饲料中添加酵母提取物对凡纳滨对虾免疫相关基因表达及抗菌机能的影响

为了评估饲料中添加酵母提取物对凡纳滨对虾免疫灵敏性和平衡性的相关基因表达的影响,以鱼粉含量为24.5%的基础饲料(对照组)及在基础饲料中添加2.5%酵母提取物的实验饲料,分别饲喂凡纳滨对虾56 d,检测两种饲料投喂的对虾在急性感染溶藻弧菌前后鳃组织Toll受体、IM D和溶菌酶mRNA表达量变化及感染后的对虾死亡情况。结果表明:摄食添加酵母提取物饲料的凡纳滨对虾鳃组织中Toll受体mRNA和溶菌酶mRNA表达量均显著高于对照组对虾(P0.05),IMD mRNA表达量与对照组无显著性差异(P0.05)。急性感染溶藻弧菌后,两组对虾鳃组织Toll受体、IMD和溶菌酶mRNA表达量随感染进程均出现显著变化,Toll受体、IMD和溶菌酶mRNA表达量峰值出现的时间分别为24、42和36 h,且摄食添加酵母提取物组对虾各基因的表达量峰值分别为对照组峰值的201.06%、481.46%和276.77%,均显著高于对照组对虾(P0.05)。摄食添加酵母提取物饲料组对虾鳃组织中Toll受体和IM D mRNA表达量在感染溶藻弧菌后12和24 h分别出现显著上调,而对照组对虾鳃组织中Toll受体和IMD mRNA表达量分别在感染弧菌后24和42 h才出现显著上调。两组对虾经溶藻弧菌人工急性感染后72 h内的累积死亡率无显著差异(P0.05)。实验表明,饲料中添加酵母提取物上调了溶藻弧菌感染前凡纳滨对虾Toll受体和溶菌酶mRNA的表达量,提早了溶藻弧菌感染后对虾Toll受体和IMD mRNA表达量上调时间,且增加了对虾Toll受体、IM D和溶菌酶mRNA表达量的峰值,在一定程度上提高了凡纳滨对虾免疫相关基因表达的灵敏性。     

饲料中花生四烯酸水平对凡纳滨对虾免疫相关基因表达及抗菌能力的影响

为了评估饲料中花生四烯酸(arachidonic acid,ARA)水平对凡纳滨对虾免疫相关基因表达及抗菌能力的影响,分别以鱼油和混合植物油为脂肪源,设计了2个系列共9组不同ARA含量的等氮等能,且具备相同饱和脂肪酸(SFA)、单不饱和脂肪酸(MUFA)、多不饱和脂肪酸(PUFA)和n-3/n-6比例的实验饲料,投喂对虾6周后,检测各组对虾在急性感染溶藻弧菌0、24、36和42 h时鳃组织中Toll受体、IMD(immune deficiency)和溶菌酶m RNA表达量,并统计感染后96 h内对虾的死亡情况。结果表明:1对虾摄食以鱼油为脂肪源的饲料[饲料DHA(22:6n-3)和EPA(20:5n-3)含量分别为5.85和3.83 mg/g饲料],其鳃组织中Toll受体、IMD和溶菌酶m RNA表达量均随饲料中ARA含量的升高呈现先升高后下降的变化;0.56(B组)和0.87 mg ARA/g饲料组(C组)对虾溶菌酶m RNA表达量显著高于0.44(A组)、1.02(D组)和0.28 mg ARA/g饲料组(E组)(P0.05);对虾摄食以植物混合油为脂肪源的饲料(DHA和EPA含量分别为3.28和1.87 mg/g饲料),其鳃组织中Toll受体、IMD和溶菌酶m RNA表达量随饲料中ARA含量的升高而升高;1.44 mg ARA/g饲料组(I组)对虾Toll受体m RNA表达量显著高于0.19 mg ARA/g饲料组(F组)(P0.05)。2人工急性感染溶藻弧菌后,各组对虾鳃组织中Toll受体、IMD和溶菌酶m RNA的表达量随感染进程均出现显著变化。摄食以鱼油为脂肪源的饲料时,对虾鳃组织中Toll受体、IMD和溶菌酶m RNA表达量峰值均出现在0.56 mg ARA/g饲料组(B组),且峰值分别出现在感染后24、42和24 h。摄食以混合植物油为脂肪源的饲料时,对虾鳃组织中Toll受体、IMD和溶菌酶m RNA表达量峰值均出现在1.44 mg ARA/g饲料组(I组),且峰值分别出现在感染后24、42和36 h。3各实验组对虾急性感染溶藻弧菌后96 h累积死亡率无显著差异。本研究表明,饲料ARA水平影响凡纳滨对虾免疫相关基因(Toll受体、IMD和溶菌酶)的表达,且ARA调控免疫基因表达的效果受饲料EPA和DHA水平的影响。     

