Intrinsic stability of episomal circles formed during human immunodeficiency virus type 1 replication

The development of surrogate markers capable of detecting residual ongoing human immunodeficiency virus type 1 (HIV-1) replication in patients receiving highly active antiretroviral therapy is an important step in understanding viral dynamics and in developing new treatment strategies. In this study, we evaluated the utility of circular forms of the viral genome for the detection of recent infection of cells by HIV-1. We measured the fate of both one-long terminal repeat (1-LTR) and 2-LTR circles following in vitro infection of logarithmically growing CD4+ T cells under conditions in which cell death was not a significant contributing factor. Circular forms of the viral genome were found to be highly stable and to decrease in concentration only as a function of dilution resulting from cell division. We conclude that these DNA circles are not intrinsically unstable in all cell types and suggest that the utility of 2-LTR circle assays in measuring recent HIV-1 infection of susceptible cells in vivo needs to be reevaluated.     

Changes in radial tree growth for Picea abies, Larix decidua, Pinus cembra and Pinus uncinata near the alpine timberline since 1750

Changes in radial growth of the four coniferous species growing in the French Alps near the upper treeline are investigated. Thirty-seven populations of Norway spruce [Picea abies (L.) Karst.], European larch (Larix decidua Mill.), Swiss stone pine (Pinus cembra L.) and mountain pine (Pinus uncinata Mill. ex Mirb.) were sampled by taking 1320 cores and analysing tree-ring widths. Sites were chosen in various climatic conditions (macroclimate and aspect) and on two kinds of bedrock in order to take into account the ecological behaviour of these species. Belledonne, Moyenne-Tarentaise, Haute-Maurienne and Briançonnais areas were sampled along increasing gradients of summer aridity and winter continentality. The calculation of time series after removing the age trend brings strong evidence for an increase in radial growth during the two last centuries, but with different stages and fluctuations for each species. This growth trend is significantly enhanced since 1860 for the spruce, and since 1920 for the two pine species. Furthermore, it also appears on Larix decidua with the same pattern despite periodical growth reduction due to attacks of the larch bud moth (Zeiraphera diniana Gn.). The analysis of ring-widths at a given cambial age reveals that this enhanced phenomenon is observed especially during the tree’s early years (25–75 years). The analysis of four regional climatic series, and three longer series of temperature (in farther single sites) reveals synchronous decadal fluctuations and an evident secular increase in minimum temperatures (especially in January and from July to October), that may be involved in tree-growth enhancement. Thermic amplitudes are significantly reduced during the whole growing period, what is more pronounced in Belledonne, the most oceanic region. Long term growth changes are well described by stepwise regression models, especially for the pine species. These models involved both a linear trend (CO₂ concentration or N-deposition) and low frequency of Turin monthly temperatures. However, they show different patterns than those observed from response functions at a yearly scale.     

Glucan synthesis by intact cotton fibres fed with different precursors at the stages of primary and secondary wall formation

Seed clusters of individual locules from fruit capsules of Gossypium arboreum L. with adhering intact fibres were fed with radioactive uridinediphosphoglucose (UDPG), guanosinediphosphoglucose (GDPG), glucose and sucrose. The incorporation into high molecular weight glucans of the fibres was studied. For primary wall fibres, UDPG at 1 mM was by far the best precursor, whereas sucrose was the best precursor for secondary wall fibres. No competition was observed between the incorporation of glucose from UDPG and from sucrose when the two were fed simultaneously to secondary wall fibres, indicating that their metabolic pathways are well separated when they are fed from the apoplast. Inhibitors of respiratory ATP-formation strongly inhibited incorporation of sucrose but not that of UDPG. Sucrose incorporation was studied at five different stages of development of the cotton fibres. At the stage of most intense secondary wall formation the incorporation rate was about 300 times that during primary wall formation (24 days post anthesis (DPA)). Incorporation from 1 mM UDPG or GDPG by secondary wall fibres (35 DPA) was less than twice that of primary wall fibres (22 DPA), indicating that the two sugar nucleotides are not readily used as precursors for secondary wall cellulose when they are fed to the exterior of intact cells. The high molecular weight non-cellulosic glucans formed from UDPG and sucrose at 5 and 1,000 M were solubilized in strongly alkaline solutions or dimethyl-sulfoxide (DMSO) and were partially characterized by degradation with an exo--1,3-glucanase. After feeding for one hour, at most 1/3 of the radioactivity in high molecular weight material was found in cellulose and at least 2/3 in -1,3-glucan. The proportions varied little for fibres in the age range of 30 to 48 DPA when sucrose was the precursor although the total incorporation varied by a factor of about four. The fact that at all stages of secondary wall formation -1,3-glucan is synthesized at a very high rate, but that the total amount in the cell wall does not exceed 2% in the later stages of wall formation, can be interpreted in terms of a high turnover of this polysaccharide if it is assumed that wound effects are negligible in the system under study.Abbreviations UDPG uridinediphosphoglucose - GDPG guanosinediphosphoglucose - HEPES N-2-hydroxyethylpiperazine-N-2-ethansulphonic acid - DMSO dimethyl-sulfoxide - DNP 2,4-dinitrophenol - DPA days post anthesis     

