An ultrastructural overview of tubificid spermatozoa

The spermatozoa of seven species of tubificid oligochaete were compared to those already described in order to supply spermatozoal characters for systematic work on Tubificidae. The spermatozoa of Clitellio arenarius (subfamily Tubificinae) resemble those of the two other known members of the subfamily (Tubifex tubifex and Tubificoides amplivasatus) in being of two different types and in showing the same morphology of acrosomes and nuclei. Among Phallodrilinae, the three gutless species Inanidrilus bulbosus, Inanidrilus leukodermatus and Olavius planus share with Bathydrilus formosus the shape of the nucleus and various characters of the acrosome, whereas Thalassodrilus prostatus is particular in having the acrosome vesicle external to the tube and the longest middle piece recorded in oligochaetes. Monopylephorus limosus and Rhizodrilus russus (Rhyacodrilinae) differ widely in acrosomal and nuclear characters: M. limosus has a twisted nucleus, whereas R. russus has an apically flanged nucleus with a conspicuous apical concavity and an endonuclear canal. The data published on spermatozoa of the Limnodriloidinae are reviewed.Department of Zoology University of Göteborg     

Evidence that biosynthesis of phosphatidylethanolamine, phosphatidylcholine, and triacylglycerol occurs on the cytoplasmic side of microsomal vesicles

Experiments were performed to localize the hepatic microsomal enzymes of phosphatidylcholine, phosphatidylethanolamine, and triacylglycerol biosynthesis to the cytoplasmic or lumenal surface of microsomal vesicles. Greater than 90 percent of the activities of fatty acid-CoA ligase (EC 6.2.1.3), sn-glycerol 3-phosphate acyltransferase (EC 2.3.1.15), lysophosphatidic acid acyltransferase, diacylglycerol acyltransferase (EC 2.3.1.20), diacylglycerol cholinephosphotransferase (EC 2.7.8.2), and diacylglycerol ethanolaminephosphotransferase (EC 2.7.8.1) was inactivated by proteolysis of intact microsomal vesicles. The phosphatidic acid phosphatase (EC 3.1.3.4) was not inactivated by any of the protease tested. Under conditions employed, <5 percent of the luminal mannose-6-phosphatase (EC 3.1.3.9) activity was lost. After microsomal integrity was disrupted with detergents, protease treatment resulted in a loss of >74 percent of the mannose-6-phosphatase activity. The latency of the mannose-6-phosphatase activity was not affected by protease treatment. Mannose-6-phosphatase latency was not decreased by the presence of the assay components of several of the lipid biosynthetic activities, indicating that those components did not disrupt the microsomal vesicles. None of the lipid biosynthetic activities appeared latent. The presence of a protease-sensitive component of these biosynthetic activities on the cytoplasmic surface of microsomal vesicles, and the absence of latency for any of these biosynthetic activities suggest that the biosynthesis of phosphatidylcholine, phosphatidylethanolamine, and triacylglycerol occurs asymmetrically on the cytoplasmic surface of the endoplasmic reticulum. The location of biosynthetic activities within the transverse plane of the endoplasmic reticulum is of particular interest for enzymes whose products may be either secreted or retained within the cell. Phosphatidylcholine, phosphatidylethanolamine, and triacylglycerol account for the vast majority of hepatic glycerolipid biosynthesis. The phospholipids are utilized for hepatic membrane biogenesis and for the formation of lipoproteins, and the triacylglycerols are incorporated into lipoproteins or accumulate within the hepatocyte in certain disease states (14). The enzymes responsible for the biosynthesis of these glycerolipids (Scheme I) from fatty acids and glycerol-3P have all been localized to the microsomal subcellular fraction (12, 16, 29, 30). Microsomes are derived from the endoplasmic reticulum and are sealed vesicles which maintain proper sidedness. (11, 22). The external surface of these vesicles corresponds to the cytoplasmic surface of the endoplasmic reticulum. Macromolecules destined for secretion must pass into the lumen of the endoplasmic reticulum (5, 23). Uncharged molecules of up to approximately 600 daltons are able to enter the lumen of rat liver microsomes, but macromolecules and charged molecules of low molecular weight do not cross the vesicle membrane (10, 11). Because proteases neither cross the microsomal membrane nor destroy the permeability barrier of the microsomal vesicles, only the enzymes and proteins located on the cytoplasmic surface of microsomal vesicles are susceptible to proteolysis unless membrane integrity is disrupted (10, 11). By use of this approach, several enzymes and proteins have been localized in the transverse plane of microsomal membranes (11). With the possible exception of cytochrome P 450, all of the enzymes and proteins investigated were localized asymmetrically by the proteolysis technique (11). By studies of this type, as well as by product localization, glucose-6-phosphate (EC 3.1.3.9) has been localized to the luminal surface of microsomal vesicles (11) and of the endoplasmic reticulum (18, 19). All microsomal vesicles contain glucose-6-phosphatase (18, 19) which can effectively utilize mannose-6-P as a substrate, provided the permeability barrier of the vesicles has been disrupted to allow the substrate access to the active site located on the lumenal surface (4). An exact correspondence between mannose- 6-phosphate activity and membrane permeability to EDTA has been established (4). The latency of mannose-6-phosphatase activity provides a quantitative index of microsomal integrity (4.) Few of the microsomal enzymes in the synthesis of phosphatidylcholine, phosphatidylethanolamine, and triacylglycerol have been solubilized and/or purified, and little is known about the topography of these enzymes in the transverse or lateral planes of the endoplasmic reticulum. An asymmetric location of these biosynthetic enzymes on the cytoplasmic or lumenal surface of microsomal vesicles may provide a mechanism for regulation of the glycerolipids to be retained or secreted by the cell, and for the biogenesis of asymmetric phospholipid bilayers. In this paper, we report investigations on the localization of all seven microsomal enzymes (Scheme I) in the biosynthesis of triacylglycerol, phosphatidylcholine, and phosphatidylethanolamine, using the protease technique with mannose-6-phosphatase serving as luminal control activity. The latency of these lipid biosynthetic enzymes was also investigated, using the latency of mannose-6-phosphatase as an index of microsomal integrity.     

