2005 Vol. 26, No. 4
Molecular Cloning and Sequence Analyses of cDNAs Encoding Seven C-type Lectin-like Protein Subunits from Daboia russellii siamensis
2005, 26(4): 337-343.
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Total mRNA was extracted from a venom gland of snake Daboia russellii siamensis following the manufacturers protocol of the PolyATtract System 1000 kit purchased from Promega Biotech. cDNAs encoding C-type lectins were amplified by RT-PCR and subcloned into a pMD18-T vector. Fourteen positive clones were selected for nucleotide sequencing and seven cDNAs encoding various snake venom C-type lectin-like protein precursors, designated as DRS-L1, DRS-L2, DRS-L3, DRS-L4 DRS-L5, DRS-L6 and DRS-L7, were obtained. Amino acid sequences of these proteins were deduced and each contains a carbohydrate recognition domain. Of all the deduced protein sequences, only DRS-L1 seemed to represent a closer sequence similarity to α subunits of other known snake venom C-type lectin-like proteins using the BLAST program. Homology comparison combined with analysis of cysteine position indicate that DRS-L1 and DRS-L2 are probably the light chain LC2 and LC1 of factor Ⅹ activator from Daboia russellii siamensis venom, respectively. DRS-L3 and DRS-L4 might be the β subunits of higher molecular weight C-type lectin-like proteins while DRS-L5 and DRS-L6 might be β subunits of lower molecular weight C-type lectin-like proteins. DRS-L7 might be the β subunit of a platelet membrane glycoprotein Ib-binding protein similar to echicetin.
2005, 26(4): 344-349.
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The relationships within Drosophila melanogaster species group are controversial from morphology, chromosomes and DNA sequences. This study utilises a molecular approach aimed at uncovering the phylogenetic relationships among 33 taxa representing 8 subgroups of D. melanogaster species groups. Mitocondrial ND4L-ND4 was sequenced in the all 8 subgroups covering a wide geographic area. MP and Bayesian analysis produced an identical tree topology with relatively strong support in most nodes. It reveals that the melanogaster species group clustered in three main lineages: 1) montium subgroup; 2) ananassae subgroup; 3) Oriental subgroups (melanogaster, ficsphila, eugracilis, elegans, suzukii and takahashii). The montium subgroup branched off first, followed by the ananassae subgroup. In the third lineage, melanogaster is the most divergent subgroup followed by ficsphila, eugracilis, elegans in that order. The suzukii and takahashii sister subgroups are the last to branch off.
Seasonal Variation of Diurnal Activity Budgets by Blue Sheep (Pseudois nayaur) with Different Age-sex Classes in Helan Mountain
2005, 26(4): 350-357.
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The daytime activity budgets of blue sheep (Pseudois nayaur) in Suyukou National Park, Helan Mountain, were studied from November 2003 to October 2004. Blue sheeps activities were recorded by instantaneous and scan sampling methods. We observed feeding, lying, moving, standing and other behaviors from the 10th to 20th days every month with a SJ-1 Event Recorder. Data were collected from 8:00-17:00 in winter, from 7:00-18:00 in spring and autumn, and from 6:00-20:00 in summer. We used the non-parametric Wilcoxon test to compare differences in the time budgets between male and female sheep. Mann-Whitney U and Kruskal-Wallis H tests were used to test activity budgets among age classes. A Kruskal-Wallis H test was also used to make comparisons of time budgets among ecological seasons. Feeding was the principal activity of blue sheep and accounted for 63.49±7.82% of their activities. Lying was the second principal activity, accounting for 19.32±6.79% of their activities. The remaining time was devoted to the other three activities (standing, moving, and other behaviours). The percentage of daytime spent feeding reached its highest level in February, and reached its lowest level in July. Sheep decreased their time spent lying to its highest level in July. During three seasons (spring, summer and autumn), blue sheep showed a morning and evening peak of feeding. Feeding remained at relatively higher levels (over 60%) in winter. Sheep spent a relatively higher percentage of time in other activities in winter than any other seasons. Adult and subadult sheep forage more than did kids; lambs spent more time in lying than did adult and subadult sheep; adult and subadult sheep devoted more time to moving and standing than did kids; kids spent more time standing than did adults. In February, the percentage of daytime spent feeding by male and female sheep reached its highest level. Feeding time spent by males remained a relatively constant level (<70%) in other months. However, time spent feeding by females remained at high levels (>70%) from November in 2003 to February in 2004. There was not a significant difference in the percentage of daytime spent lying between males and females. Male sheep spent more time moving than did females. Like other temperate ungulates, quality and quantity of forage plants, growth and physiological phases all contributed to the daytime activity budgets of blue sheep.
