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Response of lysozyme activity to Vibrio anguillarum infection in different tissues of Scapharca broughtonii

  • Corresponding author: Liqing ZHOU, zhoulq@ysfri.ac.cn
  • Received Date: 2018-12-11
    Accepted Date: 2019-04-25
  • The changes in the marine environment and overfishing have caused imbalances in the ecological environment, making the various diseases of shellfish increasingly prominent, and the diseases caused by pathogenic Vibrio have become more common. In this context, the number of wild resources of Scapharca broughtonii has been sharply reduced, and its breeding industry is in urgent need of improved germplasm. In order to effectively prevent the damage of bacteria and other pathogenic microorganisms on S. broughtonii, we observed the response process of the lysozyme activity in different tissues of S. broughtonii infected by Vibrio anguillarum, and explored the immune function of the lysozyme in the body of S. broughtonii. In this experiment, 20-month-old S. broughtonii. individuals were infected by injecting live bacteria, and 16 individuals were randomly selected. And 1 mL (about 1×109) V. anguillarum suspension was injected into the axe of each individual as the infection group. 16 randomly selected individuals were not injected with V. anguillarum as the control group. The two groups were cultivated in clean seawater for 4、12、24 and 48 h. Each group randomly selected 4 S. broughtonii individuals who were dissected the blood, mantle, gill, axe foot, hepatopancrea and adductor muscle tissues. The concentration of lysozyme was determined by ELISA kit. The results showed that for the invasion of V. anguillarum, the lysozyme content in the blood of the S. broughtonii rapidly increased from the normal low value and maintained a high level, indicating that the blood is one of the main immune tissues of the S. broughtonii body defense pathogen.In the case of no infection, the S. broughtonii mantle always maintained a high lysozyme concentration to prevent the interference of the external water environment. The concentration of lysozyme in infected gills and axe foot are significantly higher than the normal values after the injection about 24 hours, indicating that the mantle, gills and axe foot were the first barrier as the S. broughtonii body to contact with the outside world can also respond to the invasion of pathogenic bacteria, but the response was delayed compared with the blood. The lysozyme content of hepatopancrea and adductor muscle did not change significantly. And it was speculated that hepatopancrea and adductor muscle were not important immune tissues or organs of S. broughtonii. The results of this study can provide relevant parameters for the study of disease resistance and immune mechanism of S. broughtonii.
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  • [1] 王颖, 吴志宏, 李红艳, 等. 青岛魁蚶软体部营养成分分析及评价[J]. 渔业科学进展, 2013, 34(1): 133-139. doi: 10.3969/j.issn.1000-7075.2013.01.020Wang Y, Wu Z H, Li H Y, et al. Analysis and evaluation of nutrition composition in soft tissue of Anadara uropygimelana[J]. Progress in Fishery Sciences, 2013, 34(1): 133-139(in Chinese). doi: 10.3969/j.issn.1000-7075.2013.01.020
    [2] 李娟, 王滨亭, 王秀华, 等. 蚶类产业可持续性发展概述[J]. 食品工业, 2014, 35(11): 236-239.Li J, Wang B T, Wang X H, et al. Outline of Sustainable Development of Ark Shell Industrialization[J]. The Food Industry, 2014, 35(11): 236-239(in Chinese).
    [3] 公衍军, 陈吉祥, 杜萌, 等. 不同类型抗菌素对致病性鳗弧菌外膜蛋白表达的影响[J]. 中国水产科学, 2006, 13(5): 794-799. doi: 10.3321/j.issn:1005-8737.2006.05.016Gong Y J, Chen J X, Du M, et al. Effects of several antibiotics on expression of outer membrane proteins of Vibrio anguillarum W-1[J]. Journal of Fishery Sciences of China, 2006, 13(5): 794-799(in Chinese). doi: 10.3321/j.issn:1005-8737.2006.05.016
    [4] Egidius E. Vibriosis: Pathogenicity and pathology. A review[J]. Aquaculture, 1987, 67(1-2): 15-28. doi: 10.1016/0044-8486(87)90004-4
