Cypermethrin induces glutathione S-transferase activity in the shore crab,Carcinus maenas

Cypermethrin is a synthetic pyrethroid that is particularly toxic to crustacea. It is therefore applied as a chemotherapeutant in the salmonid aquaculture industry for the treatment of sea lice infestations. After use, cypermethrin is released directly into the marine environment, to be diluted by fresh seawater. The shore crab, Carcinus maenas is found in the vicinity of fish farms, and may come into contact with released cypermethrin. The detoxification enzyme, glutathione S-transferase (GST) has been implicated in cypermethrin metabolism in terrestrial arthropods, but this has not yet been demonstrated in crustacea. In this paper we investigate the response of GST activity in Carcinus to cypermethrin exposure, and also the time course of the induction process. GST activity is significantly increased in Carcinus exposed to nominal concentrations of 50 and 500 ng/l of water-borne cypermethrin. Carcinus demonstrate a significant elevation in GST activity following intra-cephalothoracic injection with 10 ng of cypermethrin. GST activity returns to basal levels after 36 h. The potential application of GST activity in Carcinus as a biomarker of cypermethrin exposure is discussed.     

Effects of beta-naphthoflavone on hepatic biotransformation and glutathione biosynthesis in largemouth bass (Micropterus salmoides)

We are investigating the effects of in vivo exposure of prototypical enzyme inducing agents on hepatic biotransformation enzyme expression in largemouth bass (Micropterus salmoides), a predatory game fish found throughout the United States and Canada. The current study targeted those genes involved in biotransformation and oxidative stress that may be regulated by Ah-receptor-dependent pathways. Exposure of bass to beta-naphthoflavone (beta-NF, 66 mg/kg, i.p.) elicited a 7-9-fold increase in hepatic microsomal cytochrome P4501A-dependent ethoxyresorufin O-deethylase (EROD) activities, but did not affect cytosolic GST catalytic activities toward 1-chloro-2,4-dinitrobenzene (CDNB) or 5-androstene-3,17-dione (ADI). Glutathione S-transferase A (GST-A) mRNA expression exhibited a transient, but non-significant increase following exposure to beta-NF, and generally tracked the minimal changes observed in GST-CDNB activities. Expression of the mRNA encoding glutamate-cysteine ligase catalytic subunit (GCLC), the rate-limiting enzyme in glutathione (GSH) biosynthesis, was increased 1.7-fold by beta-NF. Changes in GCLC mRNA expression were paralleled by increases in intracellular GSH. In summary, largemouth bass hepatic CYP1A-dependent and GSH biosynthetic pathways, and to a lesser extent GST, are responsive to exposure to beta-NF.     

苯并(a)芘、芘及其混合物暴露对梭鱼肝脏谷胱甘肽硫转移酶活性的影响

在实验生态条件下,观察苯并(a)芘、芘及其等浓度混合物暴露对梭鱼(Mugil so-iuy)肝脏谷胱甘肽硫转移酶(GST)活性的影响。结果显示,在7d的暴露中,苯并(a)芘、芘对肝脏GST活性的影响主要为诱导效应,芘对GST活性的诱导比苯并(a)芘强。混合物在15d的暴露中未观察到GST活性的诱导,而是在暴露的后期出现GST活性的抑制。实验表明,肝脏GST活性的诱导指示受到PAHs污染胁迫,而GST活性抑制则是受到较长时间或较严重的污染。     

