茜素络合物对半滑舌鳎(Cynoglossus semilaevis)苗种耳石的染色标记效果

采用茜素络合物(Alizarin Complexone,ALC)分别浸泡标记70 d和170 d半滑舌鳎(Cynoglossus semilaevis)苗种耳石,设置了5个浓度组和3个时间梯度组。结果显示,ALC对半滑舌鳎70–170 d苗种的耳石染色效果明显。综合考虑存活率和标记效果,ALC浸泡标记半滑舌鳎苗种耳石的适宜浓度为100 mg/L(36 h浸泡处理)和150 mg/L(24 h浸泡处理)。相同浸泡时间下,随着ALC浓度的增加,耳石上标记信号强度增加,但实验鱼死亡率升高;相同浸泡浓度下,随着时间的增加,耳石上标记信号强度增加,实验鱼死亡率也会升高。对标记的170 d苗种进行养殖实验,每30 d取样矢耳石进行观察,发现4个月后实验鱼生长良好,且耳石荧光信号仍可清晰检测到,表明ALC在半滑舌鳎矢耳石上形成的荧光标记可长期存续,标记效果好。研究结果可为半滑舌鳎大规模标志放流及早期生活史研究提供群体身份识别技术支持。     

茜素络合物对鲤仔鱼耳石标记特征研究

利用100 mg/L的茜素络合物（alizarin complexone,ALC）对鲤仔鱼进行48 h的水环境浸泡标记,以探讨该ALC标记方法的特征,及其对矢耳石、星耳石和微耳石的标记效果以及鱼体ALC浸泡、续养恢复与耳石ALC标记区域形成和消失的时滞进行研究。结果显示,3种耳石在可见光和荧光下均能检测到明显的标记环。其中星耳石的标记效果最佳,微耳石次之。耳石上荧光信号出现和消失的时间与鱼体ALC浸泡开始和结束的时间均存在1 d的时滞。此外,浸泡标记过的鲤仔鱼在进行了长达50 d的续养恢复后,其耳石上的ALC标记环仍清晰可见。研究表明,ALC标记法所形成的标记环在耳石上可长期存在,使用ALC对鲤仔鱼进行生态标记具有很强的可行性。     

草鱼仔鱼耳石茜素络合物标记特征研究

草鱼(Ctenopharyngodon idellus)是我国主要增殖放流经济鱼种之一，探索有效的标志方法对其增殖放流的技术保障和效果评价无疑至关重要。本研究对草鱼仔鱼耳石进行了1 d茜素络合物(ALC)荧光标记，以确认该方法的可行性及适宜条件。结果显示，微耳石、矢耳石和星耳石均能被标记。标记环在可见光及荧光下均能被观察到。其中，后者使用蓝色激发光(WBS)观察效果最佳。总体上，微耳石的标记效果最好，矢耳石次之。ALC浸泡对鱼体的生存及生长无显著影响，耳石上荧光信号出现和消失与鱼体开始浸泡和结束均存在约12 h的时滞。浸泡标记过的实验鱼在经过长达60 d的恢复续养后，标记环仍清晰可见，表明ALC标记效果明显且可长期存在。该方法对草鱼仔鱼的增殖放流标记具有较强的可行性。     

荧光染料对短须裂腹鱼早期鱼苗的浸泡标记研究

为探究短须裂腹鱼(Schizothorax wangchiachii)早期鱼苗浸泡标记的合适荧光染料及标记条件,为短须裂腹鱼大规模标记放流及其效果评估提供参考,实验使用茜素络合物(AC)、茜素红S(ARS)、盐酸四环素(TC)、酸性茜素蓝B(AAB)、茜素黄GG(AYG)和钙黄绿素(CAL)溶液分别浸泡标记短须裂腹鱼早期鱼苗(仔、稚鱼),实验结束后摘取耳石检测荧光标记。结果显示:AC、ARS均可在短须裂腹鱼耳石上形成荧光标记,标记在绿光下最明显,呈橘红色;TC、AAB、AYG、CAL均未形成有效的荧光标记;AC标记造成的死亡率和标记质量随着溶液浓度、浸泡时间的增加而升高,随着鱼体全长的增加而降低;AC标记20日龄短须裂腹鱼结束16 d后,与空白对照组相比,除浓度300 mg/L、浸泡时间24 h的这组全长显著较低外,其他各组无显著差异;AC及ARS重复标记均可形成两个橘红色标记环,首次标记时AC造成的死亡率低于ARS,再次标记时前者高于后者。综合考虑标记质量、死亡率及对生长影响等多方面因素,本研究推荐使用AC或ARS浸泡标记短须裂腹鱼早期鱼苗,具体条件为:溶液浓度50～100 mg/L,浸泡时间12～24 h。     

