ONT sequencing identifies a high prevalence of sensitive, triple mutant and single mutant parasites within an ANC population in Nigeria.

BACKGROUND

Malaria in pregnancy is a major public health issue, particularly among vulnerable populations in malaria-endemic sub-Saharan African countries. To mitigate its risks, WHO recommends sulphadoxine-pyrimethamine (SP) for chemoprevention and artemisinin-based combination therapy (ACT) to treat uncomplicated malaria. These interventions have helped to alleviate the risk associated with malaria in pregnancy; however, in the context of the emergence of SP- and ACT-resistant , maintained efficacy is under threat. Molecular surveillance is a reliable tool to monitor the emergence of resistance where molecular markers are known. Thus, the objective of the study was to use a multiplexed amplicon Oxford Nanopore sequencing approach to assess the molecular markers for antimalarial resistance among pregnant women in Nigeria.

METHODS

Dried blood spots (DBS) were collected from pregnant women who received IPTp-SP at the enrollment and follow-up visits. genomic DNA was extracted by the Chelex method and 18S qPCR was used to detect parasite DNA in each sample. With nested PCR assays, fragments of , , , , and genes were amplified and multiplexed amplicon-based sequencing was conducted on the minION Oxford Nanopore Technology.

RESULT

In total, 251 pregnant women were enrolled in the study and 457 DBS samples were collected. genomic DNA was detected in 12% (56/457) of the samples, 31 at baseline and the remaining during the follow-up visits. , , , , were successfully sequenced in a single run. Notably, k13 artemisinin resistance mutations were absent, the frequencies of and SP resistance haplotypes, for pyrimethamine resistance and IKA/IAKA associated with sulphadoxine resistance were 82% (36/44) and 64% (27/42), respectively, and the CV resistant haplotype was at approximately 22% (7/32).

CONCLUSION AND RECOMMENDATIONS

Here a multiplexed amplicon-based ONT assay established that triple mutant -IRN, double mutant -SG haplotypes and the chloroquine sensitive strain were prevalent among pregnant women in Nigeria.