Defining the role of common variation in the genomic and biological architecture of adult human height

Using genome-wide data from 253,288 individuals, we identified 697 variants at genome-wide significance that together explained one-fifth of the heritability for adult height. By testing different numbers of variants in independent studies, we show that the most strongly associated ∼2,000, ∼3,700 and ∼9,500 SNPs explained ∼21%, ∼24% and ∼29% of phenotypic variance. Furthermore, all common variants together captured 60% of heritability. The 697 variants clustered in 423 loci were enriched for genes, pathways and tissue types known to be involved in growth and together implicated genes and pathways not highlighted in earlier efforts, such as signaling by fibroblast growth factors, WNT/β-catenin and chondroitin sulfate-related genes. We identified several genes and pathways not previously connected with human skeletal growth, including mTOR, osteoglycin and binding of hyaluronic acid. Our results indicate a genetic architecture for human height that is characterized by a very large but finite number (thousands) of causal variants.2014

The humankind genome: from genetic diversity to the origin of human diseases

Genome-wide association studies have failed to establish common variant risk for the majority of common human diseases. The underlying reasons for this failure are explained by recent studies of resequencing and comparison of over 1200 human genomes and 10 000 exomes, together with the delineation of DNA methylation patterns (epigenome) and full characterization of coding and noncoding RNAs (transcriptome) being transcribed. These studies have provided the most comprehensive catalogues of functional elements and genetic variants that are now available for global integrative analysis and experimental validation in prospective cohort studies. With these datasets, researchers will have unparalleled opportunities for the alignment, mining, and testing of hypotheses for the roles of specific genetic variants, including copy number variations, single nucleotide polymorphisms, and indels as the cause of specific phenotypes and diseases. Through the use of next-generation sequencing technologies for genotyping and standardized ontological annotation to systematically analyze the effects of genomic variation on humans and model organism phenotypes, we will be able to find candidate genes and new clues for disease’s etiology and treatment. This article describes essential concepts in genetics and genomic technologies as well as the emerging computational framework to comprehensively search websites and platforms available for the analysis and interpretation of genomic data.

Genome-scale analysis of the non-cultivable Treponema pallidum reveals extensive within-patient genetic variation

Insights into the genomic adaptive traits of Treponema pallidum, the causative bacterium of syphilis, have long been hampered due to the absence of in vitro culture models and the constraints associated with its propagation in rabbits. Here, we have bypassed the culture bottleneck by means of a targeted strategy never applied to uncultivable bacterial human pathogens to directly capture whole-genome T. pallidum data in the context of human infection. This strategy has unveiled a scenario of discreet T. pallidum interstrain single-nucleotide-polymorphism-based microevolution, contrasting with a rampant within-patient genetic heterogeneity mainly targeting multiple phase-variable loci and a major antigen-coding gene (tprK). TprK demonstrated remarkable variability and redundancy, intra- and interpatient, suggesting ongoing parallel adaptive diversification during human infection. Some bacterial functions (for example, flagella- and chemotaxis-associated) were systematically targeted by both inter- and intrastrain single nucleotide polymorphisms, as well as by ongoing within-patient phase variation events. Finally, patient-derived genomes possess mutations targeting a penicillin-binding protein coding gene (mrcA) that had never been reported, unveiling it as a candidate target to investigate the impact on the susceptibility to penicillin. Our findings decode the major genetic mechanisms by which T. pallidum promotes immune evasion and survival, and demonstrate the exceptional power of characterizing evolving pathogen subpopulations during human infection.

Plasmodium vivax circumsporozoite genotypes: a limited variation or new subspecies with major biological consequences?

Background: Plasmodium vivax circumsporozoite variants have been identified in several geographical areas. The real implication of the genetic variation in this region of the P. vivax genome has been questioned for a long time. Although previous studies have observed significant association between VK210 and the Duffy blood group, we present here that evidences of this variation are limited to the CSP central portion. Methods: The phylogenetic analyses were accomplished starting from the amplification of conserved domains of 18 SSU RNAr and Cyt B. The antibodies responses against the CSP peptides, MSP-1, AMA-1 and DBP were detected by ELISA, in plasma samples of individuals infected with two P. vivax CS genotypes: VK210 and P. vivax-like. Results: These analyses of the two markers demonstrate high similarity among the P. vivax CS genotypes and surprisingly showed diversity equal to zero between VK210 and P. vivax-like, positioning these CS genotypes in the same clade. A high frequency IgG antibody against the N- and C-terminal regions of the P. vivax CSP was found as compared to the immune response to the R- and V-repetitive regions (p = 0.0005, Fisher's Exact test). This difference was more pronounced when the P. vivax-like variant was present in the infection (p = 0.003, Fisher's Exact test). A high frequency of antibody response against MSP-1 and AMA-1 peptides was observed for all P. vivax CS genotypes in comparison to the same frequency for DBP. Conclusions: This results target that the differences among the P. vivax CS variants are restrict to the central repeated region of the protein, mostly nucleotide variation with important serological consequences.