茜素红S对异齿裂腹鱼稚鱼标记效果研究

为探索茜素红S对异齿裂腹鱼稚鱼浸泡标记的适宜条件,采用半静态式水生生物急性毒性试验法确定茜素红S对异齿裂腹鱼稚鱼安全质量浓度,利用不同质量浓度的茜素红S溶液对45日龄的异齿裂腹鱼稚鱼浸泡不同时间,探究各试验组稚鱼死亡率、生长情况、标记效果和标记有效保存时间,确定茜素红S对异齿裂腹鱼稚鱼的适宜标记条件。试验结果表明：茜素红S对异齿裂腹鱼稚鱼24、48、72、96 h半致死质量浓度分别为225.85、147.72、139.21、137.66 mg/L,安全质量浓度为28.99 mg/L;0～125 mg/L的茜素红S溶液浸泡稚鱼24 h,质量浓度为75～125 mg/L稚鱼耳石标记效果可见,当茜素红S质量浓度为125 mg/L时,稚鱼死亡率显著升高(P<0.05);75 mg/L茜素红S浸泡稚鱼12～96 h,浸泡24～96 h稚鱼耳石标记效果可见,当浸泡时间为96 h时,稚鱼死亡率显著升高(P<0.05);在4个月的续养过程中,75 mg/L茜素红S溶液浸泡72～84 h, 4个月后稚鱼耳石标记效果依旧可见。综合生长、存活率、染色效果及标记保存时间,建议采用75 mg/L茜素...     

茜素络合物浸泡标记秦岭细鳞鲑发眼卵及仔鱼耳石

采用茜素络合物(ALC)在秦岭细鳞鲑(Brachymystax lenok tsinlingensis)发眼卵和仔鱼阶段进行浸泡标记试验,为确定合适浸泡浓度和持续浸泡时间,试验设置了6个的浓度组和4个时间梯度组.结果显示:在荧光显微镜下观察被浸泡的仔鱼和经浸泡处理的发眼卵而出膜后的仔鱼,其耳石都出现橘红色荧光标记.浸泡...     

茜素络合物在青鱼幼鱼耳石上的标记效果评价

放流标记技术研究是支撑增殖放流效果评估的重要内容。该研究用质量浓度为100 mg·L-1的茜素络合物(ALC)对约2月龄的青鱼(Mylopharyngodon piceus)幼鱼进行48 h浸泡,随后移入正常水体中恢复续养20 d。对其3种耳石在可见光和不同的激发光下进行比较分析,评价了茜素络合物在不同耳石上的沉积情况与标记效果。结果表明,通过在可见光和不同的激发光下进行观察,其3种耳石上均可见明显的标记区,标记率达100%。在对标记鱼恢复续养20 d后发现,耳石上的荧光标记区强度仍很高,表现出了较好的标记效果。此外,相关标记的最佳检测光源为蓝色激发光,绿色激发光次之。微耳石是用于青鱼幼鱼耳石荧光标记观察的最适类型,同时也是用于时滞检测的最适类型,其次是星耳石。此外,将鱼体浸泡入茜素络合物溶液中至其在耳石上形成标记沉积,以及将鱼体移出浸泡溶液至其耳石上标记区域消失,均存在1 d时滞。     

