Non-specific immune response of bullfrog <Emphasis Type="Italic">Rana catesbeiana</Emphasis> to intraperitoneal injection of bacterium <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>

Non-specific immune response of bullfrog Rana catesbeiana to pathogenic Aeromonas hydrophila was studied to 60 individuals in two groups. Each bullfrog in bacterium-injected group was injected intraperitoneally （i.p.） with 0.2 ml bacterial suspension at a density of 5.2 × 10^6 CFU/ml, while each one in control group injected i.p. with 0.2 ml sterile saline solution （0.85%, w/v）. Three bullfrogs in both groups were sampled at 0, 1, 3, 7, 11, 15 and 20 days post-injection （dpi） for the evaluation of non-specific immune parameters. It was observed that intraperitoneal injection of A. hydrophila significantly increased the number of leucocytes and that of NBT-positive cells in peripheral blood. Significant increases in serum bactericidal activity and serum acid phosphatase activity were also observed in the bacterium-injected frogs when compared with those in the control group. However, a significant reduction was detected in vitro in phagocytosis activity of peripheral blood phagocytes. No significant difference in changes in the number of peripheral erythrocytes, serum superoxide dismutase （SOD） activity, and lysozyme activity was detected between the two groups. It is suggested that bullfrogs may produce a series of non-specific immune reactions in response to the A. hydrophila infection.     

Immune response of Pseudosciaena crocea to the injection of Vibrio alginolyticus

For the investigation of anti-infection immune response of Pseudosciaena crocea, 160 healthy fish samples were categorized into infected and control groups. Each individual fish in the infected group was injected intraperitoneally (i.p.) with 0.2 ml bacterial suspension of Vibrio alginolyticus in density of 2×107 CFU/ml, while each individual in the control group was injected i.p. with 0.2 ml sterile saline solution (0.85%). It was observed that the artificial injection of V. alginolyticus significantly inc...     

Immune response of <Emphasis Type="Italic">Pseudosciaena crocea</Emphasis> to the injection of <Emphasis Type="Italic">Vibrio alginolyticus</Emphasis>

For the investigation of anti-infection immune response of Pseudosciaena crocea, 160 healthy fish samples were categorized into infected and control groups. Each individual fish in the infected group was injected intraperitoneally (i.p.) with 0.2 ml bacterial suspension of Vibrio alginolyticus in density of 2×10⁷ CFU/ml, while each individual in the control group was injected i.p. with 0.2 ml sterile saline solution (0.85%). It was observed that the artificial injection of V. alginolyticus significantly increased the number of erythrocytes, leucocytes, lymphocytes in peripheral blood as well as peripheral serum antibacterial activity and antibody titer of large yellow croaker, and significantly reduced the number of peripheral blood granulocytes as compared with those in the control group. No significant difference in acid phosphytase and superoxide dismutase activity of serum was detected between the two groups. It is suggested that non-specific immune factors including leucocytes and anti-bacteria substance in peripheral blood played important role at the initial stage of infection, and specific immune factors such as antibody then played important role in response to anti-infection at the latter stage. Supported by the National High Technology Research and Development Program of China (863 program, No. 2007AA09Z115) and Technology Program of Xiamen (No. 3502Z73019)     

3种免疫途径对嗜水气单胞菌灭活疫苗保护作用的影响

应用间接ELISA方法,研究经嗜水气单胞菌疫苗浸泡、口服、注射免疫后鳜皮肤、肠道黏液和血清中抗体的变化关系,揭示鳜对3种免疫途径的免疫应答效果。结果显示:黏液抗体产生快(7 d达到峰值),但持续时间短,抗体水平低(213);血清抗体产生慢(28 d方达峰值),但持续时间长,抗体水平高(225)。不同方式免疫鳜后均产生远高于对照组的抗体滴度(P<0.05)。注射组产生的抗体水平高(225),免疫保护率最理想(70.6%);口服组相对另两个实验组,其抗体峰值(215)和免疫保护率(41.2%)均较低;浸泡组在皮肤黏液产生水平和溶菌酶含量方面,产生较好的免疫效应,分别为213和178μg/mL,但相对保护率仅为47.1%,低于注射组的70.6%。浸泡和口服组可诱发局部黏膜免疫,在抗体动态变化方面表现出相似的规律且峰值时间早于注射组。初步推断浸泡和口服可以直接刺激鱼体黏膜组织产生局部的特异性抗体。     

