【作者】 胡玉伟；

【导师】 雷朝亮；

【作者基本信息】 华中农业大学 ， 农业昆虫与害虫防治， 2011， 博士

【摘要】 中华真地鳖Eupolyphaga sinensis是一种重要的药用资源昆虫。随着人们对这种昆虫的开发与利用的重视,对其质量品质要求也越来越高。不同地理种群的中华真地鳖,其品质会有差异。本文以不同地理种群的中华真地鳖为研究对象,调查不同地理种群中华真地鳖的体型大小变异及其变异机制,同时在体型大小变异基础上探索其免疫变异并分离纯化免疫成分,为这种昆虫的资源开发与利用提供依据,其主要研究结果如下：A.中华真地鳖体型大小纬度变异在中华真地鳖主要分布区14个点采集其成虫,对成虫体长,体宽和前胸背板测量,同时检测10个气候指标对成虫体型的影响。结果发现中华真地鳖成虫随纬度变化呈现U型模型,成虫体型在低纬度和高纬度较大,在中间纬度较小；体型与气候的关系也呈现上述模型。根据结果推测季节长度的变化和中华真地鳖化性在地理上的可变性是造成这种模型的原因。气候因子也对这种模型具有重要贡献,这对于生物生态学和进化学研究具有重要的意义,进而可以预测全球气候变暖可能会影响此昆虫的地理变异。B.中华真地鳖卵鞘颜色和大小纬度变异采集中华真地鳖主要分布区不同地理种群的卵鞘,观察其颜色,测量其大小(包括长度,单卵鞘含卵量及呼吸孔数量)。同时对其进行实验室相同条件饲养,观察其后代变异。结果发现不同地理种群卵鞘颜色有差异,随纬度升高,颜色从黑色到红棕色逐渐变化,但在同等条件饲养下,差异消失；其卵鞘大小随纬度变化呈现U-型模型,由此判断,卵鞘的地理变异受各种环境影响,包括母体环境、周围环境与遗传等各方面影响,其中遗传影响可能是其主要原因。C.中华真地鳖体型大小变异机制——若虫、成虫对温度的反应对中华真地鳖不同地理种群若虫进行同条件饲养,发现其体长与体宽呈显著一元线性回归关系,不同地理种群斜率(体形)存在差异。不同地理种群各龄若虫发育历期并不随龄期增长而延长。雌性若虫期发育历期天津种群显著大于其他种群,其他种群间除江苏种群外均无显著差异。雄性若虫期发育历期除天津种群外均无显著差异。同一地理种群,雌性若虫期发育历期大于雄性若虫期发育历期。结果表明,中华真地鳖不同地理种群间在形态和生物学上均有一定的差异。检测温度变化(22、25、28和31℃)对中华真地鳖不同地理种群发育历期、成虫体型和存活率的影响。结果发现发育历期和成虫体型大小随温度上升而下降,说明发育温度显著影响此昆虫的生长和生活史的一些重要特征指标。同时,不同地理种群对温度变化的反应在这三个指标上存在显著差异,表明热反应行为在种群之间具有遗传性不同。然而,不同地理种群的纬度变异不能完全解释这种遗传上的不同。发育历期、成虫体型和存活率的变异都反映出该昆虫对热环境体系的一种适应,证实温度变异对其地理变异具有重要影响。D.中华真地鳖成虫体型大小性二型变异研究——验证differential-plasticity(雌雄体型不同可塑性)假说用具有体型雌雄二型现象的中华真地鳖为材料验证Differential-plasticity假说。将中华真地鳖不同地理种群一龄若虫分别放置于四个温度下(22、25、28和31℃)饲养至成虫。结果显示随着温度上升,雌雄虫个体变小；并且在所有温度下雌虫总是大于雄虫。体型的二型现象指标在31℃下最大,而在22℃下最小；体型的性二型现象在种群间有变异,但是雌雄虫对温度的适应程度没有显著差异。结果显示中华真地鳖体型性二型现象在种群间的变异与雌雄虫对温度的适应程度无关,本研究结果不支持此假说。此外,研究结果暗示性别选择作用(遗传作用)可能导致不同种群体型上的性二型变异。E.中华真地鳖抗菌肽变异、抗菌成分及抗菌机制研究对不同地理种群的中华真地鳖抗菌肽进行凝胶层析分离纯化,发现不同地理种群的抗菌肽粗成分无显著差别。为确定其主要成分本实验通过对其初步分离,凝胶层析分离,AKTA explorer层析,高效液相色谱分析等对其抗菌肽进一步分析。分离出一种小肽,分子量在806Da左右,通过质谱分析,其结构有六种可能,有可能是抗菌肽活性片段或者完整小肽。对抗菌肽处理过的阴性细菌细胞超微结构观察发现,此抗菌肽可通过破坏细胞膜,细胞壁或细胞质从而造成细胞死亡。结果表明,细菌细胞壁和细胞膜可能是中华真地鳖抗菌肽的作用靶点,此种抗菌肽抗菌机制是多样性的。更多还原

