Phytotoxicity and accumulation of Cu in mature and young leaves of submerged macrophyte Hydrilla verticillata (L.f.) Royle.

Phytotoxicity and accumulation of Cu in mature and young leaves of submerged macrophyte Hydrilla verticillata (L.f.) Royle were investigated by analyzing the chlorophyll contents, chloroplast ultrastructure and leaf surface structure under different Cu treatments (0, 0.01, 0.05 and 0.1 f mg L-1). The results showed that 0.05 and 0.1 mg L-1 Cu treatment decreased the contents of Chl a and Chl b, and caused damage on leaf surface structure and chloroplast ultrastructure compared with control (0 mg L-1 Cu treatment). Higher concentration of Cu induced Chlorophyll decreases and the damages on the leaf surface structure and chloroplast ultrastructure were more pronounced in mature than in young leaves. It was observed that leaf Cu concentration increased almost linearly with exposure time and majority of the Cu accumulated in the cell walls. Among different cell wall fractions, the majority of Cu accumulated in cell walls was bound to the hemicellulose 1 and cellulose, followed by the pectin, hemicellulose 2. Mature leaves had significantly higher the concentrations of total Cu and bound-Cu in cell walls due to higher uronic acid content in their cell wall fractions (pectin, hemicellulose 1 and cellulose) than young leaves. Distinct cell wall composition might partially contribute to the different Cu toxicity and accumulation between mature and young leaves of submerged macrophyte H. verticillata. Our results show that mature leaves are more efficient in the uptake and accumulation of Cu than young leaves, which might explain why mature leaves sustain more severe damage.

Response of the submerged macrophytes Vallisneria natans to snails at different densities.

The study investigated the responses of the submerged macrophyte Vallisneria natans (V. natans) to snails (Bellamya aeruginosa) at different densities, with changes in physiological parameters, morphology, leaf-epiphytic bacteria community and water quality parameters examined. The changes of water quality parameters (pH, total nitrogen (TN), total phosphorus (TP) and total organic carbon (TOC)) indicated that snails secreted nutrients into water. Changes in morphological and physiological parameters (fresh weight, root length, shoot height, chlorophyll, malondialdehyde (MDA), activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD)) demonstrated that the presence of snails were beneficial to the growth of submerged macrophytes. Microbial diversity analyses indicated that snails could decrease microbial community richness and diversity. At medium densities (340 ind. m-2), an increase in snail density was beneficial to the growth of submerged macrophytes. The results of this study provide theoretical guidance and technical support for the maintenance and restoration of submerged macrophytes.

Sex-specific plasticity of reproductive allocation in response to water depth in a clonal, dioecious macrophyte.

PREMISE OF THE STUDY: Sex-specific differences in reproductive investment contribute to sexual dimorphism in dioecious plants. Along environmental gradients, males and females may plastically adjust reproductive allocation differently because of contrasting reproductive costs. In dioecious macrophytes, variation in water depth is likely to influence reproductive allocation but has not been investigated in detail. METHODS: Vallisneria spinulosa was grown in aquatic mesocosms at water depths of 50, 100 and 150 cm for 14 weeks. Plasticity in allocation was measured to investigate whether sexual dimorphism in reproductive allocation and vegetative growth changed in response to varying water depths. KEY RESULTS: Females invested a higher fraction of resources to sexual reproduction than males across all water depths and decreased proportional allocation to sexual structures in shallow and deep water compared to intermediate water depth. In contrast, males maintained similar sexual allocation across all water depths. Females displayed larger vegetative size than males, despite greater sexual investment, but decreased vegetative biomass more than males in shallow or deep water. The sexes invested similarly in clonal propagation by tubers at all water depths, but a trade-off with sexual reproduction was only evident in females. CONCLUSIONS: Our results suggest that females of V. spinulosa have mechanisms to compensate for the costs of sexual reproduction in heterogeneous environments. Compared to males, females expressed greater plasticity in biomass allocated to sexual reproduction and vegetative growth in response to water depth variation. Environmental variation in underwater light availability probably caused the sex-specific allocation strategies found in V. spinulosa.

Responses of Vallisneria natans (Lour.) Hara to the combined effects of Mn and pH.

