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Bioassay-directed identification of toxicants in sediments of Liaohe River, northeast China
Environmental Pollution xxx (2016) 1e9
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Bioassay-directed identification of toxicants in sediments of Liaohe River, northeast China* Yan He a, b, Jian Xu a, *, Changsheng Guo a, Jiapei Lv a, Yuan Zhang a, Wei Meng a, b a b
State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012, China College of Water Sciences, Beijing Normal University, Beijing 100012, China
a r t i c l e i n f o
a b s t r a c t
Article history: Received 24 February 2016 Received in revised form 31 May 2016 Accepted 22 June 2016 Available online xxx
Contaminants accumulated in sediments may directly harm benthic organisms, however, the specific contaminants responsible for adverse effects have been poorly described. In this study, a bioassaydirected analysis combined with toxicity tests and chemistry analysis was conducted to determine the compounds eliciting the greatest toxicological effect in the sediments in Liaohe River, northeast China. A total of 24 sediment samples were examined to determine their acute toxicity to midge Chironomus tentans (C. tentans). Of these samples, 15 exhibited significant toxicity, with a mortality of 23%e93% (p < 0.05). Numerous contaminants, including 16 polycyclic aromatic hydrocarbons, 32 polychlorinated biphenyls, 20 organochlorine pesticides, 6 organophosphate pesticides, 8 pyrethroids, and 5 heavy metals were analyzed. On the basis of toxic unit (TU) analysis results, pyrethroids may contribute to the toxicity of 9 of the 15 toxic samples with concentrations of >1 TU. The significant correlation between the TUs of pyrethroids and the mortality of C. tentans (r2 ¼ 0.74, p < 0.01) confirmed the major role of pyrethroids in toxicity. The selected sediment samples responding to piperonyl butoxide and low temperature with the increased toxicity exhibited the characteristics of pyrethroids. The bioassay-based screening framework provided strong evidence that pyrethroids were the primary cause of sediment toxicity in Liaohe River. Further studies should therefore be conducted to regulate this important class of pollutants. © 2016 Elsevier Ltd. All rights reserved.
Keywords: Liaohe river Bioassay-directed analysis Pyrethroids Toxicity Chironomus tentans
1. Introduction Over the past several decades, rapid economic growth and urbanization have resulted in the deterioration of water quality in aquatic ecosystems in China. As such, the central and local governments of China have focused on water quality improvements; however, authorities have placed minor considerations on aquatic sediments in terms of practical water quality management. Sediments are important components of aquatic ecosystems providing habitat and feeding and spawning areas for aquatic organisms; sediments are also major repositories of chemical pollutants. High contaminant concentrations that accumulated in sediments are harmful to benthic organisms. Moreover, contaminated sediments adversely affect aquatic resources by causing ecological and economic damages, such as habitat degradation and costly
* This paper has been recommended for acceptance by Eddy Y. Zeng. * Corresponding author. E-mail address: [email protected] (J. Xu).
remediation and disposal actions (Ho and Burgess, 2013). Therefore, sediment contamination should be characterized to promote sediment assessment and management during decision making. Liaohe River is a heavily polluted river in northeast China. It consists of four big rivers, namely, Liao River, Taizi River, Hun River, and Daliao River. The middle- and down-stream of Liaohe River are the largest industrial bases of metallurgy, machinery, petrochemical, and building materials in northeast China. The pollution status of contaminants, including polycyclic aromatic hydrocarbons (Guo et al., 2007), organochlorine pesticides (Wang et al., 2007), polychlorinated biphenyls, polybrominated diphenyl ethers, polychlorinated dibenzo-p-dioxins (Lv et al., 2015; Zhang et al., 2010), short-chain chlorinated paraffins (Gao et al., 2012), and metals (He et al., 2015), in the sediments of Liaohe River have been extensively investigated. These studies have offered a good understanding of the occurrence and distribution of contaminants in this area but have provided limited information on their potential hazards to aquatic organisms. Hence, sediment-contact tests, which involve the exposure of benthic organisms to bulk sediments, are performed to assess the integrated effects of sediment-bound
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contaminants (Chapman and Anderson, 2005). When a positive toxic result is observed in a sediment, the sources of toxicity should be identified to determine the specific contaminants that need immediate attention and thus protect aquatic organisms and ecosystems (Anderson et al., 2010; Donnachie et al., 2014). Ke et al. (2015) examined sediments from Liao River to evaluate acute toxicity in midge larvae (Chironomus riparius) and found that As and Cd are probably associated with sediment toxicity. However, the identified toxicants have not been effectively confirmed, and the link between contaminant exposures and sediment toxicity remains unclear. To the best of our knowledge, limited data are available regarding the identification of active toxicants and establishment of cause-effect relationship. Specifically acting toxicants are difficult to identify because of the complexity of sediment matrix. Limited time and resources have also prompted researchers to devote appropriate mitigation efforts on highly toxic chemicals. The primary objective of this study was to examine the spatial distribution and magnitude of sediment toxicity to midge C. tentans in Liaohe River. The specific contaminants responsible for the toxicity were further explored. Additionally, the relationship between contamination levels and toxic effects was discussed by integrating bioassays with chemical analysis. This study provided comprehensive insights into ecological hazards posed by sediments in Liaohe River, and helped screen the priority pollutants in this area. This bioassay-based screening framework is of ecological and economic importance to assist reliable mitigation and management measures. 2. Materials and methods 2.1. Sample collection Twenty-four sediment samples (0e10 cm) were collected from Liaohe River with a Van Veen grab sampler in June 2014. Among these samples, nine (L1-L9) were from Liao River, five (H1eH5) from Hun River, six (T1eT6) from Taizi River, and four (D1-D4) from Daliao River (Fig. 1). At each sampling site, three replicate samples were collected, passed through a 2 mm mesh sieve, and then homogenized. Collected sediments were placed into sealed polyethylene bags, and transported on ice to the laboratory where they were stored at 4 C for toxicity testing and at 20 C for chemical analysis, respectively. 2.2. Chemical analysis The pollutants including polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), organophosphate pesticides (OPs), pyrethroids and inorganic heavy metals were selected as representatives of hazardous pollutants, due to their widespread distribution in sediment and potential toxicity to aquatic biota. The target compounds are shown in Table S1 in the supplementary materials. In addition, concentrations of acid-volatile sulfide (AVS) and simultaneously extracted metals (SEMs), total organic carbon (TOC) contents and sediment grain size (Table S2) were also analyzed. The details of chemicals and reagents, chemical analysis procedure and quality assurance and control are described in the supplementary materials. 2.3. Bulk sediment toxicity test C. tentans (Diptera: Chironomidae), a native species in Liaohe River, was used as the test organism. The midges obtained from Ecotoxicology Laboratory of Nankai University were laboratorycultured in accordance with standard protocols (USEPA, 2000).
Sediment samples were screened for toxicity by using 10-day survival tests for C. tentans. In brief, 50 g of sediments was placed in three replicate beakers, and each beaker was filled with 200 mL of reconstituted water prepared by adding salts to Milli-Q purified water (USEPA, 2007). After sediments were settled overnight, 10 s- or third-instar larvae were randomly introduced to each beaker and fed with 6 mg of finely ground Tetrafin Goldfish Flakes per day. Tests were performed at 23 C with a 16L: 8D photoperiod, and the overlying water was renewed twice daily. Temperature and dissolved oxygen were monitored on a daily basis; conductivity, pH, and ammonia were measured at the beginning and the end of the test. The samples containing dissolved oxygen with amounts decreased to 2.5 mg/ L were gently aerated. Each test batch was accompanied by control sediments obtained from a drinking water reservoir. After 10 days, surviving larvae were sieved and counted; their ash-free dry mass (AFDM) was determined following the standard methods (USEPA, 2000). 2.4. Estimation of toxicity The contribution of each contaminant to sediment toxicity was evaluated using a toxicity unit approach. The concentrations of contaminants in sediments were converted to TUs by dividing the 10-day sediment median lethal concentration (LC50) for midges on an organic carbon-normalized basis [Eq. (1)]. Conservative sediment benchmarks were used for metals, PAHs, and PCBs because of a lack of sediment toxicity data.
