the contaminated condition (0.056 d-1 for E. densa and 0.45 d-1 for Chara sp.) and the biomass increased in the short-term (≈20 days). The control condition (i.e. no impacted water) supported the biomass increasing over time and the development of vegetative buddings with high daily rates (1.75 cm d-1 for E. densa and 0.13 cm d-1 for Chara sp). Turbidity showed a sharp decrease in 48 h and had no effects in the plants growth in the contaminated condition. The contamination affected the plants' yields in the long-term affecting the biomass development. This study provides preliminary information about the ecological consequences of a mining dam rupture aiming to collaborate with monitoring and risk assessments.

Evaluation of the feeding preference between the aquatic macrophytes Egeria densa and Chara indica by the invasive mollusk Melanoides tuberculata / Avaliação da preferência do molusco invasor Melanoides tuberculata pelas macrófitas aquáticas Egeria densa e Chara indica

Abstract This study evaluated the feeding preference of the invasive mollusk Melaniudes tuberculata between the aquatic macrophytes Egeria densa and Chara indica. The experiment consisted of twelve experimental units (glass aquariums) each of which contained three liters of water and three compartments. Fragments of E. densa and C. indica were placed in separate compartments within each unit; the third compartment, which did not contain macrophytes, was used as the control. Twenty Melanoides tuberculata individuals were placed in each unit and monitored hourly over the course of 24 hours for preferential movements. Physical and chemical water variables were measured at the beginning and end of the experiment. Habitat complexity was determined through collected macrophyte fragments and determined using the Fractop program. After 24 hours, the highest average number of individuals was observed in the treatment with Chara indica (ten individuals), which differed significantly from the treatment with E. densa (four individuals) and the control treatment (two individuals). The number of individuals between the E. densa and control treatment were similar. M. tuberculata showed a clear feeding preference for C. indica.

Resumo O presente trabalho teve como objetivo avaliar a preferência do molusco invasor M. tuberculata pelas macrófitas aquáticas Egeria densa e Chara indica. Doze unidades experimentais (aquários de vidro), com dimensões de 30×15×15 cm, subdivida em três compartimentos foram utilizadas. Em cada unidade experimental foram adicionados três litros de água, sendo que em dois compartimentos foram adicionados aleatoriamente fragmentos de E. densa e C. indica e um compartimento ficou sem planta (controle). Na parte central do aquário foram adicionados os moluscos equidistantes dos três compartimentos. O experimento teve duração de 24 horas, sendo vistoriado e contabilizado o número de organismos presentes em cada compartimento a cada hora. As variáveis ambientais foram medidas no inicio e fim do experimento. Fragmentos de macrófitas foram coletados para determinação da complexidade do habitat através do programa Fractop. Após 24 horas, os resultados mostraram um maior valor médio de indivíduos no tratamento que possuía Chara indica (dez indivíduos), que diferiu significativamente do tratamento com E. densa (quatro indivíduos) e do tratamento controle (dois indivíduos). Os tratamentos com E. densa e o controle foram semelhantes entre si evidenciando, uma possível preferência do molusco por a C. indica.

Evaluation of the feeding preference between the aquatic macrophytes Egeria densa and Chara indica by the invasive mollusk Melanoides tuberculata.

This study evaluated the feeding preference of the invasive mollusk Melaniudes tuberculata between the aquatic macrophytes Egeria densa and Chara indica. The experiment consisted of twelve experimental units (glass aquariums) each of which contained three liters of water and three compartments. Fragments of E. densa and C. indica were placed in separate compartments within each unit; the third compartment, which did not contain macrophytes, was used as the control. Twenty Melanoides tuberculata individuals were placed in each unit and monitored hourly over the course of 24 hours for preferential movements. Physical and chemical water variables were measured at the beginning and end of the experiment. Habitat complexity was determined through collected macrophyte fragments and determined using the Fractop program. After 24 hours, the highest average number of individuals was observed in the treatment with Chara indica (ten individuals), which differed significantly from the treatment with E. densa (four individuals) and the control treatment (two individuals). The number of individuals between the E. densa and control treatment were similar. M. tuberculata showed a clear feeding preference for C. indica.