菲、芘、苯并(a)芘单一暴露及分别与α-萘黄酮(ANF)联合暴露对海水青鳉(Oryzias melastigma)胚胎发育毒性效应的比较研究

为了解和探讨3~5环PAHs对海水鱼类胚胎发育的毒性及作用方式,比较研究了菲(phenanthrene,Phe)、芘(pyrene,Py)、苯并(a)芘(benzo(a)pyrene,BaP)单一暴露和三者各自与α-萘黄酮(α-naphthoflavone,ANF)联合暴露对海水青鳉(marine medaka, Oryzias melastigma)胚胎发育的毒性效应。胚胎体内EROD活性、发育畸形、孵化率和心律等毒性指标被测定,结果显示:Phe,Py和BaP对海水青鳉胚胎体内EROD活性的诱导能力大小为BaP>Py>Phe,各化合物对EROD诱导与发育畸形之间的关系较为复杂,除Phe所引起的EROD诱导与畸形指数之间呈显著相关(r=0.95,p=0.015)外,Py和BaP均无相关性;在100 μg/dm3 ANF影响下,CYP1A活性诱导被抑制,但胚胎发育的畸形指数被显著提高,ANF分别与Phe,Py和BaP的联合暴露对胚胎发育呈潜在的协同作用。本文研究初步表明,3~5环PAHs化合物对海水青鳉胚胎发育的毒性作用方式可能不同;CYP1A活性抑制在PAHs混合物对海水青鳉胚胎发育的毒性作用过程中未起到缓解毒性的作用,CYP1A抑制剂与PAH型CYP1A诱导剂的混合物对鱼类胚胎发育具有潜在的协同毒性作用,现有的PAHs混合物毒性风险评价方法可能低估了实际环境中PAHs的风险;海水青鳉早期生活阶段的心脏发育对PAHs混合物暴露较为敏感,可推荐其作为生物标志物指示PAHs或溢油污染。     

黄渤海海域贝类麻痹性贝毒的检测与分析

采用小白鼠生物测试法和高效液相色谱法对2002~2005年我国黄渤海海域采集的贝类样品进行了麻痹性贝毒毒性检测,结果显示大连海域的虾夷扇贝含有麻痹性贝毒,有毒样品均出现在5月和6月,部分虾夷扇贝样品的毒素含量已经超过食用安全标准。通过高效液相色谱法分析了有毒虾夷扇贝体内的毒素成分,共检出了6种麻痹性贝毒组分,主要以毒性较低的C1和C2毒素为主,GTX3和GTX2次之,STX和neoSTX含量很低。通过高效液相色谱法分析得到的各毒素组分毒性总和与小白鼠生物测试法毒性测试结果基本相当。     

有毒赤潮藻及其毒素的危害与检测

海洋中可引发赤潮的藻类约有300种,其中有毒赤潮藻为80种左右。现已知道的赤潮藻主要毒素有麻痹性贝毒、腹泻性贝毒、记忆缺失性贝毒、神经性贝毒、西加鱼度和溶血性毒素,前5种毒素的结构已经基本得到证实。有毒赤潮藻的毒素可以在海洋生物体内积累,人类误食含有藻毒素的食品时可能中毒,严重者还可能死亡。海洋有毒赤潮藻及其毒素的检测已经成为当今全球赤潮研究和监测的重要内容之一,可以通过形态学分类方法、分子生物学技术(遗传探针)和免疫学检测技术对有毒赤潮藻进行检测;可以通过生物学、物理化学检测方法和神经受体结合、免疫学检测技术对赤潮藻毒素进行检测。     

Effects of β-naphthoflavone on hepatic biotransformation and glutathione biosynthesis in largemouth bass (Micropterus salmoides)

We are investigating the effects of in vivo exposure of prototypical enzyme inducing agents on hepatic biotransformation enzyme expression in largemouth bass (Micropterus salmoides), a predatory game fish found throughout the United States and Canada. The current study targeted those genes involved in biotransformation and oxidative stress that may be regulated by Ah-receptor-dependent pathways. Exposure of bass to β-naphthoflavone (β-NF, 66 mg/kg, i.p.) elicited a 7–9-fold increase in hepatic microsomal cytochrome P4501A-dependent ethoxyresorufin O-deethylase (EROD) activities, but did not affect cytosolic GST catalytic activities toward 1-chloro-2,4-dinitrobenzene (CDNB) or 5-androstene-3,17-dione (ADI). Glutathione S-transferase A (GST-A) mRNA expression exhibited a transient, but non-significant increase following exposure to β-NF, and generally tracked the minimal changes observed in GST–CDNB activities. Expression of the mRNA encoding glutamate-cysteine ligase catalytic subunit (GCLC), the rate-limiting enzyme in glutathione (GSH) biosynthesis, was increased 1.7-fold by β-NF. Changes in GCLC mRNA expression were paralleled by increases in intracellular GSH. In summary, largemouth bass hepatic CYP1A-dependent and GSH biosynthetic pathways, and to a lesser extent GST, are responsive to exposure to β-NF.     

