PDF(Pubmed)
Abstract：
BACKGROUND: Rapid diagnostic tests (RDTs) play a significant role in expanding case management in peripheral healthcare systems. Histidine-rich protein-2 (HRP2) antigen detection RDTs are predominantly used to diagnose Plasmodium falciparum infection. However, the evolution and spread of P. falciparum parasite strains with deleted hrp2/3 genes, causing false-negative results, have been reported. This study assessed the diagnostic performance of HRP2-detecting RDTs for P. falciparum cases and the prevalence of pfhrp2/3 deletions among symptomatic patients seeking malaria diagnosis at selected health facilities in southern Ethiopia.
METHODS: A multi-health facilities-based cross-sectional study was conducted on self-presenting febrile patients seeking treatment in southern Ethiopia from July to September 2022. A purposive sampling strategy was used to enroll patients with microscopically confirmed P. falciparum infections. A capillary blood sample was obtained to prepare a blood film for microscopy and a RDT using the SD Bioline™ Malaria Pf/Pv Test. Dried blood spot samples were collected for further molecular analysis. DNA was extracted using gene aid kits and amplification was performed using nested PCR assay. Exon 2 of hrp2 and hrp3, which are the main protein-coding regions, was used to confirm its deletion. The diagnostic performance of RDT was evaluated using PCR as the gold standard test for P. falciparum infections.
RESULTS: Of 279 P. falciparum PCR-confirmed samples, 249 (89.2%) had successful msp-2 amplification, which was then genotyped for hrp2/3 gene deletions. The study revealed that pfhrp2/3 deletions were common in all health centres, and it was estimated that 144 patients (57.8%) across all health facilities had pfhrp2/3 deletions, leading to false-negative PfHRP2 RDT results. Deletions spanning exon 2 of hrp2, exon 2 of hrp3, and double deletions (hrp2/3) accounted for 68 (27.3%), 76 (30.5%), and 33 (13.2%) of cases, respectively. The study findings revealed the prevalence of P. falciparum parasites lacking a single pfhrp2-/3-gene and that both genes varied across the study sites. This study also showed that the sensitivity of the SD Bioline PfHRP2-RDT test was 76.5% when PCR was used as the reference test.
CONCLUSIONS: This study confirmed the existence of widespread pfhrp2/3- gene deletions, and their magnitude exceeded the WHO-recommended threshold (> 5%). False-negative RDT results resulting from deletions in Pfhrp2/3- affect a country\'s attempts at malaria control and elimination. Therefore, the adoption of non-HRP2-based RDTs as an alternative measure is required to avoid the consequences associated with the continued use of HRP-2-based RDTs, in the study area in particular and in Ethiopia in general.
METHODS: A multi-health facilities-based cross-sectional study was conducted on self-presenting febrile patients seeking treatment in southern Ethiopia from July to September 2022. A purposive sampling strategy was used to enroll patients with microscopically confirmed P. falciparum infections. A capillary blood sample was obtained to prepare a blood film for microscopy and a RDT using the SD Bioline™ Malaria Pf/Pv Test. Dried blood spot samples were collected for further molecular analysis. DNA was extracted using gene aid kits and amplification was performed using nested PCR assay. Exon 2 of hrp2 and hrp3, which are the main protein-coding regions, was used to confirm its deletion. The diagnostic performance of RDT was evaluated using PCR as the gold standard test for P. falciparum infections.
RESULTS: Of 279 P. falciparum PCR-confirmed samples, 249 (89.2%) had successful msp-2 amplification, which was then genotyped for hrp2/3 gene deletions. The study revealed that pfhrp2/3 deletions were common in all health centres, and it was estimated that 144 patients (57.8%) across all health facilities had pfhrp2/3 deletions, leading to false-negative PfHRP2 RDT results. Deletions spanning exon 2 of hrp2, exon 2 of hrp3, and double deletions (hrp2/3) accounted for 68 (27.3%), 76 (30.5%), and 33 (13.2%) of cases, respectively. The study findings revealed the prevalence of P. falciparum parasites lacking a single pfhrp2-/3-gene and that both genes varied across the study sites. This study also showed that the sensitivity of the SD Bioline PfHRP2-RDT test was 76.5% when PCR was used as the reference test.
CONCLUSIONS: This study confirmed the existence of widespread pfhrp2/3- gene deletions, and their magnitude exceeded the WHO-recommended threshold (> 5%). False-negative RDT results resulting from deletions in Pfhrp2/3- affect a country\'s attempts at malaria control and elimination. Therefore, the adoption of non-HRP2-based RDTs as an alternative measure is required to avoid the consequences associated with the continued use of HRP-2-based RDTs, in the study area in particular and in Ethiopia in general.
