Draft genome of the Leptospira interrogans strains, Acegua, RCA, Prea, and Capivara, obtained from wildlife maintenance hosts and infected domestic animals

In the present paper, we announce new draft genomes of four Leptospira interrogans strains named Acegua, RCA, Prea, and Capivara. These strains were isolated in the state of Rio Grande do Sul, Brazil, from cattle, dog, Brazilian guinea pig, and capybara, respectively.

Assessing the mitochondrial DNA diversity of the Chagas disease vector Triatoma sordida (Hemiptera: Reduviidae)

Triatoma sordida is a species that transmits Trypanosoma cruzi to humans. In Brazil, T. sordida currently deserves special attention because of its wide distribution, tendency to invade domestic environments and vectorial competence. For the planning and execution of control protocols to be effective against Triatominae, they must consider its population structure. In this context, this study aimed to characterise the genetic variability of T. sordida populations collected in areas with persistent infestations from Minas Gerais, Brazil. Levels of genetic variation and population structure were determined in peridomestic T. sordida by sequencing a polymorphic region of the mitochondrial cytochrome b gene. Low nucleotide and haplotype diversity were observed for all 14 sampled areas; π values ranged from 0.002-0.006. Most obtained haplotypes occurred at low frequencies, and some were exclusive to only one of the studied populations. Interpopulation genetic diversity analysis revealed strong genetic structuring. Furthermore, the genetic variability of Brazilian populations is small compared to that of Argentinean and Bolivian specimens. The possible factors related to the reduced genetic variability and strong genetic structuring obtained for studied populations are discussed in this paper.

Complete genome of the multidrug-resistant Acinetobacter baumannii strain KBN10P02143 isolated from Korea

Acinetobacter baumannii, a strictly aerobic, non-fermentative, Gram-negative coccobacillary rod-shaped bacterium, is an opportunistic pathogen in humans. We recently isolated a multidrug-resistant A. baumannii strain KBN10P02143 from the pus sample drawn from a surgical patient in South Korea. We report the complete genome of this strain, which consists of 4,139,396 bp (G + C content, 39.08%) with 3,868 protein-coding genes, 73 tRNAs and six rRNA operons. Identification of the genes related to multidrug resistance from this genome and the discovery of a novel conjugative plasmid will increase our understanding of the pathogenicity associated with this species.

Genome-wide association study identifies variants associated with progression of liver fibrosis from HCV infection.

Polymorphisms in IL28B were shown to affect clearance of hepatitis C virus (HCV) infection in genome-wide association (GWA) studies. Only a fraction of patients with chronic HCV infection develop liver fibrosis, a process that might also be affected by genetic factors. We performed a 2-stage GWA study of liver fibrosis progression related to HCV infection. We studied well-characterized HCV-infected patients of European descent who underwent liver biopsies before treatment. We defined various liver fibrosis phenotypes on the basis of METAVIR scores, with and without taking the duration of HCV infection into account. Our GWA analyses were conducted on a filtered primary cohort of 1161 patients using 780,650 single nucleotide polymorphisms (SNPs). We genotyped 96 SNPs with P values <5 × 10(-5) from an independent replication cohort of 962 patients. We then assessed the most interesting replicated SNPs using DNA samples collected from 219 patients who participated in separate GWA studies of HCV clearance. In the combined cohort of 2342 HCV-infected patients, the SNPs rs16851720 (in the total sample) and rs4374383 (in patients who received blood transfusions) were associated with fibrosis progression (P(combined) = 8.9 × 10(-9) and 1.1 × 10(-9), respectively). The SNP rs16851720 is located within RNF7, which encodes an antioxidant that protects against apoptosis. The SNP rs4374383, together with another replicated SNP, rs9380516 (P(combined) = 5.4 × 10(-7)), were linked to the functionally related genes MERTK and TULP1, which encode factors involved in phagocytosis of apoptotic cells by macrophages. Our GWA study identified several susceptibility loci for HCV-induced liver fibrosis; these were linked to genes that regulate apoptosis. Apoptotic control might therefore be involved in liver fibrosis.

Hepatitis C virus-linked mitochondrial dysfunction promotes hypoxia-inducible factor 1 alpha-mediated glycolytic adaptation.

