中國四種土壤跳蟲對重金屬汞的響應

劉傳棟，苗秀蓮，賈少波，張廣征，單麗英

聊城大學生命科學學院 生態學與生物多樣性山東省重點實驗室，聊城 252059

中國四種土壤跳蟲對重金屬汞的響應

劉傳棟，苗秀蓮，賈少波*，張廣征，單麗英

聊城大學生命科學學院 生態學與生物多樣性山東省重點實驗室，聊城 252059

汞；跳蟲；半致死濃度；慢性毒性實驗；低劑量刺激效應

近期由環境保護部和國土資源部發布的《全國土壤污染狀況調查公報》(2014年4月17日)表明，中國目前國土污染形勢嚴峻，其中重金屬對耕地的污染涉及糧食安全，尤其需要關注。我國污水灌溉中重金屬Hg含量的高低與相對應的灌區土壤中重金屬的累積量多少基本一致，目前受重金屬污染的耕地已經有1/6左右，如何進行耕地重金屬污染的監測、綜合防治和生態修復，成為迫切需要考慮和解決的問題[1-4]。土壤動物作為土壤生態系統中的重要組成部分和土壤健康狀況的體現者，選擇其成員作為模式動物是探索土壤生態系統毒理學的重要途徑。

1 材料和方法(Materials and methods)

1.1 儀器與試劑

儀器：RTOP多段編程人工氣候箱(杭州托普儀器有限公司)，SZX12體視顯微鏡(Olympus公司)，BX51正置顯微鏡(Olympus公司)，BSA224S電子天平(Sartorius公司)。試劑：氯化汞(HgCl2)等化學試劑為國產分析純試劑。

1.2 實驗材料

整個實驗及先期培養均在上口徑120 mm、下口徑88 mm、高65 mm的專用培養瓶(透明聚丙烯容器)中進行，培養瓶底部覆蓋一層0.5 cm厚的瓊脂培養基(1.5%)，頂部有透明聚丙烯蓋封口。

1.3 實驗方法

1.3.1 LC50急性毒性試驗(存活試驗)

暴露期間喂食含毒酵母糊，其中Hg2+含量對應6個實驗濃度組分別為0.0、0.4、2.0、10、50、100 mg·kg-1干重。初始每隔12 h觀察一次，24 h后每隔1天觀察一次跟蹤觀察到第7天，并做好記錄。死亡的判定標準是觸之不動。

1.3.2 28 d慢性毒性試驗

根據預試驗結果，設置6個濃度0.05、0.5、1.0、2.0、4.0、8.0 mg·L-1的染毒組和一個對照組(0 mg·L-1)進行28 d慢性毒性試驗。每組設3個重復。培養基染毒過程及暴露期間的實驗方法與LC50急性毒性試驗相同。同LC50急性毒性試驗，暴露期間喂食含毒酵母糊，其Hg2+干重含量分別對應染毒組濃度。每周開口通氣并補充少量蒸餾水。幼蟲計數方法采用顯微拍照定格后人工識別計數。

1.4 數據處理

數據用平均值±標準差(mean±SD)表示。采用SPSS Statistics 19.0統計分析軟件；在95%的置信水平進行單因素方差分析(One-Way ANOVA)，Duncan法對平均值進行多重比較，p<0.05為差異顯著；采用Bliss法(機率單位加權回歸法)[14-15]計算汞(Hg2+)對跳蟲的半致死濃度(LC50)。

2 結 果(Results)

2.1 LC50急性毒性實驗

2.1.1 Hg急性毒性期間的跳蟲行為觀察

2.1.2 Hg對4種跳蟲的24 h-LC50和72 h-LC50

2.1.3 急性實驗中四種跳蟲7 d內在不同Hg2+濃度下的存活情況

表1 Hg對四種跳蟲24 h, 72 h的半致死濃度Table 1 24 h, 72 h-LC50 values of Hg to four species of Collembolans

表2 不同濃度Hg對曲毛裸長()7 d存活影響的F值和p值Table 2 F values and p values of different concentrations of Hg on survival of S. curviseta for 7 d

圖1 不同濃度Hg對曲毛裸長()7 d存活的影響注：圖柱上字母的不同表示差異達到 0.05 的顯著水平。Fig. 1 Effect of different concentrations of Hg on survival of S. curviseta for 7 dNote: Different letters on columns mean significant difference at p<0.05.

