中国对虾中一种球状病毒的分离提纯与检测

从人工养殖生长比较缓慢的对虾与病虾的肝胰腺和消化道中,分离出一种大小为８０ｎｍ的球状病毒。同时采用超薄切片技术可在对虾中肠细胞中发现大量的球状病毒,这些病毒聚集成片,在同一细胞内的细胞器,如线粒体等均发生病变。此外,将分离提取的球状病毒进行感染试验,感染死亡率为６０％。并且从感染致死的对虾消化腺及肠道中,同样可以观察到大小相同、形态一致的球状病毒颗粒。     

两种病毒侵染中国对虾后细胞超微病理学变化与免疫标记

应用电子显微技术研究人工感染的中国对虾病毒病原及其宿主细胞超微病理学变化。结果显示病虾体内存在球状与杆状两种病毒病原,有时在同一病虾组织的同一细胞中可见两种病毒同时侵染现象,该现象提示存在复合感染的可能。利用胶体金免疫标记技术对感染病虾细胞质中出现的球状病毒作定位标记,初步结果表明已分离提纯的球状病毒与感染病虾细胞质中观察到的病毒粒子性质基本相同。病毒侵染后,细胞内主要的细胞器如线粒体、内质网、核糖体均发生了显著变化;侵染后期,可见溶酶体及多种膜性结构大量增生、细胞核被一些微管样结构包裹等特殊变化的发生。     

中国对虾呼肠孤样病毒感染的电镜观察

呼肠孤病毒在自然界广泛存在于脊椎动物、无脊椎动物和植物中.水产动物呼肠孤病毒感染曾见于鱼、贝、蟹.近几年随着对对虾病毒病害研究的日益重视,在对虾中也发现呼肠孤病毒的感染.Tsing等[1]最早于1987年在法国南部养殖的日本对虾(P.japonicus)幼虾中发现呼肠孤病毒感染.Krol等[2]于1990年在试验感染的南美白对虾(P.vannamei)中发现呼肠孤样病毒与对虾杆状病毒混合感染.中国大陆养殖的中国对虾自1993年全面暴发流行病以来,许多学者进行了对虾流行病的病原学、流行病学及诊断和防治方法的研究,部分学者曾在中国对虾(P.chinensis)中观察到呼肠孤样病毒颗粒[3],但未报道较详细的电子显微镜观察资料.     

罗氏沼虾苗肌肉白浊病病原的初步研究

收集罗氏沼虾肌肉白浊病典型病虾苗 ,进行细菌分离、除菌组织悬液制备及人工感染试验、超薄切片、病毒粗提取 ,结果表明 :从白浊病病虾的肌肉、肝胰腺中分离到的细菌以5× 1 0 8CFU/mL浓度浸泡感染正常沼虾苗不能复制疾病 ;病虾除菌组织过滤液以 1∶5 0、1∶2 5 0浓度浸泡感染正常虾苗 ,7d后可发病 ,并出现典型的肌肉白浊病症状 ,氯仿处理不能破坏病毒的感染力 ;病虾超薄切片电镜观察发现肌肉间隙组织中存在 2 1— 2 3nm大小的球状病毒 ,病虾组织浆经氯仿处理、PEG沉淀、磷钨酸负染后可见到大量 2 2— 2 4nm大小的球状病毒颗粒。上述结果表明罗氏沼虾肌肉白浊病是一种由病毒引起的疾病。     

中国对虾两种球状病毒的免疫检测

以江苏、河北某虾场病虾组织中提取的对虾的两种球状病毒作为抗原,制备出特异性强、效价较高的抗血清,用对流免疫电泳,酶联免疫吸附,免疫电镜技术对五个不同地区的病虾材料进行检测,结果表明：江苏、青岛、河北地区的病虾组织中均含有相应的球状病毒病原。     