Toxicokinetics of pyrene in the freshwater alga Chara rudis.

Chara has been suggested a good model to study uptake of xenobiotics into cytoplasm due to their large internode cells surrounded by a layer of cortex cells. We studied the uptake and elimination of pyrene (nominal concentration of 5 µg L(-1)) in the freshwater alga Chara rudis during 22 days in two treatments mimicking epilimnetic (warm and light) and hypolimnetic (cold and dark) conditions. The growth of Chara during the exposure was higher in epilimnetic conditions (40%) compared to both hypolimnetic pyrene exposed Chara and controls (epilimnetic and hypolimnetic, no pyrene). In the water, a more rapid dissipation of pyrene was observed in epilimnetic conditions, possibly as a result of the increased algal growth. In the cortex, pyrene, 1-OH-pyrene (minor metabolite) and an unknown hydrophobic major metabolite was measured. Pyrene amounts decreased over time, while amounts of the unknown metabolite increased. In internode cytoplasm, pyrene and 1-OH-pyrene showed initially increasing followed by decreasing trends, while the unknown metabolite was not detected. The total mass balance showed that we were able to account for the applied pyrene until 4 days of exposure. However, after this time, there was a significant decrease in amounts accounted for by fluorescence, suggesting that the metabolism of pyrene involves degradation of the ring structure. The degradation was larger in epilimnetic than hypolimnetic conditions.

Experimental evidence for enhanced top-down control of freshwater macrophytes with nutrient enrichment.

The abundance of primary producers is controlled by bottom-up and top-down forces. Despite the fact that there is consensus that the abundance of freshwater macrophytes is strongly influenced by the availability of resources for plant growth, the importance of top-down control by vertebrate consumers is debated, because field studies yield contrasting results. We hypothesized that these bottom-up and top-down forces may interact, and that consumer impact on macrophyte abundance depends on the nutrient status of the water body. To test this hypothesis, experimental ponds with submerged vegetation containing a mixture of species were subjected to a fertilization treatment and we introduced consumers (mallard ducks, for 8 days) on half of the ponds in a full factorial design. Over the whole 66-day experiment fertilized ponds became dominated by Elodea nuttallii and ponds without extra nutrients by Chara globularis. Nutrient addition significantly increased plant N and P concentrations. There was a strong interactive effect of duck presence and pond nutrient status: macrophyte biomass was reduced (by 50%) after the presence of the ducks on fertilized ponds, but not in the unfertilized ponds. We conclude that nutrient availability interacts with top-down control of submerged vegetation. This may be explained by higher plant palatability at higher nutrient levels, either by a higher plant nutrient concentration or by a shift towards dominance of more palatable plant species, resulting in higher consumer pressure. Including nutrient availability may offer a framework to explain part of the contrasting field observations of consumer control of macrophyte abundance.

Does mechanical disturbance affect the performance and species composition of submerged macrophyte communities?

Submerged macrophyte communities are frequently subjected to disturbance of various frequency and strength. However, there is still little experimental evidence on how mechanical disturbance affects the performance and species composition of such plant communities. In a greenhouse experiment, we constructed wetland communities consisting of five co-occurring clonal submerged macrophyte species (Hydrilla verticillata, Elodea canadensis, Ceratophyllum demersum, Chara fragilis, and Myriophyllum spicatum) and subjected these communities to three mechanical disturbance regimes (no, moderate and strong disturbance). Strong mechanical disturbance greatly decreased overall biomass, number of shoot nodes and total shoot length, and increased species diversity (evenness) of the total community. It also substantially decreased the growth of the most abundant species (H. verticillata), but did not affect growth of the other four species. Our data reveal that strong disturbance can have different effects on different submerged macrophyte species and thus alters the performance and species composition of submerged macrophyte communities.