Near full-length genome analysis of low prevalent human immunodeficiency virus type 1 subclade F1 in Sao Paulo, Brazil

Background: The genetic diversity of the human immunodeficiency virus type 1 (HIV-1) is critical to lay the groundwork for the design of successful drugs or vaccine. In this study we aimed to characterize and define the molecular prevalence of HIV-1 subclade F1 currently circulating in Sao Paulo, Brazil. Methods: A total of 36 samples were selected from 888 adult patients residing in Sao Paulo who had previously been diagnosed in two independent studies in our laboratory as being infected with subclade F1 based on pol subgenomic fragment sequencing. Proviral DNA was amplified from the purified genomic DNA of all 36 blood samples by 5 fragments overlapping PCR followed by direct sequencing. Sequence data were obtained from the 5 fragments of pure subclade F1 and phylogenetic trees were constructed and compared with previously published sequences. Subclades F1 that exhibited mosaic structure with other subtypes were omitted from any further analysis Results: Our methods of fragment amplification and sequencing confirmed that only 5 sequences inferred from pol region as subclade F1 also holds true for the genome as a whole and, thus, estimated the true prevalence at 0.56%. The results also showed a single phylogenetic cluster of the Brazilian subclade F1 along with non-Brazilian South American isolates in both subgenomic and the full-length genomes analysis with an overall intrasubtype nucleotide divergence of 6.9%. The nucleotide differences within the South American and Central African F1 strains, in the C2-C3 env, were 8.5% and 12.3%, respectively. Conclusion: All together, our findings showed a surprisingly low prevalence rate of subclade F1 in Brazil and suggest that these isolates originated in Central Africa and subsequently introduced to South America.

Genetic Variation among Major Human Geographic Groups Supports a Peculiar Evolutionary Trend in PAX9

A total of 172 persons from nine South Amerindian, three African and one Eskimo populations were studied in relation to the Paired box gene 9 (PAX9) exon 3 (138 base pairs) as well as its 5' and 3' flanking intronic segments (232 bp and 220 bp, respectively) and integrated with the information available for the same genetic region from individuals of different geographical origins. Nine mutations were scored in exon 3 and six in its flanking regions; four of them are new South American tribe-specific singletons. Exon3 nucleotide diversity is several orders of magnitude higher than its intronic regions. Additionally, a set of variants in the PAX9 and 101 other genes related with dentition can define at least some dental morphological differences between Sub-Saharan Africans and non-Africans, probably associated with adaptations after the modern human exodus from Africa. Exon 3 of PAX9 could be a good molecular example of how evolvability works.

Single-nucleotide polymorphism, linkage disequilibrium and geographic structure in the malaria parasite Plasmodium vivax: prospects for genome-wide association studies

Background: The ideal malaria parasite populations for initial mapping of genomic regions contributing to phenotypes such as drug resistance and virulence, through genome-wide association studies, are those with high genetic diversity, allowing for numerous informative markers, and rare meiotic recombination, allowing for strong linkage disequilibrium (LD) between markers and phenotype-determining loci. However, levels of genetic diversity and LD in field populations of the major human malaria parasite P. vivax remain little characterized. Results: We examined single-nucleotide polymorphisms (SNPs) and LD patterns across a 100-kb chromosome segment of P. vivax in 238 field isolates from areas of low to moderate malaria endemicity in South America and Asia, where LD tends to be more extensive than in holoendemic populations, and in two monkey-adapted strains (Salvador-I, from El Salvador, and Belem, from Brazil). We found varying levels of SNP diversity and LD across populations, with the highest diversity and strongest LD in the area of lowest malaria transmission. We found several clusters of contiguous markers with rare meiotic recombination and characterized a relatively conserved haplotype structure among populations, suggesting the existence of recombination hotspots in the genome region analyzed. Both silent and nonsynonymous SNPs revealed substantial between-population differentiation, which accounted for similar to 40% of the overall genetic diversity observed. Although parasites clustered according to their continental origin, we found evidence for substructure within the Brazilian population of P. vivax. We also explored between-population differentiation patterns revealed by loci putatively affected by natural selection and found marked geographic variation in frequencies of nucleotide substitutions at the pvmdr-1 locus, putatively associated with drug resistance. Conclusion: These findings support the feasibility of genome-wide association studies in carefully selected populations of P. vivax, using relatively low densities of markers, but underscore the risk of false positives caused by population structure at both local and regional levels.