Queen size dimorphism in the ant Tetramorium moravicum (Hymenoptera, Formicidae): Morphometric, molecular genetic and experimental evidence

Summary. By combining different methods we evaluate whether the ant Tetramorium rhenanum is specifically separated from T. moravicum or whether it is a conspecific microgyne form. High-precision morphometry shows a clear difference in queen size. Sequence comparison of 1031 bp of COI reveals that T. rhenanum falls into a clade with T. moravicum, which is significantly separated from T. forte and T. chefketi. T. rhenanum shares at least two haplotypes with T. moravicum and is considered as a junior synonym. Sexual behaviour and colony foundation experiments corroborate conspecificity. The queen dimorphism is discussed in the context of social parameters such as queen number and reproductive strategy.Received 10 June 2004; revised 28 October 2004; accepted 12 November 2004.     

Protein dynamics detected in a membrane-embedded potassium channel using two-dimensional solid-state NMR spectroscopy

We report longitudinal ¹⁵N relaxation rates derived from two-dimensional (¹⁵N, ¹³C) chemical shift correlation experiments obtained under magic angle spinning for the potassium channel KcsA-Kv1.3 reconstituted in multilamellar vesicles. Thus, we demonstrate that solid-state NMR can be used to probe residue-specific backbone dynamics in a membrane-embedded protein. Enhanced backbone mobility was detected for two glycine residues within the selectivity filter that are highly conserved in potassium channels and that are of core relevance to the filter structure and ion selectivity.     

Long-term adherence to antiretroviral treatment and program drop-out in a high-risk urban setting in sub-Saharan Africa: a prospective cohort study

Background

Seventy percent of urban populations in sub-Saharan Africa live in slums. Sustaining HIV patients in these high-risk and highly mobile settings is a major future challenge. This study seeks to assess program retention and to find determinants for low adherence to antiretroviral treatment (ART) and drop-out from an established HIV/ART program in Kibera, Nairobi, one of Africa''s largest informal urban settlements.

Methods and Findings

A prospective open cohort study of 800 patients was performed at the African Medical Research Foundation (AMREF) clinic in the Kibera slum. Adherence to ART and drop-out from the ART program were independent outcomes. Two different adherence measures were used: (1) “dose adherence” (the proportion of a prescribed dose taken over the past 4 days) and (2) “adherence index” (based on three adherence questions covering dosing, timing and special instructions). Drop-out from the program was calculated based on clinic appointment dates and number of prescribed doses, and a patient was defined as being lost to follow-up if over 90 days had expired since the last prescribed dose. More than one third of patients were non-adherent when all three aspects of adherence – dosing, timing and special instructions – were taken into account. Multivariate logistic regression revealed that not disclosing HIV status, having a low level of education, living below the poverty limit (US$ 2/day) and not having a treatment buddy were significant predictors for non-adherence. Additionally, one quarter of patients dropped out for more than 90 days after the last prescribed ART dose. Not having a treatment buddy was associated with increased risk for drop-out (hazard ratio 1.4, 95% CI = 1.0–1.9).

Conclusion

These findings point to the dilemma of trying to sustain a growing number of people on life-long ART in conditions where prevailing stigma, poverty and food shortages threatens the long-term success of HIV treatment.     

Coxiella burnetii infection in C57BL/6 mice aged 1 or 14 months

The objective of this study was to investigate the effects of age on infection with Coxiella burnetii, the agent of Q fever. Bacterial burden and granuloma number were increased in the spleens of 14-month-old as compared with 1-month-old mice. This increase was not the result of an anti-inflammatory macrophage response, because inflammatory and anti-inflammatory cytokines were induced in macrophages from young mice but were repressed in mature mice. In addition, macrophage microbicidal competence was similar in mature and young mice. These results suggest the importance of individual host factors in the pathophysiology of an infectious disease such as Q fever.     