2005, 26(4): 358-364.
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There are significant correlations between different taxonomic groups with respect to species richness. However, different studies yield different correlation coefficients between taxonomic groups. In this paper we analysed the impacts of some non-environmental potential factors, such as area effects, taxonomic groups, spatial scale, species distribution, avian residents and travelers on species richness covariance among taxonomic groups. Our results showed that significant covariance only occurred between the taxonomic groups with similar origins and ecologies. There were no area effects in our data set. Spatial scale affected covariance among taxonomic groups, although similar trends only occurred with similar scales. Species with different distribution patterns showed significantly different covariance. There were higher correlations in Oriental species richness than in Paleoartic species richness or between both species. The species richness of avian breeds had very significant covariance with mammals in terms of species richness rather than with other avian ecological types. Therefore, the ecological types and origins of taxonomic groups may have impacts on the covariance of species richness among these taxonomic groups.
Ecological Distributions and Community Compositions of the Soil Macro-animals in the Mid-west Plain of Jilin Province
2005, 26(4): 365-372.
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soil macro-animals in the mid-west plain of Jilin Province were investigated with the emphasis on species richness and abundance in relation to five types of land use I.e. farm garden, farmland, Three-North Forest Shelterbelt, natural secondary forest and natural mowing pasture. Soil macro-animals were collected by hand-sorting. A total of 5 842 soil macro-animal individuals were captured and classified into 3 phylums, 7 classes, 16 orders and 101 families. Lasius Tetramorium and Achaeta were the dominant groups that accounted for 49.20% of the total individuals. The results suggested that the types of land use affected the species richness and abundance, and human activity has a significant impact on the soil macro-animals community. Agricultural activity reduced soil macro-animal diversity, but did not change their vertical distributions in the soil profile. The results also showed that the differences in the composition of the soil macro-animal community between the middle plain and the west plain in Jilin Province were mainly explained by the dominant group density and rare group numbers in all habitats.
2005, 26(4): 373-378.
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During periods of October 2002 to June 2003 and November 2003 to June 2004, the study of the factors affecting dispersal of Elliots pheasants was carried out in Gutianshan National Nature Reserve, Zhejiang Province of China, using methods of radiotelemetry, GPS, SAW modelling and sampling. The results showed that both males and females dispersed in spring, and their dispersal distance varied from 1.5 to 2.1 km with a duration of 16 to 23 days. Movement routes of Elliots pheasant were selective when dispersal occurred in spring. The variables of herb coverage, number of tree species, number of shrub species, canopy height of shrubs, abundance of shrubs and abundance of herbs resulted in highly significant differences between used and unused patches (P<0.01). Five additional variables, including distance to water, slope, number of trees, canopy height of trees and number of herb species also resulted in significant differences between used and unused patches (P<0.05). The result of Conditional Logistic Regression analysis suggested that the major factors affecting dispersal of Elliots pheasant in the spring were number of shrub species, canopy height of shrubs, slope and abundance of shrubs.
2005, 26(4): 379-385.
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Ficus (Moraceae) and their species-specific pollinator wasps (Agaonidae) form a remarkable plant-insect obligate mutualism, and non-pollinators are the exploiters of the mutualism. The negative impact of exploiters on the reciprocal mutualists might disrupt the reciprocal mutualism in the process of evolution, but how the exploiters could coexist with the reciprocal mutualsits is not still clear. In this study, the diet of the five species of non-pollinators and relationship among fig wasps were analyzed on Ficus racemosa L. in Xishuangbanna from Dec. 2003 to Apr. 2004. In a controlled experiment pollinators and each species of non-pollinators oviposited and counts of wasps and seeds in mature figs were conducted. The results indicated that only Platyneura testacea Motschulsky and Platyneura mayri Rasplus are gall-makers, which can induce the ovaries into galls; Apocrypta sp., Apocrypta westwoodi Grandi and Platyneura agraensis Joseph are the parasitoids. The gall-makers and the parasitoids of pollinators have negative impacts on pollinators, but the impacts are not significant because of the influence of the ants and parasitoids of gall-makers. Additionally, the experiment excluding non-pollinators oviposition showed that the number of offspring of pollinators and seeds were not significantly different with the natural fruits. Moreover, the analysis on the natural population structure of fig wasps revealed that the pollinators are the dominant species. So in the natural condition, the abundance of gall-makers and parasitoids of pollinators are below the level needed to exclude pollinators, and thus they have a relatively weak impact on the stability of fig-pollinator mutualism and can coexist with the mutualism.