Whole-Genome Characterization of Human and Simian Immunodeficiency Virus Intrahost Diversity by Ultradeep Pyrosequencing

Rapid evolution and high intrahost sequence diversity are hallmarks of human and simian immunodeficiency virus (HIV/SIV) infection. Minor viral variants have important implications for drug resistance, receptor tropism, and immune evasion. Here, we used ultradeep pyrosequencing to sequence complete HIV/SIV genomes, detecting variants present at a frequency as low as 1%. This approach provides a more complete characterization of the viral population than is possible with conventional methods, revealing low-level drug resistance and detecting previously hidden changes in the viral population. While this work applies pyrosequencing to immunodeficiency viruses, this approach could be applied to virtually any viral pathogen.

Codon usage bias and A+T content variation in human papillomavirus genomes

We analyzed the codon usage bias of eight open reading frames (ORFs) across up to 79 human papillomavirus (HPV) genotypes from three distinct phylogenetic groups. All eight ORFs across HPV genotypes show a strong codon usage bias, amongst degenerately encoded amino acids, toward 18 codons mainly with T at the 3rd position. For all 18 degenerately encoded amino acids, codon preferences amongst human and animal PV ORFs are significantly different from those averaged across mammalian genes. Across the HPV types, the L2 ORFs show the highest codon usage bias (73.2 +/- 1.6% and the E4 ORFs the lowest (51.1 +/- 0.5%), reflecting as similar bias in codon 3rd position A + T content (L2: 76.1 +/- 4.2%; E4: 58.6 +/- 4.5%). The E4 ORF, uniquely amongst the HPV ORFs, is G + C rich, while the other ORFs are A + T rich. Codon usage bias correlates positively with A + T content at the codon 3rd position in the E2, E6, L1 and L2 ORFs, but negatively in the E4 ORFs. A general conservation of preferred codon usage across human and non-human PV genotypes whether they originate from a same supergroup or not, together with observed difference between the preferred codon usage for HPV ORFs and for genes of the cells they infect, suggests that specific codon usage bias and A + T content variation may somehow increase the replicational fitness of HPVs in mammalian epithelial cells, and have practical implications for gene therapy of HPV infection. (C) 2003 Elsevier B.V. All rights reserved.

Restriction enzyme analysis of the human cytomegalovirus genome in specimens collected from immunodeficient patients in Belém, State of Pará, Brazil

INTRODUCTION: Human cytomegalovirus is an opportunistic betaherpesvirus that causes persistent and serious infections in immunodeficient patients. Recurrent infections occur due to the presence of the virus in a latent state in some cell types. It is possible to examine the virus using molecular methods to aid in the immunological diagnosis and to generate a molecular viral profile in immunodeficient patients. The objective of this study was to characterize cytomegalovirus genotypes and to generate the epidemiological and molecular viral profile in immunodeficient patients. METHODS: A total of 105 samples were collected from immunodeficient patients from the City of Belém, including newborns, hemodialysis patients, transplant recipients and HIV+ patients. An IgG and IgM antibody study was completed using ELISA, and enzymatic analysis by restriction fragment length polymorphism (RFLP) was performed to characterize viral genotypes. RESULTS: It was observed that 100% of the patients had IgG antibodies, 87% of which were IgG+/IgM-, consistent with a prior infection profile, 13% were IgG+/IgM+, suggestive of recent infection. The newborn group had the highest frequency (27%) of the IgG+/IgM+ profile. By RFLP analysis, only one genotype was observed, gB2, which corresponded to the standard AD169 strain. CONCLUSIONS: The presence of IgM antibodies in new borns indicates that HCMV continues to be an important cause of congenital infection. The low observed genotypic diversity could be attributed to the small sample size because newborns were excluded from the RFLP analysis. This study will be continued including samples from newborns to extend the knowledge of the general and molecular epidemiology of HCMV in immunodeficient patients.

