龟纹瓢虫（Propylaea japonica）雌性生殖系统发育及人工饲料的研究
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摘要
龟纹瓢虫(Propylaea japonica Thungberg)隶属于鞘翅目(Coleoptera)、瓢虫科(Coccinellidae)、龟纹瓢虫属(Propylaea)，能捕食多种农业害虫，并且具有食量大、产卵多、适应性强、年发生代数多等特点，因此是我国农业生产中一种重要的捕食性天敌昆虫。本文研究了与生殖密切相关的龟纹瓢虫雌性生殖系统的发育和人工饲料对龟纹瓢虫生理生化及捕食等方面的影响。
本研究通过组织学方法和免疫学技术研究了龟纹瓢虫卵子发生以及卵黄蛋白的相关特性，同时运用生物化学方法，深入研究了人工饲料对龟纹瓢虫生长和繁殖性能、营养效应、体成分组成、中肠消化酶活性以及捕食行为等各方面的影响，并对其机理进行了讨论。研究结果可为丰富龟纹瓢虫生殖生物学提供基础资料，同时也为进一步研究适合龟纹瓢虫生长和繁殖的人工饲料提供数据参考。
通过对龟纹瓢虫卵子发生研究，结果表明：龟纹瓢虫卵子发生过程可明显划分为卵母细胞分化期、卵母细胞生长期、卵黄形成及卵子成熟期3个时期，并可进一步分为7个阶段。对卵子发生的分期有利于今后进一步的研究。
经PAGE和SDS-PAGE实验表明，龟纹瓢虫卵黄蛋白分子量为294.81±40.70kDa，并由分子量分别为144.68±0.03kDa和51.23±0.27 kDa的两种亚基组成。对卵黄蛋白的氨基酸组成和含量分析发现必需氨基酸总量占57.48％，略高于非必需氨基酸；其中谷氨酸(Glu)含量最高，为15.26％，而色氨酸(Trp)和蛋氨酸(Met)含量较低，分别为0.50％和0.11％。
采用间接ELISA法，系统测定了龟纹瓢虫成虫期脂肪体、血淋巴和卵巢中卵黄蛋白的动态变化。结果表明：脂肪体是卵黄原蛋白合成的场所，卵黄原蛋白的合成始于羽化后第2d；脂肪体、血淋巴和卵巢中卵黄原蛋白的滴度在羽化后第4d开始迅速上升，至成虫期的第8d左右达到高峰期。
人工饲料对龟纹瓢虫生长和繁殖的研究表明：添加橄榄油后幼虫发育历期明显增长，添加0.3％的橄榄油能显著提高幼虫的成活率；在成虫体重增长上，雌雄虫的体重均为在刚羽化后几天内体重增长最显著，然后增长变缓；并且添加蔗糖能显著提高雌虫的体重增长率而对雄虫作用不显著；在雌虫繁殖性能的研究发现，蚜虫喂养组有明显的持续时间较长的产卵高峰期，而人工饲料的产卵高峰期持续时间较短并且出现时间较晚或分为两个高峰期；添加0.3％的橄榄油能显著缩短龟纹瓢虫的产卵前期，几乎达到蚜虫喂养组的水平，而添加蔗糖在一定程度也能缩短产卵前期，同时还能显著提高龟纹瓢虫的产卵量。
Propylea japonica (Thunberg) is subordinated to Coleoptera, Coccinellidae, Propylaea. The ladybird preys on many pests in agriculture. Due to large appetite, high fecundity, adaptability and many generations annually, Propylea japonica is one of important predatory natural enemies in agriculture system. In this paper, the development of female reproductive system and the artificial diets in Propylea japonica are investigated.
The vitellogenesis and the characteristics of vitellin were studied using histological and immunological techniques. And then, by the method of biochemistry, an attempt was made to study the effect of artificial diets on the Propylea japonica, including growth and fecundity, nutritional indices, body composition, activity of digestive enzyme in midgut and the functional response to cotton aphid. The purpose is to accumulate basal information for reproductive biology of Propylea japonica, and provide reference for optimum design of artificial diets.
Oogenesis in the adult stage could be divided into 3 periods: oocytes differentiation; oocytes growth; the vitellogenesis and the oocytes maturation. And the 3 periods could be divided into 7 stages further.
The characterization of vitellin was carried out by PAGE and SDS-PAGE. The results indicated that vitellin had a molecular weight of approximately 294.81±40.70 kDa and was composed of two kinds of subunits, whose molecular weights were 144.68±0.03 kDa and 51.23±0.27 kDa respectively. The content of essential amino acid was 57.48%, which was little higher than that of non-essential amino acid. And the highest contents of amino acids made up of vitellin was Glu, its content was 15.26%, while contents of Try and Met were very low, only 0.50%, 0.11% respectively.
