【作者】 杨翠华；

【导师】 王清印； 孔杰；

【作者基本信息】 中国海洋大学 ， 海洋生物学， 2007， 博士

【摘要】 中国对虾是我国最重要的养殖经济虾类之一。但是九十年代以来,由于疾病和养殖环境的恶化,中国对虾的池塘存活率和养殖产量受到了严重的影响。为了提高养殖中国对虾的生长性状和抗白斑综合症病毒(简称WSSV)的能力,2004年中国水产科学研究院黄海水产研究所在全国水产技术推广总站的支持下,开始和挪威水产研究所遗传育种中心(AKVAFORSK, Genetics Center, AS)合作实施中国对虾的选择育种计划。本研究以中国对虾为材料,采用不平衡巢式设计方法和人工授精技术,每尾雄虾分别与2-3尾雌虾交尾,共构建了57个全同胞家系(分别属于26个父系半同胞家系)。所有家系均在控制条件下育苗、养殖和进行性状测试。分别测定了不同群体的抗WSSV能力和非特异免疫抗性,计算了这些性状的遗传力和遗传相关,并对不同群体的杂交和近交结果进行了分析。结果如下:1中国对虾早期体重和抗WSSV性状的遗传力及遗传相关测定了26个半同胞家系感染WSSV后的存活时间和死亡时的体重。结果显示,在平均体重1.56g时,中国对虾抗WSSV性状遗传力估计值(h2±S.E.)是0.10±0.03,体重的遗传力估计值(h2±S.E.)是0.18±0.04,两者的表型相关系数为0.15±0.00,检验极显著;但是,两者间的遗传相关系数为0.08±0.23,经检验相关不显著。结合成体测定的生长性状和池塘存活率,根据60:5:35(生长:池塘存活率:抗WSSV能力)的选择指数计算综合育种值,进行了家系预留种。选择群体的平均存活时间为108.83h,高于全群均值7.56h。2中国对虾不同群体及其杂交组合抗WSSV能力的比较分析根据洄游路线、越冬及其产卵地点的不同,将从不同地点采集的中国对虾的自然种群划分为不同地理群体。在遗传学上,每个地理群体代表了不同的遗传资源。为了比较乳山(RS)群体、青岛(QD)群体、日照(RZ)群体、朝鲜半岛南海岸野生群体(SKW)、朝鲜半岛南海岸养殖群体(SKC)和本实验室培育的两个选育群体(黄海一号/HH1和即抗98/JK98),以及不同地理群体及其杂交组合抗WSSV性状的差异,建立了不同地理群体及其杂交组合的家系,并在仔虾平均体重1.51g时进行人工感染WSSV试验。结果显示,SKW♂×JK98♀交尾组合感染WSSV后存活时间最长,为103.87h;纯系繁育群体中,QD群体感染病毒后存活101.73h,抗病能力最强,而RZ群体存活时间最短,仅85.53h。本试验结果表明,各个地理群体对WSSV的易感性存在差异,利用群体间杂交能提高大多数组合的抗病能力。3近交对中国对虾早期体重和抗WSSV性状的影响过量的捕捞和不合理的人工育苗措施,使中国对虾野生和养殖群体的遗传多样性均出现不同程度的降低,业已引起水产科技工作者的重视。但是迄今为止,尚未见近交对中国对虾体重和抗病性状影响的报道。本试验利用相似环境条件下养殖的40个野生对虾产生的家系和3个兄妹交配产生的家系,定量测定了平均体重1.43~1.58g时1460尾中国对虾早期体重和感染WSSV后存活时间的近交衰退系数。结果显示,野生对虾家系组的平均体重和平均存活时间分别为1.58±0.01g和100.43±0.68h,而兄妹交家系组平均体重和平均存活时间分别是1.43±0.04g和85.84±1.70h,两组间的平均体重和平均存活时间均存在极显著差异(P<0.01)。野生对虾家系组体重和存活时间的表型相关系数为0.16±0.00,而兄妹交家系组两者间的表型相关系数为0.20±0.00,两组间表型相关系数差异不显著(P>0.05)。近交系数每增加10%,体重和感染WSSV后存活时间分别衰退-3.80±0.17%和-5.81±0.11%,与近交能够降低生长和疾病抗性的观点相一致。这表明,在选择育种和种质资源保护过程中,都应该保证基础群体遗传背景最大化,并应选择合适的育种路线,从而有效控制近交。4中国对虾六项免疫相关组分的遗传力和遗传相关大量研究已证明,血液中的某些免疫组分在对虾抗病和抗逆过程中发挥着重要的调节作用。精确的估计这些性状的遗传参数,特别是遗传力和遗传相关,对于开展遗传育种工作是非常重要的。本试验在控制条件下建立了中国对虾51个全同胞家系,并养殖到平均体重7.64g。每个家系随机捕捞15尾对虾抽取血液,共测定了765尾对虾血清中的总蛋白浓度(PC)、氧合血蓝蛋白浓度(HC)、酚氧化酶(PO)、超氧化物歧化酶(SOD)、酸性磷酸酶(ACP)和碱性磷酸酶(ALP)等血液中与免疫相关酶类的活性,并以最优线性无偏估计(BLUP)理论为基础,计算了上述六项免疫相关组分的估计遗传力、各项性状间的遗传相关和表型相关。结果表明,当中国对虾平均体重7.64g时,血清中上述六项免疫组分的正常含量分别是153.88±1.65mg/ml,36.04±0.71 U/ml,165.24±4.93 U/ml,333.33±2.74 U/ml,52.51±2.15 U/100ml和61.68±2.61U/100ml。雌性和雄性中国对虾间的六种免疫相关组分差异均不显著(P>0.05)。六项血液免疫相关组分的遗传力估计值(h2)分别是0.00±0.13,0.09±0.22,0.03±0.20,0.30±0.20,0.63±0.32和0.39±0.25。对遗传参数剖分结果同时显示,在不同养殖环境下养殖的家系间性状差异较大,六项血液免疫相关组分的共同环境效应(c2)分别为0.10±0.06,0.20±0.11,0.21±0.10,0.00±0.00,0.02±0.01和0.04±0.01。本研究中,除SOD活性外,几乎所有其他性状间的遗传相关、环境相关和表型相关均为正值。结果表明,虽然中国对虾血液中某些免疫相关组分的遗传力估计值很低,但是大多数免疫性状通过选择育种能够得到改善,从而使选育群体整体非特异性免疫水平得到提高。同时,该部分利用家系估计育种值间的表型相关,估计了早期感染WSSV后存活时间与成体六项免疫组分间的遗传相关,结果显示,存活时间与PC、PO和ACP活性间表现为弱正相关,相关系数分别为0.093±0.516、0.058±0.687和0.044±0.760,而存活时间与HC、SOD和ALP活性间表现为负相关,相关系数分别为-0.179±0.209、-0.268±0.057和-0.110±0.441,该结果表明,中国对虾早期抗WSSV性状与成体免疫性状间不存在显著的相关性,这可能与两者使用的材料不同有关,也可能与对虾机体感染疾病和正常状态下免疫活性的表达不同有关。5中国对虾两个地理群体及其杂交组合血液免疫组分活性的比较利用在相似条件下建立的49个家系,比较了黄渤海(YB)群体、朝鲜半岛南海岸(SK)群体及其杂交组合对虾血液免疫组分的活性。结果显示,SK群体的各项免疫组分含量均高于YB群体,并且除HC和SOD活性外所有杂交组合的免疫组分含量均高于YB群体。几乎所有的杂交组合都显示出正的杂交优势,SK×YB组合的杂交优势均为正,并在HC、PO和ALP活性方面优于其反交组合。结果表明,YB群体和SK群体在许多免疫组分方面存在差异,这一结果为群体间杂交以提高抗逆性提供了基础资料。6控制养殖条件下中国对虾6项免疫组分的近交衰退分析定量测定了中国对虾血清中总蛋白浓度、血蓝蛋白浓度、酚氧化酶活性、过氧化物歧化酶、酸性磷酸酶活性和碱性磷酸酶活性六项免疫指标的近交衰退系数。结果显示,兄妹交配产生的F2家系,近交系数为25%,中国对虾的五项免疫指标均低于不存在近交的F1家系,并且在酚氧化酶活性、酸性磷酸酶活性和碱性磷酸酶活性方面存在极显著差异(P<0.01)。近交系数每增加10%,六项免疫指标分别衰退-1.74±0.13%、-4.75±0.23%、-10.50±0.31%、0.62±0.09%、-25.42±0.21%和-21.06±0.30%。结果表明,近交能够影响对虾免疫性状的活性。更多还原