淡水中心在淡水鱼增殖放流标记技术研究上取得新进展

<正>目前，有效的标记手段和增殖放流评价方法，仍是制约增殖放流应用工作的“瓶颈”问题。淡水渔业研究中心渔业微化学实验室选取鲢鱼和鲫鱼，通过鱼体茜素络合物（ALC）荧光标记和锶元素浸泡标记，探索相关标记方法在鱼类多类硬组织标记和效果评价上的适用性特征和优化途径。研究发现，ALC不仅能够成功标记鲢鱼幼鱼广受关注的耳石（矢耳石、星耳石、微耳石），     

锶元素对黑鲷幼鱼耳石的标记效果分析

通过开展不同锶浓度养殖水体的黑鲷(Sparus macrocephalus)幼鱼耳石浸泡标记实验,验证利用该方法开展黑鲷幼鱼耳石指纹标记的可行性。研究结果显示:经7 d的浸泡标记,4个处理组幼鱼在特定耳石区位的Sr/Ca值均显著增加,形成了明显的耳石元素指纹标记;各标记处理组的标记成功率均为100%。单因素方差分析结果显示,各处理组和对照组黑鲷幼鱼在标记实验期间的生长率和死亡率均无显著性差异,表明该标记方法不会对受试黑鲷幼鱼产生显著负影响。浸泡标记处理可使受试幼鱼肌体内的锶浓度明显提升;标记结束17 d后,各处理组幼鱼肌体内的锶浓度大幅下降,18 mg·L~(-1)标记组幼鱼肌体内的锶浓度可降至对照个体水平。     

茜素络合物对鳙耳石的标志效果研究

利用不同浓度梯度(50~150mg/L)的茜素络合物(ALC)水溶液对20日龄的鳙幼鱼浸泡8h进行标志。结果显示,ALC溶液浓度不高于100mg/L时,鳙幼鱼存活率为100%,150mg/L浓度下幼鱼全部死亡。茜素络合物在其耳石上能形成标记环带,随着ALC溶液浓度的增加,在可见光下标记色越来越明显,50mg/L ALC溶液中幼鱼耳石略见淡红色标记,75mg/L浓度组有清晰的淡红色标记环,100mg/L ALC溶液中幼鱼标记环带明显,鳙幼鱼最适的ALC标记浓度为100mg/L。     

Mass-marking of coregonid larvae and fry by tetracycline tagging of otoliths

Abstract. Larvae and juveniles of two species of Coregouns sp. were immersed in tetracycline (TC) solutions (200–800)mg/1) and fluorescent marks in the otoliths from fish were identified under a UV-light microscope. Whitefish. Coregonus Iavaretus (L.). juveniles (average weight 191 mg) immersed in TC solution retained their marking until the 127th day, hut 40% of the population lost their TC marking after wintering. Whitefish and peled. C. peled Gmelin, larvae retained TC marking from the TC solution (400 mg/1) if water temperature was higher than 5°C, Whitefish juveniles retained clear marking in the otolith after 120 days of post-immersion rearing. Peled larvae immersed in TC solution were grown in a pond and the fluorescent otolith centre of 55·4-g fish was well visible after processing. Nearly 100% of peled larvae grown for 176 days can he identified based on fluoresceent marks observed under a UV-lighl microscope.     

耳石锶标记识别鮻放流群体的可行性研究

为探究耳石锶(Sr2+)标记技术在鮻(Liza haematocheila)增殖放流群体识别的可行性，以体长5 cm左右的鮻幼鱼为研究对象，将其养殖海水中Sr2+的质量浓度分别调至50、100、200和400 mg/L，以自然海水(Sr2+质量浓度为7.9 mg/L)为对照组，人为改变鮻幼鱼生存环境中Sr2+的质量浓度标记48 h。同时开展双环标记实验(在鮻3 cm幼鱼时进行Sr2+质量浓度200 mg/L海水96 h标记，待其长至5 cm幼鱼时进行Sr2+质量浓度100 mg/L海水48 h标记)。实验结果显示，Sr2+质量浓度50~400 mg/L的海水48 h标记的鮻耳石Sr/Ca比均出现一个显著的上升阶段且面分析出现明显的“高Sr2+标记环”。综合本研究结果和标记效率，推荐采用Sr2+质量浓度为50 mg/L的海水进行鮻耳石Sr2+标记。双环标记实验鮻耳石Sr2+元素面分布结果出现层次分明的双环，且双环标记组鮻耳石2条不同标记环的线性分析结果与其相对应的单环标记组基本一致，这表明通过Sr2+标记次数来进行不同放流群体识别是可行的。     