Explaining Variation in Organizational Change: The Reform of Human Resource Management in the European Commission and the OECD

In this article we investigate the reforms of human resource management in the European Commission and the OECD by analyzing comparatively to what extent both organizations have adjusted their respective structures towards the ideal type of the so-called New Public Management (NPM). The empirical findings show that reforms towards NPM are more pronounced in the Commission than in the OECD. These findings are surprising for two reasons: First, it seems rather paradoxical that the OECD as central promoter of NPM at the international level lags behind the global trend when it comes to reforming its own structures. Second, this result is in contradiction with theoretical expectations, as they can be derived from theories of institutional isomorphism. To nevertheless account for the surprising results, it is necessary to modify and complement existing theories especially with regard to the scope conditions of their causal mechanisms.

Side effects of genome structural changes.

The first extensive catalog of structural human variation was recently released. It showed that large stretches of genomic DNA that vary considerably in copy number were extremely abundant. Thus it is conceivable that they play a major role in functional variation. Consistently, genomic insertions and deletions were shown to contribute to phenotypic differences by modifying not only the expression levels of genes within the aneuploid segments but also of normal copy-number neighboring genes. In this report, we review the possible mechanisms behind this latter effect.

Plasmodium falciparum CS protein - prime malaria vaccine candidate: definition of the human CTL domain and analysis of its variation

Studies in mice have shown that immunity to malaria sporozoites is mediated primarily by citotoxic T lymphocytes (CTL) specific for epitopes within the circumsporozoite (CS) protein. Humans, had never been shown to generate CTL against any malaria or other parasite protein. The design of a sub-unit vaccine for humans ralies on the epitopes recognized by CTL being identified and polymorphisms therein being defined. We have developed a novel technique using an entire series of overlapping synthetic peptides to define the epitopes of the Plasmodium falciparum CS protein recognized by human CTL and have analyzed the sequence variation of the protein with respect to the identified CTL epitopic domain. We have demonstrated that some humans can indeed generate CTL. against the P. falciparum CS protein. Furthermore, the extent of variation observed for the CTL recognition domain is finite and the combination of peptides necessary for inclusion in a polyvalent vaccine may be small. If ways can be found to increase immune responsiveness, then a vaccine designed to stimulate CS protein-specific CTL activity may prevent malaria.

GWAS of human bitter taste perception identifies new loci and reveals additional complexity of bitter taste genetics.

Human perception of bitterness displays pronounced interindividual variation. This phenotypic variation is mirrored by equally pronounced genetic variation in the family of bitter taste receptor genes. To better understand the effects of common genetic variations on human bitter taste perception, we conducted a genome-wide association study on a discovery panel of 504 subjects and a validation panel of 104 subjects from the general population of São Paulo in Brazil. Correction for general taste-sensitivity allowed us to identify a SNP in the cluster of bitter taste receptors on chr12 (10.88- 11.24 Mb, build 36.1) significantly associated (best SNP: rs2708377, P = 5.31 × 10(-13), r(2) = 8.9%, β = -0.12, s.e. = 0.016) with the perceived bitterness of caffeine. This association overlaps with-but is statistically distinct from-the previously identified SNP rs10772420 influencing the perception of quinine bitterness that falls in the same bitter taste cluster. We replicated this association to quinine perception (P = 4.97 × 10(-37), r(2) = 23.2%, β = 0.25, s.e. = 0.020) and additionally found the effect of this genetic locus to be concentration specific with a strong impact on the perception of low, but no impact on the perception of high concentrations of quinine. Our study, thus, furthers our understanding of the complex genetic architecture of bitter taste perception.

A genome-wide association study of early menopause and the combined impact of identified variants.

Early menopause (EM) affects up to 10% of the female population, reducing reproductive lifespan considerably. Currently, it constitutes the leading cause of infertility in the western world, affecting mainly those women who postpone their first pregnancy beyond the age of 30 years. The genetic aetiology of EM is largely unknown in the majority of cases. We have undertaken a meta-analysis of genome-wide association studies (GWASs) in 3493 EM cases and 13 598 controls from 10 independent studies. No novel genetic variants were discovered, but the 17 variants previously associated with normal age at natural menopause as a quantitative trait (QT) were also associated with EM and primary ovarian insufficiency (POI). Thus, EM has a genetic aetiology which overlaps variation in normal age at menopause and is at least partly explained by the additive effects of the same polygenic variants. The combined effect of the common variants captured by the single nucleotide polymorphism arrays was estimated to account for ∼30% of the variance in EM. The association between the combined 17 variants and the risk of EM was greater than the best validated non-genetic risk factor, smoking.