Systematics within Gyps vultures: a clade at risk

Background  

Populations of the Oriental White-backed Vulture (Gyps bengalensis) have declined by over 95% within the past decade. This decline is largely due to incidental consumption of the non-steroidal anti-inflammatory veterinary pharmaceutical diclofenac, commonly used to treat domestic livestock. The conservation status of other Gyps vultures in southern Asia is also of immediate concern, given the lack of knowledge regarding status of their populations and the continuing existence of taxonomic uncertainties. In this study, we assess phylogenetic relationships for all recognized species and the majority of subspecies within the genus Gyps. The continuing veterinary use of diclofenac is an unknown but potential risk to related species with similar feeding habits to Gyps bengalensis. Therefore, an accurate assessment of the phylogenetic relationships among Gyps vultures should aid in their conservation by clarifying taxonomic uncertainties, and enabling inference of their respective relatedness to susceptible G. bengalensis.     

Isolation, partial chemical and biological characterization of thymone B

A new approach to the study of the molecular arrangements of proteins in membranes is described. Irradiation with visible light of native erythrocytes or washed erythrocyte membranes suspended in buffers containing a) riboflavin, fluorescein or fluorescein coupled to dextran and b) ³H-labelled tryptophan resulted in incorporation of radioactivity into the membrane proteins. Polyacrylamide gel electrophoresis of solubilized membranes followed by radioactivity measurements of the separated membrane proteins revealed that in native erythrocytes the protein components known to be located at the exterior cell surface, Band 3 and the major sialoglycoproteins became specifically labelled, whereas in washed lysed cells all of the major membrane proteins were labelled.     

Postpartum reproductive performance in the multiparous mare fed to obesity

Twenty multiparous Quarter Horse mares were assigned to one of two treatment groups at 40 to 75 d of pregnancy. Group 1 was the control group and the mares were fed to maintain a moderate degree of body fat (condition score 5.5 to 7). Group 2 was the obese group and the mares were fed to achieve (prepartum) and then maintain (post partum) an extremely high degree of body fat (condition score 9). Estrous intensity was evaluated using subjective teasing scores ranging from 0 (rejection) to 4 (maximum receptivity). Mares were artificially inseminated beginning with the second postpartum ovulatory cycle; the study was terminated after 63 d of pregnancy. Duration of estrus, maximum teasing score and the number of mares exhibiting overt estrus (teasing score > 2) did not differ between treatment groups during the first and second postpartum ovulatory cycles. The intervals from foaling to first cycle ovulation, foaling to second cycle ovulation, and first to second cycle ovulation were also similar between treatment groups. All mares in both treatment groups conceived and maintained pregnancy. The first cycle conception rate and the number of cycles per conception did not differ between treatment groups. A high degree of body fat produced by overfeeding during gestation did not adversely affect postpartum reproductive performance in the multiparous mare.     

Patterns of divergence during evolution of alpha 1-proteinase inhibitors in mammals

alpha 1-Proteinase inhibitor (alpha 1-PI), a member of the serine proteinase inhibitor superfamily, has a primary role in controlling neutrophil elastase activity within the mammalian circulation. Several studies have indicated that the reactive center region of alpha 1-PI, the amino acid sequence of which is critical to recognition of and binding to target proteinases, is highly divergent within and among species. This appears to be a consequence of accelerated rates of evolution that may have been driven by positive Darwinian selection. In order to examine this and other features of alpha 1-PI evolution in more detail, we have isolated and sequenced cDNAs representing alpha 1- PI mRNAs of the mouse species Mus saxicola and Mus minutoides and have compared these with a number of other mammalian alpha 1-PI mRNAs. Relative to other mammalian mRNAs, the extent of nonsynonymous substitution is generally high throughout the alpha 1-PI mRNA molecule, indicating greater overall rates of amino acid substitution. Within and among mouse species, the 5'-half of the mRNA, but not the 3'-half, has been homogenized by concerted evolution. Finally, the reactive center is under diversifying or positive Darwinian selection in murid rodents (rats, mice) and guinea pigs yet is under purifying selection in primates and artiodactyls. The significance of these findings to alpha 1-PI function and the possible selective forces driving evolution of serpins in general are discussed.      

