Feeding resumption, morphological changes and mortality during starvation in Japanese flounder larvae

Japanese flounder Paralichthys olivaceus larvae established first feeding 3 days after hatching (DAH) at c . 17° C. Non-fed fish reached irreversible starvation at age 5 DAH. Non-fed fish showed similar feeding rate and feeding intensity as the fed fish when they were provided with prey before 5 DAH, after which the starved larvae did not feed even when prey became available. None of the six morphological measurements examined (total length, body height, eye height, head height, gut height and myotome height) showed significant differences between the non-fed and fed larvae until 5 DAH. Normal development continued only in the fed group, and the non-fed larvae showed reverse growth or body collapse after 5 DAH. Owing to the shrinkage and collapse at the top of head due to starvation, head height could be a sensitive indicator of starvation in Japanese flounder larvae. In the fed treatments, high mortality occurred from first feeding (3 DAH) to irreversible starvation (5 DAH), accounting for about two-thirds to three-quarters of the overall mortality (46–52%) throughout the experiments. This mortality was not prey density or larval density dependent. Mortality during the same period in the non-fed larvae accounted for about a third of the overall mortality (100%).     

牙鲆变态过程中的细胞凋亡

利用整体的原位TUNEL方法检测了牙鲆(Paralichthysolivaceus)变态过程中身体各器官细胞凋亡的分布及变化情况。结果如下:(1)与眼睛移动相关的脑颅骨骼的细胞凋亡右侧眼睛移动开始之后,在额骨、中筛软骨和犁骨软骨中出现细胞凋亡,并保持到眼睛移动结束;(2)中枢神经和感觉器官的细胞凋亡在眼睛移动开始之前,脊髓和脊髓鞘出现细胞凋亡,在眼睛移动开始之后,脊髓和脊髓鞘细胞凋亡停止,而在脑、眼睛和内耳出现细胞凋亡,并一直持续到眼睛移动结束;(3)与游泳、捕食和消化等功能相关的器官的细胞凋亡在眼睛移动开始后,冠状幼鳍的基部出现凋亡;在变态中后期,尾鳍基部出现细胞凋亡;下颌骨、鳃弓以及肝脏在眼睛移动开始之后,出现细胞凋亡,也一直持续到眼睛移动结束。细胞凋亡通过有序地去除多余的细胞来参与器官形态建立和重组,本研究的结果表明,在牙鲆器官功能变化过程中,细胞凋亡在与其相适应的的器官形态重塑中起着重要作用[动物学报52(2):355-361,2006]。     

Isolation and characterization of new microsatellite markers from the Japanese flounder (Paralichthys olivaceus)

The isolation and characterization of eight polymorphic microsatellite loci from a Japanese flounder partial genomic library are reported. The eight markers isolated in this study were highly polymorphic and their positions on the linkage genome map of the Japanese flounder were determined. Therefore, they are useful for ecological studies of wild populations. These markers are more effective than other markers with no information of chromosomal locations.     

几种因素对牙鲆胚胎玻璃化冷冻保存的影响

鱼类胚胎冷冻保存技术还远没有成熟, 为了寻找最佳的鱼类胚胎玻璃化冷冻保存条件, 我们以牙鲆(Paralichthys olivaceus) 胚胎为例, 研究了影响鱼类胚胎玻璃化冷冻保存的几个主要因子: 玻璃化液、麦管直径、胚胎阶段、平衡时间及平衡温度、洗脱浓度和洗脱时间。发现: (1) 含有多种抗冻剂的玻璃化液PMDD(2% PVP), 玻璃化稳定, 脱玻璃化率较低, 适宜进行玻璃化冷冻; (2) 尾芽期胚胎较其他时期耐受力强, 平衡40 min就足以使玻璃化液渗透完全, 时间延长, 成活率显著降低, 各个时期的胚胎对温度都比较敏感, 0°C与4°C下平衡的成活率显著高于15°C; (3) 洗脱浓度和洗脱时间对胚胎成活率影响不大; (4) 根据优化的条件, 对牙鲆两个时期的胚胎进行超低温冷冻保存实验, 共成活4次, 获得成活胚胎8粒, 其中7粒孵化出健康的鱼苗。本文为鱼类胚胎冷冻保存技术的建立提供基础资料, 并显示了牙鲆胚胎玻璃化冷冻保存是可行的。     

A set of polymorphic trinucleotide and tetranucleotide microsatellite markers for the Japanese flounder (Paralichthys olivaceus)

We have developed the first set of trinucleotide and tetranucleotide markers for the Japanese flounder, Paralichthys olivaceus. One hundred and sixty-seven polymorphic trinucleotide and tetranucleotide microsatellites were isolated using clones derived from two libraries. Of almost 200,000 clones analysed, 0.5% presented trinucleotide or tetranucleotide repeat regions. Among the trinucleotide repeats analysed in this study, the most frequent one was (CAG)(n) and the most common tetranucleotide repeat was (GATA)(n). The position of the new markers in the genetic linkage map was determined. Markers were evenly distributed along the P. olivaceus linkage groups, without distinction between the kinds of repeats and library of origin. The markers isolated in this study contribute significantly to the genetic linkage map of the Japanese flounder.     

