Noncytotoxic orange and red/green derivatives of DsRed-Express2 for whole-cell labeling

<p>Abstract</p> <p>Background</p> <p>Whole-cell labeling is a common application of fluorescent proteins (FPs), but many red and orange FPs exhibit cytotoxicity that limits their use as whole-cell labels. Recently, a tetrameric red FP called DsRed-Express2 was engineered for enhanced solubility and was shown to be noncytotoxic in bacterial and mammalian cells. Our goal was to create derivatives of this protein with different spectral properties.</p> <p>Results</p> <p>Building on previous studies of DsRed mutants, we created two DsRed-Express2 derivatives: E2-Orange, an orange FP, and E2-Red/Green, a dual-color FP with both red and green emission. We show that these new FPs retain the low cytotoxicity of DsRed-Express2. In addition, we show that these new FPs are useful as second or third colors for flow cytometry and fluorescence microscopy.</p> <p>Conclusion</p> <p>E2-Orange and E2-Red/Green will facilitate the production of healthy, stably fluorescent cell lines and transgenic organisms for multi-color labeling studies.</p

Genomic mapping of RNA polymerase II reveals sites of co-transcriptional regulation in human cells

BACKGROUND: Transcription by RNA polymerase II is regulated at many steps including initiation, promoter release, elongation and termination. Accumulation of RNA polymerase II at particular locations across genes can be indicative of sites of regulation. RNA polymerase II is thought to accumulate at the promoter and at sites of co-transcriptional alternative splicing where the rate of RNA synthesis slows. RESULTS: To further understand transcriptional regulation at a global level, we determined the distribution of RNA polymerase II within regions of the human genome designated by the ENCODE project. Hypophosphorylated RNA polymerase II localizes almost exclusively to 5' ends of genes. On the other hand, localization of total RNA polymerase II reveals a variety of distinct landscapes across many genes with 74% of the observed enriched locations at exons. RNA polymerase II accumulates at many annotated constitutively spliced exons, but is biased for alternatively spliced exons. Finally, RNA polymerase II is also observed at locations not in gene regions. CONCLUSION: Localizing RNA polymerase II across many millions of base pairs in the human genome identifies novel sites of transcription and provides insights into the regulation of transcription elongation. These data indicate that RNA polymerase II accumulates most often at exons during transcription. Thus, a major factor of transcription elongation control in mammalian cells is the coordination of transcription and pre-mRNA processing to define exons

Chemical abundances in the protoplanetary disk LV2 (Orion): clues to the causes of the abundance anomaly in HII regions

Optical integral field spectroscopy of the archetype protoplanetary disk LV2 in the Orion Nebula is presented, taken with the VLT FLAMES/Argus fibre array. The detection of recombination lines of CII and OII from this class of objects is reported, and the lines are utilized as abundance diagnostics. The study is complemented with the analysis of HST Faint Object Spectrograph ultraviolet and optical spectra of the target contained within the Argus field of view. By subtracting the local nebula background the intrinsic spectrum of the proplyd is obtained and its elemental composition is derived for the first time. The proplyd is found to be overabundant in carbon, oxygen and neon compared to the Orion Nebula and the sun. The simultaneous coverage over LV2 of the CIII] 1908-A and [OIII] 5007-A collisionally excited lines (CELs) and CII and OII recombination lines (RLs) has enabled us to measure the abundances of C++ and O++ for LV2 with both sets of lines. The two methods yield consistent results for the intrinsic proplyd spectrum, but not for the proplyd spectrum contaminated by the generic nebula spectrum, thus providing one example where the long-standing abundance anomaly plaguing metallicity studies of HII regions has been resolved. These results would indicate that the standard forbidden-line methods used in the derivation of light metal abundances in HII regions in our own and other galaxies underestimate the true gas metallicity.Comment: Accepted by MNRAS November 8; 16 pages, 9 figs; typos corrected, error in FWHMs in table 4 corrected in this versio

Radiocarbon data reveal contrasting sources for carbon fractions in thermokarst lakes and rivers of Eastern Canada (Nunavik, Quebec)

Greenhouse gas (GHG) emissions from permafrost organic carbon decomposition in lakes and rivers can accelerate global warming. We used radiocarbon (14C) measurements to determine the predominant sources of dissolved organic carbon (DOC), particulate organic carbon (POC), dissolved inorganic carbon (DIC), and methane (CH4) in five thermokarst lakes and three rivers in an area of widespread permafrost degradation in Northern Quebec to assess contributions from thawing permafrost and other old carbon (fixed before CE 1950) reservoirs. We compared emission pathways (dissolved gas and ebullition), seasons (summer and winter), and surface soil type (mineral and peat soils). Modern carbon (fixed after CE 1950) was the dominant source of DOC, DIC, and CH4 of non‐peatland aquatic systems, while POC and sediment carbon were predominantly fixed in the last millennia. In the peatland systems, modern and permafrost carbon were important sources of DOC, lake DIC, lake ebullition CO2, and lake dissolved CH4. In contrast, POC, lake ebullition CH4, and river DIC were dominated by millennial‐old carbon. In winter, the 14C age of DOC, DIC, and POC in the peatland lakes increased, but the 14C age of dissolved CH4 did not change. Our results point to a clearly older overall carbon source for ebullition CH4 relative to dissolved CH4 in the peatland lakes, but not the non‐peatland lakes. The younger ages of diffusive CH4 and DIC relative to DOC and POC in all lakes suggest that recent primary productivity strongly influences the large total lake CH4 emissions in this area, as diffusion fluxes greatly exceed ebullition fluxes