Continuous,high capacity reconstitution of nutrient media from concentrated intermediates

We designed an Integrated Media Preparation System (IMPS) for continuous, on-line preparation of cell culture media and delivery to intermediate storage vessels or directly to a bioreactor. Key components of the IMPS include: a high precision, continuous fluid mixing device; formulation-specific liquid medium concentrates; validated process controls and membrane filtration; and automated dispensing into large volume flexible plastic containers. The IMPS system is designed to produce sterile, single-strength liquid medium from common raw materials at a delivery rate of 1000–3000 liters per hour and will manufacture homogenous batches from several thousand liters to over 60,000 liters. Fortified nutrient media prepared from multi-component 50X concentrates have been demonstrated to accelerate bioreactor seed chains, increase product yield, and reduce the overall manufacturing cost of nutrient medium. A productivity matrix will analyze the fully-loaded costs and contrast alternative methods for media preparation against projected biological yield.Abbreviations IMPS Integrated Media Preparation System - 50X Nutrient fluid components formulated at fifty-fold final use concentration - 1X Nutrient fluid formulated at final, single-strength use concentration - cGMP Current Good Manufacturing Practices - SCADA Supervisory Control and Data Acquisition - PLC Process Logic Controller - LTI Life Technologies, Inc. - WFI Water for Injection - CIP Clean in place - SIP Sterilize in place - HPLC High performance liquid chromatography - DMEM Dulbecco's Modified Eagle's Medium     

Mouse oocyte maturation is affected by lithium via the polyphosphoinositide metabolism and the microtubule network

The incubation of mechanically denuded mouse oocytes in medium containing LiCl delayed both germinal vesicle breakdown (GVBD) and polar body extrusion in a dose-dependent and reversible manner. When myo-inositol alone was added to the culture medium, we observed that it accelerated GVBD and increased the rate of polar body extrusion, whereas, when combined with LiCl, the normal timing of GVBD was recovered. In the same way, when inositol trisphosphate (InsP₃) was microinjected into the ooplasma, we observed an important improvement of the rate of GVBD, as compared to control oocytes, and prevention of lithium inhibition. However, neither myo-inositol nor InsP₃ were able to rescue totally the oocytes from the negative effect of lithium on polar body extrusion. Moreover, lithium induced some important changes in microtubule and chromosome organizations. Before extrusion of the first polar body, the reduction of the spindle size or the appearance of short individualized chromosomes dispersed around a large aster of microtubules were often observed, whereas, after polar body extrusion, the spindle appeared smaller and chromosomes were often trapped in the midbody. Thus lithium affects mouse oocyte maturation at two different levels: GVBD and polar body extrusion. Whereas the former seems to be affected via polyphosphoinositide turnover, the latter is InsP₃-independent and seems to be influenced negatively via underdevelopment of microtubular structures. © 1994 Wiley-Liss, Inc.     

The expression of nuclear lamin A and C epitopes is regulated by the developmental stage of the cytoplasm in mouse oocytes or embryos.

The cytoplasmic regulation of changes of nuclear lamin antigens was examined by transferring 16-cell stage blastomeres into mouse oocytes. Sixteen-cell stage blastomeres were transferred to either pronuclear eggs, enucleated pronuclear eggs or metaphase II oocytes, which were subsequently activated. Pronuclei react with a monoclonal antibody to A/C lamins (J9), whereas nuclei from 16-cell stage blastomeres do not react with J9. However, after transfer of 16-cell stage nuclei to activated metaphase II oocytes, the transferred nuclei acquire the antigen. This is in contrast to 16-cell nuclei that were transferred to intact or enucleated pronuclear eggs; i.e., the nuclei only faintly acquired the A/C epitope. These results suggest that the developmental stage of the cytoplasm regulates the exposure of nuclear lamina epitopes, perhaps by limiting the supply of lamin A/C in the oocyte or because nuclear lamina assembly can only occur at the telophase transition. Furthermore, it appears that there is some exchange of the A/C epitope between (pro)nuclei within the same cell but that the majority of the A/C lamin epitope can be removed from a cell with (pro)nuclear removal.     

Ecological specialization and niche overlap of subterranean rodents inferred from DNA metabarcoding diet analysis

Knowledge of how animal species use food resources available in the environment can increase our understanding of many ecological processes. However, obtaining this information using traditional methods is difficult for species feeding on a large variety of food items in highly diverse environments. We amplified the DNA of plants for 306 scat and 40 soil samples, and applied an environmental DNA metabarcoding approach to investigate food preferences, degree of diet specialization and diet overlap of seven herbivore rodent species of the genus Ctenomys distributed in southern and midwestern Brazil. The metabarcoding approach revealed that these species consume more than 60% of the plant families recovered in soil samples, indicating generalist feeding habits of ctenomyids. The family Poaceae was the most common food resource retrieved in scats of all species as well in soil samples. Niche overlap analysis indicated high overlap in the plant families and molecular operational taxonomic units consumed, mainly among the southern species. Interspecific differences in diet composition were influenced, among other factors, by the availability of resources in the environment. In addition, our results provide support for the hypothesis that the allopatric distributions of ctenomyids allow them to exploit the same range of resources when available, possibly because of the absence of interspecific competition.