Aquatic plants play a vital role in maintaining the health and stability of ecosystems and in ecological restoration of contaminated water bodies. Herein, a 21-day-long laboratory-scale experiment was designed to explore the growth and physiological responses of Vallisneria natans (Lour.) Hara (V. natans) to the combined effects of manganese (Mn, 5, 20, and 80 mg L-1) and pH (pH 4.0, 5.5, and 7.0). Our results showed the combined toxicity intensity was closely related to Mn concentration and the toxicity exhibited by Mn gradually strengthened with the decrease of pH level. High concentration of Mn stress significantly reduced plants leaf area, final leaf number, photosynthetic pigment content, RGR (relative growth rate) and biomass accumulation, but significantly increased the contents of malondialdehyde (MDA) and hydrogen peroxide (H2O2). At the same time, V. natans plants can resist the adverse stress by activating the antioxidant defense system, including superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and polyphenol oxidase (PPO) activities. Besides, V. natans tended to adjust the biomass allocation strategy and transferred more energy to the subsurface and the ramets and stolons parts under the combined stress. This experiment also showed that the increasing pH within a certain range could largely improve the removal rate of Mn (at highest by 84.28%). This may indicate the V. natans plant species can act as a promising tool for the Mn phytoremediation in aquatic environments which needs to be further explored by longer cycle field studies.

Responses of Aquatic Organisms to Cyanobacteria and Elodea in Microcosms.

The aim of the present study is investigation of the impact of mass development of cyanobacteria and Elodea on population characteristics (abundance, biomass, fecundity) and "health" parameters (the state of embryos, heart rate, and thermal resistance) of aquatic invertebrates in experiments with controlled conditions. In July-August 2018, in four variants of microcosms, the conditions were modeled characteristic of the coastal zone of the Rybinsk Reservoir in the period of mass development of cyanobacteria. Mass species were placed in microcosms of planktonic (with domination (80%) Daphnia longispina) and bottom animals (Unio pictorum, Gmelinoides fasciatus, Asellus aquaticus), as well as Elodea canadensis. In the variants with cyanobacteria, the toxic microcystins LR were revealed (24-53% of the sum of toxins, or from 6.6-66.5 µg/L). The presence of cyanobacteria (Aphanizomenon flos-aquae, Microcystis aeruginosa, and Gloeoitrichia spp.) influenced structural parameters of plankton: the quantity to green algae decreased, small-sized crustaceans increased. Influence of cyanobacteria caused reliable changes in parameters of aquatic animal from those of the control: thermal resitance of mollusks and amphipods decreased, time of restoration of heart rate after loads (salinity test) in mollusks increased; in amphipods, individual fecundity decreased, frequency of abnormalities and of mortality of embryos increased. The presence of Elodea intensified these effects. Thus, mass development of cyanobacteria and Elodea influences ecological-physiological parameters of aquatic animals, decreasing their adaptivity to natural stresses and deteriorating the quality of the progeny.

A functional-structural model of ephemeral seagrass growth influenced by environment.

Background and Aims: Ephemeral seagrasses that respond rapidly to environmental changes are important marine habitats. However, they are under threat due to human activity and are logistically difficult and expensive to study. This study aimed to develop a new functional-structural environmentally dependent model of ephemeral seagrass, able to integrate our understanding of ephemeral seagrass growth dynamics and assess options for potential management interventions, such as seagrass transplantation. Methods: A functional-structural plant model was developed in which growth and senescence rates are mechanistically linked to environmental variables. The model was parameterized and validated for a population of Halophila stipulacea in the Persian Gulf. Key Results: There was a good match between empirical and simulated results for the number of apices, net rhizome length or net number of internodes using a 330 d simulation. Simulated data were more variable than empirical data. Simulated structural patterns of seagrass rhizome growth qualitatively matched empirical observations. Conclusions: This new model successfully simulates the environmentally dependent growth and senescence rates of our case-study ephemeral seagrass species. It produces numerical and visual outputs that help synthesize our understanding of how the influence of environmental variables on plant functional processes affects overall growth patterns. The model can also be used to assess the potential outcomes of management interventions like seagrass transplantation, thus providing a useful management tool. It is freely available and easily adapted for new species and locations, although validation with more species and environments is required.

Engineered nanoparticles interact with nutrients to intensify eutrophication in a wetland ecosystem experiment.