三丁基锡对罗氏沼虾血清中免疫酶活力的影响

将罗氏沼虾(Macrobrachium rosenbergii)在含0.1、0.2、0.4 mg/L三丁基锡(TBT)的水体中浸浴48 h,检测TBT对罗氏沼虾血清中部分免疫酶活力的影响。结果表明:12 h内高剂量(0.4 mg/L)可诱导超氧化物歧化酶(SOD),酶活力差异有统计学意义(P0.01),至48 h时,诱导作用逐步解除;12 h内各剂量组血清中酸性磷酸酶(ACP)活力均表现抑制效应,酶活力差异有统计学意义(P0.05),而高剂量组在48 h时TBT对ACP表现为诱导效应,酶活力高于对照组,差异有统计学意义(P0.05);低剂量组(0.1 mg/L)TBT对罗氏沼虾血清中AKP活力的影响无统计学意义(P0.05),而中剂量组(0.2 mg/L)和高剂量组(0.4 mg/L)AKP活力表现为先抑制后逐渐转为诱导的效应,。     

壳寡糖对吉富罗非鱼幼鱼生长性能、非特异性免疫及血液学指标的影响

用添加质量分数0.00(对照组)、0.10%、0.30%、0.50%和0.70%壳寡糖的饲料饲喂初始体重(3.81±0.23)g的吉富罗非鱼幼鱼(Oreochromis niloticus)10周,研究不同浓度壳寡糖的添加对吉富罗非鱼幼鱼的生长、非特异性免疫功能以及血脂指标的影响。结果表明:与对照组相比,在4个不同浓度添加组中,添加质量分数0.50%壳寡糖能显著提高幼鱼的增重率(P<0.05)、特定生长率(P<0.05),并降低饲料系数(P<0.05);添加质量分数0.3%和0.5%壳寡糖能显著提高幼鱼抗嗜水气单胞菌感染的能力(P<0.01);添加质量分数0.10%、0.30%、0.50%壳寡糖能显著提高幼鱼血清中碱性磷酸酶活性(P<0.05)。同时,各浓度的壳寡糖均能明显提高幼鱼血清中溶菌酶和超氧化物歧化酶活性以及抗嗜水气单胞菌感染的能力(P<0.05),并降低幼鱼血清中总胆固醇和低密度脂蛋白胆固醇水平(P<0.05)。在本实验条件下,添加壳寡糖可提高吉富罗非鱼幼鱼生长性能、饲料利用率、非特异性免疫功能和调节血脂水平,添加量以质量分数0.30%~0.50%为宜。     

Tyrosinase, a new innate humoral immune parameter in large yellow croaker （Pseudosciaena crocea R）

We evaluated the immune response to infection with a pathogen in large yellow croaker(Pseudosciaena crocea Richardson).The fish were given an intraperitoneal(i.p.) injection of Vibrio parahaemolyticus or sterile sea water(control).We collected blood sera from the fish 0.17,1,2,4,8,12,or 16 d after injection(dpi).We measured tyrosinase activity and the concentrations of lysozyme,NOS,and antibodies.Serum tyrosinase activity was significantly higher at 0.17 and 4 dpi than in the control group,and peaked at 8 d...     

Isolation and characterization of AHL-degrading bacteria from fish and pond sediment

Quorum sensing(QS)disruption is considered as a potential alternative strategy to combat bacterial diseases in aquaculture.In this study,we isolated and identified bacteria degrading QS molecules from pond sediment and fish intestine.A total of 132 strains were obtained in the enrichment culture,of which two strains were identified as Enterobacter sp.f003 and Staphylococcus sp.sw120,being isolated from the fish intestine and pond sediment,respectively.We found that strains f003 and sw120 could degrade acyl-homoserine lactones(AHLs)and cause no hemolysis of sheep red blood cells.The AHL lactonase(aiiA)homologous gene in the two strains was detected in PCR amplification and the high-degrading activity to N-hexanoyl-L-homoserine lactone(C6-HSL)and AHLs secreted from pathogenic Aeromonas hydrophila was assessed.Meanwhile,the artificial infection of cyprinid Carassius auratus gibelio with intraperitoneal injection showed that the two strains were avirulent.Therefore,the obtained indigenous bacteria are candidate probiotics against pathogenic A.hydrophila in aquaculture.     