Combined effects of cadmium and zinc on growth, tolerance, and metal accumulation in Chara australis and enhanced phytoextraction using EDTA.

Chara australis (R. Br.) is a macrophytic alga that can grow in and accumulate Cd from artificially contaminated sediments. We investigated the effects of Zn independently and in combination with Cd on C. australis growth, metal tolerance, and uptake. Plant growth was reduced at concentrations ≥ 75 mg Zn (kg soil)⁻¹. Zn also increased the concentration of glutathione in the plant, suggesting alleviation of stress. Phytotoxic effects were observed at ≥ 250 mg added Zn (kg soil)⁻¹. At 1.5mg Zn (kg soil)⁻¹, the rhizoid bioconcentration factor (BCF) was >1.0 for both Cd and Zn. This is a criterion for hyperaccumulator status, a commonly used benchmark for utility in remediation of contaminated soils by phytoextraction. There was no significant interaction between Cd and Zn on accumulation, indicating that Chara should be effective at phytoextraction of mixed heavy metal contamination in sediments. The effects of the chelator, ethylenediaminetetraacetic acid (EDTA), were also tested. Moderate levels of EDTA increased Cd and Zn accumulation in rhizoids and Cd BCF of shoots, enhancing Chara's potential in phytoremediation. This study demonstrates for the first time the potential of macroalgae to remove metals from sediments in aquatic systems that are contaminated with a mixture of metals.

Cd tolerance and accumulation in the aquatic macrophyte, Chara australis: potential use for charophytes in phytoremediation.

We investigated the potential use of the alga Chara australis (R. Br.) forphytore mediation of Cd-contaminated sediments in aquatic systems. Chara tolerated up to 20 mg added Cd (kg soil)⁻¹ in laboratory culture. Chlorophyll a and b levels were not affected even at Cd concentrations that suppressed growth. Levels of glutathione were suppressed at 2-35 mg added Cd (kg soil)⁻¹ to 200-350 nmol GSH (g DW)⁻¹, while control levels were 660 nmol GSH (g DW)⁻¹). Histochemical studies showed Cd occurred throughout cell walls and cytoplasm in plants grown in 5-20 mg Cd (kg soil)⁻¹. Quantification using ICP-MS showed the maximum concentration in shoots was 72 mg Cd (kg DW)⁻¹ at 35 mg added Cd (kg soil)⁻¹, while the maximum in rhizoids was 116 mg Cd (kg DW)⁻¹ at 25 mg added Cd (kg soil)⁻¹. The bioconcentration factor (BCF, concentration in plant/concentration in soil) exceeded 1.0, the critical value for hyperaccumulators, for shoots exposed to 35 mg Cd (kg soil)⁻¹ and rhizoids exposed to ≥25 mg Cd (kg soil)⁻¹. Translocation factors (TF, shoot concentration/rhizoid concentration) did not exceed 1.0 for any treatment. While Chara cannot be considered a hyperaccumulator, it shows promise for use in phytoremediation efforts.

The distribution of cell wall polymers during antheridium development and spermatogenesis in the Charophycean green alga, Chara corallina.

