低剂量长期铅暴露对雄性黑斑蛙精巢组织关键酶活性的影响

在实验条件下,将健康的性成熟雄性黑斑蛙暴露于0.1、0.2、0.4、0.8和1.6mg·L-1的Pb2+溶液中30d,以Ca2+-ATP酶、Na+-K+-ATP酶、Ca2+-Mg2+-ATP酶、β-葡萄糖醛酸苷酶(β-DG)、乳酸脱氢酶(LDH)和酸性磷酸酶(ACP)活性为指标,进行了长期铅暴露对雄性黑斑蛙生殖毒性的研究.结果表明:随着铅染毒剂量的增加,Ca2+-ATP酶和Na+-K+-ATP酶活性被诱导,Ca2+-Mg2+-ATP酶活性被抑制,当Ca2+-Mg2+-ATP酶的抑制程度超过Ca2+-ATP酶和Na+-K+-ATP酶的代偿机制时,会抑制精子的发生,进而导致雄性生殖毒性;在1.6mg·L-1Pb2+处理下,ACP酶被显著抑制,提示支持细胞受损,精子总数受到影响;在0.4mg·L-1Pb2+处理下,LDH酶被显著抑制,提示生精细胞受损,精子质量受到影响.     

Molecular characterization and developmental expression patterns of thyroid hormone receptors (TRs) and their responsiveness to TR agonist and antagonist in Rana nigromaculata

Considering some advantages of Rana nigromaculata as an experimental species, we propose that this species, like Xenopus laevis, could be used to assay thyroid hormone（TH） signaling disrupting actions. To validate the utilizability of R. nigromaculata, we investigated the responsiveness of R. nigromaculata to a TH receptor（TR） agonist（T3） and antagonist（amiodarone） by analyzing expression, based on characterizing TR cDNA and developmental expression patterns. With high levels of identity with the corresponding genes in X. laevis, both TRα and TRβ in R. nigromaculata exhibited roughly similar developmental expression patterns to those of X. laevis, in spite of some species-specific differences. Both TRα and TRβ expression had greater changes in the liver and intestine than in the tail and brain during metamorphosis. T3 exposure for 2 days induced more dramatic increases of TRβ expression in stage 27 than in stage34 tadpoles but not in stage 42 tadpoles, showing that the responsiveness of R. nigromaculata to TH decreased with development and disappeared at the onset of metamorphic climax.Corresponding to greater changes of TRβ expression in the liver and intestine than in the tail and brain during metamorphosis, the liver and intestine had higher responsiveness to exogenous T3 than the tail and brain. Amiodarone inhibited T3-induced TRβ expression. Our results show that R. nigromaculata can be used as a model species for assaying TH signaling disrupting actions by analyzing TRβ expression, and intestine tissues at stage 27 are ideal test materials due to high responsiveness and easy accessibility.     

异噻唑啉酮类杀菌剂对黑斑蛙胚胎和蝌蚪的急性毒性

异噻唑啉酮类杀菌剂1,2-苯并异噻唑-3-酮（BIT）和甲基异噻唑啉酮（MIT）虽已在多种行业中广泛使用,但目前有关其毒性尤其对水体中生物毒性的数据还较少。鉴于BIT和 MIT在水体中普遍存在,本文研究了这两种污染物对两栖动物黑斑蛙胚胎和蝌蚪的急性毒性。黑斑蛙胚胎和蝌蚪分别暴露系列浓度的BIT和 MIT,观察化学品对其生长、发育和运动的影响,计算96小时半数致死浓度（96 h-LC₅₀）和96小时半数致畸浓度（96 h-TC₅₀）,确定最小生长抑制浓度（MCIG）。结果发现,BIT对黑斑蛙胚胎的96 h-LC₅₀和96 h-TC₅₀分别为2.99 mg?L^-1和0.60 mg?L^-1,MCIG小于0.40 mg?L^-1,对蝌蚪的96 h-LC₅₀为6.44 mg?L^-1。MIT对黑斑蛙胚胎的96 h-LC₅₀和96 h-TC₅₀分别为5.30 mg?L^-1和2.36 mg?L^-1,MCIG为2.59 mg?L^-1,对蝌蚪的96 h-LC₅₀为7.58 mg?L^-1。根据《化学农药环境安全评价准则报批稿》中两栖动物蝌蚪急性毒性的分级标准,判定BIT和MIT的毒性等级为中等。该毒性数据为异噻唑啉酮类杀菌剂的环境管理提供参考。     

