中华卵索线虫雌雄寄生后期幼虫基因差异表达分析

索科线虫是一类具有很大生防潜力的昆虫(如棉铃虫等)天敌资源,但由于其体外培养尚未获得成功,阻碍了商业化生产和大规模应用,究其原因主要是体外培养的线虫不能完成雌雄性别分化。因此,研究该类线虫性别分化机制成为近年本领域的热点课题。该文以中华卵索线虫为实验材料,采用mRNA差异显示的方法,分析了中华卵索线虫性别分化关键时期雌、雄寄生后期幼线虫的基因表达差异。在雌雄线虫体内得到具有表达差异的基因片段20条。其中8条在雄虫体内特异性表达,12条在雌虫体内特异性表达。利用信息生物学技术对所分离到的差异片段进行了序列分析。其中,Ensembl分析发现4个片段与秀丽线虫X染色体具有可匹配片段,推测这些片段可能是影响中华卵索线虫性别分化的重要基因。这些结果将为后续研究提供思路。     

寄生于粘虫的卵索线虫属一新种——中华卵索线虫(线虫纲:索科)

本文报道从河南省驻马店地区和山东省临沂地区麦田内采集的粘虫幼虫的寄生线虫优势种卵索线虫属一新种,命名为中华卵索线虫,新种Ovomermis sinensis sp.nov.     

棉铃虫感染中华卵索线虫后血淋巴酚氧化酶活性的变化及其分离纯化

研究了中华卵索线虫Ovomermis sinensis感染棉铃虫Helicoverpa armigera幼虫后宿主体内酚氧化酶活性的变化。研究结果表明,在感染后的第1天,中华卵索线虫的侵入引起酚氧化酶活性的增加,感染组酶活性是同期对照组的1.12倍; 但在随后的寄生期间,中华卵索线虫抑制了宿主的酚氧化酶活性,其中以第5天的抑制最为强烈： 同期对照组酶活性是感染组的1.52倍。对酚氧化酶进行了初步的分离纯化,纯化倍数为41.5倍,酶得率为12.7%,比活力为4 030.6 U/mg。     

中华卵索线虫感染棉铃虫后脂肪体结构的变化

采用组织超薄切片技术,电镜下观察研究昆虫棉铃虫Helicoverpa armigera Hǖbner脂肪体细胞的超微结构,结果表明。在中华卵索线虫Ovomermis sinensisChen感染棉铃虫后的不同寄生时期内,宿主脂肪体细胞内的线粒体、内质网、脂肪球和细胞核等细胞器在形态和结构上均有较大的病理变化,这些病理变化,与这一时期中华卵索线虫的生长发育和寄生生活等特点密切相关。     

中华卵索线虫的研究与应用

中华卵索线虫Ovomermis sinensis Chen etal.作为一种昆虫病原线虫,在农业害虫的生物防治中起着重大作用。文章综述90年代以来我国有关中华卵索线虫的生物学特性、田间应用、体内外培养以及生化与分子生物学研究进展,并对中华卵索线虫今后的研究方向提出建议。     

中华卵索线虫雌雄成虫可溶性蛋白双向电泳分析

昆虫寄生线虫 (又称虫生线虫、昆虫病原线虫)是本世纪发展起来的一种有潜能的生物防治因子 ,它寄主范围广、能主动寻找寄主、对人畜及环境安全无毒。中华卵索线虫是昆虫寄生线虫的一种 ,由我国学者发现并命名的新种 (陈果等 ,1 9 91 ) ,具有广泛的寄主范围 ,可杀灭粘虫 (任慧芳等 ,1 989)、烟青虫、烟蚜 (侯茂林等 ,2 0 0 2 )及棉铃虫 (陈果 ,1 994)等危害十分严重的害虫 ,其寄生率等于宿主的死亡率。因此在生物防治手段日益突出的今天 ,中华卵索线虫无疑有着广泛的应用前景。然而 ,在线虫培养的过程中 ,无论是体内培养还是体外培养 ,我们发…     