溶藻弧菌VZ5株在凡纳滨对虾育苗中应用的研究

在凡纳滨对虾育苗涵状幼体期以不同频次添加溶藻弧菌VZ5株,并在糠虾幼体后搭配添加芽孢杆菌Bz5株,测定育苗过程中水质、水体中弧菌密度变动以及对虾幼体生长和存活状况,研究溶藻弧菌VZ5株在凡纳滨对虾育苗中应用的效果。结果表明,添加溶藻弧菌的3个处理组仔虾期氨氮浓度显著低于对照组;仔虾期幼体干重分别为0．0463±0．0009、0．0420±0．0006、0．0407±0．0009g／尾,显著高于对照组（0．0253±0．0003g／尾）。滏状幼体期每天添加溶藻弧菌的处理组幼体成活率显著高于其他处理组（P〈0．05）。溶藻弧菌VZ5株能够有效降低水体中氨氮的积累和促进对虾幼体生长。     

饲料锌添加水平对凡纳滨对虾免疫抗菌机能和溶菌酶mRNA及Toll受体mRNA表达的影响

在基础饲料中添加不同水平蛋氨酸锌(添加水平分别为0、50、150 mg Zn/kg)并饲喂凡纳滨对虾,养殖14 d后,取样测定对虾鳃组织中Toll受体mRNA和溶菌酶mRNA的表达水平以及肝胰腺、肌肉和血淋巴中超氧化物歧化酶(SOD)和溶菌酶(LSZ)活性,并进行溶藻弧菌人工急性感染试验。结果表明,凡纳滨对虾肝胰腺及肌肉中锌蓄积水平随饲料锌添加量的增加而显著增加(P<0.05),肝胰腺中锌蓄积更明显。添加50 mg Zn/kg组(锌含量为73.25 mg Zn/kg饲料)对虾鳃组织中的Toll受体mRNA和溶菌酶mRNA表达量均显著高于未添加锌组和添加150 mg Zn/kg组(P<0.05)。添加50 mg Zn/kg组对虾肌肉、肝胰腺和血淋巴中溶菌酶活性显著高于未添加锌组(P<0.05)。添加50 mg Zn/kg组对虾肝胰腺和血淋巴中的SOD活性也显著高于未添加锌组,但与添加150 mg Zn/kg组无显著差异。而肌肉中SOD活性在添加150 mg Zn/kg组中最高。经溶藻弧菌人工急性感染后,添加50 mg Zn/kg组对虾半致死时间和全致死时间大于未添加锌组和添加150 mg Zn/kg组。本研究表明,相比摄食未添加锌组饲料和添加150 mg Zn/kg组饲料,凡纳滨对虾的免疫抗菌机能在摄取添加50 mg Zn/kg(锌含量为73.25 mg Zn/kg饲料)饲料时得到改善。     

中草药对凡纳滨对虾生长、免疫功能的影响

以凡纳滨对虾（Litopenaeus vannamei）为实验对象,基础饵料中分别添加不同水平的中草药（0％、1．0％、2．0％）,结果表明,添加不同剂量的中草药能显著提高凡纳滨对虾的增重率和增长率,显著地降低对虾的饵料系数,对凡纳滨对虾的成活率有极显著提高．能极显著提高对虾血清中溶菌酶和SOD活性,能显著提高ACP活性,极显著提高对虾对溶藻弧菌的免疫保护率。     

