Abstract
Objective
The use of chlorine dioxide (ClO2) increased in the last year to prevent SARS-CoV-2 infection due to its use as disinfectant and therapeutic human treatments against viral infections. The absence of toxicological studies and sanitary regulation of this contaminant represents a serious threat to human and environmental health worldwide. The aim of this study was to evaluate the acute toxicity and sublethal effects of ClO2 on tadpoles of Trachycephalus typhonius, which is a common bioindicator species of contamination from aquatic ecosystems.
Materials and methods
Median lethal concentration (LC50), the lowest-observed effect concentration (LOEC), and the no-observed effect concentration (NOEC) were performed. Acetylcholinesterase (AChE) and glutathione-S-transferase (GST) activities, swimming behavior parameters, and cardiac rhythm were estimated on tadpoles of concentrations ≤ LOEC exposed at 24 and 96 h. ANOVA and Dunnett’s post-hoc comparisons were performed to define treatments significance (p ≤ 0.05).
Results
The LC50 of ClO2 was 4.17 mg L−1 (confidence limits: 3.73–4.66). In addition, NOEC and LOEC values were 1.56 and 3.12 mg L−1 ClO2, respectively, at 48 h. AChE and GST activities, swimming parameters, and heart rates increased in sublethal exposure of ClO2 (0.78–1.56 mg L−1) at 24 h. However, both enzyme activities and swimming parameters decreased, whereas heart rates increased at 96 h.
Conclusion
Overall, this study determined that sublethal concentrations of ClO2 produced alterations on antioxidant systems, neurotoxicity reflected on swimming performances, and variations in cardiac rhythm on treated tadpoles. Thus, our findings highlighted the need for urgent monitoring of this chemical in the aquatic ecosystems.
Graphical abstract
Similar content being viewed by others
Avoid common mistakes on your manuscript.
Introduction
Chlorine dioxide (ClO2) is a disinfection agent that is massively used due to its high oxidizing capacity that deactivates chlorine-resistant pathogens and prevents biofilm formation [1]. In the last decades, drinking water treatments and industrial processes increase the use of ClO2, which replaces other chlorine disinfectants [2, 3]. Several studies determined that the consumption of ClO2 solutions is a preventive and therapy tool against different viral human infections, such as HIV/AIDS [4, 5]. The occurrence of ClO2 had an exponential increase in surface water during the last two years due to its use as treatment and preventive therapy against SARS-CoV-2 [6, 7]. Although ClO2 lacks of scientific and sanitary approval as medical treatment against SARS-CoV-2 for human consumption, the ClO2 began to be commonly commercialized specially in Latin America [8,9,10]. Moreover, the increased use of ClO2, together with other pharmaceutical products and disinfection agents poses a serious threat to aquatic systems, animal and human health [11, 12].
In addition, treatment protocols to prevent waterborne pathogens in public water systems and drinking water was determined by the World Health Organization, including also those treatments with ClO2 water. It has been reported that a concentration higher than 0.5 mg L−1 of free chlorine and 2.19 mg L−1 ClO2 inactivate SARS viruses. In general, residual chlorine and ClO2 concentrations in wastewater treatments do not exceed 6.5 mg L−1 and 10 mg L−1, respectively [6, 13, 14]. However, solutions of 20 mg L−1 ClO2 are recommended for SARS-CoV-2 inactivation for domestic use as disinfectant, whereas solutions over 500 mg L−1 ClO2 are suggested for hospital and health unit care disinfections. Thus, maximum concentrations of 0.8 mg L−1 ClO2 and 1.0 mg L−1 chlorite ion for drinking waters were recommended by U.S.EPA recommends [15].
During COVID-19 pandemic, sanitation with chlorinated products increased in domestic wastewater, drinking water, and surface water, which pose a healthy human and environmental risk [16]. For example, concentrations of chlorine ranged between 1.5 and 4−5 mg L−1 in wastewater treatment plants were used for preventive measure in China [17]. The same study recorded residual chlorine concentration up to 0.4 mg L−1 in some Chinese lakes, where had not been detected before the actual COVID pandemic. This concentration produced acute toxicity on freshwater organism [17].
The effect of ClO2 on virus, bacteria, algae, and plankton has been analyzed during last decades [18,19,20]. However, few studies described ClO2 effect on wild organisms, such as aquatic vertebrates. The ClO2 lethal toxicity has been reported for the early stages of several fishes [21, 22], chironomid larvae [23], and nematodes [24]. It is important to note that ClO2 toxicity data obtained for common sentinel organisms of model organisms (e.g., Cyprinidae, Daphnidae, Brachionidae, and Hyalelladae) is generally used by U.S.EPA water quality criteria method [25]. The monitoring of ClO2 on aquatic environments and ecotoxicological studies that include lethally and sublethal parameters of other sentinel species is needed. To the best of our knowledge, no information exists on ClO2 toxicity on amphibian tadpoles.
Amphibians present the highest decrease of population than other vertebrates worldwide, due to their complex life cycle that involves physiological, morphological, and behavioral changes during metamorphosis. These biological traits increase amphibian vulnerability to man-made environmental modifications [26,27,28]. Contaminants (e.g., pharmaceuticals, agrochemicals, health care products) affect survival and produce several sublethal effects on amphibians [29, 30]. Death and sublethal effects at different biological levels (e.g., cell, endocrinology, metabolic and behavior alterations, metamorphosis rates, malformations, among others) were determined for amphibian tadpoles [31,32,33]. In addition, the cholinesterase (ChEs) enzyme activity is useful to indicate the presence of neurotoxic substances in amphibian tadpoles [34]. Moreover, acetylcholinesterase (AChE) activity is inhibited in presence of neurotoxic substances, leading to acetylcholine accumulation and producing alterations in nerve function that can be lethal on exposed organisms [35]. The alteration in normal AChE activity has been linked with neurotoxic effects on locomotor activity in different invertebrates and vertebrates [36, 37]. Thus, glutathione-S-transferase (GST) activity is used to assess the detoxification process of contaminants [38]. The GST activity is frequently quantified in amphibian tadpoles exposed to several contaminants, such as pharmaceutical residues [39, 40]. However, other sublethal effects of tadpole regarding to physiology, such as heart function and rhythm are poorly explored [41]. Tadpoles’ heart rate is considered novel biomarkers in amphibian ecotoxicology [32, 42] and could reflect of what happens at biochemical, physiological, and ecological levels in aquatic organisms exposed to different contaminants [43].