Despite the rapid rise in diversity and quantities of engineered nanomaterials produced, the impacts of these emerging contaminants on the structure and function of ecosystems have received little attention from ecologists. Moreover, little is known about how manufactured nanomaterials may interact with nutrient pollution in altering ecosystem productivity, despite the recognition that eutrophication is the primary water quality issue in freshwater ecosystems worldwide. In this study, we asked two main questions: (1) To what extent do manufactured nanoparticles affect the biomass and productivity of primary producers in wetland ecosystems? (2) How are these impacts mediated by nutrient pollution? To address these questions, we examined the impacts of a citrate-coated gold nanoparticle (AuNPs) and of a commercial pesticide containing Cu(OH)2 nanoparticles (CuNPs) on aquatic primary producers under both ambient and enriched nutrient conditions. Wetland mesocosms were exposed repeatedly with low concentrations of nanoparticles and nutrients over the course of a 9-month experiment in an effort to replicate realistic field exposure scenarios. In the absence of nutrient enrichment, there were no persistent effects of AuNPs or CuNPs on primary producers or ecosystem productivity. However, when combined with nutrient enrichment, both NPs intensified eutrophication. When either of these NPs were added in combination with nutrients, algal blooms persisted for >50 d longer than in the nutrient-only treatment. In the AuNP treatment, this shift from clear waters to turbid waters led to large declines in both macrophyte growth and rates of ecosystem gross primary productivity (average reduction of 52% ± 6% and 92% ± 5%, respectively) during the summer. Our results suggest that nutrient status greatly influences the ecosystem-scale impact of two emerging contaminants and that synthetic chemicals may be playing an under-appreciated role in the global trends of increasing eutrophication. We provide evidence here that chronic exposure to Au and Cu(OH)2 nanoparticles at low concentrations can intensify eutrophication of wetlands and promote the occurrence of algal blooms.

Submerged freshwater plant communities do not show species complementarity effect in wetland mesocosms.

It is a generally accepted theory that ecological functions are enhanced with increased diversity in plant communities due to species complementarity effects. We tested this theory in a mesocosm study using freshwater submerged plant beds to determine if increasing species number caused overyielding and species complementarity. We applied a maximum of four species in the plant beds corresponding to the typical species number in natural freshwater plant beds. We found no clear effects of species number (1-4) on biomass production and thus no conclusive overyielding and complementarity effect. This may be explained by low species differentiation among the four species in plant traits relevant for resource acquisition in freshwater, or that other species interactions, e.g. allelopathy, were inhibiting overyielding. The existing knowledge on species complementarity in aquatic plant communities is sparse and inconclusive and calls for more research.

Floral development of petaloid Alismatales as an insight into the origin of the trimerous Bauplan in monocot flowers.

Monocots are remarkably homogeneous in sharing a common trimerous pentacyclic floral Bauplan. A major factor affecting monocot evolution is the unique origin of the clade from basal angiosperms. The origin of the floral Bauplan of monocots remains controversial, as no immediate sister groups with similar structure can be identified among basal angiosperms, and there are several possibilities for an ancestral floral structure, including more complex flowers with higher stamen and carpel numbers, or strongly reduced flowers. Additionally, a stable Bauplan is only established beyond the divergence of Alismatales. Here, we observed the floral development of five members of the three 'petaloid' Alismatales families Butomaceae, Hydrocharitaceae, and Alismataceae. Outer stamen pairs can be recognized in mature flowers of Alismataceae and Butomaceae. Paired stamens always arise independently, and are either shifted opposite the sepals or close to the petals. The position of stamen pairs is related to the early development of the petals. In Butomaceae, the perianth is not differentiated and the development of the inner tepals is not delayed; the larger inner tepals (petals) only permit the initiation of stamens in antesepalous pairs. Alismataceae has delayed petals and the stamens are shifted close to the petals, leading to an association of stamen pairs with petals in so-called stamen-petal complexes. In the studied Hydrocharitaceae species, which have the monocot floral Bauplan, paired stamens are replaced by larger single stamens and the petals are not delayed. These results indicate that the origin of the floral Bauplan, at least in petaloid Alismatales, is closely linked to the position of stamen pairs and the rate of petal development. Although the petaloid Alismatales are not immediately at the base of monocot divergence, the floral evolution inferred from the results should be a key to elucidate the origin of the floral Bauplan of monocots.

