摘要:
为了进一步探索种群水平的生态风险评估方法,本文利用β-N-Acetyl-D-glucosaminidase (NAGase)的变化量来监测农药对摇蚊种群发育的影响。从花翅摇蚊Chironomus kiiensis体内分离纯化得到电泳纯的NAGase,并通过免疫大白兔制得NAGase的多克隆抗体。运用间接非竞争ELISA法检测抗体特异性,结果表明其与共同存在于水体的一些生物的NAGase的交叉反应率为隆线溞4.41%、老年低额溞3.12%、多刺裸腹溞3.40%、中华薄壳介4.17%、日本沼虾3.23%、白纹伊蚊7.50%、小球藻< 0.5%。运用抗体测得毒死蜱、氰戊菊酯和阿维菌素3种杀虫剂对于摇蚊NAGase释放量的12 d-EC50分别为1.2012、0.0043和0.6281μg·L-1,以NAGase活力作为测试终点,测得相应的12 d-EC50:1.4765、0.0051和0.6756 μg·L-1,两者差异不显著,但均显著低于以死亡作为测试终点的12 d-LC50:4.8171、0.0954和2.1340 μg·L-1,且毒力大小均为氰戊菊酯 > 阿维菌素 > 毒死蜱。上述结果表明,利用NAGase多克隆抗体可以特异性地检测农药对花翅摇蚊种群发育的影响。
Abstract:
Risk assessment on population level is still challenged by its proper in terms of methodology. In this work, population development of Chironomus kiiensis was assessed by quantity of β-N-Acetyl-D-glucosaminidase (NAGase) released from the larvae. To do so, the NAGase was purified from the bodies and polyclonal antibodies were obtained by immunizing rabbits with the purified enzyme. Indirect and non-competitive enzyme-linked immunosorbent assay (ELISA) was employed to analyze specificity of the antibodies, and the results showed that the cross reactions was 4.41% for Daphnia carinata, 3.12% for Simocephalus vetulus, 3.40% for Moina macrocopa, 4.17% for Dolerocypris sinensis, 3.23% for Macrobrachium nipponense, 7.50% for Aedes albopictus, and < 0.5% for Chlorella vulgaris. After being exposed for 12 days to pesticides, the EC50 based on residues of NAGase, which was determined by ELISA, was 1.201 μg·L-1 for chlorpyrifos, 0.004 μg·L-1 for fenvalerate, and 0.628 μg·L-1 for abamectin. The EC50 determined by activity of NAGase was 1.477 μg·L-1 for chlorpyrifos, 0.005 μg·L-1 for fenvalerate, and 0.676 μg·L-1 for abamectin. However, the LC50 were measured to be 4.817 μg·L-1 for chlorpyrifos, 0.095 μg·L-1 for fenvalerate, and 2.134 μg·L-1 for abamectin. Our results suggested that the impact of pesticides on population development of Chironomus kiiensis could be assessed distinctively by quantity of NAGase released by the larvae.
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