The aim of this study was to evaluate the acute and sublethal effects of ClO2 on Trachycephalus typhonius tadpoles. It has been hypothesized that the ClO2 produces different biological changes on AChE and GST activities and physiological (cardiac rhythm and swimming behavior) endpoints.
Results and discussion
Mortality
LC50 of ClO2 was 4.17 mg L−1 (CL: 3.73–4.66) at 48 h. NOEC and LOEC values were 1.56 and 3.12 mg L−1 ClO2, respectively, at 48 h. The TU for ClO2 was 23.98, meaning a high acute toxicity substance (Class IV) in the hazard classification system.
The toxicity of ClO2 has been studied in alkaline solutions when it becomes chlorate [44]. In fishes (feather minnows Pimephales promelas), the LC50 was determined to be 0.19 mg L−1 of ClO2, whereas LC50 was recorded to 63.38 mg L−1 of chlorite within 24 h of exposure [45, 46]. In amphibians, the LC50 values are only reported for chlorite concentration that varied between 65.69 mg L−1 (Lithobates pipiens) and 149.60 mg L−1 (Anaxyrus americanus). Chlorite is much less toxic than its precursor chlorine [25]. No estimation could be made regarding to the toxicity of ClO2 in terms of values of LC50 obtained in this study for T. thyphonius with respect to other amphibian species because mortality values of ClO2 are unknown for other anurans. Time of exposure, acclimation temperature, and chlorine species can be considered as factors that influence the ClO2 toxicity [46]. For example, the LC50 values of ClO2 varied between 0.41 and 0.23 mg L−1 for different instar larvae (1st to 4th) of chironomid species at 24 h [23], whereas these values were 0.022 mg L−1 for planktonic crustacean (Daphnia magna) and 47 mg L−1 for mosquitoes larvae [25].
Enzyme activities
AChE activity significantly increased (51%, F = 7.18, p < 0.01) in 0.78 mg L−1 ClO2 treated tadpoles with respect to CO (Dunnett’s test p < 0.01) at 24 h (Fig. 1A, Table S1 in supplementary material). In contrast, AChE activity significantly decreased (F = 5.16, p < 0.01) between 32.15 and 36.51% in ClO2 treated tadpoles with respect to CO (Dunnett’s test p < 0.01) after 96 h of exposure (Fig. 1B, Table S1). In addition, exposure to ClO2 also altered the AChE activity of larvae of indianmeal moth (Plodia interpunctella) [47]. Matsushita et al. [48] who recognized ClO2 as insecticide, reported that ClO2 produced a fast increase of AChE activities in P. interpunctella than those insecticide-based-malathion or -methidathion formulations. Moreover, the AChE activities decreased with increasing chlorination time in ClO2 treated T. thyphonius tadpoles at 24 and after 96 h, which may be explained in terms of enzyme synthesis as a response to initial inhibition [49]. It is important to note that AChE is involved in the cleavage of the acetylcholine neurotransmitter within the synaptic cleft. An inhibition of AChE activity leads to acetylcholine accumulation, hyperstimulation of nicotinic and muscarinic receptors, and disrupted neurotransmission, consequently affecting the animal behavior [50].
Acetylcholinesterase (AChE) and glutathione-S-transferase (GST) activities (nmol min−1 mg−1 protein) in Trachycephalus typhonius tadpoles from control (CO) and chlorine dioxide (ClO2 mg L−1) treatments, at 24 h (left) and 96 h (right) of exposure. p < 0.05 (*) and p < 0.01 (**) indicates statistical differences with respect to CO
The GST activity of ClO2 treated tadpoles significantly increased (F = 4.47, p < 0.05) in 0.78 mg L−1 and 1.56 mg L−1 concentrations with respect to CO (Dunnett’s test p < 0.05) at 24 h (87.84% and 79.41%, respectively; Fig. 1B, Table S1). There was a positive correlation between the responses of both AChE and GST enzyme activities (r Spearman = 1; p < 0.01) at 24 h. The mean GST activity significantly decreased (30%, F = 8.19, p < 0.01) in 0.78 mg L−1 ClO2 concentration with respect to CO (Dunnett’s test p < 0.01) after 96 h (Fig. 1B, Table S1).
The ClO2 is recognized as a powerful oxidant that affects the first line of antioxidant defense, which is mediated by GST in animals [51]. Besides, the GST plays a pivotal role in cellular detoxification via conjugation of their electrophilic group with the GSH nucleophilic group of harmful contaminants. In contrast to our results, Elia et al. [52] reported an increase in liver GST activity in carp fishes (Cyprinus carpio) exposed to 1.6 mg L−1 ClO2 after 20 days, suggesting that GST acts as a useful biomarker of disinfectants-mediated oxidative stress. The significant decrease in the GST activity in treated tadpoles exposed to 0.76 mg L−1 of ClO2 after 96 h, could be related to the availability of substrate of this enzyme. It is well known that ClO2 oxidizes the glutathione and produces several products that block substrates for normal reaction [53].
Swimming activity
The mean total distance of ClO2 treated tadpoles significantly increased (F = 39.73 p < 0.01) in 0.78 and 3.12 mg L−1 ClO2 concentrations (103.19% and 139.36%, respectively) with respect to CO (Dunnett’s test p < 0.01) at 24 h (Fig. 2A, Table S1). The mean speed of ClO2 treated tadpoles increased (F = 77.18, p < 0.01) in all concentrations (103.16; 78.29 and 139.36% for 0.78; 1.56 and 3.12 mg L−1, respectively; Fig. 2B, Table S1) with respect to CO (Dunnett’s test p < 0.01). The global activity significantly increased (F = 53,02, p < 0.05) at 1.56 and 3.12 mg L−1 ClO2 concentrations (456.57 and 994.48%, respectively; Fig. 2C, Table S1) with respect to CO (Dunnett’s test p < 0.05). The effects of ClO2 concentrations on global activity were negatively correlated with both enzyme activities (r Spearman = − 1; p < 0.01) at 24 h.
