【作者】 南宫自艳；

【导师】 高宝嘉；

【作者基本信息】 河北农业大学 ， 森林培育学， 2008， 博士

【摘要】 作为我国林业上的重要害虫,我国松毛虫属昆虫多达27个种(包括4个亚种),其中马尾松毛虫(Dendrolimu punctatus punctatus Walker)、油松毛虫(D.punctatustabulaeformis Tsai et Liu)、赤松毛虫(D.punctatus spectabilis Butler)、落叶松毛虫(D.superans Butler)、云南松毛虫(D.houi Lajonquiere)、思茅松毛虫(D.kikuchiiMatsumura)危害较为严重。松毛虫暴发成灾时,森林一眼望去如同火烧一般,须经过长时间才能够恢复原貌,所以给林业生产带来巨大经济损失。松毛虫自身遗传力的变异与自然环境相互作用逐渐形成不同的地理亚种或不同的生态型,探明松毛虫属昆虫在不同环境条件下的遗传适应机制,对于多种松毛虫的综合治理具有重要意义。分别进行了松毛虫属昆虫不同种之间、同种不同地理种群之间、同一地理种群内不同亚居群之间三个层次上的遗传多样性研究:(1)马尾松毛虫、落叶松毛虫、云南松毛虫、思茅松毛虫4种松毛虫之间遗传多样性研究:4种松毛虫之间存在生殖隔离,再加上长时间不同地理气候的影响,使得各种松毛虫在表型性状、蛋白质水平和分子水平均存在较大的变异。通过表型性状数值变异分析,得出蛹重、蛹长、雌虫体重、雌虫翅展、雌虫体长、雄虫体重、雄虫翅展和雄虫体长8项性状指标在4种松毛虫之间差异极显著。在所有性状中变异系数最大的是蛹重,CV=51.99;变异系数最小的是雄虫翅展,CV=5.47;变异系数平均值从大到小依次为:蛹重、雄虫体重、雌虫体重、蛹长、雄虫体长、雌虫体长、雌虫翅展和雄虫翅展。对各种松毛虫变异系数比较发现,4种松毛虫中变异系数最大的是落叶松毛虫,最小的是云南松毛虫。对8个性状做主成分分析得出前3个主分量构成的信息量为总信息量的93.92%,反映了绝大部分的信息,基本能代表这8个性状的变异。共对4种松毛虫124个个体进行了模糊聚类和系统聚类,聚类结果均为马尾松毛虫与落叶松毛虫聚为第一分支,思茅松毛虫和云南松毛虫聚为第二分支。4种松毛虫表型性状的变异与环境因子的相关性较密切:蛹重、雄虫体重与纬度呈显著正相关;雌虫体长、雄虫体长、雌虫翅展与降水量呈显著负相关;蛹重、雄虫体重、雌虫体重与年均温也呈显著负相关。表型性状欧氏距离与地理距离不存在显著的相关性。在8种等位酶标记的分析中,共检测了12个位点,其中6个为多态位点,检测到22个等位基因。4种松毛虫总体水平P=50.00%,A=1.8333,He=0.2323,平均水平上A=1.2708,P=25.00%,He=0.0828,可以看出4种松毛虫杂合度属于较高水平。根据遗传距离构建松毛虫种群系统树,4种松毛虫大致可以分为3支:马尾松毛虫和落叶松毛虫相聚为一类,思茅松毛虫和云南松毛虫各分为一支。环境因子对等位酶标记结果的影响要小得多,仅有Shannon信息指数I与海拔呈负相关趋势。遗传距离与地理距离作相关性分析,结果表明不存在显著的相关性。(2)马尾松毛虫不同地理种群间的遗传多样性研究:RAPD分子标记揭示马尾松毛虫不同地理种群平均水平A=1.252,Ae=1.069;P=25.19%;总体水平上A=1.830,Ae=1.398,P=82.95%,可以看出在马尾松毛虫群体遗传多样性已达较高水平。通过Shannon多样性指数和Nei基因多样性指数两个遗传参数显示遗传多样性变异水平最高的均为江西余江马尾松毛虫种群,最低的为马尾松毛虫地理亚种——辽宁锦州赤松毛虫种群。Shannon多样性指数显示74.90%的变异存在于种群间,基因流Nm仅为0.168;Nei基因多样性指数显示77.80%的变异存在于种群间,基因流Nm仅为0.143;二者数据分析均表明马尾松毛虫种群间的遗传分化已经达到较高水平,种群间基因交流很小。对6个马尾松毛虫种群进行聚类分析,3个马尾松毛虫的地理亚种——赤松毛虫种群相聚形成第一大类群,另外三个马尾松毛虫种群相聚形成第二大类群。以RAPD标记得来的遗传参数分别对环境因子作相关性分析,结果表明Ae、Shannon信息指数I、Nei基因多样性指数h和生境的降水量、年均温呈显著的正相关趋势。遗传距离与种群之间地理距离存在一定相关性,但相关性不显著。在6种等位酶标记的分析中,共检测了10个位点,其中3个为多态位点,检测到15个等位基因。马尾松毛虫6个种群平均水平上的遗传多样性参数A=1.233,P=21.67%,He=0.067;总体水平P=30.00%,A=1.500,He=0.089;从二者的数据结果可以看出,马尾松毛虫在种级水平上的遗传多样性均高于群体平均水平上的遗传多样性。各种群间杂合性基因多样度比率Fst的平均值为0.265,说明总遗传多样性的26.5%来自于种群间的遗传变异,种群间的遗传分化已经达到较高的水平;基因流Nm为0.692,说明马尾松毛虫各种群间基因交流较小。三个赤松毛虫种群首先相聚形成一支,三个马尾松毛虫种群也相聚在一起,形成第二支大的类群。在等位酶分析结果与环境因子相关性研究中发现遗传参数与环境因子均没有显著的相关性。遗传距离与种群之间存在一定相关性,但相关性不显著。(3)油松毛虫种群内不同亚居群间的遗传多样性研究:8对微卫星引物在平泉县油松毛虫种群内5个天然亚居群共225个个体基因组DNA中都获得较好的扩增。在油松毛虫5个亚居群的平均水平上P=80.00%,A=2.6250,He=0.3765;数据表明pop4即天然油松纯林油松毛虫亚居群杂合度最高,pop2即油松、落叶松混交林亚居群杂合度最小。油松毛虫平泉种群总体水平上P=87.50%,A=3.1250,He=0.4747,Fst=0.2159,Nm=0.9081,数据说明承德平泉县油松毛虫种群总遗传多样性的21.59%来自于亚居群间的遗传变异,亚居群间已经发生了明显的遗传分化;各亚居群间基因交流少。根据遗传距离初步推测,pop1亚居群与pop2亚居群分化时间最短,而pop2亚居群与pop3亚居群分化时间则较长。林分类型对各亚居群遗传多样性影响最明显,其次海拔、坡向及是否化防也是重要的影响因素。更多还原

【Abstract】 As the major forestry pest insects in China,Dendrolimus included Dendrolimus punctatus punctatus Walker.D.Punctatus tabulaeformis Tsai et Liu.D.punctatus spectabilis Butler.D.superans Butler.D.houi Lajonquiere and D.kikuchii Matsumura. During sequential outbreaks,economical damage can be so extensive for forest appears to be burned and unable to withstand such a long period of defoliation.We assess gene diversity and genetic structure among four species of Dendrolimus spp,among geographical populations of D.punctatus Walker and among sub-populations of D. punctatus tabulaeformis Tsai et Liu using morphological diversities,allozyme,RAPD and SSR,which provide-a powerful tool for investigation of genetic variation.The influence of ecological factors on the genetic diversity and morphological diversity is also discussed by the correlation analysis.