The use of alizarin complexone for immersion marking of the otoliths of embryos and larvae of the turbot, Scophthalmus maximus (L.): dosage and treatment time

The purpose of this work was to determine the efficacy of marking turbot, Scophthalmus maximus (L.), eggs and larvae with alizarin complexone (ALC) for use in enhancement programmes and in studies of the ecology and behaviour of the species. The main aim was to determine the optimum dosage and treatment time for large batches of larvae destined for release into the sea.Eggs and larvae were immersed in a range of ALC concentrations (0–60 mg L^–1) for various times (3–24 h). On day 8 after hatching, eggs marked with the different ALC dosages had 99% marking success and had mainly good-quality or very-good-quality marks, whereas larvae achieved 100% success and had mainly very-good-quality marks. The best results were obtained after marking larvae with ALC at 60 mg L^–1 for 6–24 h.During the first months after marking, the fluorescent rings formed in the otoliths were easily detected when directly examined with a UV microscope, no prior preparation of the otoliths was necessary, but after 7 months the marks become faint as the otoliths get thicker, so it is necessary to cut or polish the otoliths before examination.     

Validation of daily increment formation in otoliths of immature and adult Japanese anchovy <Emphasis Type="Italic">Engraulis japonicus</Emphasis>

In order to validate daily increment formation in otoliths of immature and adult Japanese anchovy Engraulis japonicus, three rearing experiments using chemical marking of otoliths were conducted on adult anchovy in summer 2004 and immature anchovy in summer 2005 and in winter 2006. In the two experiments conducted in summer, the number of otolith microincrements between alizarin complexone (ALC) marks showed that microincrements were formed daily. In the summer 2005 experiment, immature anchovy under conditions of reduced daily food rations also showed daily microincrement formation. Average increment width was 0.9 μm in adults and 1.8–3.1 μm in immature anchovy. In contrast, no clear increments were observed between ALC marks on the otoliths from the experiment in winter 2006, and scanning electron microscope (SEM) observations failed to confirm clear increment formation. We consider that low water temperatures (<13–14°C) restricted otolith growth and lowered the contrast between the discontinuous and the incremental zones of the otolith increments. For age estimation of Japanese anchovy, clear increments wider than about 1 μm in the otolith can be regarded as daily increments. However, daily age estimation of immature and adult anchovy that experience low water temperatures in winter may be difficult due to the obscurity of the increments.     

Identification of Atlantic bluefin tuna (Thunnus thynnus) stocks from putative nurseries using otolith chemistry

Chemical signatures in the otoliths of teleost fishes represent natural tags that may reflect differences in the chemical and physical characteristics of an individuals' environment. Otolith chemistry of Atlantic bluefin tuna (Thunnus thynnus) was quantified to assess the feasibility of using these natural tags to discriminate juveniles (age 0 and age 1) from putative nurseries. A suite of six elements (Li, Mg, Ca, Mn, Sr and Ba) was measured in whole otoliths using solution‐based inductively coupled plasma mass spectrometry. Otolith chemistry of age‐1 T. thynnus collected from the two primary nurseries in the Mediterranean Sea and western Atlantic Ocean differed significantly, with a cross‐validated classification accuracy of 85%. Spatial and temporal variation in otolith chemistry was evaluated for age‐0 T. thynnus collected from three nurseries within the Mediterranean Sea: Alboran Sea (Spain), Ligurian Sea (northern Italy), and Tyrrhenian Sea (southern Italy). Distinct differences in otolith chemistry were detected among Mediterranean nurseries and classification accuracies ranged from 62 to 80%. Interannual trends in otolith chemistry were observed between year classes of age‐0 T. thynnus in the Alboran Sea; however, no differences were detected between year classes in the Tyrrhenian Sea. Age‐0 and age‐1 T. thynnus collected from the same region (Ligurian Sea) were also compared and distinct differences in otolith chemistry were observed, indicating ontogenetic shifts in habitat or elemental discrimination. Findings suggest that otolith chemistry of juvenile T. thynnus from different nurseries are distinct and chemical signatures show some degree of temporal persistence, indicating the technique has considerable potential for use in future assessments of population connectivity and stock structure of T. thynnus.     