    [5] 刘志鸿, 牟海津, 王清印. 软体动物免疫相关酶研究进展[J]. 海洋水产研究, 2003, 24(3): 86-90.Liu Z H, Mou H J, Wang Q Y. Research progress of immune related enzymes in mollusca[J]. Marine Fisheries Research, 2003, 24(3): 86-90(in Chinese).
    [6] 张朝霞, 王军, 丁少雄, 等. 贝类免疫学研究新进展[J]. 厦门大学学报(自然科学版), 2006, 45(S2): 90-96.Zhang Z X, Wang J, Ding S X, et al. Recent progresses of shellfish’s immunology[J]. Journal of Xiamen University (Natural Science), 2006, 45(S2): 90-96(in Chinese).
    [7] 刘世良, 麦康森. 贝类免疫系统和机理的研究进展[J]. 海洋学报, 2003, 25(2): 95-105. doi: 10.3321/j.issn:0253-4193.2003.02.011Liu S L, Mai K S. The progress of studies on molluscs Immunological system and mechanism-a review[J]. Acta Oceanologica Sinica, 2003, 25(2): 95-105(in Chinese). doi: 10.3321/j.issn:0253-4193.2003.02.011
    [8] 吴宁, 陈梦玫, 王素芳. 贝类免疫机制的研究进展[J]. 药物生物技术, 2017, 24(1): 68-71.Wu N, Chen M M, Wang S F. Research progress of shellfish immune mechanism[J]. Pharmaceutical Biotechnology, 2017, 24(1): 68-71(in Chinese).
    [9] 刘可君, 刘阳, 潘宝平. 鳗弧菌侵染对青蛤溶菌酶和超氧化物岐化酶活性的影响[J]. 四川动物, 2011, 30(5): 802-804. doi: 10.3969/j.issn.1000-7083.2011.05.036Liu K J, Liu Y, Pan B P. Effect of Vibrio anguillarum on activity of lysozgme and superoxide dismutase in Cyclina sinensis[J]. Sichuan Journal of Zoology, 2011, 30(5): 802-804(in Chinese). doi: 10.3969/j.issn.1000-7083.2011.05.036
    [10] Zhou L Q, Yang A G, Liu Z H, et al. Changes in hemolymph characteristics of ark shell Scapharaca broughtonii dealt with Vibrio anguillarum challenge in vivo and various of anticoagulants in vitro[J]. Fish & Shellfish Immunology, 2017, 61: 9-15.
    [11] 孙虎山, 李光友. 双壳贝类参与免疫防御的体液因子[J]. 海洋科学, 2001, 25(4): 35-36. doi: 10.3969/j.issn.1000-3096.2001.04.011Sun H S, Li G Y. Humoral factors of bivalves participating in immunity defence[J]. Marine Sciences, 2001, 25(4): 35-36(in Chinese). doi: 10.3969/j.issn.1000-3096.2001.04.011
    [12] Cheng T C, Rodrick G E, Foley D A, et al. Release of lysozyme from hemolymph cells of Mercenaria mercenaria during phagocytosis[J]. Journal of Invertebrate Pathology, 1975, 25(2): 261-265. doi: 10.1016/0022-2011(75)90076-2
    [13] Cheng T C. The role of lysosomes in molluscan inflammation[J]. American Zoologist, 1983, 23(1): 129-144. doi: 10.1093/icb/23.1.129
    [14] 魏玉西, 汪靖超, 程殿林, 等. 鳗弧菌(Vibrio anguillarum)胞外产物中蛋白酶的纯化及其性质[J]. 应用与环境生物学报, 2002, 8(4): 414-418. doi: 10.3321/j.issn:1006-687X.2002.04.017Wei Y X, Wang J C, Cheng D L, et al. Purification and property of extracellular proteases from Vibrio Anguillarum[J]. Chinese Journal of Applied and Environmental Biology, 2002, 8(4): 414-418(in Chinese). doi: 10.3321/j.issn:1006-687X.2002.04.017
    [15] 周丽青, 杨爱国, 王清印, 等. 魁蚶血细胞分类及其免疫功能的初步分析[J]. 水产学报, 2013, 37(4): 599-606.Zhou L Q, Yang A G, Wang Q Y, et al. Studies on the hemocytes types and their immunological functions in bloody clam (Scapharca broughtonii)[J]. Journal of Fisheries of China, 2013, 37(4): 599-606(in Chinese).
    [16] 周丽青, 杨爱国, 王清印, 等. 鳗弧菌对魁蚶血细胞形态及免疫功能的影响[J]. 海洋与湖沼, 2014, 45(3): 536-541. doi: 10.11693/hyhz20130300009Zhou L Q, Yang A G, Wang Q Y, et al. Effect of vibrio anguillarum on morphology and Immunological function of blood cells in Scapharca broughtonii[J]. Oceanologia et Limnologia Sinica, 2014, 45(3): 536-541(in Chinese). doi: 10.11693/hyhz20130300009
    [17] Anderson R S, Giam C S, Ray L E, et al. Effects of environmental pollutants on immunological competency of the clam Mercenaria mercenaria: Impaired bacterial clearance[J]. Aquatic Toxicology, 1981, 1(3-4): 187-195. doi: 10.1016/0166-445X(81)90014-X
    [18] Itoh N, Okada Y, Takahashi K G, et al. Presence and characterization of multiple mantle lysozymes in the Pacific oyster, Crassostrea gigas[J]. Fish & Shellfish Immunology, 2010, 29(1): 126-135.
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Response of lysozyme activity to Vibrio anguillarum infection in different tissues of Scapharca broughtonii