Effects of combinations of malathion and cypermethrin on survivability and time of metamorphosis of tadpoles of Indian cricket frog (Fejervarya limnocharis)

This study was undertaken to determine the effect of environmentally realistic concentrations of two commonly used pesticides viz., malathion and cypermethrin, using a fully 3 × 3 factorial experiments on the survivability and time of metamorphosis in a common rice paddy field frog (cricket frog) Fejervarya limnocharis under laboratory conditions. The results suggest that cypermethrin is more toxic than malathion and combinations of higher concentrations of cypermethrin (50 μg/L) with malathion (250 and 500 μg/L) are more deleterious to the survivability of tadpoles. With increasing cypermethrin concentration, the survivability of tadpole decreased (r = ?0.986, P = 0.108). But cypermethrin alone induced early metamorphosis among the surviving tadpoles. However, there was a delay in the time required for metamorphosis induced by malathion and its combination with cypermethrin. The delay in metamorphosis may indicate the altered physiological fitness of the individual. The emergent froglets will be subjected to environmental stressors like high temperature and less humidity of post-monsoon tropical climate that could enhance negative influence triggered by pesticides.     

异噻唑啉酮类杀菌剂对黑斑蛙胚胎和蝌蚪的急性毒性

异噻唑啉酮类杀菌剂1,2-苯并异噻唑-3-酮（BIT）和甲基异噻唑啉酮（MIT）虽已在多种行业中广泛使用,但目前有关其毒性尤其对水体中生物毒性的数据还较少。鉴于BIT和 MIT在水体中普遍存在,本文研究了这两种污染物对两栖动物黑斑蛙胚胎和蝌蚪的急性毒性。黑斑蛙胚胎和蝌蚪分别暴露系列浓度的BIT和 MIT,观察化学品对其生长、发育和运动的影响,计算96小时半数致死浓度（96 h-LC₅₀）和96小时半数致畸浓度（96 h-TC₅₀）,确定最小生长抑制浓度（MCIG）。结果发现,BIT对黑斑蛙胚胎的96 h-LC₅₀和96 h-TC₅₀分别为2.99 mg?L^-1和0.60 mg?L^-1,MCIG小于0.40 mg?L^-1,对蝌蚪的96 h-LC₅₀为6.44 mg?L^-1。MIT对黑斑蛙胚胎的96 h-LC₅₀和96 h-TC₅₀分别为5.30 mg?L^-1和2.36 mg?L^-1,MCIG为2.59 mg?L^-1,对蝌蚪的96 h-LC₅₀为7.58 mg?L^-1。根据《化学农药环境安全评价准则报批稿》中两栖动物蝌蚪急性毒性的分级标准,判定BIT和MIT的毒性等级为中等。该毒性数据为异噻唑啉酮类杀菌剂的环境管理提供参考。     

