Fluorescent Plasmodium berghei sporozoites and pre-erythrocytic stages:: a new tool to study mosquito and mammalian host interactions with malaria parasites
Fluorescent Plasmodium berghei sporozoites and pre-erythrocytic stages:: a new tool to study mosquito and mammalian host interactions with malaria parasites
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DOI：
10.1046/j.1462-5822.2001.00117.x
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发表时间：
2001-06-01
影响因子：
3.4
通讯作者：
Vernick, KD
中科院分区：
文献类型：
--
作者：
Natarajan, R;Thathy, V;Vernick, KD
为了在蚊子和哺乳动物宿主内追踪疟疾寄生虫进行生物学研究，我们构建了一个稳定转化的伯氏疟原虫无性系PbFluspo，其中造孢子性和红细胞前期肝期寄生虫是自主荧光的。从环子孢子(CS)蛋白基因的5‘和3’侧翼序列中表达了FACS适应绿色荧光蛋白突变体2(GFPmu2)的结构基因，并在内源CS基因座上进行了整合和表达。携带CS野生型副本的重组寄生虫产生高荧光卵囊和子孢子，这些卵囊和子孢子侵入蚊子唾液腺，并正常传播给啮齿动物宿主。寄生虫在体外感染培养的肝细胞，在那里它们发育成荧光的前红细胞形式。被这些寄生虫感染的哺乳动物细胞可以通过荧光激活细胞分类器(FACS)分析从未感染的细胞中分离出来。这些荧光昆虫和哺乳动物阶段的伯氏疟原虫应该有助于研究它们各自宿主的表型，以及识别在这些寄生虫阶段表达的新基因。
To track malaria parasites for biological studies within the mosquito and mammalian hosts, we constructed a stably transformed clonal line of Plasmodium berghei, PbFluspo, in which sporogonic and pre-erythrocytic liver-stage parasites are autonomously fluorescent. A cassette containing the structural gene for the FACS-adapted green fluorescent protein mutant 2 (GFPmut2), expressed from the 5' and 3' flanking sequences of the circumsporozoite (CS) protein gene, was integrated and expressed at the endogenous CS locus. Recombinant parasites, which bear a wild-type copy of CS, generated highly fluorescent oocysts and sporozoites that invaded mosquito salivary glands and were transmitted normally to rodent hosts. The parasites infected cultured hepatocytes in vitro, where they developed into fluorescent pre-erythrocytic forms. Mammalian cells infected by these parasites can be separated from non-infected cells by fluorescence activated cell sorter (FACS) analysis. These fluorescent insect and mammalian stages of P. berghei should be useful for phenotypic studies in their respective hosts, as well as for identification of new genes expressed in these parasite stages.