渤海湾对虾发生量与补充量动态特征的研究

每年５月中旬和８月初,在中国对虾（Ｐｅｎａｅｕｓｃｈｉｎｅｎｓｉｓ）的主要产卵场———渤海湾进行产卵场和栖息地调查,分别获取卵子／幼体和幼虾相对资源量,把卵子／幼体密度视为亲体数量（Ｓｔ）的一个相对数值,把幼虾相对资源量视为补充量（Ｒｔ）的一个相对数值,用Ｒｉｃｋｅｒ模式（Ｒｔ＝ａＳｔｅ－ｂＳｔ）拟合资料,有：ａ＝２９２２４３;ｂ＝００６９;Ｒ２＝０４７;Ｐ＜０００５。结果表明：对虾的发生量对其补充量的控制程度近５０％。通过回归分析,确证了对虾早期发育阶段栖息地的降雨量、大风、日照和黄河径流量等环境因素对卵子／幼体的成活率乃至对补充量的重要影响     

温度对中国对虾（Penaeus orientalis）越冬亲虾性腺发育与存活率的影响

本文根据１１ａ来中国对虾越冬亲虾生产性培育试验资料的统计分析结果得出：亲虾性腺发育的起点温度为７．９±０．３℃,性腺发育成熟的有效积温为２５０．４±３３．６日度,预测性腺发育成熟所需日期公式为。升高或降低越冬期间的基础水温可控制亲虾产卵时间迟早。产卵期间间歇升降温或连续升温,可控制亲虾集中产卵的次数和亲虾产卵持续时间。亲虾越冬期间,有利于提高亲虾存活率的水温,宜控制在８．０－１３．０℃,不宜超过１３．５℃,升降温幅度不宜超过１．５℃,以１．０℃左右为好。亲虾产卵期间,有利于亲虾存活率、产后卵子孵化率和亲虾再次产卵率的适宜水温为１２．０－１５．０℃,不宜超过１６．０℃,升降温幅度不宜超过１．０℃,以１．０℃以下为好。     

中国对虾卵子活化及卵裂的激光扫描共聚焦显微镜研究(英文)

有报导,与其它动物不同,对虾的卵子一接触海水、不经过受精,就可以被活化。但是,只有受精卵才能卵裂发育。而且,各种对虾卵子内皮质棒及皮层颗粒的形态结构、活化过程各有不同。本文就中国对虾卵子的活化及卵裂过程进行了激光扫描共聚焦显微镜（ＬＳＣＭ）研究。该技术可以清晰地显示细胞的立体图像。采集中国对虾,人工诱导产卵,未受精卵取自产卵虾体内,受精卵分期分批取样,固定。供试样品经ＰＢＳ漂洗,ＡＯ、ＦＩＴＣ和ＰＩ染色,ＬＳＣＭ观察、图像处理。刚产出的、未活化卵子形态不规则,皮层内富含皮质棒和皮层颗粒（Ｆｉｇ１）。不论受精与否,产入海水的卵子都能被激活并恢复减数分裂。活化后的卵子释放出皮层囊泡内的皮质棒（Ｆｉｇｓ２＆３）。释放出的皮质棒转变为均质的胶质层（Ｆｉｇｓ４＆５）。随后,皮层颗粒释放并在卵子表面转化为均质化的孵化膜。卵子开始举起,与孵化膜之间形成卵周隙。卵周隙内具有絮状物质（Ｆｉｇｓ６,７＆８）。卵子产出后笫８～１０ｍｉｎ,经过减数分裂,第一极体排放,并随着孵化膜的举起而被推向卵外（Ｆｉｇ５）。笫２０ｍｉｎ,第二极体排放,并由卵子表面到达孵化膜的内缘（Ｆｉｇ６）。在２０℃水温条件下,受精卵产入海水中６０     

对虾白斑综合征杆状病毒体内增殖模型的建立

应用对虾白斑综合征杆状病毒（WSSV）,对淡水克氏螯虾、罗氏沼虾、日本沼虾和两种淡水蟹（中华绒螯蟹、长江华溪蟹）进行人工感染实验。结果除淡水克氏螯虾之外,其它受试的虾蟹均不能感染WSSV。克氏螯虾3个不同剂量级感染至12d平均死亡率为94％。从发病或死亡个体采集血淋巴,经电镜负染色可观察到完整的病毒粒子,其形态大小、靶细胞组织病理均与从中国对虾中分离的WSSV相似或相同。同时,通过原位杂交技术进一步证明该实验的可靠性。克氏螯虾重复感染效果良好,有可能成为研究WSSV的一种理想的病毒体内增殖模型。     

