三种蚤生殖系统的细微结构:雄性外生殖器的发育

本文研究了缓慢细蚤Leptopsylla segnis(Schonherr),不等单蚤Monopsyllus anisus(Rothschild)和猫栉首蚤指名亚种Ctenocephalides felis felis(Bouche)雄性外生殖器的结构,观察了从幼虫、前蛹、蛹至成虫各发育时期的雄性外生殖器的内部结构变化.对有争议的雄蚤上抱器的起源,雄蚤生殖孔的位置,雄性外生殖器芽内陷的腹节以及射精管横切面的细胞数目和阳茎背、腹杆的结构等问题进行了详细的观察和探讨.     

三种蚤生殖系统发育的细微结构:雌性生殖腺的发育

本文研究的缓慢细蚤Leptopsylla segnis (Schonherr,1811),不等单蚤Monopsyllus anisus (Rotbschild,1907)和猫栉首蚤指名亚种Ctenocephalides felis felis (Bouchè,1835)均采自贵阳市区,鼠、猫宿主体上。通过饲养与繁殖,制作从三龄幼虫至成虫成熟各发育时间的连续切片。预察了各发育时间生殖腺发育的内部结构变化,依据卵巢芽的发育、分化,末端最大卵母细胞形态和大小的变化,卵泡上皮细胞形态和大小的变化,卵泡间组织的出现,卵黄的产生、积累及卵壳物质的分泌等,将雌性生殖腺的发育分为四期10个阶段,并对雌性生殖腺发育内部结构的变化,雌蚤卵黄沉淀初期的“空位现象”等进行了分析析和讨论。     

两种鼠蚤在新羽化和吸血后不同时间三种酶的组织化学研究

采用组织化学技术,研究了不等单蚤Monopsyllus anisus (Rothschild)和缓慢细蚤Leptopsylla segnis (Schonherr)新羽化和吸血后24 h、48 h和72 h碱性磷酸酶、酸性磷酸酶和三磷酸腺苷酶的分布和活性。显微摄影及定量图像分析结果显示：新羽化蚤碱性和酸性磷酸酶主要存在于中肠、神经细胞核、精子束头部、精巢附腺、射精管、输卵管和受精囊附腺中;三磷酸腺苷酶各组织中均有分布。吸血消化后,两种蚤中肠3种酶活性均有增强;除碱性磷酸酶在消化72 h 后酶活性有所下降外,其余不同消化时间酶活性增强程度上不存在显著差异。两种蚤卵母细胞发育成熟过程中3种酶活性亦见增强。     

果蝇睾丸基因敲除突变体的构建及表型分析

生殖系统功能的正常维持是物种繁衍的基础,需要多基因协同作用,但其中许多基因的具体功能和作用机制并不清楚。本研究选取了果蝇(Drosophila melanogaster)中8个在睾丸中表达、功能未知且与人(Homo sapiens)和小鼠(Mus musculus)高度同源的基因(CG4161、CG11475、CG2921、CG10541、CG7276、CG3800、CG8117和CG16779),分析了它们在不同组织中的表达水平,并分别检测了它们在雄性生殖系统中的功能。在这8个基因中,前5个为睾丸优势表达基因,其余3个为全身性表达。首先,利用CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9)技术结合同源定向修复(homology-directed repair, HDR)在果蝇中对8个候选基因逐一进行敲除,建立了纯合的基因敲除突变品系;然后对这些品系的雄蝇进行了生育力测试及睾丸细胞学观察。结果显示,CG7276和CG3800基因敲除果蝇出现部分雄蝇不育且可育雄蝇后代数量较野生型显著下降。睾丸解剖观察显示CG7276和CG3800的功能缺失可导致雄蝇分别出现不同程度的精囊缩小及精原干细胞减少和细胞分布混乱;染色结果也提示CG7276和CG3800在精子的成熟过程中发挥一定作用。其他6个基因突变并未导致雄蝇育性变化或睾丸形态异常。这些突变体的获得及表型的初步分析为进一步研究基因功能及机制提供了良好的动物模型及基础。     

三种蚤生殖系统的细微结构:雌性外生殖器的发育

本文研究了缓慢细蚤 Leptopsylla segnis(Schōnherr),不等单蚤 Monopsyllus anisus(Rothschild)和猫栉首蚤指名亚种Ctenocephalides felis felis(Bouché)雌性外生殖器的结构,观察了从幼虫、前蛹、蛹至成虫各发育时期的雌性外生殖器的内部结构变化.对一直悬而未决的雌蚤中输卵管等的起源问题,进行了详细的观察和探讨.认为从晚期3龄幼虫开始出现的雌性外生殖器芽,在前蛹期成为四个部分:1.第7腹板后缘腹壁内陷形成的一对外胚层囊;2.紧接外胚层囊后并延伸至第8腹板的外胚层增厚;3.在第8腹板后部,外胚层增厚两侧的产卵器芽;4.第8腹板后缘腹壁内陷形成的受精囊芽.并认为,这三种蚤的中输卵管由一对外胚层囊和其后的外胚层增厚前端的一小部分内陷形成,阴道由外胚层增厚的大部分和第8、9腹板腹壁内陷形成,受精囊由受精囊芽内陷形成.     