Directional asymmetry of the zebrafish epithalamus guides dorsoventral innervation of the midbrain target

The zebrafish epithalamus, consisting of the pineal complex and flanking dorsal habenular nuclei, provides a valuable model for exploring how left-right differences could arise in the vertebrate brain. The parapineal lies to the left of the pineal and the left habenula is larger, has expanded dense neuropil, and distinct patterns of gene expression from the right habenula. Under the influence of Nodal signaling, positioning of the parapineal sets the direction of habenular asymmetry and thereby determines the left-right origin of habenular projections onto the midbrain target, the interpeduncular nucleus (IPN). In zebrafish with parapineal reversal, neurons from the left habenula project to a more limited ventral IPN region where right habenular axons would normally project. Conversely, efferents from the right habenula adopt a more extensive dorsoventral IPN projection pattern typical of left habenular neurons. Three members of the leftover-related KCTD (potassium channel tetramerization domain containing) gene family are expressed differently by the left and right habenula, in patterns that define asymmetric subnuclei. Molecular asymmetry extends to protein levels in habenular efferents, providing additional evidence that left and right axons terminate within different dorsoventral regions of the midbrain target. Laser-mediated ablation of the parapineal disrupts habenular asymmetry and consequently alters the dorsoventral distribution of innervating axons. The results demonstrate that laterality of the dorsal forebrain influences the formation of midbrain connections and their molecular properties.     

Responses of Antarctic Iridaea cordata (Rhodophyta) tetraspores exposed to ultraviolet radiation

In Antarctica ozone depletion is highest during spring, coinciding with the reproduction of many seaweed species. Propagules are the life-stage of an alga most susceptible to environmental perturbations. Therefore, fertile thalli of Iridaea cordata (Turner) Bory (Rhodophyta) were collected in the eulittoral of King George Island (Antarctica) to examine spore susceptibility to ultraviolet radiation (UVR). In the laboratory, freshly released tetraspores were exposed to photosynthetically active radiation (PAR) (400–700 nm), PAR+UV-A (320–700 nm) or PAR+UV-A+UV-B (280–700 nm). Photosynthetic efficiency was measured during 1–8 h of exposure and after 48 h of recovery. Additionally, mycosporine-like amino acids (MAAs) and DNA damage were determined. Saturating irradiance of photosynthesis of freshly released tetraspores was 57 µmol photons m⁻² s⁻¹. Exposure to increasing fluence of PAR reduced photosynthetic efficiency. UVR further decreased the photosynthetic efficiencies of the tetraspores but spores were able to recover completely after UVR exposure and 2 days post-cultivation under low PAR. DNA damage was minimal and lesions were effectively repaired under photoreactivating light. Concentrations of the MAAs shinorine and palythine were higher in tetraspores treated with UVR than in spores only exposed to PAR. Generally, the tetraspores show a good UV tolerance. This flexible response of the tetraspores of this species to changing radiation conditions enables the alga to grow along a considerable depth gradient from the sublittoral to the eulittoral where they can be exposed to enhanced UVBR under conditions of stratospheric ozone depletion.     

Variety‐specific Epidemiology of Cercospora beticola Sacc. and Consequences for Threshold‐based Timing of Fungicide Application in Sugar Beet

In Central Europe, fungicides to control leaf spot disease in sugar beet caused by Cercospora beticola are applied based on thresholds of disease incidence (DI, per cent of infected plants). As variety‐specific fungicide application was not analyzed to date, the epidemiology of C. beticola and its effect on white sugar yield (WSY) in varieties with different susceptibility were investigated at seven sites in Germany and Austria in 2004 and 2005. All varieties reached the summary thresholds 5 / 15 / 45% DI in all environments. Fitting a logistic growth curve to DI revealed significant differences among varieties. At high disease pressure, susceptible varieties reached a considerably higher disease severity (DS, per cent of infected leaf area) at harvest and a larger area under disease progress curve (AUDPC) than resistant varieties. Fitting a logistic growth curve to DS showed an increasing differentiation among varieties with time. The growth rate estimated based on the logistic growth curve was the only variable that performed equally well in differentiating varieties under low and high disease pressure. With increasing disease pressure, varieties differed considerably in WSY, but differences between susceptible and resistant varieties were significant only in some environments. The disease‐loss relation between AUDPC and relative WSY was variety‐specific. Resistant varieties had an approximately identical WSY with and without infection and compensated for negative infection effects even at higher AUDPC. Therefore, at high disease pressure, resistant varieties had a higher relative yield compared to susceptible ones. However, our results indicate that there is no need to develop variety‐specific thresholds, but resistant varieties reach the established thresholds later than susceptible ones. Consequently, the time of fungicide application can be delayed in resistant varieties. This will help to reduce the use of fungicides to the bare essentials as requested for the integrated crop protection management.