对虾白斑综合征杆状病毒体内增殖模型的建立

应用对虾白斑综合征杆状病毒(WSSV),对淡水克氏螯虾、罗氏沼虾、日本沼虾和两种淡水蟹(中华绒螯蟹、长江华溪蟹)进行人工感染实验.结果除淡水克氏螯虾之外,其它受试的虾蟹均不能感染WSSV.克氏螯虾3个不同剂量组感染至12 d,平均死亡率为94%.从发病或死亡个体采集血淋巴,经电镜负染色可观察到完整的病毒粒子,其形态大小、靶细胞组织病理均与从中国对虾中分离的WSSV相似或相同.同时,通过原位杂交技术进一步证明该实验的可靠性.克氏螯虾重复感染效果良好,有可能成为研究WSSV的一种理想的病毒体内增殖模型.     

Development of gene transfer technology for black tiger shrimp, Penaeus monodon

An effective foreign gene transfer method for shrimp would have several potential uses in the shrimp culture industry, such as in preventing infectious diseases. We evaluated two gene transfer methods and used black tiger shrimp, Penaeus monodon, as a model target species. For a promoter, we used the 1,592-bp promoter region of the EF-1alpha gene, a house-keeping gene, of kuruma shrimp Marsupenaeus japonicus. The promoter region was linked to either the gene for green fluorescence protein (GFP) or the gene for chloramphenicol acetyl transferase (CAT). The fusion genes were designated pJEF-GFP and pJEF-CAT, respectively. The pJEF-GFP gene was introduced into fertilized eggs of black tiger shrimp by microinjection and particle gun bombardment. The survival rate of the microinjected eggs was 17.6%, and 1.0% of the treated embryos were found to be GFP-positive. However, the GFP-positive embryos were damaged and embryogenesis did not progress. The survival rate of the particle-bombarded eggs was 60.6%, and 0.42% of the treated embryos were found to be GFP-positive. Ubiquitous GFP expression was observed from 8 hr post-fertilization and these embryos developed and hatched normally. The pJEF-CAT gene was introduced into fertilized eggs of black tiger shrimp using the optimized conditions of the particle gun bombardment. CAT activity was observed from 1 to 7 days post-fertilization, with the highest activities being observed at 5 and 7 days post-hatching.     

White spot syndrome virus (WSSV) detected by PCR in rotifers and rotifer resting eggs from shrimp pond sediments

White spot syndrome virus (WSSV) was detected by PCR-dot blot hybridization in rotifer resting eggs from shrimp Penaeus chinensis culture-pond sediments. It was also detected in rotifers hatched from those eggs. Surface disinfection before analysis indicated that WSSV was probably present within the resting eggs. Results suggested that rotifer resting eggs may be an overwintering reservoir for WSSV in shrimp ponds.     

Nutritional value of brine shrimp cysts as a factitious food for Orius laevigatus (Heteroptera: Anthocoridae)

