Deep-level diagnostic value of the rDNA-ITS region

The similarity of certain reported angiosperm rDNA internal transcribed spacer (ITS) region sequences to those of green algae prompted our analysis of the deep-level phylogenetic signal in the highly conserved but short 5.8S and hypervariable ITS2 sequences. We found that 5.8S sequences yield phylogenetic trees similar to but less well supported than those generated by a ca. 10-fold longer alignment from rDNA-18S sequences, as well as independent evidence. We attribute this result to our finding that, compared to 18S, the 5.8S has a higher proportion of sites subject to vary and greater among-site substitution rate homogeneity. We also determined that our phylogenetic results are not likely affected by intramolecular compensatory mutation to maintain RNA secondary structure nor by evident systematic biases in base composition. Despite historical homology, there appears to be no ITS2 primary sequence similarity shared sufficient similarity to cluster correctly on the basis of alignability. Our results indicate that groups, however, share sufficient similarity to cluster correctly on the basis of alignability. Our results indicate that ITS region sequences can diagnose organismal origins and phylogenetic relationships at many phylogenetic levels and provide a useful paradigm for molecular evolutionary study.      

拉萨河谷上段江孜沙棘Hippophea gyantsensis(Rousi) Lian种群的海拔梯度性变异

江孜沙棘[Hippophea gyantsensis(Rousi)Lian]是青藏高原特有的一种广生态幅的小乔木,在拉萨河谷地区的海拔3500~4200m范围内均有分布。前人工作多集中在江孜沙棘果实的开发利用方面,对其基础生态学研究较少。本研究旨在探讨江孜沙棘沿海拔梯度的群落组成和表型变异的规律。为此,在拉萨河谷上段沿海拔梯度由东向西设置了4个样带:3850m、3950m、4050m和4200m,每个样带设置2至3个10m×10m的样方进行研究。首先,详细记录了每个样方内林下维管植物的物种组成、样方内的沙棘盖度、海拔、样方与河岸的实际距离,并用DCA[detrended correspondence analysis(去势对应分析)]排序方法对群落及其组成物种进行排序分析。随机抽取了每个样方内的20个江孜沙棘植株个体,测定其胸径、基径、株高和叶片长度,用回归分析法分析这些变量和海拔之间的关系。研究结果表明,江孜沙棘在拉萨河谷内的主要生境分为4种类型,即:河边砾石滩地、河阶草滩、河边草甸和河边林缘,样方排序结果主要受海拔的影响;同时,江孜沙棘植株的基径、胸径和高度都随着海拔的升高而显著减小,而叶片长度与海拔之间无显著相关。本文研究结果表明,对江孜沙棘而言,海拔所代表的综合环境因子对其分布和表型有显著的影响,而局部光照可能也是影响其表型特征的重要生态因子。     

Production of Serine Proteases by the Oyster Pathogen Perkinsus marinus (Apicomplexa) In Vitro

ABSTRACT. Analysis of the cell-free supernatants of Perkinsus marinus cultures by sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and silver staining revealed the presence of as many as 17 bands ranging in molecular weight from 239 to 32 kDa. These bands were not present in un-inoculated medium. Moreover, P. marinus produces extracellular proteins that possess proteolytic activities; the cell-free supernatants of P. marinus cultures could digest a variety of proteins including gelatin, casein, fibronectin and laminin. Oyster plasma was also digested by cell-free culture supernatants. The proteolytic activity in cell-free culture supernatants was detected 24 h post-inoculation, while no proteolytic activity could be detected in cell lysates. The proteolytic activities were characterized using substrate-impregnated sodium dodecylsulfate-polyacrylamide gels and had approximate molecular weights ranging from 55 to 35 kDa. The proteolytic activity of cell-free culture supernatants was inhibited by the serine protease inhibitors phenylmethylsulphonyl fluoride, 3,4-dichloroisocoumarin and soybean trypsin inhibitor. In contrast, inhibitors (i.e. trans-epoxysuccinyll-leucylamido(4-guanidino)-butane, 1, 10-phenanthroline, captopril, ethylenediaminetetracetic acid, pepstatin A or diazoacetyl-DL-norleucine methyl ester) from the other three classes of proteases had no effect. It was concluded that the P. marinus proteases in cell-free culture supernatants are serine proteases.     

