Development of amplicon deep sequencing markers and data analysis pipeline for genotyping multi-clonal malaria infections

摘要: Amplicon deep sequencing permits sensitive detection of minority clones and improves discriminatory power for genotyping multi-clone Plasmodium falciparum infections. New amplicon data analysis protocols are needed in epidemiological studies drug efficacy trials P. falciparum. Targeted molecular marker csp novel cpmp was conducted duplicate on mixtures parasite culture strains 37 field samples. A protocol allowing to multiplex up 384 samples a single run applied. Software “HaplotypR” developed analysis. Cpmp highly diverse (He = 0.96) contrast (He = 0.57). Minority were robustly detected if their frequency >1%. False haplotype calls owing errors observed below that threshold. To reliably detect haplotypes at very low frequencies, experiments best performed should aim coverage >10′000 reads/amplicon. When compared length polymorphic msp2, multiplexed displayed greater sensitivity detecting clones.