Comparative Analysis of Field-Isolate and Monkey-Adapted <i>Plasmodium vivax</i> Genomes

<div><p>Significant insights into the biology of <i>Plasmodium vivax</i> have been gained from the ability to successfully adapt human infections to non-human primates. <i>P</i>. <i>vivax</i> strains grown in monkeys serve as a renewable source of parasites for <i>in vitro</i> and <i>ex vivo</i> experimental studies and functional assays, or for studying <i>in vivo</i> the relapse characteristics, mosquito species compatibilities, drug susceptibility profiles or immune responses towards potential vaccine candidates. Despite the importance of these studies, little is known as to how adaptation to a different host species may influence the genome of <i>P</i>. <i>vivax</i>. In addition, it is unclear whether these monkey-adapted strains consist of a single clonal population of parasites or if they retain the multiclonal complexity commonly observed in field isolates. Here we compare the genome sequences of seven <i>P</i>. <i>vivax</i> strains adapted to New World monkeys with those of six human clinical isolates collected directly in the field. We show that the adaptation of <i>P</i>. <i>vivax</i> parasites to monkey hosts, and their subsequent propagation, did not result in significant modifications of their genome sequence and that these monkey-adapted strains recapitulate the genomic diversity of field isolates. Our analyses also reveal that these strains are not always genetically homogeneous and should be analyzed cautiously. Overall, our study provides a framework to better leverage this important research material and fully utilize this resource for improving our understanding of <i>P</i>. <i>vivax</i> biology.</p></div

Genomic relationships among <i>P</i>. <i>vivax</i> isolates.

<p>Principal component analysis based on 81,328 SNVs in four field isolates (C127, C08, M08 and M15) and seven monkey-adapted strains (Salvador-I, Brazil-I, Belem, Chesson, North Korea, India-VII and Mauritania-I) for which an entire haploid genome sequence could be reconstructed. The sample names are colored by their geographic origin: blue for Central and South America, green for Asia and red for Africa.</p

Genomic distribution of the nucleotide differences between the clones present in the Mauritania-I sample.

<p>Each grey bar represents one single nucleotide difference between the two clones detected in the Mauritania-I genome sequence data and is displayed according to its position (x-axis, in bp) along one of the <i>P</i>. <i>vivax</i> chromosome (from chromosome 1 on top to chromosome 14 at the bottom). Note that 1,969 out of the 2,255 nucleotides differences (87%) between the two clones were clustered in 153 regions accounting for 3.78 Mb (or 20% of the genome).</p

Shared telomeric deletion.

<p>The figure shows a ~120 kb deletion indicated by the decrease in sequence coverage (y-axis, in reads per bp) at the telomeric end of chromosome 7 (x-axis in 1,000 bp). The sequence coverage is displayed, from top to bottom, for three monkey-adapted strains (Belem, Brazil-I and North Korea) and one Cambodian field isolate (C15). The bottom track shows the variation in GC content along this region. The lower coverage in North Korea and C15 indicates that only some of the parasites carry the deletion. Note also that the deletion boundary is different in different samples.</p

Distribution of the Reference Allele Frequency (RAF) in monkey-adapted strains sequenced to date.

<p>The graph shows the number of variable positions (y-axis) in a given sample according to the proportion of reads carrying the reference (i.e., Salvador I) allele (x-axis, in %). For most monkey-adapted strains the distribution is U shaped consistent with the present of a single haploid clone. However, the RAF distributions for Mauritania-I (in dotted red) and Chesson (in dotted green) indicate the presence of a second clone. The RAF for the human isolates mentioned in the manuscript is presented in <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003566#pntd.0003566.s003" target="_blank">S1 Fig.</a></p

Complexity of infection in the Mauritania-I and Mauritania-II samples.

<p>The top panel shows the passage history of the Mauritania-I (starting with the initial patient infection of February 12, 1995) and Mauritania-II (starting with the October 2 relapse) strains (adapted from Collins et al., 1998). The six samples analyzed in this study are indicated in red. Solid black lines represent infections propagated in either <i>Aotus nancymaae</i> (AI, AO and WR) or <i>Saimiri boliviensis boliviensis</i> (SI) monkeys through injections of infected erythrocytes. Dashed lines represent passage through mosquitoes and propagation by sporozoites. The lower panel shows genotypes of the different clones present in each of the Mauritania samples analyzed (all samples are monkey-adapted strains but the “Patient Relapse” which is a clinical sample). The height of each allele represents its relative frequency in each sample and the alleles are organized based on the haplotypes inferred for each clone. Note that the allele frequencies in WR-1714 likely indicate the presence of one or more supplementary clones in addition to P1, P2 and P3.</p

Maternal Nutrition Induces Pervasive Gene Expression Changes but No Detectable DNA Methylation Differences in the Liver of Adult Offspring

<div><p>Aims</p><p>Epidemiological and animal studies have shown that maternal diet can influence metabolism in adult offspring. However, the molecular mechanisms underlying these changes remain poorly understood. Here, we characterize the phenotypes induced by maternal obesity in a mouse model and examine gene expression and epigenetic changes induced by maternal diet in adult offspring.</p><p>Methods</p><p>We analyzed genetically identical male mice born from dams fed a high- or low-fat diet throughout pregnancy and until day 21 postpartum. After weaning, half of the males of each group were fed a high-fat diet, the other half a low-fat diet. We first characterized the genome-wide gene expression patterns of six tissues of adult offspring - liver, pancreas, white adipose, brain, muscle and heart. We then measured DNA methylation patterns in liver at selected loci and throughout the genome.</p><p>Results</p><p>Maternal diet had a significant effect on the body weight of the offspring when they were fed an obesogenic diet after weaning. Our analyses showed that maternal diet had a pervasive effect on gene expression, with a pronounced effect in liver where it affected many genes involved in inflammation, cholesterol synthesis and RXR activation. We did not detect any effect of the maternal diet on DNA methylation in the liver.</p><p>Conclusions</p><p>Overall, our findings highlighted the persistent influence of maternal diet on adult tissue regulation and suggested that the transcriptional changes were unlikely to be caused by DNA methylation differences in adult liver.</p></div

Body weight of male offspring.

<p>Average offspring weight (in g, ±SE) is presented from one week through six months of age. Maternal diet is denoted by line color (light gray: LF, black: HF) and adult diet is shown by line style (dashed: LF, solid: HF).</p

Overview of the experimental design.

<p>Dams are displayed in dark grey, offspring in light grey. LF and HF designate, respectively, low fat and high fat diet. Double-sided dashed arrows indicate pair-wise comparisons testing the effect of the diet post-weaning (on fixed maternal diet), solid arrows indicate the comparisons testing the effect of the maternal diet (on fixed post-weaning diet).</p

Influence of the maternal diet and diet after weaning on body size and metabolism regulation.

<p>LF and HF stand for, respectively, low fat and high fat diet. p Mat and p Post indicate, respectively, the p-values associated with the influence of the maternal diet and diet after weaning. The influence of the maternal diet on dams' phenotypes and offspring's body weight at weaning was assessed using a Student's t-test. For all other offspring's phenotypes, the respective influence of the maternal and post-weaning diet was determined using a two-way ANOVA. After correcting for animal weight, many of the phenotypes did not reach statistical significance for the effect of post-weaning or maternal diet.</p