Quantitative genome re-sequencing defines multiple mutations conferring chloroquine resistance in rodent malaria

<p>Abstract</p> <p>Background</p> <p>Drug resistance in the malaria parasite <it>Plasmodium falciparum </it>severely compromises the treatment and control of malaria. A knowledge of the critical mutations conferring resistance to particular drugs is important in understanding modes of drug action and mechanisms of resistances. They are required to design better therapies and limit drug resistance.</p> <p>A mutation in the gene (<it>pfcrt</it>) encoding a membrane transporter has been identified as a principal determinant of chloroquine resistance in <it>P</it>. <it>falciparum</it>, but we lack a full account of higher level chloroquine resistance. Furthermore, the determinants of resistance in the other major human malaria parasite, <it>P. vivax</it>, are not known. To address these questions, we investigated the genetic basis of chloroquine resistance in an isogenic lineage of rodent malaria parasite <it>P. chabaudi </it>in which high level resistance to chloroquine has been progressively selected under laboratory conditions.</p> <p>Results</p> <p>Loci containing the critical genes were mapped by Linkage Group Selection, using a genetic cross between the high-level chloroquine-resistant mutant and a genetically distinct sensitive strain. A novel high-resolution quantitative whole-genome re-sequencing approach was used to reveal three regions of selection on chr11, chr03 and chr02 that appear progressively at increasing drug doses on three chromosomes. Whole-genome sequencing of the chloroquine-resistant parent identified just four point mutations in different genes on these chromosomes. Three mutations are located at the foci of the selection valleys and are therefore predicted to confer different levels of chloroquine resistance. The critical mutation conferring the first level of chloroquine resistance is found in <it>aat1</it>, a putative aminoacid transporter.</p> <p>Conclusions</p> <p>Quantitative trait loci conferring selectable phenotypes, such as drug resistance, can be mapped directly using progressive genome-wide linkage group selection. Quantitative genome-wide short-read genome resequencing can be used to reveal these signatures of drug selection at high resolution. The identities of three genes (and mutations within them) conferring different levels of chloroquine resistance generate insights regarding the genetic architecture and mechanisms of resistance to chloroquine and other drugs. Importantly, their orthologues may now be evaluated for critical or accessory roles in chloroquine resistance in human malarias <it>P. vivax </it>and <it>P. falciparum</it>.</p

Experimental evolution, genetic analysis and genome re-sequencing reveal the mutation conferring artemisinin resistance in an isogenic lineage of malaria parasites

<p>Abstract</p> <p>Background</p> <p>Classical and quantitative linkage analyses of genetic crosses have traditionally been used to map genes of interest, such as those conferring chloroquine or quinine resistance in malaria parasites. Next-generation sequencing technologies now present the possibility of determining genome-wide genetic variation at single base-pair resolution. Here, we combine <it>in vivo </it>experimental evolution, a rapid genetic strategy and whole genome re-sequencing to identify the precise genetic basis of artemisinin resistance in a lineage of the rodent malaria parasite, <it>Plasmodium chabaudi</it>. Such genetic markers will further the investigation of resistance and its control in natural infections of the human malaria, <it>P. falciparum</it>.</p> <p>Results</p> <p>A lineage of isogenic <it>in vivo </it>drug-selected mutant <it>P. chabaudi </it>parasites was investigated. By measuring the artemisinin responses of these clones, the appearance of an <it>in vivo </it>artemisinin resistance phenotype within the lineage was defined. The underlying genetic locus was mapped to a region of chromosome 2 by Linkage Group Selection in two different genetic crosses. Whole-genome deep coverage short-read re-sequencing (Illumina^®Solexa) defined the point mutations, insertions, deletions and copy-number variations arising in the lineage. Eight point mutations arise within the mutant lineage, only one of which appears on chromosome 2. This missense mutation arises contemporaneously with artemisinin resistance and maps to a gene encoding a de-ubiquitinating enzyme.</p> <p>Conclusions</p> <p>This integrated approach facilitates the rapid identification of mutations conferring selectable phenotypes, without prior knowledge of biological and molecular mechanisms. For malaria, this model can identify candidate genes before resistant parasites are commonly observed in natural human malaria populations.</p

Contribution a l'etude de l'extraction d'une huile aromatique et de la rectification du miscella

SIGLEAvailable from INIST (FR), Document Supply Service, under shelf-number : T 82943 / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Methanogenic octadecene degradation by syntrophic enrichment culture from brackish sediments

A microbial enrichment culture from brackish sediments was able to grow on octadec-1-ene (an unsaturated aliphatic hydrocarbon) as sole source of carbon and energy, under methanogenic conditions. Octadecene degradation is stopped either when bromoethanesulfonic acid, a selective inhibitor of methanogenesis is introduced, or when hydrogen is introduced. In the presence of bromoethanesulfonic acid, the degradation is restored by the addition of a hydrogenotrophic sulfate-reducing microorganism with sulfate. Results of molecular biodiversity, which revealed the presence of bacteria as well as of acetoclastic and hydrogenotrophic methanogens, are consistent with a syntrophic degradation involving Bacteria and Archaea. This is the first demonstration of syntrophic alkene degradation by microbial communities, showing that syntrophy is more widespread than we could have thought so far. These results highlight the need for a better understanding of microbial interactions and their role in the organic-matter degradation in polluted environments