Hepatitis C virus (HCV) infection induces a state of oxidative stress by affecting mitochondrial-respiratory-chain activity. By using cell lines inducibly expressing different HCV constructs, we showed previously that viral-protein expression leads to severe impairment of mitochondrial oxidative phosphorylation and to major reliance on nonoxidative glucose metabolism. However, the bioenergetic competence of the induced cells was not compromised, indicating an efficient prosurvival adaptive response. Here, we show that HCV protein expression activates hypoxia-inducible factor 1 (HIF-1) by normoxic stabilization of its alpha subunit. In consequence, expression of HIF-controlled genes, including those coding for glycolytic enzymes, was significantly upregulated. Similar expression of HIF-controlled genes was observed in cell lines inducibly expressing subgenomic HCV constructs encoding either structural or nonstructural viral proteins. Stabilization and transcriptional activation of HIF-1alpha was confirmed in Huh-7.5 cells harboring cell culture-derived infectious HCV and in liver biopsy specimens from patients with chronic hepatitis C. The HCV-related HIF-1alpha stabilization was insensitive to antioxidant treatment. Mimicking an impairment of mitochondrial oxidative phosphorylation by treatment of inducible cell lines with oligomycin resulted in stabilization of HIF-1alpha. Similar results were obtained by treatment with pyruvate, indicating that accumulation of intermediate metabolites is sufficient to stabilize HIF-1alpha. These observations provide new insights into the pathogenesis of chronic hepatitis C and, possibly, the HCV-related development of hepatocellular carcinoma.

Genome-wide association study identifies two loci strongly affecting transferrin glycosylation.

Polysaccharide sidechains attached to proteins play important roles in cell-cell and receptor-ligand interactions. Variation in the carbohydrate component has been extensively studied for the iron transport protein transferrin, because serum levels of the transferrin isoforms asialotransferrin + disialotransferrin (carbohydrate-deficient transferrin, CDT) are used as biomarkers of excessive alcohol intake. We conducted a genome-wide association study to assess whether genetic factors affect CDT concentration in serum. CDT was measured in three population-based studies: one in Switzerland (CoLaus study, n = 5181) and two in Australia (n = 1509, n = 775). The first cohort was used as the discovery panel and the latter ones served as replication. Genome-wide single-nucleotide polymorphism (SNP) typing data were used to identify loci with significant associations with CDT as a percentage of total transferrin (CDT%). The top three SNPs in the discovery panel (rs2749097 near PGM1 on chromosome 1, and missense polymorphisms rs1049296, rs1799899 in TF on chromosome 3) were successfully replicated , yielding genome-wide significant combined association with CDT% (P = 1.9 × 10(-9), 4 × 10(-39), 5.5 × 10(-43), respectively) and explain 5.8% of the variation in CDT%. These allelic effects are postulated to be caused by variation in availability of glucose-1-phosphate as a precursor of the glycan (PGM1), and variation in transferrin (TF) structure.

Genetic relationships between suicide attempts, suicidal ideation and major psychiatric disorders: A genome-wide association and polygenic scoring study.

Epidemiological studies have recognized a genetic diathesis for suicidal behavior, which is independent of other psychiatric disorders. Genome-wide association studies (GWAS) on suicide attempt (SA) and ideation have failed to identify specific genetic variants. Here, we conduct further GWAS and for the first time, use polygenic score analysis in cohorts of patients with mood disorders, to test for common genetic variants for mood disorders and suicide phenotypes. Genome-wide studies for SA were conducted in the RADIANT and GSK-Munich recurrent depression samples and London Bipolar Affective Disorder Case-Control Study (BACCs) then meta-analysis was performed. A GWAS on suicidal ideation during antidepressant treatment had previously been conducted in the Genome Based Therapeutic Drugs for Depression (GENDEP) study. We derived polygenic scores from each sample and tested their ability to predict SA in the mood disorder cohorts or ideation status in the GENDEP study. Polygenic scores for major depressive disorder, bipolar disorder and schizophrenia from the Psychiatric Genomics Consortium were used to investigate pleiotropy between psychiatric disorders and suicide phenotypes. No significant evidence for association was detected at any SNP in GWAS or meta-analysis. Polygenic scores for major depressive disorder significantly predicted suicidal ideation in the GENDEP pharmacogenetics study and also predicted SA in a combined validation dataset. Polygenic scores for SA showed no predictive ability for suicidal ideation. Polygenic score analysis suggests pleiotropy between psychiatric disorders and suicidal ideation whereas the tendency to act on such thoughts may have a partially independent genetic diathesis. © 2014 Wiley Periodicals, Inc.

