Isolation and characterization of microsatellites in Pacific white shrimp Penaeus (Litopenaeus) vannamei

Five polymorphic microsatellite loci were characterized for Penaeus (Litopenaeus) vannamei. Loci were isolated using a partial Sau3A1 genomic library by the sequencing of randomly selected clones and by a biotinylated (CT)₁₀ and (GT)₁₀ probes screening procedure. The last strategy resulted in the most useful data. About 40% of the clones showed a previously reported satellite/microsatellite (PVS1), reducing the chance of finding new microsatellite regions. Whereas two of the microsatellite loci with more than 10 alleles will be useful for mating analysis in a breeding program, the others might prove useful for population genetic studies.     

Proteomic analysis of differentially expressed proteins in Penaeus vannamei hemocytes upon Taura syndrome virus infection

To understand molecular responses of crustacean hemocytes to virus infection, we applied 2-DE proteomics approach to investigate altered proteins in hemocytes of Penaeus vannamei during Taura syndrome virus (TSV) infection. At 24 h postinfection, quantitative intensity analysis and nano-LC-ESI-MS/MS revealed 11 forms of 8 proteins that were significantly up-regulated, whereas 9 forms of 5 proteins were significantly down-regulated in the infected shrimps. These altered proteins play important roles in host defense (hemocyanin, catalase, carboxylesterase, transglutaminase, and glutathione transferase), signal transduction (14-3-3 zeta), carbohydrate metabolism (acetylglucosamine pyrophosphorylase), cellular structure and integrity (beta-tubulin, beta-actin, tropomyosin, and myosin), and ER-stress response (protein disulfide isomerase). Semiquantitative RT-PCR and Western blot analysis confirmed the upregulation of 14-3-3 at both mRNA and protein levels. Interestingly, several altered protein spots were identified as fragments of hemocyanin. Mass spectrometric analysis showed that the hemocyanin spots at acidic and basic regions represented the C- and N-terminal hemocyanin fragments, respectively. As three-quarters of C-terminal fragments were up-regulated, whereas two-thirds of N-terminal hemocyanin fragments were down-regulated, we therefore hypothesize that C- and N-terminal hemocyanin fragments may have differential roles in hemocytes. Further investigation of these data may lead to better understanding of the molecular responses of crustacean hemocytes to TSV infection.     

凡纳滨对虾肠道红杆菌科细菌富集碳源筛选及其定向分离

【目的】红杆菌科(Rhodobacteraceae)细菌为凡纳滨对虾肠道微生物的优势类群,在健康对虾肠道中具有较高的相对丰度,是指示对虾健康的关键类群,探究对虾肠道红杆菌科细菌定向富集和分离方法,可为对虾养殖益生菌菌剂的研发提供基础。【方法】利用16S rRNA基因高通量测序技术研究不同碳源添加对凡纳滨对虾肠道中红杆菌科细菌的富集作用,筛选对红杆菌科细菌有显著富集作用的碳源;利用纯培养技术从红杆菌科细菌富集的样品中定向分离红杆菌科细菌,并对其进行鉴定和遗传多样性分析。【结果】添加短链脂肪酸(乙酸、丙酸、丁酸、戊酸)和碳酸氢钠对红杆菌科细菌有显著富集作用,主要富集到Cribrihabitans、Tritonibacter、Rhodovulum、Ruegeria、Sagittula和Thalassobius属相关菌株;对红杆菌科细菌相对丰度最高的样品进行稀释涂布培养,共分离纯化出303株细菌,分属于2门12科,其中红杆菌科细菌为主导类群共119株,主要包括Tritonibacter (90株)、Phaeobacter (25株)、Sulfitobacter (1株)、Ruegeria (1...     

