Anthropogenic toxic chemicals can easily contaminate water bodies, and exert negative impacts on the ecosystems. Reference indexes for standard test organisms, such as NOEC (No observed effect concentration) and EC50 (50% effective concentration, meaning the concentration estimated to immobilize 50% of individuals) values obtained from laboratory toxicity tests (OECD 2004), have been used for the estimation of direct toxic impacts on target trophic level biocenose. On the other hand, community level experiments have elucidated indirect impacts of the chemicals through the modification of biological interactions (Preston 2002, Van Wijngaarden et al. 2005, Relyea and Hoverman 2006, Sarma and Nandini 2006).
Lake ecosystems are composed of many different species at different trophic levels. Cladocerans (zooplankton) are one of the groups most sensitive to insecticides. They are, however, superior herbivores in food competition with rotifers (MacIsaac and Gilbert 1989). Community level mesocosm/enclosure experiments have demonstrated that rotifer density increases after disruption of cladoceran populations at insecticide-contaminated sites (Chang et al. 2005). This is because rotifers are far more tolerant than cladocerans to insecticides (Havens and Hanazato 1993). Within the cladoceran group, results from mesocosm/enclosure experiments have suggested that large cladocerans tend to be more sensitive to insecticides than smaller ones (e.g.,
In contrast to the community level experiments, however, results from the individual-level laboratory tests have suggested that
The contradiction between community and individual level studies indicates that the vulnerability of each cladoceran species to insecticide can change greatly when a variety of species coexist and interact in a complex manner. However, there is insufficient information to explain the phenomenon.
Insecticides often affect the plankton communities indirectly by the modification of predator-prey interactions (Lurling and Scheffer 2007). For instance, increased swimming speed of the rotifer,
A carbamate insecticide, carbaryl (1-naphthyl-Nmethylcarbamate), was used in the present study since its toxicity to cladocerans had been well tested both in species- and community level studies (Hanazato 2001). Here, we investigated the following two hypotheses by a microcosm experiment: (1) the response to carbaryl of each cladoceran changes in the presence of competitive or predatory interactions; (2) disturbance of the predatory interactions is a possible cause yielding the inconsistency between community- and individual-level studies.
In order to compare the tolerances of plankton communities with different structures, two different zooplankton communities, dominated by
Six 20 L cylindrical polyethylene tanks (diameter, 30 cm; height, 31 cm) were used as microcosms for the preparation of zooplankton communities 14 days before the treatments (day -14). The zooplankton community was established from the resting eggs (or the resting stages of the animals) in bottom mud obtained from the central area of a eutrophic lake, Lake Suwa (36˚2′ N, 138˚5′E) Japan. In Lake Suwa, the zooplankton community is dominated by bosminid species, and no
clone, originally from Lake Kasumigaura Japan) were added into each of the 9 tanks (Fig. 1).
Carbaryl applications were conducted on day 11. The insecticide (>99% grade, CAS 63-25-2) was purchased from Wako Pure Chemical Industries Ltd., Japan. A stock solution of the reagent (1000 mg/L) was prepared by dissolving the chemical in 99% ethanol to a final volume of 10 mL, then diluted to the required concentrations with dechlorinated tap water, and introduced to the tanks. The established zooplankton communities were exposed to following five different treatments, each having three replications: (1) control (no carbaryl and no
To measure the absolute concentrations of carbaryl, 100 mL water was collected from each tank at 0, 3, 6 and 9 days after the application (day 11 to 20). Each water sample containing carbaryl was filtered through a Whatman GF/C filter, and passed through a solid-phase cartridge (PS-2 plus; Waters, Milford, MA, USA). The samples were eluted with 5 mL acetonitrile. The acetonitrile solutions were dried under a gentle nitrogen stream and redissolved in 500 μL acetonitrile for HPLC analysis. Parent carbaryl concentrations were determined by HPLC (LC-10A series; Shimadzu, Kyoto, Japan) with UV-VIS detector (SPD-10A; Shimadzu, Kyoto, Japan) equipped with an ODS column (Mightysil RP-18 GP 150 mm × 2.0 mm ？ (5 μm); Kanto Chemical Co., Inc., Tokyo, Japan).
