Whole Genome Sequencing of Field Isolates Reveals Extensive Genetic Diversity in <i>Plasmodium vivax</i> from Colombia

<div><p><i>Plasmodium vivax</i> is the most prevalent malarial species in South America and exerts a substantial burden on the populations it affects. The control and eventual elimination of <i>P. vivax</i> are global health priorities. Genomic research contributes to this objective by improving our understanding of the biology of <i>P. vivax</i> and through the development of new genetic markers that can be used to monitor efforts to reduce malaria transmission. Here we analyze whole-genome data from eight field samples from a region in Cordóba, Colombia where malaria is endemic. We find considerable genetic diversity within this population, a result that contrasts with earlier studies suggesting that <i>P. vivax</i> had limited diversity in the Americas. We also identify a selective sweep around a substitution known to confer resistance to sulphadoxine-pyrimethamine (SP). This is the first observation of a selective sweep for SP resistance in this species. These results indicate that <i>P. vivax</i> has been exposed to SP pressure even when the drug is not in use as a first line treatment for patients afflicted by this parasite. We identify multiple non-synonymous substitutions in three other genes known to be involved with drug resistance in <i>Plasmodium</i> species. Finally, we found extensive microsatellite polymorphisms. Using this information we developed 18 polymorphic and easy to score microsatellite loci that can be used in epidemiological investigations in South America.</p></div

A selective sweep around <i>dhps</i>.

<p>Two measures of genetic diversity (y-axis), were calculated in overlapping windows across a portion of Chromosome 14 (positions in x-axis). The dashed path under the x-axis represents the position of exons in the current annotation of the <i>P. vivax</i> reference genome. Exons of <i>dhps</i> are shaded black, all others are shaded gray. Dashed lines represent the genome-wide average of each diversity measure.</p

Summary of genomic data.

<p>Segments from outside to inside: <i>P. vivax</i> chromosomes with exonic regions shaded green; regions excluded from variant calling in this study; heatmap of <math><msub><mi>θ</mi><mi>w</mi></msub><mo>^</mo></math> in 10kb windows, high-diversity regions are darker and the maximum value is 0.0018; heatmap of <math><mi>π</mi><mo>^</mo></math> in 10kb windows; mean number of samples covered per base in 10kb windows.</p

Reference base frequency spectra.

<p>The first eight panels represent the distribution of the frequency of reference bases in sequencing reads produced from our samples across all sites in the <i>P. vivax</i> genome. In the final panel the same distribution is graphed for reads generated from Sal-I, a known single infection. Red triangles represent the frequency expected if one non-reference containing read was mapped to a site with average sequencing coverage for that sample (given in parentheses after the sample name). The majority of sites from all samples contain only non-reference bases; these sites were removed to allow clearer visualization of these distributions.</p

Allele sharing among <i>P. vivax</i> populations.

<p>Note: Ellipses for Peruvian [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0004252#pntd.0004252.ref018" target="_blank">18</a>] Cambodian and Madagascan [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0004252#pntd.0004252.ref031" target="_blank">31</a>] populations include only those SNVs reported in the cited studies and not included in a region that was excluded in our study.</p

Table_4_Transcriptional programming of immunoregulatory responses in human Langerhans cells.csv

Human epidermal Langerhans cells (LCs) maintain immune homeostasis in the skin. To examine transcriptional programming of human primary LCs during homeostasis, we performed scRNA-seq analysis of LCs before and after migration from the epidermis, coupled with functional assessment of their regulatory T cell priming capabilities. The analysis revealed that steady-state LCs exist in a continuum of maturation states and upregulate antigen presentation genes along with an immunoregulatory module including the genes IDO1, LGALS1, LAMTOR1, IL4I, upon their migration. The migration-induced transition in genomic state is accompanied by the ability of LCs to more efficiently prime regulatory T cell responses in co-culture assays. Computational analyses of the scRNAseq datasets using SCENIC and Partial Information Decomposition in Context identified a set of migration-induced transcription factors including IRF4, KLF6 and RelB as key nodes within a immunoregulatory gene regulatory network. These findings support a model in which efficient priming of immunoregulatory responses by LCs is dependent on coordinated upregulation of a migration-coupled maturation program with a immunoregulation-promoting genomic module.</p

Table_6_Transcriptional programming of immunoregulatory responses in human Langerhans cells.csv