TU ¼
Contaminant concentration in sediment=TOC LC50 or sediment benchmark=TOC
(1)
The TUs of metals were calculated on the basis of consensusbased probable effect concentrations (PECs). AVS-SEM models were also used to evaluate the toxicity contributed by metals (Table S3). The TUs of PAHs were estimated in terms of equilibriumpartitioning sediment benchmarks (SESBTUs; Table S4). For PCBs, the PEC of total PCBs was considered (Table S5). The sediment LC50 of pesticides was obtained from previous studies and listed in Tables S6eS8. For the TUs of hexachlorocyclohexane (HCH), the concentration of gamma isomer was used because other isomers exhibit lower aquatic toxicity (Eaton and Klaassen, 2001). LC50 of allethrin was unavailable; as such, a screening-level toxicity value was computed by using an equilibrium partitioning approach based on a water-column 24 h LC50 value (ECOTOX database) (Moran et al., 2011). The TUs of heavy metals were expressed as the average of five metals. For each class of organics, the TUs of individual contaminant were added to assess mixture toxicity. Moreover, the combined toxicity of pesticides was estimated based on concentration addition (Belden et al., 2007). The approximation of concentration was 0.5 TU when mortality likely occurred; this value was also considered a threshold, that is, at >0.5 TU, a contaminant was assumed potentially responsible for mortality (Weston et al., 2004). 2.5. Confirmation Several lines of evidence were utilized to confirm possible toxicants. One approach was statistical correlation; in this approach, the estimated TUs of contaminants were plotted against the observed mortality. TU evaluation and statistical analysis suggested that pyrethroids may be the primary cause of toxicity; therefore, a focused toxicity identification evaluation (TIE), including piperonyl butoxide (PBO) addition and temperature manipulation, was applied to confirm their toxicity. PBO enhances
Please cite this article in press as: He, Y., et al., Bioassay-directed identification of toxicants in sediments of Liaohe River, northeast China, Environmental Pollution (2016), http://dx.doi.org/10.1016/j.envpol.2016.06.052
Y. He et al. / Environmental Pollution xxx (2016) 1e9
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Fig. 1. Sampling sites in Liaohe River, northeast China.
the toxicity of pyrethroids by inhibiting their detoxification (Amweg et al., 2006a; Mehler et al., 2011). Toxicity tests were conducted with and without PBO (200 mg/L) in the overlying water. Approximately 80% of the water was replaced daily with fresh PBO solution or water; methanol (10 mL/L) as a solvent control was also included. As temperature is decreased, pyrethroid toxicity increases because of the slow pyrethroid metabolism and increased nerve sensitivity (Harwood et al., 2009). The tests were performed at 23 C, and a concurrent test was conducted at 17 C. The larvae cultured at 23 C, were acclimated to 17 C by reducing the temperature by 1 C/h and incubated overnight. Three replicates were tested for each treatment, and larval survival was determined after 10 days.
2.6. Data analysis The normality and homogeneity of data variances were evaluated by conducting Shapiro-Wilk’s and Levene’s tests, respectively. Differences in survival between samples and controls were determined through one-way ANOVA followed by Dunnett’s multiple comparison tests. Percent survival data were arcsine square roottransformed when necessary to satisfy the assumptions of the normality and homogeneity of variance. A sample was considered toxic if a significantly different response from controls was detected and if the difference in organisms’ response was greater than or equal to 20% relative to the controls. Individual TIE treatment results were compared with the unamended ones by performing a
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paired t-test when parametric assumptions were satisfied; otherwise, Wilcoxon rank sum tests were carried out. A probability level of p < 0.05 indicated statistical significance.
and urbanization.
3. Results and discussion
A series of sediment-associated contaminants was analyzed. The concentrations of metals, AVS, and SEM were determined (Table S3). A detailed discussion was presented in our previous study (He et al., 2015). Sixteen PAHs in the priority list were detected in the sediments, and total concentrations ranged from 82.5 ng/g to 25,400 ng/g (Table S4). The levels of PCBs were extremely lower (0.4e41.8 ng/g; Table S5) than those of PAHs. The identified PCB congeners were predominated by triCBs and tetraCBs, and this result is consistent with the production and usage of PCBs in China (Xing et al., 2005). Although most OCPs have been banned for 30 years, they remain detectable in sediments of Liaohe River. The total concentrations of OCPs varied from 1.3 ng/g to 94.8 ng/g, and high residues of HCHs and DDTs were found at most sites (Table S6). The concentrations of six commonly used OPs ranged from 2.8 ng/g to 45.5 ng/g (Table S7). Among the detected OPs, dichlorvos was the most abundant possibly because of its heavy consumption in China (He, 2008). High pyrethroid residues with concentrations of 2.2e102.5 ng/g were also detected in the sediments (Table S8). Among these residues, cypermethrin yielded the highest concentrations because its relatively low price has resulted in its widespread use (Whittle, 2010). Among the three classes of pesticides in Liaohe River, pyrethroids contributed the highest amounts to the total concentration and constituted an average of 56%, followed by OCPs (24%) and OPs (20%) (Fig. S1). OPs and pyrethroids are currently used pesticides as substitutes for OCPs. The concentration of pyrethroid residues in sediments has increased because of the high hydrophobicity and long half-life of this contaminant (Feo et al., 2010). OPs accumulate to a less extent than pyrethroids do because of their relatively high water solubility and fast degradation rates. OCPs levels were also slightly high due to their extensively historical application in Liaoning Province and prolonged persistence.
3.1. Sediment toxicity The sediments from Liaohe River were examined for their acute toxicity to midge C. tentans. Throughout the test, the water quality parameters were within acceptable ranges: dissolved oxygen, >80% saturation; temperature, 21 Ce24 C; pH, 6.9e8.1; conductivity, 378e586 ms/cm; and ammonia, 0.1e1.5 mg/L. The control survivals in all of the tests were 90%e95%, with an average AFDM of >0.48 mg (USEPA, 2000). Of the 24 sediment samples, 15 were significantly toxic to C. tentans. The highest toxicity, with 93% mortality, was observed at site H3 in Shenyang City. In Fig. 2, the levels of sediment toxicity within the entire watershed spatially varied. The sediments from the upstream of Liao River caused low mortality (7%e30%). At the downstream sites, the sediment toxicity increased and the sediments at site L9 in Panjin city resulted in a high mortality of 54%. The sediments from three other rivers flowing through many large- or medium-sized industrial cities exhibited a relatively higher toxicity than those from Liao River. In Hun River, four sediments were particularly toxic, with mortalities ranging from 30% to 93%, except the sediments at site H1 in Dahuofang Reservoir (10% mortality). Likewise, four sediments from Taizi River, where some industrial factories are situated, were significantly toxic with 27%e73% mortality. Among these sediments, those at site T2 were highly toxic because of large amounts of industrial and domestic wastewater discharged from Benxi City. The sediments collected from Daliao River elicited significant toxicity, with a mean mortality of 30%. Site D3 is in Yingkou City, an important coastal industrial base and traffic hub, where notable larval death was observed (40% mortality). Overall, the majority of toxic sites were located in large cities. This finding suggested that sediment toxicity may be closely associated with industrialization
3.2. Sediment chemistry
Fig. 2. The percent survival of C. tentans exposed to sediments collected from Liaohe River. Aasterisks (*) denote a significant difference from the controls (p < 0.05) and the difference in organisms’ response >20% relative to the controls.