Swimming activity of Trachycephalus typhonius tadpoles of control (CO) and chlorine dioxide (ClO2 mg L−1) treatments at 24 h (left) and after 96 h (right) of exposure: A Total distance moved (cm); B Mean speed (cm s−1); C Global activity (cm2), showing the trajectory of individual in each concentration. p < 0.05 (*) and p < 0.01 (**) indicates statistical differences with respect to CO
The mean total distance significantly decreased (F = 44.03, p < 0.05) in 1.56 and 3.12 mg L−1 ClO2 treated tadpoles (43.67 and 66.51%, respectively) with respect to CO (Dunnett’s test p < 0.05) after 96 h (Fig. 2A, Table S1). Also, the mean speed significantly decreased (F = 18.04, p < 0.05) in the same concentrations (44.4 and 58.34%, respectively) with respect to CO (Dunnett’s test p < 0.05; Fig. 2B, Table S1)). Similarly, the global activity significantly decreased (F = 25.8, p < 0.05) in 0.78 mg L−1 ClO2 treated tadpoles (64.76%) with respect to CO (Dunnett’s test p < 0.05) after 96 h (Fig. 2C, Table S1).
The observed responses of swimming activity parameters indicated that ClO2 affects the mechanisms involved in swimming behavior at different times (24 and after 96 h). For example, swimming behavior has been related to AChE activity in invertebrates, fish, and amphibian larvae exposed to different contaminants [32, 34, 39, 54, 55]. In this sense, swimming behavior observed in ClO2 treated tadpoles at 24 h may be related to neurotoxicity through AChE over-activation [50]. Conversely, decrease in swimming behavior observed in ClO2 treated tadpoles after 96 h could be related to the disruption of the normal nervous system function by inhibition of the AChE activity and the resulting transduction signal [56, 57]. Regarding to these behavior disparities, a higher expression of AChE genes may occur as a response to acetylcholine neurotransmitters accumulation [58].
Yonkos et al. [45] described gill pathology with epithelial lifting, hypertrophy, hyperplasia, lamellar fusion, and necrosis on feather minnow P. promelas exposed to ClO2. Histological alterations in gills may produce alteration in swimming activity due to its demands of oxygen [59]. In this sense, the decrease of swimming parameters after 96 h of exposure may be related to structural changes in the body of tadpoles such as gills that are related to physiological changes [60]. Moreover, Orme et al. [61] reported a reduced level of thyroxine in Sprague Dawley rats exposed to ClO2 during development and demonstrated anti-thyroid and neuro-behavioral effects during their development. Thyroid hormones play important roles in neuro-behavioral functioning during anuran development [32], however, studies regarding to thyrotoxicosis and neuro-behavioral performance are scarce.
Cardiac rhythm
The mean heart rate (heartbeats per minute) in 0.78 and 3.12 mg L−1 ClO2 treated tadpoles significantly increased (84.61% and 70.19%; F = 5.323, p < 0.01) with respect to CO (Dunnett’s test p < 0.01) at 24 h (Fig. 3, Table S1). Heart rates were positively related (r Spearman = 0.8; p < 0.01) to some swimming parameters (total distance, mean speed) and GST activities.
Heart rates (beats min−1) of Trachycephalus typhonius tadpoles of control (CO) and chlorine dioxide (ClO2 mg L−1) treatments at 24 h (left) and after 96 h (right) of exposure. p < 0.05 (*) and p < 0.01 (**) indicates statistical differences with respect to CO
Heart rate showed a significant (F = 7.25, p < 0.01) increase in treated tadpoles exposed to 0.78, 1.56 and 3.12 mg L−1 ClO2 concentrations after 96 h (75.88%, 84.93% and 69.64%, respectively) respect to CO (Dunnett’s test p < 0.01; Fig. 3, Table S1). Heart rate showed a negative correlation with AChE (r Spearman = − 1; p < 0.01). Results of Spearman correlation between biological endpoints were summarized in Table S2 (supplementary material). The cardiac rhythm is recognized as effective biomarkers for amphibian’s health in several ecotoxicological investigations [32, 39, 41, 62]. In the present study, heart rate increased in ClO2 treated tadpoles at 24 and after 96 h. In contrast, reduction of heartbeats was reported in ClO2 treated larvae of P. interpunctella [47]. The relation among heart rhythm, oxidative stress, and GST activities were determined in tadpoles exposed to other chemicals such as pyriproxyfen, however, mechanisms involved in these relations remain unclear [32]. A study on pigeons (Columba livia) suggested that ClO2 may increase the risk of cardiac rhythm alteration due to an increase in plasma cholesterol levels and size of thrombocytes in blood vessels [63].
Materials and methods
Chemical
Chlorine dioxide was synthesized as described by Hey et al. [64] and Chhetri et al. [65]. Demineralized water (400 mL) was mixed with 25 mL of 4% HCl and 25% NaClO2. This mixture was diluted to 1000 mL in demineralized water after an overnight reaction. This procedure produced an approximately 1 g L−1 chlorine dioxide solution, which was used for three replicates of stock solutions of 100 mg L−1 ClO2. The stability of concentration of stock solutions was verified by chlorine dioxide Sensing Strip (Insta-TEST, LaMotte®-USA) [66, 67] in lab conditions immediately after preparation and at every 24 h for three consecutive days. These observations were used for defining the renewal time of treatment solutions. The initial concentration (0 h) was reduced by 30% at 24 h, and more than half at 48 h, so it was decided to renew the solutions at every 48 h.
