Molecular cloning and functional analysis of cathepsin B in nutrient metabolism during larval development in Meretrix meretrix

Seed rearing is an important part in large scale clam culture industry. Since the nutritional history affects early development in bivalve, the condition of larval nutrition plays a key role in successful seed rearing. So far, the molecular mechanism of nutrient uptake in bivalve larvae is unclear. As one of the important proteolytic enzymes, cathepsin B of several organisms has been reported to be involved in digestion. We intended to analyze whether cathepsin B is involved in larval nutrient metabolism in the economic bivalve, clam Meretrix meretrix. The full length of M. meretrix cathepsin B (MmeCB) cDNA was cloned, which is 1647 bp with an open reading frame of 1014 bp. The deduced amino acid sequence encoded a preproenzyme of 337 residues with Cys-114, His-282 and Asn-302 composing cathepsin B activity center. The temporal and spatial expressions of MmeCB mRNA were examined from trochophore to post larva stages by whole mount in situ hybridization. In trochophore stage, no detectable signal was found. In the later three stages, MmeCB mRNA was detected in the digestive gland, suggesting a possible role of MmeCB in digestion. Moreover, MmeCB mRNA was also observed in the epidermal cells in D-veligers. Cathepsin B specific inhibitor (CA074 methyl ester) was applied to block the activity of cathepsin B in unfed larvae. The average shell lengths of treated larvae were smaller than that in control groups. The results of mRNA epidermal distribution and inhibitor treatment in D-veligers indicated that MmeCB may be also associated with other pathway of nutrient metabolism in larval epidermis. The overall results in this paper revealed that MmeCB might play a role in larval nutrient metabolism. (C) 2008 Elsevier B.V. All rights reserved.

Effects of rearing temperature and density on growth, survival and development of sea cucumber larvae, Apostichopus japonicus (Selenka)

In laboratory conditions, effects of rearing temperature and stocking density were examined on hatching of fertilized egg and growth of auricularia larvae of Apostichopus japonicus respectively. Data series like larval length and density, metamorphic time, and survival rate of the larvae were recorded. Statistics showed that for A. japonicus, survival rate (from fertilized egg to late auricularia) decreased significantly with the increasing rearing temperature (P < 0.05). At different temperatures SGR was statistically significant as well (P < 0.05) from day 1, and maximal SGR was found on day 9 at 24A degrees C (159.26 +/- 3.28). This study clearly indicated that at low temperature (< 24A degrees C), metamorphic rate was remarkably higher than at higher temperature (> 26A degrees C). Hatching rate was significantly different between 0.2-5 ind./ml groups and 20-50 ind./ml groups. Rearing larvae at the higher density had the smaller maximal-length, whereas needed longer time to complete metamorphosis. This study suggested that 21A degrees C and 0.4 ind./ml can be used as the most suitable rearing temperature and stocking density for large -scale artificial breeding of A. japonicus's larvae.

Structure of a fig wasp community: Temporal segregation of oviposition and larval diets

Observations were made on six fig wasp species on Ficus racemosa growing in the Xishuangbanna Tropical Botanic Garden, Yunnan Province, China. The oviposition sequence was determined for Apocryptophagus testacea, Apocrypta sp2, Apocryptophagus mayri, Cera

Effects of Abnormal Early Rearing Environments on Fear Memory in Adult Rats

为了探讨早期饲养环境对成年后大鼠条件化恐惧的影响,实验采用巴氏恐惧条件化的方法对不同饲养环境下的大鼠进行了行为检测.具体操作为:把断奶后的大鼠饲养在3个不同的人工控制的环境中(丰富环境、社群环境和贫瘠环境);8周后进行恐惧条件化训练和测试(指标为僵直百分比),以及检测不同饲养环境下的大鼠的体重、自发活动和足部电击敏感性.结果显示:与社群组相比,丰富组的大鼠条件化僵直水平明显增加,而贫瘠组的明显减少;丰富和贫瘠环境明显的影响了大鼠的体重;不同环境对自发活动和足部电击敏感性没有影响.这些结果说明幼年期的丰富环境能提高成年后大鼠的由声音诱发的条件化恐惧反应,而贫瘠环境则削弱了这种反应.

Mass rearing of Aphidius gifuensis (Hymenoptera : Aphidiidae) for biological control of Myzus persicae (Homoptera : Aphididae)