Source and profile of paralytic shellfish poisoning toxins in shellfish in Daya Bay, South China Sea

Changes in cell density and cyst flux of Alexandrium tamarense, paralytic shellfish poisoning (PSP) toxin contents in shellfishes, and environmental parameters were measured in two stations in Daya Bay, South China Sea from March 2005 to July 2006. Vegetative cells of A. tamarense occurred sporadically; however, they presented abundantly during the winter months. Meanwhile, cyst flux reached its maximum level just following the peak abundance of motile cells. The PSP contents in shellfish were generally low, but higher in winter with the maximum of 14,015 μg STX equiv./kg. The majority of toxins were found in digestive glands, with a maximum of 66,227 μg STX equiv./kg. There were significant positive relationships between toxin level and vegetative cell density and cyst flux. This indicates that vegetative cells and cysts of Alexandrium significantly influenced PSP level, and could be an important source of PSP toxins in shellfish during winter.     

麻痹性贝毒毒素的应用研究进展

由于有害赤潮的发生日趋频繁，对养殖业、自然生态系统和人类健康的危害越来越大，造成的经济损失也逐渐增加，因而有害赤潮问题成为人们关注的焦点。藻毒素是有害赤潮致害的重要因子之一，而麻痹性贝毒(Paralytic Shellfish Poisoning，PSP)毒素又是藻毒素中分布最广、危害最大的一类毒素。随着科学家们对PSP毒素的来源(Steidinger，1993)、结构和作用方式(Penzotti et al.,1998)的深入研究，PSP毒素在赤潮研究、分子生物学和神经生物学基础研究、医药、军事防化等的应用也逐渐受到重视。本文作者针对这些问题进行了综合评述，以期为深入开展PSP毒素应用研究提供参考依据。     

Assessment of cytochrome P450 1A in harbour seals (Phoca vitulina) using a minimally-invasive biopsy approach

Biomarkers of organochlorine exposure, such as the induction of cytochrome P450 1A (CYP1A), can be used to assess the impact of environmental contaminants on the health of free-ranging marine mammal populations. The objective of the present study was to measure CYP1A in skin and liver biopsies obtained from live harbour seals (Phoca vitulina). Twelve harbour seal pups, aged three to five weeks, were captured from the Fraser River estuary, British Columbia, Canada, and temporarily held in captivity. Skin ( approximately 60 mg) and liver ( approximately 40 mg) biopsies, obtained while seals were under general anaesthesia, yielded sufficient tissue for the measurement of CYP1A by immunoblot analysis and ethoxyresorufin O-deethylase activity. A short-term exposure experiment, in which harbour seals (n=3) were treated orally with beta-naphthoflavone (BNF), resulted in increased hepatic and cutaneous CYP1A protein levels, consistent with observations in other mammals. This study is the first to measure CYP1A in skin and liver biopsies from live harbour seals and to report in vivo BNF-associated CYP1A induction in a marine mammal. The results demonstrate that microsamples collected using minimally-invasive techniques can provide toxicologically-relevant information form marine mammals.     

Comparison of short-term toxicity of 14 common phycotoxins (alone and in combination) to the survival of brine shrimp Artemia salina

Toxic harmful algal blooms(HABs) can cause deleterious effects in marine organisms, threatening the stability of marine ecosystems. It is well known that different strains, natural populations and growth conditions of the same toxic algal species may lead to different amount of phycotoxin production and the ensuing toxicity. To fully assess the ecological risk of toxic HABs, it is of great importance to investigate the toxic effects of phycotoxins in marine organisms. In this study, the short-te...     