Effects of the decomposing liquid of Cladophora oligoclona on Hydrilla verticillata turion germination and seedling growth.

Excessive proliferation of filamentous green algae (FGA) has been considered an important factor resulting in the poor growth or even decline of submerged macrophytes. However, there is a lack of detailed information regarding the effect of decaying FGA on submerged macrophytes. This study aimed to investigate whether the decomposing liquid from Cladophora oligoclona negatively affects Hydrilla verticillata turion germination and seedling growth. The results showed that the highest concentrations of decomposing liquid treatments inhibited the turion germination rate, which was the lowest than other treatments, at only 84%. The chlorophyll a fluorescence (JIP test) and physiological indicators (chlorophyll a content, soluble sugars, Ca2+/Mg2+-ATPase and PAL activity) were also measured. The chlorophyll a content in the highest concentration (40% of original decomposing liquid) treatment group decreased by 43.53% than that of the control; however, soluble sugars, Ca2+/Mg2+-ATPase, and PAL activity increased by 172.46%, 271.19%, and 26.43% respectively. The overall results indicated that FGA decay has a considerable effect on submerged macrophyte turion germination and seedling growth, which could inhibit their expansion and reproduction. This study emphasized the need to focus on effects of FGA decomposition on the early growth stages of submerged macrophytes and offered technological guidance for submerged vegetation restoration in lakes and shallow waters.

Phytotoxic effects of Cu, Cd and Zn on the seagrass Thalassia hemprichii and metal accumulation in plants growing in Xincun Bay, Hainan, China.

Seagrasses play an important role in coastal marine ecosystems, but they have been increasingly threatened by human activities. In recent years, seagrass communities have rapidly degenerated in the coastal marine ecosystems of China. To identify the reasons for the decline in seagrasses, the phytotoxic effects of trace metals (Cu, Cd and Zn) on the seagrass Thalassia hemprichii were investigated, and the environmental contents of the metals were analyzed where the seagrass grows. The results showed that leaf necrosis in T. hemprichii exposed to 0.01-0.1 mg L-1 of Cu2+ for 5 days was more serious than that in plants exposed to the same concentrations of Cd2+ and Zn2+. The chlorophyll content in T. hemprichii declined in a concentration-dependent manner after 5 days of exposure to Cu2+, Cd2+ and Zn2+. The evident reduction in ΔF/Fm' in T. hemprichii leaves was observed at day 1 of exposure to 0.01-1.0 mg L-1 of Cu2+ and at day 3 of exposure to 0.1-1.0 mg L-1 of Cd2+. The antioxidant enzyme activities (SOD, POD and CAT) in T. hemprichii leaves exposed to the three metal ions also showed significant changes. In seawater from Xincun Bay (Hainan, China), where T. hemprichii grows, Cu had reached a concentration (i.e., 0.01 mg L-1) that could significantly reduce chlorophyll content and ΔF/Fm' in T. hemprichii leaves. Our results indicate that Cu influences the deterioration of seagrasses in Xincun Bay.

Effects of Planted Versus Naturally Growing Vallisneria natans on the Sediment Microbial Community in West Lake, China.

Submerged macrophytes play an important role in aquatic ecosystems, which has led to an increase in studies on vegetation recovery in polluted lakes from which submerged macrophytes have disappeared. The comparison of microbial communities in sediment cloned with planted and naturally growing submerged macrophytes is an interesting but rarely studied topic. In this investigation, Maojiabu and Xilihu, two adjacent sublakes of West Lake (Hangzhou, China), were selected as aquatic areas with planted and naturally growing macrophytes, respectively. Sediment samples from sites with/without Vallisneria natans were collected from both sublakes. The results showed that sediment total nitrogen and organic matter were significantly lower in the plant-covered sites than that in the non-plant sites in Maojiabu. Additionally, the sediment microbial community characterized by 16S ribosomal RNA (rRNA) sequencing differed more significantly for Maojiabu than for Xilihu. The relative abundances of microbes involved in C, N, and S elemental cycling were significantly higher in the sediments with plants than in those without. Results from both fatty acid methyl ester analysis and 16S rRNA sequencing indicated that vegetation significantly influenced the sulfate-reducing bacteria (SRB). Thus, the gene copies and composition of SRB were explored further. The relative gene abundance of SRB was 66% higher with natural vegetation colonization but was not influenced by artificial colonization. An increase in dominant SRB members from the families Syntrophobacteraceae and Thermodesulfovibrionaceae contributed to the increase of total SRB. Thus, macrophyte planting influences sediment nutrient levels and microbial community more than natural growth does, whereas the latter is more beneficial to sediment SRB.

