Molecular and cellbiological studies on gametocytogenesis of a malaria parasite, Plasmodium falciparum

以恶性疟原虫野生分离株为材料，从细胞生物学和分子生物学的角度研究了恶性疟原虫配子体发生的诱导机制。野生菌株配子体发生的诱导：研究了从缅甸和印度尼西亚采集的野生菌株的配子体发生诱导方法。当寄生虫密度达到约5%时，观察到配子体发生，然后在比正常培养条件高3-4倍的RBC浓度下培养。适合于体外培养的野生菌株均能形成配子体，转化率达50-70%。缺糖诱导配子体生成：由于配子体生成是在较高的RBC浓度下培养诱导的，推测营养缺乏，特别是葡萄糖的缺乏，可能在触发这种现象中起重要作用。事实上，在培养液中添加葡萄糖抑制了配子体发生的诱导。此外，在正常培养条件下，添加2-脱氧葡萄糖也能诱导其生长，但寄生虫血症明显减少。这些结果表明，葡萄糖缺乏可能在诱导配子细胞发生中起重要作用。已建立菌株的配子体诱导试验：利用Fcr-3和K-L菌株，在野生菌株中进行了类似的配子体诱导试验，但没有形成配子体，这表明在这些菌株中可能缺失了诱导配子体发生的重要基因。诱导配子体发生的主控基因(S)的鉴定：用减法从配子体发生诱导和非诱导的寄生虫中分离到可能调节配子体发生的主控基因(S)。然而，消减后得到的所有克隆都只是热休克蛋白基因，并没有检测到预期的基因，如cAMP依赖蛋白激酶。

Induction mechanism of gametocytogenesis in a malaria parasite, Plasmodium falciparum was investigated cell-biologically and molecularly, using wild isolates.1. Induction of gametocytogenesis in wild isolates : Induction methods of gametocytogenesis was studied, using wild isolates collected from Myanmar and Indonesia.Gametocytogenesis was observed when the parasite density reached to ca. 5% and then they were cultivated at 3-4 times higher concentrations of RBC than normal culture condition. All wild isolates adapted for in vitro cultivation were capable in forming gametocytes, and their transformation rates reached to 50-70%.2. Induction ofgametocytogenesis by glucose deficiency : As gametocytogenesis was induced by cultivation at higher RBC concentration, it was speculated that nutrient deficiency, particularly glucose, may play important role in triggering this phenomenon. In fact, addition of glucose into the medium inhibited the induction of gametocytogenesis. Furthermore, addition of 2-deoxy glucose was capable in inducing it under a normal culture condition, although a great decrease ofparasitaemia was observed. These results indicated that glucose deficiency might play an important role in inducing gametocytogenesis.3. Trials of induction ofgametocytogenesis in established strains : Using FCR-3 and K-l strains, induction of gametocytogenesis was examined as like in wild isolates, but no gametocyte was formed, suggesting that important genes for induction of gametocytogenesis might be deleted in these strains.4. Identification of master gene(s) for induction of gametocytogenesis : Using a subtraction method between mRNA isolated from gametocyogenesis-induced and non-induced parasites, master gene(s) which may regulate gametocytogenesis was investigated. However, all clones obtained after the subtraction were only heat shock protein genes, and expected genes, such as cAMP-dependent kinases, were not detected.