Yengo Bernis Neneyoh, Ruszin Victoria, Dieng Cheikh Cambel, Kipayko Canelle, Mdluli Nontokozo, Ayukenchengamba Bate, Bisong Ebai Calvin, Sona Doris Tabi, Nematchoua Weyou Zidedine, Sumbele Irene, Kimbi Helen Kuokuo, Lo Eugenia
Department of Microbiology and Immunology, Drexel University, College of Medicine, Philadelphia, PA, USA.
Department of Animal Biology and Conservation (ABC), Faculty of Science, University of Buea, Buea, Cameroon.
J Infect Dis. 2025 Jun 12. doi: 10.1093/infdis/jiaf322.
Vivax malaria, once thought rare in Duffy-negative Africans, is now reported in various parts of Africa, suggesting alternate invasion mechanisms and parasite adaptability to Duffy-null cells. One hypothesis is that copy number variation (CNV) of genes involved in erythrocyte invasion may impact parasite invasion capability and/or host immune evasion particularly in Duffy-negative individuals.
Using novel digital PCR, we assessed CNV of three key erythrocyte-binding genes of Pv isolated from DN individuals in three eco-epidemiological zones of Cameroon. For a subset of samples, we compare dPCR results with qPCR and PCR diagnostic approaches.
PvDBP1 duplications were detected in ∼92% of DN Pv samples, compared to ∼10% of the samples with multi-copy PvEBP/DBP2 and PvRBP2b. A significant positive correlation was detected between PvDBP1 CNV and parasite load among the samples. Both Malagasy- and Cambodian-type PvDBP1 duplications were detected. About one-third of the samples harbored both duplication types and these samples were exclusively in northern highland of Cameroon, suggesting either polyclonal infections or a third duplication type.
Multi-copy PvDBP1 across all study sites may imply significant parasite adaptability, and improved parasite invasion, potentially influencing parasitemia. This was confirmed with the significantly higher parasitemia observed in samples with Cambodian and those with both duplication types compared to ones with no duplication. Further, our data showed CNV analysis by qPCR may not be as precise as dPCR particularly for low-parasitemia DN Pv samples. Predominantly low PvEBP/DBP2 and PvRBP2b copy raise question to their role in parasite adaptation to invading DN erythrocytes.
间日疟原虫曾被认为在达菲阴性的非洲人中罕见，现在非洲各地均有报道，这表明存在其他入侵机制以及寄生虫对达菲阴性细胞的适应性。一种假说认为，参与红细胞入侵的基因的拷贝数变异（CNV）可能会影响寄生虫的入侵能力和/或宿主免疫逃避，特别是在达菲阴性个体中。
我们使用新型数字PCR，评估了从喀麦隆三个生态流行病学区域的达菲阴性个体中分离出的间日疟原虫的三个关键红细胞结合基因的CNV。对于一部分样本，我们将数字PCR结果与定量PCR和PCR诊断方法进行了比较。
在约92%的达菲阴性间日疟原虫样本中检测到间日疟原虫DBP1（PvDBP1）重复，相比之下，多拷贝间日疟原虫红细胞结合蛋白/DBP2（PvEBP/DBP2）和间日疟原虫RBP2b（PvRBP2b）的样本约为10%。在样本中检测到PvDBP1 CNV与寄生虫载量之间存在显著正相关。检测到马达加斯加型和柬埔寨型PvDBP1重复。约三分之一的样本同时含有两种重复类型，且这些样本仅在喀麦隆北部高地，这表明要么是多克隆感染，要么是第三种重复类型。
所有研究地点的多拷贝PvDBP1可能意味着寄生虫具有显著的适应性，并改善了寄生虫入侵，可能影响寄生虫血症。与无重复的样本相比，在柬埔寨型样本以及同时具有两种重复类型的样本中观察到的寄生虫血症显著更高，这证实了上述结论。此外，我们的数据表明，定量PCR进行的CNV分析可能不如数字PCR精确，特别是对于低寄生虫血症的达菲阴性间日疟原虫样本。PvEBP/DBP2和PvRBP2b的拷贝数主要较低，这对它们在寄生虫适应入侵达菲阴性红细胞中的作用提出了疑问。