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Y. He et al. / Environmental Pollution xxx (2016) 1e9 Table 1 The toxic units (TUs) of heavy metals, polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), organophosphate pesticides (OPs) and pyrethroids in sediments of Liaohe River.
L1 L2 L3 L4 L5 L6 L7 L8 L9 H1 H2 H3 H4 H5 T1 T2 T3 T4 T5 T6 D1 D2 D3 D4 mean
Metals
SPAH
SPCB
SOCP
SOP
Spyrethroid
0.09 0.08 0.09 0.08 0.13 0.15 0.08 0.08 0.19 0.15 0.39 0.33 0.22 0.18 0.19 0.20 0.18 0.23 0.13 0.13 0.08 0.21 0.17 0.13 0.16
0.12 0.05 0.07 0.04 0.08 0.11 0.03 0.08 0.17 0.04 0.32 0.21 0.91 0.12 0.10 0.71 0.50 0.46 0.11 0.44 0.35 0.20 0.13 0.38 0.24
<0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 0.06 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01
0.24 0.02 0.14 0.05 0.24 0.07 0.02 0.11 0.15 0.06 0.30 0.16 0.07 0.42 0.08 0.28 0.10 0.24 0.12 0.42 0.08 0.03 0.03 0.09 0.15
0.02 0.06 <0.01 0.05 0.07 0.02 0.03 0.21 0.02 <0.01 0.03 0.02 <0.01 <0.01 0.01 <0.01 <0.01 0.01 0.02 0.01 0.02 0.13 0.15 0.01 0.05
0.30 0.66 1.65 0.55 0.55 0.69 0.42 1.65 3.00 0.64 0.36 0.22 0.26 2.77 1.24 0.63 0.46 1.34 0.82 1.79 1.27 1.21 1.94 1.33 1.07
3.3. Causes of sediment toxicity A TU approach was employed in our study to assess the contribution of contaminants to the observed toxicity and to determine the causes of toxicity. The estimated TUs of each group of contaminants are listed in Table 1. The average TUs of the five metals ranged from 0.08 to 0.39, with an average of 0.16; this result indicated their minimal contribution to toxicity. The AVS-SEM models further confirmed that the potential toxicity of metals to aquatic life is unexpected at most sites (He et al., 2015). On the basis
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of ESBs for PAHs mixtures, their predicted TUs were between 0.03 and 0.91, with an average of 0.24. Three sediments (H4, T2 and T3) contained more than 0.5 TU; this result suggested that PAHs slightly contributed to the observed mortality, but their risk should be considered at the three sites. Compared with PAHs, PCBs were present at very low concentrations; as such, PCBs could not be accounted for the toxicity because their TUs were <0.01. Among the target OCPs, g-HCHs, endosulfan, and endrin yielded TUs of >0.01, but their combined contribution was <0.5 TU at all of the sites. Therefore, OCPs unlikely influenced the mortality of C. tentans. Chlorpyrifos was the only OP with a TU of >0.01. Indeed, OPs were unlikely the cause of acute toxicity because of their low TUs (0.01e0.21). The highest values were observed at most sites when pyrethroid concentrations were converted to TUs. Of the 24 sediments, 11 yielded 1 TU and 10 caused a significantly reduced survival of C. tentans. Conversely, the sediments at site T6 elicited low toxicity with 23.3% mortality at 1.79 TU. The overestimation of toxicity may be attributed to the reduced bioavailability of pyrethroids in sediments because of grain size or organic carbon type (You et al., 2008; Li et al., 2013b). This sediment contained high percent of fine grains (silt and clay fractions, 90%). Pyrethroids are strongly adsorbed onto fine particles; as such, they are less available to C. tentans (Laskowski, 2002). Low mortality was observed in the 13 remaining sediments containing <0.5 TU except at the three sites (H2, H3, and T2), where a very high mortality may be linked to the unanalyzed contaminants. Overall, 9 of the 15 toxic samples (out of the 24 samples) yielded at least 1 TU. This finding suggested that pyrethroids mostly accounted for the observed toxicity. When TUs were apportioned among individual pyrethroids, cypermethrin apparently contributed the greatest amount to pyrethroid TUs and yielded an average of 77% (Fig. 3). Of the 11 toxic samples, cypermethrin TUs exceeded the threshold of 0.5. This result indicated the major role of cypermethrin in toxicity. Deltamethrin was the second-largest contributor, with approximately 13% of TUs. For the other pyrethroids, their total contributions to toxicity were minimal and accounted for 10% of the overall TUs.
Fig. 3. Comparison of mean pyrethroid TUs in sediments of Liaohe River, and the contribution of the various pyrethroids to total TUs.
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A concentration yielding 1 TU should cause 50% mortality, but mortality of <50% was noted in this study. This phenomenon usually occurs; previous studies also revealed that sediments with 1e3 pyrethroid TUs cause mortality that is lower than the predicted value (Weston et al., 2005; Amweg et al., 2006b; Hintzen et al., 2009). This discrepancy is possibly attributed to the variability of LC50 of pyrethroids among different types of sediments (Amweg et al., 2005; Maund et al., 2002). Furthermore, a TU approach based on concentration addition may overestimate mixture toxicity, but differences between observed and predicted effect concentrations do not exceed a factor of 3 (Faust et al., 2003). Therefore, TU analysis suggested that pyrethroids are probably the major cause of sediment toxicity in Liaohe River.
3.4. Confirmation of pyrethroid-related toxicity Three lines of evidence were applied to determine whether pyrethroids are responsible for the toxicity of C. tentans. One approach was statistical correlation; in this approach, the sum or individual pyrethroid TUs were regressed against the observed mortality after the three outliers were eliminated. For the three sediments (H2, H3, and T2), non-pyrethroids apparently contributed to the toxicity. In Fig. 4, total pyrethroid TUs were significantly correlated with C. tentans toxicity (Spearman’s rank correlation, r2 ¼ 0.74, p < 0.01). This finding confirmed that pyrethroids were primarily responsible for mortality. In terms of individual pyrethroid, cypermethrin was the main contributor to toxicity, as
Fig. 4. Relationship between pyrethroid TUs and observed toxicity in sediments of Liaohe River (Spearman’s rank correlation).
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indicated by a highly significant correlation (r2 ¼ 0.71, p < 0.01). TIEs, including PBO addition and temperature manipulation, were effective tools to characterize pyrethroid-related toxicity. In this study, 11 of 24 sediments were evaluated on the basis of the focused TIEs. The control sediments with PBO in the overlying water exhibited 80%e90% survival; this result indicated that PBO did not induce toxicity. In Fig. 5, toxicity was increased in all of the samples by the added PBO, and 9 of the samples exhibited notable differences (p < 0.05). At sites L7 and H1, the added PBO slightly increased sediment toxicity possibly because of the relatively low pyrethroid contents. By contrast, the added PBO significantly increased the toxicity at site L1 (p < 0.05), although the pyrethroid concentrations were low (TU ¼ 0.30). This phenomenon probably
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occurred because other contaminants, such as PAHs, in sediments may be synergized by PBO and thus contributed to the toxicity (Weinstein and Garner, 2008). Sediment toxicity increased at all of the sites except site L7 when the test temperature was reduced from 23 C to 17 C (Fig. 5). In the sediments at site L7, the TUs of the analyzed pollutants were very low; this finding suggested the presence of some undetermined toxicants, and their toxicities decreased at low temperature. Similar studies have also revealed that low temperature occasionally performs inconsistently and induces a reduction in toxicity (Weston et al., 2009; Weston and Lydy, 2010). Of the 11 samples, only three exhibited a significant increase in toxicity (p < 0.05). The slight temperature response was consistent with that described in a previous study, which revealed
Fig. 5. The effect of the various TIE manipulations on the toxicity of sediments from Liaohe River. Aasterisks (*) indicate significant differences between amended sediments and unamended ones (p < 0.05).
