Li Jiangyan, Tao Zhiyong, Li Qian, Brashear Awtum, Wang Ying, Xia Hui, Fang Qiang, Cui Liwang
Department of Microbiology and Parasitology, Bengbu Medical College, Bengbu, China.
Anhui Key Laboratory of Infection and Immunity, Bengbu Medical College, Bengbu, China.
Malar J. 2017 May 17;16(1):201. doi: 10.1186/s12936-017-1855-3.
Isolation of Plasmodium-infected red blood cells (iRBCs) from clinical blood samples is often required for experiments, such as ex vivo drug assays, in vitro invasion assays and genome sequencing. Current methods for removing white blood cells (WBCs) from malaria-infected blood are time-consuming or costly. A prototype non-woven fabric (NWF) filter was developed for the purification of iRBCs, which showed great efficiency for removing WBCs in a pilot study. Previous work was performed with prototype filters optimized for processing 5-10 mL of blood. With the commercialization of the filters, this study aims to evaluate the efficiency and suitability of the commercial NWF filter for the purification of Plasmodium vivax-infected RBCs in smaller volumes of blood and to compare its performance with that of Plasmodipur filters.
Forty-three clinical P. vivax blood samples taken from symptomatic patients attending malaria clinics at the China-Myanmar border were processed using the NWF filters in a nearby field laboratory. The numbers of WBCs and iRBCs and morphology of P. vivax parasites in the blood samples before and after NWF filtration were compared. The viability of P. vivax parasites after filtration from 27 blood samples was examined by in vitro short-term culture. In addition, the effectiveness of the NWF filter for removing WBCs was compared with that of the Plasmodipur filter in six P. vivax blood samples.
Filtration of 1-2 mL of P. vivax-infected blood with the NWF filter removed 99.68% WBCs. The densities of total iRBCs, ring and trophozoite stages before and after filtration were not significantly different (P > 0.05). However, the recovery rates of schizont- and gametocyte-infected RBCs, which were minor parasite stages in the clinical samples, were relatively low. After filtration, the P. vivax parasites did not show apparent morphological changes. Culture of 27 P. vivax-infected blood samples after filtration showed that parasites successfully matured into the schizont stage. The WBC removal rates and iRBC recovery rates were not significantly different between the NWF and Plasmodipur filters (P > 0.05).
When tested with 1-2 mL of P. vivax-infected blood, the NWF filter could effectively remove WBCs and the recovery rates for ring- and trophozoite-iRBCs were high. P. vivax parasites after filtration could be successfully cultured in vitro to reach maturity. The performance of the NWF and Plasmodipur filters for removing WBCs and recovering iRBCs was comparable.
从临床血样中分离疟原虫感染的红细胞（iRBCs）常用于体外药物检测、体外侵袭试验和基因组测序等实验。目前从疟疾感染血液中去除白细胞（WBCs）的方法耗时或成本高昂。已开发出一种用于纯化iRBCs的原型无纺布（NWF）过滤器，在一项初步研究中显示出去除WBCs的高效性。先前的工作是使用针对处理5 - 10毫升血液优化的原型过滤器进行的。随着过滤器的商业化，本研究旨在评估商业NWF过滤器在较小体积血液中纯化间日疟原虫感染红细胞的效率和适用性，并将其性能与疟原虫净化过滤器的性能进行比较。
在中国 - 缅甸边境疟疾诊所就诊的有症状患者采集的43份临床间日疟原虫血样，在附近的现场实验室使用NWF过滤器进行处理。比较NWF过滤前后血样中WBCs和iRBCs的数量以及间日疟原虫寄生虫的形态。通过体外短期培养检测27份血样过滤后的间日疟原虫寄生虫的活力。此外，在6份间日疟原虫血样中比较了NWF过滤器与疟原虫净化过滤器去除WBCs的有效性。
用NWF过滤器过滤1 - 2毫升间日疟原虫感染的血液可去除99.68%的WBCs。过滤前后总iRBCs、环状体和滋养体阶段的密度无显著差异（P > 0.05）。然而，裂殖体和配子体感染的红细胞（临床样本中的次要寄生虫阶段）的回收率相对较低。过滤后，间日疟原虫寄生虫未显示明显的形态变化。对27份过滤后的间日疟原虫感染血样进行培养表明，寄生虫成功成熟为裂殖体阶段。NWF过滤器和疟原虫净化过滤器之间的WBC去除率和iRBC回收率无显著差异（P > 0.05）。
当用1 - 2毫升间日疟原虫感染的血液进行测试时，NWF过滤器可有效去除WBCs，环状体和滋养体iRBCs的回收率较高。过滤后的间日疟原虫寄生虫可在体外成功培养至成熟。NWF过滤器和疟原虫净化过滤器在去除WBCs和回收iRBCs方面的性能相当。