Accumulation and oxidative stress biomarkers in Japanese flounder larvae and juveniles under chronic cadmium exposure

This study investigated how Cd exposure affected oxidative biomarkers in Japanese flounder, Paralichthys olivaceus, at early life stages (ELS). Fish were exposed to waterborne Cd (0–48 µg L^− 1) from embryonic to juvenile stages for 80 days. Growth, Cd accumulation, activities of superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6), glutathione S-transferase (GST, EC 2.5.1.18), and levels of glutathione (GSH) and lipid peroxidation (LPO) were investigated at three developmental stages. Flounder growth decreased and Cd accumulation increased with increasing Cd concentration. In metamorphosing larvae, CAT and SOD activities were inhibited and GSH level was elevated, while LPO was enhanced by increasing Cd concentrations. CAT and GST activities of settling larvae were inhibited but GSH level was elevated at high Cd concentrations. In juveniles, SOD activity and LPO level were increased but GST activity was inhibited as Cd concentration increased. Antioxidants in flounder at ELS were able to develop ductile responses to defend against oxidative stress, but LPO fatally occurred due to Cd exposure. These biochemical parameters could be used as effective oxidative biomarkers for evaluating Cd contamination and toxicity in marine environments: CAT, SOD, GSH, and LPO for metamorphosing stage; CAT, GSH, and GST for settling stage; and SOD, GST, and LPO for juvenile stage.     

蛋白质限制后恢复投喂对牙鲆幼鱼生长的影响

2004年4月4日~5月22日,在(23.0±0.5)℃条件下研究了牙鲆(平均体重,(8.80±0.18)g)继蛋白质限制后恢复投喂对其生长的影响。整个实验期间对照组(C组)连续48d饱食投喂含能17.29kJ/g,含粗蛋白50.00%的饵料。蛋白质限制阶段(1~18d)处理组T30和T40的饵料蛋白含量分别为27.95%和40.47%,但饵料含能与对照组相同,在恢复投喂阶段(19~48d)各处理组均投喂与对照组相同的饵料。实验结果表明,在蛋白质限制阶段,饵料蛋白质水平下降显著降低了处理组鱼的体重、特定生长率、饵料湿重、干重以及能量转化率(p<0.05),而摄食率、蛋白质效率和饵料蛋白质转化率随饵料蛋白水平降低而显著升高(p<0.05)。恢复投喂结束后,处理组上述各指标均恢复至对照组水平(p>0.05)。此外,除实验结束时T30组鱼体脂肪含量显著低于对照组(p<0.05)外,整个实验期间各处理组的表观消化率及鱼体成分与对照组相比均无显著差异(p>0.05)。实验结果表明,牙鲆幼鱼继蛋白质限制后的恢复投喂阶段出现了完全补偿生长效应。     

Demand-feeding behaviour and diel pattern of feeding activity in Oncorhynchus mykiss held under different photoperiod regimes

Demand-feeding behaviour was studied in Oncorhynchus mykiss held at 7–10°C in groups of 30 individuals, under a 16:8, 12:12 or 8 : 16 light : dark regime, using self-feeders connected to a computer. The daily food demand was variable between tanks from 1.5 to 5% of the body weight, the specific growth rate ranged between 1.4 and 2.6% of body weight increase per day, and the feed: gain ratio ranged between 0.84 and 3.17, regardless of the photoperiod length. More than 98% of the feeding demand occurred during the photophase, regardless of the photoperiod, with a main peak at dawn and an occasional peak at dusk.     