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an increased toxicity in 4 of 13 samples at low temperature (Weston et al., 2013). This result indicated that low temperature treatment slightly affected pyrethroid toxicity. The two TIE response profiles were consistently indicative of pyrethroid toxicity. In summary, the focused TIE manipulations combined with TU estimation and strong correlations provided sufficient evidence that pyrethroids were the main causative toxicant in the majority of sediments from Liaohe River. However, the cause of toxicity in a small number of samples has yet to be determined. Emerging contaminants should be identified in future studies. Similar researches on the causes of sediment toxicity have been previously conducted in California (Holmes et al., 2008), Texas (Hintzen et al., 2009), Illinois (Ding et al., 2010) and Oregon/ Washington (Weston et al., 2011) in the United States (USA) and Pearl River Delta (Mehler et al., 2011) and Guangzhou (Li et al., 2013a) in China. In those studies, pyrethroids played a major role in the observed toxicity. As for the individual pyrethroid, bifenthrin was the main toxic contributor in USA, while in China cypermethrin was the dominant acting toxicant. This difference is due to the applicator preference in pyrethroid usage in the two countries (Wang et al., 2012). In USA, most of the households have private lawns, where bifenthrin is primarily applied for structural pest control and lawn maintenance (Weston et al., 2005). In contrast, cypermethrin is extensively used in agriculture, mosquito control and landscape maintenance in China due to its relatively low price (Li et al., 2011). In the current study, Liaohe River had slightly lower pyrethroid concentrations and less toxicity as compared to Pearl River Delta and Chebei Creek, Guangzhou in south China (Mehler et al., 2011; Li et al., 2013a). This result can be explained by the different land-use types and urbanization level. In south China, highly developed economy has created substantial urbanization areas. Dense populations and municipal landscaping lead to the intensive use of pyrethroids. In comparison, Liaohe River in northeast China has a lot of agricultural land, which is less urbanized than south China. Similar studies have also revealed that total pyrethroid concentrations were significantly related to the percent urban land (Kuivila et al., 2012). Moreover, the climatic conditions may affect the amounts of pyrethroid use. Humid climate in south China promotes the breeding of mosquitoes, resulting in increased pyrethroid consumption (Li et al., 2011). 4. Conclusions This study fully demonstrated the acute toxic effects of contaminants in the sediments of Liaohe River. Although none of the sediment samples caused 100% mortality to C. tentans, the survival of approximately half of the samples significantly decreased. Pyrethroids, particularly cypermethrin, were identified as the primary cause of the toxicity, as indicated by TU analysis. The cause-effect relationship between pyrethroid concentrations and acute toxicity in Liaohe River was established on the basis of multiple lines of evidence. The proposed bioassay-directed analysis can be applied to screen pollutants in Liaohe River and thus be used as a basis for subsequent pollution control and mitigation. However, other contaminants should be determined because the causes of toxicity have yet to be fully elucidated. Moreover, other organisms should be subjected to toxicity tests to facilitate a reliable identification of priority pollutants. Acknowledgments This work was financially supported by Major Science and Technology Program for Water Pollution Control and Treatment (2012ZX07501-001) and gs2:National Science Foundation of China (51178438).
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Joint algal toxicity of 16 dissimilarly acting chemicals is predictable by the concept of independent action. Aquat. Toxicol. 63, 43e63. , D., 2010. Determination of pyrethroid insecticides in Feo, M.L., Eljarrat, E., Barcelo environmental samples. TrAC, Trends Anal. Chem. 29, 692e705. Gao, Y., Zhang, H., Fan, S., Tian, Y., Chen, J., 2012. Environmental occurrence and distribution of short chain chlorinated paraffins in sediments and soils from the liaohe river basin. Environ. Sci. Technol. 46, 3771e3778. Guo, W., He, M., Yang, Z., Lin, C., Quan, X., Wang, H., 2007. Distribution of polycyclic aromatic hydrocarbons in water, suspended particulate matter and sediment from Daliao River watershed, China. Chemosphere 68, 93e104. Harwood, A.D., You, J., Lydy, M.J., 2009. Temperature as a toxicity identification evaluation tool for pyrethroid insecticides: toxicokinetic confirmation. Environ. Toxicol. Chem. 28, 1051e1058. He, H., 2008. Current situation and development trend of organophosphorous insecticides industry. World Pestic. 30, 29e30. He, Y., Meng, W., Xu, J., Zhang, Y., Liu, S., Guo, C., 2015. Spatial distribution and toxicity assessment of heavy metals in sediments of Liaohe River, northeast China. Environ. Sci. Pollut. Res. 22, 14960e14970. Hintzen, E.P., Lydy, M.J., Belden, J.B., 2009. Occurrence and potential toxicity of pyrethroids and other insecticides in bed sediments of urban streams in central Texas. Environ. Pollut. 157, 110e116. Ho, K.T., Burgess, R.M., 2013. What’s causing toxicity in sediments? Results of 20 years of toxicity identification and evaluations. Environ. Toxicol. Chem. 32, 2424e2432. Holmes, R.W., Anderson, B.S., Phillips, B.M., Hunt, J.W., Crane, D.B., Mekebri, A., Connor, V., 2008. Statewide investigation of the role of pyrethroid pesticides in sediment toxicity in California’s urban waterways. Environ. Sci. Technol. 42, 7003e7009. Ke, X., Gao, L., Huang, H., Kumar, S., 2015. Toxicity identification evaluation of sediments in Liaohe River. Mar. Pollut. Bull. 93, 259e265. Kuivila, K.M., Hladik, M.L., Ingersoll, C.G., Kemble, N.E., Moran, P.W., Calhoun, D.L., Nowell, L.H., Gilliom, R.J., 2012. Occurrence and potential sources of pyrethroid insecticides in stream sediments from seven US metropolitan areas. Environ. Sci. Technol. 46, 4297e4303. Laskowski, D.A., 2002. Physical and chemical properties of pyrethroids. In: Reviews of Environmental Contamination and Toxicology. Springer New York, pp. 49e170. Li, H., Mehler, W.T., Lydy, M.J., You, J., 2011. Occurrence and distribution of sediment-associated insecticides in urban waterways in the Pearl River Delta, China. Chemosphere 82, 1373e1379. Li, H., Sun, B., Lydy, M.J., You, J., 2013a. Sediment-associated pesticides in an urban stream in Guangzhou, China: implication of a shift in pesticide use patterns. Environ. Toxicol. Chem. 32, 1040e1047. Li, H., Sun, B., Chen, X., Lydy, M.J., You, J., 2013b. Addition of contaminant bioavailability and species susceptibility to a sediment toxicity assessment: application in an urban stream in China. Environ. Pollut. 178, 135e141. Lv, J., Zhang, Y., Zhao, X., Zhou, C., Guo, C., Luo, Y., Meng, W., Zou, G., Xu, J., 2015. Polybrominateddiphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs) in sediments of Liaohe River: levels, spatial and temporal distribution, possible
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Y. He et al. / Environmental Pollution xxx (2016) 1e9 sources, and inventory. Environ. Sci. Pollut. Res. 22, 4256e4264. Maund, S.J., Hamer, M.J., Lane, M.C., Farrelly, E., Rapley, J.H., Goggin, U.M., Gentle, W.E., 2002. Partitioning, bioavailability, and toxicity of the pyrethroid insecticide cypermethrin in sediments. Environ. Toxicol. Chem. 21, 9e15. Mehler, W.T., Li, H., Lydy, M.J., You, J., 2011. Identifying the causes of sedimentassociated toxicity in urban waterways of the Pearl River Delta, China. Environ. Sci. Technol. 45, 1812e1819. Moran, P.W., Calhoun, D.L., Nowell, L.H., Kemble, N.E., Ingersoll, C.G., Hladik, M.L., Kuivila, K.M., Falcone, J.A., Gilliom, R.J., 2011. Contaminants in stream sediments from seven US metropolitan areas: data summary of a national pilot study. U. S. Geol. Surv. Sci. Investig. Rep. 5092, 22. USEPA, 2000. Methods for Measuring the Toxicity and Bioaccumulation of Sediment-associated Contaminants with Freshwater Invertebrates. EPA/600/R99/064, second ed. U.S. Environmental Protection Agency, Office of Research and Development, Washington, D.C. USEPA, 2007. Sediment Toxicity Identification Evaluation (TIE) Phases I, II, and III Guidance Document. EPA/600/R-07/080. U.S. Environmental Protection Agency, Office of Research and Development, Washington, D.C. Wang, H., He, M., Lin, C., Quan, X., Guo, W., Yang, Z., 2007. Monitoring and assessment of persistent organochlorine residues in sediments from the Daliaohe River Watershed, Northeast of China. Environ. Monit. Assess. 133, 231e242. Wang, J.Z., Li, H.Z., You, J., 2012. Distribution and toxicity of current-use insecticides in sediment of a lake receiving waters from areas in transition to urbanization. Environ. Pollut. 161, 128e133. Weinstein, J.E., Garner, T.R., 2008. Piperonylbutoxide enhances the bioconcentration and photoinduced toxicity of fluoranthene and benzo [a] pyrene to larvae of the grass shrimp (Palaemonetespugio). Aquat. Toxicol. 87, 28e36. Weston, D.P., Asbell, A.M., Hecht, S.A., Scholz, N.L., Lydy, M.J., 2011. Pyrethroid insecticides in urban salmon streams of the Pacific Northwest. Environ. Pollut.