Zooplankters were sampled before carbaryl application on days 0 and 11, and every third day thereafter by using a column sampler (diameter 5.5 cm, length 50 cm) with a hydraulically operated flap at the bottom (total volume: 1 L). Just before the samplings, DO and pH were monitored for each microcosm tanks. Collected water was filtered through a 40-μm mesh and the residue was fixed with sugar-containing formalin at a final concentration of 4% (Haney and Hall 1973). The fixed samples were concentrated to 5 mL by settling for over 6 h. Aliquots of 1 mL were used for counting rotifers and copepod nauplii. Whole samples were used for counting cladocerans and copepods. Zooplankton were identified to species (or genus) level and counted using a microscope. In addition, morphotypes (antennule types) of adult females of
Effects of the treatments on the repeatedly sampled zooplankton abundances, the morphologies of
For each microcosm, a time-weighted average (WA) was calculated for each response variable (carbaryl concentration, densities of zooplankton, relative abundance of “
Model selections using a generalized linear model (GLM) with WA variables were performed to assess the effects of carbaryl and Daphnia on response variables (WARES), using:
[Fig. 2.] Carbaryl concentrations (average ± SE) in microcosms: (a) Measured concentrations by using HPLC. Decomposition rate of carbaryl (b) was approximately 5% per day, irrespective of either the presence of Daphnia in the community or the initial concentration of the insecticide.
to the other zooplankton (MacIsaac and Gilbert 1989).
To analyze the differences in species compositions and those abundances between no-carbaryl tanks and carbaryl- introduced tanks, principal response curves (PRCs) were calculated based on a redundancy analysis (RDA) (Van den Brink and Ter Braak 1999, Hens et al. 2005) by using “Vegan” package in R 2.15.2 (R Core Development Team 2011) . Biotic data (abundance) were log(+1) transformed, centered over time and standardized before the analysis. The environmental variable was each treatment, with sampling time as the co-variable. PRCs were derived by plotting the canonical coefficients (Cdt) against time. The line at y = 0 represents the mean of the controls and the Cdt’s of the treated microcosms indicate their deviation from the controls for each sampling date. Accompanying species scores allow an interpretation nat the species level. Data sets for no-
All statistical analyses excepting repeated-measures ANOVA were performed using the statistical software R (version 2.15.2).
During the experiment, DO and pH in tanks ranged between 3-7 mg O2 L-1 and 6.7 to 7.2, respectively. However, the values did not differ significantly among the treatments (
Carbaryl application caused a downdrift of the cladoceran density dependent on the concentrations (Fig. 3a). Contrary to the cladocerans, total herbivorous rotifer density increased markedly in the “high carbaryl” treatment (Fig. 3b). However, densities of the carnivorous zooplankton did not differ between the treatments (Fig. 3c and 3d).
Cladoceran communities were mainly composed of two
We performed the model selections using a generalized linear model (GLM) with time-weighted average (WA) of response variables to assess the effects of carbaryl and
[Fig. 3.] Changes in density of herbivorous (a and b) and carnivorous (c and d) zooplankton in each treatment (average ± SE). Asterisks (*) show the effects of explanatory variables (*, P < 0.05; **, P < 0.01 with repeated-measures ANOVA. No asterisk means P ≥ 0.05).
[Fig. 4.] Changes in density (average ± SE) of herbivorous cladocerans (a-d) and rotifers (e-g), copepod nauplii (h) and relative abundance of “pellucida (defensive)”-morphed B. longirostris in adult females (i) in each treatment. Note that Y-axes rotifer and nauplii (e-h) are represented as individual numbers per 200 mL. Asterisks (*) show the effects of explanatory variables (*, P < 0.05; **, P < 0.01 with repeated-measures ANOVA. No asterisk means P ≥ 0.05).
