Lumefantrine pressure selects nonsynonymous mutation in cysteine desulfurase IscS gene in the rodent malaria parasite Plasmodium berghei ANKA

Beatrice Gachie, Jean Chepngetich, Brenda Muriithi, Kelvin Thiong’o, Jeremiah Gathirwa, Francis Kimani, Peter Mwitari, Gabriel Magoma, Daniel Kiboi
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Abstract

Background: Lumefantrine (LM), piperaquine (PQ), and amodiaquine (AQ) are the essential long-acting partner drugs in the artemisinin-based combination therapies (ACTs) treatment regimens globally. The recent report on the emergence of artemisinin-resistant parasites portends an imminent failure of the partner drug in clearing the high residual parasite densities. Understanding the resistance mechanisms to partner drugs remains critical for tracking resistant parasites. Cysteine desulfurase IscS (nfs1), one of the proteins involved in the iron-sulfur (FeS) biogenesis pathway, has been implicated in mediating malaria parasite drug resistance. Methods: Using the rodent malaria parasites Plasmodium berghei ANKA in mice, we assessed whether the nfs1 gene is associated with LM, PQ, and AQ resistance. We first verified the stability of the LM, PQ, and AQ-resistant parasites in the standard 4-Day Suppressive Test. By means of PCR and sequencing analysis, we probed for single nucleotide polymorphisms (SNPs) in the nfs1 gene. Using qPCR, we then measured the expression of the nfs1 gene in resistant parasites relative to the drug-sensitive parent parasites. Results: Our analyses of nfs1 reveal a non-synonymous Gln142Arg mutation in the LM and PQ-resistant parasites. This mutation was not detected in the AQ-resistant parasites. The mRNA quantification of the nfs1 gene reveals differential expression in both LM and PQ-resistant parasites. Conversely, nfs1 expression remained unchanged in the AQ-resistant parasites. Conclusion: Our data suggest that LM and PQ selection pressure induces nonsynonymous mutation and differential expression of the nfs1 gene in Plasmodium berghei. Collectively, these findings provide a premise for investigating LM and PQ resistance mechanisms in both P. berghei and P. falciparum.
甲苯胺压力在啮齿动物疟原虫伯氏疟原虫中选择半胱氨酸脱硫酶IscS基因的非同义突变
背景:氨芳汀(LM)、哌喹(PQ)和阿莫地喹(AQ)是全球以青蒿素为基础的联合疗法(ACTs)治疗方案中必不可少的长效伴用药。最近关于出现青蒿素耐药寄生虫的报告预示着伴侣药物在清除高残留寄生虫密度方面即将失败。了解对伴侣药物的耐药机制对于追踪耐药寄生虫仍然至关重要。半胱氨酸脱硫酶IscS (nfs1)是参与铁硫(FeS)生物发生途径的蛋白之一,在介导疟疾寄生虫耐药性过程中发挥了重要作用。方法:采用小鼠感染的伯氏疟原虫ANKA检测nfs1基因是否与LM、PQ和AQ耐药相关。我们首先在标准的4天抑制试验中验证了LM、PQ和aq耐药寄生虫的稳定性。通过PCR和测序分析,我们检测了nfs1基因的单核苷酸多态性(SNPs)。然后,我们使用qPCR检测了耐药寄生虫中nfs1基因相对于药物敏感亲本寄生虫的表达。结果:我们对nfs1的分析显示,在LM和pq抗性寄生虫中存在非同义的Gln142Arg突变。该突变未在抗aq寄生虫中检测到。nfs1基因的mRNA定量分析显示,在LM和pq抗性寄生虫中,nfs1基因的表达存在差异。相反,nfs1的表达在抗aq寄生虫中保持不变。结论:我们的数据表明,LM和PQ选择压力诱导了伯格氏疟原虫nfs1基因的非同义突变和差异表达。总之,这些发现为研究柏氏疟原虫和恶性疟原虫的LM和PQ耐药机制提供了前提。
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