Organisms and experimental design
Tadpoles (Gosner Stage 28–32) of the veined tree frog Trachycephalus typhonius were sampled from temporary small ponds from the Paraná River floodplain, Argentina (31° 11′ 31″ S, 60° 9′ 29″ W). Authorization for tadpole sampling was given by the Ministerio de Ambiente de la Provincia de Santa Fe (No 02101–0018518-1). Site of tadpole sampling was contamination-free, as determined in a previous research of our lab [68]. Tadpoles were transported immediately to laboratory in dechlorinated tap water (DTW) and acclimatized. Tadpoles’ acclimation period was performed under controlled lab conditions of 12/12 h (light N100 Lx/dark photoperiod cycles) at 24 ± 2 °C for two days. Tadpole behavior such as eat and swim was checked daily.
The experiments followed regulations of ASIH [69]. Tadpoles were euthanized following the protocol of the Animal Euthanasia Guide proposed by the Institutional Animal Care and Use Committee and the Advisory Committee on Ethics and Safety in research of the Facultad de Bioquímica y Ciencias Biológicas of the Universidad Nacional del Litoral (No 388/06).
There is lack of environmental concentration data on aquatic ecosystems and doubts related to the fate of ClO2 on surface freshwater. For this reason, a first approximation to the toxicity of ClO2 on tadpoles was performed to determine the median lethal concentration (LC50), similarly as previously performed for different contaminants [39]. A first bioassay was carried out by exposing three replicate solutions each containing 12 tadpoles. The nominal concentrations of ClO2 assayed were: 12.5, 6.25, 3.125, 1.56, and 0.78 mg L−1. Mortality was recorded at 24 and 48 h to estimate LC50, lowest-observed effect concentration (LOEC), and no-observed effect concentration (NOEC) values.
The sublethal effects (enzyme activities, swimming behavior, and heart rhythm as biomarkers) were evaluated in tadpoles of treatments with ≤ LOEC at 24 and after 96 h of exposure. These effects were analyzed in subsequent bioassays with the same lab conditions. The solutions were renewed at 48 h in the 96 h experiment.
Biomarkers
Enzyme activities
Tadpoles (n = 7 per treatment) were weighed (g), and homogenized (1:10, w/v) in ice-cold 25 mM sucrose, 20 mMTris-HCl buffer (pH ¼ 7.4) containing 1 mM EDTA, using a polytron tissue grinder. The homogenates were centrifuged at 10,000 g for 15 min at 4 ± 1 °C, and stored at − 80 °C until measurements. Enzyme activities were quantified after considering total protein concentration by the Biuret method according to Kingsley [70]. AChE activity was determined colorimetrically following Ellman et al. [71] and expressed as nmol min−1 mg−1protein (MEC = 13.6 × 103 M−1 cm−1). The reaction mixture consisted on 25 mM Tris–HCl and contained 1 mM CaCl2 (pH = 7.6), 10 mL 20 μM acetylthiocholineiodide, and 50 μL 5,5-dithio-bis-(2-nitrobenzoic acid) (3 × 104 M, final concentration). The variation in optical density was measured in duplicate at 410 nm at 25 °C for 1 min, using a Jenway 6405 UV–vis spectrophotometer. GST activity was measured spectrophotometrically according to Habig et al. [72] with modifications of Habdous et al. [73] for mammal serum GST activity. The measurement was performed with 340 nm in 100 mM Na–phosphate buffer (pH 6.5), 20 μL of 0.2 mM 1-chloro-2,4-dinitrobenzene, 50 μL of 5 mM reduced glutathione, and the sample homogenates. GST activity was expressed as nmol min−1 mg−1protein (molar extinction coefficient, MEC = 9.61 × 103 M−1 cm−1).
Swimming activity
The swimming activity of individual tadpoles was recorded for one minute using a digital video camera (Motic®, 10.0 M pixel) that was mounted in a tripod and placed just above the petri dish. The petri dish was filled with 200 ml of different treatment solutions, following procedures previously established in our lab [39, 74]. For each treatment, three tadpoles were evaluated independently.
Total distance moved (cm), mean speed (cm segˉ1), and global activity (cm2) were evaluated on each tadpole. These behavior parameters were quantified and analyzed using video-tracking software Smart (3.0.02 PanLab Harvard Apparatus ®).
Cardiac rhythm
Cardiac rhythm was evaluated following the methodology described for other native amphibian species [32, 39]. The body of tadpoles (n = 5 per treatment) was located in a thin concave plate in ventral side up position. Continual trans-illumination on heart area (front quarter body) was applied bottom-up with a spot-led light (Luxeon Rebel 3 wat Led). Videos were recorded in completely immobile tadpoles with a remote-triggered portable USB Digital Microscope (Video capture resolution: 640 × 480, 30 fps) at lab conditions (constant 24 °C) during 15 s.
The heart rate (HR; beats. min−1) was quantified from slow-speed digital videos by direct visual examination of maximum systole ventricle beating [39, 75]. The number of beats measured in the recorded 15 s was multiplied by four to obtain the value of a complete one min [76, 77].
Data analyzes
The LC50 value and its respective 95% confidence limits (95% CL) were calculated by the trimmed Spearman–Karber method [78]. Post-hoc comparison Dunnett’s test was used to analyze means of mortality of LOEC and NOEC values [79]. The toxicity of this compound was estimated according to the hazard classification system for wastewater discharged into the aquatic environment because the effects of ClO2 were not previously studied for tadpoles [80]. The obtained LC50 value was transformed into Toxic Units (TU) following the criterion of Lajmanovich et al. [81]: TU = 100 / LC50. The values were classified into five classes: I (No acute toxicity) = TU < 0.4; II (Slight acute toxicity) = 0.4 < TU < 1; III (Acute toxicity): 1 < TU < 10; IV (High acute toxicity): 10 < TU < 100; V (very high acute toxicity): TU > 100.
The data of biomarkers were expressed as means ± standard deviation (SD). Kolmogorov–Smirnov test and Levene test were used to confirm normality and homogeneity of variances, respectively. Differences in enzymatic activity of CO and ClO2 treated tadpoles were analyzed by one-way ANOVA and Dunnett’s test for post-hoc comparisons. GraphPad InStad® software was used for statistical analyzes. In addition, Spearman correlations were used to evaluate correlation between all pairs of variables (biochemical, behavioral, and cardiac endpoints, shown in table S2 from Supplementary material) for each treatment at 24 and after 96 h exposure. Values were considered significant at p < 0.05. No significant differences were found among replicates (p > 0.05), so, replicates from each treatment were pooled.