(1)Genetic diversities and morphological diversities among D.punctatus punctatus Walker.D.superans Butler.D.houi Lajonquiere and D.kikuchii Matsumura:Morphological diversities among four species of Dendrolimus spp.were analyzed including pupae weight,pupae length,female weight,female wing length,female length, male weight,male wing length and male length.Analysis of variance for all characteristics was significantly different among species.Coefficient of variation of morphological traits showed that pupae weight is significantly higher than other traits.Pupae length,female weight and female wing length were the main traits account for the phenotypic variations according to the Principal Component Analysis(PCA)and disciminte analysis.Basing on the UPGMA cluster,four species of Dendrolimus could be divided into two groups:the first branch in the dendrogram included D.punctatus punctatus Walker and D.superans Butler,and D.houi Lajonquiere and D.kikuchii Matsumura were grouped together into second branch.The correlations between the phenotypic traits and ecological and geographic factors are significant.There was no significant relationship between the Euclidean distance and geographic distance among species of Dendrolimus spp.During allozyme method on the sampled species,a total of 12 presumed loci and 22 alleles were scored in this study,which 6 loci were polymorphic.At population level,the mean number of alleles per locus(A)was 1.2708,the Percentage of polymorphic loci(P) was 25.00%,and the mean expected heterozygosity per locus(He)was 0.0828.At species level,A=1.8333,P=50.00%,He=0.2323.So,the genetic diversity at species level was rather high than that at populations level.A UPGMA dendrogram was generated based on Nei’s genetic distance showed 4 species of Dendrolimus were clustered into three groups: the first branch in the dendrogram included D.punctatus punctatus Walker and D. superans Butler,and D.houi Lajonquiere and D.kikuchii Matsumura became the second and the third branch independently.The correlations between the genetic data and ecological factors are not significant except for Shannon information index and altitude. There was no significant relationship between the genetic distance and geographic distance.(2)Genetic diversities among 6 geographical populations of D.punctatus Walker:At population level,the percentage of polymorphic loci(P)was 25.19%,the mean number of alleles per locus(A)was 1.252,Ae=1.069,the mean Shannon’s information index(I)was 0.090.For the species level,P=82.95%,A=1.830,Ae=1.398 in RAPD analysis method.The coefficient of genetic differentiation between populations based on Shannon information index was 0.7490,based on Nei’s genetic diversity was 0.7780, which revealed a very high level of genetic differentiation among populations.The number of migrants per generation among populations(Nm)was 0.168 based on Shannon information index,Nm was 0.143 based on Nei’s genetic diversity,which revealed a very low gene flow among populations.Shannon information index and Nei’s genetic diversity all showed