The rhizosphere is the interface between the root system of a plant and its surrounding soil. The microbiome of the rhizosphere, which is the totality of all microbes present there, represents a ...complex microbial ecosystem that nourishes the terrestrial biosphere. To untangle the complexity of the rhizosphere, and of the rhizospheric microbiome in particular, an integrated multi-omics approach can be applied to reveal the composition of the rhizospheric microbiome (through 16S ribosomal amplicons and metagenomics), the functional properties of the microbiome (through metatranscriptomics and metaproteomics), and the signaling network within the rhizosphere (through metametabolomics). The successful application of integrated multi-omics to rhizospheric science depends on the availability of rhizosphere-specific data and on the appropriate software used to analyze -omics data from the rhizosphere. In this review, we analyze the availability of software suites that are normally applied to surrogate disciplines (e.g., soil and plants) but which can be used for rhizospheric science. We also identify potential issues, challenges, and opportunities for rhizosphere science.
Genetic polymorphism in Plasmodium falciparum is a considerable obstacle to malaria intervention. Parasites have repeatedly evolved to overcome every front-line antimalarial deployed throughout ...history, and artemisinin resistant populations are expanding in Southeast Asia. Promising vaccine candidates routinely fail when challenged by the genetic diversity of natural parasite populations, and a recent trial using a blood-stage antigen showed immunity was allele specific. Modern sequencing technologies have revolutionized our understanding of parasite genomics and population genetics by providing access to single nucleotide variation, but characterizing more complex polymorphism remains a key challenge. Solving this problem is important because the selective pressures from drugs and host immunity often create complex polymorphism in the most clinically relevant genes that is missed using standard genotyping methods. In three sections, this thesis is a narrative about 1) encountering complex variation, 2) overcoming it with novel tools, and then 3) innovatively applying those tools to old and new questions. I first show examples of complex variation in a vaccine candidate (EBA-175) and a drug resistance gene (pfcrt) while reporting SNP based analyses of Kenyan and Tanzanian field isolates. While introducing this complex variation I also describe biological insights discovered in these populations. In Kenya I show evidence that chloroquine resistance selects for parasites that are primaquine sensitive, use a GWAS approach to discover new drug resistance loci, and catalogue variation in known resistance genes. In Tanzania I describe the population structure and allele frequencies of parasites from two geographic regions. In the second section of the thesis I develop methods for accessing complex variation and demonstrate their utility by producing de novo assemblies of eba-175, pfcrt, ama1, and msp3.4 from thousands of sequenced samples. Finally, in the third section I apply these tools in depth to eba-175. I comprehensively characterize the SNP and structural variation in eba-175 using an alignment of 1419 de novo assemblies. I use this resource to illustrate the profiles of positive selection across the gene, and corroborate these signals of balancing selection by showing the geographic distribution of the F/C indels and a lesser known 6bp indel positioned between the DBL domains. I then use the alignments to design Sequenom genotyping assays that facilitate a genome wide association study, testing for human associations with the eba-175 indels in the infecting parasite. I close by reporting a potential association on human chromosome 14 with the 6bp indel in eba-175.