饲料中添加壳聚糖季铵盐对凡纳滨对虾生长及非特异免疫的影响

实验研究壳聚糖季铵盐对凡纳滨对虾生长性能及非特异性免疫能力的影响。在饲料中分别添加0、0.05%、0.10%、0.15%和0.20%的壳聚糖季铵盐, 制成5组等氮等能饲料。将900尾[初体质量(3.820.34) g]健康的凡纳滨对虾随机分成5组(45尾4平行), 养殖时间56d。结果表明:饲料中添加壳聚糖季铵盐显著影响凡纳滨对虾的生长, 0.15%实验组凡纳滨对虾的增重率和特定生长率最佳(P0.05)。饲料中添加壳聚糖季铵盐0.1%、0.15%和0.20%能显著提高凡纳滨对虾血清溶菌酶和碱性磷酸酶及酚氧化酶的活性(P0.05)。饲料中添加壳聚糖季铵盐可显著提高凡纳滨对虾抗副溶血弧菌感染的能力(P0.05), 0.15%组的保护效果最好, 其相对免疫保护率为33.24%。壳聚糖季铵盐能显著提高凡纳滨对虾的生长性能和抗病能力, 本实验条件下适宜的添加量为0.15%。     

亚热带河口养殖塘沉积物有机碳矿化及其影响因子

亚热带河口的养殖塘是温室气体二氧化碳(CO2)的重要排放源,但其沉积物有机碳矿化动力学特征目前尚未厘清.以我国东南沿海亚热带闽江、木兰溪以及九龙江河口区的6个凡纳滨对虾(Penaeus vannamei)养殖塘为研究对象,对沉积物取样,并进行为期60 d的室内厌氧培养,对养殖塘沉积物CO2累积产量曲线利用一阶动力学模型...     

模拟阴天对南美白对虾养殖水体理化及其生长指标的影响

为探索阴天对南美白对虾(Penaeus vannamei Boon)养殖水体理化及其生长指标的影响, 实验采用黑色遮荫网模拟阴天弱光环境对南美白对虾进行遮荫实验, 设置遮荫组与对照组各5个平行, 实验周期7周。遮荫组采用黑色遮荫网交替性遮荫, 即第1、第3、第5、第7周遮荫, 第2、第4、第6周曝光, 对照组正常养殖。结果显示: 遮荫组与对照组的pH、DO、Chl. a、 -N、NH₃-N、 -N、 -N及 -P的浓度在遮荫处理下具有显著性差异(P<0.05), Chl. a、 -N及 -P的浓度具有明显的波动性, 表现为遮荫时Chl. a的浓度极显著降低(P<0.01), -N、 -P的浓度极显著升高(P<0.01); 曝光时遮荫组Chl. a的浓度较遮荫时升高, -N的浓度极显著降低(P<0.01), -P浓度的增长幅度降低, 此时遮荫组与对照组Chl. a、 -N及 -P的浓度无显著差异。 -N的浓度在实验前5周, 遮荫组高于对照组, 第6、7周对照组高于遮荫组。 -N浓度在整个实验周期中则是遮荫组高于对照组。南美白对虾生长指标结果显示: 两个组生长指标具有显著性差异。研究表明: 妨害南美白对虾生长的主要是NH₃-N, 然后是 -N; 藻类可间接促进南美白对虾生长。遮荫处理抑制藻类吸收养殖水体中的有毒离子、恶化南美白对虾的生长环境并危害对虾的生存、生长。     

回转式凡纳滨对虾温室集约化养殖池氨化菌与硝化菌数演变状况

研究了回转式凡纳滨对虾温室集约化养殖池中氨化菌与硝化菌的演变状况.在回转式凡纳滨对虾温室集约化养殖试验中,养殖全过程不换水、不用药,利用净水网草片、复合微生物制剂、水循环与水生植物净化水质,适时排污,使养殖水体的微生物指标与水化学指标有效控制在虾生长合适范围内.试验期间,248号池水中主要微生物指标平均值如下：氨化细菌为1080000±2411556 cfu/mL,亚硝化细菌898±1124cfu/mL,硝化细菌为1700±759 cfu/L;250号池水中主要微生物指标平均值如下：氨化细菌为881000±1793815 cfu/mL,亚硝化细菌1306±1550 cfu/mL,硝化细菌为2250±2047 cfu/L.同时,实验表明,利用净水草网片能取得良好的净化水质的效果和具有抗不利环境冲击的能力.利用这种养殖模式和相应的管理,养殖用水得到有效的控制,养殖效果良好.     