棉铃虫感染中华卵索线虫后血淋巴游离氨基酸含量的变化

中华卵索线虫Ovomermis sinensis感染棉铃虫Helicoverpa armigera 1天后,棉铃虫幼虫血淋巴中游离氨基酸总量大幅度下降,各种游离氨基酸的含量也是如此变化。感染2～4天后的棉铃虫血淋巴中游离氨基酸总量变化不大,各种游离氨基酸的含量有升有降。感染5～6天后的棉铃虫血淋巴中游离氨基酸总量和各种氨基酸的含量都急剧上升。研究表明,中华卵索线虫寄生棉铃虫时,棉铃虫血淋巴中游离氨基酸含量的变化朝着有利于线虫生长发育的方向进行。     

中华卵索线虫的体外培养

在研究中华卵索线虫的体外培养方法的同时,对其在不同培养基中的生长发育情况进行了观察。结果表明：以培养基TC－199加20％热灭活胎牛血清的培养效果较为理想,大多数线虫可存活3个月,最大虫体长55．1mm,宽204．13um,其发育程度大致与该种索线虫在宿主粘虫体内寄生8－9天的情况相近,培养期间观察到2次蜕皮;第一次蜕皮在卵内,第二次在培养6－8天之后,口针消失,虫体内滋养物体发育明显,尾部附器已经形成,没有观察到生殖原基的发育。     

对稻飞虱寄生天敌——两索线虫的初步观察

<正> 经田间与室内观察,初步查明两索线虫Amphimermis sp.是寄生稻飞虱的较好天敌。褐稻虱被两索线虫寄生后双晚稻田第五、六代发生量可减少34.5±5.72％。现将观察结果整理如下。 一、两索线虫在稻田土壤内的分布 两索线虫除寄生期幼虫在寄主稻飞虱体内     

五种索科线虫RAPD亲缘关系分析

采用RAPD技术构建了索科线虫4属5种的指纹图谱。从47个引物中筛选出12个稳定性好、多态性高的引物,共扩增出161条谱带,其中150条谱带具遗传多态性,占93·17%。所获片段长度大小为200~3200bp,单个引物扩增的条带数在11~16之间,平均为13·42条。采用RAPDistance软件及MEGA程序,计算Nei氏相似系数和遗传距离,建立UPGMA和NJ聚类图,两个聚类图拓扑结构相同,将5种索科线虫分为两大分支:同属于蚊幼寄生罗索属线虫的食蚊罗索线虫(Romanomermisculicivorax)与武昌罗索线虫(R.wuchangensis)亲缘关系最近,先聚在一起,再与同翅目(Homoptera)寄生长沙多索线虫(Agamermischang-shaensis)聚为一支;鳞翅目(Lepidoptera)寄生中华卵索线虫(Ovomermissinensis)和同翅目寄生两索属线虫(Amphimermissp·)亲缘关系较近,两者聚为一支。5种索线虫属内种间的遗传距离较小,食蚊罗索线虫与武昌罗索线虫之间遗传距离仅为0·1789;而属间遗传距离较大,在0·4471~0·5488之间。上述结果表明:RAPD技术可以应用于索科线虫亲缘关系的分析,能够反映出不同线虫间的遗传差距,从而成功地进行属、种的分类及进化问题研究。     