摘要：
背景：快速诊断测试（RDT）在扩展外围医疗保健系统中的病例管理中起着重要作用。富含组氨酸的蛋白2（HRP2）抗原检测RDT主要用于诊断恶性疟原虫感染。然而,hrp2/3基因缺失的恶性疟原虫的进化和传播，导致假阴性结果，已被报道。这项研究评估了在埃塞俄比亚南部选定的医疗机构寻求疟疾诊断的有症状患者中，HRP2检测RDT对恶性疟原虫病例的诊断性能以及pfhrp2/3缺失的患病率。
方法：对2022年7月至9月在埃塞俄比亚南部寻求治疗的自我表现的发热患者进行了一项基于多医疗机构的横断面研究。使用有目的的抽样策略来招募具有显微镜确认的恶性疟原虫感染的患者。使用SDBioline™疟疾Pf/Pv测试获得毛细管血液样品以制备用于显微镜检查的血膜和RDT。收集干燥的血斑样品用于进一步的分子分析。使用基因辅助试剂盒提取DNA,并使用巢式PCR测定进行扩增。hrp2和hrp3的外显子2是主要的蛋白质编码区，用于确认其删除。使用PCR作为恶性疟原虫感染的金标准测试来评估RDT的诊断性能。
结果：在279个恶性疟原虫PCR证实的样本中，249例（89.2%）msp-2扩增成功，然后对其进行hrp2/3基因缺失的基因分型。研究表明，pfhrp2/3缺失在所有健康中心都很常见，据估计，所有医疗机构中有144名患者(57.8%)有pfhrp2/3缺失，导致PfHRP2RDT结果假阴性。跨越hrp2外显子2、hrp3外显子2的缺失和双缺失(hrp2/3)占68(27.3%),76（30.5%），33例(13.2%),分别。研究发现揭示了缺乏单个pfhrp2-/3-基因的恶性疟原虫寄生虫的患病率，并且两个基因在研究地点都有所不同。本研究还表明，以PCR作为参考测试时，SDBiolinePfHRP2-RDT测试的灵敏度为76.5％。
结论：这项研究证实存在广泛的pfhrp2/3-基因缺失，其大小超过WHO推荐的阈值(>5%)。Pfhrp2/3缺失导致的假阴性RDT结果会影响一个国家控制和消除疟疾的尝试。因此,需要采用基于非HRP2的RDT作为替代措施，以避免与继续使用基于HRP-2的RDT相关的后果，在研究领域，特别是在埃塞俄比亚。
方法：对2022年7月至9月在埃塞俄比亚南部寻求治疗的自我表现的发热患者进行了一项基于多医疗机构的横断面研究。使用有目的的抽样策略来招募具有显微镜确认的恶性疟原虫感染的患者。使用SDBioline™疟疾Pf/Pv测试获得毛细管血液样品以制备用于显微镜检查的血膜和RDT。收集干燥的血斑样品用于进一步的分子分析。使用基因辅助试剂盒提取DNA,并使用巢式PCR测定进行扩增。hrp2和hrp3的外显子2是主要的蛋白质编码区，用于确认其删除。使用PCR作为恶性疟原虫感染的金标准测试来评估RDT的诊断性能。
结果：在279个恶性疟原虫PCR证实的样本中，249例（89.2%）msp-2扩增成功，然后对其进行hrp2/3基因缺失的基因分型。研究表明，pfhrp2/3缺失在所有健康中心都很常见，据估计，所有医疗机构中有144名患者(57.8%)有pfhrp2/3缺失，导致PfHRP2RDT结果假阴性。跨越hrp2外显子2、hrp3外显子2的缺失和双缺失(hrp2/3)占68(27.3%),76（30.5%），33例(13.2%),分别。研究发现揭示了缺乏单个pfhrp2-/3-基因的恶性疟原虫寄生虫的患病率，并且两个基因在研究地点都有所不同。本研究还表明，以PCR作为参考测试时，SDBiolinePfHRP2-RDT测试的灵敏度为76.5％。
结论：这项研究证实存在广泛的pfhrp2/3-基因缺失，其大小超过WHO推荐的阈值(>5%)。Pfhrp2/3缺失导致的假阴性RDT结果会影响一个国家控制和消除疟疾的尝试。因此,需要采用基于非HRP2的RDT作为替代措施，以避免与继续使用基于HRP-2的RDT相关的后果，在研究领域，特别是在埃塞俄比亚。