Human epidermal Langerhans cells (LCs) maintain immune homeostasis in the skin. To examine transcriptional programming of human primary LCs during homeostasis, we performed scRNA-seq analysis of LCs before and after migration from the epidermis, coupled with functional assessment of their regulatory T cell priming capabilities. The analysis revealed that steady-state LCs exist in a continuum of maturation states and upregulate antigen presentation genes along with an immunoregulatory module including the genes IDO1, LGALS1, LAMTOR1, IL4I, upon their migration. The migration-induced transition in genomic state is accompanied by the ability of LCs to more efficiently prime regulatory T cell responses in co-culture assays. Computational analyses of the scRNAseq datasets using SCENIC and Partial Information Decomposition in Context identified a set of migration-induced transcription factors including IRF4, KLF6 and RelB as key nodes within a immunoregulatory gene regulatory network. These findings support a model in which efficient priming of immunoregulatory responses by LCs is dependent on coordinated upregulation of a migration-coupled maturation program with a immunoregulation-promoting genomic module.</p

Table_5_Transcriptional programming of immunoregulatory responses in human Langerhans cells.csv

Human epidermal Langerhans cells (LCs) maintain immune homeostasis in the skin. To examine transcriptional programming of human primary LCs during homeostasis, we performed scRNA-seq analysis of LCs before and after migration from the epidermis, coupled with functional assessment of their regulatory T cell priming capabilities. The analysis revealed that steady-state LCs exist in a continuum of maturation states and upregulate antigen presentation genes along with an immunoregulatory module including the genes IDO1, LGALS1, LAMTOR1, IL4I, upon their migration. The migration-induced transition in genomic state is accompanied by the ability of LCs to more efficiently prime regulatory T cell responses in co-culture assays. Computational analyses of the scRNAseq datasets using SCENIC and Partial Information Decomposition in Context identified a set of migration-induced transcription factors including IRF4, KLF6 and RelB as key nodes within a immunoregulatory gene regulatory network. These findings support a model in which efficient priming of immunoregulatory responses by LCs is dependent on coordinated upregulation of a migration-coupled maturation program with a immunoregulation-promoting genomic module.</p

Table_3_Transcriptional programming of immunoregulatory responses in human Langerhans cells.csv

Human epidermal Langerhans cells (LCs) maintain immune homeostasis in the skin. To examine transcriptional programming of human primary LCs during homeostasis, we performed scRNA-seq analysis of LCs before and after migration from the epidermis, coupled with functional assessment of their regulatory T cell priming capabilities. The analysis revealed that steady-state LCs exist in a continuum of maturation states and upregulate antigen presentation genes along with an immunoregulatory module including the genes IDO1, LGALS1, LAMTOR1, IL4I, upon their migration. The migration-induced transition in genomic state is accompanied by the ability of LCs to more efficiently prime regulatory T cell responses in co-culture assays. Computational analyses of the scRNAseq datasets using SCENIC and Partial Information Decomposition in Context identified a set of migration-induced transcription factors including IRF4, KLF6 and RelB as key nodes within a immunoregulatory gene regulatory network. These findings support a model in which efficient priming of immunoregulatory responses by LCs is dependent on coordinated upregulation of a migration-coupled maturation program with a immunoregulation-promoting genomic module.</p

DataSheet_1_Transcriptional programming of immunoregulatory responses in human Langerhans cells.pdf

Human epidermal Langerhans cells (LCs) maintain immune homeostasis in the skin. To examine transcriptional programming of human primary LCs during homeostasis, we performed scRNA-seq analysis of LCs before and after migration from the epidermis, coupled with functional assessment of their regulatory T cell priming capabilities. The analysis revealed that steady-state LCs exist in a continuum of maturation states and upregulate antigen presentation genes along with an immunoregulatory module including the genes IDO1, LGALS1, LAMTOR1, IL4I, upon their migration. The migration-induced transition in genomic state is accompanied by the ability of LCs to more efficiently prime regulatory T cell responses in co-culture assays. Computational analyses of the scRNAseq datasets using SCENIC and Partial Information Decomposition in Context identified a set of migration-induced transcription factors including IRF4, KLF6 and RelB as key nodes within a immunoregulatory gene regulatory network. These findings support a model in which efficient priming of immunoregulatory responses by LCs is dependent on coordinated upregulation of a migration-coupled maturation program with a immunoregulation-promoting genomic module.</p