Comparative studies on histological and ultra-structure of the pituitary of different ploidy level fishes

The histological and ultra-structure of the pituitary in diploid red crucian carp(Carassius auratus red var.),triploid crucian carp and allotetraploid hybrids within and after the breeding season were comparatively studied.The result showed that there were six endocrine cell types in the pituitary of these three kinds of fishes,and there was an obvious difference in cell size among different ploidy level fishes.As for the same type of pituitary cells,the cell size was increased gradually with the in- creasing ploidy level.In the breeding season,the allotetraploid hybrids had higher proportion of go- nadotropin cells(GTH)than triploids,and the triploids had higher proportion of GTH than diploids.The results were related to the earlier sexual maturity of allotetraploid hybrids and sterility of triploid cru- cian carp.On the other hand,among the three kinds of fishes,the proportion of somatotropin(STH) cells in triploids crucian carp was the highest,whereas that in allotetraploid hybrids was the lowest. The results might be connected with the faster growth rate of triploids and slower growth rate of al- lotetraploid hybrids.In addition,in GTH cells of meso-adenohypophysis after the breeding season, there were many endocrine particles in triploids,while those endocrine particles were released from the cells in allotetraploids and diploids.This result showed that the sterility of triploid crucian carp might be related to the hormone which was not released from the GTH cells.In a word,the present study indicated that the differences in the structure of pituitary among different ploidy level fishes contributed to their difference in the growth rate and gonadal development.     

The cloning of Dmc1 cDNAs and a comparative study of its expression in different ploidy cyprinid fishes

Dmc1 (disrupted meiotic cDNA) is a functionally specific gene, which was firstly discovered in yeast and then found to encode a protein required for homologous chromosome synapsis during the process of meiosis. In this investigation, we cloned the partial cDNAs of Dmc1 of diploid red crucian carp, Japanese crucian carp, common carp, triploid crucian carp and allotetraploid hybrids by using a pair of degenerate primers based on the conservative sequence of amino acids of the DMC1 protein in yeast, mouse and human. The full length cDNAs were then obtained by rapid amplification of cDNA ends (RACE). Our data showed that the full length cDNAs of Dmc1 in the three diploid fishes are all 1375 bp long, while it is 1383 bp long in triploids and 1379 bp long in allotetraploids. And despite of the variation in length, all the cDNAs encode a protein of 342 amino acids. A high homology of 97.3% of the DMC1 protein can be drawn by comparing the amino acid sequences in the three diploids, which is also of 86%, 86% and 95% similarity to human, mouse and zebrafish, respectively. A comparative study of the expression pattern of Dmc1 was carried out by RT-PCR using specific primers against the same sequences of coding regions in different ploidy cyprinid fishes, from which it was showed that Dmc1 was expressed only in gonads of these five kinds of fishes. The expression pattern of Dmc1 in both ovaries and testes from different ploidy fishes within breeding season was also studied by Real-time PCR, and the results showed that the expression of this gene was greatly different among the three different ploidy fishes, which was the highest of triploid and lowest of allotetraploids. The histological sections data showed matured gonads of both diploid red crucian carp and allotetraploids in breeding season, although the latter demonstrated a higher maturation, and no gonadal maturation could be observed in triploids. In conclusion, we suggest that Dmc1 is specifically expressed in the period of meiosis in all the ploidy cyprinid fishes and directly related with the development of gonad in a manner of ploidy-independent way. And further, the high expression of Dmc1 in female triploids might be associated with abnormal meiosis and sterility.     

The cloning of Dmc1 cDNAs and a comparative study of its expression in different ploidy cyprinid fishes