The green peach aphid, Myzus persicae, is a major pest of tobacco, Nicotiana tabacum, in Yunnan province, China, where its control still depends on the use of insecticides. In recent years, the local government and farmers have sought to improve the biological control of this tobacco pest. In this paper, we present methods for mass rearing Aphidius gifuensis, a dominant endoparasitoid of M. persicae on tobacco plants in this region. The tobacco cultivar K326 (N. tabacum) was used as the host plant and M. persicae as the host insect. In the greenhouse, we collected tobacco seedlings for about 35 days (i.e., until the six-true-leaf stage), transferred them to 7.5-cm diameter pots, and kept these plants in the greenhouse for another 18 days. These pots were then transferred to an insectary-greenhouse, where the tobacco seedlings were inoculated with five to seven wingless adult M. persicae per pot. After 3 days, the infested seedlings were moved to a second greenhouse to allow the aphid population to increase, and after an additional 4 +/- 1 days when 182 +/- 4.25 aphid adults and nymphs were produced per pot, they were inoculated with A. gifuensis. With this rearing system, we were able to produce 256 +/- 8.8 aphid mummies per pot, with an emergence rate of 95.6 +/- 2.45%; 69% were females. The daily cost of parasite production (recurring costs only) was US$ 0.06 per 1000 aphid mummies. With this technique, we released 109 800 parasitoids in 1998, 196 000 in 1999, 780 000 in 2000, and 5 600 000 in 2001 during a 2-month period each year This production method is discussed with respect to countrywide usage in biological control and integrated control of M. persicae.

Effects of pure microcystin-LR on the transcription of immune related genes and heat shock proteins in larval stage of zebrafish (Danio rerio)

Microcystins (MCs) are cyanobacterial toxins in water blooms that have received increasing attention as a public biohazard for human and animal health. Previous studies were mainly focused on the toxic effects on adult fish, rather than juvenile or larvae, and the response of fish immune system were usually neglected. This paper presents the first data of the effects of microcystin-LR (MC-LR) on transcription of several genes essential for early lymphoid development (Rag1, Rag2, Ikaros, GATA1, Lck and TCR alpha) and heat shock proteins (HSP90, HSP70, HSP60, HSP27) in zebrafish larvae. Relative changes of mRNA transcription were analyzed by real time PCR. The transcription of Rag1, Rag2, Ikaros, GATA1, Lck and TCR alpha were up-regulated when following exposure to 800 mu g/L MC-LR, which may indicate that specific lymphocytes differentiation and TCR/lg arrangement are induced to counteract the toxic effects of MC-LR. It was also interesting to note the dramatically increased transcription of HSP90. HSP70, HSP60 and HSP27, which may indicate their important roles as molecular chaperones under oxidative stress. (C) 2009 Elsevier B.V. All rights reserved.

Feeding rates affect heat shock protein levels in liver of larval white sturgeon (Acipenser transmontanus)

A two-week trial was conducted to study the effect of feeding rates on heat shock protein levels in larval white sturgeon. The larvae (30 day post hatch, 230 mg initial body weight) were fed a commercial feed (12.6% moisture, 49.5% crude protein. 20.7% Crude fat, and 8.6% ash) at 5, 15. or 25% body weight per clay (BW d(-1)). Liver heat shock proteins (Hsp) were measured before and after the larvae were subjected to a heat shock from 18 to 26 degrees C at 1 degrees C/15 min and maintained at 26 degrees C for 4 h thereafter. Before heat shock, larvae fed 5% BW d(-1) had significantly (P<0.05) lower final body weight, RNA/DNA ratio, whole body lipid and protein content, and Hsp60 and Hsp70 levels but higher protein efficiency ratio, and whole body moisture content than larvae fed the two higher feeding rates. Heat shock significantly induced Hsp60 and Hsp70 levels in the liver of all fish but they were lower in larvae fed the 5% than those fed 15 and 25% BW d(-1). Hsp70 level increased much more than Hsp60 after the heat shock Suggesting that Hsp70 is a more sensitive biomarker under our experimental conditions. (c) 2008 Elsevier B.V. All rights reserved.

Apo-14 is required for digestive system organogenesis during fish embryogenesis and larval development

Apo-14 is a fish-specific apolipoprotein and its biological function remains unknown. In this study, CagApo-14 was cloned from gibel carp (Carassius auratus gibelio) and its expression pattern was investigated during embryogenesis and early larval development. The CagApo-14 transcript and its protein product were firstly localized in the yolk syncytial layer at a high level during embryogenesis, and then found to be restricted to the digestive system including liver and intestine in later embryos and early larvae. Immunofluorescence staining in larvae and adults indicated that CagApo-14 protein was predominantly synthesized in and excreted from sinusoidal endothelial cells of liver tissue. Morpholino knockdown of CagApo-14 resulted in severe disruption of digestive organs including liver, intestine, pancreas and swim bladder. Moreover, yolk lipid transportation and utilization were severely affected in the CagApo-14 morphants. Overall, this data indicates that CagApo-14 is required for digestive system organogenesis during fish embryogenesis and larval development.

Dose-dependent effects of extracted microcystins on embryonic development, larval growth and histopathological changes of southern catfish (Silurus meridionalis)

A laboratory toxic experiment was conducted to examine dose-dependent effects of extracted microcystins (MCs) on embryonic development, larval growth and histopathological changes of southern catfish (Silurus meridionalis). Fertilized eggs were incubated in solutions with four concentrations of MCs (0, 1, 10, 100 mu g MC-LReq l(-1)). Higher MCs retarded egg development (2-10 h delays) and larval growth, reduced hatching rate (up to 45%), and caused high malformation rate (up to 15%) and hepatocytes damage (characterized by disorganization of cell structure and a loss of adherence between hepatocytes, cellular degeneration with vacuolar hepatocytes and marginal nuclei, even hepatocellular necrosis). A 10 mu g MC-LReql(-1) is close to a high concentration in natural cyanobacterial blooms, suggesting a possible existence of such toxic effects in eutrophic waters. (c) 2007 Elsevier Ltd. All rights reserved.