蚤类染色体的制片方法及分带技术

本文首次介绍了蚤类染色体的空气干燥制片方法和分带技术。以蚤成熟三龄幼虫和成蚤生殖腺为材料,采用体外短期培养、空气干燥、Giemsa染色技术制片,具取材容易、操作简便、中期分裂相多且染色佳、标本不需封片即可长期保存等优点,克服了以往醋酸地农红压片法染色体分散不佳、杂质多、染色易褪色及标本需封片才能保存等缺点。在印鼠客蚤等四种蚤的染色体研究中均获得满意效果,成功率高,重复性好,尤以生殖腺染色体制片成功率为100％。在此基础上还摸索建立了蚤类染色体的C-分带、G-分带和银染技术。     

苯醚威作用于印鼠客蚤的组织学研究

【目的】通过研究苯醚威对印鼠客蚤 Xenopsylla cheopis (Rothschild,1903)的早 3 龄幼虫和未吸血新羽化成虫的组织学变化,探讨其灭蚤机理,为鼠疫媒介蚤种的防治提供基础资料。【方法】以微量点滴法将苯醚威施药于印鼠客蚤早3龄幼虫和未吸血新羽化成虫,采用组织学、显微摄影及统计学方法观察组织变化。【结果】经苯醚威作用后,印鼠客蚤的早 3 龄幼虫的表皮增厚、卵巢芽生殖细胞萎缩、睾丸芽精原细胞间质减少;未吸血新羽化成虫的睾丸塞消失快、唾液腺细胞破坏严重、中肠上皮细胞萎缩。【结论】(1)苯醚威通过干扰印鼠客蚤幼虫的变态,引起幼虫表皮、生殖芽异常改变,不能发育为成虫而死亡;(2)苯醚威可加速印鼠客蚤新羽化雄性成虫的睾丸塞吸收;(3)苯醚威可破坏印鼠客蚤新羽化成虫的唾液腺细胞,并引起中肠上皮细胞萎缩。     

蚤类染色体的制片方法及分带技术

本文首次介绍了蚤类染色体的空气干燥制片方法和分带技术。以蚤成熟三龄幼虫和成蚤生殖腺为材料,采用体外短期培养、空气干燥、Giemsa染色技术制片,具取材容易、操作简便、中期分裂相多且染色佳、标本不需封片即可长期保存等优点,克服了以往醋酸地衣红压片法染色体分散不佳、杂质多、染色易褪色及标本需封片才能保存等缺点。在印鼠客蚤等四种蚤的染色体研究中均获得满意效果,成功率高,重复性好,尤以生殖腺染色体制片成功率为100%。在此基础上还摸索建立了蚤类染色体的C一分带、G一分带和银染技术。     

大型枝角类蚤状溞对小型热带湖泊浮游植物群落影响的研究

2006年4月10日至2006年4月22日,采用原位吊瓶实验的方法,在暨南大学校园明湖中进行了大型枝角类蚤状溞(Daphnia pulex)对浮游植物种群变化和群落结构影响的实验。实验初期,明湖浮游植物群落的组成是以飞燕角甲藻(Ceratium hirundinella)和单角盘星藻(Pediastrum simplex)为优势种。在室内将蚤状溞培养至体长2mm以上,将蚤状溞加入到装满湖水的4.5L的透明瓶中,设置4个梯度:0ind(A,对照),10ind.(B),20ind(C)和30ind(D),每个梯度有3个平行,将实验瓶置于水表层50cm处。12d后实验结束,不同处理之间的浮游植物种群数量和组成有了明显的差异。与对照组相比,飞燕角甲藻和颗粒直链藻(Meclosira granulata)密度在实验组B、C和D中下降明显;小环藻(Cyclotella)和栅藻(Scenedesmus)等小型浮游植物在牧食压力比较大的C和D瓶中密度低于对照组,而在牧食压力相对较低的B瓶中,它们的密度高于对照组,这可能是由于蚤状溞加速了水体的营养盐循环反过来促进这些小型浮游植物的生长。处理组中绿藻门的盘星藻数量没有明显的下降,这与盘星藻不能被浮游动物直接滤食有关。实验结果表明蚤状溞对浮游植物的群落数量和组成的影响程度与其自身的种群密度密切相关,也与浮游植物群落结构有关。由于飞燕角甲藻是一种细胞较大的种类,蚤状溞对它的抑制作用主要是通过机械伤害作用实现的。     