Dmc1 (disrupted meiotic cDNA) is a functionally specific gene, which was firstly discovered in yeast and then found to encode a protein required for homologous chromosome synapsis during the process of meiosis. In this investigation, we cloned the partial cDNAs of Dmc1 of diploid red crucian carp, Japanese crucian carp, common carp, triploid crucian carp and allotetraploid hybrids by using a pair of degenerate primers based on the conservative sequence of amino acids of the DMC1 protein in yeast, mouse and human. The full length cDNAs were then obtained by rapid amplification of cDNA ends (RACE). Our data showed that the full length cDNAs of Dmc1 in the three diploid fishes are all 1375 bp long, while it is 1383 bp long in triploids and 1379 bp long in allotetraploids. And despite of the variation in length, all the cDNAs encode a protein of 342 amino acids. A high homology of 97.3% of the DMC1 protein can be drawn by comparing the amino acid sequences in the three diploids, which is also of 86%, 86% and 95% similarity to human, mouse and zebrafish, respectively. A comparative study of the expression pattern of Dmc1 was carried out by RT-PCR using specific primers against the same se-quences of coding regions in different ploidy cyprinid fishes, from which it was showed that Dmc1 was expressed only in gonads of these five kinds of fishes. The expression pattern of Dmc1 in both ovaries and testes from different ploidy fishes within breeding season was also studied by Real-time PCR, and the results showed that the expression of this gene was greatly different among the three different ploidy fishes, which was the highest of triploid and lowest of allotetraploids. The histological sections data showed matured gonads of both diploid red crucian carp and allotetraploids in breeding season, although the latter demonstrated a higher maturation, and no gonadal maturation could be observed in triploids. In conclusion, we suggest that Dmc1 is specifically expressed in the period of meiosis in all the ploidy cyprinid fishes and directly related with the development of gonad in a manner of ploidy-independent way. And further, the high expression of Dmc1 in female triploids might be associ-ated with abnormal meiosis and sterility.     

Distant hybridization leads to different ploidy fishes

Distant hybridization makes it possible to transfer the genome of one species to another, which results in changes in phenotypes and genotypes of the progenies. This study shows that distant hybridization or the combination of this method with gynogenesis or androgenesis lead to different ploidy fishes with genetic variation, including fertile tetraploid hybrids, sterile triploid hybrids, fertile diploid hybrids, fertile diploid gynogenetic fish, and their derived progenies. The formations of the different ploidy fishes depend on the genetic relationship between the parents. In this study, several types of distant hybridization, including red crucian carp (Carassius auratus red var.) (2n=100, abbreviated as RCC) (♀)×common carp (Cyprinus carpio L.) (2n=100, abbreviated as CC) (♂), and RCC (2n=100) (♀)×blunt snout bream (Megalobrama amblycephala) (2n=48, abbreviated as BSB) (♂) are described. In the distant hybridization of RCC (♀)×CC (♂), bisexual fertile F3–F18 allotetraploid hybrids (4n=200, abbreviated as 4nAT) were formed. The diploid hybrid eggs and diploid sperm generated by the females and males of 4nAT developed into diploid gynogenetic hybrids and diploid androgenetic hybrids, respectively, by gynogenesis and androgenesis, without treatment for doubling the chromosome. Improved tetraploid hybrids and improved diploid fishes with genetic variation were derived from the gynogenetic hybrid line. The improved diploid fishes included the high-body RCC and high-body goldfish. The formation of the tetraploid hybrids was related to the occurrence of unreduced gametes generated from the diploid hybrids, which involved in premeiotic endoreduplication, endomitosis, or fusion of germ cells. The sterile triploid hybrids (3n=150) were produced on a large scale by crossing the males of tetraploid hybrids with females of diploid fish (2n=100). In another distant hybridization of RCC (♀)×BSB (♂), different ploidy fishes were obtained, including diploid bisexual fertile natural gynogenetic fish (2n=100), sterile triploid hybrids (3n=124), and bisexual fertile tetraploid hybrids (4n=148). Furthermore, two kinds of pentaploid hybrids (5n=172 and 5n=198) were formed. The biological characteristics and the mechanisms of formation of the different ploidy fish were compared and discussed at the cellular and molecular level. The results indicated distant hybridization or the combination of this method with gynogenesis or androgenesis affects the formation of different ploidy fish with genetic variation.     

异源四倍体鲫鲤、三倍体湘云鲫和它们父母本线粒体DNA 12S rRNA基因遗传变异的分析

采用质粒克隆测序方法,获得了异源四倍体鲫鲤5个个体、异源四倍体鲫鲤雌核发育二倍体后代2个个体、三倍体湘云鲫2个个体及红鲫、湘江野鲤和日本白鲫各1个个体的线粒体DNA 12S rRNA基因的全序列。经对比发现,异源四倍体5个个体共享2种单元型,异源四倍体鲫鲤雌核发育二倍体后代2个个体、三倍体湘云鲫2个个体以及红鲫、湘江野鲤和日本白鲫各1个个体分别共享1种单元型。用MEGA 1.0 软件分析了它们的碱基组成和核苷酸序列差异,用邻接法构建系统进化树。它们间的序列同源性在95%~99%之间,异源四倍体鲫鲤、三倍体湘云鲫和它们母本（分别为红鲫和日本白鲫）之间的序列同源性大于异源四倍体鲫鲤、三倍体湘云鲫和它们父本（分别为湘江野鲤和异源四倍体鲫鲤）之间的序列同源性,结果表明：异源四倍体鲫鲤和三倍体湘云鲫在线粒体DNA 12S rRNA基因上具有母性遗传特征。本研究另一值得注意地方的是异源四倍体鲫鲤经过9代(F3-F11)繁殖后,在5个个体中发现了2种单元型,说明在四倍体基因库中存在遗传多样性,为四倍体基因库的繁殖、保护和种群复壮提供了一些有价值的信息。     

