Addis Ababa University, Aklilu Lemma Institute of Pathobiology, Addis Ababa, Ethiopia.
Department of Bioinformatics and Genomics, University of North Carolina at Charlotte, Charlotte, NC, 28223, USA.
Malar J. 2021 Feb 23;20(1):109. doi: 10.1186/s12936-021-03629-x.
Rapid diagnostic tests (RDTs) targeting histidine rich protein 2(HRP2) are widely used for diagnosis of Plasmodium falciparum infections. Besides PfHRP2, the PfHRP3 antigen contributes to the detection of P. falciparum infections in PfHRP2 RDTs. However, the performance HRP2-based RDT is affected by pfhrp2/3 gene deletions resulting in false-negative test results. The objective of this study was to determine the presence and prevalence of pfhrp2/3 gene deletions including the respective flanking regions among symptomatic patients in Assosa zone, Northwest Ethiopia.
A health-facility based cross-sectional study was conducted in febrile patients seeking a malaria diagnosis in 2018. Blood samples were collected by finger-prick for microscopic examination of blood smears, malaria RDT, and molecular analysis using dried blood spots (DBS) prepared on Whatman filter paper. A total of 218 P. falciparum positive samples confirmed by quantitative PCR were included for molecular assay of pfhrp2/3 target gene.
Of 218 P. falciparum positive samples, exon 2 deletions were observed in 17.9% of pfhrp2 gene and in 9.2% of pfhrp3 gene. A high proportion of deletions in short segments of pfhrp2 exon1-2 (50%) was also detected while the deletions of the pfhrp3 exon1-2 gene were 4.1%. The deletions were extended to the downstream and upstream of the flanking regions in pfhrp2/3 gene (above 30%). Of eighty-six PfHRP2 RDT negative samples, thirty-six lacked pfhrp2 exon 2. Five PfHRP2 RDT negative samples had double deletions in pfhrp2 exon 2 and pfhrp3 exon2. Of these double deletions, only two of the samples with a parasite density above 2000 parasite/µl were positive by the microscopy. Three samples with intact pfhrp3 exon2 in the pfhrp2 exon2 deleted parasite isolates were found to be positive by PfHRP2 RDT and microscopy with a parasite density above 10,000/µl.
This study confirms the presence of deletions of pfhrp2/3 gene including the flanking regions. Pfhrp2/3 gene deletions results in false-negative results undoubtedly affect the current malaria control and elimination effort in the country. However, further countrywide investigations are required to determine the magnitude of pfhrp2/3 gene deletions and its consequences on routine malaria diagnosis.
快速诊断检测（RDT）针对富含组氨酸蛋白 2（HRP2），广泛用于诊断恶性疟原虫感染。除 PfHRP2 外，PfHRP3 抗原有助于在 PfHRP2 RDT 中检测到恶性疟原虫感染。然而，基于 HRP2 的 RDT 的性能受到 pfhrp2/3 基因缺失的影响，导致假阴性检测结果。本研究的目的是确定在埃塞俄比亚西北部阿索萨地区有症状患者中 pfhrp2/3 基因缺失的存在和流行情况，包括侧翼区域。
2018 年，在发热患者中进行了一项基于卫生机构的横断面研究，他们在寻求疟疾诊断时采集指血样本来进行血涂片显微镜检查、疟疾 RDT 和使用 Whatman 滤纸制备的干血斑（DBS）进行分子分析。总共纳入了 218 个由定量 PCR 确认的恶性疟原虫阳性样本，用于对 pfhrp2/3 靶基因进行分子分析。
在 218 个恶性疟原虫阳性样本中，pfhrp2 基因的外显子 2 缺失在 17.9%的样本中观察到，pfhrp3 基因的缺失在 9.2%的样本中观察到。还检测到 pfhrp2 exon1-2 短片段缺失的比例很高（50%），而 pfhrp3 exon1-2 基因的缺失为 4.1%。pfhrp2/3 基因的缺失延伸到侧翼区域的下游和上游（超过 30%）。在 86 个 PfHRP2 RDT 阴性样本中，有 36 个样本缺乏 pfhrp2 外显子 2。5 个 PfHRP2 RDT 阴性样本在 pfhrp2 外显子 2 和 pfhrp3 外显子 2 中存在双重缺失。在这些双重缺失中，只有两个寄生虫密度超过 2000 个寄生虫/µl 的样本通过显微镜检查呈阳性。在 pfhrp2 外显子 2 缺失寄生虫分离物中，pfhrp3 外显子 2 完整的 3 个样本通过 PfHRP2 RDT 和显微镜检查呈阳性，寄生虫密度超过 10,000/µl。
本研究证实了 pfhrp2/3 基因包括侧翼区域的缺失。pfhrp2/3 基因缺失导致假阴性结果，无疑会影响该国目前的疟疾控制和消除工作。然而，需要在全国范围内进行进一步的调查，以确定 pfhrp2/3 基因缺失的程度及其对常规疟疾诊断的影响。