Improved in vitro embryo development using in vivo matured oocytes from heifers superovulated with a controlled preovulatory LH surge

In bovine in vitro embryo production, the IVM step is rather successful with 80% of the oocytes reaching the MII stage. However, the extent to which the process limits the yield of viable embryos is still largely unknown. Therefore, we compared embryonic developmental capacity during IVC of IVF oocytes which had been matured in vitro with those matured in vivo. In vitro maturation was carried out for 22 h using oocytes (n = 417) obtained from 2- to 8-mm follicles of ovaries collected from a slaughterhouse in M199 with 10% fetal calf serum (FCS), 0.01 IU/mL LH, and 0.01 IU/mL FSH. In vivo matured oocytes (n = 219) were aspirated from preovulatory follicles in eCG/PG/anti-eCG-superovulated heifers 22 h after a fixed time GnRH-induced LH surge; endogenous release of the LH surge was suppressed by a Norgestomet ear implant. This system allowed for the synchronization of the in vitro and in vivo maturation processes and thus for simultaneous IVF of both groups of oocytes. The in vitro developmental potential of in vivo matured oocytes was twice as high (P < 0.01) as that of in vitro matured oocytes, with blastocyst formation and hatching rates 11 d after IVC of 49.3 +/- 6.1 (SEM; n = 10 heifers) vs 26.4 +/- 1.0% (n = 2 replicates), and 39.1 +/- 5.1% vs 20.6 +/- 1.4%, respectively. It is concluded that IVM is a major factor limiting in the in vitro production of viable embryos, although factors such as the lack of normal preovulatory development of IVM oocytes contributed to the observed differences.     

Effects of aglépristone, a progesterone receptor antagonist, administered during the early luteal phase in non-pregnant bitches

Aglépristone, a progesterone receptor antagonist, was administered to six non-pregnant bitches in the early luteal phase in order to determine its effects on the duration of the luteal phase, the interestrous interval, and plasma concentrations of progesterone and prolactin. Aglépristone was administered subcutaneously once daily on two consecutive days in a dose of 10 mg/kg body weight, beginning 12 +/- 1 days after ovulation. Blood samples were collected before, during, and after administration of aglépristone for determination of plasma progesterone and prolactin concentrations. The differences in mean plasma concentration of progesterone and of prolactin before, during, and after treatment were not significant. Also, the duration of the luteal phase in the six treated bitches (72 +/- 6 days) did not differ significantly from that in untreated control dogs (74 +/- 4 days ). However, the intervals during which plasma progesterone concentration exceeded 64 and 32 nmol/l were significantly shorter in the six treated bitches than in untreated control dogs. The interestrous interval was significantly shorter in beagle bitches treated with aglépristone (158 +/- 16 days) than in the same group prior to treatment (200 +/- 5 days ). It is concluded that administration of aglépristone during the early luteal phase in the non-pregnant bitch affects progesterone secretion, but not sufficiently to shorten the luteal phase. The shortening of the interestrous interval suggests that aglépristone administered in the early luteal phase influences the hypothalamic-pituitary-ovarian axis.     

Fetometry and fetal heart rates between Day 35 and 108 in bovine pregnancies resulting from transfer of either MOET, IVP-co-culture or IVP-SOF embryos

The Large Offspring Syndrome has frequently been reported for in vitro produced calves. The objective of this study was to determine whether any differences in body dimensions (biparietal diameter of the cranium (BPD), cross-section of the abdomen at the insertion of the umbilical cord (CAU)) and heart rate (FHR) can be detected during the first 108 days of gestation between bovine foetuses derived from different methods of embryo production. Three groups of pregnancies with calvings at term resulted from non-surgical transfers of three types of embryos: recipients carrying an embryo obtained by standard MOET procedures (n = 25); recipients carrying an embryo produced in vitro from OPU-derived oocytes, using co-culture-medium (n = 14) or SOF-medium (n = 22). Transrectal ultrasonographic examinations were performed weekly. Ultrasound images were recorded and during off-line analysis FHR, BPD and CAU were determined. For each foetus a curve was fitted and the estimates on fixed time intervals were used as dependent variables in an analysis of variance to detect differences between the three pregnancy groups. Neither gestation length nor birth weight differed significantly between the three pregnancy groups, nor could any differences with respect to BPD, CAU or FHR be detected between Days 35 and 108 of gestation. It is concluded that no differences exist between the early development of bovine foetuses, derived from MOET, IVP-co-culture or IVP-SOF embryos, and resulting in calves with normal birth weights.