Decapsulated cysts of the brine shrimp Artemia franciscana were assessed as a factitious food for rearing the anthocorid predator Orius laevigatus. Developmental and reproductive traits of O. laevigatus reared for a single generation on A. franciscana from three geographical locations or on gamma-irradiated eggs of the Mediterranean flour moth Ephestia kuehniella were compared. There was no effect of diet on nymphal survival but nymphal period on E. kuehniella eggs (12.2 days) was 0.7-1.6 days shorter than on the Artemia diets. The predator developed 0.5-1 day faster on cysts from San Francisco Bay (USA) than on cysts from Great Salt Lake (USA) or Macau (Brazil). Fecundity on brine shrimp cysts from different locations was similar to that on flour moth eggs (142-187 eggs/female). The biochemical composition of decapsulated cysts from San Francisco Bay was compared with that of E. kuehniella eggs. Depending on the type of analysis, Artemia cysts contained higher or similar amounts of protein as compared with E. kuehniella eggs, but amino acid patterns were generally similar. Flour moth eggs were almost three times richer in fatty acids than brine shrimp cysts, with some marked differences in fatty acid profiles. Because nutrient imbalances in a diet may be expressed only after several generations of rearing, the predator was cultured for three consecutive generations on A. franciscana cysts from San Francisco Bay. In the third generation on brine shrimp cysts, nymphs took 18% longer to develop, and adults were shorted-lived and about 60% less fecund than those maintained on E. kuehniella eggs. Brine shrimp cysts may be used as a supplement in the mass production of O. laevigatus but may not be a suitable food for long-term culturing of the predator.     

Acquisition of epibiotic bacteria along the life cycle of the hydrothermal shrimp Rimicaris exoculata

The caridean shrimp Rimicaris exoculata dominates the fauna at several Mid-Atlantic Ridge hydrothermal vent sites. This shrimp has an enlarged gill chamber, harboring a dense ectosymbiotic community of chemoautotrophic bacteria associated with mineral oxide deposits. Until now, their acquisition is not fully understood. At three hydrothermal vent sites, we analyzed the epibionts diversity at different moult stages and also in the first stages of the shrimp life (eggs, hatched eggs (with larvae) and juveniles). Hatched eggs associated with young larvae were collected for the first time directly from gravid females at the Logachev vent site during the Serpentine cruise. An approach using 16S rRNA clone libraries, scanning and transmission electron microscopy, and fluorescent in situ hybridization was used. Molecular results and microscope observations indicated a switch in the composition of the bacterial community between early R. exoculata life cycle stage (egg libraries dominated by the Gammaproteobacteria) and later stages (juvenile/adult libraries dominated by the Epsilonproteobacteria). We hypothesized that the epibiotic phylotype composition could vary according to the life stage of the shrimp. Our results confirmed the occurrence of a symbiosis with Gammaproteobacteria and Epsilonproteobacteria, but more complex than previously assumed. We revealed the presence of active type-I methanotrophic bacteria colonizing the cephalothorax of shrimps from the Rainbow site. They were also present on the eggs from the Logachev site. This could be the first ‘epibiotic'' association between methanotrophic bacteria and hydrothermal vent crustacean. We discuss possible transmission pathways for epibionts linked to the shrimp life cycle.     

三峡水库秀丽白虾生长与繁殖生物学特征研究

2012 年11 月至2013 年10 月逐月对三峡水库木洞江段秀丽白虾的生长和繁殖生物学特征进行了研究。结果显示秀丽白虾是一年生的虾类, 寿命为1214 个月。雌性个体的平均体长(37.727.92) mm 显著大于雄性(37.086.59) mm, 具有明显的雌雄异形现象。秀丽白虾的渐近体长(L)为59.33 mm, 生长系数(K)是1.6/年, 生长方程为:Lt=59.33-e-1.6(t+0.081)。雌虾繁殖季节从4 月上旬持续至9 月下旬, 繁殖高峰期为4 月中旬至6 月中旬, 由越冬虾和当年虾两个世代的繁殖群体组成。抱卵雌虾的体长为27.0753.71 mm, 平均体长(L50)为37.06 mm。雌雄性比月变化为1.112.36, 平均为1.46, 群体中雌性个体数量显著多于雄性(P 0.001)。绝对繁殖力(FA)为31294 粒, 相对繁殖力(FR)为(11522)粒/g。绝对繁殖力与体长呈幂函数关系(FA=0.0031L2.8632, r=0.72, n=106), 而与体重呈直线关系(FA=104.63W+9.9534, r=0.79, n=106)。卵径(D)平均值为(1215102) m。为合理保护和利用三峡水库秀丽白虾种群资源, 建议加强虾类捕捞管理, 繁殖期间(49 月)禁止捕捞虾类或限制捕捞强度。       