凡纳滨对虾弧菌病及其防治方法研究进展

近年来,凡纳滨对虾(Litopenaeus vannamei)养殖生产中弧菌病的发病率越来越高,严重阻碍了我国海水养殖业的发展.文章介绍了凡纳滨对虾养殖过程中容易发生的几种弧菌病及其防治方法的相关研究,并对常见的哈维氏弧菌、溶藻弧菌、霍乱弧菌和副溶血弧菌这4种弧菌的研究进展进行了综述,以期为海水养殖中弧菌病的防治和控制...     

亚硝酸氮和副溶血弧菌对凡纳滨对虾部分免疫指标的影响

在养殖水体中含不同浓度亚硝酸氮的情况下,用副溶血弧菌浸泡感染的方式对凡纳滨对虾(Litopenaeus vannamzei)进行毒性实验;并检测了凡纳滨对虾在亚硝酸氮和弧菌胁迫下的部分免疫指标的变化,研究亚硝酸氮和副溶血弧菌对凡纳滨对虾免疫力的影响.结果表明,实验Ⅰ:亚硝酸氮浓度为0.75、1.50、3.00 mg/L组分别胁迫健康凡纳滨对虾,对虾的血细胞数、血清酚氧化酶活力、酸性磷酸酶活力、超氧化物歧化酶活力和过氧化氢酶活力均显著低于对照组(0.005 mg/L)(P<0.05);10 d时,0.005、0.75、1.50、3.00 mg/L组的对虾死亡率分别达到0%、16.7%、20.0%、26.7%.实验Ⅱ:亚硝酸氮胁迫同时,用副溶血弧菌感染对虾,与实验Ⅰ相比,对虾的各项免疫指标受到的影响更大,其各项免疫指标与相应的实验Ⅰ相比,均有显著的差异(P<0.05);第10天时,0.005、0.75、1.50、3.00 mg/L对虾死亡率分别达到33.3%、40.0%、46.7%、50.0%.可见亚硝酸氮浓度越高,弧菌对对虾免疫力的破坏性越大,对虾的死亡率也越高.     

凡纳滨对虾健康育苗技术的研究

本实验通过水体与育苗池消毒、合理投喂、科学防病以及育苗池水质理化因子(DO、pH、NH 4+-N、NO 2--N、COD)和病原生物(弧菌、WSSV等)的实时监测,对凡纳滨对虾高健康育苗模式做了一定的探索。实验结果显示,整个育苗期间育苗池水溶解氧保持在4.2～5.8mg/L(平均5.01±0.63mg/L),pH保持在8.04～8.38(平均8.13±0.11);NH 4+-N控制在0.15～1.21mg/L(平均0.51±0.40mg/L),NO 2--N控制在0.15～1.21mg/L(平均0.05±0.02mg/L),COD控制在1.56～7.02mg/L(平均4.75±2.18mg/L)。异养细菌数200～91000cfu/mL,弧菌0～6980cfu/mL。投放无节幼体4600×104尾,收获虾苗1280×104尾,成活率达27.8%,且虾苗体质健康,活力旺盛,无携带病毒。     

凡纳滨对虾中发光坎氏弧菌的分离、鉴定及致病性

为研究凡纳滨对虾死亡的致病机理,实验采用TCBS培养基从濒死凡纳滨对虾肝胰腺分离到浅绿色、直径3～7 mm的发荧光菌落;革兰氏染色为阴性短杆菌;经API 20E鉴定,分离株PvL-1与坎氏弧菌参考菌株CAIM249相似度为90％;fitZ序列与坎氏弧菌CP020076相似度为99.31％;gapA序列与坎氏弧菌EF59...     