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159, 3051e3056. Weston, D.P., Ding, Y., Zhang, M., Lydy, M.J., 2013. Identifying the cause of sediment toxicity in agricultural sediments: the role of pyrethroids and nine seldommeasured hydrophobic pesticides. Chemosphere 90, 958e964. Weston, D.P., Holmes, R.W., You, J., Lydy, M.J., 2005. Aquatic toxicity due to residential use of pyrethroid insecticides. Environ. Sci. Technol. 39, 9778e9784. Weston, D.P., Lydy, M.J., 2010. Focused toxicity identification evaluations to rapidly identify the cause of toxicity in environmental samples. Chemosphere 78, 368e374. Weston, D.P., You, J., Harwood, A.D., Lydy, M.J., 2009. Whole sediment toxicity identification evaluation tools for pyrethroid insecticides: III. Temperature manipulation. Environ. Toxicol. Chem. 28, 173e180. Weston, D.P., You, J., Lydy, M.J., 2004. Distribution and toxicity of sedimentassociated pesticides in agriculture-dominated water bodies of California’s Central Valley. Environ. Sci. Technol. 38, 2752e2759. Whittle, B., 2010. Communities in Peril: Asian Regional Report on Community Monitoring of Highly Hazardous Pesticide Use. Pesticide Action Network Asia and the Pacific, Penang. Jothee Printers, Penang, Malaysia, pp. 1e150. Xing, Y., Lu, Y., Dawson, R.W., Shi, Y., Zhang, H., Wang, T., Liu, W., Ren, H., 2005. A spatial temporal assessment of pollution from PCBs in China. Chemosphere 60, 731e739. You, J., Pehkonen, S., Weston, D.P., Lydy, M.J., 2008. Chemical availability and sediment toxicity of pyrethroid insecticides to Hyalella azteca: application to field sediment with unexpectedly low toxicity. Environ. Toxicol. Chem. 27, 2124e2130. Zhang, H., Zhao, X., Ni, Y., Lu, X., Chen, J., Su, F., Zhao, L., Zhang, N., Zhang, X., 2010. PCDD/Fs and PCBs in sediments of the Liaohe River, China: levels, distribution, and possible sources. Chemosphere 79, 754e762.
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Bioassay-directed identification of toxicants in sediments of Liaohe River, northeast China* Yan He a, b, Jian Xu a, *, Changsheng Guo a, Jiapei Lv a, Yuan Zhang a, Wei Meng a, b a b
State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012, China College of Water Sciences, Beijing Normal University, Beijing 100012, China
a r t i c l e i n f o
a b s t r a c t
Article history: Received 24 February 2016 Received in revised form 31 May 2016 Accepted 22 June 2016 Available online xxx
Contaminants accumulated in sediments may directly harm benthic organisms, however, the specific contaminants responsible for adverse effects have been poorly described. In this study, a bioassaydirected analysis combined with toxicity tests and chemistry analysis was conducted to determine the compounds eliciting the greatest toxicological effect in the sediments in Liaohe River, northeast China. A total of 24 sediment samples were examined to determine their acute toxicity to midge Chironomus tentans (C. tentans). Of these samples, 15 exhibited significant toxicity, with a mortality of 23%e93% (p < 0.05). Numerous contaminants, including 16 polycyclic aromatic hydrocarbons, 32 polychlorinated biphenyls, 20 organochlorine pesticides, 6 organophosphate pesticides, 8 pyrethroids, and 5 heavy metals were analyzed. On the basis of toxic unit (TU) analysis results, pyrethroids may contribute to the toxicity of 9 of the 15 toxic samples with concentrations of >1 TU. The significant correlation between the TUs of pyrethroids and the mortality of C. tentans (r2 ¼ 0.74, p < 0.01) confirmed the major role of pyrethroids in toxicity. The selected sediment samples responding to piperonyl butoxide and low temperature with the increased toxicity exhibited the characteristics of pyrethroids. The bioassay-based screening framework provided strong evidence that pyrethroids were the primary cause of sediment toxicity in Liaohe River. Further studies should therefore be conducted to regulate this important class of pollutants. © 2016 Elsevier Ltd. All rights reserved.
Keywords: Liaohe river Bioassay-directed analysis Pyrethroids Toxicity Chironomus tentans
1. Introduction Over the past several decades, rapid economic growth and urbanization have resulted in the deterioration of water quality in aquatic ecosystems in China. As such, the central and local governments of China have focused on water quality improvements; however, authorities have placed minor considerations on aquatic sediments in terms of practical water quality management. Sediments are important components of aquatic ecosystems providing habitat and feeding and spawning areas for aquatic organisms; sediments are also major repositories of chemical pollutants. High contaminant concentrations that accumulated in sediments are harmful to benthic organisms. Moreover, contaminated sediments adversely affect aquatic resources by causing ecological and economic damages, such as habitat degradation and costly
* This paper has been recommended for acceptance by Eddy Y. Zeng. * Corresponding author. E-mail address: [email protected] (J. Xu).