敌百虫对中国林蛙蝌蚪生长发育的毒性效应

为评价水域环境中敌百虫(trichlorfon)污染对两栖类幼体的急性毒性,将中国林蛙(Rana chensinensis)28～29期(Gos-ner)蝌蚪分别暴露于10～30 mg· L-1敌百虫5个不同浓度的水体中,分别在24、48、72和96 h统计蝌蚪的死亡率,计算半致死浓度(LC50).结果显示,暴露24、48、72和96 h,敌百虫对蝌蚪的LC50分别为14250±3.23、49.19±128、25.68±2.04、1555±1.93 mg·L-1,安全浓度(SC)为156±0.19 mg· L-1.蝌蚪中毒后尾部多呈弯曲状,仰翻,外观浮肿.对死亡蝌蚪的解剖表明,其鳃腔内充水,内鳃萎缩,肝脏、肠管和肾脏呈灰白色.另外,将28～29期蝌蚪分别暴露于0.2 ～ 2.0 mg·L-1敌百虫4个不同浓度的水体中进行慢性暴露实验,检测蝌蚪暴露28和42 d时的体重和体长以及75％个体变态所需的时间.结果表明,蝌蚪在低剂量敌百虫水体中持续暴露,其生长发育受到明显抑制,并可导致蝌蚪身体扭曲、尾部强直性弯曲等畸型发生,蝌蚪的死亡率显著增高,作用强度呈现剂量和时间的累积效应.慢性暴露实验证明SC以下的敌百虫水体仍威胁着蝌蚪的生存.     

Effects of flow regimes altered by dams on survival, population declines, and range-wide losses of California river-breeding frogs

Widespread alteration of natural hydrologic patterns by large dams combined with peak demands for power and water delivery during summer months have resulted in frequent aseasonal flow pulses in rivers of western North America. Native species in these ecosystems have evolved with predictable annual flood-drought cycles; thus, their likelihood of persistence may decrease in response to disruption of the seasonal synchrony between stable low-flow conditions and reproduction. We evaluated whether altered flow regimes affected 2 native frogs in California and Oregon (U.S.A.) at 4 spatial and temporal extents. We examined changes in species distribution over approximately 50 years, current population density in 11 regulated and 16 unregulated rivers, temporal trends in abundance among populations occupying rivers with different hydrologic histories, and within-year patterns of survival relative to seasonal hydrology. The foothill yellow-legged frog (Rana boylii), which breeds only in flowing water, is more likely to be absent downstream of large dams than in free-flowing rivers, and breeding populations are on average 5 times smaller in regulated rivers than in unregulated rivers. Time series data (range = 8 - 19 years) from 5 populations of yellow-legged frogs and 2 populations of California red-legged frogs (R. draytonii) across a gradient of natural to highly artificial timing and magnitude of flooding indicate that variability of flows in spring and summer is strongly correlated with high mortality of early life stages and subsequent decreases in densities of adult females. Flow management that better mimics natural flow timing is likely to promote persistence of these species and others with similar phenology.     

双酚A对中国林蛙蝌蚪生长发育的毒性效应

为评价双酚A(BPA)对中国林蛙(Rana chensinensis)蝌蚪的急性毒性,将26期的蝌蚪暴露于浓度为2.4×10-5mol·L-1～4.2×10-5mol·L-1BPA的水体中进行急性毒性实验.结果表明,24、48、72、96h蝌蚪的死亡几率(y)与浓度对数(x)的回归方程分别为y=16.915x-4.1157、y=22.11x-6.1905、y=20.766x-5.3871、y=20.715x-5.351;半数致死浓度(LC50)分别为3.47×10-5、3.28×10-5、3.20×10-5、3.16×10-5mol·L-1.安全浓度(SC)为0.88×10-5mol·L-1.为探讨在安全浓度以下BPA对中国林蛙蝌蚪生长发育的影响,将林蛙蝌蚪分别于10-5、10-6、10-7mol·L-1BPA的水体中连续暴露直至完全变态,并设10-8、10-9mol·L-1雌二醇(E2)的阳性对照和空白对照,分别测定并统计蝌蚪发育所需的发育时间、体长和体质量.结果表明,蝌蚪对10-5、10-6、10-7mol·L-1BPA与10-8、10-9mol·L-1E2的效应相似,可延缓林蛙幼体的发育时间,导致体长和体质量降低.推测低浓度BPA抑制蝌蚪生长发育的机制之一是干扰了正常的内分泌活动.     