【Abstract】 Eupolyphaga sinensis Walker (Blattaria) is an insect of medical importance. The high quality is required along with the market demand of it. However, the quality is different between populations. This paper focused on variation in body size, ootheca size, sexual dimorphism and antimicrobial peptides in E. sinensis among populations and we also showed the mechanism.A. Latitudinal shifts in body size in cockroach, Eupolyphaga sinensis (Blattaria)In the present study we investigated the geographic variation in body size of the Chinese cockroach, Eupolyphaga sinensis, which has a variable life cycle length. Adults collected from fourteen localities across temperate zone to subtropical zone in China were measured by using body length, body width and pronotum width and 10 variables were examined to test whether climatic factors affect body size. We found that the body size of E. sinensis varied considerably with latitude, demonstrating a U-shaped pattern. Cockroaches were larger at low and high latitudes, but smaller at intermediate latitudes. Thus the relationship between climate and body size conformed to a U-shaped pattern. Results indicate that two factors were significantly associated with body size clines: season length and variability in life cycle length with latitude. Our results also demonstrated that climate variables contribute to latitudinal clines in body size, which has important ecological and evolutionary implications. It can be expected that global climate change may alter latitudinal clines in body size of E. sinensis.B. Geographic variation in ootheca size along a latitudinal gradient from ChinaBody size has a considerable effect on offspring. In the first chapter, body size of E. sinensis varies with latitude and we expect that there is a relationship between ootheca size and latitude. We observed the colour of ootheca size and measured latitude-related ootheca size variation in field-collected E. sinensis individuals. We selected seven major collection locations that are the main distribution regions. We measured field-collected oothecas in terms of ootheca length, eggs per ootheca and crest numbers that are correlated with oothecas, and found that ootheca size demonstrated a U-shaped pattern with latitude, showing large size at low and high latitudes and small at intermediate latitudes. We conclude that there is clinal variation in ootheca size of E. sinensis and the variation is likely to be the result of maternal effect or local adaptation to environment. C. Effect of temperatures on development time and body size in Eupolyphaga sinensis along a latitudinal gradient from ChinaThe development of six population of Eupolyphaga sinensis were studied at 31℃in the laboratory. The relationship of the body length and the body width was the univaniate correlation of linear regression, which was extremely significance in the six population as well as the slopes (shape). The results showed that the duration of the nymph did not increased as the instar stage increased. The female developmental duration of Tianjin population was significantly greater than other populations, and there were no significant difference lies among the other populations except Jiangsu population. The male developmental duration of Tianjin was significant higher than the other five populations which had no significant difference. As far as the same population, the developmental duration of female was longer than that of the male. The results suggested that the six population had some difference in morphology and biology.We examined the effects of various temperatures (of 22,25,28 and 31℃) on development time, adult body size and pre-adult survivorship in three populations of the cockroach, E. sinensis, collected at different latitudes. We found substantial temperature-induced plasticity in development time, body size and pre-adult survivorship, indicating that developmental temperatures have strong impacts on growth and life history traits of E. sinensis. Genetic differences for development time, body size and pre-adult survivorship were detected among populations, and the three traits exhibited highly significant variations in the responses of different populations to various temperature conditions, indicating genetic differences among populations in terms of thermal reaction norms. We also found that two populations seem to support the beneficial acclimation hypothesis whereas the third mid-latitude population does not. The results are likely due to differences in season length and voltinism, indicating that not only temperature regime but also its interactions with generation time (and development time), voltinism, and season length are likely to have considerable effects on insect development time and body size. Overall, changes in development time, body size and pre-adult survivorship in E. sinensis can all be regarded as adaptations to changing thermal regimes.D. Variation in sexual size dimorphism among populations:testing the differential-plasticity hypothesisWe test the differential-plasticity hypothesis that sex-differential plasticity to environmental variables generates among-population variation in the degree of sexual dimorphism in this study. We examined the effects of rearing animals at various temperatures (of 22,25,28 and 31℃) on sexual dimorphism in four populations of the cockroach, Eupolyphaga sinensis, collected at different latitudes. We found that females were larger than males at all temperatures and the degree of this dimorphism was largest at the highest temperature (31℃) but smallest at the lowest temperatures (22℃). There is variation in the degree of sexual size dimorphism among population (sex×population interaction), but differences between the sexes in their plastic responses (sex×temperature interaction) were not observed for body size. Our results indicated that sex-differential plasticity to temperature was not the cause of differences among populations in the degree of sexual dimorphism in body size.E. Antimicrobial peptides and antimicrobial mechanisms of Eupolyphaga sinensisWe have confirmed that body size varied among populations along latitude. Body size is considered as one of the most important traits of an organism, because it influences nearly every aspect of the biology of the organism, especially in immunoreaction. Are antimicrobial peptides different among populations in Eupolophyga sinensis? Antimicrobial peptides were isolated from the E. sinensis by a four-step protocol including one step Sephadex G-50, one step Sephadex G-25 and two steps of RP-HPLC. We found that there seem to be no difference in antimicrobial peptides among populations. The antimicrobial mechanisms were also investigated. They may exert their antimicrobial functions by various means, including forming lamellar mesosome-like structures, and peeling off the cell walls, forming pores.更多还原