A Method for Amplicon Deep Sequencing of Drug Resistance Genes in Plasmodium falciparum Clinical Isolates from India

Author:

Rao Pavitra N.1,Uplekar Swapna1,Kayal Sriti2,Mallick Prashant K.3,Bandyopadhyay Nabamita3,Kale Sonal3,Singh Om P.3,Mohanty Akshaya4,Mohanty Sanjib5,Wassmer Samuel C.6,Carlton Jane M.1

Affiliation:

1. Center for Genomics and Systems Biology, Department of Biology, New York University, New York, New York, USA

2. National Institute of Technology, Raurkela, Odisha, India

3. National Institute of Malaria Research, Indian Council of Medical Research, Dwarka, New Delhi, India

4. Institute of Life Sciences, Bhubaneswar, Odisha, India

5. Ispat General Hospital, Raurkela, Odisha, India

6. Department of Microbiology, Division of Parasitology, New York University School of Medicine, New York, New York, USA

Abstract

ABSTRACT A major challenge to global malaria control and elimination is early detection and containment of emerging drug resistance. Next-generation sequencing (NGS) methods provide the resolution, scalability, and sensitivity required for high-throughput surveillance of molecular markers of drug resistance. We have developed an amplicon sequencing method on the Ion Torrent PGM platform for targeted resequencing of a panel of six Plasmodium falciparum genes implicated in resistance to first-line antimalarial therapy, including artemisinin combination therapy, chloroquine, and sulfadoxine-pyrimethamine. The protocol was optimized using 12 geographically diverse P. falciparum reference strains and successfully applied to multiplexed sequencing of 16 clinical isolates from India. The sequencing results from the reference strains showed 100% concordance with previously reported drug resistance-associated mutations. Single-nucleotide polymorphisms (SNPs) in clinical isolates revealed a number of known resistance-associated mutations and other nonsynonymous mutations that have not been implicated in drug resistance. SNP positions containing multiple allelic variants were used to identify three clinical samples containing mixed genotypes indicative of multiclonal infections. The amplicon sequencing protocol has been designed for the benchtop Ion Torrent PGM platform and can be operated with minimal bioinformatics infrastructure, making it ideal for use in countries that are endemic for the disease to facilitate routine large-scale surveillance of the emergence of drug resistance and to ensure continued success of the malaria treatment policy.

Funder

HHS | NIH | National Institute of Allergy and Infectious Diseases

Publisher

American Society for Microbiology

Subject

Microbiology (medical)

Reference88 articles.

1. WHO/UNICEF. 2015. Achieving the malaria MDG target: reversing the incidence of malaria 2000–2015. World Health Organization Geneva Switzerland. http://apps.who.int/iris/bitstream/10665/184521/1/9789241509442_eng.pdf?ua=1.

2. WHO. 2015. India drives down malaria rates sets sights on elimination. World Health Organization Geneva Switzerland. http://www.who.int/features/2015/india-programme-end-malaria/en/.

3. A better resolution for integrating methods for monitoring Plasmodium falciparum resistance to antimalarial drugs

4. Plasmodium falciparum:Detection of Polymorphisms in theDihydrofolate ReductaseandDihydropteroate SynthetaseGenes by PCR and Restriction Digestion

5. Use of Molecular Beacons To Detect an Antifolate Resistance-Associated Mutation in Plasmodium falciparum

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