Abstract
Conventional farming uses a large volume of pesticides that may reach aquatic ecosystems. This is also the case for the insecticide fipronil and the herbicide 2,4-D, which are widely used in many crops. This study aimed at evaluating the individual and mixture toxicity of these pesticides to the tropical amphipod Hyalella meinerti. To this end, acute toxicity tests (96 h) were conducted. Chronic bioassays (10 days) were also carried out, in which the body length and dry biomass were evaluated as endpoints. In addition, a complete factorial mixture chronic toxicity test was carried out. H. meinerti was sensitive to fipronil in the acute toxicity tests, with a LC50-96-h of 0.86 μg L−1 (95% CI 0.26–0.46), and no acute effects were observed after 2,4-D exposure even at the highest test concentration of 100 mg L−1. In the chronic toxicity tests, all tested concentrations of both pesticides decreased the growth of H. meinerti, in which losses on biomass reached 45% and 65% for 2,4-D and fipronil, respectively. The pesticide mixture indicated antagonism although it still significantly decreased the body growth. The results obtained indicate a high sensitivity of H. meinerti exposed to environmentally realistic concentrations, demonstrating that there are risks for the species in real field conditions.
Similar content being viewed by others
Explore related subjects
Discover the latest articles, news and stories from top researchers in related subjects.Avoid common mistakes on your manuscript.
Introduction
In conventional farming, a large amount of pesticides is used in crops, and despite the fact that these molecules are developed to be toxic to specific pests, they may also exert effects on non-target species in terrestrial and aquatic environments (Bartlett et al. 2016). In Brazil, herbicides are the largest consumed pesticide class, followed by fungicides and insecticides. The 2,4-dichlorophenoxyacetic acid (2,4-D) is the second most used herbicide in the country (48921 ton), and fipronil the sixth most used insecticide (1689.7 ton) (IBAMA 2018). In addition, 2,4-D is one of the most consumed herbicides in the world, and fipronil is almost exclusively used in underdeveloped countries where its use has not yet been banned (Brennan et al. 2009; Weston and Lydy 2014; Islam et al. 2018). 2,4-D is a growth inhibitor for dicotyledonous plants that was introduced on the market in 1940 (Tomlin 1994; Islam et al. 2018). Fipronil belongs to the phenylpyrazole chemical group, developed to block the GABA-regulated chloride channels in insects and was introduced as a substitute for organochlorine and organophosphate insecticides (Tomlin 1994; Brennan et al. 2009; Weston and Lydy 2014).
2,4-D has a low affinity with soil particles and a high water solubility (Koc = 20–280) that increases its transport potential to aquatic environments (Barriuso et al. 1992; DeQuattro and Karasov 2016). On the other hand, fipronil has a high affinity for organic matter (Koc = 825) and may be carried from crops to aquatic systems associated with organic particles from the soil (Ying and Kookana 2001; Mize et al. 2008). Both pesticides are allowed to be used on sugarcane, soybean, maize, and rice crops, as well as five other crops (MAPA 2019). The half-life of 2,4-D and fipronil in soils range from 17 to 39 days and 6 to 33 days, respectively (Bolan and Baskaran 1996; Bobé et al. 1998; Ying and Kookana 2002; Zhu et al. 2004). Thus, the relative persistence of both pesticides in soils and their application on the same crops increases the probability of their combined occurrence in the soil and edge-of-field water bodies in agroecosystems.
Despite the risks associated with pesticides for non-target species, monitoring these molecules in freshwater environments is still scarce in Brazil, even considering their high consumption. In a critical review, Albuquerque et al. (2016) found only 29 studies dealing with pesticide monitoring in surface water environments. Fipronil was the insecticide most commonly found with occurrences in 54% of 251 samples, and 2,4-D was found in 14% of 210 samples (Albuquerque et al. 2016). In São Paulo State (Brazil), fipronil occurred in 62% of the samples, and 2,4-D was detected in less than 20%. Besides, the occurrence of both pesticides together was reported in water bodies around sugarcane crops (CETESB 2019). Concentrations ranging from 0.1 to 465 μg L−1 of fipronil and 1.14 to 366 μg L−1 of 2,4-D have been reported in freshwater environments in Brazil (Marchesan et al. 2010; Pinheiro et al. 2010; CETESB 2018). The occurrence of these molecules in the environment, alone or together, may imply lethal or sublethal effects to exposed organisms. Recent studies have demonstrated the effects of both compounds (alone and in a mixture) on different groups of aquatic biota, such as algae (Moreira et al. 2020b), cladocerans (Silva et al. 2020), chironomids (Park et al. 2010; Monteiro et al. 2019), and fish (Moreira et al. 2021). However, there is still a lack of information related to the risks of fipronil and 2,4-D, alone or in a mixture, for tropical species.
Amphipods have often been used in ecotoxicological studies, and Hyalella and Gammarus are the most used genera (Mugni et al. 2011; Nguyen et al. 2012; Dalhoff et al. 2018). Due to their detritivorous-herbivorous habits, these organisms are involved in the energy flux along the food chain and are an important link between primary producers and high trophic levels (Giusto et al. 2012). H. azteca is widely used as a test species; however, the use of indigenous tropical species brings more realistic results regarding the effects (and potential risks) of pesticides in tropical regions (Giusto et al. 2012). Hyalella meinerti Stebbing, 1899 is an amphipod largely distributed in Brazil and found in other South American countries such as Venezuela, Colombia, Peru, and Ecuador (González and Watling 2003), and was used as a test organism in this study. The general aim was to evaluate the individual and mixture toxicity of the fipronil and the 2,4-D to the macroinvertebrate H. meinerti. The pesticides were evaluated as their commercial products since they are used in this form in real-world agricultural practices. The specific aims were (i) to evaluate the effects of fipronil and 2,4-D on the survival and growth of the amphipod H. meinerti in single and mixture exposures and (ii) to evaluate whether single and mixture toxicity was different between the parameters analyzed.
Methods
Test organisms
Cultures of H. meinerti were maintained at the Nucleus of Ecotoxicology and Applied Ecology (NEEA), at the Centre for Water Resources and Environmental Studies (CRHEA), located in the municipality of Itirapina, São Paulo State, Brazil. Organisms were kept according to procedures described in ABNT NBR 15470 (2013) at 25 ± 1 °C and 12 h:12 h (light: dark) in glass aquariums with 4 L of culture water (well water, pH 7–7.5, conductivity 150–160 μS cm−1, and hardness 40–48 mgCaCO3 L−1), under constant aeration and 100 males and 100 females per aquarium. The aquatic macrophytes Myriophyllum aquaticum and Egeria densa were added as substratum and vegetal food supply. The organisms were fed ad libitum daily with compound food (Tetramin® suspension and biological yeast, 0.025 mL per organism). The cultures were renewed weekly when the adults and macrophytes were transferred to a new aquarium. When the cultures were renewed, the water from the old cultures containing the juveniles (0–7 days old) was gently poured through a mesh (145 μm), and the retained organisms were transferred to a new aquarium and maintained for 1 week until the toxicity tests were carried out.
Chemicals and test solutions
In the present study, two commercial formulations containing fipronil and 2,4-D were evaluated. The insecticide Regent® 800 WG (BASF S.A., Brazil) contains 80% (w/w) of the active ingredient fipronil and 20% (m/m) of inert ingredients. The herbicide DMA® 806 BR (Dow AgroSciences Industrial Ltda, Brazil) contains 67% w/v of the active ingredient 2,4-D (acid equivalent) and 41.9% w/v of inert ingredients.
