Contamin H, Fandeur T, Bonnefoy S, Skouri F, Ntoumi F, Mercereau-Puijalon O
Unité de Parasitologie Expérimentale, Institut Pasteur, Paris, France.
J Clin Microbiol. 1995 Apr;33(4):944-51. doi: 10.1128/jcm.33.4.944-951.1995.
We report on an analysis of the constraints of PCR typing of field Plasmodium falciparum isolates by using a few highly polymorphic markers, MSA-1, MSA-2, TRAP, and CS. We show that the reactions are specific for the P. falciparum species. The detection threshold (minimum number of parasites required to detect a visible band by ethidium bromide) differed from one marker to the other and, within one locus, from one primer combination to the other. Importantly, the various MSA-1 and MSA-2 reference alleles were amplified with the same efficiency. Amplification from reconstituted allele mixtures indicated that at certain allele ratios, the most abundant allele interfered with the amplification of the less abundant one. An analysis of nine isolates collected from patients with acute malaria in Dielmo, Senegal, during a transmission season when the inoculation rate was one infective bite every second night is presented and discussed. All samples contained more than one parasite type. A significant polymorphism was observed for the four markers. Novel TaqI restriction fragment length polymorphisms were found for the TRAP gene, and TRAP gene typing alone allowed a distinction between the various isolates. MSA-1 and MSA-2 gave multiple band patterns specific for each sample.
我们报告了一项关于利用少数高度多态性标记物（MSA-1、MSA-2、TRAP和CS）对野外恶性疟原虫分离株进行PCR分型的限制因素分析。我们表明这些反应对恶性疟原虫物种具有特异性。检测阈值（通过溴化乙锭检测可见条带所需的最低寄生虫数量）因标记物不同而不同，并且在一个位点内，因引物组合不同而不同。重要的是，各种MSA-1和MSA-2参考等位基因以相同的效率进行扩增。从重组等位基因混合物中扩增表明，在某些等位基因比例下，最丰富的等位基因会干扰较不丰富等位基因的扩增。本文展示并讨论了在塞内加尔迪耶尔莫一个传播季节从急性疟疾患者中收集的9株分离株的分析结果，该季节的接种率为每两晚一次感染性叮咬。所有样本都包含不止一种寄生虫类型。在这四个标记物上观察到显著的多态性。在TRAP基因中发现了新的TaqI限制性片段长度多态性，仅TRAP基因分型就可以区分各种分离株。MSA-1和MSA-2产生了每个样本特有的多条带模式。