    Corresponding author: Liqing ZHOU, zhoulq@ysfri.ac.cn
  • 1. Key Laboratory of Sustainable Development of Marine Fisheries, Ministry of Agriculture and Rural Affairs, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China
  • 2. Shanghai Ocean University, Shanghai 201306, China
  • 3. Key Laboratory of Exploitation and Utilization of East China Sea and Ocean Fishery Resources, Ministry of Argriculture and Rual Areas villages, Shanghai 200090, China
  • 4. Qingdao National Laboratory of Marine Science and Technology Marine Fishery Science and Food Production Process Function Laboratory, Qingdao 266071, China

Abstract: The changes in the marine environment and overfishing have caused imbalances in the ecological environment, making the various diseases of shellfish increasingly prominent, and the diseases caused by pathogenic Vibrio have become more common. In this context, the number of wild resources of Scapharca broughtonii has been sharply reduced, and its breeding industry is in urgent need of improved germplasm. In order to effectively prevent the damage of bacteria and other pathogenic microorganisms on S. broughtonii, we observed the response process of the lysozyme activity in different tissues of S. broughtonii infected by Vibrio anguillarum, and explored the immune function of the lysozyme in the body of S. broughtonii. In this experiment, 20-month-old S. broughtonii. individuals were infected by injecting live bacteria, and 16 individuals were randomly selected. And 1 mL (about 1×109) V. anguillarum suspension was injected into the axe of each individual as the infection group. 16 randomly selected individuals were not injected with V. anguillarum as the control group. The two groups were cultivated in clean seawater for 4、12、24 and 48 h. Each group randomly selected 4 S. broughtonii individuals who were dissected the blood, mantle, gill, axe foot, hepatopancrea and adductor muscle tissues. The concentration of lysozyme was determined by ELISA kit. The results showed that for the invasion of V. anguillarum, the lysozyme content in the blood of the S. broughtonii rapidly increased from the normal low value and maintained a high level, indicating that the blood is one of the main immune tissues of the S. broughtonii body defense pathogen.In the case of no infection, the S. broughtonii mantle always maintained a high lysozyme concentration to prevent the interference of the external water environment. The concentration of lysozyme in infected gills and axe foot are significantly higher than the normal values after the injection about 24 hours, indicating that the mantle, gills and axe foot were the first barrier as the S. broughtonii body to contact with the outside world can also respond to the invasion of pathogenic bacteria, but the response was delayed compared with the blood. The lysozyme content of hepatopancrea and adductor muscle did not change significantly. And it was speculated that hepatopancrea and adductor muscle were not important immune tissues or organs of S. broughtonii. The results of this study can provide relevant parameters for the study of disease resistance and immune mechanism of S. broughtonii.