Biannual versus annual mass azithromycin distribution and malaria seroepidemiology among preschool children in Niger: a sub-study of a cluster randomized trial.

BACKGROUND: Biannual mass azithromycin administration to preschool children reduces all-cause mortality, but the mechanism for the effect is not understood. Azithromycin has activity against malaria parasites, and malaria is a leading cause of child mortality in the Sahel. The effect of biannual versus annual azithromycin distribution for trachoma control on serological response to merozoite surface protein 1 (MSP-119), a surrogate for malaria incidence, was evaluated among children in Niger. METHODS: Markers of malaria exposure were measured in two arms of a factorial randomized controlled trial designed to evaluate targeted biannual azithromycin distribution to children under 12 years of age compared to annual azithromycin to the entire community for trachoma control (N = 12 communities per arm). Communities were treated for 36 months (6 versus 3 distributions). Dried blood spots were collected at 36 months among children ages 1-5 years, and MSP-119 antibody levels were assessed using a bead-based multiplex assay to measure malaria seroprevalence. RESULTS: Antibody results were available for 991 children. MSP-119 seropositivity was 62.7% in the biannual distribution arm compared to 68.7% in the annual arm (prevalence ratio 0.91, 95% CI 0.83 to 1.00). Mean semi-quantitative antibody levels were lower in the biannual distribution arm compared to the annual arm (mean difference - 0.39, 95% CI - 0.05 to - 0.72). CONCLUSIONS: Targeted biannual azithromycin distribution was associated with lower malaria seroprevalence compared to that in a population that received annual distribution. Trial Registration Clinicaltrials.gov NCT00792922

Faint recombination lines in Galactic PNe with [WC] nucleus

We present spatially resolved high-resolution spectrophotometric data for the planetary nebulae PB8, NGC2867, and PB6. We have analyzed two knots in NGC2867 and PB6 and one in PB8. The three nebulae are ionized by [WC] type nuclei: early [WO] for PB6 and NGC2867 and [WC 5-6] in the case of PB8. Our aim is to study the behavior of the abundance discrepancy problem (ADF) in this type of PNe. We measured a large number of optical recombination (ORL) and collisionally excited lines (CEL), from different ionization stages (many more than in any previous work), thus, we were able to derive physical conditions from many different diagnostic procedures. We determined ionic abundances from the available collisionally excited lines and recombination lines. Based on both sets of ionic abundances, we derived total chemical abundances in the nebulae using suitable ionization correction factors. From CELs, we have found abundances typical of Galactic disk planetary nebulae. Moderate ADF(O++) were found for PB8 (2.57) and NGC2867 (1.63). For NGC2867, abundances from ORLs are higher but still consistent with Galactic disk planetary nebulae. On the contrary, PB8 presents a very high O/H ratio from ORLs. A high C/O was obtained from ORLs for NGC2867; this ratio is similar to C/O obtained from CELs and with the chemical composition of the wind of the central star, indicating that there was no further C-enrichment in the star, relative to O, after the nebular material ejection. On the contrary, we found C/O<1 in PB8. Interestingly, we obtain (C/O)ORLs/(C/O)CELs < 1 in PB8 and NGC2867; this added to the similarity between the heliocentric velocities measured in [OIII] and OII lines for our three objects, argue against the presence of H-deficient metal-rich knots coming from a late thermal pulse event.Comment: 25 pages, 13 Tables, 4 Figures Accepted for publication in A&A. First page is blank for obscure latex reason

Treatment seeking and antibiotic use for urinary tract infection symptoms in the time of COVID-19 in Tanzania and Uganda