凡纳滨对虾饲料添加剂的研究进展

凡纳滨对虾(Litopenaeus vannamei)俗称南美白对虾,是目前世界养殖产量最高的三大虾种之一,具有对水环境抗逆能力强、营养要求低、生长速度快,虾体含肉量大、肉质鲜美、营养丰富等诸多优点,自1998年来,在广东、广西、海南等省(区)广为     

Dietary administration of soybean isoflavones enhances the immunity of white shrimp Litopenaeus vannamei and its resistance against Vibrio alginolyticus

The growth response, total hemocyte count (THC), respiratory burst (release of superoxide anion), phenoloxidase (PO) activity, superoxide dismutase (SOD) activity, nitric oxide synthase (NOS) activity and resistance to the pathogen Vibrio alginolyticus were measured in white shrimp (Litopenaeus vannamei) which had been fed diets supplemented with soybean isoflavones (SI) at 0, 5, 10, 20, 40 mg kg^?1 feed for 8 weeks. Dietary SI had no significant influence on survival rate of the shrimp. Significantly greater SGR was observed in treatments with 10, 20 and 40 mg kg^?1 SI supplement than that in control (P < 0.05). Shrimp fed a diet with 20 mg kg^?1 SI showed significant increase in THC, PO activity, respiratory burst activity, SOD activity as well as NOS activity (P < 0.05). Increased THC, respiratory burst activity, SOD activity and NOS activity were also noticed in shrimp fed SI at 10 or 40 mg kg^?1 (P < 0.05). Significant lower cumulative mortality after challenge was observed in shrimp fed diets with SI supplement (10.0–13.3%) than that in control group (36.7%) (P < 0.05). It is therefore concluded that oral administration of SI at 20 mg kg^?1 for 8 weeks could enhance the immune ability of shrimp and improve production in shrimp farming.     

注射溶藻弧菌疫苗对南美白对虾免疫功能的影响

应用溶藻弧菌疫苗注射南美白对虾,测定了弧菌疫苗对南美白对虾免疫指标的影响以及疫苗的免疫保护作用。结果表明,注射溶藻弧菌疫苗后,血清凝集效价增强,24 h时活力达到最高值,尤其是1.08×108cell/ml组的凝集效价升高最明显,为对照组的6.0倍;在24 h时,3个试验组的凝集效价均明显高于对照组(P<0.05)。溶菌活力注射后6 h即达到最高值,1.08×107、1.08×108cell/ml组的溶菌活力明显高于对照组(P<0.05)。抗菌活力在注射后3 h时最高,明显高于对照组。溶血活力在注射疫苗后有所增强,注射后6 h时1.08×106、1.08×107cell/ml组的活力达到最高,而1.08×108cell/ml组12 h时活力达到峰值。注射疫苗对血清超氧化物歧化酶活力基本没有影响。1.08×106、1.08×107、1.08×108cell/ml对南美白对虾的免疫保护率分别为46.3%、61.0%和75.6%。     

Effects of dietary xylooligosaccharides on growth performance,immunity and Vibrio alginolyticus resistance of juvenile Litopenaeus vannamei

A 60‐day feeding trial was conducted to assess the effects of xylooligosaccharides on growth performance, immunity and Vibriosis alginolyticus resistance of juvenile Litopenaeus vannamei. Five diets were formulated to contain 0 (T0), 1 (T1), 2 (T2), 4 (T4) and 6 (T6) g/kg of xylooligosaccharides, triplicate groups of 40 shrimps with initial weight of 0.10 ± 0.01 g were fed to apparent satiation four times daily. At the end of the feeding trail, ten shrimps from each tank were challenged with V. alginolyticus. Results showed that shrimps fed xylooligosaccharides containing diets had higher (p < 0.05) feed efficiency, survival rate and intestinal villi length and area than those of T0. There was no significant difference in final body weight, weight gain rate and chemical compositions of whole shrimp among treatments. Shrimps fed T4 and T6 diets had higher (p < 0.05) phenoloxidase and lysozyme activities than those in T0. Concentration of albumin, phenoloxidase activity and survival rate of shrimps decreased (p < 0.05) after 72 hr of challenge, with SR being greater (p < 0.05) in T2, T4 and T6 than that of in T0. Results demonstrate that supplementing 4–6 g/kg of xylooligosaccharides in diets increased feed efficiency, survival rate, intestinal villi length and area, and improved the resistance of juvenile L. vannamei against V. alginolyticus. This study suggests that xylooligosaccharides could be a potential feed additive in practical diet of juvenile L. vannamei.     

