评估 2018-2019 年刚果民主共和国治疗失败患者分离株中 pfk13-推进器、pfcrt 和 pfmdr1 基因中的疟原虫抗疟药物耐药标志物。
Assessment of Plasmodium falciparum anti-malarial drug resistance markers in pfk13-propeller, pfcrt and pfmdr1 genes in isolates from treatment failure patients in Democratic Republic of Congo, 2018-2019.
机构信息
Department of Basic Sciences, Faculty of Medicine, University of Kinshasa, Kinshasa, Democratic Republic of Congo.
School of Public Health and Research Institute of Health and Society, Catholic University of Louvain, 1200, Brussels, Belgium.
出版信息
Malar J. 2021 Mar 11;20(1):144. doi: 10.1186/s12936-021-03636-y.
BACKGROUND
The national policy for malaria treatment of the Democratic Republic of Congo recommends two first-line artemisinin-based combinations for the treatment of uncomplicated malaria: artesunate-amodiaquine and artemether-lumefantrine. This study investigated the presence of markers associated with resistance to the current first-line artemisinin-based combination therapy (ACT) in isolates of Plasmodium falciparum from treatment failure patients in the Democratic Republic of Congo.
METHODS
From November 2018 to November 2019, dried blood spots were taken from patients returning to health centres for fever within 28 days after an initial malaria treatment in six sentinel sites of the National Malaria Control Programme across Democratic Republic of Congo. The new episode of malaria was first detected by a rapid diagnostic test and then confirmed by a real-time PCR assay to define treatment failure. Fragments of interest in pfk13 and pfcrt genes were amplified by conventional PCR before sequencing and the Pfmdr1 gene copy number was determined by a TaqMan real-time PCR assay.
RESULTS
Out of 474 enrolled patients, 364 (76.8%) were confirmed positive by PCR for a new episode of P. falciparum malaria, thus considered as treatment failure. Of the 325 P. falciparum isolates obtained from 364 P. falciparum-positive patients and successfully sequenced in the pfk13-propeller gene, 7 (2.2%) isolates carried non-synonymous mutations, among which 3 have been previously reported (N498I, N554K and A557S) and 4 had not yet been reported (F506L, E507V, D516E and G538S). Of the 335 isolates successfully sequenced in the pfcrt gene, 139 (41.5%) harboured the K76T mutation known to be associated with chloroquine resistance. The SVMNT haplotype associated with resistance to amodiaquine was not found. None of the isolates carried an increased copy number of the pfmdr1 gene among the 322 P. falciparum isolates successfully analysed.
CONCLUSION
No molecular markers currently known to be associated with resistance to the first-line ACT in use were detected in isolates of P. falciparum from treatment failure patients. Regular monitoring through in vivo drug efficacy and molecular studies must continue to ensure the effectiveness of malaria treatment in Democratic Republic of Congo.
背景
刚果民主共和国的国家疟疾治疗政策推荐两种青蒿素为基础的联合疗法（ACT）一线药物治疗无并发症疟疾：青蒿琥酯-阿莫地喹和青蒿素-哌喹。本研究调查了在刚果民主共和国六个国家疟疾控制规划哨点从治疗失败的患者中分离出的恶性疟原虫分离株中，与当前一线青蒿素为基础的联合疗法（ACT）耐药相关的标记物的存在情况。
方法
2018 年 11 月至 2019 年 11 月，从刚果民主共和国国家疟疾控制规划的六个哨点中返回卫生中心发热的初始疟疾治疗后 28 天内的患者中采集了干血斑。通过快速诊断检测首次检测到新的疟疾发作，然后通过实时 PCR 检测确认治疗失败。在测序之前，通过常规 PCR 扩增 pfk13 和 pfcrt 基因中的感兴趣片段，并用 TaqMan 实时 PCR 检测 Pfmdr1 基因拷贝数。
结果
在 474 名入组患者中，364 名（76.8%）通过 PCR 检测到新的恶性疟原虫疟疾发作，因此被认为是治疗失败。从 364 名恶性疟原虫阳性患者中获得的 325 株恶性疟原虫分离株中，有 7 株（2.2%）分离株携带非同义突变，其中 3 株已被报道（N498I、N554K 和 A557S），4 株尚未报道（F506L、E507V、D516E 和 G538S）。在成功测序的 335 株 pfcrt 基因中，有 139 株（41.5%）携带与氯喹耐药相关的 K76T 突变。未发现与阿莫地喹耐药相关的 SVMNT 单倍型。在成功分析的 322 株恶性疟原虫分离株中，没有一株携带 pfmdr1 基因的拷贝数增加。
结论
在治疗失败患者的恶性疟原虫分离株中未发现与目前使用的一线 ACT 耐药相关的已知分子标记物。必须通过体内药物疗效和分子研究进行定期监测，以确保刚果民主共和国的疟疾治疗效果。
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