圖2 不同濃度Hg對小原等節()7 d存活的影響Fig. 2 Effect of different concentrations of Hg on survival of P. minuta for 7 d

暴露時間TimeofexposureF值Fvaluesp值pvalues12h168.750.0001d2064.500.0003d686.330.0005d893.10.0007d1014.600.000

表4 不同濃度Hg對四刺泡角()7 d存活影響的F值和p值Table 4 F values and p values of different concentrations of Hg on survival of C. duplicispinosa for 7 d

圖3 不同濃度Hg對四刺泡角()7 d存活的影響Fig. 3 Effect of different concentrations of Hg on survival of C. duplicispinosa for 7 d

圖4 不同濃度Hg對茉莉花長角()7 d存活的影響Fig. 4 Effect of different concentrations of Hg on survival of Entomobrya sp. for 7 d

暴露時間TimeofexposureF值Fvaluesp值pvalues12h16.240.0001d31.380.0003d60.590.0005d91.090.0007d58.320.000

2.2 28 d慢性毒性試驗

圖5 不同濃度Hg對茉莉花長角()28 d的影響Fig. 5 Effect of different concentrations of Hg on survival of Entomobrya sp.for 28 days

圖6 不同濃度Hg對小原等節()28 d的影響Fig. 6 Effect of different concentrations of Hg on survival of P. minuta for 28 days

暴露時間TimeofexposureF值Fvaluesp值pvalues1d12.250.0007d6.980.00114d23.070.00021d13.100.00028d6.880.001

圖7 不同汞濃度下28 d時茉莉花長角()和小原等節()的幼蟲數量與成蟲數量比注：折線圖上的**表示差異達到 0.01 的極顯著水平。Fig. 7 Ratios of live offspring to adults of Entomobrya sp. and P. minuta on Hg concentrations for 28 daysNote: ** means significant difference at p<0.01.

3 討 論(Discussion)

3.1 不同Hg2+濃度對跳蟲7 d內存活的影響

3.2 低濃度汞對跳蟲存活和繁殖的影響

3.3 跳蟲的24 h，72 h-LC50變化及其適應性意義

[1] 宋 偉, 陳百明, 劉 琳. 中國耕地土壤重金屬污染概況[J]. 水土保持研究, 2013, 20(2): 293-298

Song W, Chen B M, Liu L. Soil Heavy Metal pollution of cultivated land in China [J]. Research of Soil and Water Conservation, 2013, 20(2): 293-298 (in Chinese)

[2] 宰松梅, 王朝輝, 龐鴻賓. 污水灌溉的現狀與展望[J]. 土壤, 2006, 38(6): 805-813

Zai S M, Wang Z H, Pang H B. Situation and prospect of sewage irrigation in agriculture [J]. Soils, 2006, 38(6): 805-813 (in Chinese)

[3] 王 靜, 林春野, 陳瑜琦, 等. 中國村鎮耕地污染現狀、原因及對策分析[J]. 中國土地科學, 2012,26(2): 25-30,43

Wang J, Lin C Y, Chen Y Q, et al. Cultivated land pollution at township level in China: Situation, factors and measures [J]. China Land Sciences, 2012, 26(2): 25-30, 43 (in Chinese)

[4] 張乃明, 陳建軍, 常曉冰. 污灌區土壤重金屬累積影響因素研究[J]. 土壤, 2002, 34(2): 90-93

[5] 陳建秀, 麻智春, 嚴海娟, 等. 跳蟲在土壤生態系統中的作用[J]. 生物多樣性, 2007, 15(2): 154-161

Chen J X, Ma Z C, Yan H J, et al. Roles of springtails in soil ecosystem [J]. Biodiversity Science, 2007, 15(2): 154-161 (in Chinese)

[6] Kim S W, An Y J. Jumping behavior of the springtail Folsomia candida as a novel soil quality indicator in metal-contaminated soils [J]. Ecological Indicators, 2014, 38: 67-71

[7] Santorufoa L, Gestelb V C, Maisto G. Sampling season affects conclusions on soil arthropod community structure responses to metal pollution in Mediterranean urban soils [J]. Geoderma, 2014, 226: 47-53

[8] 劉玉榮, 賀紀正, 鄭袁明. 跳蟲在土壤污染生態風險評價中的應用[J]. 生態毒理學報, 2008, 3(4): 323-330

Liu Y R, He J Z, Zheng Y M. A review of application of springtails in ecological risk assessment of contaminated soils [J]. Asian Journal of Ecotoxicology, 2008, 3(4): 323-330 (in Chinese)

[9] US EPA. Framework for Ecological Risk Assessment [R]. Washington DC: US EPA. 1992. EPA/630/R-92/001

[10] Amorim M J B, Pereira C, Menezes-Oliveira V B, et al. Assessing single and joint effects of chemicals on the survival and reproduction of Folsomia candida(Collembola) in soil [J]. Environmental Pollution, 2012, 160(1): 145-152

[11] An Y J, Kim S W, Lee W M. The collembola Lobella sokamensis juvenile as a new soil quality indicator of heavy metal pollution [J]. Ecological Indicators, 2013, 27: 56-60

