A comprehensive analysis of drug resistance molecular markers and Plasmodium falciparum genetic diversity in two malaria endemic sites in Mali
A comprehensive analysis of drug resistance molecular markers and Plasmodium falciparum genetic diversity in two malaria endemic sites in Mali
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DOI：
10.1186/s12936-019-2986-5
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发表时间：
2019-11-12
期刊：
影响因子：
3
通讯作者：
Diakite, Mahamadou
中科院分区：
文献类型：
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作者：
Diakite, Seidina A. S.;Traore, Karim;Diakite, Mahamadou
背景耐药性是马里疟疾控制方案面临的最大挑战之一。下一代测序（NGS）技术的最新进展为追踪非洲的抗药性疟疾寄生虫提供了新的有效方法。在马里的Dangassa和Nioro-du-Sahel这两个疟疾传播模式不同的地点评估了恶性疟原虫抗药性分子标记的多样性和流行率。丹加萨省的季节性疟疾传播频繁，而萨赫勒地区的季节性疟疾传播不稳定且时间较短。方法2016年从恶性疟原虫阳性患者中采集了多达270份干血斑样本（Dangassa 214份，Nioro-du-Sahel 56份）。样品在Mass MassARRAY（R）iPLEX平台上进行分析。靶向Pfcrt、Pfmdr 1、Pfdhfr和Pfdhps、Pfarps 10、Pfferredoxin、Pfexonuclease和Pfmdr 2基因中的特定密码子。采用桑格氏101-SNPs-条形码法对恶性疟原虫的遗传多样性进行了分析，并对疟原虫种属进行了鉴定。结果Dangassa和Nioro-du-Sahel地区的Pfcrt_76T氯喹耐药基因型检出率分别为64.4%和45.2%（p = 0.025）。Pfdhfr_51I-59 R-108 N乙胺嘧啶抗性基因型在丹加萨和尼奥罗-杜-萨赫勒分别为14.1%和19.6%。Pfdhps_S436-A437-K540-A581- 613 A硫代嘌呤抗性基因的突变在Dangassa中比在Nioro-du-Sahel中显著更普遍（p = 0.035）。高达17.8%的分离物从Dangassa与7%从Nioro-du-Sahel窝藏至少两个密码子替换在这个单倍型。仅在三个样本中发现了阿莫地喹耐药性Pfmdr1_N86Y突变（两个在Dangassa，一个在Nioro-du-Sahel）。在Dangassa和Nioro-du-Sahel分别有39.9%和48.2%的样本中发现了本芴醇敏感性降低的Pfmdr1_Y184F突变。在Dangassa中发现了一个哌喹耐药的Exo_E415G突变，而没有发现青蒿素耐药的遗传背景。观察到恶性疟原虫的高多样性，但在两个研究地点都没有发现明显的遗传聚集。相对于Nioro-du-Sahel，使用COIL（p = 0.04）和真实的McCOIL（p = 0.02）方法在Dangassa中观察到更高的感染复数。结论本研究揭示了高流行率的氯喹和乙胺嘧啶耐药标记，以及高密码子替换率的磺胺嘧啶耐药基因。恶性疟原虫的遗传多样性较高。这些观察结果表明，青蒿素的使用与丹加萨和萨赫勒地区的尼奥罗有关。
Background Drug resistance is one of the greatest challenges of malaria control programme in Mali. Recent advances in next-generation sequencing (NGS) technologies provide new and effective ways of tracking drug-resistant malaria parasites in Africa. The diversity and the prevalence of Plasmodium falciparum drug-resistance molecular markers were assessed in Dangassa and Nioro-du-Sahel in Mali, two sites with distinct malaria transmission patterns. Dangassa has an intense seasonal malaria transmission, whereas Nioro-du-Sahel has an unstable and short seasonal malaria transmission. Methods Up to 270 dried blood spot samples (214 in Dangassa and 56 in Nioro-du-Sahel) were collected from P. falciparum positive patients in 2016. Samples were analysed on the Agena MassARRAY (R) iPLEX platform. Specific codons were targeted in Pfcrt, Pfmdr1, Pfdhfr, and Pfdhps, Pfarps10, Pfferredoxin, Pfexonuclease and Pfmdr2 genes. The Sanger's 101-SNPs-barcode method was used to assess the genetic diversity of P. falciparum and to determine the parasite species. Results The Pfcrt_76T chloroquine-resistance genotype was found at a rate of 64.4% in Dangassa and 45.2% in Nioro-du-Sahel (p = 0.025). The Pfdhfr_51I-59R-108N pyrimethamine-resistance genotype was 14.1% and 19.6%, respectively in Dangassa and Nioro-du-Sahel. Mutations in the Pfdhps_S436-A437-K540-A581-613A sulfadoxine-resistance gene was significantly more prevalent in Dangassa as compared to Nioro-du-Sahel (p = 0.035). Up to 17.8% of the isolates from Dangassa vs 7% from Nioro-du-Sahel harboured at least two codon substitutions in this haplotype. The amodiaquine-resistance Pfmdr1_N86Y mutation was identified in only three samples (two in Dangassa and one in Nioro-du-Sahel). The lumefantrine-reduced susceptibility Pfmdr1_Y184F mutation was found in 39.9% and 48.2% of samples in Dangassa and Nioro-du-Sahel, respectively. One piperaquine-resistance Exo_E415G mutation was found in Dangassa, while no artemisinin resistance genetic-background were identified. A high P. falciparum diversity was observed, but no clear genetic aggregation was found at either study sites. Higher multiplicity of infection was observed in Dangassa with both COIL (p = 0.04) and Real McCOIL (p = 0.02) methods relative to Nioro-du-Sahel. Conclusions This study reveals high prevalence of chloroquine and pyrimethamine-resistance markers as well as high codon substitution rate in the sulfadoxine-resistance gene. High genetic diversity of P. falciparum was observed. These observations suggest that the use of artemisinins is relevant in both Dangassa and Nioro-du-Sahel.