Feeding behaviour of Helicoverpa armigera larvae on insect-resistant transgenic cotton and non-transgenic cotton

Abstract: Feeding behaviour of Helicoverpa armigera Hübner (Lep.; Noctuidae) larvae on non‐transgenic Bacillus thuringiensis (Bt) cotton (Gossypium hirsutum L.), Zhong 30, and transgenic cowpea trypsin inhibitor (CpTI)‐Bt cotton, SGK 321, and non‐transgenic cotton, Shiyuan 321, was investigated in both choice tests and no‐choice tests. The results of choice tests suggested that neonates have the ability to detect and avoid transgenic cotton. In the choice tests of neonates with both transgenic and non‐transgenic cotton leaves, a significantly greater proportion of larvae and higher consumption were observed on non‐transgenic cotton than on the transgenic Bt or CpTI‐Bt cotton. In the choice tests with leaves of two transgenic cotton lines, the proportion of neonates on leaf discs of the two lines was not significantly different, but there was significantly higher consumption on CpTI‐Bt transgenic cotton than that on Bt transgenic cotton. In addition, significantly more neonates were found away from the leaf discs, lower consumption and higher mortality were achieved in the choice test with two transgenic cotton leaves than in the choice tests containing non‐transgenic cotton leaves. Leaves and buds were examined in choice tests of fourth instars. It appeared that fourth instars were found in equal numbers on transgenic and non‐transgenic cotton, except when larvae were exposed to leaves for 3 h. However, the total consumption on transgenic cotton was lower than that of the non‐transgenic cotton, so fourth instars may still have the capacity to detect transgenic cotton and reduce feeding on it, although they showed no preference on either transgenic or non‐transgenic cotton. More larvae were found off diet in the treatments with leaves than that of buds, and the number of injured leaf discs by per fourth instar was significantly higher than that of buds in choice tests, suggesting that leaf is a less preferred organ for H. armigera larvae, elicited more larval movements. Similarly, in no‐choice tests of fifth instars, significantly fewer feeding time and more moving time occurred on leaf than that of bud, boll and petal. When cotton line was considered, compared with non‐transgenic cotton, significantly lower feeding time and higher resting time occurred on the two transgenic cottons. Overall, H. armigera larvae have the ability to detect the transgenic Bt and CpTI‐Bt cottons or the less preferred organs and selectively feed more on the non‐transgenic cotton or the preferred organs, especially the neonates, which have a high capacity for avoiding transgenic cotton.     

Feeding behaviour of artificially reared Romane lambs

A consequence of increasing litter size in sheep is that a portion of the lambs have to be reared artificially. Detailed information about the pattern of milk consumption of artificially reared lambs would help improve their management. The purpose of this study is to describe the individual and group feeding behaviour of 94 Romane artificially reared lambs from 5 to 28 days of age using an electronic automatic lamb feeder. Animals were located in four pens of 8 to 15 lambs of similar age with one teat per pen. They were fed ad libitum. In our experimental situation (group rearing, continuous lightning) on average a lamb made 1.4±0.7 visits to the teat per meal and 9.5±3 meals per day. Mean meal duration was 247±158 s and the mean daily time spent feeding was 38±25 min. The mean quantity of milk intake was 176±132 ml per meal and 1.68±0.8 l per day. With age, the number of daily meals and their duration decreased while the quantity of milk consumed per meal and per day increased. Females tended to make more visits to the teat per meal and perform more meals per day but their milk consumption per meal was lower. The feed conversion ratio was 1.36±0.2. Synchrony in feeding (group meal) was estimated as the percentage of lambs that wanted to access the teat within the same short period (relative group meal size). On average 65% of lambs in the pen wanted to access the teat within the same period, but for 35% of group meals the relative group meal size was >90%. There was no consistency in the order in which lambs accessed the teat during a group meal. Our evaluation suggested that electronic automatic lamb feeders are tools that can provide, on a large scale, data describing the feeding behaviour of artificially reared lambs. It is then possible to study factors influencing these traits in order to improve the outcome of artificially reared lambs.     

Copulation behaviour of Lygocoris pabulinus under laboratory conditions

As a prerequisite to the elucidation of the sex pheromone of the green capsid bug Lygocoris pabulinus (L.), we studied the bug's reproductive development and behaviour. When kept under long-day conditions (L18:D6) at 25 °C, both males and females start to copulate 4 days after the final moult. Second matings occur in some females, even on consecutive days. The premating period is 10 min on average and copulation lasts 1 to 2 min. At long range, when males are attracted to traps baited with live females, we did not observe a specific calling position of the females. At short range, a characteristic courtship behaviour of the females was not observed either. Males showed a characteristic vibration of the abdomen, which was repeated several times before copulation. This behaviour can be used as a bioassay to test potential sex pheromone compounds.     