BACKGROUND AND AIMS: The production of multicellular gametangia in green plants represents an early evolutionary development that is found today in all land plants and advanced clades of the Charophycean green algae. The processing of cell walls is an integral part of this morphogenesis yet very little is known about cell wall dynamics in early-divergent green plants such as the Charophycean green algae. This study represents a comprehensive analysis of antheridium development and spermatogenesis in the green alga, Chara corallina. METHODS: Microarrays of cell wall components and immunocytochemical methods were employed in order to analyse cell wall macromolecules during antheridium development. KEY RESULTS: Cellulose and pectic homogalacturonan epitopes were detected throughout all cell types of the developing antheridium including the unique cell wall protuberances of the shield cells and the cell walls of sperm cell initials. Arabinogalactan protein epitopes were distributed only in the epidermal shield cell layers and anti-xyloglucan antibody binding was only observed in the capitulum region that initially yields the sperm filaments. During the terminal stage of sperm development, no cell wall polymers recognized by the probes employed were found on the scale-covered sperm cells. CONCLUSIONS: Antheridium development in C. corallina is a rapid event that includes the production of cell walls that contain polymers similar to those found in land plants. While pectic and cellulosic epitopes are ubiquitous in the antheridium, the distribution of arabinogalactan protein and xyloglucan epitopes is restricted to specific zones. Spermatogenesis also includes a major switch in the production of extracellular matrix macromolecules from cell walls to scales, the latter being a primitive extracellular matrix characteristic of green plants.

[Allelopathic effect of Corallina pilulifera on Heterosigma akashiwo and its responses to UV-B irradiation].

By the method of co-culture and using cell density as the main indicator, this paper studied the allelopathic effect of Corallina pilulifera on Heterosigma akashiwo and its responses to UV-B irradiation. Under normal condition, the fresh tissue and aqueous extracts of C. pilulifera had significant inhibitory effects on the growth of H. akashiwo (P < 0.05), indicating their allopathic effect on H. akashiwo, while the dry power and culture media filtrate of C. pilulifera had less effect (P > 0.05). After pre-treated with different dose UV-B radiation and then co-cultured with H. akashiwo, C. pilulifera had some changes in the allelopathic activity of its fresh tissue, dry powder, and aqueous extracts. High-dose UV-B radiation (3.0 J x m(-2)) induced the decrease of the allelopathic effect, whereas low-dose UV-B radiation (0.9 J x m(-2)) was in adverse (P < 0.05).

Calcium pectate chemistry causes growth to be stored in Chara corallina: a test of the pectate cycle.

Calcium pectate chemistry was reported to control the growth rate of cells of Chara corallina, and required turgor pressure (P) to do so. Accordingly, this chemistry should account for other aspects of growth, particularly the ability of plants to compensate for brief exposure to low P, that is, to 'store' growth. Live Chara cells or isolated walls were attached to a pressure probe, and P was varied. Low P caused growth to be inhibited in live cells, but when P returned to normal (0.5 MPa), a flush of growth completely compensated for that lost at low P for as long as 23-53 min. This growth storage was absent in isolated walls, mature cells and live cells exposed to cold, indicating that the cytoplasm delivered a metabolically derived growth factor needing P for its action. Because the cytoplasm delivered pectate needing P for its action, pectate was supplied to isolated walls at low P as though the cytoplasm had done so. Growth was stored while otherwise none occurred. It was concluded that a P-dependent cycle of calcium pectate chemistry not only controlled growth rate and new wall deposition, but also accounted for stored growth.

[Allelopathic effects of Corallina pilulifera on red tide microalgae Heterosigma akashiwo].

Different concentration methanol-, acetone-, ether-, and chloroform extracts of Corallina pilulifera were used to study their growth inhibitory effects on red tide microalgae Heterosigma akashiwo. The results showed that methanol extract at relatively higher concentrations had the highest growth inhibitory activity and killed all H. akashiwo cells, while the other three kinds of organic solvent extracts had no apparent inhibitory effects, suggesting that the growth inhibitory substances in C. pilulifera had relatively high polarity. The methanol extract was partitioned to petroleum ether phase, ethyl acetate phase, butanol phase, and distilled water phase by liquid-liquid fractionation, and the bioassays on the activity of each fraction were carried out on H. akashiwo. It was found that petroleum ether phase and ethyl acetate phase had strong algicidal effects on H. akashiwo, suggesting that the fatty acids in C. pilulifera tissues might be one of the main allelochemicals.

Tension required for pectate chemistry to control growth in Chara corallina.