The stock solutions of Regent® 800 WG (800 μg fipronil L−1) and DMA® 806 BR (1 g 2,4-D L−1) were diluted in distilled water. The test concentrations in the acute and chronic bioassays were obtained by diluting the stock solutions with culture water. The stock and test solutions were prepared immediately before the experiments to avoid possible pesticide degradation.
Acute toxicity tests
The bioassays were carried out using the same water, temperature, and illumination as described for the cultures in accordance with the conditions described in ABNT NBR 15470 (2013). Static acute toxicity tests were carried out in non-toxic plastic bottles (Copaza®) with 175 mL of the test solution and four replicates with ten organisms each (7–14 days old) per concentration. Four nominal concentrations of fipronil (1.0, 1.5, 2.0, 2.5, and 3.0 μg L−1) and 2,4-D (6.25, 12.50, 25, 50, 100 mg L−1) were tested. A nylon screen (4 cm × 15 cm, 0.25 mesh size) was added as a substrate, and organisms were fed at the beginning and post 48 h (0.025 mL of compound food per organism). The duration of the test was 96 h, and the solutions were not renewed. The test was considered valid if the survival in control was higher than 80%, and the experiments were repeated three times with different broods (ABNT NBR 15470 2013).
Chronic bioassays
Chronic bioassays were carried out using 250 mL of test solutions, a nylon screen as a substrate, and eight replicates containing ten organisms each (7–10 days old) in the same conditions described for the acute tests. The highest concentration of fipronil was chosen based on the lethal concentration—LC20-96 h obtained in the acute tests. The tested concentrations were 0.0875, 0.1750, 0.3500, 0.7000, and 1.4000 μg fipronil L−1. The concentrations of 2,4-D (22.5, 45, 90, 180, 360 μg a.i. L−1) were selected based on the values quantified in streams in São Paulo state, Brazil (CETESB 2018), as no acute effects were observed for the herbicide (see topic 3.1). The organisms were fed every 48 h, and the test solution was renewed every 96 h. At the end of the tests (10 days), the living organisms were counted, anesthetized with phenoxyethanol, and preserved in 70% ethanol. The organisms were photographed, and the body length was measured using a free software called Kinovea 0.8.15 (https://www.kinovea.org/) that was calibrated using graph paper. The body size was measured as the dorsal line contour from the base of the first antenna into the third uropod base.
The mixture experiment was carried out under the same conditions as the chronic bioassays, but with four replicates per treatment aiming to allow a greater number of treatments (Pavlaki et al. 2011). The full factorial modeling (Moreira et al. 2020a) was applied as the experiment design: one control, ten isolated concentrations (five for fipronil and 2,4-D each), and 25 combinations of the two pesticides (five of fipronil x five of 2,4-D). The concentrations used were the same as those evaluated in the chronic bioassays. All combinations are presented in the Supplementary material (Table S1). At the end of the tests, the surviving organisms were counted, preserved in 70% ethanol, photographed, and evaluated as described above.
Water parameters
At the beginning and end of the acute and chronic tests, the water conductivity (Orion 145), pH (micronal B374), dissolved oxygen, and water temperature (YSI55-25 ft) were measured. In addition, at the end of the chronic tests, the hardness (ABNT NBR 12621 1995) and the ammonium ion concentration (Hansen and Koroleff 2007) were determined.
Body length-biomass estimation
Fifty organisms were obtained randomly from cultures at different stages of the cultures (0–7 until > 40 days) to estimate the body length-biomass. The selected organisms were maintained in clean water for 24 h to allow gut clearing. Individuals were then killed with phenoxyethanol, photographed to measure the body length as described, dried at 60 °C for 24 h, and weighed (0.01 mg, Mettler–AE 240) to establish the dry biomass. The relationship between the body size and dry biomass for H. meinerti was determined, and the dry biomass of the organisms in the chronic bioassays was estimated based on the measured body length.
Chemical analyses
Samples from the stock solutions of fipronil (800 μg L−1) and 2,4-D (1 g L−1) and from all dilutions at the beginning of the chronic tests were taken for chemical analyses. All analyses were carried out at the Environmental Chemistry Laboratory at the Institute of Chemistry at the State University of Campinas (UNICAMP), according to the methodology described in Goulart et al. (2020). The samples from 2,4-D tests were diluted with methanol (200 μL methanol and 800 μL of the sample) and filtered through a syringe filter (PTFE 0.22 μm). For fipronil analysis, the samples were filtered (syringe PTFE with a filter of 0.22 μm) and prepared by online solid phase extraction by passing 500 mL of the sample through a 500 mg HLB Oasis cartridge (Waters Corporation—Milford, USA) with a flow rate of 7 mL min−1. After that, the cartridges were dried, the analytes were eluted, and the extracts were prepared for the analysis, as described in Goulart et al. (2020). The chemical analyses were carried out by liquid chromatography coupled with mass spectrometry (Agilent Technologies QqQ 1200, 6410B, Santa Clara, USA) with electrospray ionization in negative mode. For the chromatographic separation, the Zorbax SB-C18 column (2.1 × 30 mm, particle size of 3.5 μm) and the Poroshehell 120 EC-C18 column (3.0 × 50 mm, particle size of 2.7 μm) at 30 °C were used for 2,4-D and fipronil analysis, respectively. The volume injected was 10 μL and the mobile phase (aqueous 0.01% ammonium hydroxide and methanol) had a flow rate of 0.3 mL min−1.
The limit of detection (LOD) and quantification (LOQ) were determined using the signal-to-noise ratio method (SNR). The analytical signal in the low concentrations of both compounds in ultrapure water was compared with the noise in the baseline. The SNR proportion was 10:1 for LOQ and 3:1 for LOD. The LOD and LOQ were 0.001 and 0.005 μg L−1 for fipronil and 0.5 and 1.0 μg L−1 for 2,4-D, respectively. The determination coefficient was 0.996 and 0.998, and the sensitivity of the analytical curve was 384.6 and 463.5, for 2,4-D and fipronil, respectively. The calibration curve was determined by using seven different concentrations ranging from 5 to 300 ng mL−1 for both pesticides (Goulart et al. 2020).
Mixture interactions
Interactions of the fipronil and 2,4-D mixture were assessed based on the predicted and observed effects at the mixture, which were calculated according to the methodology described by Gottardi et al. (2017), considering the independent action model (Jonker et al. 2005; Loureiro et al. 2010). The predicted effect was estimated by the following equation:
where D, F, and C are the mean responses (e.g., survival) registered for 2,4-D (D) and fipronil (F) alone and control (C) for a specific combination. The observed effect in a combination was calculated as the ratio of the effect observed in the mixture (M) and the untreated control (Observed effect = M/C). The relationship between the predicted and observed effects may determine the mixture interaction and was calculated only for those combinations that were statistically different from the control or the individual fipronil or 2,4-D treatments. When the observed effect was lower than the predicted effect, the mixture effect was considered synergic, and when the observed effect was higher than the predicted effect, antagonism was assumed (Gottardi et al. 2017).
Data analyses
All data analyses were performed in R version 3.6.0 (2009) using the RStudio version 1.2.1335 (2019). The lethal concentrations post 96 h (LCx-96 h) were calculated by the non-linear estimation with the logistic model. In the chronic tests, the survival and growth (body length and biomass) were compared with controls by generalized linear models (GLM) (Lopes et al. 2018; Scherer et al. 2020; Figueirêdo et al. 2020). Survival was analyzed by GLM using the binomial family with the logit-link function. The effects on growth were assessed by GLM with the Gaussian family and the identity-link function. In the mixture study, GLM was used as described for the chronic tests, and effects were evaluated by comparing the combinations with the control and the respective concentrations of fipronil and 2,4-D alone. In all analyses, the data met the assumption of independence between treatments and the confidence level was 95% (p value of 0.05). A regression analysis was used to establish the relationship between the body length and dry biomass. The biomass of organisms was estimated from the established equation. All results are presented as the overall mean of the three independent experiments that were validated.