Genetic polymorphism in Plasmodium falciparum is a considerable obstacle to malaria intervention. Parasites have repeatedly evolved to overcome every front-line antimalarial deployed throughout ...history, and artemisinin resistant populations are expanding in Southeast Asia. Promising vaccine candidates routinely fail when challenged by the genetic diversity of natural parasite populations, and a recent trial using a blood-stage antigen showed immunity was allele specific. Modern sequencing technologies have revolutionized our understanding of parasite genomics and population genetics by providing access to single nucleotide variation, but characterizing more complex polymorphism remains a key challenge. Solving this problem is important because the selective pressures from drugs and host immunity often create complex polymorphism in the most clinically relevant genes that is missed using standard genotyping methods. In three sections, this thesis is a narrative about 1) encountering complex variation, 2) overcoming it with novel tools, and then 3) innovatively applying those tools to old and new questions. I first show examples of complex variation in a vaccine candidate (EBA-175) and a drug resistance gene (pfcrt) while reporting SNP based analyses of Kenyan and Tanzanian field isolates. While introducing this complex variation I also describe biological insights discovered in these populations. In Kenya I show evidence that chloroquine resistance selects for parasites that are primaquine sensitive, use a GWAS approach to discover new drug resistance loci, and catalogue variation in known resistance genes. In Tanzania I describe the population structure and allele frequencies of parasites from two geographic regions. In the second section of the thesis I develop methods for accessing complex variation and demonstrate their utility by producing de novo assemblies of eba-175, pfcrt, ama1, and msp3.4 from thousands of sequenced samples. Finally, in the third section I apply these tools in depth to eba-175. I comprehensively characterize the SNP and structural variation in eba-175 using an alignment of 1419 de novo assemblies. I use this resource to illustrate the profiles of positive selection across the gene, and corroborate these signals of balancing selection by showing the geographic distribution of the F/C indels and a lesser known 6bp indel positioned between the DBL domains. I then use the alignments to design Sequenom genotyping assays that facilitate a genome wide association study, testing for human associations with the eba-175 indels in the infecting parasite. I close by reporting a potential association on human chromosome 14 with the 6bp indel in eba-175.
Over the past century, the significance of the rhizosphere as a complex, biological system, comprised of vast, interconnected networks of microbial organisms that interact directly with their plant ...hosts (e.g., archæa, bacteria, fungi, eukaryotes, and viruses) has been increasingly recognized by the scientific community. Providing a nutritional base to the terrestrial biosphere, the rhizosphere is integral to plant growth, crop production and ecosystem health. Lack of mechanistic understanding of the rhizosphere constitutes a critical knowledge gap, inhibiting our ability to predict and control the terrestrial ecosystem in order to achieve desirable outcomes (e.g., bioenergy production, crop yield maximization, and soilbased carbon sequestration). Application of multi-omics has the potential to significantly advance our knowledge of rhizospheric science. This review covers: cutting- and bleeding-edge, multi-omic techniques and technologies; methods and protocols for specific rhizospheric science questions; and, challenges to be addressed during this century of rhizospheric science.
Background Drug resistance remains a chief concern for malaria control. In order to determine the genetic markers of drug resistant parasites, we tested the genome-wide associations (GWA) of ...sequence-based genotypes from 35 Kenyan P. falciparum parasites with the activities of 22 antimalarial drugs. Methods and Principal Findings Parasites isolated from children with acute febrile malaria were adapted to culture, and sensitivity was determined by in vitro growth in the presence of anti-malarial drugs. Parasites were genotyped using whole genome sequencing techniques. Associations between 6250 single nucleotide polymorphisms (SNPs) and resistance to individual anti-malarial agents were determined, with false discovery rate adjustment for multiple hypothesis testing. We identified expected associations in the pfcrt region with chloroquine (CQ) activity, and other novel loci associated with amodiaquine, quinazoline, and quinine activities. Signals for CQ and primaquine (PQ) overlap in and around pfcrt, and interestingly the phenotypes are inversely related for these two drugs. We catalog the variation in dhfr, dhps, mdr1, nhe, and crt, including novel SNPs, and confirm the presence of a dhfr-164L quadruple mutant in coastal Kenya. Mutations implicated in sulfadoxine-pyrimethamine resistance are at or near fixation in this sample set. Conclusions/Significance Sequence-based GWA studies are powerful tools for phenotypic association tests. Using this approach on falciparum parasites from coastal Kenya we identified known and previously unreported genes associated with phenotypic resistance to anti-malarial drugs, and observe in high-resolution haplotype visualizations a possible signature of an inverse selective relationship between CQ and PQ.
