Unraveling the Complexity of Imported Malaria Infections by Amplicon Deep Sequencing.

Xi He,Luyi Zhao,Liwang Cui,Weiling Zeng,Zheng Xiang,Guiyun Yan,Maohua Pan,Siqi Wang,Liang Pi,Yanrui Wu,Daibin Zhong,Chunyan Zou,Xi Chen,Yu Si,Yaming Huang,Zhaoqing Yang,Yucheng Qin

doi:10.3389/fcimb.2021.725859

Abstract

Imported malaria and recurrent infections are becoming an emerging issue in many malaria non-endemic countries. This study aimed to determine the molecular patterns of the imported malaria infections and recurrence. Blood samples were collected from patients with imported malaria infections during 2016–2018 in Guangxi Zhuang Autonomous Region, China. Next-generation amplicon deep-sequencing approaches were used to assess parasite genetic diversity, multiplexity of infection, relapse, recrudescence, and antimalarial drug resistance. A total of 44 imported malaria cases were examined during the study, of which 35 (79.5%) had recurrent malaria infections within 1 year. The majority (91.4%) had one recurrent malaria episode, whereas two patients had two recurrences and one patient had three recurrences. A total of 19 recurrence patterns (the species responsible for primary and successive clinical episodes) were found in patients returning from malaria epidemic countries. Four parasite species were detected with a higher than usual proportion (46.2%) of non-falciparum infections or mixed-species infections. An increasing trend of recurrence infections and reduced drug treatment efficacy were observed among the cases of imported malaria. The high recurrence rate and complex patterns of imported malaria from Africa to non-endemic countries have the potential to initiate local transmission, thereby undermining efforts to eliminate locally acquired malaria. Our findings highlight the power of amplicon deep-sequencing applications in molecular epidemiological studies of the imported malaria recurrences.

Highlights

Malaria remains a major public health problem in sub-Saharan Africa
For cytochrome b (CYTB), no polymorphism was detected within any species except for P. ovale, which includes two subspecies P. ovale curtisi and P. ovale wallikeri, whereas high polymorphism was found in the small subunit ribosomal RNA (SSU) marker with the number of haplotypes ranging from 1 to 5 in the five parasite species and similarity >99% compared to sequences in GenBank
Six out of the 15 mixed species were identified by the SSU marker, which showed various frequencies of different haplotypes, and most of them had predominant P. ovale curtisi mixed with less abundant P. falciparum (Supplementary Figure 1A, B)

Summary

Introduction

Malaria remains a major public health problem in sub-Saharan Africa. Guangxi had no indigenous malaria cases reported since 2013. Imported malaria is increasing due to increasing travel for tourism and trade in high-risk areas; Guangxi remains the area of China at highest risk for imported malaria (Lin et al, 2017; Jianhai et al, 2020). During 2011 and 2018 in Guangxi, a total of 3,943 imported malaria cases were reported and most of them originated from Africa (e.g., Ghana, Congo, and Cameroon), where Plasmodium falciparum is the predominant parasite species (Lin Kang-Ming et al, 2019). The increase in imported malaria cases poses a major challenge for the malaria elimination program in China (Lai et al, 2019)

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