Results
Chemical analysis
The concentrations quantified in the stock solutions by LC-MS/MS analysis were 536 μg L−1 for fipronil and 0.90 g L−1 for 2,4-D, corresponding, respectively, with nominal concentrations of 800 μg L−1 and 1.0 g L−1. These quantified values ranged more than 20% from the intended values; thus, the measured concentrations are presented and were used to calculate the toxicity values (OECD 2011).
Regarding the initial concentrations of the chronic tests, the detected values were 0.2, 0.3, 0.4, 0.5, and 0.9 μg a.i. L−1 for fipronil relative to the nominal concentrations of 0.0875, 0.1750, 0.3500, 0.7000, and 1.400 μg a.i. L−1. For the herbicide 2,4-D, the concentrations were quantified as 29, 66, 112, 221, and 426 μg a.i. L−1, relative to the nominal concentrations of 22.5, 45, 90, 180, and 360 μg a.i. L−1. Similar to that described above, for the chronic bioassays, these measured values are presented and used for toxicity threshold calculations.
Water quality parameters
At the beginning and end of the acute and chronic tests, the water quality parameters were compared with those under culture conditions. The pH and electrical conductivity of the water ranged between 6.5–7.5 and 160–190 μS cm−1, respectively. The oxygen concentrations were higher than 6.5 mg L−1 in all experiments. At the end of the chronic tests, the concentration of ammonium ion ranged between 0.8 and 1.7 mg L−1 and the hardness of the water between 44 and 46 mg CaCO3 L−1.
Acute toxicity tests
In the acute toxicity tests, no lethal effects were observed for H. meinerti exposed to 2,4-D even at the highest test concentration of 100 mg L−1. Conversely, fipronil was toxic to the species in low concentrations. The average lethal concentrations are shown in Table 1.
Chronic bioassays
In the chronic toxicity tests, the survival of the control group was greater than 90% in all bioassays. The higher concentration of fipronil (0.9 μg L−1) decreased the survival (80 ± 9%) compared to the control (94 ± 3%, p < 0.05). For the other concentrations of fipronil, as well as all treatments containing 2,4-D, survival is similar to the controls (p > 0.05, Fig. 1).
The regression analysis is well adjusted (F = 426, p < 0.05, and R2 = 0.92, Fig. S1), and therefore, the dry biomass of H. meinerti in the chronic experiment is estimated from the body length data. All concentrations of the herbicide 2,4-D decreased the body length and biomass related to the control (p < 0.05, Fig. 2), in which losses in the biomass ranged from 36% at 29 μg L−1 to 45% at 112 μg L−1. No dose-dependent effect was observed for 2,4-D, and decreases were similar at all concentrations tested (p > 0.05). The lowest observed effect concentration (LOEC) and no observed effect concentration (NOEC) for 2,4-D were obtained from the GLM analysis. The LOEC was 29 μg L−1 and the NOEC was lower than 29 μg L−1. In the same way, fipronil decreases the body length and biomass of H. meinerti at all test concentrations (p < 0.05, Fig. 2), in which losses in the biomass ranged from 30% at 0.3 μg L−1 to 65% at 0.9 μg L−1. The LOEC was 0.2 μg L−1, and the NOEC was lower than 0.2 μg L−1. Contrary to that observed for 2,4-D, organisms exposed to fipronil increased the losses in growth with increasing doses of the insecticide (p < 0.05), except for the lowest concentration (0.2 μg L−1) in which the losses were higher than those denoted in the 0.3 μg L−1 treatment (Fig. 2).
Mixture toxicity tests
The mixture test was validated with 100% of survival in controls. Fipronil alone decreases the survival at 0.9 μg L−1 (35 ± 17%, p < 0.05), and no effects occur for 2,4-D alone at any concentration tested (p > 0.05, Fig. 3). Regarding the mixture combinations, survival was decreased compared with the control (p < 0.05) for 0.9 μg L−1 of fipronil combined with 2,4-D at 29 μg L−1 (65 ± 10%), 66 μg L−1 (60 ± 22%), and 426 μg L−1 (40 ± 23%). For the other combinations, the survival is greater than 80%, and no effects occur compared with the control (p > 0.05, Fig. 3). No synergism occurred for any mixture combination, and all mixture treatments that had significant differences from the control or the individual compounds were antagonistic.
Table 2 presents the interaction of pesticide mixtures for those combinations that were statistically different from the control or compounds alone regarding the body length and biomass parameters. Growth (body length and biomass) of H. meinerti exposed to fipronil and 2,4-D alone was decreased at all tested concentrations compared with the control (p < 0.05). The results of all the combinations for both parameters are presented in Fig. 4. The combinations of 0.2 μg L−1 fipronil with 66 μg L−1 2,4-D and 0.4 μg L−1 fipronil with 29 μg L−1 2,4-D had an abnormal growth, contrary to the corresponding isolated compounds and the other mixture combinations. The combination of 0.2 μg L−1 fipronil with 29, 66, and 112 μg L−1 2,4-D and 0.4 μg L−1 of the insecticide with 29, 112, and 426 μg L−1 2,4-D showed an antagonistic interaction, which had observed effects that were 1.0 to 3.6 times higher than the predicted values. In the same way, the combinations of 0.3, 0.5, and 0.9 μg L−1 fipronil with all concentrations of 2,4-D were also antagonistic, with observed effects 1.1 to 3.3 times higher than those predicted. Synergism did not occur at any tested combination.
Discussion
The herbicide 2,4-D and the insecticide fipronil are widely used in Brazil, and concentrations reaching 366 μg L−1 and 465 μg L−1, respectively, have already been reported in streams located in areas that have a predominance of sugarcane crops in São Paulo State (CETESB 2018). The occurrence of fipronil in concentrations ranging from 0.1 to 26.2 μg L−1 and 2,4-D ranging from 1.14 to 74.5 μg L−1 has also been reported in the south of Brazil in rivers in rice cultivation areas (Grützmacher et al. 2008; Marchesan et al. 2010; Pinheiro et al. 2010). Outside Brazil, both compounds have also been detected in several surface waters worldwide, in which concentrations ranged from 0.062 to 12 μg L−1 for 2,4-D and 0.0007 to 4.07 μg L−1 for fipronil (Demcheck 2003; Islam et al. 2018; Fang et al. 2019).
The concentrations of fipronil quantified in surface waters may imply a lethal risk for H. meinerti when considering the LC50-96 h obtained for this species. Acute toxicity values reported in the literature indicated that H. azteca presented sensitivity close to H. meinerti, in which LC50-96 h values ranged from 0.32 to 1.7 μg fipronil L−1 (Lizotte et al. 2009; Weston and Lydy 2014), and the shrimp Macrobrachium rosenbergii (LC50-96 h 0.98 μg fipronil L−1) presented higher sensitivity (Shan et al. 2003). H. meinerti had the highest sensitivity to fipronil as compared to the cladoceran species, such as Ceriodaphnia silvestrii (LC50-48 h 3.90 μg L−1), C. reticulata (LC50-48 h 8.83 μg L−1), C. dubia (LC50-48 h 18.53 μg L−1), and D. magna (LC50-48 h 88.30 μg L−1), which are widely used as a standard test organism in ecotoxicological tests, including those conducted for regulatory purposes (Konwick et al. 2006; Hayasaka et al. 2012; Silva et al. 2020).