渤海南部牙鲆的食性及摄食的季节性变化

1051尾渤海南部牙鲆的胃含物分析表明,渤海南部牙鲆为捕食性(游泳生物食性)鱼类,其食物成分有30余种,以底栖虾类中的口虾蛄、鲜明鼓虾、日本鼓虾、脊尾褐虾,软体动物的日本枪乌贼及鱼类中的斑(鱼祭)、鳀鱼、黄鲫、六丝矛尾虾虎鱼及焦氏舌鳎等动物为主要食物。牙鲆除冬季外,其余时间均强烈摄食,而且其摄食的季节性变化明显:春季以鱼类(重量占80.9％)及甲壳类(18.6％)为主要食物;夏季以鱼类(83.5％)及软体动物类(13.1％)为主要食物;秋季主要摄食鱼类(87.9％);冬季则主要摄食鱼类(73.2％)及甲壳类(19.7％)。     

Predatory behaviour ofScatophaga stercoraria under laboratory conditions

Laboratory experiments tested the effectiveness ofScatophaga stercoraria (L.) as a predator of various insect species. Photophases, ages and gender of predators and effects of dung presence on predation rates were compared for either sex using counts of prey cadavers, behavioral observations and oviposition rates. Males consistently killed more prey than females, but longer photophases did not significantly increase daily predation rates. Nine to 10-day-old flies approaching sexual maturity achieved highest predation rates. Dung was found to affect predatory behaviour but not cumulative predation. Predatory behaviour of sexually mature males was more affected than female behaviour by the presence of dung, whereas sexually immatures flies showed no response to dung presence. Preconditioning ofS. stercoraria adults using 3 prey species of different sizes did not affect the prey chosen in subsequent random choice experiments. Larger prey species such asMusca domestica (L.) orDelia antiqua (Meigen) were preferred toDrosophila sp. by both sexes ofS. stercoraria. This species may be a useful predator for future integrated pest management techniques in vegetable crops or control of house flies in barns.      

Interactive effect between dietary valine and isoleucine on growth performance,blood parameters and resistance against low salinity stress of Japanese flounder Paralichthys olivaceus

The interaction between dietary valine (Val) and isoleucine (Ile) on growth, blood parameters, and tolerance against low salinity stress of Japanese flounder Paralichthys olivaceus was studied in a 60-day feeding trial. Fish (initial body weight: 3.57 ± 0.05 g) were fed six experimental diets containing different proportions of Ile and Val, two levels of Ile 0% (LI) and 2% diet (HI), combine with three levels of Val 0% (LV), 0.83% (MV), and 2.27% diet (HV), respectively. The results revealed that significant interactive effect of Ile and Val was found on body weight gain and feed conversion ratio, also stimulative effect was found in high Ile groups. Meanwhile, interactive effect of these two amino acids was also observed on high-density lipoprotein (HDL), the maximum of Ile with maximum of Val group has the highest concentration of HDL, which was significantly higher than other groups (p < .05). Moreover, no interaction was found on whole body proximate composition, muscle Ile and Val concentration, and tolerance ability against low salinity stress (reflected by LT₅₀). But high level of Ile combined with high level of Val diet altered tolerance ability against low salinity stress of Japanese flounder. It would be indicated that dietary Ile and Val interacted in the diet of Japanese flounder, and the flounders fed the diet supplemented with 2% Ile and 2.27% Val was superior to the other groups on growth and blood parameters, and tolerance against low salinity stress.     

Feeding behaviour,midgut distension and ovarian development in Phlebotomus papatasi (Diptera: Psychodidae)

Phlebotomus papatasi females were fed through membranes or from cotton wool soaked in blood, water, sucrose or sodium chloride solutions. In membrane-fed flies, all diets were routed to the midgut and not to the crop. Following the meals that went to the midgut, females showed ovarian development at least 3 times greater than in sucrose-fed, autogenous control flies. Neither small quantities of water arriving in the midgut following drinking from soaked cotton wool, nor piercing of a membrane without feeding, stimulated ovarian development. Flies exhibited different feeding behaviour namely, blood feeding, sugar feeding, and water drinking. The blood-feeding behaviour was typical of flies ingesting any of the experimental diets through membranes, or blood or saline from cotton wool. The other two types of behaviour were observed in flies which fed from soaked cotton wool. The type of behaviour was characterized by the depth of penetration of the mouthparts into the substrate, the deployment of the palps and the degree of contact between the palps and the surface. It is suggested that the stimuli which control the routing of meals to the crop or to the midgut are derived from these types of behaviour.     

