SopD acts cooperatively with SopB during Salmonella enterica serovar Typhimurium invasion

The intracellular bacterial pathogen, Salmonella enterica serovar Typhimurium (S. typhimurium), causes disease in a variety of hosts. To invade and replicate in host cells, these bacteria subvert host molecular machinery using bacterial proteins, called effectors, which they translocate into host cells using specialized protein delivery systems. One of these effectors, SopD, contributes to gastroenteritis, systemic virulence and persistence of S. typhimurium in animal models of infection. Recently, SopD has been implicated in invasion of polarized epithelial cells and here we investigate the features of SopD-mediated invasion. We show that SopD plays a role in membrane fission and macropinosome formation during S. typhimurium invasion, events previously shown to be mediated by the SopB effector. We further demonstrate that SopD acts cooperatively with SopB to promote these events during invasion. Using live cell imaging we show that a SopD-GFP fusion does not localize to HeLa cell cytosol as previously described, but instead is membrane associated. Upon S. typhimurium infection of these cells, SopD-GFP is recruited to the invasion site, and this recruitment required the phosphatase activity of SopB. Our findings demonstrate a role for SopD in manipulation of host-cell membrane during S. typhimurium invasion and reveal the nature of its cooperative action with SopB.     

6-Azathymidine-4'-thionucleosides: synthesis and antiviral evaluation

The synthesis of dideoxy-6-azathymidine 4'-thionucleoside 1-(2,3-dideoxy-4-thio-beta-D-erythro-pentofuranosyl)-(6-azathymidine) (2), and the L-nucleoside, 1-(4-thio-beta-L-erythro-pentofuranosyl)-(6-azathymidine) (3) and their evaluation against a wide panel of antiviral assays are described. The L-thionucleoside (3) was devoid of antiviral activity. The dideoxy-thionucleoside (2) was moderately active against vaccinia virus (VV) and the herpes simplex virus strains HSV-1 (strain KOS) and HSV-2 (strain G) (MIC 12 microM) and retained inhibitory activity vs a thymidine kinase-deficient strain HSV-1/TK(-), suggesting that (2) is not dependent on viral TK-catalysed phosphorylation for antiviral activity and/or may use an alternative metabolic activation pathway.     

Budding yeast Rif1 binds to replication origins and protects DNA at blocked replication forks

Despite its evolutionarily conserved function in controlling DNA replication, the chromosomal binding sites of the budding yeast Rif1 protein are not well understood. Here, we analyse genome‐wide binding of budding yeast Rif1 by chromatin immunoprecipitation, during G1 phase and in S phase with replication progressing normally or blocked by hydroxyurea. Rif1 associates strongly with telomeres through interaction with Rap1. By comparing genomic binding of wild‐type Rif1 and truncated Rif1 lacking the Rap1‐interaction domain, we identify hundreds of Rap1‐dependent and Rap1‐independent chromosome interaction sites. Rif1 binds to centromeres, highly transcribed genes and replication origins in a Rap1‐independent manner, associating with both early and late‐initiating origins. Interestingly, Rif1 also binds around activated origins when replication progression is blocked by hydroxyurea, suggesting association with blocked forks. Using nascent DNA labelling and DNA combing techniques, we find that in cells treated with hydroxyurea, yeast Rif1 stabilises recently synthesised DNA. Our results indicate that, in addition to controlling DNA replication initiation, budding yeast Rif1 plays an ongoing role after initiation and controls events at blocked replication forks.     

Potential effects of ethnicity in genetic and environmental sources of variability in the stature, mass, and body mass index of children

We examined sources of variability in stature, body mass, and body mass index (BMI) in families of black and white elementary schoolchildren from Philadelphia, Pennsylvania. The sample consisted of 445 black and 379 white children, 7-13 years old, and their parents (total n = 2016). The sample was distributed among 596 nuclear families, each representing an independent pedigree. Maximum-likelihood-based variance decomposition methods were used to simultaneously estimate ethnic group-specific effects of genes, sex, age by sex, and unmeasured environmental factors on stature, body mass, and BMI. Likelihood ratio tests were performed to assess the significance of h2 estimates and differences in sigma g and sigma e between black and white families. Genes account for moderate proportions of the phenotypic variance (h2) of these traits in black and white children. In black and white children, respectively, h2 estimates were 0.37 and 0.53 for stature, 0.37 and 0.31 for body mass, and 0.38 and 0.24 for BMI (p < 0.0005). Although the differences in h2 between ethnic groups were not significant (stature, p = 0.23; body mass, p = 0.49; BMI, p = 0.14), black children exhibited a significantly greater total residual phenotypic standard deviation (sigma e and sigma g) in body mass and BMI and a significantly greater sigma e for stature compared with white children. The larger residual phenotypic variance in the black sample is likely due to exposure to unmeasured environmental factors that are not accounted for in this model. Given that sigma g for stature is not significantly different between ethnic groups, the slightly lower estimates in black children are due to the increased contribution of the environment to the phenotypic variance in this trait.     