  • 魁蚶(Scapharca broughtonii)也称“赤贝”,属软体动物门(Mollusca)、瓣鳃纲(Lamellibranchia)、蚶目(Arcoida)、蚶科(Arcidae)、毛蚶属(Scapharca),是一种高蛋白、低脂肪食品[1]。其个大质厚、肉嫩味鲜,自古就被视为珍品,又因含有多种营养的保健功能和独特的风味更受人们青睐。但近年来,随着人们生活水平的提高,对野生贝类资源需求量越来越大,多年无节制的采捕和日益严重的海洋污染,致使贝类的养殖产量和面积日趋减少,魁蚶的种质资源也受到严重威胁[1-2]。随着蚶类养殖业的发展,蚶类养殖在构筑现代海洋产业体系、推动贝类传统产业高端化发展、集约高效利用海洋资源、增强海洋经济可持续发展方面起重要的拉动和示范作用。因此,魁蚶滩涂养殖发展迅猛起来,其养殖面积和产量不断扩大,魁蚶已成为我国海洋养殖贝类的主要品种。然而,高密度养殖水体易滋生大量病原菌,其中致病性鳗弧菌(Vibrio anguillarum)引起的水产动物疾病较为严重,且在世界范围流行[3-4]。因此,魁蚶养殖业的健康可持续发展亟需选育出抗逆抗病能力强的魁蚶品种。

    大量研究认为软体动物缺乏免疫球蛋白,其体液免疫主要是通过血清中的非特异性酶和免疫因子进行的[5-8]。溶菌酶属非特异性免疫因子,是生物体内一种具有杀菌作用的重要抗菌蛋白,在机体内起到防御病菌和消化的作用。目前,国内已有学者对青蛤(Cyclina sinensis)溶菌酶活性变化进行研究,并对其免疫作用机制进行了相关报道[9]。有关鳗弧菌侵染魁蚶后对其不同组织溶菌酶活性变化的研究尚未见有正式报道。本文旨在分析魁蚶不同组织中溶菌酶活性对鳗弧菌侵染的响应情况,通过观察溶菌酶在鳗弧菌侵染后不同时间的表达量来探究魁蚶不同组织对于细菌侵入的防御能力,进一步确定魁蚶的主要免疫组织及其在免疫功能中发挥的作用。本实验的结果可为魁蚶抗病优良品种的选育提供有效的参数。

    • 实验用20月龄魁蚶于2018年4月4日购自青岛生鲜市场,选取无病无伤、应激反应灵敏、吃食和排便正常的个体约40只,个体大小规格差异小,称量其体质量平均值12.14 g,平均壳长43.21 mm,平均壳宽33.56 mm,平均壳高29.14 mm。将其暂养于120 cm3充气过滤海水养殖槽中,每隔24 h换一次海水,每次换水1/2,用软管虹吸吸除残便等杂质,水温18 °C左右,盐度30,海水pH 8.0。暂养期间,投喂三角褐指藻(Phaeodactylum tricornutum Bohlin),暂养一周后进行实验。

    • 鳗弧菌菌种由中国海洋大学海洋生命学院提供,采用2216E海水培养基28 °C振荡培养24 h获得高浓度菌液,镜检确定鳗弧菌扩繁成功,培养菌液放置冰箱4 °C保存备用。注射前用高压灭菌的0.8%生理盐水稀释菌液至1×109个鳗弧菌/mL。