In aquatic ecosystems, fipronil may occur together with its degradation products, which presents higher persistence compared with its parental compound (Maul et al. 2008; Brennan et al. 2009; Qu et al. 2016). Toxicity studies have reported similar or higher toxicity of the degradation products than fipronil, indicating an increased or at least prolonged risk to the exposed organisms. For example, for H. azteca, the LC50-96 h for fipronil was 1.6 μg L−1, 1.4 μg L−1 for fipronil-sulfide, and 0.43 μg L−1 for fipronil-sulfone (Weston and Lydy 2014). For the crayfish Procambarus clarkia, the LC50-96 h was 63.7 μg L−1 for fipronil, and 34 μg L−1, 73.7 μg L−1, and 149.7 μg L−1 for the degradation products fipronil-sulfone, sulfide, and desulfinyl, respectively (Schlenk et al. 2001).
Regarding 2,4-D, acute risks are not expected for H. meinerti exposed to environmental-realistic concentrations. Contrary to our study, the amphipods H. azteca (LC50-96 h 0.6 mg 2,4-D L−1), Gammarus pulex (LC50-96 h 2.3 mg 2,4-D L−1), and G. fasciatus (LC50-96 h 3.8 mg 2,4-D L−1); the cladocerans D. lumholtzi (LC50-48 h 10 mg 2,4-D L−1) and D. magna (LC50-48 h 25 mg 2,4-D L−1); and the crayfish Astacus leptodactylus (LC50-96 h 32.6 mg 2,4-D L−1) presented lethal responses to 2,4-D at concentrations lower than 100 mg L−1 (Sanders 1970; Kader et al. 1976; Seuge et al. 1970; George et al. 1982; Alexander et al. 1985; Paul et al. 2006; Benli et al. 2007). Crustaceans had a clear resistance to 2,4-D at short time exposure, with LC50 values at doses that are not environmentally relevant (mg L−1 range), except for H. azteca. For this reason, concentrations higher than 100 mg 2,4-D L−1 were not tested in the present study.
In the chronic exposure tests, only the highest concentration of fipronil decreased the survival of the test species. As previously discussed, the insecticide is highly toxic to amphipods, and the mortality may be associated with its neurotoxicity (Tomlin 1994) and the prolonged exposure time. The impairments of growth of H. meinerti exposed to fipronil and 2,4-D may imply consequences for the species, mainly considering the concentrations already registered in Brazilian superficial water bodies. Contrarily to our study, the shrimp Palaemonetes pugio had no impairments on weight and length when exposed up to 0.2 μg fipronil L−1 despite effects on survival (Volz et al. 2003). Regarding 2,4-D, no effects were observed on the growth of the cladoceran D. magna at concentrations ranging from 10 to 100 μg L−1 (Kashian and Dodson 2002). In the same way, the exposure of the cladoceran Ceriodaphnia silvestrii to 1.3 μg fipronil L−1 and 163 μg 2,4-D L−1 did not lead to impairments of body length (Moreira et al. 2020c). The effects of fipronil and 2,4-D on impairment of growth for other animal groups have previously been studied, e.g., for chironomids (Maul et al. 2008; Monteiro et al. 2019), fishes (Fairchild et al. 1997; Li et al. 2017), and tadpoles (Freitas et al. 2019). To the best of our knowledge, however, there are no studies evaluating the effects of both pesticides on the growth of crustaceans, and our results thus bring new findings about the implications of prolonged exposure for the Hyalella genus.
Organisms stressed by the prolonged exposure to pesticides, such as fipronil and 2,4-D, will have less energy available to invest in growth, compared with the non-exposed ones, leading to implications on the population level and increasing the risks of local extinction (Liber et al. 1996). Thus, the reduced growth of H. meinerti may have resulted from a reallocation of energy reserve as compensatory mechanisms for the detoxification of the pesticides. Fipronil indeed enhanced the energy demand for detoxification mechanisms of the fish Dicentrarchus labrax during exposure and after the depuration period (Dallarés et al. 2020). In the same way, the insecticide increased the metabolism of the midge Chironomus riparius, which was accompanied by decreases in larval growth (Monteiro et al. 2019). Several studies have also shown the influence of the herbicide 2,4-D on increases in the detoxification mechanisms (Park et al. 2010; Banaoui et al. 2015; Gaaied et al. 2019; Arcaute et al. 2019).
Besides alterations in energy allocation, the impairments in growth may also be attributed to losses in nutrient absorption or alterations in the behavior of exposed organisms with consequences for the uptake of food and allocation of energy (Richardi et al. 2018). Previous studies have shown the effects of both pesticides on the behavior of a range of organisms. Exposure to 2,4-D leads to changes in the locomotion of the crab Astacus leptodactylus, having implications for its foraging behavior (Benli et al. 2007). The crayfish Orconectes rusticus exposed to the herbicide spends more time finding a source of food, leading to decreases in the rates of food consumed (Browne and Moore 2014). Fish exposed to 2,4-D changed their swimming behavior and exhibited other alterations including anorexia, edemas, hemorrhages, breathing difficulties, and congestion, whereas larvae of Culex pipiens fatigans exhibited impairments on respiration and swimming (Farah et al. 2004). Fipronil decreased the swimming performance of the fishes Pimephales promelas and Danio rerio at sublethal concentrations (Stehr et al. 2006; Beggel et al. 2010; Chaulet et al. 2019). The neurotoxic character of fipronil may induce feeding inhibition and a reduction in activities of the exposed organisms, with consequences on growth (Monteiro et al. 2019). To the best of our knowledge, there has not been any study yet evaluating the inhibition of feeding behavior of species exposed to the insecticide.
Effects on growth may have implications on the population level as the brood size of amphipods is directly correlated with the adult body length (Cooper 1965). This pattern was observed in natural populations of H. bonariensis and H. carstica in the south and southeast of Brazil, where the female size was positively correlated with the number of eggs produced, and males and females amplexed were larger than non-paired organisms (Torres et al. 2015; Castiglioni et al. 2018). In the same way, the size of females was positively correlated with the number of produced eggs of H. pleoacuta and H. castroi under laboratory conditions (Castiglioni and Bond-Buckup 2007). Greater females of Gammarus minus were related to more offspring per brood at ten natural populations in North America streams (Glazier 2000). Thus, impairments on survival and growth are thus likely to lead to declines in natural populations of amphipods in freshwater ecosystems receiving low doses of fipronil and 2,4-D.
The occurrence of pesticides in freshwater ecosystems, mainly in streams surrounding agriculture landscapes, is a current problem worldwide, and exposure to these compounds most often includes two or three molecules (Schreiner et al. 2016). The co-occurrence of fipronil and 2,4-D has already been registered in areas with predominance of sugarcane crops in São Paulo State, Brazil (CETESB 2018), showing evidence of the importance of understanding the interaction of these compounds for indigenous species. In the present study, the mixture of both pesticides had an antagonistic response on the survival of H. meinerti. In the same way, an antagonistic interaction was observed for the survival of the shrimp Procambarus clarkii exposed to a mixture of the herbicide 2,4-D with the herbicide monosodium methanearsonate (MSMA) (Green and Abdelghani 2004).