Wolf predation and snow cover as mortality factors in the ungulate community of the Bialowieża National Park,Poland

Summary Wolf-ungulate interactions were studied in the pristine deciduous and mixed forests of the Bialowiea National Park in 1985–1989. The study period included two severe and two mild winters. The community of ungulates inhabiting Bialowiea National Park consisted of red deer Cervus elaphus, 55% of all ungulates; wild boar Sus scrofa, 42%; and roe deer Capreolus capreolus, moose Alces alces, and European bison Bison bonasus, about 1% each. The average size of red deer groups increased from 2.7 (SD 2.35) in spring and summer to 6.9 (SD 6.84) in autumn and winter. In winter the group size of red deer was positively correlated with the depth of snow cover and negatively correlated with the mean daily temperature. Average group size of wild boar did not change significantly between seasons; it was 6.8 (SD 5.16) in spring and summer and 5.7 (SD 4.67) in autumn and winter. Analysis of 144 wolf scats showed that wolves preyed selectively on red deer. In October–April, Cervidae (mostly red deer) constituted 91% of biomass consumed by wolves, while wild boar made up only 8%. In May–September deer formed 77% of prey biomass, and the share of wild boar increased to 22%. In all seasons of the year wolves selected juveniles from deer and boar populations: 61% of red deer and 94% of wild boar of determined age recovered from wolves' scats were young <1 year old. Analysis of 117 carcasses of ungulates found in Bialowiea National Park showed that predation was the predominant mortality factor for red deer (40 killed, 10 dead from causes other than predation) and roe deer (4 killed, none dead). Wild boar suffered most from severe winter conditions (8 killed, 56 dead). The percentage of ungulates that had died from undernutrition and starvation in the total mortality was proportional to the severity of winter.     

Adding Content to Contacts: Measurement of High Quality Contacts for Maternal and Newborn Health in Ethiopia,North East Nigeria,and Uttar Pradesh,India

BackgroundFamilies in high mortality settings need regular contact with high quality services, but existing population-based measurements of contacts do not reflect quality. To address this, in 2012, we designed linked household and frontline worker surveys for Gombe State, Nigeria, Ethiopia, and Uttar Pradesh, India. Using reported frequency and content of contacts, we present a method for estimating the population level coverage of high quality contacts.ConclusionsMeasuring content of care to reflect the quality of contacts can reveal missed opportunities to deliver best possible health care.     

Electron crystallography reveals the structure of metarhodopsin I

Rhodopsin is the prototypical G protein-coupled receptor, responsible for detection of dim light in vision. Upon absorption of a photon, rhodopsin undergoes structural changes, characterised by distinct photointermediates. Currently, only the ground-state structure has been described. We have determined a density map of a photostationary state highly enriched in metarhodopsin I, to a resolution of 5.5 A in the membrane plane, by electron crystallography. The map shows density for helix 8, the cytoplasmic loops, the extracellular plug, all tryptophan residues, an ordered cholesterol molecule and the beta-ionone ring. Comparison of this map with X-ray structures of the ground state reveals that metarhodopsin I formation does not involve large rigid-body movements of helices, but there is a rearrangement close to the bend of helix 6, at the level of the retinal chromophore. There is no gradual build-up of the large conformational change known to accompany metarhodopsin II formation. The protein remains in a conformation similar to that of the ground state until late in the photobleaching process.     