微囊藻毒素-LR对雄性黑斑蛙精巢中CYP46A1,CYP2H2和CYP2G1 mRNA表达的影响

微囊藻毒素-LR(microcystin-LR,MC-LR)是分布最广泛和毒性最强的一种微囊藻毒素,对水生动物造成潜在的健康威胁。大量科学研究证实,动物体中的细胞色素P450酶(CYP)参与内源性物质及外源毒性物质的代谢过程。为探究两栖动物生殖器官中的CYP酶对低剂量MC-LR生殖毒效应的调节作用,选择雄性黑斑蛙(Rana nigromaculata)为受试动物,采用静态置换法和体内暴露方法,分别暴露于0、0.1、1和10μg·L~(-1)MC-LR溶液0、7和14 d;用实时荧光定量PCR方法检测精巢中CYP46A1、CYP2H2和CYP2G1的mRNA表达水平。结果表明,暴露于0.1、1和10μg·L~(-1)MC-LR 14 d后,CYP46A1在mRNA水平分别上调了1.86、1.65和1.22倍,CYP2H2在mRNA水平分别上调了4.62、1.80和1.04倍,CYP2G1的mRNA水平分别上调了2.63、2.16和1.56倍。MC-LR在1μg·L~(-1)剂量下暴露7 d后,CYP46A1、CYP2H2和CYP2G1 mRNA水平均出现显著上调。上述研究表明,微囊藻毒素对黑斑蛙精巢3种CYP基因在mRNA水平上都存在低剂量刺激效应。低剂量MC-LR能诱导黑斑蛙精巢中CYP46A1转录水平变化,促进胆固醇转化为24S-羟化胆固醇,潜在破坏雄性黑斑蛙精巢中胆固醇水平的平衡; MC-LR也能够诱导精巢中CYP2H2和CYP2G1转录水平的变化,潜在调节CYP2H2和CYP2G1转录水平,进而影响MC-LR的代谢作用。     

氟离子慢性暴露对中国林蛙蝌蚪生长发育的毒性效应研究

氟元素以不同的结合形式广泛存在于自然界中,适量氟的摄取有益于龋齿预防和骨骼发育,然而过量氟的摄取会对动物及人体健康造成危害。近年来,大量的人类活动导致水环境中氟含量持续升高。为探究水域氟污染对中国林蛙的毒性影响,本研究以中国林蛙(Rana chensinensis)胚胎为试验材料,对卵黄栓期(G12期)胚胎进行了0、0.7、4.2、19.4、42.8 mg·L~(-1)F-慢性水体暴露直至胚胎发育到变态高峰期(G42期)的研究。分别于暴露25 d和40 d后取样测定了蝌蚪全长、体长、体重和发育分期;此外,分析了F-慢性暴露对变态率、G42期蝌蚪的全长、体长、体重和后肢长以及G42期蝌蚪骨骼发育的影响。结果表明:暴露25 d时,4.2 mg·L~(-1)F-处理组促进了林蛙蝌蚪的生长发育,而42.8 mg·L~(-1)F-处理组显著抑制了蝌蚪的生长发育;暴露40 d时,19.4 mg·L~(-1)F-和42.8 mg·L~(-1)F-处理组蝌蚪的生长发育均受到显著抑制。持续进行慢性暴露78 d后,4.2 mg·L~(-1)F-处理组蝌蚪的变态率显著升高,而42.8 mg·L~(-1)F-处理组蝌蚪的变态率受到了显著抑制。此外,42.8 mg·L~(-1)F-处理组G42期蝌蚪形态指标(全长、体长和后肢长)以及骨骼发育均受到抑制。依据G42期中国林蛙蝌蚪的生长发育指标和变态率为观察指标,氟离子慢性暴露对中国林蛙蝌蚪的最低可观察效应浓度(LOEC)为0.7 mg·L~(-1)。研究表明,水环境中高浓度的氟污染会对中国林蛙蝌蚪的生长发育、变态和骨骼发育等造成潜在的不利影响,水体氟污染的生态毒性效应理应引起高度重视。