Clonal outbreak of Plasmodium falciparum infection in eastern Panama

Abstract

Identifying the source of resurgent parasites is paramount to a strategic, successful intervention for malaria elimination. Although the malaria incidence in Panama is low, a recent outbreak resulted in a 6-fold increase in reported cases. We hypothesized that parasites sampled from this epidemic might be related and exhibit a clonal population structure. We tested the genetic relatedness of parasites, using informative single-nucleotide polymorphisms and drug resistance loci. We found that parasites were clustered into 3 clonal subpopulations and were related to parasites from Colombia. Two clusters of Panamanian parasites shared identical drug resistance haplotypes, and all clusters shared a chloroquine-resistance genotype matching the pfcrt haplotype of Colombian origin. Our findings suggest these resurgent parasite populations are highly clonal and that the high clonality likely resulted from epidemic expansion of imported or vestigial cases. Malaria outbreak investigations that use genetic tools can illuminate potential sources of epidemic malaria and guide strategies to prevent further resurgence in areas where malaria has been eliminated.

Keywords: Plasmodium falciparum; drug resistance; molecular surveillance; outbreak, epidemic; tropical diseases.

Figure 1.

Study sites in Panama and Colombia where Plasmodium falciparum isolates were collected during 2003–2008. Sites are color coded by clonal groups identified by molecular barcoding (group I = blue, group II = green, group III = red, and Colombian = orange), and province and country boundaries are denoted by gray lines. The diameter of each circle corresponds to the number of samples collected at each site. The base map for this figure was generated with Python 2.7.3, using shapefiles (http://www.diva-gis.org/gdata).

Figure 2.

Epidemiologic and genotyping data for Panamanian and Colombian isolates. Isolate sample names are color coded by collection site (left: Darien = purple, Panamá = green, and Kuna Yala = blue). Epidemiologic data are shown with genotypes for each sample. Groups I (blue), II (green), and III (red) and the Colombian group (orange) are indicated on the right.

Figure 3.

Membership fraction plot of eastern Panamanian and Colombian subpopulations. Structure analysis showing K = 2 (A) and K = 5 (B) clustering. The genotype of each sample is represented by a single vertical line partitioned into segments in proportion of the estimated membership in the different subpopulations. The uppermost level of structure among Panamanian isolates (ie, the most likely number of clusters [K = 2]; A) and the second-most-likely number of clusters (K = 5; B) are shown. The sample name is color coded by region (Darien = purple, Panamá = green, Kuna Yala = blue, coastal = red, and Colombia = orange).

Figure 4.

Maximum clade credibility tree of Plasmodium falciparum isolates from eastern Panama and Colombia. Isolates are color coded at terminal branch arms on the basis of study site. Groups I (blue), II (green), and III (red) and the Colombian group (orange) are distinguished by triangles. Branch nodes are labeled by the median height at each node for recalculated genetic distances, based on 10 000 bootstrap replicates.

Figure 5.

Two-dimensional plots of principal components analysis (PCA) of Panamanian Plasmodium falciparum isolates are shown, using 24 alleles of the molecular barcode for the Panamanian and Colombian isolates alone (A), with PC1 explaining 39% of the variance and PC2 explaining 18% of the variance, or with the Senegalese outgroup (B), with PC1 explaining 26% of the variance and PC2 explaining 14% of the variance. Samples are color coded by group and region (group I = blue, group II = green, group III = red, the Colombian group = orange, and the Senegalese group = gray).