牙鲆一株弹状病毒病原的分离与鉴定

从患病牙鲆中分离鉴定了一株弹状病毒（Paralichthys olivaceus rhabdovirus,PoRV）。用过滤除菌后的患病牙鲆组织匀浆液,接种不同的鱼类细胞,其中有7种鱼类细胞出现明显的病变在对病毒进行挑斑分离后,测定了PoRV的滴度,显示PoRV在敏感鱼类细胞（Grass Carp Ovary,GCO）中的滴度达到106.5TCID50/mL;绘制了PoRV生长曲线;经蔗糖密度梯度离心提纯PoRV,负染及宿主细胞超薄切片的电镜观察,显示PoRV大小约为60nm×200nm。测定了PoRV的理化性质,显示该病毒对有机溶剂和温度敏感,但对DNA抑制剂阿糖胞苷（Ara-c）不敏感。经SDS-PAGE电泳,对PoRV的蛋白图谱进行了分析,表明该病毒有5种主要蛋白带,其分子量大小分别约为：250kD、67kD、44kD、30kD、23kD。     

Relationship between stable cesium concentration and body size of Japanese flounder Paralichthys olivaceus and the effect of a size-dependent shift in diet

To understand the relationship between the radioactive cesium (Cs) concentration in muscle of Japanese flounder Paralichthys olivaceus and the species' biological characteristics (size, sex, and age) under conditions of ecological equilibrium (i.e., distributed among ecosystem components over sufficient time, and with nearly constant ratios of Cs concentration in organisms to the concentration in water) as existed before the accident at the Fukushima Dai-ichi Nuclear Power Station (FDNPS), Japan, in 2011, we examined stable Cs, as it is thought to exist in equilibrium in the environment and behave similarly to radioactive Cs in aquatic animals. The concentration of stable Cs in 241 P. olivaceus (range 216–782 mm total length [TL]) collected in Sendai Bay, approximately 90 km north of the FDNPS, in June–July 2015 was expressed as an exponential function with size as an independent variable; the results show the concentration of stable Cs doubled with an increase in TL of 442 mm. Next, to evaluate the cause of the size-dependent change in stable Cs concentration, we examined 909 individuals (200–770 mm TL) collected in September 2013–July 2015 to determine their feeding habit based on size. Analysis of the frequency of occurrence of prey organisms in stomach contents showed that sand lance Ammodytes japonicus (55–180 mm standard length [SL]) was the most consistently consumed across size classes. Analysis on a wet-mass basis showed that A. japonicus and anchovy Engraulis japonicus (65–130 mm SL) were the main food of P. olivaceus sized 200–599 mm TL, whereas chub mackerel Scomber japonicus (120–230 mm SL) and two species of flatfishes (180–205 mm SL) were abundant in the diet of P. olivaceus sized ≥600 mm TL. All these prey items were presumed to have similar concentrations of stable Cs. Based on the above, the effect of diet on the relationship between stable Cs in muscle and fish size was considered negligible. That the diet of P. olivaceus largely did not change with size was also confirmed by C and N stable isotope ratios in P. olivaceus and their prey species. Therefore, the Cs–size relationship is probably determined by changes in the balance between the rate of Cs intake from food and seawater and the excretion rate during growth, both of which change as functions of body mass. Values of stable Cs concentrations among environmental components and animals appear to be a valid indicator for understanding the radioactive Cs distribution in the marine environment and aquatic animals under the equilibrium state, as existed before the 2011 nuclear accident.     

Effects of temperature and delayed initial feeding on the survival and growth of Japanese flounder larvae

The effects of the timing of initial feeding (0, 1, 2, 3 and 4 days after yolk exhaustion) and temperature (15, 18 and 21° C) on the point‐of‐no‐return (PNR), survival and growth of laboratory‐reared Japanese flounder Paralichthys olivaceus larvae were studied under controlled conditions. The larvae reached PNR on 7·7, 5·2 and 4·2 days‐post‐hatching (dph) at 15, 18 and 21° C, respectively. At each temperature, larval growth did not differ significantly among the delayed initial feedings 1 day before PNR but decreased significantly in larvae first fed after that. In the treatments where initial feeding was equally delayed, larvae grew significantly faster at 18 and 21° C than at 15° C. The larvae survived apparently better at 15 and 18° C than at 21° C when initial feeding was equally delayed. At each temperature, survival of the larvae first fed before PNR did not differ noticeably, while delayed initial feeding after that apparently reduced their survival. These results indicated that there existed a negatively temperature‐dependent PNR in the Japanese flounder larvae. Survival and growth of the larvae strongly depended on temperature as well as the timing of initial feeding. High temperature accelerated the yolk exhaustion and growth of the larvae and thus reduced their starvation tolerance and survival. To avoid potential starvation mortality and obtain good growth, the Japanese flounder larvae must establish successful initial feeding within 2 days after yolk exhaustion at 15° C and within 1 day at both 18 and 21° C.