Bulk segregant linkage mapping for rodent and human malaria parasites

Abstract

In 2005 Richard Carter's group surprised the malaria genetics community with an elegant approach to rapidly mapping the genetic basis of phenotypic traits in rodent malaria parasites. This approach, which he termed "linkage group selection", utilized bulk pools of progeny, rather than individual clones, and exploited simple selection schemes to identify genome regions underlying resistance to drug treatment (or other phenotypes). This work was the first application of "bulk segregant" methodologies for genetic mapping in microbes: this approach is now widely used in yeast, and across multiple recombining pathogens ranging from Aspergillus fungi to Schistosome parasites. Genetic crosses of human malaria parasites (for which Richard Carter was also a pioneer) can now be conducted in humanized mice, providing new opportunities for exploiting bulk segregant approaches for a wide variety of malaria parasite traits. We review the application of bulk segregant approaches to mapping malaria parasite traits and suggest additional developments that may further expand the utility of this powerful approach.

Keywords: Bulk segregant analysis; Genetic crosses; Genotype; Linkage group selection; Phenotype.

Fig. 1.

Comparison of classical linkage mapping and bulk segregant analysis. IC50, half maximal inhibitory concentration; RSA, ring-stage survival assay; LOD score, logarithm of the odds.

Fig. 2.

A brief history of bulk segregant analysis (BSA). Green boxes indicate terminologies used in different organisms, such as linkage group selection (LGS) in rodent malaria parasites and extreme-QTL mapping (X-QTL) in yeast; and blue boxes are representative analysis approaches. Arrows mark the publication year of each BSA method.

Fig. 3.

QTL mapping and gene identification using bulk segregant analysis. A, outcome of competition between 3D7 and NHP4026 under different culture conditions. Culture conditions: AlbuMAX, medium contained only AlbuMAX; Ser:Alb, medium contained serum and AlbuMAX at a 50:50 ratio; Serum, culture medium contained only serum. B, genetic cross and segregant pools. The cross was between 3D7 and NHP4026, and progeny pools were grown in different media (serum or AlbuMAX). C, comparison of 3D7 allele frequency between bulks maintained in culture contain serum and bulks maintained in AlbuMAX. D, QTL mapping by G’ approach. QTL regions are indicated with red arrows in panel C and D. Panels in Fig. 3 were adapted or modified from [77].

Fig. 4.

Future prospects for human malaria bulk segregant analysis (BSA). A, apply BSA to other selectable phenotypic traits, such as host specificity, immunity, temperatures (simulating malaria-induced fever), pH of culture media (acidosis), and parasite-host interaction. B, apply the extended BSA approach (BSR-SEq. [102], combined BSA with RNAseq) to human malaria parasites. C, combine BSA with single-cell sequencing technologies.