10.1 Overview of the Scaled-Up Model Ecosystem

10.1.1 Species Composition

A scaled-up model ecosystem can be constructed by introducing submerged plants, fish, bivalve mollusks, shrimp, and other organisms into the microcosm N-system , which consists of conventional producers , predators, and decomposers (Takagi et al. 1994; Takamatsu et al. 1995; Inamori and Takamatsu 1995). The model ecosystem with aquatic animals and plants is shown in Fig. 10.1. Representative test organisms in temperate areas include Egeria densa, a submerged plant; Anodonta woodiana, a freshwater bivalve mollusk; and the fish Rhodeus ocellatus and Pseudorasbora parva. With the coexistence of these organisms, creating an ecosystem through the combination of submerged plants and aquatic animals, the construction of a scaled-up model for co-culturing large aquatic organisms was conducted. It was then possible to analyze the influence of chemical substances on the selected model ecosystem. The species used in this scaled-up model ecosystem included Potamogeton malaianus, Vallisneria denseserrulata, Elodea nuttallii, Potamogeton dentatus, and Potamogeton pusillus as producers ; the fish Rhinogobius flumineus; and the arthropod predator Caridina multidentata . A stable model ecosystem hydrosphere can be constructed using the appropriate combination of species.

Fig. 10.1
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Overview of scaled-up model ecosystem

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10.1.2 Culture Vessel and Culture Conditions

An example of an ecosystem model using aquatic animals and plants is shown in Fig. 10.2. The experimental apparatus should contain glass beads (1 L) for planting submerged plants and 4 L of lake water in a 5 L beaker. The apparatus should be installed in a thermostatic chamber at 20 °C that blocks light from the outside. An irradiance of 5000 lux and fluorescent lamps that can be adjusted to create day and night (L/D) cycles should be used.

Fig. 10.2
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Water plants and animals supplied to scaled-up model ecosystem and its culture system

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10.2 Chemical Substance Addition Experiment

10.2.1 LAS

An experiment was performed for the purpose of constructing a standard by planning the construction of the model ecosystem (scaled-up model ecosystem) hydrosphere using larger aquatic organisms in addition to the microcosm N-system, evaluating the risk associated with the introduction of chemical substances in the model ecosystem. To construct an optimum, scaled-up model hydrosphere ecosystem, which consisted of plants, fish, bivalve mollusks, and crustaceans, the abundance input organisms, submerged plants, feed quantity, and bottom sediment quality were selected. Additionally, experiments were conducted to determine whether it is possible to maintain the number of initial input organisms with a constant abundance of individuals under the 24-h light condition without setting an L/D cycle. As a result, there were periods in which input organisms could survive continuously for more than 1 week, but high mortality indicates that the experimental aquatic ecosystem was far from stable. Therefore, when the abundance of organisms varied due to mortality, an experiment for evaluating the number of viable organisms was conducted without introducing new organisms. As a result, there were no fatalities for 10 days for three Rhodeus ocellatus, two Rhinogobius sp., four Caridina multidentata, and two Anodonta woodiana at the time that the powdered potato feed was supplied under the experimental condition with the highest survivorship. We determined abundance to be a viable input. Furthermore, the viable abundance under the L/D = 12 hr./12 hr. condition that was similar to natural conditions was evaluated, and the model ecosystem hydrosphere in the 5 L plastic tank (i.e., the scaled-up model ecosystem) included three Rhodeus ocellatus, one Rhinogobius sp., three Caridina multidentata, and two Anodonta woodiana based on laboratory findings. Vallisneria denseserrulata was planted, supplied with powder feed, and subjected to filling by pollutants + humus and black soil + sand.

To evaluate the influence of the chemical substance LAS on the model ecosystem hydrosphere, LAS was added to the system at 0, 1, 5, and 10 mg/L. Based on fluctuations in the P/R ratio and the persistence of aquatic organisms, an impact assessment of the model ecosystem was conducted. As shown in Fig. 10.3, it was assumed that the functional parameter (P/R ratio) in the experiment with the powder feed supply, which fluctuated rapidly only when 10 mg/L was added, and some other effects were latent. Also, in the structural parameters (i.e., persistence of aquatic animals), from the first day to the third day after the addition of 10 mg/L, two Rhodeus ocellatus, one Rhinogobius sp., and one Anodonta woodiana died. The risk at 10 mg/L became obvious, and the deaths of one Anodonta woodiana on the first day at 5 mg/L and one Rhodeus ocellatus on the seventh day at 1 mg/L were also confirmed. The results of the P/R ratio of the experiment without the supply of powder feed exhibited no change with the addition of any amount of LAS , and it was determined that there was no influence. With respect to the persistence of the aquatic animals, one Rhodeus ocellatus each died on the first and second day with the 10 mg/L addition and with the 1 mg/L addition, and one Anodonta woodiana and one Caridina multidentata died on the second and third day, respectively.

