Optimizing bulk segregant analysis of drug resistance using Plasmodium falciparum genetic crosses conducted in humanized mice
Katelyn Vendrely Brenneman,
Xue Li,
Sudhir Kumar,
Elizabeth Delgado,
Lisa A. Checkley,
Douglas A. Shoue,
Ann Reyes,
Biley A. Abatiyow,
Meseret T. Haile,
Rupam Tripura,
Tom Peto,
Dysoley Lek,
Katrina A. Button-Simons,
Stefan H.I. Kappe,
Mehul Dhorda,
François Nosten,
Standwell C. Nkhoma,
Ian H. Cheeseman,
Ashley M. Vaughan,
Michael T. Ferdig,
Tim J.C. Anderson
Affiliations
Katelyn Vendrely Brenneman
Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, USA
Xue Li
Program in Disease Intervention and Prevention, Texas Biomedical Research Institute, San Antonio, TX, USA
Sudhir Kumar
Center for Global Infectious Disease Research, Seattle Children’s Research Institute, Seattle, WA, USA
Elizabeth Delgado
Program in Disease Intervention and Prevention, Texas Biomedical Research Institute, San Antonio, TX, USA
Lisa A. Checkley
Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, USA
Douglas A. Shoue
Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, USA
Ann Reyes
Program in Disease Intervention and Prevention, Texas Biomedical Research Institute, San Antonio, TX, USA
Biley A. Abatiyow
Center for Global Infectious Disease Research, Seattle Children’s Research Institute, Seattle, WA, USA
Meseret T. Haile
Center for Global Infectious Disease Research, Seattle Children’s Research Institute, Seattle, WA, USA
Rupam Tripura
Mahidol-Oxford Tropical Medicine Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand; Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine Research Building, University of Oxford Old Road Campus, Oxford, UK
Tom Peto
Mahidol-Oxford Tropical Medicine Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand; Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine Research Building, University of Oxford Old Road Campus, Oxford, UK
Dysoley Lek
National Center for Parasitology, Entomology and Malaria Control, Phnom Penh, Cambodia; School of Public Health, National Institute of Public Health, Phnom Penh, Cambodia
Katrina A. Button-Simons
Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, USA
Stefan H.I. Kappe
Center for Global Infectious Disease Research, Seattle Children’s Research Institute, Seattle, WA, USA; Department of Pediatrics, University of Washington, Seattle, WA, USA
Mehul Dhorda
Mahidol-Oxford Tropical Medicine Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand; Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine Research Building, University of Oxford Old Road Campus, Oxford, UK
François Nosten
Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine Research Building, University of Oxford Old Road Campus, Oxford, UK; Shoklo Malaria Research Unit, Mahidol-Oxford Tropical Medicine Research Unit, Faculty of Tropical Medicine, Mahidol University, Mae Sot, Thailand
Standwell C. Nkhoma
BEI Resources, American Type Culture Collection (ATCC), Manassas, VA, USA
Ian H. Cheeseman
Program in Host Pathogen Interactions, Texas Biomedical Research Institute, San Antonio, TX, USA
Ashley M. Vaughan
Center for Global Infectious Disease Research, Seattle Children’s Research Institute, Seattle, WA, USA; Department of Pediatrics, University of Washington, Seattle, WA, USA; Corresponding author
Michael T. Ferdig
Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, USA; Corresponding author
Tim J.C. Anderson
Program in Disease Intervention and Prevention, Texas Biomedical Research Institute, San Antonio, TX, USA; Corresponding author
Summary: Classical malaria parasite genetic crosses involve isolation, genotyping, and phenotyping of progeny parasites, which is time consuming and laborious. We tested a rapid alternative approach—bulk segregant analysis (BSA)—that utilizes sequencing of bulk progeny populations with and without drug selection for rapid identification of drug resistance loci. We used dihydroartemisinin (DHA) selection in two genetic crosses and investigated how synchronization, cryopreservation, and the drug selection regimen impacted BSA success. We detected a robust quantitative trait locus (QTL) at kelch13 in both crosses but did not detect QTLs at four other candidate loci. QTLs were detected using synchronized, but not unsynchronized progeny pools, consistent with the stage-specific action of DHA. We also successfully applied BSA to cryopreserved progeny pools, expanding the utility of this approach. We conclude that BSA provides a powerful approach for investigating the genetic architecture of drug resistance in Plasmodium falciparum.