    • 海水培养基:50 mL新鲜过滤海水,酵母膏0.05 g,蛋白胨0.25 g,高压灭菌;1×PBS (0.01 mol磷酸缓冲液);灭菌蒸馏水;MAS(modified alserver’s solution)抗凝剂:葡萄糖20.80 g,柠檬酸钠8.00 g,乙二胺四乙酸二钠(EDTA)含量为3.36 g,氯化钠22.50 g,溶于1 000 mL蒸馏水,pH值调至7.5,4 °C保存备用。本研究团队前期对血液抗凝剂进行了筛选,发现MAS抗凝剂既能较好的保存血细胞形态,有效抗凝,又能有效保护免疫相关酶的活性[10],因此,本研究采用MAS抗凝剂来保存采集的血液样品。江莱生物贝类溶菌酶酶联免疫分析(enzyme-linked immunosorbent assay,ELISA)试剂盒:封板膜;96孔酶标被板;0.3 mL的标准品6管,浓度依次为0、0.5、1、2、4、8 U/L,10 mL的酶标试剂;6 mL的样品稀释液;6 mL的显色剂A液;6 mL的显色剂B液;6 mL的终止液;25 mL 20× 浓缩洗涤液。酶标板微孔底部包被有兔抗鸡的多克隆抗体,所采用的一抗是鸡蛋清中纯化的全长天然溶菌酶蛋白,二抗可以特异性地捕获外源溶菌酶,再与HRP标记的检测抗体结合,形成抗体-抗原-酶标抗体复合物,经过彻底洗涤后加底物TMB的方法进行显色检测。

    • 暂养一周后,取32只魁蚶阴干,平均分成2组,即感染组和对照组,每组16只个体。感染组中每只魁蚶的斧足处分别注射鳗弧菌液1 mL(生理盐水稀释至约1×109个鳗弧菌),对照组注射高压灭菌的0.8%生理盐水1 mL,然后将这2组魁蚶分开放回养殖槽内暂养。

    • 分别于注射4、12、24和48 h后,随机取4只感染组魁蚶个体和4只对照组魁蚶个体,从每只个体的前闭壳肌血窦中吸取魁蚶血液,然后依次取外套膜、鳃、斧足、肝胰腺及闭壳肌等样品,分别装入1.5 mL离心管。血液加等量MAS抗凝剂,其他组织经PBS洗涤之后,用液氮迅速冷冻保存,再转移至−80 °C超低温冰箱中保存备用。标本融化后,用匀浆器在冰上将标本匀浆充分。按照试剂盒流程要求,以3 000 r/min离心20 min左右,仔细收集上清液,用于溶菌酶活性的测定。

    • 魁蚶溶菌酶活性的测定采用江莱生物公司的ELISA-Kit试剂盒,应用双抗体夹心法测定魁蚶不同组织中溶菌酶吸光度值(OD值),试剂盒中含有溶菌酶标准溶液,用于制作标准曲线,先用标准液的浓度与OD值计算出标准曲线的直线回归方程式,再将样品的OD值代入方程式,计算出样品浓度,最后乘以稀释倍数,即为样品中溶菌酶的实际浓度,详细测定步骤参照试剂盒说明书。

    • 实验数据输入Excel和Prism进行数据整理和分析,对魁蚶的6种组织进行单因素方差分析和多重比较,P<0.05表示差异显著。

    2.   结果
    • 对照组魁蚶外套膜的溶菌酶含量最高,血液中溶菌酶含量偏低,其他组织的溶菌酶含量略低于外套膜的溶菌酶含量,且不同个体各组织溶菌酶含量在一定范围内上下波动(图1)。

      Figure 1.  Lysozyme concentration in different tissues of S. broughtonii

    • 魁蚶在注射鳗弧菌溶液后,不同时间段血液的溶菌酶含量变化结果显示,12 h时感染组溶菌酶含量迅速升高,24 h时感染组溶菌酶含量高于对照组,在48 h时溶菌酶达到最高活性,感染组明显高于对照组(图2)。