remediation and disposal actions (Ho and Burgess, 2013). Therefore, sediment contamination should be characterized to promote sediment assessment and management during decision making. Liaohe River is a heavily polluted river in northeast China. It consists of four big rivers, namely, Liao River, Taizi River, Hun River, and Daliao River. The middle- and down-stream of Liaohe River are the largest industrial bases of metallurgy, machinery, petrochemical, and building materials in northeast China. The pollution status of contaminants, including polycyclic aromatic hydrocarbons (Guo et al., 2007), organochlorine pesticides (Wang et al., 2007), polychlorinated biphenyls, polybrominated diphenyl ethers, polychlorinated dibenzo-p-dioxins (Lv et al., 2015; Zhang et al., 2010), short-chain chlorinated paraffins (Gao et al., 2012), and metals (He et al., 2015), in the sediments of Liaohe River have been extensively investigated. These studies have offered a good understanding of the occurrence and distribution of contaminants in this area but have provided limited information on their potential hazards to aquatic organisms. Hence, sediment-contact tests, which involve the exposure of benthic organisms to bulk sediments, are performed to assess the integrated effects of sediment-bound
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contaminants (Chapman and Anderson, 2005). When a positive toxic result is observed in a sediment, the sources of toxicity should be identified to determine the specific contaminants that need immediate attention and thus protect aquatic organisms and ecosystems (Anderson et al., 2010; Donnachie et al., 2014). Ke et al. (2015) examined sediments from Liao River to evaluate acute toxicity in midge larvae (Chironomus riparius) and found that As and Cd are probably associated with sediment toxicity. However, the identified toxicants have not been effectively confirmed, and the link between contaminant exposures and sediment toxicity remains unclear. To the best of our knowledge, limited data are available regarding the identification of active toxicants and establishment of cause-effect relationship. Specifically acting toxicants are difficult to identify because of the complexity of sediment matrix. Limited time and resources have also prompted researchers to devote appropriate mitigation efforts on highly toxic chemicals. The primary objective of this study was to examine the spatial distribution and magnitude of sediment toxicity to midge C. tentans in Liaohe River. The specific contaminants responsible for the toxicity were further explored. Additionally, the relationship between contamination levels and toxic effects was discussed by integrating bioassays with chemical analysis. This study provided comprehensive insights into ecological hazards posed by sediments in Liaohe River, and helped screen the priority pollutants in this area. This bioassay-based screening framework is of ecological and economic importance to assist reliable mitigation and management measures. 2. Materials and methods 2.1. Sample collection Twenty-four sediment samples (0e10 cm) were collected from Liaohe River with a Van Veen grab sampler in June 2014. Among these samples, nine (L1-L9) were from Liao River, five (H1eH5) from Hun River, six (T1eT6) from Taizi River, and four (D1-D4) from Daliao River (Fig. 1). At each sampling site, three replicate samples were collected, passed through a 2 mm mesh sieve, and then homogenized. Collected sediments were placed into sealed polyethylene bags, and transported on ice to the laboratory where they were stored at 4 C for toxicity testing and at 20 C for chemical analysis, respectively. 2.2. Chemical analysis The pollutants including polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), organophosphate pesticides (OPs), pyrethroids and inorganic heavy metals were selected as representatives of hazardous pollutants, due to their widespread distribution in sediment and potential toxicity to aquatic biota. The target compounds are shown in Table S1 in the supplementary materials. In addition, concentrations of acid-volatile sulfide (AVS) and simultaneously extracted metals (SEMs), total organic carbon (TOC) contents and sediment grain size (Table S2) were also analyzed. The details of chemicals and reagents, chemical analysis procedure and quality assurance and control are described in the supplementary materials. 2.3. Bulk sediment toxicity test C. tentans (Diptera: Chironomidae), a native species in Liaohe River, was used as the test organism. The midges obtained from Ecotoxicology Laboratory of Nankai University were laboratorycultured in accordance with standard protocols (USEPA, 2000).
Sediment samples were screened for toxicity by using 10-day survival tests for C. tentans. In brief, 50 g of sediments was placed in three replicate beakers, and each beaker was filled with 200 mL of reconstituted water prepared by adding salts to Milli-Q purified water (USEPA, 2007). After sediments were settled overnight, 10 s- or third-instar larvae were randomly introduced to each beaker and fed with 6 mg of finely ground Tetrafin Goldfish Flakes per day. Tests were performed at 23 C with a 16L: 8D photoperiod, and the overlying water was renewed twice daily. Temperature and dissolved oxygen were monitored on a daily basis; conductivity, pH, and ammonia were measured at the beginning and the end of the test. The samples containing dissolved oxygen with amounts decreased to 2.5 mg/ L were gently aerated. Each test batch was accompanied by control sediments obtained from a drinking water reservoir. After 10 days, surviving larvae were sieved and counted; their ash-free dry mass (AFDM) was determined following the standard methods (USEPA, 2000). 2.4. Estimation of toxicity The contribution of each contaminant to sediment toxicity was evaluated using a toxicity unit approach. The concentrations of contaminants in sediments were converted to TUs by dividing the 10-day sediment median lethal concentration (LC50) for midges on an organic carbon-normalized basis [Eq. (1)]. Conservative sediment benchmarks were used for metals, PAHs, and PCBs because of a lack of sediment toxicity data.
TU ¼
Contaminant concentration in sediment=TOC LC50 or sediment benchmark=TOC
(1)
The TUs of metals were calculated on the basis of consensusbased probable effect concentrations (PECs). AVS-SEM models were also used to evaluate the toxicity contributed by metals (Table S3). The TUs of PAHs were estimated in terms of equilibriumpartitioning sediment benchmarks (SESBTUs; Table S4). For PCBs, the PEC of total PCBs was considered (Table S5). The sediment LC50 of pesticides was obtained from previous studies and listed in Tables S6eS8. For the TUs of hexachlorocyclohexane (HCH), the concentration of gamma isomer was used because other isomers exhibit lower aquatic toxicity (Eaton and Klaassen, 2001). LC50 of allethrin was unavailable; as such, a screening-level toxicity value was computed by using an equilibrium partitioning approach based on a water-column 24 h LC50 value (ECOTOX database) (Moran et al., 2011). The TUs of heavy metals were expressed as the average of five metals. For each class of organics, the TUs of individual contaminant were added to assess mixture toxicity. Moreover, the combined toxicity of pesticides was estimated based on concentration addition (Belden et al., 2007). The approximation of concentration was 0.5 TU when mortality likely occurred; this value was also considered a threshold, that is, at >0.5 TU, a contaminant was assumed potentially responsible for mortality (Weston et al., 2004). 2.5. Confirmation Several lines of evidence were utilized to confirm possible toxicants. One approach was statistical correlation; in this approach, the estimated TUs of contaminants were plotted against the observed mortality. TU evaluation and statistical analysis suggested that pyrethroids may be the primary cause of toxicity; therefore, a focused toxicity identification evaluation (TIE), including piperonyl butoxide (PBO) addition and temperature manipulation, was applied to confirm their toxicity. PBO enhances
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Y. He et al. / Environmental Pollution xxx (2016) 1e9
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Fig. 1. Sampling sites in Liaohe River, northeast China.
the toxicity of pyrethroids by inhibiting their detoxification (Amweg et al., 2006a; Mehler et al., 2011). Toxicity tests were conducted with and without PBO (200 mg/L) in the overlying water. Approximately 80% of the water was replaced daily with fresh PBO solution or water; methanol (10 mL/L) as a solvent control was also included. As temperature is decreased, pyrethroid toxicity increases because of the slow pyrethroid metabolism and increased nerve sensitivity (Harwood et al., 2009). The tests were performed at 23 C, and a concurrent test was conducted at 17 C. The larvae cultured at 23 C, were acclimated to 17 C by reducing the temperature by 1 C/h and incubated overnight. Three replicates were tested for each treatment, and larval survival was determined after 10 days.