Sound speed and attenuation measurements within a seagrass meadow from the water column into the seabed.

In situ measurements of sound speed and attenuation at 50 kHz were conducted in a Thalassia testudium meadow. Measurements were obtained at discrete depths in the water column, in the seagrass canopy, and in the sediment beneath the seagrass. Measurements were also obtained in bare sediment located a few meters away. Sediment biomass abundance was measured from cores collected at each site. Even though the measurements were obtained in the dormant season (winter), significant differences in sound speed and attenuation were observed in the sediment beneath the seagrass bed compared to the bare sediment.

Mowing Submerged Macrophytes in Shallow Lakes with Alternative Stable States: Battling the Good Guys?

Submerged macrophytes play an important role in maintaining good water quality in shallow lakes. Yet extensive stands easily interfere with various services provided by these lakes, and harvesting is increasingly applied as a management measure. Because shallow lakes may possess alternative stable states over a wide range of environmental conditions, designing a successful mowing strategy is challenging, given the important role of macrophytes in stabilizing the clear water state. In this study, the integrated ecosystem model PCLake is used to explore the consequences of mowing, in terms of reducing nuisance and ecosystem stability, for a wide range of external nutrient loadings, mowing intensities and timings. Elodea is used as a model species. Additionally, we use PCLake to estimate how much phosphorus is removed with the harvested biomass, and evaluate the long-term effect of harvesting. Our model indicates that mowing can temporarily reduce nuisance caused by submerged plants in the first weeks after cutting, particularly when external nutrient loading is fairly low. The risk of instigating a regime shift can be tempered by mowing halfway the growing season when the resilience of the system is highest, as our model showed. Up to half of the phosphorus entering the system can potentially be removed along with the harvested biomass. As a result, prolonged mowing can prevent an oligo-to mesotrophic lake from becoming eutrophic to a certain extent, as our model shows that the critical nutrient loading, where the lake shifts to the turbid phytoplankton-dominated state, can be slightly increased.

New data on the effect of ionizing radiation on the growth of the aquatic plant Elodea canadensis in the laboratory.

Toxicological experiments with radioactive bottom sediments and extrinsic γ-irradiation have demonstrated that the growth of common elodea roots is suppressed by irradiation at doses several times lower than the established threshold dose. The effect of γ-irradiation on the growth of elodea stems has not been observed at any dose used. The data obtained show that elodea could be recommended for use as an indicator of biological effects of radiation in the range of low radiation doses.

Integrating subsistence practice and species distribution modeling: assessing invasive elodea's potential impact on Native Alaskan subsistence of Chinook salmon and whitefish.

Alaska has one of the most rapidly changing climates on earth and is experiencing an accelerated rate of human disturbance, including resource extraction and transportation infrastructure development. Combined, these factors increase the state's vulnerability to biological invasion, which can have acute negative impacts on ecological integrity and subsistence practices. Of growing concern is the spread of Alaska's first documented freshwater aquatic invasive plant Elodea spp. (elodea). In this study, we modeled the suitable habitat of elodea using global and state-specific species occurrence records and environmental variables, in concert with an ensemble of model algorithms. Furthermore, we sought to incorporate local subsistence concerns by using Native Alaskan knowledge and available statewide subsistence harvest data to assess the potential threat posed by elodea to Chinook salmon (Oncorhynchus tshawytscha) and whitefish (Coregonus nelsonii) subsistence. State models were applied to future climate (2040-2059) using five general circulation models best suited for Alaska. Model evaluations indicated that our results had moderate to strong predictability, with area under the receiver-operating characteristic curve values above 0.80 and classification accuracies ranging from 66 to 89 %. State models provided a more robust assessment of elodea habitat suitability. These ensembles revealed different levels of management concern statewide, based on the interaction of fish subsistence patterns, known spawning and rearing sites, and elodea habitat suitability, thus highlighting regions with additional need for targeted monitoring. Our results suggest that this approach can hold great utility for invasion risk assessments and better facilitate the inclusion of local stakeholder concerns in conservation planning and management.