寡糖对三黄鸡生产性能及免疫机能的影响

用96只1日龄广东三黄鸡,研究了饲料添加0.2%或0.3%的寡糖对肉鸡生产性能与免疫机能的影响。结果表明:饲料中添加0.2%的寡糖能显著提高11~30日龄肉鸡的增重、免疫器官指数及新城疫H I抗体滴度(P<0.05),血液白细胞吞噬率和血清IgG含量亦高于对照组,但组间差异不显著;添加0.2%或0.3%的寡糖能明显改善全期的饲料转化率(P<0.05)。     

不同温度下罗非鱼人工感染海豚链球菌后血液生化指标及组织病理

在28℃、33℃两种温度下,用海豚链球菌人工感染尼罗罗非鱼后,测定尼罗罗非鱼血液生化指标。结果显示:两个温度水平病鱼的血清总蛋白(TP)随时间延长都呈现升高趋势;谷丙转氨酶(ALT)、谷草转氨酶(AST)活性随时间延长先升高后降低再升高,33℃实验组ALT、AST活性比28℃实验组达峰值时间短,并显著高于对照组(p<0.05);尿素、钾(K+)含量随时间延长逐渐升高后降低,钠(Na+)含量先升高后降低再升高,肌酐含量呈上升趋势;血清碱性磷酸酶(AKP)活性28℃实验组呈先升高后降低趋势,33℃实验组呈升高趋势;超氧化歧化酶(SOD)活性随时间延长先升高后降低再升高,33℃实验组各个时间点基本高于28℃实验组;球蛋白(GLO)随感染后时间延长呈现升高趋势,至120 h,显著高于对照组(p<0.05),但温度组间各时间点没有显著差异(p>0.05)。组织病理学观察发现,33℃实验组肝细胞、肾小管上皮细胞水肿变性;28℃实验组肝细胞出现嗜酸性小体;肾小管上皮细胞出现嗜酸性的红色小体,颗粒变性。表明感染海豚链球菌的尼罗罗非鱼肝脏、肾脏均有损伤,造成功能障碍,高温时损伤更严重。     

Analysis of IL-2-1ike factor in lymphocyte culture supernatant of olive flounder, Paralichthys oliveaceus

To study immune mechanism of fish lymphocyte we performed a proliferation assay and ELISA using monoclonal antibody against human IL-2. The result showed that an interleukin-2 (IL-2)-like factor was detected in the supernatant of plant haemoglutinin (PHA)-stimulated lymphocyte culture from peripheral blood, spleen and head kidney of olive flounder,Paralichthys olivaceus. The quantities of IL-2-like factor in the supernatant from different lymphoid tissues were quite different. The IL-2 like factor in the supernatant from cultured head kidney lymphocytes was much higher than those of peripheral blood lymphocytes and spleen lymphocytes (P<0.01). The IL-2 activity was found in either mouse thymocyte proliferation assay or flounder head kidney lymphocyte proliferation assay and shown to have obvious enhancing effect on proliferation of the above two types of cell. The recombinant human IL-2, (rhIL-2) was able to stimulate flounder thymocyte proliferation and used to detect the IL-2 receptor (IL-2R) on the surface of flounder lymphocyte. The cross-reaction between the lymphocytes of flounder peripheral blood and CD25(IL-2R) was detected with flow cytometry and shown that the percentage of CD25-positive cell in peripheral blood was 7.74±0.67%. This work was supported by National “973” Project G1999012003, G19999012006.     