Conclusion
The present study determined the toxicity of ClO2 in amphibian tadpoles for first time worldwide. These data would be included in the maximum concentration criterion of chlorine compound toxicity assessment that has only been determined for freshwater fish and invertebrates. Our findings indicate a high risk of ClO2 exposure to anuran tadpoles that present alterations of AChE and GST enzymes, swimming behavior, and cardiac rhythm. The relevance of our study resides in the fact that this compound that extensively and continuously input to water become a threat for aquatic organisms and aquatic ecosystems functions and services, many of which are essential to human well—being.
Availability of data and material
Data presented in this study are available on request from the corresponding author.
References
Vertova A, Miani A, Lesma G, Rondinini S, Minguzzi A, Falciola L, Ortenzi MA (2019) Chlorine dioxide degradation issues on metal and plastic water pipes tested in parallel in a semi-closed system. Int J Environ Res Public Health 16(22):4582. https://doi.org/10.3390/ijerph16224582
Gordon G, Rosenblatt AA (2005) Chlorine dioxide: the current state of the art. Ozone Sci Eng 27(3):203–207
Chatuev BM, Peterson JW (2010) Analysis of the sporicidal activity of chlorine dioxide disinfectant against Bacillus anthracis (Sterne strain). J Hosp Infect 74(2):178–183. https://doi.org/10.1016/j.jhin.2009.09.017
Ogata N, Shibata T (2009) Effect of chlorine dioxide gas of extremely low concentration on absenteeism of schoolchildren. Int J Med Med Sci 1(7):288–289. https://doi.org/10.5897/IJMMS.9000077
Kály-Kullai K, Wittmann M, Noszticzius Z, Rosivall L (2020) Can chlorine dioxide prevent the spreading of coronavirus or other viral infections? Med Hypotheses Physiol Int 107(1):1–11. https://doi.org/10.1556/2060.2020.00015
Wang J, Shen D, Ye X, Yan Y, Zhang W et al (2020) Disinfection technology of hospital wastes and wastewater: suggestions for disinfection strategy during coronavirus Disease 2019 (COVID-19) pandemic in China. Environ Pollut 262:114665. https://doi.org/10.1016/j.envpol.2020.114665
Trigo MS, Kurmanaev A, Cabrera JML (2020) Coronavirus en América Latina: algunas autoridades respaldan tratamientos cuestionables. NY Times. Available online at: https://www.nytimes.com/es/2020/07/23/espanol/america-latina/bolivia-cloro-coronavirus-ivermectina.html. Accessed 11 Aug 2021
Bendezu-Quispe G, Rodríguez-Zúñiga MJM, Roman YM, Mori-Llontop LM, Peralta V, Fiestas F (2020) Agentes potencialmente terapéuticos contra el SARS-CoV-2: revisión rápida de la evidencia. Rev Peru Med Exp Salud Pública 37(2):320–326. https://doi.org/10.17843/rpmesp.2020.372.5409
Burela A, Hernández-Vásquez A, Comandé D, Peralta V, Fiestas F (2021) Dióxido de cloro y derivados del cloro para prevenir o tratar la COVID-19: revisión sistemática. Rev Peru Med Exp Salud Publica 37:605–610. https://doi.org/10.17843/rpmesp.2020.374.6330
Mostajo-Radji MA (2021) Pseudoscience in the times of crisis: how and why chlorine dioxide consumption became popular in Latin America during the COVID-19 pandemic. Front Polit Sci 3:25. https://doi.org/10.3389/fpos.2021.621370
García-Ávila F, Valdiviezo-Gonzales L, Cadme-Galabay M, Gutiérrez-Ortega H, Altamirano-Cárdenas L, Zhindón-Arévalo C, Flores del Pino L (2020) Considerations on water quality and the use of chlorine in times of SARS-CoV-2 (COVID-19) pandemic in the community. Case Stud CSCEE 2:100049. https://doi.org/10.1016/j.cscee.2020.100049
Quevedo R, Bastías JM, Espinoza T, Ronceros B, Balic I, Muñoz O (2020) Inactivation of Coronaviruses in food industry: the use of inorganic and organic disinfectants, ozone, and UV radiation. Scientia Agropecuaria 11(2):257–266
Kitajima M, Ahmed W, Bibby K, Carducci A, Gerba CP, Hamilton KA, Haramoto E, Rose JB (2020) SARS-CoV-2 in wastewater: state of the knowledge and research needs. Sci Total Environ 739:139076. https://doi.org/10.1016/j.scitotenv.2020.139076
Environmental Assessment for Food Contact Notification No. 1804. Available at https://www.fda.gov/food/environmental-decisions/environmental-decision-memo-food-contact-notification-no-1804
Toxicological profile for chlorine dioxide and chlorite. Available at: https://www.atsdr.cdc.gov/toxprofiles/tp160.pdf
Dewey HM, Jones JM, Keating MR, Budhathoki-Uprety J (2021) Increased use of disinfectants during the COVID-19 pandemic and its potential impacts on health and safety. ACS Chem Health Saf
Chu W, Fang C, Deng Y, Xu Z (2020) Intensified disinfection amid COVID-19 pandemic poses potential risks to water quality and safety. Environ Sci Technol 55(7):4084–4086
Junli HLW, Nenqi R, Li LX, Fun SR, Guanle Y (1997) Disinfection effect of chlorine dioxide on viruses, algae, and animal planktons in water. Wat Res 31:455–460. https://doi.org/10.1016/S0043-1354(96)00276-X
Wigginton KR, Pecson BM, Sigstam T, Bosshard F, Kohn T (2012) Virus inactivation mechanisms: impact of disinfectants on virus function and structural integrity. Environ Sci Technol 46(21):12069–12078. https://doi.org/10.1021/es3029473