雌激素对中华鳖性腺分化及DMRT1、SOX9基因表达的影响

中华鳖（Pelodiscus sinensis）性别决定的方式一直存在较大的争议,分子机制更是不清楚。在大部分脊椎动物中,雌激素在性别决定和性腺分化中扮演重要的调控作用。实验通过对性别分化前胚胎进行雌二醇（E2）和芳香化酶抑制剂（AI）处理,研究雌激素在中华鳖性腺分化中的作用及机理。实验结果显示,与对照组（雌性比例49%）相比,E2处理组中雌性中华鳖仔鳖比例显著增加,高达92.3%;而在AI处理组中,雌性比例显著下调至13.1%。HE染色分析表明,ZZ（雄性）和ZW（雌性）胚胎分别经过E2和AI处理后,ZZ和ZW性腺结构呈现明显的雌性化和雄性化特征。同时,通过RT-PCR和免疫荧光染色发现,E2能显著降低雄性性别关键因子DMRT1和SOX9 mRNA和蛋白表达水平;AI则表现相反的调节作用。综上所述,雌激素通过抑制雄性性别关键因子DMRT1和SOX9的表达来抑制雄性分化,促进雌性分化,揭示雌激素在中华鳖雌性性别分化中起着重要的调控作用。       

中华鳖Dmrt1基因的克隆、表达及在性逆转中的变化分析

在大部分脊椎动物中,Dmrt1基因在雄性性别决定和性腺分化中起重要的调控作用.本文从m RNA和蛋白水平分析Dmrt1基因的组织差异性表达、在不同发育阶段性腺中的细胞定位及在性逆转中的表达变化,研究Dmrt1基因在中华鳖性别分化中的调控作用.Rapid-amplification of c DNA ends(RACE)结果显示,Dmrt1基因c DNA序列全长2409 bp,其中5′非编码区为230 bp,3′非编码区为1072 bp,开放阅读框为1107 bp,编码368个氨基酸,具有一个高度保守的DM结构域.荧光定量PCR和免疫组化结果显示,Dmrt1在性腺分化之前的第16期雄性性腺中开始表达,先于Amh和Sox9基因表达.随着性腺的发育,Dmrt1蛋白主要定位于性腺Sertoli细胞的细胞核上,在雌性性腺发育过程中并未见其表达.此外,在雌二醇诱导的雄性转雌性性逆转胚胎性腺中,Dmrt1表达显著下调;在芳香化酶抑制剂诱导的雌性转雄性性腺中,Dmrt1表达则显著上升.上述研究表明,Dmrt1基因是中华鳖雄性特异性基因,参与雄性性腺的发育过程,可能在中华鳖早期性别决定中起重要的调控作用.     

Ontogeny of sexual development in the roach (Rutilus rutilus) and its interrelationships with growth and age

The roach (Rutilus rutilus) has become a sentinel species for the study of sexual disruption in wild fish populations as a consequence of exposure to endocrine disrupting chemicals (EDCs). Little is known, however, about the normal ontogeny of sexual development in this species. Here, we analyzed the ontogeny of sexual development in captive‐bred roach and assessed how growth rate and fish size affected the timing of both sexual differentiation and sexual development over a 2‐year period. Ovarian differentiation was first recorded at 68 days post‐fertilization (dpf) and this preceded testicular differentiation (first recorded at 98 dpf). In contrast, sexual maturation occurred at an earlier age in males (300 dpf) compared with females (728 dpf). No differences in body size (length or weight) were recorded between male and female roach until the fish were 415 dpf. Studies on three populations of roach which grew at different rates showed that the timing of sexual differentiation was highly variable and more related to fish size than to fish age. Time to sexual maturation was also variable among populations but, subsequent to their first year of life, gonadal status was less well associated with fish size. Interestingly, the sex ratio of the population was biased towards females in populations that grew more rapidly during early life. The findings presented here provide a valuable foundation of work to support both field‐ and laboratory‐based assessments on the effects of EDCs, and other stressors, on sexual differentiation and development in the roach. J. Morphol., 2008. © 2008 Wiley‐Liss, Inc.     