Generalized linear model (GLM) used to estimate the effects of carbaryl concentration and Daphnia density on the response variables.
was not significant. The positive effect of carbaryl on rotifers was not significant for species level values.
PRC provided the information about the changes in species (taxon) composition due to the carbaryl applications (Fig. 6). The first components of PRC in no-
[Fig. 6.] Principal response curves (PRC) with species weights for the zooplankton data set, (a) no-Daphnia treatments, (b) Daphnia-introduced treatments, indicating the effects of low and high dose of the insecticide. The canonical coefficients (Cdt’s, first principal component of treatment effects) of the treated microcosms indicate their deviation of species composition from the controls for each sampling date. 71.3% and 69.5% of variance were explained by the first components in the left and right panels, respectively.
Population dynamics of zooplankton in lakes and ponds are controlled by complex combinations of biotic interactions and abiotic environmental factors. Vulnerability to anthropogenic toxic chemicals differs between each species, so the chemicals should induce different community structures. Several researchers have concluded that large cladocerans tend to be more sensitive to insecticides than smaller ones based on mesocosm/enclosure experiments (Moore and Folt 1993, Hanazato 1998). On the other hand, the opposite relationship has been observed in the laboratory toxicity tests (Passino and Novak 1984, Sakamoto et al. 2005). However, the “size-dependent-tolerance” is not always applicable to broader taxonomic scale comparison (Mano et al. 2010).
Results of the present study did not coincide with the previous community-level studies. The population densities of the small cladocerans (
A possible reason for the above inconsistency is the rapid recovery of the small cladocerans by hatching from resting eggs in the enclosure studies. Hanazato (1991a) reported that
The difference in the results may also have been due to the different locations of the experiments. Carbaryl seems to be photolysed easily (Perez-Ruiz et al. 2003) and the light intensity would be higher in field than in the laboratory. Hanazato and Yasuno (1990) reported that 89% of carbaryl applied to the outdoor enclosures was decomposed within 1 day. In our experiment, however, the absolute reduction of the concentration was only 5% per day irrespective of the initial concentration (Fig. 2). Different decomposition rates of carbaryl can lead to different results, because the damaged reproductivity of the animals can recover if the concentration of toxic chemicals decreases.
One conspicuous effect of insecticide application on zooplankton communities is an increase of rotifer density (Chang et al. 2005). This is because rotifers are far more tolerant to the insecticides than cladocerans. Cladocerans show superior exploitative competition to rotifers, and therefore chemical pollution can free the latter from food shortage (Hanazato 2001). The same phenomenon was also observed in the present microcosm experiment. Total rotifer density was positively correlated with the carbaryl concentration (Table 1). However, this was not the case in the presence of
Negative correlation with
Zooplankton community structure diverged depending on the pollutant level (Fig. 6). In the high-dose treatments (Fig. 6), the canonical coefficient (Cdt) of PRCs increased subsequent to the carbaryl application. This indicates that the decrease of sensitive species due to the acute toxicity of carbaryl and the incidental increase of rotifers occurred rapidly. In the low dose treatment, on the other hand, Cdt value did not fluctuate from zero until day 14, meaning that the direct lethal effect was weak. Sakamoto et al. (2009) have reported that the population growth rate of
Insecticides not only affect the individual survival or the reproduction rate, but also disturb the prey-predator interactions by enhancing or inhibiting the induction of morphological defences of prey animals (Lurling and Scheffer 2007). For example, carbamate and organophosphorus insecticides enhance the development of protuberant morphologies of
Some lake ecosystems have many real problems due to anthropogenic impacts. Chemical pollution is one contributing factor to these problems, pesticides having great impact on the survival of the organisms in the system. Accurate evaluation of the impact of anthropogenic toxic chemicals on ecosystems is one of the most important goals for ecotoxicologists. It has been shown that the genera