A NOVEL MUTATION IN BCS1L IN A PATIENT WITH AN ISOLATED MITOCHONDRIAL COMPLEX III DEFICIENCY

Background: Isolated complex III deficiencies are caused by mutations in the mitochondrial CytB gene, in the BCS1L gene coding for a CIII assembly factor and in the UQCRQ gene that codes for the ubiquinone binding protein of complex III. Objective: Description of clinical features, mitochondrial function and molecular genetic analysis in a patient with an isolated complex III deficiency. Patient: A 17 year old boy, born to consanguineous parents who presented with hypoglycemia, glycosuria, deafness, growth retardation, Fanconi Syndrome and severe lactic acidosis in the neonatal period. Methods: Activities and assembly of OXPHOS complexes were investigated spectrophotometrically and by BN-PAGE. mt-DNAwas screened for deletions. Cytochrome b (CytB) and the BCS1L gene were sequenced. Results: Isolated complex III deficiency was detected in the patient's skeletal muscle. Using BN-PAGE blotting a complex III of lower molecular weight was detected. Staining the 2D reveals a missing subunit. No mutation was detected in the mitochondrial CytB gene. Sequence analysis of BCS1L revealed a novel homozygous point mutation p.M48V. Conclusion: The patients decreased complex III activity is most likely caused by incomplete assembly of complex III due to the homozygous p. M48V mutation in the BCS1L gene.

Choice of an adequate promoter for efficient complementation in Saccharomyces cerevisiae: a case study.

Conservation of the function of open reading frames recently identified in fungal genome projects can be assessed by complementation of deletion mutants of putative Saccharomyces cerevisiae orthologs. A parallel complementation assay expressing the homologous wild type S. cerevisiae gene is generally performed as a positive control. However, we and others have found that failure of complementation can occur in this case. We investigated the specific cases of S. cerevisiae TBF1 and TIM54 essential genes. Heterologous complementation with Candida glabrata TBF1 or TIM54 gene was successful using the constitutive promoters TDH3 and TEF. In contrast, homologous complementation with S. cerevisiae TBF1 or TIM54 genes failed using these promoters, and was successful only using the natural promoters of these genes. The reduced growth rate of S. cerevisiae complemented with C. glabrata TBF1 or TIM54 suggested a diminished functionality of the heterologous proteins compared to the homologous proteins. The requirement of the homologous gene for the natural promoter was alleviated for TBF1 when complementation was assayed in the absence of sporulation and germination, and for TIM54 when two regions of the protein presumably responsible for a unique translocation pathway of the TIM54 protein into the mitochondrial membrane were deleted. Our results demonstrate that the use of different promoters may prove necessary to obtain successful complementation, with use of the natural promoter being the best approach for homologous complementation.

Mapping of a gene coding for a major late structural polypeptide on the vaccinia virus genome.

Cell-free translation of total RNA isolated from vaccinia virus-infected cells late in infection results in a complex mixture of polypeptides. A monospecific antibody directed against one of the major structural proteins of the virus particle immunoprecipitated a single polypeptide with a molecular weight of 11,000 (11K) from this mixture. Immunoprecipitation was therefore used to identify the structural polypeptide among the in vitro translation products of RNA purified by hybridization selection to restriction fragments of the vaccinia virus genome. This allowed us to map the mRNA coding for the 11K polypeptide to the extreme left-hand end of the HindIII E fragment. Detailed transcriptional mapping of this region of the genome by nuclease S1 analysis revealed the presence of a late RNA transcribed from the rightward-reading strand. Its 5' end mapped at ca. 130 base pairs to the left of the HindIII site at the junction between the HindIII F and E fragments. The map position of this RNA coincided precisely with the map position of the late message coding for the 11K polypeptide.