栉孔扇贝体内麻痹性贝毒的累积与排出过程研究

通过室内摄食实验,研究了栉孔扇贝(Chlamys farreri)对麻痹性贝毒(PSP)产毒藻的敏感性及其累积和排出毒素的特征.有毒微小亚历山大藻(Alexandrium minutum)在短时间内可对栉孔扇贝的摄食有一定的抑制作用,且投喂毒藻的浓度越大,对摄食作用的抑制越明显.栉孔扇贝对PSP贝毒的累积能力很强,实验中,48 h后内脏累积毒素就高达5000μg STXeq100g-1,但其毒素排出速率较慢,在24d解毒阶段后内脏中的毒素还没有下降至国际贝毒食用安全标准水平的80μg STXeq 100g-1.各毒素组成比在整个累积与排除过程中有较大变化,GTX1,GTX4在累积阶段比例下降,同时GTX2,GTX3比例上升;内脏中GTX2:GTX3的比值逐渐增加.毒藻中PSP贝毒毒素组成和贝体中PSP贝毒毒素组成之间的主要差别是GTX4含量的减少和GTX1含量的升高;而扇贝粪便中的毒素组成却与对数生长期的毒藻极为接近.     

镉胁迫条件下大弹涂鱼(Boleophthalmus pectinirostris)外周血微核标记及肝脏过氧化物酶标记的变化

采用静态毒性实验方法,研究重金属镉胁迫条件下大弹涂鱼外周血微核率和肝脏过氧化物酶活性的变化。结果表明,在0.05mg/L Cd2 浓度组,10d时检测到微核率极显著升高;0.5mg/L和5mg/L组5d时就可检测到微核率极显著升高,到10d时微核率达到最高值;10d时转入清洁海水后,3个组在15d再次出现一个微核率的最高值。大弹涂鱼肝脏过氧化物酶标记包括谷胱甘肽过氧化物酶(GSH-Px)、谷胱甘肽转硫酶(GST)。3个组的GSH-Px酶活性和5mg/L浓度组GST酶活性在12h后显示出极显著变化,3个组GST酶活性在24h检测到极显著差异;在转入清洁海水5d时,3个组GSH-Px酶活性和对照组相比可检测到极显著差异,而3个组的GST酶活性均无显著差异。研究结果表明,大弹涂鱼外周血微核标记和肝脏过氧化物酶标记能够灵敏地指示水环境中的镉污染;大弹涂鱼外周血微核标记和肝脏内过氧化物酶标记具有一定的互补作用。     

Study on impact of dinoflagellate Alexandrium tamarense on life activities of marine bivalves

The effects of a PSP producing dinoflagellate Alexandrium tamarense on marine bivalvesat their several important life stages: egg, D- shape larva, eyespot larva, juvenile and adult, were studied. The results show that the hatching, survival, activity, filtration and growth were adversely affected by the alga and the impact was significantly increased with the increase of algal density. The inhibitory effect on egg hatching was most significant, which the hatching rate was only 30% of the control when exposed to the alga at 100 cell/cm3 after 36 h. Further experiments show that the algal culture, re-suspended cells and cell fragments had the inhibitory effect, while no such effect was from the cell-free medium, cell contents and standard STX. The results indicate that the alga could produce unknown toxins, rather than PSP, associated with the cell surface.     

塔玛亚历山大藻对海产双壳类生命活动的影响

通过一种重要有毒赤潮藻——产麻痹性贝毒毒素PSP的塔玛亚历山大藻对几种海产双壳类主要发育阶段:受精卵、D形幼虫、眼点幼虫、稚贝、仔贝和成贝生命活动的影响研究,发现该藻对双壳类孵化率、存活率、运动能力、滤食率和生长都有影响,毒性效应与藻密度有明显的相关关系.该藻对受精卵的抑制作用在100个/cm3的密度下最显著,36h后孵化率仅为对照组的30%.进一步的逐项毒性筛选实验表明此毒性来源于藻液、藻细胞和细胞碎片,而与去藻过滤液、藻细胞内容物、标准毒素STX无关,由此可以证实塔玛亚历山大藻确能产生一类非PSP毒素,并与细胞表面的未知毒素相关.     