Effects of dietary β‐glucan and glycyrrhizin on non‐specific immunity and disease resistance of white shrimp,Litopenaeus vannamei (Boone) challenged with Vibrio alginolyticus

The white shrimp Litopenaeus vannamei, fed immunostimulant‐free, 0.2%β‐glucan and 0.06% glycyrrhizin diets for 18 days, respectively, were challenged with Vibrio alginolyticus at 6.4 × 10⁴ CFU shrimp^?1. The total haemocyte count (THC), phenoloxidase (PO) activity, respiratory burst (RB) and superoxide dismutase (SOD) activity changes for a 120‐h period were investigated, and shrimp mortality was also recorded. The results showed that PO activity, RB and SOD activity were significantly higher in shrimp fed the two immunostimulant diets after 18 days than those in shrimp fed immunostimulant‐free diets. The THC and SOD activity decreased significantly from 0 to 24 h post challenge, and then reverted to normal levels at 96 and 72 h respectively. The values for PO activity and RB increased from 0 to 48 h post challenge. Compared with those fed the control diets, shrimp fed immunostimulants had significantly higher PO activity and RB values at 120 h post challenge. Mortalities after challenge with V. alginolyticus were significantly lower in shrimp fed with β‐glucan or glycyrrhizin than in those fed with a diet without immunostimulants. It was concluded that dietary β‐glucan and glycyrrhizin increased the shrimp immunity. Furthermore, β‐glucan caused an increase in some immune parameters 12 h earlier than glycyrrhizin after V. alginolyticus challenge.     

饲料添加益生菌对凡纳滨对虾肠道菌群、Toll受体及溶菌酶基因表达及抗感染的影响

以初体重为(7.20±1.38)g的凡纳滨对虾(Litopenaeus vannamei)为研究对象,在室内养殖箱进行3周的养殖实验和2周的哈维氏弧菌(Vibrio harveyi)人工感染实验,其中对照组每日投喂普通商品饲料,实验组每日投喂在普通商品饲料中添加地衣芽孢杆菌(Bacillus licheniformis)1.0×10~8 CFU/m L配制成的实验饲料。目的是研究饲料中添加益生菌对凡纳滨对虾肠道菌群、Toll受体及溶菌酶基因表达量和抗哈维氏弧菌能力的影响。实验结果表明,在饲料中添加地衣芽孢杆菌可显著提高对虾抗哈维氏弧菌感染的能力,其相对免疫保护率为22.22%。与对照组相比,实验组可显著降低对虾肠道内弧菌数量(P0.05)。感染哈维氏弧菌后,实验组溶菌酶(lysozyme,LZM)m RNA的相对表达量在12 h、18 h、24 h、36 h、48 h、72 h均显著高于对照组(P0.05)。实验组感染哈维氏弧菌后在6 h、12 h、18 h、24 h、36 h、48 h、72 h、7 d的Toll受体(Toll receptor)m RNA的相对表达量均显著高于对照组(P0.05)。实验结果提示:饲料中添加地衣芽孢杆菌可有效提高对虾抗哈维氏弧菌感染的能力,这种能力的提高可能是通过降低对虾肠道内的弧菌量,并提高抗病相关基因的表达量实现的。     

Dietary protein level and natural food management in the culture of blue (Litopenaeus stylirostris) and white shrimp (Litopenaeus vannamei) in microcosms