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.     

Nucleotide polymorphism at the alcohol dehydrogenase locus of pocket gophers, genus Geomys

Using the strictly neutral model as a null hypothesis, we tested for deviations from expected levels of nucleotide polymorphism at the alcohol dehydrogenase locus (Adh-1) within and among four species of pocket gophers (Geomys bursarius major, G. knoxjonesi, G. texensis llanensis, and G. attwateri). The complete protein-encoding region was examined, and 10 unique alleles, representing both electromorphic and cryptic alleles, were used to test hypotheses (e.g., the neutral model) concerning the maintenance of genetic variation. Nineteen variable sites were identified among the 10 alleles examined, including 9 segregating sites occurring in synonymous positions and 10 that were nonsynonymous. Several statistical methods, including those that test for within-species variation as well as those that examine variation within and among species, failed to reject the null hypothesis that variation (both within and between species of Geomys) at the Adh locus is consistent with the neutral theory. However, there was significant heterogeneity in the ratio of polymorphism to divergence across the gene, with polymorphisms clustered in the first half of the coding region and fixed differences clustered in the second half of the gene. Two alternative hypotheses are discussed as possible explanations for this heterogeneity: an old balanced polymorphism in the first half of the gene or a recent selective sweep in the second half of the gene.      

In vivo uptake of a haem analogue Zn protoporphyrin IX by the human malaria parasite P. falciparum‐infected red blood cells

The cellular traffic of haem during the development of the human malaria parasite Plasmodium falciparum, through the stages R (ring), T (trophozoite) and S (schizonts), was investigated within RBC (red blood cells). When Plasmodium cultures were incubated with a fluorescent haem analogue, ZnPPIX (Zn protoporphyrin IX) the probe was seen at the cytoplasm (R stage), and the vesicle‐like structure distribution pattern was more evident at T and S stages. The temporal sequence of ZnPPIX uptake byP. falciparum‐infected erythrocytes shows that at R and S stages, a time‐increase acquisition of the porphyrin reaches the maximum fluorescence distribution after 60 min; in contrast, at the T stage, the maximum occurs after 120 min of ZnPPIX uptake. The difference in time‐increase acquisition of the porphyrin is in agreement with a maximum activity of haem uptake at the T stage. To gain insights into haem metabolism, recombinant PfHO (P. falciparum haem oxygenase) was expressed, and the conversion of haem into BV (biliverdin) was detected. These findings point out that, in addition to haemozoin formation, the malaria parasite P. falciparum has evolved two distinct mechanisms for dealing with haem toxicity, namely, the uptake of haem into a cellular compartment where haemozoin is formed and HO activity. However, the low Plasmodium HO activity detected reveals that the enzyme appears to be a very inefficient way to scavenge the haem compared with the Plasmodium ability to uptake the haem analogue ZnPPIX and delivering it to the food vacuole.     

A consideration of factors underlying the selection of methods in the assessment of skeletal maturity

The assessment of skeletal maturity is basically a method. How one makes this assessment or what one does with the assessment is dictated by the research and/or clinical problem at hand. A variety of methods have been suggested for the assessment of skeletal maturity, although the two most commonly used are the inspectional and bone specific approaches. Several of the proposed methods are considered, with primary emphasis upon the problems, factors, and/or alternatives related to the choice of one method over another.     

Reduced natural selection associated with low recombination in Drosophila melanogaster

Synonymous codons are not used equally in many organisms, and the extent of codon bias varies among loci. Earlier studies have suggested that more highly expressed loci in Drosophila melanogaster are more biased, consistent with findings from several prokaryotes and unicellular eukaryotes that codon bias is partly due to natural selection for translational efficiency. We link this model of varying selection intensity to the population-genetics prediction that the effectiveness of natural selection is decreased under reduced recombination. In analyses of 385 D. melanogaster loci, we find that codon bias is reduced in regions of low recombination (i.e., near centromeres and telomeres and on the fourth chromosome). The effect does not appear to be a linear function of recombination rate; rather, it seems limited to regions with the very lowest levels of recombination. The large majority of the genome apparently experiences recombination at a sufficiently high rate for effective natural selection against suboptimal codons. These findings support models of the Hill-Robertson effect and genetic hitchhiking and are largely consistent with multiple reports of low levels of DNA sequence variation in regions of low recombination.