Kingsley DH, Pérez-Pérez RE, Niemira BA, Fan X (2018) Evaluation of gaseous chlorine dioxide for the inactivation of Tulane virus on blueberries. Int J Food Microbiol 273:28–32. https://doi.org/10.1016/j.ijfoodmicro.2018.01.024
Hose JE, Di Fiore D, Parker HS, Sciarrotta T (1989) Toxicity of chlorine dioxide to early life stages of marine organisms. Bull Environ Contam Toxicol 42(3):315–319. https://doi.org/10.1007/BF01699954
Svecevicius G, Syvokiene J, Stasiŭnaite P, Mickeniene L (2005) Acute and chronic toxicity of chlorine dioxide (ClO2) and chlorite (ClO2-) to rainbow trout (Oncorhynchus mykiss). Environ Sci Pollut Res Int 5:302–305. https://doi.org/10.1065/espr2005.04.248
Sun XB, Cui FY, Guo ZH (2007) Toxicity and influencing factors of chlorine dioxide to chironomid larvae. Huan Jing KeXue 11:2569–2573
Kos J, Brmež M, Markić M, Sipos L (2019) The mortality of nematodes in drinking water in the presence of ozone, chlorine dioxide, and chlorine. Ozone Sci Eng 42:120–127
Burton DT, Fisher DJ (2001) Chlorine dioxide: the state of science, regulatory, environmental issues, and case histories. In: Proceedings of the 4th international symposium, Las Vegas, NV
Blaustein AR, Bancroft BA (2007) Amphibian population declines: evolutionary considerations. Bioscience 57(5):437–444. https://doi.org/10.1641/B570517
Hayes TB, Falso P, Gallipeau S, Stice M (2010) The cause of global amphibian declines: a developmental endocrinologist's perspective. J Exp Biol Mar 213(6):921–933. https://doi.org/10.1242/jeb.040865
Blaustein AR, Han BA, Relyea RA, Johnson PT, Buck JC, Gervasi SS, Kats LB (2011) The complexity of amphibian population declines: understanding the role of cofactors in driving amphibian losses. Ann N Y Acad Sci 1223:108–119. https://doi.org/10.1111/j.1749-6632.2010.05909.x
Egea-Serrano A, Relyea RA, Tejedo M, Torralva M (2012) Understanding of the impact of chemicals on amphibians: a meta-analytic review. Ecol evol 2(7):1382–1397. https://doi.org/10.1002/ece3.249
Connon RE, Geist J, Werner I (2012) Effect-based tools for monitoring and predicting the ecotoxicological effects of chemicals in the aquatic environment. Sensors 12(9):12741–12771. https://doi.org/10.3390/s120912741
Brodeur JC, Poliserpi MB, D’Andrea MF, Sánchez M (2014) Synergy between glyphosate- and cypermethrin-based pesticides during acute exposures in tadpoles of the common South American Toad Rhinella arenarum. Chemosphere 112:70–76. https://doi.org/10.1016/j.chemosphere.2014.02.065
Lajmanovich RC, Peltzer PM, Martinuzzi CS, Attademo AM, Bassó A, Colussi CL (2019) Insecticide pyriproxyfen (Dragón®) damage biotransformation, thyroid hormones, heart rate, and swimming performance of Odontophrynusamericanus tadpoles. Chemosphere 220:714–722. https://doi.org/10.1016/j.chemosphere.2018.12.181
Bahl MF, Brodeur JC, Costa CS, D’Andrea MF, Sansiñena JA, Marino DJ, Natale GS (2021) Lethal and sublethal effects of the natural and healthy spinosad-based formulation TracerTM on tadpoles of two neotropical species. Environ Sci Pollut Res 28(11):13524–13535. https://doi.org/10.1007/s11356-020-09808-8
Peltzer PM, Junges CM, Attademo AM, Bassó A, Grenón P, Lajmanovich RC (2013) Cholinesterase activities and behavioral changes in Hypsiboas pulchellus (Anura: Hylidae) tadpoles exposed to glufosinate ammonium herbicide. Ecotoxicology 22(7):1165–1173. https://doi.org/10.1007/s10646-013-1103-8
Nunes B (2011) The use of cholinesterases in ecotoxicology. In: Whitacre DM (ed) Reviews of environmental contamination and toxicology. Springer, New York, pp 29–60
Walker CH, Hopkin SP, Sibly RM, Peakall DB (2001) Principles of ecotoxicology, 2nd edn. Taylor and Francis, New York
Denoël M, D’Hooghe B, Ficetola GF, Brasseur C, De Pauw E, Thomé JP, Kestemont P (2012) Using sets of behavioral biomarkers to assess short-term effects of pesticide: a study case with endosulfan on frog tadpoles. Ecotoxicology 21:1240–1250
Prokić MD, Borković-Mitić SS, Krizmanić II, Mutić JJ, Gavrić JP, Despotović SG, Saičić ZS (2017) Oxidative stress parameters in two Pelophylax esculentus complex frogs during pre-and post-hibernation: arousal vs heavy metals. Comp Biochem Physiol Part C Toxicol Pharmacol 202:19–25
Peltzer PM, Lajmanovich RC, Martinuzzi C, Attademo AM, Curi LM, Sandoval MT (2019) Biotoxicity of diclofenac on two larval amphibians: Assessment of development, growth, cardiac function and rhythm, behavior and antioxidant system. Sci Total Environ 683:624–637. https://doi.org/10.1016/j.scitotenv.2019.05.275
Fernández LP, Brasca R, Attademo AM, Peltzer PM, Lajmanovich RC, Culzoni MJ (2020) Bioaccumulation and glutathione S-transferase activity on Rhinellaarenarum tadpoles after short-term exposure to antiretrovirals. Chemosphere 246:125830. https://doi.org/10.1016/j.chemosphere.2020.125830
Costa MJ, Monteiro DA, Oliveira-Neto AL, Rantin FT, Kalinin AL (2008) Oxidative stress biomarkers and heart function in bullfrog tadpoles exposed to Roundup Original. Ecotoxicology 17(3):153–163. https://doi.org/10.1007/s10646-007-0178-5
Perrichon P, Pasparakis C, Mager EM, Stieglitz JD, Benetti DD, Grosell M, Burggren WW (2017) Morphology and cardiac physiology are differentially affected by temperature in developing larvae of the marine fish mahi-mahi (Coryphaena hippurus). Biology open 6(6):800–809