【Abstract】 Chinese shrimp Fenneropenaeus chinensis was one of the important penaeus. However, since 90 times the pond survival and the total output of F. chinensis have been in a low level because of the breakout of many diseases and the deterioration of the cultural environment. Nowadays the reasonable selection breeding program should be built immediately to product seedling with the rapid growth, the strong resistance to disease and adverse. In 2004, Yellow Sea Fisheries Research Institute of Chinese Fishery Sciences Academy (YSFRI) in collaboration with National Fisheries Technical Extension Center (NFTEC) and Akvaforsk Genetic Center (AFGC) initiated“Selective breeding program of Fenneropenaeus chinensis”to demonstrate the advanced selective breeding technology, as applied to develop the Norwegian salmon in Norway and GIFT tilapia in the Philippines. 57 full-sib families (representing 26 paternal half-sib families), which were cultured in the controlled environment were challenged with White Spot Syndrome Virus (WSSV). In this thesis many key questions, i.e. WSSV resistance and the hemolymph analytes, of different populations were tested. And heritability and genetic correlations of many characters were evaluated. Moreover, effects of outbreeding and inbreeding were compared in this study. The following is the results in detail.1 Genetic (co)variation of body weight and resistance to WSSV in young F. chinensisWSSV has been one of the most economically significant pathogens affecting hatchery-reared and wild shrimp world-wide. The survival time, i.e. from all animals contacted virus to their death, and their body weight were recorded in this study. The estimate of heritability (h2±S.E.) for WSSV resistance was 0.10±0.03 when the average body weight of F.Chinensis was 1.56g. Estimated heritability of body weight was 0.18±0.04. Phenotypic correlation between survival time and body weight was different significantly, but genetic correlation between them was not significant. And their correlation coefficients were 0.15±0.00 and 0.08±0.23, respectively. Combining with the results of growth and pond survival ratio, composite breeding value had been computed by the selective index 60:5:35(body: pond survival: resistance to WSSV). The average survival time of all families was 108.83h, which was higher than that of the selected familes 101.27h.2 Comparison resistance to WSSV among the different geographic populations and their hybridization of F. chinensisChinese shrimp is distributed along the coasts of the northern seacoast of China and the Korean Peninsula. According to their migration routes, spawning and overwintering places, F. chinensis are divided into many geographic variety populations. These populations represent potentially different genetic resources in their economical traits important to aquaculture and for their contribution to hybridization breeding programs. To determine their characteristics of resistance to WSSV, mates were made between and within five populations (Rushan population (RS), Qingdao population (QD), Rizhao population (RZ), the wild population in south coast of Korean Peninsula (SKW) and the cultured population in south coast of Korean Peninsula (SKC)) belong to four geographic variety populations, two breeding populations (Huanghai NO.1 (HH1) and Jikang98 (JK98)), and the offspring was reared to 1.51g. The results indicated