Similar to the effects observed for survival, the pesticide mixture had an antagonistic interaction on the body length and dry biomass. We emphasize that despite the antagonistic interaction observed, in most combinations of fipronil and 2,4-D, growth was reduced compared to control. For example, the losses in biomass at the highest mixture concentrations reach 80% and are greater than the losses observed for the individual compounds (44 and 70% for 2,4-D and fipronil, respectively, Fig. 4). These results demonstrate that the mixture of fipronil and 2,4-D still implies risks for the species since all tested concentrations were environmentally relevant, as previously discussed. Besides, the decreases in biomass may have cascading effects on aquatic food webs due to the importance of amphipods for secondary productivity of freshwater environments (Giusto et al. 2012).
Antagonism was also observed for the reproduction of C. silvestrii exposed to the same formulations as those tested in the present study. On the other hand, for the acute toxicity tests, antagonism occurred for C. silvestrii in doses lower than the EC50 values of fipronil and 2,4-D, and synergism at higher doses (Silva et al. 2020). We highlight that synergism was observed at higher concentrations of 2,4-D than those evaluated in our study (350 to 650 times, respectively). In addition, in the study by Silva et al. (2020), the EC50 for C. silvestrii exposed to the insecticide fipronil was 25 times higher than the LC50 value derived in the present study for H. meinerti. Antagonistic interactions of fipronil and 2,4-D mixed with other pesticides have previously been reported in the literature (Oruç and Üner 2000; Ullah et al. 2017; Van Meter et al. 2018; Levchenko and Silivanova 2019). On the other hand, there is still a lack of information on the mixture effects of fipronil and 2,4-D for aquatic organisms.
Finally, we highlight that the bioassays were carried out with the formulated products which contain other substances besides the active ingredients, and these ingredients (the inert ingredients) can evidently also contribute to the observed effects. Besides, the presence of these other ingredients in the formulation may explain the abnormal growth in some combinations. For example, Carvalho et al. (2020) demonstrated that different formulations containing 2,4-D presented differences in toxicity to the fish C. decemmaculatus. In the same study, it was observed that a formulation of glyphosate mixed with two different 2,4-D-based formulations had different interactions (synergism or antagonism) depending on the formulated product (Carvalho et al. 2020). Adding a surfactant to the mixture of the herbicides 2,4-D and MSMA altered the interaction between the compounds from antagonistic to synergetic (Green and Abdelghani 2004). Many studies have demonstrated that formulated products may have a greater toxicity compared to the active ingredient they contain (Nagy et al. 2020 and references therein). The Brazilian regulation for packaging and labeling commercial pesticides requires indicating the name and percentage of each active ingredient; for the excipients, only the amount present in the formulation is mandatory (Brazil 1989, art. 7). This may be why the composition of the other components of the Regent® 800 WG BASF S.A. and DMA® 806 BR formulations is unknown. Only the active ingredients (fipronil and 2,4-D, respectively) are known.
Conclusions
The insecticide fipronil and the herbicide 2,4-D are widely used on crops in Brazil and other countries worldwide. In the present study, the commercial formulations of both pesticides implied toxicity to the crustacean H. meinerti, including decreases in survival after acute exposure to the insecticide, and impairments of growth after chronic exposure to low concentrations of both pesticides. Crustaceans are highly sensitive to the insecticide, and amphipods present the highest sensitivity to both pesticides when compared with other groups such as cladocerans that are largely used in ecotoxicological studies.
The mixture effects of both pesticides were antagonistic, although the observed effects still demonstrated impairments on growth. All observed effects at the single and mixed exposure were obtained at environmentally realistic concentrations, pointing at risks for the species and other sensitive aquatic organisms in real world field settings.
References
ABNT NBR 12621 (1995) Waters - determination of total hardness by EDTA-Na titulometric method - Method of test
ABNT NBR 15470 (2013) Aquatic ecotoxicology — acute and chronic toxicity — method for assessing the toxicity of sediment using Hyalella spp (Amphipod)
Albuquerque AF, Ribeiro JS, Kummrow F, Nogueira AJA, Montagner CC, Umbuzeiro GA (2016) Pesticides in Brazilian freshwaters: a critical review. Environ Sci Process Impacts 18:779–787. https://doi.org/10.1039/C6EM00268D
Alexander HC, Gersich FM, Mayes MA (1985) Acute toxicity of four phenoxy herbicides to aquatic organisms. Bull Environ Contam Toxicol 35:314–321. https://doi.org/10.1007/BF01636516
Arcaute CR, Ossana NA, Pérez-Iglesias JM et al (2019) Auxinic herbicides induce oxidative stress on Cnesterodon decemmaculatus (Pisces: Poeciliidae). Environ Sci Pollut Res 26:20485–20498. https://doi.org/10.1007/s11356-019-05169-z
Banaoui A, El Hamidi F, Kaaya A et al (2015) Assessment of multimarker responses in Perna perna, Mytilus galloprovincialis and Donax trunculus bivalves exposed to malathion and 2,4- dichlorophenoxyacetic acid pesticides. J Mater Environ Sci 6:1678–1683
Barriuso E, Feller C, Calvet R, Cerri C (1992) Sorption of atrazine, terbutryn and 2,4-D herbicides in two Brazilian Oxisols. Geoderma 53:155–167. https://doi.org/10.1016/0016-7061(92)90028-6
Bartlett AJ, Struger J, Grapentine LC, Palace VP (2016) Examining impacts of current-use pesticides in Southern Ontario using in situ exposures of the amphipod Hyalella azteca. Environ Toxicol Chem 35:1224–1238. https://doi.org/10.1002/etc.3265
Beggel S, Werner I, Connon RE, Geist JP (2010) Sublethal toxicity of commercial insecticide formulations and their active ingredients to larval fathead minnow (Pimephales promelas). Sci Total Environ 408:3169–3175. https://doi.org/10.1016/j.scitotenv.2010.04.004
Benli AÇK, Sarıkaya R, Sepici-Dincel A, Selvi M, Şahin D, Erkoç F (2007) Investigation of acute toxicity of (2,4-dichlorophenoxy) acetic acid (2,4-D) herbicide on crayfish (Astacus leptodactylus Esch. 1823). Pestic Biochem Physiol 88:296–299. https://doi.org/10.1016/j.pestbp.2007.01.004
Bobé A, Meallier P, Cooper J-F, Coste CM (1998) Kinetics and mechanisms of abiotic degradation of fipronil (hydrolysis and photolysis). J Agric Food Chem 46:2834–2839. https://doi.org/10.1021/jf970874d