that there was highest genetic diversity in population of D.punctatus punctatus Walker of YJM,the lowest genetic diversity in population of D.punctatus spectabilis Butler of JZC.Three populations of D.punctatus punctatus Walker and three populations of D.punctatus spectabilis Butler were clustered into one branch independently by using Nei’s unbiased genetic distances.The correlations between the genetic index and ecological and geographic factors are significant.There was no significant relationship between the genetic distance and geographic distance among populations.In allozyme method,a total of 10 presumed loci and 15 alleles resolved were scored in this study,which 3 loci were polymorphic.At population level,the mean number of alleles per locus(A)was 1.233,the percentage of polymorphic loci(P)was 21.67%,and the mean expected heterozygosity per locus(He)was 0.067.At species level,A=1.500, P=30.00%,He=0.089.So,the genetic diversity at species level was rather high than that at populations level.From the summary of F-statistics at polymorphic loci of 6 populations, we could found the Fst was 0.265,which mean there show high genetic diversity among populations.The number of migrants per generation among populations(Nm),based on Fst calculations,was 0.692,suggesting the occurrence of rather low gene flow among populations.A UPGMA dendrogram was generated showed 6 populations of D.punctatus Walker were clustered into two groups:the first branch in the dendrogram included three populations of D.punctatus spectabilis Butler,and the remaining three populations of D. punctatus punctatus Walker were grouped together into second branch.The correlations between the genetic data and ecological factors are not significant.There was no significant relationship between the genetic distance and geographic distance among populations,the same to the RAPD data.(3)Genetic diversities among 5 sub-populations of D.punctatus tabulaeformis Tsai et Liu:Polymorphic bands were produced using 8 primers in SSR analysis method.The percentage ofpolymorphic loci(P)was 80.00%,the mean number of alleles per locus(A) was 2.6250,and the mean expected heterozygosity per locus(He)was 0.3765.For the species level,P=87.50%,A=3.1250,He=0.4747.From the summary of F-statistics at polymorphic loci of 5 sub-populations,we could found the Fst was 0.2159,which mean there show high genetic diversity among sub-populations.There were the highest genetic diversity in sub-population of pure Pinus tabulaeformis Carr,the lowest genetic diversity in sub-population of the artificial mixed forest of P.tabulaeformis Carr and Larix principis-rupprechtii Mayr.The number of migrants per generation among populations (Nm)was 0.9081,suggesting the occurrence of rather low gene flow among sub-populations.A UPGMA dendrogram based on Nei’s genetic distance showed 5 sub-populations of D.punctatus tabulaeformis Tsai et Liu were clustered into two groups: the first branch in the dendrogram included pop1 and pop2,and the remaining three sub-populations were grouped together into second branch.The genetic index has positive relationship with ecological factors but not significant.Stand type,slope,aspect and altitude were the main traits account for the genetic diversities according to the PCA.更多还原