冬虫夏草菌和蛹虫草菌的研究现状、问题及展望

冬虫夏草菌和蛹虫草菌是两种最著名的虫草菌。从分类学地位、分布、生活史及有性生殖类型、寄主范围、遗传多样性、分子遗传学和基因组学、生态学、人工栽培及产品开发等方面对冬虫夏草菌和蛹虫草菌的研究现状进行总结,指出了研究中存在的一些问题,并对研究前景进行展望。     

Sexual differentiation in three unconventional mammals: spotted hyenas, elephants and tammar wallabies

The present review explores sexual differentiation in three non-conventional species: the spotted hyena, the elephant and the tammar wallaby, selected because of the natural challenges they present for contemporary understanding of sexual differentiation. According to the prevailing view of mammalian sexual differentiation, originally proposed by Alfred Jost, secretion of androgen and anti-Mullerian hormone (AMH) by the fetal testes during critical stages of development accounts for the full range of sexually dimorphic urogenital traits observed at birth. Jost's concept was subsequently expanded to encompass sexual differentiation of the brain and behavior. Although the central focus of this review involves urogenital development, we assume that the novel mechanisms described in this article have potentially significant implications for sexual differentiation of brain and behavior, a transposition with precedent in the history of this field. Contrary to the "specific" requirements of Jost's formulation, female spotted hyenas and elephants initially develop male-type external genitalia prior to gonadal differentiation. In addition, the administration of anti-androgens to pregnant female spotted hyenas does not prevent the formation of a scrotum, pseudoscrotum, penis or penile clitoris in the offspring of treated females, although it is not yet clear whether the creation of masculine genitalia involves other steroids or whether there is a genetic mechanism bypassing a hormonal mediator. Wallabies, where sexual differentiation occurs in the pouch after birth, provide the most conclusive evidence for direct genetic control of sexual dimorphism, with the scrotum developing only in males and the pouch and mammary glands only in females, before differentiation of the gonads. The development of the pouch and mammary gland in females and the scrotum in males is controlled by genes on the X chromosome. In keeping with the "expanded" version of Jost's formulation, secretion of androgens by the fetal testes provides the best current account of a broad array of sex differences in reproductive morphology and endocrinology of the spotted hyena, and androgens are essential for development of the prostate and penis of the wallaby. But the essential circulating androgen in the male wallaby is 5alpha androstanediol, locally converted in target tissues to DHT, while in the pregnant female hyena, androstenedione, secreted by the maternal ovary, is converted by the placenta to testosterone (and estradiol) and transferred to the developing fetus. Testicular testosterone certainly seems to be responsible for the behavioral phenomenon of musth in male elephants. Both spotted hyenas and elephants display matrilineal social organization, and, in both species, female genital morphology requires feminine cooperation for successful copulation. We conclude that not all aspects of sexual differentiation have been delegated to testicular hormones in these mammals. In addition, we suggest that research on urogenital development in these non-traditional species directs attention to processes that may well be operating during the sexual differentiation of morphology and behavior in more common laboratory mammals, albeit in less dramatic fashion.     

Organizational–Activational Concept Revisited: Sexual Differentiation in an Atherinomorph Teleost

Because of its numerous sexually dimorphic characters, the Western mosquitofishGambusia affinis affinisis an excellent vertebrate system for addressing questions concerning sexual differentiation. In this review, the actions and limits of gonadal sex steroids, specifically testosterone, on the development of the sexually dimorphic anal fin and its axial and appendicular support are described. Data from our laboratory show that the critical period in this species for the anterior transposition of the male anal fin and its appendicular support occurs during the late embryonic period and that this differentiation of the male phenotypic genital areaduring the critical period is regulated by androgen.     

Potential contribution of progesterone receptors to the development of sexual behavior in male and female mice

We previously showed that estradiol can have both defeminizing and feminizing effects on the developing mouse brain. Pre- and early postnatal estradiol defeminized the ability to show lordosis in adulthood, whereas prepubertal estradiol feminized this ability. Furthermore, we found that estradiol upregulates progesterone receptors (PR) during development, inducing both a male-and female-typical pattern of PR expression in the mouse hypothalamus. In the present study, we took advantage of a newly developed PR antagonist (ZK 137316) to determine whether PR contributes to either male- or female-typical sexual differentiation. Thus groups of male and female C57Bl/6j mice were treated with ZK 137316 or OIL as control: males were treated neonatally (P0–P10), during the critical period for male sexual differentiation, and females were treated prepubertally (P15–P25), during the critical period for female sexual differentiation. In adulthood, mice were tested for sexual behavior. In males, some minor effects of neonatal ZK treatment on sexual behavior were observed: latencies to the first mount, intromission and ejaculation were decreased in neonatally ZK treated males; however, this effect disappeared by the second mating test. By contrast, female mice treated with ZK during the prepubertal period showed significantly less lordosis than OIL-treated females. Mate preferences were not affected in either males or females treated with ZK during development. Taken together, these results suggest a role for PR and thus perhaps progesterone in the development of lordosis behavior in female mice. By contrast, no obvious role for PR can be discerned in the development of male sexual behavior.     