A review of ploidy in the genus Prosopis (Leguminosae)

The genus Prosopis contains 44 species of trees and shrubs, the majority of which originate in the Americas. Most species are reported to be diploid, with a somatic chromosome number of 2 n  = 28. There are rare reports of polyploidy, although it is thought that these may represent polysomaty in root tissues. However, flow cytometry has recently indicated that P. juliflora is entirely tetraploid with a somatic chromosome number of 2 n  = 56. In order to clarify the situation, an extensive review of ploidy in Prosopis was undertaken, the first of its kind. The ploidy levels of 124 samples of Prosopis from 21 countries, including both the natural and introduced ranges, were analysed using flow cytometry. In addition, a comprehensive literature review was carried out, examining 305 published ploidy values and covering 32 of the 44 species of Prosopis . Flow cytometry analysis suggests that P. juliflora is the only tetraploid species, with a somatic chromosome number of 2 n  = 4 x  = 56, whilst the remainder of the species analysed are diploid with 2 n  = 2 x  = 28, including the first report for P. articulata (2 n  = 28). A critical review of published ploidy values shows that all species of Prosopis are reported to be entirely diploid, except P. glandulosa , P. juliflora , and P. koelziana , for which both diploid and tetraploid values have been recorded. © 2008 The Linnean Society of London, Botanical Journal of the Linnean Society , 2008, 156 , 425–438.     

NORs and counterstain-enhanced fluorescence studies in Cyprinidae of different ploidy level

The ribosomal RNA gene expression in the genomes of evolutionary diploid (Scardinius erythrophthalmus, Leucaspius delineatus, Tinca tinca) and polyploid species (Cyprinus carpio, Carassius carassius, Carassius auratus gibelio, Carassius auratus auratus) of Cyprinidae has been investigated by means of a silver nitrate technique. The diploid species investigated exhibited only one pair of chromosomes with nucleolus organizers (NOR). Higher numbers of rRNA-expressing chromosomal sites in several evolutionary polyploid species (Carassins) gave evidence against a complete functional diploidization, at least with regard to the NOR bearing chromosomes in these species. The NORs displayed a heterochromatic brilliant chromomycin A₃ fluorescence. No distamycin-A/DAPI-bright heterochromatic blocks were detected in the genomes of the Cyprinidae.     

Comparative and Evolutionary Analysis of the Cytochrome b Sequences in Cyprinids with Different Ploidy Levels Derived from Crosses

The mitochondrial cyt b genes in the allotetraploid and triploid crucian carp as well as triploid common carp were isolated and completely sequenced. Their DNA sequences were compared with those derived from the cyt b genes of the red crucian carp, Japanese crucian carp, and common carp with MEGA 1.0 software. Phylogenetic analysis revealed the sister relationships between allotetraploid and diploid red crucian carp, between the triploid crucian carp and diploid Japanese crucian carp, and between triploid common carp and diploid common carp. Our results indicated the cyt b genes in the allotetraploid, triploid crucian carp, and triploid common carp were maternally inherited. Through maternal inheritance, the cyt b gene in the F11 tetraploid displayed extremely high similarity to that in the female parent red crucian carp after 11 generations (from F1 to F11 hybrids). Since the establishment of the new tetraploid stocks has great significance in analyzing evolutionary theory of vertebrate and in improving aquaculture industry, analysis of the cyt b gene and the elucidation of the variation of the cyt b gene DNA in different cyprinids prove that cyt b is a useful genetic marker to monitor the variations in the progeny of the crosses.     