      Figure 2.  Changes of lysozyme concentration in blood of S. broughtonii in control group and infected group at different times

    • 魁蚶在注射鳗弧菌溶液后,不同时间段外套膜的溶菌酶含量变化结果显示,鳗弧菌侵染魁蚶后,外套膜内的溶菌酶含量没有明显的变化规律,在24 h时,感染组与对照组出现显著差异(P<0.05),外套膜感染组溶菌酶含量整体低于对照组,且2组均在24 h时达到峰值(图3)。

      Figure 3.  Changes of lysozyme concentration in mantle of S. broughtonii in control group and infected group at different times

    • 鳗弧菌侵染对魁蚶鳃部,其溶菌酶含量变化显示,鳃部感染鳗弧菌后其溶菌酶活性有明显先上升然后下降的趋势,24 h时溶菌酶含量达到最大值,48 h时溶菌酶含量降低,有恢复正常鳃部溶菌酶含量的趋势(图4)。但单因素方差分析和多重比较结果显示,对照组和感染组没有显著性差异。

      Figure 4.  Changes of lysozyme concentration in gill of S. broughtonii in control group and infected group at different times

    • 魁蚶注射鳗弧菌溶液后,不同时间段斧足的溶菌酶含量变化结果显示,鳗弧菌侵染魁蚶后,感染组斧足内的溶菌酶浓度也有明显先上升后下降的趋势,在24 h时达到峰值,且高于正常魁蚶的溶菌酶含量。对照组斧足溶菌酶含量呈现逐渐升高的趋势,因而在12和48 h时,感染组与对照组出现显著性差异(P<0.05)(图5)。

      Figure 5.  Changes of lysozyme concentration in axe foot of S.broughtonii in control group and infected group at different times

    • 鳗弧菌侵染后魁蚶肝胰腺溶菌酶含量变化结果显示,对照组魁蚶和感染组魁蚶溶菌酶活性整体先呈上升趋势,在24 h时,溶菌酶含量都达到最大值,几乎与对照组的溶菌酶含量持平,24 h后,两组溶菌酶含量均逐渐下降。但单因素方差分析和多重比较结果显示,对照组和感染组也没有显著性差异(图6)。

      Figure 6.  Changes of lysozyme concentration in hepatopancreas of S. broughtonii in control group and infected group at different times

    • 感染组魁蚶闭壳肌溶菌酶活性的含量变化整体呈上升趋势,在48 h时闭壳肌溶菌酶含量达到最高,在4、12和24 h时,感染组溶菌酶含量均低于对照组。实验期间,对照组与感染组差异不明显,感染组溶菌酶含量的整体平均值接近正常魁蚶个体闭壳肌的溶菌酶含量值。单因素方差分析和多重比较结果同样显示,对照组和感染组没有显著性差异(图7)。

      Figure 7.  Changes of lysozyme concentration in adductor muscle of S. broughtonii in control group and infected group at different times

    3.   讨论
    • 溶菌酶是软体动物体内参与免疫防御的重要酶类[11],在贝类的炎症反应中起核心作用[12-13]。本研究发现魁蚶血液在无感染的情况下溶菌酶含量相对偏低,一旦有病原微生物侵入,溶菌酶含量会应激性升高,且随着时间的延长,感染组溶菌酶含量逐渐高于对照组溶菌酶含量,进一步印证了魏玉西等[14]的观点,即血液在病原微生物入侵后其溶菌酶含量会增加,来水解鳗弧菌中的碱性蛋白。由此可见,溶菌酶在消灭病原微生物中起比较重要的免疫作用,是魁蚶体内一种重要的免疫蛋白酶。本研究中血液溶菌酶含量变化的结果与周丽青等[15]研究血细胞免疫功能的结果是一致的,发现魁蚶在注射病原菌 12 h后白细胞数目明显增多,且白细胞对入侵的病原菌有很强的吞噬及消化分解能力[16],因此,推测溶菌酶主要存在于血液中的白细胞内。Anderson等[17]发现,栖息在受污染海域的硬壳蛤(Mercenaria mercenaria)的血细胞中溶菌酶含量高于其他组织细胞溶菌酶含量,同时还发现血细胞对病原菌的清除能力会受海水污染程度的影响,说明血液组织中血细胞在抵御病原菌侵染时起重要的作用,溶菌酶是杀死和清除病原菌的重要免疫蛋白,但血细胞和溶菌酶等清除病原菌的能力受生化环境影响是有限的。

