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

doi:10.1016/j.isci.2022.104095

Optimizing bulk segregant analysis of drug resistance using Plasmodium falciparum genetic crosses conducted in humanized mice

iScience. 2022 Mar 16;25(4):104095. doi: 10.1016/j.isci.2022.104095. eCollection 2022 Apr 15.

Authors

Katelyn Vendrely Brenneman¹, Xue Li², Sudhir Kumar³, Elizabeth Delgado², Lisa A Checkley¹, Douglas A Shoue¹, Ann Reyes², Biley A Abatiyow³, Meseret T Haile³, Rupam Tripura^{4

5}, Tom Peto^{4

5}, Dysoley Lek^{6

7}, Katrina A Button-Simons¹, Stefan H I Kappe^{3

8}, Mehul Dhorda^{4

5}, François Nosten^{5

9}, Standwell C Nkhoma¹⁰, Ian H Cheeseman¹¹, Ashley M Vaughan^{3

8}, Michael T Ferdig¹, Tim J C Anderson²

Affiliations

¹ Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, USA.
² Program in Disease Intervention and Prevention, Texas Biomedical Research Institute, San Antonio, TX, USA.
³ Center for Global Infectious Disease Research, Seattle Children's Research Institute, Seattle, WA, USA.
⁴ Mahidol-Oxford Tropical Medicine Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand.
⁵ Centre for Tropical Medicine and Global Health, Nuffield Department of Medicine Research Building, University of Oxford Old Road Campus, Oxford, UK.
⁶ National Center for Parasitology, Entomology and Malaria Control, Phnom Penh, Cambodia.
⁷ School of Public Health, National Institute of Public Health, Phnom Penh, Cambodia.
⁸ Department of Pediatrics, University of Washington, Seattle, WA, USA.
⁹ Shoklo Malaria Research Unit, Mahidol-Oxford Tropical Medicine Research Unit, Faculty of Tropical Medicine, Mahidol University, Mae Sot, Thailand.
¹⁰ BEI Resources, American Type Culture Collection (ATCC), Manassas, VA, USA.
¹¹ Program in Host Pathogen Interactions, Texas Biomedical Research Institute, San Antonio, TX, USA.

Abstract

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.

Keywords: Biological sciences; Genomics; Parasitology.

Abstract

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