麻疯树种子发育过程中脂类物质变化的显微和超微结构观察

用石蜡切片、半薄切片和超薄切片方法研究了麻疯树种子发育过程中脂类物质的变化。结果表明:脂类物质主要储存于胚乳当中,当种子发育成熟时胚乳中迅速积累了大量的油脂;种皮在发育过程中脂类物质含量较多,成熟时外种皮硬化,内种皮含有大量的油脂;种子成熟时胚中含有少量的脂类物质。细胞内脂类物质含量比较多时,内质网、线粒体、质体和高尔基体数量也会较多。种子完全成熟时进行采收加工是最为合适的。     

The effect of ploidy level on fitness in parthenogenetic flatworms

Although polyploidy plays an important role in speciation, its impact on fitness is still debated. One problem is that its adaptive significance can only be inferred by comparing forms with different ploidy that are identical in all other traits. This situation is uncommon, presumably because ploidy types often differ in reproduction mode, genetic background or habitat. Here we compare fitness in a system of triploid and tetraploid karyotypes of the planarian flatworm Schmidtea polychroa . Both types have the same type of sperm-dependent parthenogenesis and share the same genetic background and habitat. Hence, fitness differences, if any, can be attributed to different ploidy levels only. Contrary to the general assumption of a positive correlation between fitness and ploidy level, we showed that triploids produced 58% more offspring than tetraploids. Within each ploidy type, we identified groups of highly related clones using microsatellites. Significant variation among clonal groups in body size, offspring and cocoon number and hatching time indicated a genetic basis for variance in these traits. A small model shows that despite low fitness of tetraploids, stable coexistence of triploids and tetraploids can be explained by the recurrent origin of triploids from tetraploids and vice versa.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 85 , 191–198.     

稻飞虱酵母类胞内共生菌的组织学研究进展

褐飞虱Nilaparvata lugens体内酵母类共生菌在褐飞虱生长发育和繁殖中起着重要作用,并影响着褐飞虱对寄主植物的致害性变异。该文系统总结了国内外已有的研究成果,讨论了褐飞虱共生菌的分类地位、形态特征、在寄主发育中的功能、侵染途径等,提出了开展褐飞虱生物型形成过程中体内共生菌变异的研究建议,以便利用共生菌来监测田间不同生物型褐飞虱的发生情况,最终实现控菌防虫和对不同生物型的准确预测。     

Stomatal length and frequency as a measure of ploidy level in black wattle, Acacia mearnsii (de Wild)

The length and frequency of stomata on leaf surfaces were examined as rapid techniques for future identification of ploidy level of Acacia mearnsii (de Wild). Diploid (2 n  = 2 x  = 26) and tetraploid (2 n  = 4 x  = 52) plants were germinated from chipped seed at 25°C and grown under nursery conditions. After one month, measurements showed that the mean stomatal length was 27.17 ± 0.474 µm for diploids and 40.24 ± 0.521 µm for tetraploids and these differed significantly from each other ( P  < 0.001). The frequency of stomata per leaf surface was shown to decrease significantly ( P  < 0.001) as the ploidy level increased, with a mean of 22.11 ± 0.495 for diploids and 10.26 ± 0.495 for tetraploids. It was concluded that stomatal length and stomatal frequency are rapid indirect methods to identify ploidy level in black wattle.  © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society , 2003, 141 , 177–181.     

Effect of gonadotropin-releasing hormone on estrogen receptor messenger ribonucleic acid level in perifused pituitary cells

Summary 1. We examined the potential effect of GnRH pulses on pituitary estrogen receptor mRNA level.2. The treatment of perifused pituitary cell aggregates with four hourly pulses of GnRH (10 nM/1 min/h) resulted in a marked increase in the steady-state level of ER mRNA (25%vs unstimulated control, n = 3).3. No changes were observed for the LH ß mRNA. Data suggest, for the first time, that a cross-talk between the GnRH and nuclear ER may occur in the gonadotrope cells.     

沙棘木蠹蛾生殖系统形态学和组织学观察

沙棘木蠹蛾Holcocerus hippophaecolus Hua,Chou,Fang et Chen属鳞翅目(Lepidoptera)木蠹蛾科(Cossidae),是我国的特有种.2001年以来,沙棘木蠹蛾在我国大面积爆发,是屹今为止我国沙棘林最大的蛀干害虫,给我国的沙棘产业造成了巨大的损失.本论文从形态学和组织学2个方面确定了沙棘木蠹蛾雌、雄生殖系统的特征,同时,对比了雄性交尾前后生殖系统各部位的差异.未交尾雄蛾的生殖系统贮精囊、附腺管和射精管呈白色,其内包含大量的精子束待交尾,已交尾雄蛾的生殖系统呈透明或半透明状.除雄性附腺外,在交尾后的精巢、贮精囊和射精管中都能观察到真核精子束.通过上述对雄性生殖系统的形态学和组织学观察,尤其是对贮精囊和射精管中精子束观察,可判断精子束的填充状态,从而判断雄蛾交尾与否,能够检验性信息素引诱剂对沙棘木蠹蛾的防治效果,确立判断雄蛾交尾与否的标准对指导沙棘木蠹蛾防治具有重要意义.     