    • 外套膜和斧足是魁蚶与外界接触的第一道屏障,其组织细胞中需要维持较高常量的溶菌酶含量及其他免疫相关因子以应对环境中微生物的不良刺激。长牡蛎(Crassostrea gigas)外套膜中发现至少有2种类型的溶菌酶在不同pH值、离子强度和温度条件下,其抗菌活性不同,二者共同担负起防御细菌侵染的作用[18]。鳃丝位置在外套膜之后,鳗弧菌感染魁蚶后,鳃丝中所测溶菌酶含量的变化似乎没有规律,感染组和对照组也没有显著性差异。估计是实验操作时,鳃组织的剖取在抽提血液之后,鳃组织中残留的血液量不同会导致不同魁蚶个体鳃部溶菌酶含量存在差异,同时也说明是流经鳃丝的血液在免疫防御中起重要作用。闭壳肌的主要功能是运动,维持贝壳的开合,肝胰腺的主要功能是饵料消化,二者都在贝壳、外套膜、斧足和鳃等组织的防线之内。当鳗弧菌经过外套膜和血细胞等的防御后,侵入闭壳肌、肝胰腺的病原菌数量就会减少。因此,闭壳肌和肝胰腺感染组的溶菌酶含量相对对照组含量偏低。综上所述,魁蚶组织因与外界不良刺激所接触的频率和强度不同,组织细胞中生成溶菌酶的含量也就不同,对病原菌的防御和抵抗能力也有所差异。

    • 本研究在不同时间段里,感染组溶菌酶含量整体呈先上升后下降的趋势,这可能与Cheng等[14]提出的有机体在接触细菌后,溶酶体酶诱导体液免疫因子并释放到血清的滞后作用有关。但随着贝类体内病原物被凝聚、分解、吞噬等作用的完成,其酶的活力逐步回落[9]。此外,对照组的溶菌酶含量也会发生变化,有时甚至高于感染组,这在外套膜、斧足、肝胰腺和闭壳肌中表现得尤为明显,因此,此结果一方面进一步证实这4种组织不是主要的免疫组织,另一方面也证明魁蚶不同组织在不同的时间段中溶菌酶含量可能存在生理性的变化,而并非一成不变的,这为今后探讨魁蚶溶菌酶在不同发育阶段或不同生存环境中的变化提供了参考价值。

    4.   结论
    • 在魁蚶抵抗病原菌入侵过程中,血液是最主要的免疫组织。外套膜、鳃、斧足这3种组织器官,作为魁蚶与外界接触的第一道防线,需维持一定量溶菌酶或其他免疫因子来抵抗外界不良因素的刺激,在鳗弧菌等病原微生物的刺激下,魁蚶机体适当增长的溶菌酶含量会产生一定的防御作用,以保护机体免受病原菌侵害。肝胰腺和闭壳肌对照组的溶菌酶含量变化波动较大,且随病原菌侵入,感染组溶菌酶含量变化并不表现出明显的规律,且与对照组差异不显著,这可能与生理性变化相关,说明肝胰腺和闭壳肌不是魁蚶的重要免疫组织或器官。综上所述,本研究结果可为魁蚶在免疫机理方面的研究提供重要的参考价值。

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