Impact of Oecophylla smaragina on the Percentage Number of Offspring of Pollinator and Nonpollinating Wasps on Ficus racemosa
2005, 26(4): 386-390.
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We performed field observations and ant exclusion experiments to determine whether Oecophylla smaragina tending homopterans on the fruit (fig) of Ficus racemosa Linn. Affects the reproduction of F. racemosa in Xishuangbanna from December 2003 to May 2004. The results of preliminary observations revealed that pollinator of F. racemosa oviposit inside the figs; five species of non-pollinating wasps attacked figs from the exterior; the number of non-pollinating wasps oviposition attempt can be quantified by counting the number of scars they made on the figs. The results of preliminary experiments showed the sticky glue barrier we used to exclude ants has no effect on oviposition of six species of wasps. When there were young figs on the sample trees, we selected two branches (a pair) which were similar in size, position and developmental phase and carried out experiments on eight randomly sampled trees. We excluded O. smaragina from one branch of each pair using sticky glue. We collected figs from trees just prior to the time that wasp progeny were due to emerge. We then counted the number of individuals of each wasp species per fig. The results showed that foraging workers of ants attacked non-pollinating wasps severely when which ovipositing on the figs. But they could not interfere with pollinators, which could enter the fig from the ostioles in a few seconds. The result of two-way ANOVA revealed that O. smaragina has significant impacts on the percentage (N%) of six wasp species breeding in the fig (n=82, F1,80>9, P<0.02). Ants present the N% of offspring of pollinator was 73.02. Ants excluded the N% of offspring of pollinator reduced to 2.14. Except for Platyneura agraensis, when ants were excluded the N% of offspring of other species of non-pollinating also increased. Ceratosolen fusciceps was sole pollinator of F. racemosa, while nonpollinating wasps competed with pollinator for galls or parasitized pollinator but play no role in pollination process. So O. smaragina can benefit F. racemosa indirectly by increasing the N% of pollinator while decreasing the number of non-pollinating wasps .
Effects of Intraperitoneal Injection of Polychlorinated Biphenyls During Pregnancy on Sexual Behavior of F1 Rats
2005, 26(4): 391-396.
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50 time-mated pregnant rats were divided into five groups and injected daily from gestational days 7 to 18 with either 2,2′,4,4′tetrachlorobiphenyl (PCB 47) at the dosage of 1.00 or 20.00 mg/kg body weight; or 3,3′,4,4′ tetrachlorobiphenyl (PCB 77) at the dosage of 0.25 or 1.00 mg/kg body weight; or sesame oil (control) to investigate the effects of fetal and lactational PCB exposure on reproductive behavior in male and female laboratory rats. Offspring were then tested for male sexual behavior: mount frequency, MF; mount latency, ML; intromission frequency, IF; intromission latency, IL; ejaculation latency, EL; post ejaculatory interval, PEI; hit rate and female sexual behavior; approach latency, AL; mount return latency, MRL; intromission return latency, IRL; post ejaculatory refractory period, PER; lordosis quotient, LQ. Measures were made at the age of postnatal days 70 to 91, and 97 to 101, respectively. The results showed that exposure to both PCB 77 and PCB 47 significantly reduced the LQ (84.4% for control, 76.0%, 67.8% for PCB 47 groups and 64.4%, 53.3% for PCB 77 groups; P<0.05) in the female offspring. There were no significant effects on AL, MR, IRL, PER (P>0.05) of the female offspring, or on MF, ML, IF, IL, EL, PEI and hit rate (P>0.05) of the male offspring.
Molecular Cloning and Comparison of Ubiquitin Fusion Protein and Ribosomal Protein L30 from Ophiophagus hannah
2005, 26(4): 397-403.
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Total RNA was extracted from the venom gland of snake Ophiophagus hannah. A king cobra venom gland cDNA library was then constructed using purified mRNA from the total RNA. By random sequencing of 200 independent clones, two conserved cDNA sequences of house-keeping genes were obtained. One is ubiquitin fusion protein (GenBank accession number AF297036); the other is ribosomal protein L30 (GenBank accession number AF297033). The former has an open reading frame of 387 bp, encoding a 128 amino acid ubiquitin fusion protein precursor composed of a 76 amino acid ubiquitin domain and followed by a 52 amino acid ribosomal protein L40 domain. The deduced precursor protein from the nucleotide sequence is basic. The C-terminus of the ubiquitin fusion protein contains the zinc-finger motif. The latter has an open reading frame of 348 bp, encoding for 115 amino acid ribosomal protein L30 precursor. The predicted amino acid sequence of ubiquitin fusion protein is highly conserved when compared to the sequences of homologous proteins from sixteen diverse species.