氨氮胁迫对日本蟳免疫生理指标及器官结构的影响

采用生物酶测定及组织学方法,研究了水体中不同浓度氨氮胁迫下日本蟳免疫相关指标的变化,以及对鳃、肝胰腺和胃等器官结构的影响。结果表明:在氨氮胁迫下,各实验组的血细胞密度(DHC)、血蓝蛋白含量、超氧化物歧化酶(SOD)和过氧化氢酶(CAT)活力,以及低浓度胁迫组的酚氧化酶(PO)和溶菌酶(LSZ)活力呈"先升后降"的变化趋势;高浓度组LSZ活力持续下降。在低浓度氨氮胁迫下,过氧化代谢产物丙二醛(MDA)含量呈"先升后降再升",而高浓度氨氮胁迫导致MDA含量持续上升。胁迫第15天,除低浓度组的DHC、血蓝蛋白含量和PO活力略高于对照组外,其他实验组所测指标均低于对照组;PO、LSZ、SOD和CAT等酶活力以及DHC均与氨氮胁迫浓度呈显著负相关,MDA含量则与胁迫浓度呈显著正相关(P0.01)。高浓度氨氮胁迫会导致鳃几丁质膜变薄、断裂,鳃上皮排列紊乱、染色质异固缩;鳃腔中血淋巴减少,密度降低,血细胞凝集、质膜破裂,胞质严重空泡化;肝胰腺上皮形态不规则,B细胞减少,腺细胞出现大量空泡,染色质凝聚;胃几丁质膜断裂,胃上皮排列不规则,胞质中出现大量残余体。研究表明高浓度氨氮胁迫对日本蟳免疫相关指标和器官结构产生显著影响,SOD活力和MDA含量长期变化情况可作为衡量日本蟳在氨氮胁迫下免疫状态的指标。     

Pituitary somatostatin receptors: dissociation at the pituitary level of receptor affinity and biological activity for selective somatostatin analogs

High affinity and saturable binding of [125I-Tyr11]somatostatin (SS) is described in membrane homogenates from a pituitary transplantable tumor (GH4C1) rich in somatotrophs (KD for SS = 0.67 nM; Bmax = 30 fmol/mg protein). Binding characteristics and pharmacology are similar to those measured on normal pituitary membranes. The potency of various SS analogs highly correlates with that measured in in vitro bioassay for growth hormone. This suggests that those GH4C1 membranes are a good model for SS receptors on somatotrophs. Interestingly however, analogs in which the Asn5 is deleted (Des-Asn5) or D-Ser replaces Ser13 show dissociated potencies between the various assays: [D-Ser13] analogs are more potent in pituitary than in GH4C1 membranes binding assay. Des-Asn5-modified analogs are much more potent in both pituitary binding assays than in the bioassay. This could reflect a multiplicity of SS receptor subtypes in pituitary.     

Effects of ploidy and sex-locus genotype on gene expression patterns in the fire ant Solenopsis invicta

Males in many animal species differ greatly from females in morphology, physiology and behaviour. Ants, bees and wasps have a haplodiploid mechanism of sex determination whereby unfertilized eggs become males while fertilized eggs become females. However, many species also have a low frequency of diploid males, which are thought to develop from diploid eggs when individuals are homozygous at one or more sex determination loci. Diploid males are morphologically similar to haploids, though often larger and typically sterile. To determine how ploidy level and sex-locus genotype affect gene expression during development, we compared expression patterns between diploid males, haploid males and females (queens) at three developmental timepoints in Solenopsis invicta. In pupae, gene expression profiles of diploid males were very different from those of haploid males but nearly identical to those of queens. An unexpected shift in expression patterns emerged soon after adult eclosion, with diploid male patterns diverging from those of queens to resemble those of haploid males, a pattern retained in older adults. The finding that ploidy level effects on early gene expression override sex effects (including genes implicated in sperm production and pheromone production/perception) may explain diploid male sterility and lack of worker discrimination against them during development.