Funding: CARE: COVID-19 and Antimicrobial Resistance in East Africa – impact and response is a Global Effort on COVID-19 (GECO) Health Research Award (MR/V036157/1) funded by UK Research and Innovation (Medical Research Council) and the Department of Health and Social Care (National Institute for Health Research).Background There is still little empirical evidence on how the outbreak of coronavirus disease 2019 (COVID-19) and associated regulations may have disrupted care-seeking for non-COVID-19 conditions or affected antibiotic behaviours in low- and middle-income countries (LMICs). We aimed to investigate the differences in treatment-seeking behaviours and antibiotic use for urinary tract infection (UTI)-like symptoms before and during the pandemic at recruitment sites in two East African countries with different COVID-19 control policies: Mbarara, Uganda and Mwanza, Tanzania. Methods In this repeated cross-sectional study, we used data from outpatients (pregnant adolescents aged >14 and adults aged >18) with UTI-like symptoms who visited health facilities in Mwanza, Tanzania and Mbarara, Uganda. We assessed the prevalence of self-reported behaviours (delays in care-seeking, providers visited, antibiotics taken) at three different time points, labelled as ‘pre-COVID-19 phase’ (February 2019 to February 2020), ‘COVID-19 phase 1’ (March 2020 to April 2020), and ‘COVID-19 phase 2’ (July 2021 to February 2022). Results In both study sites, delays in care-seeking were less common during the pandemic than they were in the pre-COVID phase. Patients in Mwanza, Tanzania had shorter care-seeking pathways during the pandemic compared to before it, but this difference was not observed in Mbarara, Uganda. Health centres were the dominant sources of antibiotics in both settings. Over time, reported antibiotic use for UTI-like symptoms became more common in both settings. During the COVID-19 phases, there was a significant increase in self-reported use of antibiotics like metronidazole (<30% in the pre-COVID-19 phase to 40% in COVID phase 2) and doxycycline (30% in the pre-COVID-19 phase to 55% in COVID phase 2) that were not recommended for treating UTI-like symptoms in the National Treatment Guidelines in Mbarara, Uganda. Conclusions There was no clear evidence that patients with UTI-like symptoms attending health care facilities had longer or more complex treatment pathways despite strict government-led interventions related to COVID-19. However, antibiotic use increased over time, including some antibiotics not recommended for treating UTI, which has implications for future antimicrobial resistance.Publisher PDFPeer reviewe

Deep Echelle Spectrophotometry of S 311, a galactic H II Region Outside the Solar Circle

We present echelle spectrophotometry of the Galactic H II region S 311. The data have been taken with the Very Large Telescope UVES echelle spectrograph in the 3100 to 10400 angstroms range. We have measured the intensities of 263 emission lines, 178 are permitted lines of H0, D0 (deuterium), He0, C0, C+, N0, N+, O0, O+, S+, Si0, Si+, Ar0 and Fe0; some of them are produced by recombination and others mainly by fluorescence. Physical conditions have been derived using different continuum and line intensity ratios. We have derived He+, C++ and O++ ionic abundances from pure recombination lines as well as abundances from collisionally excited lines for a large number of ions of different elements. We have obtained consistent estimations of t2 applying different methods. We have found that the temperature fluctuations paradigm is consistent with the t2(He I) vs. t2(H I) relation for H II regions, in contrast with what has been found for planetary nebulae. We report the detection of deuterium Balmer lines up to D6 in the blue wings of the hydrogen lines, which excitation mechanism seems to be continuum fluorescence.Comment: 13 pages, 12 tables, 5 figures, accepted for publication in MNRA

Partitioning the Heritability of Tourette Syndrome and Obsessive Compulsive Disorder Reveals Differences in Genetic Architecture

The direct estimation of heritability from genome-wide common variant data as implemented in the program Genome-wide Complex Trait Analysis (GCTA) has provided a means to quantify heritability attributable to all interrogated variants. We have quantified the variance in liability to disease explained by all SNPs for two phenotypically-related neurobehavioral disorders, obsessive-compulsive disorder (OCD) and Tourette Syndrome (TS), using GCTA. Our analysis yielded a heritability point estimate of 0.58 (se = 0.09, p = 5.64e-12) for TS, and 0.37 (se = 0.07, p = 1.5e-07) for OCD. In addition, we conducted multiple genomic partitioning analyses to identify genomic elements that concentrate this heritability. We examined genomic architectures of TS and OCD by chromosome, MAF bin, and functional annotations. In addition, we assessed heritability for early onset and adult onset OCD. Among other notable results, we found that SNPs with a minor allele frequency of less than 5% accounted for 21% of the TS heritability and 0% of the OCD heritability. Additionally, we identified a significant contribution to TS and OCD heritability by variants significantly associated with gene expression in two regions of the brain (parietal cortex and cerebellum) for which we had available expression quantitative trait loci (eQTLs). Finally we analyzed the genetic correlation between TS and OCD, revealing a genetic correlation of 0.41 (se = 0.15, p = 0.002). These results are very close to previous heritability estimates for TS and OCD based on twin and family studies, suggesting that very little, if any, heritability is truly missing (i.e., unassayed) from TS and OCD GWAS studies of common variation. The results also indicate that there is some genetic overlap between these two phenotypically-related neuropsychiatric disorders, but suggest that the two disorders have distinct genetic architectures