2005, 26(4): 404-409.
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A single chain protein with an apparent molecular weight of 33 kDa was purified from skin of Bombina maxima by a combination of ion exchange and gel filtration chromatography steps. N-terminal amino acid sequence determination indicated that it shares 70%, 64% and 56% identity with those of annexin Ⅱ from the African claw toad, red jungle fowl and human, respectively. The purified protein from B. maxina inhibits stejnulxin (a specific platelet agonist via platelet membrane glycoprotein Ⅵ receptor) induced platelet aggregation in a Ca2+ dependent manner. Maximal inhibition rate reaches 48%. Based on the N-terminal amino acid sequence BLAST search results and the fact that its activity is strictly Ca2+ dependent, the purified protein might be structurally and functionally related to the annexin protein family.
2005, 26(4): 410-415.
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Two antimicrobial peptides were purified from Penus chinensis by Sephadex G-50 gel filtration and reverse phase high performance liquid chromatography. The molecular weights were 1 071 and 1 311 Da respectively, determined by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. They inhibited the growth of Gram-positive and Gram-negative bacteria. The antimicrobial peptides strengthened the contractile response of isolated mice ileum. Serine proteases did not affect hydrolytic activities in our current experiments. These antimicrobial peptides play an important role in preventing microbial infection in P. chinensis.
Experimental Research on the Construction of Cloned Embryos Through Human-rabbit Inter-species Nuclear Transfer
2005, 26(4): 416-421.
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Therapeutic cloning is one of our major research objectives, and therapeutic cloning is based on human somatic cell nuclear transfer. Though inter-species nuclear transfer has been introduced to construct human somatic cell cloned embryos, the effects of type, passage and preparation method of donor cells on embryo development remain to be determined. In our experiment, cloned embryos were reconstructed with different passage and preparation methods of ossocartilaginous cell, skin fibroblast and cumulus cells. The cumulus cell embryos show significantly higher development rates than the other two (P<0.05). The development rate of embryos reconstructed with skin fibroblasts of different passage number and somatic cells of different chilling durations show no significant difference. Also, FISH were conducted to detect nuclear derivation of the embryos. The result showed that the nuclei of the inter-species cloned embryo cells came from human. We conclude (1) that cloned embryos can be constructed through human-rabbit inter-species nuclear transfer; (2) different kinds of somatic cell result in different efficiency of nuclear transfer, while in vitro passage of the donor does not influence embryo development; (3) refrigeration is a convenient and efficient donor cell preparation method. Finally, it is feasible to detect DNA genotype through FISH.
Seventy-five female red-spotted grouper (Epinephelus akaara) of 2-3 years old were divided into three groups [control group ?, high (H) and low dose group (L), 25 fish per group] . Each group was treated as follows: high dose group [300 μg luteinizing-hormone releasing-hormone analogue (LHRH-A)/kg body weight], low dose group (100 μg LHRHA/kg body weight) and control (no LHRH-A). Blood samples were collected on day 0, 10, 20, 30 and 40 after implantation and serum estradiol-17β (E2) and testosterone (T) were determined by radioimmunoassay (RIA). The fish were killed and the gonads were removed and weighed for calculation of the gonadosomatic index (GSI). The microstructure and ultrastructure of ovary, liver and pituitary were examined by both light microscope and transmission electron microscope. The results showed that the ovulation rate in the LHRH-A implantation group was significantly higher than that in the control group; that the ovulation rate of the high dose group was significantly higher than that in low dose group. The GSI was 1.055% and migrated nucleus was observed on day 10 in high dose group, ovary developed slowly in low dose group. The GSI reached a high of 1.858% and a low of 0.987% at day 20 in the mature ovary in the LHRH-A implanted group. Serum T and E2 increased significantly after the LHRH-A implantation at day 10 and declined significantly at day 20. Serum T was significantly lower than the control group. Hypertrophied GTH cells, distinct degranulation and marked dilatation of cisternase of the rough endoplasmic reticulum suggest the release of hormone from the GTH cells. These results suggested that sustained administration of LHRH-A significantly induced ovarian maturation and ovulation in the red-spotted grouper. Sex steroid hormone is correlated with vitellogenesis, not ovulation.
2005, 26(4): 429-434.