Denoël M, Libon S, Kestemont P, Brasseur C, Focant JF, De Pauw E (2013) Effects of a sublethal pesticide exposure on locomotor behavior: a video-tracking analysis inlarval amphibians. Chemosphere 90:945e951
Couri D, Abdel-Rahman MS, Bull RJ (1982) Toxicological effects of chlorine dioxide, chlorite and chlorate. Envir Health Persp 46:13
Yonkos LT, Fisher DJ, Wright DA, Kane AS (2000) Pathology of fathead minnows (Pimephales promelas) exposed to chlorine dioxide and chlorite. Mar Environ Res 50(1–5):267–271. https://doi.org/10.1016/S0141-1136(00)00048-9
López-Galindo C, Vargas-Chacoff L, Nebot E, Casanueva JF, Rubio D, Solé M, Mancera JM (2010) Biomarker responses in Soleasenegalensis exposed to sodium hypochlorite used as antifouling. Chemosphere 78(7):885–893. https://doi.org/10.1016/j.chemosphere.2009.11.022
Kim M, Kwon H, Kim W, Kim Y (2016) Enhanced Acetylcholinesterase activity of the Indianmeal Moth, Plodia interpunctella, under chlorine dioxide treatment and altered negative phototaxis behavior. Kor J Appl Entomol 55(1):27–33
Matsushita T, Fujita Y, Omori K, Huang Y, Matsui Y, Shirasaki N (2020) Effect of chlorination on anti-acetylcholinesterase activity of organophosphorus insecticide solutions and contributions of the parent insecticides and their oxons to the activity. Chemosphere 261:127743
Carvalho CS, Utsunomiya HS, Pasquoto T, Costa MJ, Fernandes MN (2017) Cholinesterase activity as potential biomarkers: Characterization in bullfrog tadpole’s brain after exposure to metals. In: XVII safety, health and environment world congress, Vila Real, Portugal, pp 86–88
Barillet S, Adam-Guillermin C, Palluel O, Porcher JM, Devaux A (2011) Uranium bioaccumulation and biological disorders induced in zebrafish (Danio rerio) after a depleted uranium waterborne exposure. Environ Pollut 159(2):495–502
Pozzetti L, Broccoli M, Potenza G, Canestro D, Affatato A, Sapone A, Antelli A, D’Amico E, Vangelisti S, Cantelli-Forti G, Paolini M, Ferrara G, Elia AC, Do¨rr AJM, Taticchi MI, Mantilacci L, Natali M, (2003) Tecnicheenzimatiche per valutare la tossicita` di acquepotabilizzate. Acqua Aria 4:64–68
Elia AC, Anastasi V, Dörr AJM (2006) Hepatic antioxidant enzymes and total glutathione of Cyprinus carpio exposed to three disinfectants, chlorine dioxide, sodium hypochlorite and peracetic acid, for superficial water potabilization. Chemosphere 64(10):1633–1641
Ison A, Odeh IN, Margerum DW (2006) Kinetics and mechanisms of chlorine dioxide and chlorite oxidations of cysteine and glutathione. Inorg Chem 45(21):8768–8775
Bonansea RI, Wunderlin DA, Amé MV (2016) Behavioral swimming effects and acetylcholinesterase activity changes in Jenynsia multidentata exposed to chlorpyrifos and cypermethrin individually and in mixtures. Ecotox Environ Safe 129:311–319
Ren Q, Zhao R, Wang C, Li S, Zhang T, Ren Z, Wang X (2017) The role of AChE in swimming behavior of Daphnia magna: correlation analysis of both parameters affected by deltamethrin and methomyl exposure. J Toxicol. https://doi.org/10.1155/2017/3265727
Ballesteros ML, Durando PE, Nores ML, Díaz MP, Bistoni MA, Wunderlin DA (2009) Endosulfan induces changes in spontaneous swimming activity and acetylcholinesterase activity of Jenynsia multidentata (Anablepidae, Cyprinodontiformes). Environ Poll 157(5):1573–1580
Pan H, Zhang X, Ren B, Yang H, Ren Z, Wang W (2017) Toxic assessment of cadmium based on online swimming behavior and the continuous AChE activity in the gill of zebrafish (Danio rerio). Water Air Soil Pollut 228(9):1–9
Kaufer D, Friedman A, Seidman S, Soreq H (1998) Acute stress facilitates long-lasting changes in cholinergic gene expression. Nature 393(6683):373–377
Choi E, Alsop D, Wilson JY (2018) The effects of chronic acetaminophen exposure on the kidney, gill and liver in rainbow trout (Oncorhynchus mykiss). Aquat Toxicol 198:20–29
Lajmanovich RC, Izaguirre MF, Casco VH (1998) Paraquat tolerance and alteration of internal gill structure of Scinax nasica tadpoles (Anura: Hylidae). Arch Environ Contam Toxicol 34:364–369
Orme J, Taylor DH, Laurie RD, Bull RJ (1985) Effects of chlorine dioxide on thyroid function in neonatal rats. J Toxicol Environ Health A Curr Issues 15(2):315–322
Dal-Medico SE, Rissoli RZ, Gamero FU, Victório JA, Salla RF, Abdalla FC, Costa MJ (2014) Negative impact of a cadmium concentration considered environmentally safe in Brazil on the cardiac performance of bullfrog tadpoles. Ecotoxicol Environ Safe 104:168–174
Revis NW, McCauley P, Bull R, Holdsworth G (1986) Relationship of drinking water disinfectants to plasma cholesterol and thyroid hormone levels in experimental studies. PNAS 83(5):1485–1489
Hey G, Ledin A, Jansen JLC, Andersen HR (2012) Removal of pharmaceuticals in biologically treated wastewater by chlorine dioxide or peracetic acid. Environ Technol 33:1041–1047. https://doi.org/10.1080/09593330.2011.606282
Chhetri RK, Baun A, Andersen HR (2019) Acute toxicity and risk evaluation of the CSO disinfectants performic acid, peracetic acid, chlorine dioxide and their by-products hydrogen peroxide and chlorite. Sci Total Environ 677:1–8. https://doi.org/10.1016/j.scitotenv.2019.04.350