The effect of dietary protein level and natural food management on the production parameters of blue and white shrimp, as well as on water quality, was evaluated in a microcosms system (plastic pools simulating aquaculture ponds). Two experimental trials were carried out in the facilities of DICTUS, University of Sonora, Northwest México. Treatment with low protein diet (LP) consisted of a low protein input (diet with 250 g kg^?1 crude protein) through the culture period; treatment with high protein diet (HP) consisted of a high protein input (diet with 400 g kg^?1 crude protein) through the trial, and finally treatment VP consisted of an adjustment of protein input (diets with 250, 350 or 400 g kg^?1 crude protein), depending on the abundance of biota (zooplankton and benthos) in the system. Each species responded differently to the treatments. For blue shrimp, low protein input resulted in the lowest final body weight (12.9 ± 0.6 g) and biomass (696.0 g pool^?1). Survival and feed conversion ratio were similar in the three treatments. For white shrimp, the best growth, biomass and food conversion ratio were obtained in the low protein input treatment. Water quality parameters such as nitrate, ammonia and organic matter during the two trials, were better for LP and VP treatments. White shrimp seems to have lower protein requirements than blue shrimp. For the blue shrimp culture, adjusting protein input according to natural food abundance (zooplankton and benthos) in the system, seems to be advantageous because of the possibility of getting a production similar to that obtained with a high protein input through the farming period, but at lower feed cost, and with a lower environmental impact. It is concluded that a high protein input through the whole farming period is not the best feeding strategy for any of the two species.     

氨氮胁迫下白斑综合征病毒对凡纳滨对虾的致病性

为了评价养殖水环境中氨氮(NH_4-N)对凡纳滨对虾(Litopenaeus vannamei)的危害性,开展了NH_4-N胁迫对凡纳滨对虾感染白斑综合征病毒(WSSV)后的死亡率、WSSV增殖速率和对虾主要免疫相关酶活性影响的实验。在NH_4-N胁迫质量浓度为15.6 mg·L-1,分别注射2×105和2×106个WSSV粒子,结果显示,NH_4-N胁迫下注射2×105个WSSV粒子的凡纳滨对虾第144小时死亡率达到53.3%,显著高于无胁迫组(40.0%)。对虾鳃组织WSSV荧光定量PCR检测结果显示,NH_4-N胁迫下凡纳滨对虾鳃组织内WSSV的增殖加快。此外,免疫相关酶活性结果显示,NH_4-N浓度突变会促使对虾血清中酚氧化酶(PO)、酸性磷酸酶(ACP)和碱性磷酸酶(AKP)活性短暂升高后持续降低。由此可见,NH_4-N胁迫会加快WSSV在患病凡纳滨对虾体内的增殖,导致更高死亡率,这可能是因为胁迫造成了对虾免疫相关酶活性降低和抗病原感染能力下降。     

Effects of Vibrio harveyi on plasma clotting protein (coagulogen) of white shrimp Litopenaeus vannamei

Blood clotting exhibits various important functions, including the prevention of body fluid loss and invasion of pathogens in shrimp. The effects of pathogenic Vibrio harveyi on plasma of white shrimp (Litopenaeus vannamei) in vitro and in vivo were investigated in this study. The clotting protein (coagulogen) in plasma of white shrimp pre‐incubated with extracellular products (ECP) of V. harveyi was found apparently decreased and fast‐migrated in crossed immunoelectrophoresis (CIE) gels. In addition, the coagulogen had been degraded to many low molecular‐weight protein bands in plasma pre‐incubated with ECP on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) gels. When pre‐challenged with bacterial cells and ECP of V. harveyi, the white shrimp began to die at about 30 and 16 h respectively. Moreover, plasma coagulogen was decreased more obvious in shrimp challenged with ECP than that with bacterial cells as visualized in CIE gels, and total plasma protein in both group of shrimp were all decreased. Haemolymph withdrawn from moribund shrimp pre‐challenged with V. harveyi or its ECP was observed unclottable. However, the addition of clotting factors (transglutaminase and/or Ca²⁺) to these unclottable plasma could apparently promote their re‐clotting ability as jelly‐like solid observed in microtubes. The recovery of clotting ability of plasma from moribund shrimp was due to the reformation of coagulogen (200 kDa) after adding the two clotting factors as shown on CIE and SDS‐PAGE gels. The present results suggest that the infection of V. harveyi in white shrimp may not only degrade coagulogen but also influence the presence of transglutaminase and Ca²⁺ ion.