A smaller particle size improved the oral bioavailability of monkey head mushroom, Hericium erinaceum, powder resulting in enhancement of the immune response and disease resistance of white shrimp, Litopenaeus vannamei

The effects of different particle sizes (100–150, 74–100, and <74 μm) of powder of the dried and ground stipe from the monkey head mushroom, Hericium erinaceum, on the immune response and disease resistance of white shrimp, Litopenaeus vannamei, against the pathogen, Vibrio alginolyticus, were examined. Mushroom powder with a particle size of <74 μm had a significantly higher effect on the disease resistance of shrimp compared to particle sizes of >74 μm. Mortality of shrimp after being injected with V. alginolyticus was particle size-dependent, increasing from 66.7% ± 3.3%–93.3% ± 3.3% with diets containing stipe particle sizes of <74 and 100–150 μm, respectively. The mortality of shrimp fed the diet containing <74-μm stipe powder for 28 days was significant lower than that of shrimp fed with the control diet and the diet containing 74–100-μm stipe powder after being challenged by V. alginolyticus. The optimal concentration of the <74-μm mushroom powder for enhancing the immune response and disease resistance of shrimp was 0.2 μg (g shrimp)^?1 day^?1. No significant change in the total hemocyte count, differential hemocyte count, glutathione reductase, or phagocytic activity was found in shrimp fed the control diet and mushroom powder-containing diet at a level of up to 0.2 μg (g shrimp)^?1 day^?1. Shrimp fed 0.2 μg (g shrimp)^?1 day^?1 of a mushroom-containing diet had a significantly higher disease resistance to V. alginolyticus via an increase in phenoloxidase activity, respiratory bursts, superoxide dismutase activity, and glutathione peroxidase activity. Therefore, a diet containing the stipe powder of monkey head mushroom with a particle size <74 μm at a level of 0.2 μg (g shrimp)^?1 day^?1 was found to enhance the immunity and disease resistance of shrimp.     

Molecular characterization and high expression during oocyte development of a shrimp ovarian cortical rod protein homologous to insect intestinal peritrophins

Penaeoid shrimp oocytes nearing the completion of oogenesis are enveloped in an acellular vitelline envelope and possess extracellular cortical rods (CRs) that extended into the cortical cytoplasm. These cortical specializations are precursors of the jelly layer (JL) of the egg. In searching for highly expressed mRNAs during oogenesis in the marine shrimp (Penaeus semisulcatus), two related cDNAs have been isolated that encode a mature protein of 250 amino acid residues. The deduced amino acid sequences revealed the presence of repeated cysteine-rich domains that are related to the chitin-binding domains of insect intestinal peritrophins. Similar cysteine-rich domains were reported in insect intestinal mucin, crustacean tachycitin, and invertebrate chitinases. The shrimp ovarian peritrophin (SOP) is glycosylated and can bind chitin when extracted from CRs. Its apparent molecular mass in SDS-PAGE is 29-35 kDa and 33-36 kDa, under nonreducing or reducing conditions, respectively. SOP is a major protein of CRs and the JL, and was immunodetected in ovaries; purified CRs; fertilized eggs that were surrounded by a JL matrix; and in the cloudy, whitish flocculent material appearing in sea water immediately after spawning. Immunolocalization in tissue sections determined that SOP was present in oocyte cytoplasm and in extraoocytic CRs. Shrimp expressed SOP mRNA in ovaries at all oocyte developmental stages, whereas expression in the hepatopancreas was restricted to vitellogenic stages. SOP mRNA was abundant in the shrimp ovary and was detected before the presence of the corresponding protein. This is the first demonstration that a protein with similar features to insect intestinal peritrophins is a component of CRs and is therefore a main precursor of the JL of spawned shrimp eggs.     