TCDD对斑马鱼脂质过氧化作用的初步研究

将150条斑马鱼随机等分为5组:2,3,7,8-四氯代二苯并二噁英(TCDD)4个染毒组(染毒剂量分别为0.1μg/L、0.2μg/L、0.4μg/L和0.8μg/L)和空白对照组,水质接触染毒5d后,采用分光光度法测定其肝组织中丙二醛(MDA)含量和超氧化物歧化酶(SOD)、谷胱甘肽-S-转移酶(GST)的活力,研究TCDD对斑马鱼的脂质过氧化作用。结果发现,TCDD各剂量染毒组MDA含量均有所增加,其中0.2μg/L组差异显著(P<0.05),0.4μg/L和0.8μg/L剂量组差异极显著(P<0.01);各染毒组SOD活性均有所下降,其中0.2μg/L、0.4μg/L和0.8μg/L剂量组差异极显著(P<0.01);各染毒组GST活力均降低且差异极显著(P<0.01)。一定剂量的TCDD能引起MDA含量增加,使SOD和GST活力降低,对斑马鱼具有脂质过氧化作用。     

Effects of Dietary Vitamin C on the Immune Function of Shrimps, Penaeus chinensis

Prepared in this experiment were six groups of diets, i.e. VC0, VC1, VC2, VC3, VC4 and VC5 with the contents of vitamin C (VCmg(100g)-1 diet) of 0, 100, 200, 400, 800, and 1200 respectively. It was found that vitamin C increased the concentrations of immunoglobulin-like (IgG-like, IgA-like and IgM-like) substances in the serum of Penaeus chinensis after a feeding period of 3 weeks. The differences among groups were significant (P＜ 0.01), but there was no difference in the contents of complement3-1ike and complement4-1ike substances in the serum (P ＞0.05). Phenoloxidase (PO) activity in the serum of VC3 group shrimps was higher than that of VC0 and other groups, but no significant difference was observed between VC0 group and other groups. Furthermore, bactericidal activity of the serum to Vibrio parahaemolyticus in shrimps fed with the VC1 diet was higher than that in the other groups (P＜ 0.01), while no difference was demonstrated among all groups for the bactericidal activity to Vibrio alginolyticus (P ＞0.05). It is, therefore, suggested that vitamin C (100-400 mg(100 g) -1 diets) could be used as an immunostimulant of P. chinensis.     

Roles of polysaccharide from <Emphasis Type="Italic">Branchiostoma belcheri</Emphasis> in anti-DNA oxidation and anti-tumor activity in S180 mice

In this study, we isolated a polysaccharide from Branchiostoma belcheri (PBB) by enzymatic protein hydrolysis and alcohol precipitation. We investigated the effects of PBB supplementation on DNA oxidation and growth of the transplanted tumor cells Sarcoma (S₁₈₀) in mice. Sixty healthy Kunming mice weighing between 18 and 25 g were randomly assigned to 6 groups, each consisting of 10 animals. All the mice, except for the blank control group, were inoculated with S180 sarcoma cells into the axilla of the left foreleg. PBB was given to mice by gavage at doses of 0 (model control), 25, 50, or 100 mg/kg b.w. in 0.2 ml saline for 30 days. The fifth group of S₁₈₀-mice was given cytoxan (50 mg/kg) by peritoneal injection as a positive control group. The animals had free access to food and water. The mice were sacrificed after the final treatment and blood was quickly collected. Spontaneous and oxidized DNA damage of peripheral lymphocytes induced by H₂O₂ were analyzed by SCGE. O⁶-methyl-guanine (O⁶-MeG) was measured by high-performance capillary zone electrophoresis. The average tumor weights (0.856–1.118 g) of the three PBB groups were significantly lower than that of the model control group (1.836 g) (p<0.05). The tumor inhibition ratios of the PBB groups were 39.1%–53.4% and similar to the cytoxan positive group (57.5%). There were no significant differences in spontaneous DNA damage in peripheral lymphocytes among the groups. The oxidative DNA damage induced by 10 μmol/L H₂O₂ in the 50 and 100 mg/kg b.w. groups were 246.1 AU and 221.7 AU, respectively, both of which were significantly lower than that in the model group (289.0 AU; p<0.05). The plasma concentrations of O⁶-MeG in the 25, 50, and 100 mg/kg supplemented groups were 2.09 μmol/L, 1.86 μmol/L, and 1.63 μmol/L, respectively, all of which were significantly lower than that of the model group (2.67 μmol/L; p<0.05). These results indicated that PBB may have antioxidative activity and thus reduce oxidation-induced DNA damage.     