Tissue slice technology for assessing alterations in fish hepatic phase I and phase II XME activity

Alterations to hepatic xenobiotic metabolizing enzymes (XMEs) is an important biomarker of contaminant exposure in aquatic toxicology. Measurement of XMEs in fish liver slices in vitro is an emerging tool for examining enzyme activity and response within the intact 3-D architecture of the liver tissue. We examined integrated phase I/phase II, and phase II metabolism of XMEs from liver slices in control and B[a]P-treated rainbow trout and channel catfish. Fluorescent assay substrates to measure rates of metabolism included 7-methoxycoumarin (7-MC), 7-ethoxycoumarin (7-EC) and 7-hydroxycoumarin (7-HC). Time-dependent increases in metabolism, and a lower rate of 7-MC metabolism compared with 7-EC metabolism, were observed at all time points for both fish species. In rainbow trout, B[a]P pretreatment caused a 10-fold increase in phase I metabolism of both 7-MC and 7-EC, and a 1.6-fold increase in phase II metabolism of 7-HC. Phase I activity in channel catfish was not notably altered by B[a]P pretreatment. However, B[a]P pretreatment in channel catfish caused a 48% decrease in phase II metabolism of 7-HC. These results indicate differences in baseline and B[a]P-altered XME profiles between rainbow trout and channel catfish.     

Estrogenic and CYP1A response of mummichogs and sunshine bass to sewage effluent

Recent studies demonstrating feminization of effluent-exposed wild-caught male fish in the UK have prompted much research regarding the estrogenic activity of effluent from municipal sewage treatment plants (MSTPs). To investigate the estrogenicity and cytochrome P450 1A (CYP1A) induction potency of MSTP effluent, two species of fish, adult male mummichogs, Fundulus heteroclitus, and juvenile sunshine bass, Morone saxatilis x Morone chrysops, were exposed to un-chlorinated effluent (75% effluent, 25% seawater) from a large MSTP in Yonkers, NY, USA. After a 21-day static-daily (75%) renewal exposure, significant elevations over controls were observed in levels of vitellogenin (VtG) in plasma (1730%) and liver (131%) in effluent-exposed sunshine bass. In contrast, hepatic VtG was not elevated in mummichogs; plasma VtG was not measured in this species. Effluent exposure elevated hepatic CYP1A protein (140-145%) and ethoxyresorufin-O-deethylase (EROD) activity (408-598%) in both species. These findings suggest ontogenetic and/or species differences in response to estrogenic compounds in MSTP effluent. Furthermore, the elevation of CYP1A in response to sewage effluent exposure indicates the presence of additional compounds that may alter xenobiotic and/or steroid biotransformation in fish.     

Biomarker induction in tropical fish species on the Northwest Shelf of Australia by produced formation water

Normal operation of oil well platforms results in the discharge of produced formation water (PFW). The expression of CYP1A, CYP2M1- and 2K1-like proteins was examined for use as possible biomarkers of PFW exposure. A pilot study on the Northwest Shelf of Australia had indicated that PFW contamination possibly contributes to induction of CYP1A-like proteins in Gold-Spotted Trevally (Carangoides fulvoguttatus). The pilot study samples were re-examined for CYP1A, and, in addition, CYP2K1/2M1-like proteins. In a subsequent caged fish study in the same location a second species, Stripey seaperch (Lutjanus carponotatus), caught at a clean site, were distributed to three caging sites in a PFW gradient from the Harriet A production platform: A (near-field), B (far-field) and C (a non-impacted reference site). Fish were sampled at time (T) T = 0, T = 3 and T = 10 days. Significant increases of CYP1A, one CYP2K1- and two CYP2M1-like proteins were noted at Site A at T = 10d. For another CYP2K1-like protein, a significant increase was observed at Site A only at T = 3d. These results support a previous study indicating that CYP1A protein is sensitive to PFW exposure. Importantly, statistically significant environmental induction of both CYP2M1- and CYP2K1-like proteins in tropical fish due to PFW exposure had not previously been described and induction of enzymes in the CYP2 family suggest new biomarkers for PFW. In addition, the novel response of one CYP2K-like protein requires further verification, but offers promise for improved monitoring of sub-lethal responses in marine organisms.