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The structural features of normal and abnormal embryo attachment systems of the Chinese mitten crab Eriocheir sinensis were investigated by scanning electron microscopy. The results showed that the volume of incubation fluid in the brooding chamber increased 15 minutes after egg-laying, immersing the embryos in the incubation fluid and thus the egg envelop became very sticky immediately. The egg stalk was largely formed within 30 minutes and completed within 80 to 140 minutes after egg-laying, so that the egg envelop became strongly adherent to the ovigerous hairs. In normal embryos, the egg stalks were highly twisted and the embryos were attached to the ovigerous hair by the egg stalk or funiculus. Many villiform were found on egg stalks and a lot of mucus was also found on the ovigerous seta. In abnormal egg stalks, villiform were never found, though numerous epibionts were attached to the embryo surface and egg stalk. The mucus of ovigerous hair and the villiform of egg stalks may contribute to egg attachment. Because of the epibionts the structure of egg investment coat egg stalk and funiculus were changed, enhancing the attrition between embryos and making embryo respiration difficult. The death and loss of embryos occur because of the above reasons.
Age-specific Changes in Estrogen Receptors α in Testis and Epididymis of Mandarin Voles (Microtus mandarinus)
2005, 26(4): 435-441.
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In order to investigate the age-specific effect of estrogen receptor α (Erα) in testis and epididymis of mandarin voles (Microtus mandarinus), the expression of Erα from five age groups [1 (neonatal), 10, 25, 45, and 60 (adult) days of age] was observed using immunohistochemistry. There were several kinds of Erα-immunoreactive cells in the testes and epididymis of normal voles, such as Leydig cells, spermatogenic cells, as well as myoid cells and epithelial cells of the epididymis. The distributions and staining intensity of Erα-immunoreactive cells changed greatly with age. Statistics showed that: the expression of Erα in pros-permatogonium cells and gonocyte was weak in the neonatal vole. The expression of Erα in spermatids appeared at postnatal day 25, as well as day 45 of age. The expression of Erα was detected in every developmental stage of germ cell in the adult vole. The expression of Erα in Leydig cells appeared in the neonatal vole followed by a decrease in intensity at postnatal day but increasing at postnatal day 25. The expression intensities of Erα in Leydig cells peaked at day 45 but decreased in the adult. The expression of Erα in epithelial cells was found in the neonatal vole and was detected in myoid cells at postnatal days 10 and 25. In the epididymes of 45 day old and adult voles, myoid cells and epithelial cells expressed Erα positively. Those results imply a molecular mechanism of Erα action. Specifically, Erα may affect the secretion of androgen in Leydig cells as special receptors during postnatal development and spermatogenesis of mandarin voles.
2005, 26(4): 442-445.
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Previous studies have shown that the ventral part of prefrontal cortex (vPFC) plays an important role in drug addiction. The increased interest in γ oscillations, now widely regarded as functionally relevant signals of the brain, underlines the importance of the concept of event-related oscillations for bridging the gap between single neurons and neural assemblies. In the present study, we examined the dynamics of γ oscillation activity (or γ-band power) associated with the development of morphine dependence in the vPFC by using electrophysiological techniques after injecting morphine (0, 0.5, 1.6, 5.0 and 8.0 mg/kg) in rhesus monkeys. The results show that the γ-band power had a long-term decrease associated with the development of morphine dependence, and the decrease in γ power had a negative relationship with the concentration of morphine. Those results suggest strongly that γ oscillation activities in the vPFC reflect morphine addiction, and indicate that the EEG γ oscillation may reflect the processes underlying morphine addiction.
Differentiation of Subspecies of Tamiops mcclellandi (Horsfield, 1840) (Sciuridae，Rodentia) in Southwestern China
2005, 26(4): 446-452.
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Tamiops mcclellandi is a species endemic to southeastern Asia. We obtained data on 15 cranial characteristics from three currently recognized subspecies of T. mcclellandi that used to be classified by pelage characteristics in China (T. m. mcclellandi,Tmm;T. m. collinus,Tmc;T. m. inconstans,Tmi) and which are distributed in southeastern Tibet, and in the south and southwest of Yunnan). We analyzed the data with SPSS 11.0 (Multivariate Analysis) in order to examine whether significant differences exist in cranial characteristics among three subspecies, to evaluate their subspecies validities, and to examine relationships among them. The results indicate: 1. The three groups are all valid subspecies. 2. Among 15 cranial characteristics, eight characteristics (ONL, ROB, ZOB, IOB, MTB, BN, MDL, HM) between Tmm and Tmc were significantly different; seven characteristics (GLS,ONL, ZOB, IOB, MTB, MH, HM) between Tmm and Tmi were significantly different; and four characteristics (ROB, MTB, BN, MH) between Tmc and Tmi were significantly different. 3. Relationships between Tmc and Tmi are closer than those between each of the two subspecies and Tmm.