Using indirect ELISA, the contents of vitellin in Propylea japonica fat body, haemolymph and ovary were detected. The results suggested that the vitellogenin was synthesized in fat body at the second day after eclosion. The contents of vitellogenin in fat body, haemolymph and ovary increased quickly on the fourth day after eclosion and reached the peak stage on the eighth day approximately in adult stage.
本研究通过组织学方法和免疫学技术研究了龟纹瓢虫卵子发生以及卵黄蛋白的相关特性，同时运用生物化学方法，深入研究了人工饲料对龟纹瓢虫生长和繁殖性能、营养效应、体成分组成、中肠消化酶活性以及捕食行为等各方面的影响，并对其机理进行了讨论。研究结果可为丰富龟纹瓢虫生殖生物学提供基础资料，同时也为进一步研究适合龟纹瓢虫生长和繁殖的人工饲料提供数据参考。
通过对龟纹瓢虫卵子发生研究，结果表明：龟纹瓢虫卵子发生过程可明显划分为卵母细胞分化期、卵母细胞生长期、卵黄形成及卵子成熟期3个时期，并可进一步分为7个阶段。对卵子发生的分期有利于今后进一步的研究。
经PAGE和SDS-PAGE实验表明，龟纹瓢虫卵黄蛋白分子量为294.81±40.70kDa，并由分子量分别为144.68±0.03kDa和51.23±0.27 kDa的两种亚基组成。对卵黄蛋白的氨基酸组成和含量分析发现必需氨基酸总量占57.48％，略高于非必需氨基酸；其中谷氨酸(Glu)含量最高，为15.26％，而色氨酸(Trp)和蛋氨酸(Met)含量较低，分别为0.50％和0.11％。
采用间接ELISA法，系统测定了龟纹瓢虫成虫期脂肪体、血淋巴和卵巢中卵黄蛋白的动态变化。结果表明：脂肪体是卵黄原蛋白合成的场所，卵黄原蛋白的合成始于羽化后第2d；脂肪体、血淋巴和卵巢中卵黄原蛋白的滴度在羽化后第4d开始迅速上升，至成虫期的第8d左右达到高峰期。
人工饲料对龟纹瓢虫生长和繁殖的研究表明：添加橄榄油后幼虫发育历期明显增长，添加0.3％的橄榄油能显著提高幼虫的成活率；在成虫体重增长上，雌雄虫的体重均为在刚羽化后几天内体重增长最显著，然后增长变缓；并且添加蔗糖能显著提高雌虫的体重增长率而对雄虫作用不显著；在雌虫繁殖性能的研究发现，蚜虫喂养组有明显的持续时间较长的产卵高峰期，而人工饲料的产卵高峰期持续时间较短并且出现时间较晚或分为两个高峰期；添加0.3％的橄榄油能显著缩短龟纹瓢虫的产卵前期，几乎达到蚜虫喂养组的水平，而添加蔗糖在一定程度也能缩短产卵前期，同时还能显著提高龟纹瓢虫的产卵量。
Propylea japonica (Thunberg) is subordinated to Coleoptera, Coccinellidae, Propylaea. The ladybird preys on many pests in agriculture. Due to large appetite, high fecundity, adaptability and many generations annually, Propylea japonica is one of important predatory natural enemies in agriculture system. In this paper, the development of female reproductive system and the artificial diets in Propylea japonica are investigated.
The vitellogenesis and the characteristics of vitellin were studied using histological and immunological techniques. And then, by the method of biochemistry, an attempt was made to study the effect of artificial diets on the Propylea japonica, including growth and fecundity, nutritional indices, body composition, activity of digestive enzyme in midgut and the functional response to cotton aphid. The purpose is to accumulate basal information for reproductive biology of Propylea japonica, and provide reference for optimum design of artificial diets.
Oogenesis in the adult stage could be divided into 3 periods: oocytes differentiation; oocytes growth; the vitellogenesis and the oocytes maturation. And the 3 periods could be divided into 7 stages further.
The characterization of vitellin was carried out by PAGE and SDS-PAGE. The results indicated that vitellin had a molecular weight of approximately 294.81±40.70 kDa and was composed of two kinds of subunits, whose molecular weights were 144.68±0.03 kDa and 51.23±0.27 kDa respectively. The content of essential amino acid was 57.48%, which was little higher than that of non-essential amino acid. And the highest contents of amino acids made up of vitellin was Glu, its content was 15.26%, while contents of Try and Met were very low, only 0.50%, 0.11% respectively.
Using indirect ELISA, the contents of vitellin in Propylea japonica fat body, haemolymph and ovary were detected. The results suggested that the vitellogenin was synthesized in fat body at the second day after eclosion. The contents of vitellogenin in fat body, haemolymph and ovary increased quickly on the fourth day after eclosion and reached the peak stage on the eighth day approximately in adult stage.
引文
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