Bolan NS, Baskaran S (1996) Biodegradation of 2,4-D herbicide as affected by Its adsorption-desorption behaviour and microbial activity of soils. Soil Res 34:1041–1053. https://doi.org/10.1071/sr9961041
Brazil (1989) Law no 7.802 from 11 July 1989
Brennan AA, Harwood AD, You J, Landrum PF, Lydy MJ (2009) Degradation of fipronil in anaerobic sediments and the effect on porewater concentrations. Chemosphere 77:22–28. https://doi.org/10.1016/j.chemosphere.2009.06.019
Browne AM, Moore PA (2014) The effects of sublethal levels of 2,4-dichlorophenoxyacetic acid herbicide (2,4-D) on feeding behaviors of the crayfish O. rusticus. Arch Environ Contam Toxicol 67:234–244. https://doi.org/10.1007/s00244-014-0032-8
Carvalho WF, Ruiz de Arcaute C, Torres L, de Melo e Silva D, Soloneski S, Larramendy ML (2020) Genotoxicity of mixtures of glyphosate with 2,4-dichlorophenoxyacetic acid chemical forms towards Cnesterodon decemmaculatus (Pisces, Poeciliidae). Environ Sci Pollut Res 27:6515–6525. https://doi.org/10.1007/s11356-019-07379-x
Castiglioni D d S, Bond-Buckup G (2007) Reproductive strategies of two sympatric species of Hyalella Smith, 1874 (Amphipoda, Dogielinotidae) in laboratory conditions. J Nat Hist 41:1571–1584. https://doi.org/10.1080/00222930701464604
Castiglioni D d S, Streck MT, Rodrigues SG et al (2018) Reproductive strategies of a population of a freshwater amphipod (Crustacea, Amhipoda, Hyalellidae) from southern Brazil. Biota Neotropica 18. https://doi.org/10.1590/1676-0611-bn-2017-0470
CETESB (2018) Qualidade das águas interiores no estado de São Paulo 2017. Série Relatórios / CETESB, São Paulo
CETESB (2019) Diagnóstico da contaminação de águas superficiais, subterrâneas e sedimentos por agrotóxicos. CETESB, São Paulo
Chaulet F da C, Barcellos HH de A, Fior D, et al (2019) Glyphosate- and fipronil-based agrochemicals and their mixtures change zebrafish behavior. Arch Environ Contam Toxicol 77:443–451. https://doi.org/10.1007/s00244-019-00644-7
Cooper WE (1965) Dynamics and production of a natural population of a freshwater amphipod, Hyalella azteca. Ecol Monogr 35:377–394. https://doi.org/10.2307/1942147
Dalhoff K, Gottardi M, Rinnan Å, Rasmussen JJ, Cedergreen N (2018) Seasonal sensitivity of Gammarus pulex towards the pyrethroid cypermethrin. Chemosphere 200:632–640. https://doi.org/10.1016/j.chemosphere.2018.02.153
Dallarés S, Dourado P, Sanahuja I, Solovyev M, Gisbert E, Montemurro N, Torreblanca A, Blázquez M, Solé M (2020) Multibiomarker approach to fipronil exposure in the fish Dicentrarchus labrax under two temperature regimes. Aquat Toxicol 219:105378. https://doi.org/10.1016/j.aquatox.2019.105378
Demcheck DK (2003) Fipronil and degradation products in the rice-producing areas of the Mermentau River Basin, Louisiana, February-September 2000 [electronic resource] / [by Dennis K. Demcheck and Stanley C. Skrobialowski]. U.S. Dept. of the Interior, U.S. Geological Survey, [Reston, Va.]
DeQuattro ZA, Karasov WH (2016) Impacts of 2,4-dichlorophenoxyacetic acid aquatic herbicide formulations. Environ Toxicol Chem 35:1478–1488. https://doi.org/10.1002/etc.3293
Fairchild JF, Ruessler DS, Haverland PS, Carlson AR (1997) Comparative sensitivity of Selenastrum capricornutum and Lemna minor to sixteen herbicides. Arch Environ Contam Toxicol 32:353–357. https://doi.org/10.1007/s002449900196
Fang W, Peng Y, Muir D, Lin J, Zhang X (2019) A critical review of synthetic chemicals in surface waters of the US, the EU and China. Environ Int 131:104994. https://doi.org/10.1016/j.envint.2019.104994
Farah MA, Ateeq B, Ali MN, Sabir R, Ahmad W (2004) Studies on lethal concentrations and toxicity stress of some xenobiotics on aquatic organisms. Chemosphere 55:257–265. https://doi.org/10.1016/j.chemosphere.2003.10.063
Figueirêdo LP, Athayde DB, Daam MA et al (2020) Impact of temperature on the toxicity of Kraft 36 EC® (a.s. abamectin) and Score 250 EC® (a.s. difenoconazole) to soil organisms under realistic environmental exposure scenarios. Ecotoxicol Environ Saf 194:110446. https://doi.org/10.1016/j.ecoenv.2020.110446
Freitas JS, Girotto L, Goulart BV, Alho LOG, Gebara RC, Montagner CC, Schiesari L, Espíndola ELG (2019) Effects of 2,4-D-based herbicide (DMA® 806) on sensitivity, respiration rates, energy reserves and behavior of tadpoles. Ecotoxicol Environ Saf 182:109446. https://doi.org/10.1016/j.ecoenv.2019.109446
Gaaied S, Oliveira M, Le Bihanic F et al (2019) Gene expression patterns and related enzymatic activities of detoxification and oxidative stress systems in zebrafish larvae exposed to the 2,4-dichlorophenoxyacetic acid herbicide. Chemosphere 224:289–297. https://doi.org/10.1016/j.chemosphere.2019.02.125
George JP, Hingorani HG, Rao KS (1982) Herbicide toxicity to fish-food organisms. Environ Pollut Ser A Ecol Biol 28:183–188. https://doi.org/10.1016/0143-1471(82)90074-5
Giusto A, Somma LA, Ferrari L (2012) Cadmium toxicity assessment in juveniles of the Austral South America amphipod Hyalella curvispina. Ecotoxicol Environ Saf 79:163–169. https://doi.org/10.1016/j.ecoenv.2011.12.020
Glazier DS (2000) Is fatter fitter? Body storage and reproduction in ten populations of the freshwater amphipod Gammarus minus. Oecologia 122:335–345. https://doi.org/10.1007/s004420050039
González ER, Watling L (2003) A new species of Hyalella from Brazil (Crustacea: Amphipoda: Hyalellidae), with redescriptions of three other species in the genus. J Nat Hist 37:2045–2076. https://doi.org/10.1080/00222930210133237
Gottardi M, Birch MR, Dalhoff K, Cedergreen N (2017) The effects of epoxiconazole and α-cypermethrin on Daphnia magna growth, reproduction, and offspring size. Environ Toxicol Chem 36:2155–2166. https://doi.org/10.1002/etc.3752
Goulart BV, Vizioli BDC, Espindola ELG, Montagner CC (2020) Matrix effect challenges to quantify 2,4-D and fipronil in aquatic systems. Environ Monit Assess 192:797. https://doi.org/10.1007/s10661-020-08776-3
Green RM, Abdelghani AA (2004) Toxicity of a mixture of 2,4-dichlorophenoxyacetic acid and monosoduim methanearsonate to the red swamp crawfish, Procambarus clarkii. Int J Environ Res Public Health 1:35–38. https://doi.org/10.3390/ijerph2004010035
Grützmacher D, Grützmacher A, Agostinetto D et al (2008) Monitoring of pesticides in two water sources in southern Brazil. Revista Brasileira de Engenharia Agrícola e Ambiental 12:632–637. https://doi.org/10.1590/S1415-43662008000600010
Hansen HP, Koroleff F (2007) Determination of nutrients. In: Methods of Seawater Analysis. Wiley, pp 159–228
Hayasaka D, Korenaga T, Suzuki K, Sánchez-Bayo F, Goka K (2012) Differences in susceptibility of five cladoceran species to two systemic insecticides, imidacloprid and fipronil. Ecotoxicology 21:421–427. https://doi.org/10.1007/s10646-011-0802-2