Studies on Antiviral and Immuno-Regulation Activity of Low Molecular Weight Fucoidan from Laminaria japonica

The antiviral activity in vitro and in vivo and the effect of the immune system of two fucoidan fractions with low molecular weight and different sulfate content from Laminaria japonica(LMW fucoidans) were investigated in order to examine the possible mechanism. In vitro, I-type influenza virus, adenovirus and Parainfluenza virus I were used to infect Hep-2, Hela and MDCK cells, respectively. And 50% tissue culture infective dose was calculated to detect the antiviral activity of two LMW fucoidans. The results indicated that compared with the control group, 2 kinds of LMW fucoidans had remarkable antiviral activity in vitro in middle and high doses, while at low doses, the antiviral activity of 2 kinds of LMW fucoidans was not statistically different from that in the blank control group. And there was no statistically difference between two LMW fucoidans in antiviral activity. In vivo, LMW fucoidans could prolong the survival time of virus-infected mice, and could improve the lung index of virus-infected mice significantly, which have statistical differences with the control group significantly(p 0.01). However, the survival time of the two LMW fucoidans was not statistically significant(p 0.05). In this study, it was shown that both of two LMW fucoidans(LF1, LF2) could increase the thymus index, spleen index, phagocytic index, phagocytosis coefficient and half hemolysin value in middle and high doses, which suggested that LMW fucoidans could play an antiviral role by improving the quality of immune organs, improving immune cell phagocytosis and humoral immunity.     

CpG-DNA对异育银鲫(Carassius auratus gibelio)免疫细胞活性的诱导作用

探索含CpG基序的未甲基化寡脱氧核苷酸(ODNs)对鱼类的免疫调节,将人工合成的鱼免疫刺激剂寡脱氧核苷酸序列1670(含一个CpG基序)、1670-D(含两个CpG基序)加入到离体培养的异育银鲫的头肾巨噬细胞和外周血白细胞中,采用MTT法测定其对鲤外周血白细胞增殖的影响,NBT还原法和Griess试剂显色法测定对头肾巨噬细胞的呼吸爆发的影响。结果显示,1670可显著(浓度为20.0μg·mL-1)或极显著(浓度为0.1、1.0、10.0μg·mL-1)提高异育银鲫巨噬细胞的氧呼吸爆发活性,但对氮呼吸爆发活性和外周血白细胞的增殖无显著作用;1670-D可显著(浓度为0.1μg·mL-1)或极显著(浓度为1.0、10.0、20.0μg·mL-1)增强异育银鲫巨噬细胞的氧呼吸爆发活性和外周血白细胞的增殖,且可极显著(浓度为1.0、10.0、20.0μg·mL-1)提高异育银鲫巨噬细胞的氮呼吸爆发活性。表明特定序列、特定作用剂量的CpG-DNA可增强体外培养的异育银鲫巨噬细胞和外周血白细胞的非特异性免疫应答。     

10种CpG-ODNs对罗非鱼抗海豚链球菌感染的作用

采用人工合成的10种未甲基化CpG-ODNs序列分别注射健康的吉富罗非鱼Oreochromis niloticus,48 h后注射海豚链球菌(Streptococcus iniae)进行人工感染,分别于感染24 h和48 h后检测鱼体血清溶菌酶(Lysozyme,LSZ)、超氧化物歧化酶(Superoxide dismutase,SOD)和碱性磷酸酶(Alkaline phosphatase,AKP)的活性,并记录死亡率。结果显示:注射ODN-205的罗非鱼LSZ、AKP和SOD活性变化均不显著(P>0.05),推测注射ODN-205后罗非鱼能较好的维持机体的平衡,增强机体的抗菌能力;注射ODN-1681、1670、2133和2006的罗非鱼LSZ活性逐渐降低,而注射ODN-1826、2143、1668、2102、1669的罗非鱼LSZ活性呈现先升高后降低的趋势。AKP活性的变化趋势同LSZ活性变化大体相似,而SOD活性前后变化不显著(p值>0.05)。注射CpG-ODNs的实验组抑制海豚链球菌感染的作用差异显著(p值<0.05),其中以注射ODN-205组的死亡率最低(18%),其次为注射ODN-1670、1681、2006和2133组,死亡率均低于30%,而ODN-1669组和生理盐水组的死亡率均超过90%。实验表明,注射一定量的ODN-205、1670、1681、2006和2133能明显提高吉富罗非鱼抗海豚链球菌感染能力。     