大劣按蚊雄蚊的交配能力和日龄及交配次数对其生殖系统形态学影响的观察

本文在实验室条件下,观察大劣按蚊(Anopheles dirus)雄蚊交配能力和日龄、交配及交配后间歇期对其生殖系统形态学的影响.羽化后1小时可见丝状精子,22小时尾器全部旋转180°.3—12日龄雄蚊交配能力最强.随着日龄增加,精囊数逐渐减少,睾丸内成熟精子占据整个睾丸的比例逐渐增加,附腺透明区宽度变窄直至消失.多次交配比一次交配后,其睾丸内精子量减少、透明区宽度增加明显;交配后经过一定间歇期后,生殖系统能重新恢复活力,但多次交配后其恢复程度和速度远比交配一次者为慢.大劣按蚊雄蚊生殖系统的变化可初步判断其日龄和交配史.     

THE DEVELOPMENT OF THE SENSILIUM OF THREE FLEA SPECIES

Abstract The result of a study on the internal structures during the development of sensilium and anal segment from first instar larva to adult of Leptopsylla segnis (Schinherr 1811), Monopsyllus anisus (Rothschild 1907) and Ctenocephalides felis felis (Bouche 1835) is reported and summarized as follows: (1) The development of the sensilium and anal segment of these fleas is essentially the same. (2) The first sensilium rudiment appears in the early third instar larva on the anterior one-second part of the tergite of the tenth abdominal segment, and in prepupa the eleventh abdominal segment which is composed of the dorsal and ventral anal lobes is formed, and in the pupa the sensilium and its frame are fully developed. (3) The differenced of opinions on the terminal segments of the flea are discussed.     

Sperm maturation screening and the effect of ecdysone on sperm development of silkworm Bombyx mori

There are two sperm morphs of silkworm, the nucleated spermatozoa (eupyrene) and anucleated spermatozoa (apyrene). Eupyrene sperm cannot complete fertilization successfully without the apyrene sperm. Here a modified rapid and efficient method for sperm identification was developed, after 10 s of fixation in paraformaldehyde and 30 s of 4′6-diamidino-2-phenylindole (DAPI) or propidium Iodide (PI) staining, the sperm bundles can be detected easily using a fluorescence microscope. Sperm maturation process of silkworm from the fifth instar larvae to the adult was described with the above method, the precise time of earliest elongate apyrene bundles was detected on day 2 of pre-pupation, with a ratio of 5% in total sperm bundles, after which the percentage of apyrene sperm bundles increased rapidly and attained a relatively stable ratio of 75% at the end of pupation and nearly 80% after eclosion. Delayed mating leads to apyrene sperm accumulation and damaged fertilization. Previous study showed that ecdysone can increase the frequency of apyrene sperm bundles in vitro. Here 20-hydroxyecdysone (20E) was injected into hemolymph of the 2-d-old fifth instar larvae, the worms entered into mounting period after three days injection, but no apyrene sperm bundles were induced unless day 2 of pre-pupation. Interestingly, maturation of eupyrene sperm bundles were accelerated, and the ratio of eupyrene sperm bundles increased and exhibited a dose-dependent effect after 20E injection, which indicated that the development of eupyrene sperm can be accelerated by ecdysone before pupation of silkworm in vivo. These results will provide new clues for lepidopteran pest control.     

Effect of constant light on male sterility in the cotton leafworm Spodoptera littoralis

Females of the cotton leafworm (Spodoptera littoralis) reared in long day conditions (LD 16:8 h) and mated to males kept throughout the whole period of development in continuous light (LL) oviposit very small numbers of mostly sterile eggs. It was found that in control males reared from the first larval instar in long day conditions there was a large accumulation of euperene sperm bundles in their testes on day 1 after imaginal moult. On day 10 of adult life the number of the sperm bundles was very small. In males kept from the first instar in continuous light there was also high number of sperm bundles on day 1 after imaginal moult but it did not decrease significantly on day 10 as was observed in controls. Transfer of different developmental stages of S. littoralis from long day conditions to continuous light resulted in a big difference in the density of eupyrene sperm bundles in their testes. In control insects reared through the whole of their development in long day conditions there was a significant decrease in the density of eupyrene sperm bundles on day 10 of adult life. By contrast, in males in continuous light, regardless of their developmental stage when transferred, there was either no change in density of sperm bundles in day 10 adults or there was a significant increase in comparison with day 1 adults. The highest density of eupyrene sperm bundles was observed in day 10 adults when they were transferred to continuous light shortly before moulting to the last instar (as day 4 larvae in the last stadium or day 1 pupae). Generally, the density of eupyrene sperm bundles on day 10 of adult development was about 2–2.5 times higher in males in continuous light then those under long day conditions. The results presented here indicate that the last larval instar and the pupa are the stages most sensitive to constant light treatment, which greatly reduces the amount of eupyrene sperm bundles released from the testes.     