IBAMA (2018) Relatórios de comercialização de agrotóxicos (Boletim 2018)
Islam F, Wang J, Farooq MA, Khan MSS, Xu L, Zhu J, Zhao M, Muños S, Li QX, Zhou W (2018) Potential impact of the herbicide 2,4-dichlorophenoxyacetic acid on human and ecosystems. Environ Int 111:332–351. https://doi.org/10.1016/j.envint.2017.10.020
Jonker MJ, Svendsen C, Bedaux JJM, Bongers M, Kammenga JE (2005) Significance testing of synergistic/antagonistic, dose level-dependent, or dose ratio-dependent effects in mixture dose-response analysis. Environ Toxicol Chem 24:2701–2713. https://doi.org/10.1897/04-431R.1
Kader HA, Thayumanavan B, Krishnaswamy S (1976) The relative toxicities of ten biocides on Spicodiaptomus chelospinus. Comp Physiol Ecol 3:78–82
Kashian DR, Dodson SI (2002) Effects of common-use pesticides on developmental and reproductive processes in Daphnia. Toxicol Ind Health 18:225–235. https://doi.org/10.1191/0748233702th146oa
Konwick BJ, Garrison AW, Black MC, Avants JK, Fisk AT (2006) Bioaccumulation, biotransformation, and metabolite formation of fipronil and chiral legacy pesticides in rainbow trout. Environ Sci Technol 40:2930–2936. https://doi.org/10.1021/es0600678
Levchenko MA, Silivanova EA (2019) Synergistic and antagonistic effects of insecticide binary mixtures against house flies (Musca domestica). Regul Mech Biosyst 10:75–82. https://doi.org/10.15421/021912
Li K, Wu J-Q, Jiang L-L, Shen LZ, Li JY, He ZH, Wei P, Lv Z, He MF (2017) Developmental toxicity of 2,4-dichlorophenoxyacetic acid in zebrafish embryos. Chemosphere 171:40–48. https://doi.org/10.1016/j.chemosphere.2016.12.032
Liber K, Call DJ, Dawson TD, Whiteman FW, Dillon TM (1996) Effects of Chironomus tentans larval growth retardation on adult emergence and ovipositing success: implications for interpreting freshwater sediment bioassays. Hydrobiologia 323:155–167. https://doi.org/10.1007/BF00007844
Lizotte RE, Knight SS, Shields FD, Bryant CT (2009) Effects of an atrazine, metolachlor and fipronil mixture on Hyalella azteca (Saussure) in a modified backwater wetland. Bull Environ Contam Toxicol 83:836–840. https://doi.org/10.1007/s00128-009-9850-1
Lopes LF d P, Agostini VO, Guimarães SS, Muxagata E (2018) Evaluation of the effect of antimicrobials in marine cultures, using the copepod Acartia tonsa as a bioindicator. Chem Ecol 34:747–761. https://doi.org/10.1080/02757540.2018.1482886
Loureiro S, Svendsen C, Ferreira ALG, Pinheiro C, Ribeiro F, Soares AMVM (2010) Toxicity of three binary mixtures to Daphnia magna: comparing chemical modes of action and deviations from conceptual models. Environ Toxicol Chem 29:1716–1726. https://doi.org/10.1002/etc.198
MAPA (2019) AGROFIT. In: Brazil. Ministry of Agriculture Livestock and Food Supply. http://agrofit.agricultura.gov.br/agrofit_cons/principal_agrofit_cons. Accessed 29 Jun 2019
Marchesan E, Sartori GMS, de Avila LA et al (2010) Residues of pesticides in the water of the depression central rivers in the State of Rio Grande do Sul, Brazil. Ciênc Rural 40:1053–1059. https://doi.org/10.1590/S0103-84782010005000078
Maul JD, Brennan AA, Harwood AD, Lydy MJ (2008) Effect of sediment-associated pyrethroids, fipronil, and metabolites on Chironomus tentans growth rate, body mass, condition index, immobilization, and survival. Environ Toxicol Chem 27:2582–2590. https://doi.org/10.1897/08-185.1
Mize SV, Porter SD, Demcheck DK (2008) Influence of fipronil compounds and rice-cultivation land-use intensity on macroinvertebrate communities in streams of southwestern Louisiana, USA. Environ Pollut 152:491–503. https://doi.org/10.1016/j.envpol.2007.03.021
Monteiro HR, Pestana JLT, Novais SC, Leston S, Ramos F, Soares AMVM, Devreese B, Lemos MFL (2019) Assessment of fipronil toxicity to the freshwater midge Chironomus riparius: molecular, biochemical, and organismal responses. Aquat Toxicol 216:105292. https://doi.org/10.1016/j.aquatox.2019.105292
Moreira RA, Araújo CVM, Junio da Silva Pinto T et al (2021) Fipronil and 2,4-D effects on tropical fish: Could avoidance response be explained by changes in swimming behavior and neurotransmission impairments? Chemosphere 263:127972. https://doi.org/10.1016/j.chemosphere.2020.127972
Moreira RA, de Araujo GS, Silva ARRG, Daam MA, Rocha O, Soares AMVM, Loureiro S (2020a) Effects of abamectin-based and difenoconazole-based formulations and their mixtures in Daphnia magna: a multiple endpoint approach. Ecotoxicology. 29:1486–1499. https://doi.org/10.1007/s10646-020-02218-z
Moreira RA, Rocha GS, da Silva LCM, Goulart BV, Montagner CC, Melão MGG, Espindola ELG (2020b) Exposure to environmental concentrations of fipronil and 2,4-D mixtures causes physiological, morphological and biochemical changes in Raphidocelis subcapitata. Ecotoxicol Environ Saf 206:111180. https://doi.org/10.1016/j.ecoenv.2020.111180
Moreira RA, Rocha O, Pinto TJ d S et al (2020c) Life-history traits response to effects of fish predation (kairomones), fipronil and 2,4-d on neotropical cladoceran Ceriodaphnia silvestrii. Arch Environ Contam Toxicol 79:298–309. https://doi.org/10.1007/s00244-020-00754-7
Mugni H, Ronco A, Bonetto C (2011) Insecticide toxicity to Hyalella curvispina in runoff and stream water within a soybean farm (Buenos Aires, Argentina). Ecotoxicol Environ Saf 74:350–354. https://doi.org/10.1016/j.ecoenv.2010.07.030
Nagy K, Duca RC, Lovas S, Creta M, Scheepers PTJ, Godderis L, Ádám B (2020) Systematic review of comparative studies assessing the toxicity of pesticide active ingredients and their product formulations. Environ Res 181:108926. https://doi.org/10.1016/j.envres.2019.108926
Nguyen LTH, Muyssen BTA, Janssen CR (2012) Single versus combined exposure of Hyalella azteca to zinc contaminated sediment and food. Chemosphere 87:84–90. https://doi.org/10.1016/j.chemosphere.2011.11.066
OECD (2011) Test No. 235: Chironomus sp., acute immobilisation test
Oruç EÖ, Üner N (2000) Combined effects of 2,4-D and azinphosmethyl on antioxidant enzymes and lipid peroxidation in liver of Oreochromis niloticus. Comp Biochem Physiol C: Pharmacol Toxicol Endocrinol 127:291–296. https://doi.org/10.1016/S0742-8413(00)00159-6
Park K, Park J, Kim J, Kwak I-S (2010) Biological and molecular responses of Chironomus riparius (Diptera, Chironomidae) to herbicide 2,4-D (2,4-dichlorophenoxyacetic acid). Comparat Biochemi Physiol C Toxicol Pharmacol 151:439–446. https://doi.org/10.1016/j.cbpc.2010.01.009
Paul E, Johnson S, Skinner KM (2006) Fish and invertebrate sensitivity to the aquatic herbicide Aquakleen®. J Freshw Ecol 21:163–168. https://doi.org/10.1080/02705060.2006.9664109
Pavlaki MD, Pereira R, Loureiro S, Soares AMVM (2011) Effects of binary mixtures on the life traits of Daphnia magna. Ecotoxicol Environ Saf 74:99–110. https://doi.org/10.1016/j.ecoenv.2010.07.010