Sexual differentiation in the spiny softshell turtle (Apalone spinifera), a species with genetic sex determination

It is hypothesized on the basis of sex determination theory that species exhibiting genetic sex determination (GSD) may undergo sexual differentiation earlier in development than species with environmental sex determination (ESD). Most turtle species exhibit a form of ESD known as temperature-dependent sex determination (TSD), and in such species the chronology of sex differentiation is well studied. Apalone spinifera is a species of softshell turtle (Trionychidae) that exhibits GSD. We studied sexual differentiation in this species in order to facilitate comparison to TSD species. Eggs were incubated at two different temperatures and embryos were harvested at various stages of mid to late development. Gonad length was measured with image analysis software, then prepared histologically. Indifferent gonads have differentiated in stage 19 embryos. Histological details of gonadogenesis follow the same pattern as described for other reptiles. Regression of the male paramesonephric duct closely follows testicular differentiation. Gonad lengths are longer at the warmer incubation temperature, and ovaries are generally longer than testes at each stage and for each temperature. Although sexual differentiation takes place at about the same stage as in other turtles with TSD (18-20), in A. spinifera this differentiation is irreversible at this stage, while in some of the TSD species sex is reversible until about stage 22. This immutable, definitive sexual differentiation may support the hypothesis of an accelerated chronology of sex differentiation for this species. We also note that sexual dichromatism at hatching is known in this species and may provide additional evidence of early differentiation. J. Exp. Zool. 290:190-200, 2001.     

中国小达氏涡虫和大变杰氏涡虫的生物学特性

对单肠目(Rhabdocoela)达氏科(Dalyelliidae)的中国小达氏涡虫(Microdalyellia sinensis)和大变杰氏涡虫(Gieysztoria macrovariata9-spinosa)进行了长期饲养与观察,了解和比较这两种涡虫的习性、繁殖、发育及组织学特点。结果表明,中国小达氏涡虫比大变杰氏涡虫反应敏捷,生殖器官的形态位置有明显区别;中国小达氏涡虫产卵周期约5 d,每期产卵10.5枚;大变杰氏涡虫产卵周期约10 d,每期产卵22.3枚;两种涡虫卵孵化期约60 h,发育成熟期9 d左右;最后探讨了卵胚非正常发育与雌雄生殖器官成熟期,以及涡虫的淡水生态学意义等问题。     

The novel putative transporter NPT1 plays a critical role in early stages of Plasmodium berghei sexual development

Transmission of Plasmodium species from a mammalian host to the mosquito vector requires the uptake, during an infected blood meal, of gametocytes, the precursor cells of the gametes. Relatively little is known about the molecular mechanisms involved in the developmental switch from asexual development to sexual differentiation or the maturation and survival of gametocytes. Here, we show that a gene coding for a novel putative transporter, NPT1, plays a crucial role in the development of Plasmodium berghei gametocytes. Parasites lacking NPT1 are severely compromised in the production of gametocytes and the rare gametocytes produced are unable to differentiate into fertile gametes. This is the earliest block in gametocytogenesis obtained by reverse genetics and the first to demonstrate the role of a protein with a putative transport function in sexual development. These results and the high degree of conservation of NPT1 in Plasmodium species suggest that this protein could be an attractive target for the development of novel drugs to block the spread of malaria.