that the mating combination of SKW♂×JK98♀is best in resistance to WSSV among all populations, and the average survival time is 103.87h post infecting WSSV. QD population is best in resistance to WSSV among intra-population, and the average survival time is 101.73h. RZ population is most sensitive to WSSV in all populations, and the average survival time is 85.53h. The results of this study suggest that hybridization is an important tool to increase resistance to WSSV of cultured shrimps.3 Effects of inbreeding on growth and WSSV resistance of the juvenile Chinese shrimpTo date, few studies had investigated the effects of inbreeding on growth and disease resistance, although inbreeding depression of such traits could play an important role in the evolution and ecology in Chinese shrimp. Effects of inbreeding on body weight and survival time post infection WSSV of juvenile shrimp were quantified on this study. All families were reared under the similar environment conditions, and the differences of growth time between two groups were not significant. The mean body weight and mean survival time of the wild shrimp group were 1.58±0.01g and 100.43±0.68h, and those of the shrimp group with inbreeding coefficient 25% were 1.43±0.04g and 85.84±1.70h, respectively. The differences of body weight and survival time post infection WSSV between two groups were all extremely significant (P<0.01). The difference of phenotypic correlation was not significant (P>0.05) between body weight and WSSV resistance of the wild shrimp group, which is 0.16±0.00, and the inbred shrimp group, which is 0.20±0.00. On this study the estimated inbreeding depression coefficient was -3.80±0.17% for body weight and -5.81±0.11% for survival time post infection WSSV per 10% increase in inbreeding coefficient. Results were consistent with viewpoint that inbreeding could reduce growth and disease resistance. Those highlight the need to maximize the genetic diversity in base population, and consider rate of inbreeding in selective breeding programs and wild resource protection.4 The heritability and genetic correlation of six hemolymph analytes in Chinese shrimpThe modulation of some hemolymph analytes may function as an important indicator of disease resistance, where precise assessment of genetic parameters is crucial to ensure the success of a selective breeding program. In this study, 51 full-sib families (representing 23 paternal half-sib families) were produced and cultured in the controlled conditions, and the genetic parameters, including the heritability, genetic and phenotypic correlation, of total serum protein concentration (PC), hemocyanin concentration (HC), phenoloxidase (PO) activity, superoxide dismutase (SOD) activity, acid phosphatase (ACP) activity and alkaline phosphatase (ALP) activity in the serum of shrimp F. chinensis were investigated. The results showed that when the average body weight of the shrimp was 7.64g, the concentration of six hemolymph analytes were 153.88±1.65mg/ml, 36.04±0.71 U/ml, 165.24±4.93 U/ml, 333.33±2.74 U/ml, 52.51±2.15 U/100ml and 61.68±2.61U/100ml, respectively. And there were not differences between male and female of all immune parameters and their variation coefficients (P>0.05). Evaluations of heritability (h2) were 0.00±0.13, 0.09±0.22, 0.03±0.20, 0.30±0.20, 0.63±0.32 and 0.39±0.25, respectively, for the traits