基于浸泡法的牙鲆耳石锶标记技术研究

为了探究耳石微化学标记技术在牙鲆增殖放流中的应用,以全长(1.56±0.16)cm的牙鲆幼鱼为研究对象,在其养殖水中分别添加0.5 mg/L、2 mg/L、8 mg/L、32 mg/L SrCl_2·6H_2O,以无添加SrCl_2·6H_2O组为对照组,人为改变牙鲆幼鱼生存环境中Sr~(2+)浓度72 h,研究不同浓度SrCl_2·6H_2O对牙鲆耳石锶标记效果。标记期间每24 h换水1次,换水量为50%,换水后补加SrCl_2·6H_2O至原始浓度,记录各浓度组标记期间及标记后的鱼苗死亡数量。利用X射线电子探针微区分析仪(EPMA)对耳石样本进行分析,定量线分析结果表明, 0.5 mg/L、2 mg/L SrCl_2·6H_2O浸染不能形成牙鲆耳石锶标记。8 mg/L浓度组出现Sr峰区, Sr/Ca (Sr:Ca×1000)均值为4.50～6.60, Sr峰值为5.49～8.49。32 mg/L浓度组Sr峰区均值为4.83～7.55, Sr峰值为5.62～11.04。这说明8 mg/L、32 mg/L SrCl_2·6H_2O可对牙鲆耳石进行锶标记。但8 mg/L浓度组出现未标记样本,且Sr峰区均值较32 mg/L浓度组低。32 mg/L浓度组Sr/Ca比值显著增高,即锶标记的Sr/Ca比值为6.61±0.86,与标记前和对照组Sr/Ca比值差异均显著(P0.05),锶标记率100%。面分布分析(mapping analysis)结果表明,从8 mg/L SrCl_2·6H_2O浓度组开始,耳石出现红色"高锶标记环",与耳石本底颜色对比明显,且随浓度升高,红色"高锶标记环"由浅变深,锶标记效果明显增强。单因素方差分析和Duncan多重比较结果显示,各浓度组全长、体重与对照组差异不显著(P0.05),不同浓度的SrCl_2·6H_2O对牙鲆幼鱼生长无影响。经χ2检验,各浓度组在浸泡标记期间死亡率及标记后养殖90 d的累积死亡率无显著性差异(P0.05),不同浓度的SrCl_2·6H_2O对牙鲆幼鱼死亡率无显著影响。因此, 32 mg/L SrCl_2·6H_2O为最佳标记浓度。     

Alizarin marking of whitefish, Coregonus lavaretus otoliths during egg incubation

Alizarin red S from 200 to 5000 mg L^?1 was used to mark the otoliths of whitefish embryos Coregonus lavaretus (L.) from 5 days after fertilization until shortly before hatching, to develop a method for evaluating the effect of stocking whitefish. Mark quality depended on developmental stage at the time of immersion. The best results were obtained when the otolith primordia had formed and had started aggregating to form the innermost part of the nucleus. This was verified by in vitro staining of otoliths where the primordia were more intensely stained than the remaining part of the nucleus. The volume ratio eggs to ARS solution, which was 1:10 in all but two treatments, must be observed when comparing fluorochrome concentrations, as mark quality decreased at higher ratios of 1:5 and 1:2. An average mortality rate of 35% in the most successful treatments was considered tolerable for a mass‐marking programme being used under commercial hatchery conditions. This method for marking otoliths of whitefish embryos with ARS during egg incubation is recommended for all cases where marking of larvae is not possible.