Pinheiro A, Silva M, Kraisch R (2010) Presença de pesticidas em águas superficiais e subterrâneas na bacia do Itajaí, SC. REGA 7:17–26. https://doi.org/10.21168/rega.v7n2.p17-26
Qu H, Ma R, Liu D, Gao J, Wang F, Zhou ZQ, Wang P (2016) Environmental behavior of the chiral insecticide fipronil: Enantioselective toxicity, distribution and transformation in aquatic ecosystem. Water Res 105:138–146. https://doi.org/10.1016/j.watres.2016.08.063
Richardi VS, Vicentini M, Morais GS, Rebechi D, da Silva TA, Fávaro LF, Navarro-Silva MA (2018) Effects of phenanthrene on different levels of biological organization in larvae of the sediment-dwelling invertebrate Chironomus sancticaroli (Diptera: Chironomidae). Environ Pollut 242:277–287. https://doi.org/10.1016/j.envpol.2018.06.091
Sanders HO (1970) Toxicities of some herbicides to six species of freshwater crustaceans. J Water Pollut Control Fed 42:1544–1550
Scherer C, Wolf R, Völker J, Stock F, Brennhold N, Reifferscheid G, Wagner M (2020) Toxicity of microplastics and natural particles in the freshwater dipteran Chironomus riparius: Same same but different? Sci Total Environ 711:134604. https://doi.org/10.1016/j.scitotenv.2019.134604
Schlenk D, Huggett DB, Allgood J et al (2001) Toxicity of fipronil and its degradation products to Procambarus sp.: field and laboratory studies. Arch Environ Contam Toxicol 41:325–332. https://doi.org/10.1007/s002440010255
Schreiner VC, Szöcs E, Bhowmik AK, Vijver MG, Schäfer RB (2016) Pesticide mixtures in streams of several European countries and the USA. Sci Total Environ 573:680–689. https://doi.org/10.1016/j.scitotenv.2016.08.163
Seuge J, Bluzat R, Rodriguez-Ruiz FJ (1970) Effets d’un melange herbicide (2,4-D ET 2,4,5-T): toxicite aigue sur 4 especes d’invertebres liminiques; toxicite chronique chez le mollusque pulmone Lymnea. Environ Pollut 16:87–104. https://doi.org/10.1016/0013-9327(78)90124-6
Shan Z, Wang L, Cai D, Gong R, Zhu Z, Yu F (2003) Impact of fipronil on crustacean aquatic organisms in a paddy field-fishpond ecosystem. Bull Environ Contam Toxicol 70:0746–0752. https://doi.org/10.1007/s00128-003-0046-9
Silva LCM, Moreira RA, Pinto TJS, Ogura AP, Yoshii MPC, Lopes LFP, Montagner CC, Goulart BV, Daam MA, Espíndola ELG (2020) Acute and chronic toxicity of 2,4-D and fipronil formulations (individually and in mixture) to the Neotropical cladoceran Ceriodaphnia silvestrii. Ecotoxicology. 29:1462–1475. https://doi.org/10.1007/s10646-020-02275-4
Stehr CM, Linbo TL, Incardona JP, Scholz NL (2006) The developmental neurotoxicity of fipronil: notochord degeneration and locomotor defects in zebrafish embryos and larvae. Toxicol Sci 92:270–278. https://doi.org/10.1093/toxsci/kfj185
Tomlin C (1994) The Pesticide Manual, 10th edn. The royal society of chemistry, Cambridge
Torres SHS, Bastos-Pereira R, Bueno AA d P et al (2015) Reproductive aspects of Hyalella carstica (Amphipoda: Hyalellidae) in a natural environment in southeastern Brazil. Nauplius 23:159–165. https://doi.org/10.1590/S0104-64972015002325
Ullah S, Ejaz M, Ali Shad S (2017) Study of synergism, antagonism, and resistance mechanisms in insecticide-resistant Oxycarenus hyalinipennis (Hemiptera: Lygaeidae). J Econ Entomol 110:615–623. https://doi.org/10.1093/jee/tow302
Van Meter RJ, Glinski DA, Purucker ST, Henderson WM (2018) Influence of exposure to pesticide mixtures on the metabolomic profile in post-metamorphic green frogs (Lithobates clamitans). Sci Total Environ 624:1348–1359. https://doi.org/10.1016/j.scitotenv.2017.12.175
Volz DC, Wirth EF, Fulton MH, Scott GI, Strozier E, Block DS, Ferry JL, Walse SS, Chandler GT (2003) Effects of fipronil and chlorpyrifos on endocrine-related endpoints in female grass shrimp (Palaemonetes pugio). Bull Environ Contam Toxicol 71:0497–0503. https://doi.org/10.1007/s00128-003-8920-z
Weston DP, Lydy MJ (2014) Toxicity of the insecticide fipronil and its degradates to benthic macroinvertebrates of urban streams. Environ Sci Technol 48:1290–1297. https://doi.org/10.1021/es4045874
Ying G-G, Kookana R (2002) Laboratory and field studies on the degradation of fipronil in a soil. Soil Res 40:1095–1102. https://doi.org/10.1071/sr02018
Ying GG, Kookana RS (2001) Sorption of fipronil and its metabolites on soils from South Australia. J Environ Sci Health B 36:545–558. https://doi.org/10.1081/PFC-100106184
Zhu G, Wu H, Guo J, Kimaro FME (2004) Microbial degradation of fipronil in clay loam soil. Water Air Soil Pollut 153:35–44. https://doi.org/10.1023/B:WATE.0000019928.67686.b1
Data and material availability
All data generated or analyzed during this study are included in this published article (and its supplementary information files) and are available from the corresponding author on reasonable request.
Funding
The work was supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP, Brazil, grant n. 2015/18790-3). T.J.S.P., L.C.M.S., M.P.C.Y., and B.V.G. have a Ph.D. scholarship from the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES). R.A.M. has a post-doctoral grant from FAPESP (2017/24126-4) and P.D.F and V.L.S.R have scientific initiation grants (FAPESP 2019/04198-6 and 2016/25611-0, respectively). Financial support was also provided to M.A.D. by the Portuguese government (Fundação para a Ciência e Tecnologia; FCT) through the research unit UIDB/04085/2020 (CENSE).
Author information
Authors and Affiliations
Contributions
Thandy Junio da Silva Pinto: Conceptualization, Methodology, Formal analysis, Investigation, Writing—Original Draft
Raquel Aparecida Moreira: Writing—Review and Editing
Laís Conceição Menezes da Silva: Investigation
Maria Paula Cardoso Yoshii: Investigation
Bianca Veloso Goulart: Resources, Investigation
Priscille Dreux Fraga: Investigation
Victor Luiz da Silva Rolim: Investigation
Cassiana Carolina Montagner: Resources; Writing—Review and Editing
Michiel Adriaan Daam: Conceptualization, Methodology, Writing—Review and Editing
Evaldo Luiz Gaeta Espindola: Conceptualization; Methodology, Writing—Review and Editing; Project administration; Funding acquisition
Corresponding author
Ethics declarations
Ethics approval and consent to participate
Not applicable.
Consent for publication
Not applicable.
Competing interests
Not applicable.
Additional information
Responsible Editor: Bruno Nunes
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
ESM 1
(DOCX 156 kb)
Rights and permissions
About this article
Cite this article
da Silva Pinto, T.J., Moreira, R.A., da Silva, L.C.M. et al. Toxicity of fipronil and 2,4-D formulations (alone and in a mixture) to the tropical amphipod Hyalella meinerti. Environ Sci Pollut Res 28, 38308–38321 (2021). https://doi.org/10.1007/s11356-021-13296-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11356-021-13296-9