Colorimetric reagent dipstick (CRD) for leukocyte esterase (LE) has shown potential for diagnosing and ruling out bacterial meningitis. Potential advantages over traditional cerebrospinal fluid (CSF) ...analysis include the small quantity of CSF required, rapid results, and easy interpretation. Our study aimed to determine whether clinicians in LMICs could accurately diagnose bacterial meningitis using CRD at the bedside.
A convenience sample of 143 patients requiring lumbar puncture for possible meningitis were enrolled from 1 October 2018 to 31 December 2019 at three hospitals, one each in rural Burundi, the Democratic Republic of Congo, and Kenya. CSF was analyzed using CRD followed by traditional laboratory-based analysis by technicians blinded to bedside results. Results were analyzed for concordance rates, sensitivity/specificity, positive and negative predictive values and impact on clinical decision-making.
One hundred and one patients were included in the analysis. The prevalence of bacterial meningitis in the convenience sample was 35% (35/101) as defined by microscopy or positive Gram stain. Using a threshold of “any positivity” for LE on the CRD, bedside testing correctly identified 33/35 cases (sensitivity 94.3%) and had a NPV of 92%. When only a clearly positive (≥ “+” for LE) CRD criterion was used, sensitivity and NPV were 77.1% and 86.2%, respectively.
Despite considerable promise, in our study, color reagent dipstick analysis of CSF did not perform well enough to rule out meningitis or screen samples for the need for microscopy. The development of a CSF-specific dipstick should be considered.
Insufficient or excessive thyroid hormone (TH) levels during fetal development can cause long-term neurological and cognitive problems. Studies in animal models of perinatal hypo- and hyperthyroidism ...suggest that these problems may be a consequence of the formation of maladaptive circuitry in the cerebral cortex, which can persist into adulthood. Here we used mouse models of maternal hypo- and hyperthyroidism to investigate the long-term effects of altering thyroxine (T4) levels during pregnancy (corresponding to embryonic days 6.5-18.5) on thalamocortical (TC) axon dynamics in adult offspring. Because perinatal hypothyroidism has been linked to visual processing deficits in humans, we performed chronic two-photon imaging of TC axons and boutons in primary visual cortex (V1). We found that a decrease or increase in maternal serum T4 levels was associated with atypical steady-state dynamics of TC axons and boutons in V1 of adult offspring. Hypothyroid offspring exhibited axonal branch and bouton dynamics indicative of an abnormal increase in TC connectivity, whereas changes in hyperthyroid offspring were indicative of an abnormal decrease in TC connectivity. Collectively, our data suggest that alterations to prenatal T4 levels can cause long-term synaptic instability in TC circuits, which could impair early stages of visual processing.
A series of novel aminoacyl adenylate mimics has been prepared and evaluated for their inhibitory activity against aminoacyl-tRNA synthetases. Several of these thiazole derivatives displayed potent ...and selective enzyme activity against both Gram-positive and Gram-negative bacteria.
A series of novel aminoacyl adenylate mimics has been prepared and evaluated for inhibitory activity against aminoacyl-tRNA synthetases. Several of these thiazole derivatives displayed potent and selective enzyme activity against both Gram-positive and Gram-negative bacteria.
Europe Lindbloom, Jason R; Balan, Violeta I; Nieuwveld, Lisa Bench ...
The International lawyer,
07/2009, Letnik:
43, Številka:
2
Journal Article
Recenzirano
Introduction The compliance and operating costs resulting from twenty-seven different private limited liability company regimes throughout the European Union makes the operation of foreign ...subsidiaries of SMEs considerably more expensive than domestic subsidiaries. ... EU SMEs that exercise their freedom of estabUshment rights (e.g., by incorporating as a UK private limited liability company while maintaining their central administrative office in another Member State), probably encounter a degree of distrust from creditors and banks.\n Finally, some modifications of the Corporation Tax Law, approved by the Royal Decree Law 4/2004 of March 5,74 are made in order to adapt its provisions to the new accounting framework.
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