Effects of Hypoxia on Dopamine Concentration and the Immune Response of White Shrimp （Litopenaeus vannamei）

Effects of hypoxia on the dopamine concentration and the immune response of White Shrimp Litopenaeus vannamei were studied. The results showed that hypoxia had significant effects on the concentration of dopamine (DA) in the haemolymph, haemocyte count, phenoloxidase activity, phagocytic activity of haemocytes and bacteriolytic and antibacterial activity in the haemolvmph (P<0.05). The concentration of the dopamine in haemolymph reached its maximum in the 3.0 and 1.5mgL-1 DO groups at 12h and 6h, and then returned to normal after 24h and 12h, respectively. All immune parameters decreased with the reduction of dissolved oxygen. Total haemocyte count (THC), the hyaline cells and semi-granular cells in the 3.0mgL-1 DO group became stable after 12h,while granular cells did so after 24h, The THC and different haemocyte count (DHC) in the 1.5mgL-1 DO group became stable alter 24h. Phenoloxidase activity and bacteriolytic activity in the 3.0 and 1.5mgL-1 DO groups reached their stable levels after 24 h and 12 h respectively, while phagocytic activity and antibacterial activity became stable after 24 and 12, and 36 and 24 It, respec-tively. It was also indicated that the changes of dopamine concentrations in haemolymph, haemocyte count and phenoloxidase activ-ity were obviously related to the exposure time under hypoxic conditions.     

Effects of dietary vitamin C on the immune function of shrimps, Penaeus chinensis

Prepared in this experiment were six groups of diets, i.e. VC0, VC1, VC2, VC3, VC4 and VC5 with the contents of vitamin C (VCmg(100 g)⁻¹ diet) of 0, 100, 200, 400, 800, and 1200 respectively. It was found that vitamin C increased the concentrations of immunoglobulin-like (IgG-like, IgA-like and IgM-like) substances in the serum of Penaeus chinensis after a feeding period of 3 weeks. The differences among groups were significant (P<0.01), but there was no difference in the contents of complement3-like and complement4-like substances in the serum (P>0.05). Phenoloxidase (PO) activity in the serum of VC3 group shrimps was higher than that of VC0 and other groups, but no significant difference was observed between VC0 group and other groups. Furthermore, bactericidal activity of the serum to Vibrio parahaemolyticus in shrimps fed with the VC1 diet was higher than that in the other groups (P<0.01), while no difference was demonstrated among all groups for the bactericidal activity to Vibrio alginolyticus (P>0.05). It is, therefore, suggested that vitamin C (100–400 mg(100 g)⁻¹ diets) could be used as an immunostimulant of P. chinensis.     

Effects of water-current speed on hematological,biochemical and immune parameters in juvenile tinfoil barb,Barbonymus schwanenfeldii(Bleeker,1854)

This study examines the effect of water-current speed on hematological,biochemical and immune parameters in juvenile tinfoil barb(B arbonymus schwanenfeldii). Blood samples were taken on days 1,23 and 45 from control fish and from two training groups maintained at current speeds of 0.06 bl/s(body length per second),0.66 bl/s,and 1.92 bl/s,respectively. Significantly increased red-blood-cell counts and hematocrit were observed in the post-training groups on days 23 and 45. Significantly increased hemoglobin concentrations were also observed in the 1.92 bl/s group on days 23 and 45. In contrast,values of mean corpuscular volume were significantly lower in the 1.92 bl/s group than in the other groups on day 45. Nitroblue-tetrazolium-positive cells and lysozyme and superoxidase dismutase activities in the plasma increased significantly with increasing training intensity on days 23 and 45. Antibacterial activities were significantly increased in the trained groups compared with the control group on day 23; significantly elevated alkaline phosphatase activity was observed in the 1.92 bl/s groups on day 45. Therefore,training intensities of 0.66 and 1.92 bl/s enhanced the blood oxygen-carrying capability and plasma immune parameters of juvenile tinfoil barbs.