Molecular identification and anti-malarial drug resistance profile of Plasmodium falciparum from patients attending Kisoro Hospital, southwestern Uganda

Godfrey Manirakiza,Ivan Mugisha Taremwa,Fredrick Byarugaba,Kennedy Kassaza,Joel Bazira

doi:10.1186/s12936-021-04023-3

Godfrey Manirakiza, Ivan Mugisha Taremwa + Show 3 more

Open Access

https://doi.org/10.1186/s12936-021-04023-3

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Abstract

BackgroundThe evolution of malaria infection has necessitated the development of highly sensitive diagnostic assays, as well as the use of dried blood spots (DBS) as a potential source of deoxyribonucleic acid (DNA) yield for polymerase chain reaction (PCR) assays. This study identified the different Plasmodium species in malaria-positive patients, and the anti-malarial drug resistance profile for Plasmodium falciparum using DBS samples collected from patients attending Kisoro Hospital in Kisoro district, Southwestern Uganda.MethodsThe blood samples were prospectively collected from patients diagnosed with malaria to make DBS, which were then used to extract DNA for real-time PCR and high-resolution melting (HRM) analysis. Plasmodium species were identified by comparing the control and test samples using HRM-PCR derivative curves. Plasmodium falciparum chloroquine (CQ) resistance transporter (pfcrt) and kelch13 to screen the samples for anti-malarial resistance markers. The HRM-PCR derivative curve was used to present a summary distribution of the different Plasmodium species as well as the anti-malarial drug profile.ResultsOf the 152 participants sampled, 98 (64.5%) were females. The average age of the participants was 34.9 years (range: 2 months–81 years). There were 134 samples that showed PCR amplification, confirming the species as Plasmodium. Plasmodium falciparum (N = 122), Plasmodium malariae (N = 6), Plasmodium ovale (N = 4), and Plasmodium vivax (N = 2) were the various Plasmodium species and their proportions. The results showed that 87 (71.3%) of the samples were sensitive strains/wild type (CVMNK), 4 (3.3%) were resistant haplotypes (SVMNT), and 31 (25.4%) were resistant haplotypes (CVIET). Kelch13 C580Y mutation was not detected.ConclusionThe community served by Kisoro hospital has a high Plasmodium species burden, according to this study. Plasmodium falciparum was the dominant species, and it has shown that resistance to chloroquine is decreasing in the region. Based on this, molecular identification of Plasmodium species is critical for better clinical management. Besides, DBS is an appropriate medium for DNA preservation and storage for future epidemiological studies.

Highlights

The evolution of malaria infection has necessitated the development of highly sensitive diagnostic assays, as well as the use of dried blood spots (DBS) as a potential source of deoxyribonucleic acid (DNA) yield for polymerase chain reaction (PCR) assays
Sample collection and analyses Venous blood samples from patients who tested positive for malaria between March and August, 2020 and DBS was prepared on Whatmann® 903TM filter paper (Ref: 10530143 Rev.AA) by putting a drop of the blood sample from the finger prick into each cycle of the filter paper
This study found a high prevalence of pan Plasmodium species in the community served by Kisoro Hospital, confirming the previously reported rising malaria burden in this study setting

Summary

Introduction

The evolution of malaria infection has necessitated the development of highly sensitive diagnostic assays, as well as the use of dried blood spots (DBS) as a potential source of deoxyribonucleic acid (DNA) yield for polymerase chain reaction (PCR) assays. Manirakiza et al Malaria Journal (2022) 21:21 deaths. The total percentage of malaria deaths among children under 5 years was 67% in 2019 [2]. The burden of malaria is higher among children and pregnant women, but people of all ages are at risk of infection [3, 4]. In 2019, an estimated 215 million malaria cases in the World Health Organization (WHO) Africa region, accounting for 94% of the global malaria cases and this contributed to 51% of malaria deaths globally [2]. Malaria accounts for 25 to 40% of all outpatient visits to health facilities in Uganda, and it is responsible for nearly half of all inpatient paediatric deaths [5, 6]. While rain fall determines the availability of breeding habitats for mosquito vectors, temperature determines the length of mosquito larvae development and the rate of growth of the malaria parasites inside the vector [7,8,9,10]

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