Beyond Symbiosis: Cleaner Shrimp Clean Up in Culture

Cleaner organisms exhibit a remarkable natural behaviour where they consume ectoparasites attached to “client” organisms. While this behaviour can be utilized as a natural method of parasitic disease control (or biocontrol), it is not known whether cleaner organisms can also limit reinfection from parasite eggs and larvae within the environment. Here we show that cleaner shrimp, Lysmata amboinensis, consume eggs and larvae of a harmful monogenean parasite, Neobenedenia sp., in aquaculture. Shrimp consumed parasite eggs under diurnal (63%) and nocturnal (14%) conditions as well as infectious larvae (oncomiracidia) diurnally (26%). Furthermore, we trialled the inclusion of cleaner shrimp for preventative parasite management of ornamental fish, Pseudanthias squamipinnis, and found shrimp reduced oncomiracidia infection success of host fish by half compared to controls (held without shrimp). Fish held without cleaner shrimp exhibited pigmentation changes as a result of infection, possibly indicative of a stress response. These results provide the first empirical evidence that cleaner organisms reduce parasite loads in the environment through non-symbiotic cleaning activities. Our research findings have relevance to aquaculture and the marine ornamental trade, where cleaner shrimp could be applied for prophylaxis and control of ectoparasite infections.     

Reproductive output of the ornamental shrimp Lysmata vittata (Stimpson, 1860) (Decapoda: Caridea) in wild populations and under different maturation diets

ABSTRACT

Reproductive output parameters (fecundity and egg volume) of Lysmata vittata were examined in a population in northeastern Brazil. Effect of maturation diets on the reproductive output of these shrimp under laboratory conditions was evaluated. Reproductive output was estimated for 25 shrimp collected in the wild. Another 45 pairs of shrimp were used for diet experiments, 15 pairs per treatment (T1: industrialized food, T2: fresh food, T3: mixed diets). For wild population, mean fecundity and egg volume were differed between developmental stages of the eggs. Fecundity was significantly lower in the specimens subjected to T1 diet (267 ± 141 eggs) compared with shrimp from the wild population (393 ± 183 eggs). Egg volume was significantly lower in shrimp subjected to three diets tested compared with those from wild population. Egg loss during embryonic development in L. vittata may be caused by several factors (e.g. aborted development and maternal cannibalism). Fresh food proved to be important for improving reproductive output in L. vittata reared in culture. We emphasize the significance of improving nutritional value and palatability of diets to improve cultivation efficiency. Lysmata vittata can be used as a model organism for future studies aiming to improve the cultivation techniques for shrimp of the genus.

Abbreviations PSH: protandric simultaneous hermaphroditism; ARS: artificial refuge structures; CL: carapace length; EV: egg volume; L: largest diameter of the egg; S: smallest diameter of the egg; T1: industrialized pellets; T2: fresh ingredients; T3: mixed diets; ANCOVA: analysis of covariancr; SD: standard deviation.     

Effect of drying period and soil moisture on egg hatch of the tadpole shrimp (Notostraca: Triopsidae).

Tadpole shrimp (Triops spp.) are potential biological control agents against larval mosquitoes in temporary ponds and flood-irrigated fields. In some rice field situations, however, they may become pests that uproot and eat young rice plants. In cursory observations, it has been reported that tadpole shrimp eggs do not readily hatch on flooding when the soil or substrate containing eggs is moist before flooding. The relationship between drying (moisture content) of soil and tadpole shrimp hatch was determined in studies conducted in mesocosms at the University of California Aquatic and Vector Control Research facilities at Riverside and at Oasis in the Coachella Valley of southern California. In laboratory hatching trials, an increase in hatch of Triops longicaudatus (LeConte) with declining soil moisture content was demonstrated (t = 8.4, P less than 0.001; r2 = 0.76). In field trials in mesocosms at Riverside, egg hatch increased with increased drying period and declining soil moisture content (G = 29.8, P less than 0.01). No hatch of eggs occurred after 3 d of drying, when soil moisture content was high, but a high level of hatching occurred after 7 and 14 d of drying, when soil moisture declined to low levels. At Oasis, soil moisture did not decrease with drying time because of porous soil and capillary action of water from adjacent flooded mesocosms and thick vegetation covering the pond bottoms. Therefore, hatch rates at Oasis were not associated with the length of the drying period (G = 35, P greater than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)