KNOCKDOWN OF ATPsyn‐b CAUSED LARVAL GROWTH DEFECT AND MALE INFERTILITY IN Drosophila

The ATPsyn‐b encoding for subunit b of ATP synthase in Drosophila melanogaster is proposed to act in ATP synthesis and phagocytosis, and has been identified as one of the sperm proteins in both Drosophila and mammals. At present, its details of functions in animal growth and spermatogenesis have not been reported. In this study, we knocked down ATPsyn‐b using Drosophila lines expressing inducible hairpin RNAi constructs and Gal4 drivers. Ubiquitous knockdown of ATPsyn‐b resulted in growth defects in larval stage as the larvae did not grow bigger than the size of normal second‐instar larvae. Knockdown in testes did not interrupt the developmental excursion to viable adult flies, however, these male adults were sterile. Analyses of testes revealed disrupted nuclear bundles during spermatogenesis and abnormal shaping in spermatid elongation. There were no mature sperm in the seminal vesicle of ATPsyn‐b knockdown male testes. These findings suggest us that ATPsyn‐b acts in growth and male fertility of Drosophila.     

Ecdysteroid production by testes of the tobacco budworm,Heliothis virescens,from last larval instar to adult

Testes of Heliothis virescens were examined in vitro for the ability to produce ecdysteroids. Production was detected in testes removed from larvae at mid and late periods in the last instar, in testes from pupae after the third day of purpal development, and in testes taken from young adult males. The testes in the isolated abdomens do not fuse during later stages of spermatogenesis as they do in intact insects. Spermatocysts in testes of larval abdomens isolated from the upper body early in the last instar developed to mature sperm despite low haemolyph and testis ecdysteroid titres. Gonadectomized animals developed and moulted normally, indicating that ecdysteroids from the testes are not essential to general development.     

玉竹雄配子的发育——着重阐明质体在生殖细胞和营养细胞中的分布和变化

玉竹(Polygonatum simizui Kitag)小孢子在分裂前,质体极性分布导致分裂后形成的生殖细胞不含质体,而营养细胞包含了小孢子中全部的质体。生殖细胞发育至成熟花粉时期,及在花粉管中分裂形成的两个精细胞中始终不含质体。虽然生殖细胞和精细胞中都存在线粒体,但细胞质中无DNA类核。玉竹雄性质体的遗传为单亲母本型。在雄配子体发育过程中,营养细胞中的质体发生明显的变化。在早期的营养细胞质中,造粉质体增殖和活跃地合成淀粉。后期,脂体增加而造粉质体消失。接近成熟时花粉富含油滴。对百合科的不同属植物质体被排除的机理及花粉中贮藏的淀粉与脂体的转变进行了讨论。     

PROTOXYLEM MATURATION IN THE SEEDLING LEAF OF WHEAT

Protoxylem (PX) maturation was followed in vascular bundles of the first foliage leaf (L#1) of wheat seedlings, using clearings and sectioned materials to assess development changes. L#1 contained seven to 11 longitudinal bundles. The seven bundles with largest diameters contained PX vessels, but the number of bundles with PX varied with leaf length. Repeated vessel maturation maintained PX continuity as older vessels collapsed due to leaf extension. Thus, the number of PX vessels in each bundle was also a function of leaf length, and the PX content of a bundle was a function of the stage of leaf development and how soon during leaf development the bundle attained mature PX. Distance from the leaf tip to the start point for PX was greater for bundles later to mature. Thus, the pattern of distal start points in mature leaves reflected the progression of PX maturation from the midrib to bundles of lower rank. It is suggested that the involvement of more and more bundles in PX maturation as the leaf ages is a general occurrence in grass leaves, and that assessing how many bundles will have PX requires observations late in leaf development.     

Correlation between spermathecal morphology and mating systems in spiders

This study tested predictions regarding male mating preferences which were based on some aspects of female reproductive morphology which may influence sperm precedence patterns in six species of spiders. Males of two species, whose 'conduit' spermathecal design has been associated in previous studies with first male sperm precedence, showed the predicted preference for associating with immature females about to moult to maturity rather than mature females. Those of a third species, however, associated indiscriminately with mature and penultimate instar females. As predicted, males of three other species with 'cul-de-sac' spermathecal morphology did not associate preferentially with immature females. Immature females were avoided in two of the species, but not in the third. One of the species with cul-de-sac spermathecae showed, as predicted, lack of a strong first male advantage in sperm precedence. These data give only limited confirmation of the predictions.