Fig. 10.3
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Time course of P/R ratio in LAS added scaled-up model ecosystem

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Catabolite suppression was considered to have possibly occurred due to the difference between experiments with a feed supply and those without a feed supply. With catabolite restraint, rapid metabolism of organic matter is a phenomenon known to restrain the generation of the enzyme necessary for stable organic metabolism. Powder feed quantity and the inspection of the metabolic rate of LAS are necessary to determine whether feeding resulted in more sudden changes in the P/R ratio than when there was no feed supplied. Additionally, as a result of changing the quality of the glass beads on the bottom and having performed a similar experiment, it was determined that, when the P/R ratio and the mortality of aquatic organisms were assessed, there was a risk for LAS = 10 mg/L. Plasticity was required, but it was made clear that the NOEC of the LAS estimated was less than 5 mg/L.

10.2.2 AE

A scaled-up microcosm as a model aquatic ecosystem, including aquatic animals and plants, which was nearer to the conditions of a real ecosystem than was the microcosm N-system was constructed, and the ecosystem risk posed by a nonionic surfactant , alcohol ethoxylate (AE) , as a chemical substance was assessed. As a result of examining the appropriate combination of large aquatic animals and plants, a stable, large-scale model ecosystem was constructed as the ecological impact assessment system in which Rhodeus ocellatus and Rhinogobius sp. (aquatic fish) and Egeria densa (a submerged plant) coexisted as consumers and a producer , and the P/R ratio was approximately constant at 1. Additionally, it may be useful to construct a stable ecosystem index using the P/R ratio. The NOEC of AE in this scaled-up aquatic microcosm was estimated as 2 mg/L. As a result of AE addition to the scaled-up microcosm, aquatic animals were not affected up to 2 mg/L of AE, and the submerged plant was not affected up to 5 mg/L of AE. The influence of AE was felt in the scaled-up microcosm at additions of 3–5 mg/L, and the P/R ratio increased above 1, exhibiting similar behavior to the system that included submerged plants only.

Using consecutive measurements of the DO in the scaled-up microcosm, ecological balance and a steady state can be informed, and the persistence of aquatic populations can be judged. An activity state can be quickly derived from the changing pattern (inspection of the recovery accuracy) of the DO level or the P/R ratio. The possibility of evaluating ecological risk from the DO pattern and the P/R ratio, without measuring the population of the aquatic organisms and using a continual measurement of DO, was illustrated. The NOEC of this scaled-up microcosm was nearly equal to the m-NOEC of the N-system, which was correlated with the flask microcosm test as shown in Fig. 10.4. This indicates that, even if the diversity and hierarchical structure of the organisms change, there is no substantial difference in the influence concentration. More specifically, the validity and effectiveness of the microcosm N-system was demonstrated by the experimental results of the scaled-up microcosm system (Fig. 10.4).

Fig. 10.4
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Time course of P/R ratio in AE added scaled-up model ecosystem

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10.3 Comparison with the Microcosm N-System

In the reports of Sugiura (Sugiura 2009, 2010), it was presumed that, in a microcosm where the production and respiration can be balanced, even if the diversity of the constituent species and the hierarchical structure are changed somewhat, there is no substantial difference in the influence of the concentration. The correspondence between the NOEC in the flask-scale microcosm and the large-scale ecosystem model, with larger aquatic animals and plants, suggests that it is possible to predict the difference in the influence of the concentration across different ecosystem scales and species compositions by collecting basic data. Furthermore, in the large-scale ecosystem model, it is possible to evaluate not only the fish (predators) but also the influence of aquatic plants (producers ) on ecosystem functions. It is a great advantage that the survivorship/mortality of the aquatic animal populations can be quickly determined from the changes in the DO value or the P/R ratio. By collecting such basic data, it is possible to ensure the accuracy of the correlation between these model ecosystems and natural ecosystems. As a result, it is expected that the NOEC of chemical substances can be accurately predicted for natural ecosystems using large-scale models of aquatic ecosystem. It is also expected to be useful as a tool for managing chemical substances, including surfactants .

Based on the results of experiments with the addition of LAS and AE , the ecosystem function parameters (DO and P/R ratio) in the scaled-up model ecosystem were consistent with the microcosm N-system. Therefore, the microcosm N-system adequately reflects the natural ecosystem, and it was shown that it is extremely useful as a model of ecosystem functions. In the OECD test method, one representative species from each trophic level of the ecosystem was selected (e.g., Selenastrum capricornutum (producer ), Ceriodaphnia dubia (primary consumer), Danio rerio (high-order consumer ), bacterial luminescence inhibition test (decomposer ), etc.) to evaluate toxicity. The ecosystem effect is then calculated based on the toxicity data for the most sensitive species. In natural ecosystems, biological interactions, material circulation, and energy flows exist and differ greatly from the physiological activities of test organisms in single-species tests. The evaluation of toxicity under different conditions of physiological activity is inadequate, especially for impact assessments of ecosystem function; thus, the necessity of multi-species tests is noted at the OECD. Meanwhile, mesocosm tests, in which natural environmental water is sealed in a container or in which part of a natural ecosystem is isolated, suffer from problems in terms of cost, reproducibility, and handling. The mesocosm test seems to be meaningful in predicting the behavior of chemical substances evaluated in the presence or absence of higher predators in the ecosystem. However, the test method in which fish and other organisms are added is considered to not be useful because it is inferior to the flask microcosm in terms of cost, speed, and reliability. Considering these points, it can be said that the particle size of the microcosm N-system is extremely useful as a multi-species coexisting system, with high reproducibility and stability.