[12] Hamda N T, Forbes V E, Stark J D, et al. Stochastic density-dependent matrix model for extrapolating individual-level effects of chemicals to the population: Case study on effects of Cd on Folsomia candida [J]. Ecological Modelling, 2014, 280: 53-64

[13] Liu Y R, Zheng Y M, Zhang L M, et al. Effects of mercury on reproduction, avoidance, and heat shock protein gene expression of the soil springtail Folsomia candida [J]. Environmental Toxicology and Chemistry, 2010, 29(3): 654-659

[14] 黎七雄, 汪暉, 肖清秋, 等. 半數致死量(LD50)Bliss法的評價及計算[J]. 數理醫藥學雜志, 1995, 8(4): 318-320

[15] 周一平. 用SPSS軟件計算新藥的LD50[J]. 藥學進展, 2003, 27(5): 314-316

Zhou Y P. The calculation of LD50in new drugs with software SPSS [J]. Progress in Pharmaceutical Sciences, 2003, 27(5): 314-316 (in Chinese)

[16] 岳麗蕊, 賈少波, 趙巖, 等. 重金屬銅對3種跳蟲的影響[J]. 生物學通報, 2011, 46(1): 51-54

[17] 申開麗, 俞潔, 林廣, 等. 低劑量刺激效應在污染物風險評價中的意義及前景[J]. 環境與職業醫學, 2012, 29(3): 183-186

Shen K L, Yu J, Lin G, et al. Importance and Potential application of hormetic effect in risk assessment of environmental pollutants [J]. Journal of Environmental & Occupational Medicine, 2012, 29(3): 183-186 (in Chinese)

[18] Calabrese E J. Evidence that hormesis represents an "overcompensation" response to a disruption in homeostasis[J]. Ecotoxicology Environment Safety, 1999, 42(2):135-137

[19] Stebbing A R. Growth hormesis: A by-product of control [J]. Health Physics, 1987, 52(5): 543-547

◆

ResponsesofFourSpeciesofChineseSoilCollembolatoHg

Liu Chuandong, Miao Xiulian, Jia Shaobo*, Zhang Guangzheng, Shan Liying

Shandong Key Laboratory of Ecology and Biodiversity, School of Life Sciences, Liaocheng University, Liaocheng 252059, China

15 May 2014accepted19 June 2014

To study the toxic effects of Hg on diverse species of Collembola, four Chinese dominant species of collembolans, Sinella curviseta, Ceratophysella duplicispinosa, Proisotoma minuta and Entomobrya sp. were exposed to six different concentrations of Hg for 24 h, 72 h LC50measurement. The LC50values for S. curviseta, C. duplicispinosa, P. minuta and Entomobrya sp. were calculated according to the Bliss method to be 92.42, 36.85, 34.93, and 47.56 mg·L-1, and 95% confidence limits were 68.57～168.6, 29.61～43.53, 25.57～44.94 and 34.33～65.11 mg·L-1, respectively, while the corresponding 72 h-LC50values were 21.92, 20.17, 18.44, and 15.99 mg L-1, and 95% confidence limits were 16.18～29.82, 6.64～29.81, 13.87～26.45 and 10.03～21.37 mg·L-1. At the same time, the Proisotoma minuta and Entomobrya sp. were exposed to Hg of six different lower concentrations for 28 d survival and reproduction experiment to determine the numbers of adults and offspring. The result shows that Entomobrya sp. and S. curviseta have higher tolerance for Hg, followed with P. minuta, while C. duplicispinosa ranks last. In addition, P. minuta display some hormesis at about 0.05 mg·L-1of Hg2+.

Hg; collembola; LC50; chronic toxicity test; hormesis

國家自然科學基金(No. 31272383) ；山東省自然科學基金(No. ZR2010CM018)

劉傳棟(1976-)，男，碩士，研究方向為動物生態毒理學，E-mail: lcliucd@189.cn；

*通訊作者(Corresponding author)，E-mail: jiasbio@163.com

10.7524/AJE.1673-5897-20140515008

2014-05-15錄用日期：2014-06-19

1673-5897(2014)5-964-08

： X171.5

： A

賈少波(1959—)，男，生態學博士，教授，主要研究方向為動物學及生態學。

劉傳棟, 苗秀蓮, 賈少波, 等. 中國四種土壤跳蟲對重金屬汞的響應[J]. 生態毒理學報，2014, 9(5): 964-971

Liu C D, Miao X L, Jia S B, et al. Responses of four species of Chinese soil collembola to Hg [J]. Asian Journal of Ecotoxicology, 2014, 9(5): 964-971 (in Chinese)