Interaction of Tritium with Elodea canadensis and Lemna minor.

Tritium is the least toxic radionuclide. The main contribution into the total tritium content in ecosystems is made by technogenic tritium, which is due to the operation of nuclear fuel cycle enterprises. The tritium content in the ecosystem of the River Yenisei is connected with its background values as well as with tritium entering the water ecosystem as a result of the operation of the Mining and Chemical Combine, MCC Rosatom. Presented here are the investigations of the possible transformation of tritium interacting with certain species of aqueous plants - submerged rnacrophyte Elodea canadensis and an aqueous plant floating on the surface of water reservoirs Lemna minor. Elodea sampling was made in a real water reservoir - the River Yenisei, while lemna was grown in the laboratory conditions. The experiments show that with the chronic exposure of young elodea shoots to tritium, the latter transforms from HTO to OBT. Optimal conditions were also obtained for the maximum transformation of tritium ≈35% from the total content: at 25°C and the light period 6/18 (day/night). In the experiments with duckweed, observed a significant increase in area of fronds in introducing tritium into the system.

Acclimation of Hydrilla verticillata to sediment anoxia in vegetation restoration in eutrophic waters.

Sediment anoxia generally results from intense organic enrichment and is a limiting factor in the restoration of vegetation in eutrophic waters. To investigate the effect of sediment anoxia on a typical pollution-tolerant submerged macrophyte species, Hydrilla verticillata, and acclimation mechanisms in the plant, a gradient of sediment anoxia was simulated with additions of sucrose to the sediment, which can stimulate increased concentrations of total nitrogen, NH4(+) and Fe in pore water. H. verticillata growth was significantly affected by highly anoxic conditions, as indicated by reduced total biomass in the 0.5 and 1% sucrose treatments. However, slight anoxia (0.1% sucrose addition) promoted growth, and the shoot biomass was 22.64% higher than in the control. In addition to morphologic alterations, H. verticillata showed physiological acclimations to anoxia, including increased anaerobic respiration and changes in carbon and nitrogen metabolism in roots. The soluble protein and soluble carbohydrate contents in roots of the 1% treatment were both significantly higher compared with those in the control. The increase in alcohol dehydrogenase activity and pyruvate content in the roots suggested that H. verticillata has a well-developed capacity for anaerobic fermentation. This study suggests that highly anoxic sediments inhibit the growth of H. verticillata and the species has a degree of tolerance to anoxic conditions. Further in situ investigations should be conducted on the interactions between sediment conditions and macrophytes to comprehensively evaluate the roles of sediment in the restoration of vegetation in eutrophic waters.

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.

Cumulative effects of fish disturbance and vegetative propagules reduce the growth advantage of Vallisneria natans.

Benthivorous fish disturbance and density-dependent competition from adjacent macrophytes are two important biotic factors that significantly impact the growth of submerged macrophyte pioneer species, which is crucial for the success of eutrophication lake restoration. We conducted an outdoor mesocosm experiment to explore the individual and combined effects of these two factors on water quality and the growth of Vallisneria natans. The experiment involved two levels of fish (Misgurnus anguillicaudatus) disturbance crossed with two levels of Hydrilla verticillata vegetative propagule (shoot) intensity. The results showed that fish disturbance significantly increased the water column total nitrogen (TN), ammonia nitrogen (N-NH4), total phosphorus (TP), and phosphate­phosphorus (P-PO4). V. natans exhibited restricted plant height elongation and decreased soluble carbohydrate (SC) and starch concentration in fish treatments. Fish disturbance inhibited the growth advantage of V. natans by increasing the extinction coefficient of the water column. There was no statistical significance in total biomass between the two macrophytes in increased vegetative propagule and fish treatments. H. verticillata exhibited a higher relative growth rate (RGR) and summed dominance ratio (SDR3) than V. natans in four treatments and the treatment with three shoots of H. verticillata and one M anguillicaudatus, respectively. Fish disturbance and vegetative propagules showed cumulative effects that negatively affected the RGR_V.H (V. natans relative to H. verticillata). Our findings indicated that benthivorous fish disturbance and vegetative propagules could individually and cumulatively reduce the growth advantage of the pioneer species, V. natans. Our study sheds light on the accumulated effects of multiple disturbances that simultaneously occur in lakes, which holds theoretical and practical importance for lake restoration efforts.