studied. The results also manifested that rearing the families separately in different tanks affected greatly the common cultured condition, indicated by the effects common to full-sibs other than additive genetics as c2, which were 0.10±0.06, 0.20±0.11, 0.21±0.10, 0.00±0.00, 0.02±0.01 and 0.04±0.01 of the six hemolymph analytes, respectively. In this study, the genetic, environmental and phenotypic correlations among the traits were generally positive except those of SOD activity. Results indicated that immunological parameters of Chinese shrimp could be increased by selective breeding programs though heritability of some traits was quite low. Moreover, genetic correlations between the survival time post affecting WSSV of the young prawn and six immunity analytes of the adult were evaluated according to the phenotypic correlations of their breeding values. Results showed that genetic correlations between the survival time and PC, PO and ACP activities were weak positive. The correlative coefficients were 0.093±0.516, 0.058±0.687 and 0.044±0.760, respectively. And genetic correlations between the survival time and HC, SOD and ALP activities were negative. The correlative coefficients were -0.179±0.209, -0.268±0.057 and -0.110±0.441, respectively. These indicated that the correlation between the young resistance-WSSV and the adult immunity was not significant. These had maybe related to the different tested samples, also maybe related to the different expressed mechanism while being affected and the normal grown.5 Heterosis of Hemolymph Analytes of Two Geographic Populations in Chinese ShrimpTo evaluate heterosis of PC, HC, PO, SOD, ACP and ALP activities, mating was produced between and within Yellow and Bo sea (YB) population and southern seacoast of Korean Peninsula (SK) population, and the offspring of 49 families was reared to 8.09g in the controlled environment. Results showed that the content of all hemolymph analytes of SK population was higher than that of YB population, and the differences were significant except of PC and SOD activity. And the immunocompetence of all hybridizations was higher than that in YB♂×YB♀except of HC and SOD activity in YB♂×SK♀. Almost hybridizations had manifested positive heterosis in 6 hemolymph analytes. Heterosis of SK×YB was all positive, and this hybridization was better than YB×SK in HC, PO and ALP activities. The results suggested that the hemolymph analytes of YB and SK populations had many genetic differences, and hybridization was an important tool to increase immune reaction and adverse resistance.6 Inbreeding impacts on some hemato-immunological parameters of F. chinensis, reared under controlled environmentThis study quantified the effects of inbreeding on PC, HC, PO, SOD, ACP and ALP activity of F. rearing in the standard condition. Results showed all traits of shrimp with inbreeding coefficient 25% were lower than those of shrimp without inbreeding, and their difference was highly significant in PO activity, ACP activity and ALP activity (P<0.01). The estimated inbreeding depression coefficients of six traits were -1.74±0.13%, -4.75±0.23%, -10.50±0.31%, 0.62±0.09%,-25.42±0.21% and -21.06±0.30% per 10% increase in inbreeding coefficient, respectively. Results indicated that inbreeding could cause the depression of immunological vigor as that of growth and disease resistance.更多还原