2.6. Data analysis The normality and homogeneity of data variances were evaluated by conducting Shapiro-Wilk’s and Levene’s tests, respectively. Differences in survival between samples and controls were determined through one-way ANOVA followed by Dunnett’s multiple comparison tests. Percent survival data were arcsine square roottransformed when necessary to satisfy the assumptions of the normality and homogeneity of variance. A sample was considered toxic if a significantly different response from controls was detected and if the difference in organisms’ response was greater than or equal to 20% relative to the controls. Individual TIE treatment results were compared with the unamended ones by performing a
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paired t-test when parametric assumptions were satisfied; otherwise, Wilcoxon rank sum tests were carried out. A probability level of p < 0.05 indicated statistical significance.
and urbanization.
3. Results and discussion
A series of sediment-associated contaminants was analyzed. The concentrations of metals, AVS, and SEM were determined (Table S3). A detailed discussion was presented in our previous study (He et al., 2015). Sixteen PAHs in the priority list were detected in the sediments, and total concentrations ranged from 82.5 ng/g to 25,400 ng/g (Table S4). The levels of PCBs were extremely lower (0.4e41.8 ng/g; Table S5) than those of PAHs. The identified PCB congeners were predominated by triCBs and tetraCBs, and this result is consistent with the production and usage of PCBs in China (Xing et al., 2005). Although most OCPs have been banned for 30 years, they remain detectable in sediments of Liaohe River. The total concentrations of OCPs varied from 1.3 ng/g to 94.8 ng/g, and high residues of HCHs and DDTs were found at most sites (Table S6). The concentrations of six commonly used OPs ranged from 2.8 ng/g to 45.5 ng/g (Table S7). Among the detected OPs, dichlorvos was the most abundant possibly because of its heavy consumption in China (He, 2008). High pyrethroid residues with concentrations of 2.2e102.5 ng/g were also detected in the sediments (Table S8). Among these residues, cypermethrin yielded the highest concentrations because its relatively low price has resulted in its widespread use (Whittle, 2010). Among the three classes of pesticides in Liaohe River, pyrethroids contributed the highest amounts to the total concentration and constituted an average of 56%, followed by OCPs (24%) and OPs (20%) (Fig. S1). OPs and pyrethroids are currently used pesticides as substitutes for OCPs. The concentration of pyrethroid residues in sediments has increased because of the high hydrophobicity and long half-life of this contaminant (Feo et al., 2010). OPs accumulate to a less extent than pyrethroids do because of their relatively high water solubility and fast degradation rates. OCPs levels were also slightly high due to their extensively historical application in Liaoning Province and prolonged persistence.
3.1. Sediment toxicity The sediments from Liaohe River were examined for their acute toxicity to midge C. tentans. Throughout the test, the water quality parameters were within acceptable ranges: dissolved oxygen, >80% saturation; temperature, 21 Ce24 C; pH, 6.9e8.1; conductivity, 378e586 ms/cm; and ammonia, 0.1e1.5 mg/L. The control survivals in all of the tests were 90%e95%, with an average AFDM of >0.48 mg (USEPA, 2000). Of the 24 sediment samples, 15 were significantly toxic to C. tentans. The highest toxicity, with 93% mortality, was observed at site H3 in Shenyang City. In Fig. 2, the levels of sediment toxicity within the entire watershed spatially varied. The sediments from the upstream of Liao River caused low mortality (7%e30%). At the downstream sites, the sediment toxicity increased and the sediments at site L9 in Panjin city resulted in a high mortality of 54%. The sediments from three other rivers flowing through many large- or medium-sized industrial cities exhibited a relatively higher toxicity than those from Liao River. In Hun River, four sediments were particularly toxic, with mortalities ranging from 30% to 93%, except the sediments at site H1 in Dahuofang Reservoir (10% mortality). Likewise, four sediments from Taizi River, where some industrial factories are situated, were significantly toxic with 27%e73% mortality. Among these sediments, those at site T2 were highly toxic because of large amounts of industrial and domestic wastewater discharged from Benxi City. The sediments collected from Daliao River elicited significant toxicity, with a mean mortality of 30%. Site D3 is in Yingkou City, an important coastal industrial base and traffic hub, where notable larval death was observed (40% mortality). Overall, the majority of toxic sites were located in large cities. This finding suggested that sediment toxicity may be closely associated with industrialization
3.2. Sediment chemistry
Fig. 2. The percent survival of C. tentans exposed to sediments collected from Liaohe River. Aasterisks (*) denote a significant difference from the controls (p < 0.05) and the difference in organisms’ response >20% relative to the controls.
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Y. He et al. / Environmental Pollution xxx (2016) 1e9 Table 1 The toxic units (TUs) of heavy metals, polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), organophosphate pesticides (OPs) and pyrethroids in sediments of Liaohe River.
L1 L2 L3 L4 L5 L6 L7 L8 L9 H1 H2 H3 H4 H5 T1 T2 T3 T4 T5 T6 D1 D2 D3 D4 mean
Metals
SPAH
SPCB
SOCP
SOP
Spyrethroid
0.09 0.08 0.09 0.08 0.13 0.15 0.08 0.08 0.19 0.15 0.39 0.33 0.22 0.18 0.19 0.20 0.18 0.23 0.13 0.13 0.08 0.21 0.17 0.13 0.16
0.12 0.05 0.07 0.04 0.08 0.11 0.03 0.08 0.17 0.04 0.32 0.21 0.91 0.12 0.10 0.71 0.50 0.46 0.11 0.44 0.35 0.20 0.13 0.38 0.24
<0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 0.06 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01 <0.01
0.24 0.02 0.14 0.05 0.24 0.07 0.02 0.11 0.15 0.06 0.30 0.16 0.07 0.42 0.08 0.28 0.10 0.24 0.12 0.42 0.08 0.03 0.03 0.09 0.15
0.02 0.06 <0.01 0.05 0.07 0.02 0.03 0.21 0.02 <0.01 0.03 0.02 <0.01 <0.01 0.01 <0.01 <0.01 0.01 0.02 0.01 0.02 0.13 0.15 0.01 0.05
0.30 0.66 1.65 0.55 0.55 0.69 0.42 1.65 3.00 0.64 0.36 0.22 0.26 2.77 1.24 0.63 0.46 1.34 0.82 1.79 1.27 1.21 1.94 1.33 1.07
3.3. Causes of sediment toxicity A TU approach was employed in our study to assess the contribution of contaminants to the observed toxicity and to determine the causes of toxicity. The estimated TUs of each group of contaminants are listed in Table 1. The average TUs of the five metals ranged from 0.08 to 0.39, with an average of 0.16; this result indicated their minimal contribution to toxicity. The AVS-SEM models further confirmed that the potential toxicity of metals to aquatic life is unexpected at most sites (He et al., 2015). On the basis
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of ESBs for PAHs mixtures, their predicted TUs were between 0.03 and 0.91, with an average of 0.24. Three sediments (H4, T2 and T3) contained more than 0.5 TU; this result suggested that PAHs slightly contributed to the observed mortality, but their risk should be considered at the three sites. Compared with PAHs, PCBs were present at very low concentrations; as such, PCBs could not be accounted for the toxicity because their TUs were <0.01. Among the target OCPs, g-HCHs, endosulfan, and endrin yielded TUs of >0.01, but their combined contribution was <0.5 TU at all of the sites. Therefore, OCPs unlikely influenced the mortality of C. tentans. Chlorpyrifos was the only OP with a TU of >0.01. Indeed, OPs were unlikely the cause of acute toxicity because of their low TUs (0.01e0.21). The highest values were observed at most sites when pyrethroid concentrations were converted to TUs. Of the 24 sediments, 11 yielded 1 TU and 10 caused a significantly reduced survival of C. tentans. Conversely, the sediments at site T6 elicited low toxicity with 23.3% mortality at 1.79 TU. The overestimation of toxicity may be attributed to the reduced bioavailability of pyrethroids in sediments because of grain size or organic carbon type (You et al., 2008; Li et al., 2013b). This sediment contained high percent of fine grains (silt and clay fractions, 90%). Pyrethroids are strongly adsorbed onto fine particles; as such, they are less available to C. tentans (Laskowski, 2002). Low mortality was observed in the 13 remaining sediments containing <0.5 TU except at the three sites (H2, H3, and T2), where a very high mortality may be linked to the unanalyzed contaminants. Overall, 9 of the 15 toxic samples (out of the 24 samples) yielded at least 1 TU. This finding suggested that pyrethroids mostly accounted for the observed toxicity. When TUs were apportioned among individual pyrethroids, cypermethrin apparently contributed the greatest amount to pyrethroid TUs and yielded an average of 77% (Fig. 3). Of the 11 toxic samples, cypermethrin TUs exceeded the threshold of 0.5. This result indicated the major role of cypermethrin in toxicity. Deltamethrin was the second-largest contributor, with approximately 13% of TUs. For the other pyrethroids, their total contributions to toxicity were minimal and accounted for 10% of the overall TUs.