Lee YB, Kim WS (2020) ClO2 Dipping treatment inhibits gray mold on cut rose flowers during storage. Hort J UTD 138
Han GD, Na J, Chun YS, Kumar S, Kim W, Kim Y (2017) Chlorine dioxide enhances lipid peroxidation through inhibiting calcium-independent cellular PLA2 in larvae of the Indianmeal moth, Plodia interpunctella. Pestic Biochem Physiol 143:48–56
Lajmanovich RC, Attademo AM, Lener G, Cuzziol Boccioni AP, Peltzer PM, Martinuzzi CS, Repetti MR (2021) Glyphosate and glufosinate ammonium, herbicides commonly used on genetically modified crops, and their interaction with microplastics: Ecotoxicity in anuran tadpoles. STOTEN 150177
ASIH (2004) Guidelines for use of live amphibians and reptiles in field and laboratory research. Lawrence, Kansas: Herpetological Animal Care and Use Committee (HACC) of the American Society of Ichthyologists and Herpetologists
Kingsley GR (1942) The direct biuret method for the determination of serum proteins as applied to photoelectric and visual colorimetry. J Lab Clin Med 27:840–845
Ellman GL, Courtney KD, JrV A, Featherstone RM (1961) A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem Pharmacol 7(2):88–95
Habig WH, Pabst MJ, Jakoby WB (1974) Glutathione Stransferase: the first step in mercapturic acid formation. J Biol Chem 249:7130–7139
Habdous M, Vincent-Viry M, Visvikis S, Siest G (2002) Rapid spectrophotometric method for serum glutathione S-transferases activity. Clin Chim Acta 326(1–2):131–142
Junges CM, Maglianese MI, Lajmanovich RC, Peltzer PM, Attademo AM (2017) Acute toxicity and etho-toxicity of three insecticides used for mosquito control on amphibian tadpoles. Water Air Soil Pollut 228(4):143
Chan PK, Lin CC, Cheng SH (2009) Non invasive technique for measurement of heartbeat regularity in zebrafish (Danio rerio) embryos. BMC Biotechnol 9(1):11. https://doi.org/10.1186/1472-6750-9-11
Kang R, Jeong JS, Yoo JC, Lee JH, Choi SJ, Gwak MS, Ko JS (2018) Effective dose of intravenous dexmedetomidine to prolong the analgesic duration of interscalene brachial plexus block: a single-center, prospective, double-blind, randomized controlled trial. Reg Anesth Pain Med 43(5):488–495
Hoage T, Ding Y, Xu X (2012) Quantifying cardiac functions in embryonic and adult zebrafish. In: Cardiovascular development, Humana Press, Totowa, New Jersey, pp 11–20
Hamilton MA, Russo RC, Thurston RV (1977) Trimmed spearman-karber method for estimating median lethal concentrations in toxicity bioassays. Environ Sci Technol 11:714–719
US EPA (U.S. Environmental Protection Agency). Short-term methods for estimating the chronic toxicity of effluents and receiving waters to fresh water organisms; Report No. EPA/600/ 4–89/001. Environmental Protection Agency: Cincinnati, OH, 1989
Persoone G, Marsalek B, Blinova I, Torokne A, Zarina D, Manusadzianas L, Nalecz-Jawecki G, Tofan L, Stepanova N, Tothova L, Kolar B (2003) A practical and user-friendly toxicity classification system with microbiotests for natural waters and wastewaters. Environ Toxicol 18(6):395–402. https://doi.org/10.1002/tox.10141
Lajmanovich RC, Peltzer PM, Martinuzzi CS, Attademo AM, Colussi CL, Basso A (2018) Acute toxicity of colloidal silicon dioxide nanoparticles on amphibian larvae: emerging environmental concern. Int J Environ Sci 12(3):269–278
Acknowledgements
We acknowledge Boccioni L.S. for supplying the chemical compounds for test solutions preparation and Lajmanovich R.F. for providing information regarding to the erroneous human ClO2 use as “miracle mineral solution”. We also thanks to ANPCyT FONCyT and UNL for providying subsidies (PICT 2017 N 1069-RCL; CAI+D 2020-PMP). Finally, we acknowledge reviewers for their comments and suggestions.
Funding
This study was supported in part by National Agency for Promotion of Science and Technology, Argentine (ANPCyT FONCyT PICT, No. 1069), and Course of Action for Research and Science Promotion, Argentine (CAI D-UNL, PIC No. 100004 LI).
Author information
Authors and Affiliations
Contributions
Conceptualization: RL and PP; Methodology: PP, ACB, CM, AA and CC; Formal analysis and investigation: PP, ACB; Writing—original draft preparation and Writing—review and editing: PP, ACB and RL.
Corresponding authors
Ethics declarations
Conflict of interest
Paola M. Peltzer, Ana P. Cuzziol Boccioni, Andrés M. Attademo, Candela S. Martinuzzi, Carlina L. Colussi, and Rafael C. Lajmanovich declare that we have no conflict of interest.
Ethics approval
Animals were treated according the Institutional Committee for the Care and Use of Animals (IACUC), and approval was obtained from the bioethics committee of the FBCB-UNL (Res. No. 388/06).
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Peltzer, P.M., Cuzziol Boccioni, A.P., Attademo, A.M. et al. Risk of chlorine dioxide as emerging contaminant during SARS-CoV-2 pandemic: enzyme, cardiac, and behavior effects on amphibian tadpoles. Toxicol. Environ. Health Sci. 14, 47–57 (2022). https://doi.org/10.1007/s13530-021-00116-3
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s13530-021-00116-3