Rearing of Procypris rabaudi during early life-history stages

The early life-history of Chinese rock carp Procypris rabaudi was investigated during a 56-day rearing period: 318 artificially propagated P. rabaudi larvae were reared throughout metamorphosis in a small-scale recirculation system (345 L water volume, 10 x 18 L rearing tanks, 150 L storage and filter compartment with bioballs, 20-30 larvae L-1) at the Institute of Hydrobiology, Wuhan, Hubei Province, China. The newly hatched larvae had an initial total length of 8.93 +/- 0.35 mm SD (n = 10) at 3 days post-hatch and reached an average total length of 33.29 mm (+/- 1.88 mm SD, n = 10) 56 days after hatching. Length increment averaged 0.45 mm day(-1), resulting in a mean growth of 24.4 mm within the 56-day period. High mortality rates of up to 92% derived from an introduced fungus infection and subsequent treatment stress with malachite green. Our results indicate that Chinese rock carp can be raised successfully from artificially fertilized eggs. We therefore assume this species to be a candidate for commercial aquaculture.

The toxic effects of microcystin-LR on embryo-larval and juvenile development of loach, Misguruns mizolepis Gunthe

Microcystin-LR, a specific and potent hepatotoxin, was tested for its effects oil loach embryo-larval and juvenile development, The results of this study showed that loach embryos were more sensitive when exposed to microcystin-LR at a later than at an earlier stage of development, Juveniles were far less sensitive to MC-LR than were embryos and larvae. Mortality and developmental abnormality were proven to be dose-dependent and to be stage-specific sensitive. Among the abnormal changes noted were: pericardial edema and tubular heart, bradycardia, homeostasis, poor yolk resumption. small head, curved body and tail, and abnormal hatching, Liver and heart were the main targets of microcystin-LR toxicity. Ultrastructural analysis documented a complex set of sublethal effects of microcystin-LR on loach hepatocytes, chiefly including morphological alteration in nuclear and RER of loach liver cells. fit addition, microcystin-LR was lethal to loach juvenile in the subacute (7 days) exposure (LC50) = 593.3 mug/l). (C) 2002 Elsevier Science Ltd. All rights reserved.

EFFECT OF DENSITY ON LARVAL DEVELOPMENT AND FEMALE PRODUCTIVITY OF TISBE-HOLOTHURIAE (COPEPODA, HARPACTICOIDA) UNDER LABORATORY CONDITIONS

POPULATION-DYNAMICS; FOOD; FISH

Larval metamorphosis and morphological characteristic analysis of triploid shrimp Fenneropenaeus chinensis (Osbeck, 1765)

Both MI and MII triploids were successfully produced by heat shock in Chinese shrimp Fenneropenaeus chinensis. The inducing conditions for MI and MII triploids were optimized. The highest inducing rate obtained for MI triploids reached more than 90%, and that for MII triploids reached nearly 100% at the nauplius stage as evaluated using flow cytometry. Comparisons of survival rates at larval stages between triploids and diploids or diploids experiencing treatment and diploids without treatment were performed. At larval stage from nauplii to postlarvae, heat shocks lowered survival at larval stages even if the ploidy was not changed. Ploidy did not affect shrimp larvae survival, and no significant difference was found in the survival of shrimp larvae between MI and MII triploids. Highly significant differences were observed in the morphology of triploids and diploids, and no apparent difference was found in the morphology of MI and MII triploids at the grow-out stages. Discriminating formulae for triploid and diploid shrimp at grow-out stage were developed and could be used to distinguish triploids from diploids based on morphological parameters. MI and MII triploids of shrimp have the potential to be used in aquaculture.

Induction of triploidy in large yellow crocker Pseudosciaena crocea (Richardson, 1846): effects of pressure shocks and growth performance in the first rearing year

The precociously sexual maturation in large yellow crocker Pseudosciaena crocea has become a serious problem. In an attempt to solve this problem, the production of sterile triploids could be an effective strategy. In this study, triploid P. crocea was obtained by subjecting fertilized eggs to pressure shock. Flow-cytometry analysis was used to assess ploidy level. In terms of triploid rate and hatching rate, the optimal conditions of pressure shock for triploidy induction in P. crocea were 7500 psi for 3 min shock at 3 min after fertilization at 20 degrees C. With the application of these parameters, 100% triploid fish were produced. During the first rearing year, triploid P. crocea had a similar growth performance compared with its diploid counterpart before the age of 8 months and showed a significant advantage at the age of 10 and 12 months in body weight and body length (P < 0.05). At the age of 12 months, the carcass weight of triploids was markedly higher than that of diploid control, and gonadal somatic index was significantly lower than that of their diploid control. During the first rearing year, survival in triploid group was 76.44%, inferior to its diploid control (83.21%).