Recent work showed that polygalacturonate (pectate) chemistry controlled the growth rate of the large-celled alga Chara corallina when turgor pressure (P) was normal (about 0.5 MPa). The mechanism involved calcium withdrawal from the wall by newly supplied pectate acting as a chelator. But P itself can affect growth rate. Therefore, pectate chemistry was investigated at various P. A pressure probe varied P in isolated walls, varying the tension on the calcium pectate cross-links bearing the load of P. When soluble pectate was newly supplied, the wall grew irreversibly but the pectate was inactive below a P of 0.2 MPa, indicating that tension was required in the existing wall before new pectate acted. It was suggested that the tension distorted some of the wall pectate (the dominant pectin), weakening its calcium cross-links and causing the calcium to be preferentially lost to the new pectate, which was not distorted. The preferential loss provided a molecular mechanism for loosening the wall structure, resulting in faster growth. However, the resulting relaxation of the vacated wall pectate would cause calcium to be exchanged with load-bearing calcium pectate nearby, auto-propagating throughout the wall for long periods. There is evidence for this effect in isolated walls. In live cells, there is also evidence that auto-propagation is controlled by binding the newly supplied pectate (now calcium pectate) to the wall and/or by additional Ca(2+) entering the wall structure. A tension-dependent cycle of pectate chemistry thus appeared to control growth while new wall was deposited as a consequence.

Calcium pectate chemistry controls growth rate of Chara corallina.

Pectin, a normal constituent of cell walls, caused growth rates to accelerate to the rates in living cells when supplied externally to isolated cell walls of Chara corallina. Because this activity was not reported previously, the activity was investigated. Turgor pressure (P) was maintained in isolated walls or living cells using a pressure probe in culture medium. Pectin from various sources was supplied to the medium. Ca and Mg were the dominant inorganic elements in the wall. EGTA or pectin in the culture medium extracted moderate amounts of wall Ca and essentially all the wall Mg, and wall growth accelerated. Removing the external EGTA or pectin and replacing with fresh medium returned growth to the original rate. A high concentration of Ca2+ quenched the accelerating activity of EGTA or pectin and caused gelling of the pectin, physically inhibiting wall growth. Low pH had little effect. After the Mg had been removed, Ca-pectate in the wall bore the longitudinal load imposed by P. Removal of this Ca caused the wall to burst. Live cells and isolated walls reacted similarly. It was concluded that Ca cross-links between neighbouring pectin molecules were strong wall bonds that controlled wall growth rates. The central role of Ca-pectate chemistry was illustrated by removing Ca cross-links with new pectin (wall "loosening"), replacing vacated cross-links with new Ca2+ ("Ca2+-tightening"), or adding new cross-links with new Ca-pectate that gelled ("gel tightening"). These findings establish a molecular model for growth that includes wall deposition and assembly for sustained growth activity.

Growth towards light as an adaptation to high light conditions in Chara branches.

Growth of plants or plant organs towards more light is commonly interpreted as an adaptation to low light conditions. Here, we show for the first time, in a study of charophyte branches, a growth-based orientation towards light functioning as a mechanism to protect the plant from excessive light. Two Chara species were exposed to five different intensities of photosynthetically active radiation and species traits and pigmentation were measured. Branches of plants exposed to higher light intensities were convergent and pointed steeply upwards, whereas those exposed to lower light intensities grew nearly straight and were less inclined. Only branches that increased in length during the experiments reacted to differences in light intensity. This indicates that branch orientation is determined by a light-dependent growth reaction. Orientation of charophyte branches towards light is accompanied by a decrease in chlorophyll a (Chla) content and a lower Chla : carotenoid ratio, which clearly indicates that the plant is taking protective measures against potentially damaging excess light conditions. We suggest that the growth-based orientation of Chara branches towards light may protect sexual organs, which grow on adaxial branch sides, from light damage. In addition, the upward orientation of branches might lead to increased light transmission within dense charophyte beds, thus enabling an enhanced gross production.