Fig. 3. Comparison of mean pyrethroid TUs in sediments of Liaohe River, and the contribution of the various pyrethroids to total TUs.
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A concentration yielding 1 TU should cause 50% mortality, but mortality of <50% was noted in this study. This phenomenon usually occurs; previous studies also revealed that sediments with 1e3 pyrethroid TUs cause mortality that is lower than the predicted value (Weston et al., 2005; Amweg et al., 2006b; Hintzen et al., 2009). This discrepancy is possibly attributed to the variability of LC50 of pyrethroids among different types of sediments (Amweg et al., 2005; Maund et al., 2002). Furthermore, a TU approach based on concentration addition may overestimate mixture toxicity, but differences between observed and predicted effect concentrations do not exceed a factor of 3 (Faust et al., 2003). Therefore, TU analysis suggested that pyrethroids are probably the major cause of sediment toxicity in Liaohe River.
3.4. Confirmation of pyrethroid-related toxicity Three lines of evidence were applied to determine whether pyrethroids are responsible for the toxicity of C. tentans. One approach was statistical correlation; in this approach, the sum or individual pyrethroid TUs were regressed against the observed mortality after the three outliers were eliminated. For the three sediments (H2, H3, and T2), non-pyrethroids apparently contributed to the toxicity. In Fig. 4, total pyrethroid TUs were significantly correlated with C. tentans toxicity (Spearman’s rank correlation, r2 ¼ 0.74, p < 0.01). This finding confirmed that pyrethroids were primarily responsible for mortality. In terms of individual pyrethroid, cypermethrin was the main contributor to toxicity, as
Fig. 4. Relationship between pyrethroid TUs and observed toxicity in sediments of Liaohe River (Spearman’s rank correlation).
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indicated by a highly significant correlation (r2 ¼ 0.71, p < 0.01). TIEs, including PBO addition and temperature manipulation, were effective tools to characterize pyrethroid-related toxicity. In this study, 11 of 24 sediments were evaluated on the basis of the focused TIEs. The control sediments with PBO in the overlying water exhibited 80%e90% survival; this result indicated that PBO did not induce toxicity. In Fig. 5, toxicity was increased in all of the samples by the added PBO, and 9 of the samples exhibited notable differences (p < 0.05). At sites L7 and H1, the added PBO slightly increased sediment toxicity possibly because of the relatively low pyrethroid contents. By contrast, the added PBO significantly increased the toxicity at site L1 (p < 0.05), although the pyrethroid concentrations were low (TU ¼ 0.30). This phenomenon probably
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occurred because other contaminants, such as PAHs, in sediments may be synergized by PBO and thus contributed to the toxicity (Weinstein and Garner, 2008). Sediment toxicity increased at all of the sites except site L7 when the test temperature was reduced from 23 C to 17 C (Fig. 5). In the sediments at site L7, the TUs of the analyzed pollutants were very low; this finding suggested the presence of some undetermined toxicants, and their toxicities decreased at low temperature. Similar studies have also revealed that low temperature occasionally performs inconsistently and induces a reduction in toxicity (Weston et al., 2009; Weston and Lydy, 2010). Of the 11 samples, only three exhibited a significant increase in toxicity (p < 0.05). The slight temperature response was consistent with that described in a previous study, which revealed
Fig. 5. The effect of the various TIE manipulations on the toxicity of sediments from Liaohe River. Aasterisks (*) indicate significant differences between amended sediments and unamended ones (p < 0.05).
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an increased toxicity in 4 of 13 samples at low temperature (Weston et al., 2013). This result indicated that low temperature treatment slightly affected pyrethroid toxicity. The two TIE response profiles were consistently indicative of pyrethroid toxicity. In summary, the focused TIE manipulations combined with TU estimation and strong correlations provided sufficient evidence that pyrethroids were the main causative toxicant in the majority of sediments from Liaohe River. However, the cause of toxicity in a small number of samples has yet to be determined. Emerging contaminants should be identified in future studies. Similar researches on the causes of sediment toxicity have been previously conducted in California (Holmes et al., 2008), Texas (Hintzen et al., 2009), Illinois (Ding et al., 2010) and Oregon/ Washington (Weston et al., 2011) in the United States (USA) and Pearl River Delta (Mehler et al., 2011) and Guangzhou (Li et al., 2013a) in China. In those studies, pyrethroids played a major role in the observed toxicity. As for the individual pyrethroid, bifenthrin was the main toxic contributor in USA, while in China cypermethrin was the dominant acting toxicant. This difference is due to the applicator preference in pyrethroid usage in the two countries (Wang et al., 2012). In USA, most of the households have private lawns, where bifenthrin is primarily applied for structural pest control and lawn maintenance (Weston et al., 2005). In contrast, cypermethrin is extensively used in agriculture, mosquito control and landscape maintenance in China due to its relatively low price (Li et al., 2011). In the current study, Liaohe River had slightly lower pyrethroid concentrations and less toxicity as compared to Pearl River Delta and Chebei Creek, Guangzhou in south China (Mehler et al., 2011; Li et al., 2013a). This result can be explained by the different land-use types and urbanization level. In south China, highly developed economy has created substantial urbanization areas. Dense populations and municipal landscaping lead to the intensive use of pyrethroids. In comparison, Liaohe River in northeast China has a lot of agricultural land, which is less urbanized than south China. Similar studies have also revealed that total pyrethroid concentrations were significantly related to the percent urban land (Kuivila et al., 2012). Moreover, the climatic conditions may affect the amounts of pyrethroid use. Humid climate in south China promotes the breeding of mosquitoes, resulting in increased pyrethroid consumption (Li et al., 2011). 4. Conclusions This study fully demonstrated the acute toxic effects of contaminants in the sediments of Liaohe River. Although none of the sediment samples caused 100% mortality to C. tentans, the survival of approximately half of the samples significantly decreased. Pyrethroids, particularly cypermethrin, were identified as the primary cause of the toxicity, as indicated by TU analysis. The cause-effect relationship between pyrethroid concentrations and acute toxicity in Liaohe River was established on the basis of multiple lines of evidence. The proposed bioassay-directed analysis can be applied to screen pollutants in Liaohe River and thus be used as a basis for subsequent pollution control and mitigation. However, other contaminants should be determined because the causes of toxicity have yet to be fully elucidated. Moreover, other organisms should be subjected to toxicity tests to facilitate a reliable identification of priority pollutants. Acknowledgments This work was financially supported by Major Science and Technology Program for Water Pollution Control and Treatment (2012ZX07501-001) and gs2:National Science Foundation of China (51178438).
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