Immune responses of Litopenaeus vannamei to thermal stress: a comparative study of shrimp in freshwater and seawater conditions

The effects of hypothermal (22–16?°C) and hyperthermal (22–28?°C) stress on the immune system responses of the shrimp Litopenaeus vannamei cultured in either freshwater or seawater were measured and compared. The following immune system indicators were measured for comparison: total hemocyte count (THC), activity of phenoloxidase (PO), nitric oxide synthase (NOS), superoxidase (SOD), and malondialdehyde (MDA) content. Thermal stress significantly decreased THC in both freshwater and seawater shrimp within 6–12?h (P?0.05). After hypothermal stress, all shrimp had a significantly lower THC level than their prechallenge levels (P?0.05). Under both types of thermal stress, the activity of PO, NOS, and SOD first increased and then decreased. After 48?h of thermal stress, shrimp PO and NOS activity decreased in both freshwater and seawater. After 48?h of thermal stress, the reduction in the SOD activity in the hemolymph of freshwater shrimp was greater than that in seawater shrimp. During exposure to stress, the MDA content in freshwater shrimp was significantly higher than in seawater shrimp, which demonstrated that lipids in freshwater shrimp were more susceptible to peroxidation than those in seawater shrimp, particularly at low temperatures. Large temperature fluctuations, particularly sudden cooling, should be avoided when rearing L. vannamei because of high rates of lipid peroxidation and decreased immunity. These effects are more marked in freshwater than in seawater.     

饲料中铜水平对凡纳滨对虾免疫相关基因表达和抗菌能力的影响

在饲料中添加0、30和50 mg Cu/kg饲料的蛋氨酸铜,投喂凡纳滨对虾(Litopenaeus vannamei)7、14和21d,检测对虾体组织铜蓄积、免疫相关基因(Toll受体mRNA和溶菌酶mRNA)表达水平和免疫抗菌能力的变化。结果表明:凡纳滨对虾肝胰腺铜含量随着饲料蛋氨酸铜添加量增加及投喂时间延长而显著增加(P0.05);对虾肌肉的铜含量显著低于肝胰腺的铜含量。饲料中铜水平对凡纳滨对虾肌肉、血淋巴及肝胰腺中溶菌酶活性无显著影响(P0.05)。对虾组织SOD活性因饲料中铜水平和投喂时间变化显著,添加30 mgCu/kg组对虾肌肉、血淋巴和肝胰腺中SOD活性在第21天时显著高于其他两组(P0.05)。饲料中铜水平对凡纳滨对虾鳃组织中溶菌酶mRNA表达水平无显著影响,但显著影响鳃组织Toll受体mRNA表达水平(P0.05)。第7天时凡纳滨对虾Toll受体mRNA表达水平随着饲料铜水平升高而显著升高(P0.05);第14和第21天时,Toll受体mRNA表达水平在摄食添加30 mg Cu/kg组最高。人工急性感染溶藻弧菌(Vibrioalginolyticus)实验表明,第7天时,摄食添加50 mg Cu/kg组凡纳滨对虾全致死时间和半致死时间长于未添加铜组和添加30 mgCu/kg组,但在第14天,摄食添加30 mg Cu/kg组的全致死时间和半致死时间最长。研究表明饲料铜添加水平不但影响组织中铜的蓄积,还影响凡纳滨对虾SOD活性和Toll受体mRNA表达水平,从而影响机体的抗弧菌能力。     

Population Genetic Structure of Pacific White Shrimp (<Emphasis Type="Italic">Litopenaeus vannamei</Emphasis>) from Mexico to Panama: Microsatellite DNA Variation

Genetic variation and population structure of wild white shrimp (Litopenaeus vannamei) from 4 geographic locations from Mexico to Panama were investigated using 5 microsatellite DNA loci. The genetic diversity between populations was indicated by the mean number of alleles per locus and mean observed heterozygosity, which ranged from 7.4 to 8.6 and from 0.241 to 0.388, respectively. Significant departures from Hardy-Weinberg equilibrium were found at most locations at each locus, with the exception Guatemala at Pvan0013, and were caused by high heterozygote deficiencies. Genetic differences between localities were detected by pairwise comparison based on allelic and genotypic frequencies, with the exception of locus Pvan1003. Significant pairwise F _ST values between locations and total F _ST showed that the white shrimp population is structured into subpopulations. However, population differentiation does not follow an isolation-by-distance model. Knowledge of the genetic diversity and structure of L.vannamei populations will be of interest for aquaculture and fisheries management to utilize and preserve aquatic biodiversity.