Unique genome replication mechanism of the archaeal virus AFV1.

The exceptional genomic content and genome organization of the Acidianus filamentous virus 1 (AFV1) that infects the hyperthermophilic archaeon Acidianus hospitalis suggest that this virus might exploit an unusual mechanism of genome replication. An analysis of replicative intermediates of the viral genome by two-dimensional (2D) agarose gel electrophoresis revealed that viral genome replication starts by the formation of a D-loop and proceeds via strand displacement replication. Characterization of replicative intermediates using dark-field electron microscopy, in combination with the 2D agarose gel electrophoresis data, suggests that recombination plays a key role in the termination of AFV1 genome replication through the formation of terminal loops. A terminal protein was found to be attached to the ends of the viral genome. The results allow us to postulate a model of genome replication that relies on recombination events for initiation and termination.

Contribution of genome-wide significant single-nucleotide polymorphisms and antiretroviral therapy to dyslipidemia in HIV-infected individuals : a longitudinal study

Rapport de synthèse :Les individus HIV-positifs constituent une population à risque pour les maladies cardiovasculaires telles que |'infarctus cardiaque ou cérébrale. Celles-ci découlent d'une formation accélérée d'athéroscIérose. Ces pathologies s'expliquent en grande partie par une dyslipidémie observée au sein de cette population et qui sont dues à des facteurs externes tels que : l'immunosuppression avancée, la virémie non-contrôlée, et les effets de la thérapie antirétrovirale. Récemment, des polymorphismes nucléotidiques simples (SNP) associés à la dyslipidémie ont été mis en évidence d'une manière globale par des Genome-Wide Association Studies (GWAS). Le but principal de cette étude est d'éva|uer et de valider |'effet cumulatif des SNP identifiés dans ces GWAS pour la dyslipidémie chez des patients HIV-positifs. De plus, |'identification des facteurs non-génétiques qui contribuent à la dyslipidémie démontrent |'importance des facteurs externes, tels que mentionnés ci- dessus, et en particulier à ceux de la thérapie antirétrovirale.Les participants de l'étude proviennent de trois groupes: 426 personnes sélectionnées pour une étude précédente, 222 personnes sélectionnées de façon arbitraire dans la "Cohorte HIV Suisse" et 103 personnes sélectionnées avec un "New-Onset Diabetes mellitus" identifiées lors d'études précédentes. Ces individus ont contribué à plus de 34'000 mesures de lipides sur une durée moyenne supérieure à 7 ans. Pour l'étude, 33 SNP identifiés dans des GWAS et 9 SNP identifiés dans d'autres études publiées dans la littérature non-couverte par des GWAS ont été repris. Le génotypage a été complété pour 745 (99.2%) des 751 participants. Pour les analyses statistiques, les thérapies antirétrovirales ont été divisées en trois groupes (favorisant peu, moyennement et fortement la dyslipidémie), et trois scores génétiques ont été créés (profil favorable, moyennement favorable, non favorable/favorisant la dyslipidémie). Dans un premier temps, l'effet sur la valeur des lipides d'un ou deux allèles variants a été analysé au moyen d'un modèle de régression pour chaque SNP en ajustant le modèle pour les variables non- génétiques. Dans un deuxième temps, les SNP ayant une valeur p >= à 0.2 ont été repris dans un model Multi-SNP, ce modèle est également ajusté pour les variables non-génétiques. Puisque cette étude se base sur des SNP précédemment identifiés, celle-ci évalue uniquement l'association établie entre chaque SNP et les critères qui ont été établis au préalable, tels que : Cholestérol totale, HDL Cholestérol, non-HDL Cholestérol ou Triglycérides. Les résultats trouvés lors de |'étude confirment les résultats de la littérature. Cette étude montre que les SNP associés à la dyslipidémie doivent être analysés dans le contexte d'une thérapie antirétrovirale en tenant compte de la démographie et en considérant les valeurs du HIV (CD4+, virémie). Ces SNP montrent une tendance à prédire une dyslipidémie prolongée chez l'individu. En effet, un patient avec une thérapie antirétrovirale favorisant la dyslipidémie et un patrimoine génétique non-favorable a un risque qui est 3-f0is plus important d'avoir un Non-HDL- Cholestérol élevé, 5-fois plus important d'avoir un HDL-Cholestérol abaissé, et 4 à 5-fois plus important d'avoir une hypertriglycéridémie qu'un patient qui suit une thérapie antirétrovirale favorisant peu la dyslipidémie qui a un patrimoine génétique favorable. Vu la corrélation entre les SNP et la thérapie antirétrovirale, les cliniciens devraient intégrer les informations génétiques afin de choisir une thérapie antirétrovirale en fonction du patrimoine génétique.