Infectious molecular clone of a recently transmitted pediatric human immunodeficiency virus clade C isolate from Africa: evidence of intraclade recombination

Although human immunodeficiency virus type 1 (HIV-1) clade C continues to dominate the pandemic, only two infectious clade C proviral DNA clones have been described (N. Mochizuki, N. Otsuka, K. Matsuo, T. Shiino, A. Kojima, T. Kurata, K. Sakai, N. Yamamoto, S. Isomura, T. N. Dhole, Y. Takebe, M. Matsuda, and M. Tatsumi, AIDS Res. Hum. Retrovir. 15:1321-1324, 1999; T. Ndung'u, B. Renjifo, and M. Essex, J. Virol. 75:4964-4972, 2001). We have generated an infectious molecular clone of a pediatric clade C strain, HIV1084i, which was isolated from a Zambian infant infected either intrapartum or through breastfeeding. HIV1084i is an R5, non-syncytium-inducing isolate that bears all known clade C signatures; gag, pol, and env consistently mapped within clade C. Interestingly, gag resembled Asian isolates, whereas pol and env resembled African isolates, indicating that HIV1084i probably arose from an intraclade recombination. As a recently transmitted clade C strain, HIV1084i will be a useful vaccine development tool.     

Long-term productive human immunodeficiency virus infection of CD1a-sorted myeloid dendritic cells

Myeloid, CD1a-sorted dendritic cells (MDC) productively replicated human immunodeficiency virus strains encoding envelope genes of either primary X4R5 or R5 strains for up to 45 days. Cell-free supernatant collected from long-term infected MDC, which had been exposed to an X4R5 virus 45 days earlier, was still infectious when placed over activated T cells. These data imply that DC can act as a persistent reservoir of infectious virus.     

At Limits of Life: Multidisciplinary Insights Reveal Environmental Constraints on Biotic Diversity in Continental Antarctica

Multitrophic communities that maintain the functionality of the extreme Antarctic terrestrial ecosystems, while the simplest of any natural community, are still challenging our knowledge about the limits to life on earth. In this study, we describe and interpret the linkage between the diversity of different trophic level communities to the geological morphology and soil geochemistry in the remote Transantarctic Mountains (Darwin Mountains, 80°S). We examined the distribution and diversity of biota (bacteria, cyanobacteria, lichens, algae, invertebrates) with respect to elevation, age of glacial drift sheets, and soil physicochemistry. Results showed an abiotic spatial gradient with respect to the diversity of the organisms across different trophic levels. More complex communities, in terms of trophic level diversity, were related to the weakly developed younger drifts (Hatherton and Britannia) with higher soil C/N ratio and lower total soluble salts content (thus lower conductivity). Our results indicate that an increase of ion concentration from younger to older drift regions drives a succession of complex to more simple communities, in terms of number of trophic levels and diversity within each group of organisms analysed. This study revealed that integrating diversity across multi-trophic levels of biotic communities with abiotic spatial heterogeneity and geological history is fundamental to understand environmental constraints influencing biological distribution in Antarctic soil ecosystems.     

Synthesis of 3′-O-Propargylthymidine as a Candidate Antiretroviral Agent

Abstract

3′-O-Propargylthymidinc, which may be viewed as a stnictural analogue of the potent antiretroviral agent 3′-azido-3′-deoxythymidine (AZT), was synthesized from 5′-O-(4,4′-dimethoxytritylthymidine by reaction with propargyl bromide followed by gentle acidolysis. The 3′-O-propargyl derivative was tested for antiretroviral activity in SC-1 mouse fibroblasts infected with Rauscher murine leukemia virus (MuLV). No inhibition of MuLV proliferation was observed at concentrations of 3′-O-propargylthymidine from 0.001 to 100 μM. whereas the IC₅₀ against the host cells was 30 μM. By comparison, AZT had an IC₅₀ for MuLV growth of 0.01 μM and an IC₅₀ for cell growth of >100 μM. Thus, replacement of the 3′-N-N≡N group in AZT by a 3′-OCH₂C≡CH group increased cytotoxicity but decreased antiretroviral activity relative to AZT.