Genetics for clinicians: from candidate genes to whole genome scans (technological advances).

Human genetics has progressed at an unprecedented pace during the past 10 years. DNA microarrays currently allow screening of the entire human genome with high level of coverage and we are now entering the era of high-throughput sequencing. These remarkable technical advances are influencing the way medical research is conducted and have boosted our understanding of the structure of the human genome as well as of disease biology. In this context, it is crucial for clinicians to understand the main concepts and limitations of modern genetics. This review will describe key concepts in genetics, including the different types of genetic markers in the human genome, review current methods to detect DNA variation, describe major online public databases in genetics, explain key concepts in statistical genetics and finally present commonly used study designs in clinical and epidemiological research. This review will therefore concentrate on human genetic variation analysis.

Genometric analyses of the organization of circular chromosomes: a universal pressure determines the direction of ribosomal RNA genes transcription relative to chromosome replication.

Selective pressures related to gene function and chromosomal architecture are acting on genome sequences and can be revealed, for instance, by appropriate genometric methods. Cumulative nucleotide skew analyses, i.e., GC, TA, and ORF orientation skews, predict the location of the origin of DNA replication for 88 out of 100 completely sequenced bacterial chromosomes. These methods appear fully reliable for proteobacteria, Gram-positives, and spirochetes as well as for euryarchaeotes. Based on this genome architecture information, coorientation analyses reveal that in prokaryotes, ribosomal RNA (rRNA) genes encoding the small and large ribosomal subunits are all transcribed in the same direction as DNA replication; that is, they are located along the leading strand. This result offers a simple and reliable method for circumscribing the region containing the origin of the DNA replication and reveals a strong selective pressure acting on the orientation of rRNA genes similar to the weaker one acting on the orientation of ORFs. Rate of coorientation of transfer RNA (tRNA) genes with DNA replication appears to be taxon-specific. Analyzing nucleotide biases such as GC and TA skews of genes and plotting one against the other reveals a taxonomic clusterization of species. All ribosomal RNA genes are enriched in Gs and depleted in Cs, the only so far known exception being the rRNA genes of deuterostomian mitochondria. However, this exception can be explained by the fact that in the chromosome of the human mitochondrion, the model of the deuterostomian organelle genome, DNA replication, and rRNA transcription proceed in opposite directions. A general rule is deduced from prokaryotic and mitochondrial genomes: ribosomal RNA genes that are transcribed in the same direction as the DNA replication are enriched in Gs, and those transcribed in the opposite direction are depleted in Gs.

Evidence for polygenic adaptation to pathogens in the human genome.

Most approaches aiming at finding genes involved in adaptive events have focused on the detection of outlier loci, which resulted in the discovery of individually "significant" genes with strong effects. However, a collection of small effect mutations could have a large effect on a given biological pathway that includes many genes, and such a polygenic mode of adaptation has not been systematically investigated in humans. We propose here to evidence polygenic selection by detecting signals of adaptation at the pathway or gene set level instead of analyzing single independent genes. Using a gene-set enrichment test to identify genome-wide signals of adaptation among human populations, we find that most pathways globally enriched for signals of positive selection are either directly or indirectly involved in immune response. We also find evidence for long-distance genotypic linkage disequilibrium, suggesting functional epistatic interactions between members of the same pathway. Our results show that past interactions with pathogens have elicited widespread and coordinated genomic responses, and suggest that adaptation to pathogens can be considered as a primary example of polygenic selection.