Mkulama Mtawa A P, Chishimba Sandra, Sikalima Jay, Rouse Petrica, Thuma Philip E, Mharakurwa Sungano
The Malaria Institute at Macha, Namwala Road, Choma, Zambia.
Malar J. 2008 May 21;7:87. doi: 10.1186/1475-2875-7-87.
In Zambia the first-line treatment for uncomplicated malaria is artemisinin combination therapy (ACT), with artemether-lumefantrine currently being used. However, the antifolate regimen, sulphadoxine-pyrimethamine (SP), remains the treatment of choice in children weighing less than 5 kg and also in expectant mothers. SP is also the choice drug for intermittent preventive therapy in pregnancy and serves as stand-by treatment during ACT stock outs. The current study assessed the status of Plasmodium falciparum point mutations associated with antifolate drug resistance in the area around Macha.
A representative sample of 2,780 residents from the vicinity of Macha was screened for malaria by microscopy. At the same time, blood was collected onto filter paper and dried for subsequent P. falciparum DNA analysis. From 188 (6.8%) individuals that were thick film-positive, a simple random sub-set of 95 P. falciparum infections were genotyped for DHFR and DHPS antifolate resistance mutations, using nested PCR and allele-specific restriction enzyme digestion.
Plasmodium falciparum field samples exhibited a high prevalence of antifolate resistance mutations, including the DHFR triple (Asn-108 + Arg-59 + Ile-51) mutant (41.3%) and DHPS double (Gly-437 + Glu-540) mutant (16%). The quintuple (DHFR triple + DHPS double) mutant was found in 4 (6.5%) of the samples. Levels of mutated parasites showed a dramatic escalation, relative to previous surveys since 1988. However, neither of the Val-16 and Thr-108 mutations, which jointly confer resistance to cycloguanil, was detectable among the human infections. The Leu-164 mutation, associated with high grade resistance to both pyrimethamine and cycloguanil, as a multiple mutant with Asn-108, Arg-59 and (or) Ile-51, was also absent.
This study points to escalating levels of P. falciparum antifolate resistance in the vicinity of Macha. Continued monitoring is recommended to ensure timely policy revisions before widespread resistance exacts a serious public health toll.
在赞比亚，单纯性疟疾的一线治疗方法是青蒿素联合疗法（ACT），目前使用的是蒿甲醚-本芴醇。然而，抗叶酸方案，即磺胺多辛-乙胺嘧啶（SP），仍然是体重不足5公斤的儿童以及孕妇的首选治疗方法。SP也是孕期间歇性预防治疗的首选药物，并且在ACT药物短缺期间作为备用治疗药物。本研究评估了马查周边地区与抗叶酸药物耐药性相关的恶性疟原虫点突变情况。
通过显微镜检查对来自马查附近地区的2780名居民的代表性样本进行疟疾筛查。与此同时，采集血液到滤纸上并干燥，以便后续进行恶性疟原虫DNA分析。从188名（6.8%）厚血膜阳性个体中，通过简单随机抽样选取95例恶性疟原虫感染样本，使用巢式PCR和等位基因特异性限制性内切酶消化法对二氢叶酸还原酶（DHFR）和二氢蝶酸合酶（DHPS）抗叶酸耐药性突变进行基因分型。
恶性疟原虫现场样本显示抗叶酸耐药性突变的发生率很高，包括DHFR三重突变（Asn-108 + Arg-59 + Ile-51）（41.3%）和DHPS双重突变（Gly-437 + Glu-540）（16%）。四重突变（DHFR三重突变+DHPS双重突变）在4例（6.5%）样本中被发现。相对于自1988年以来的先前调查，突变寄生虫的水平呈现出急剧上升。然而，在人类感染中未检测到共同赋予对环氯胍耐药性的Val-16和Thr-108突变。与对乙胺嘧啶和环氯胍的高度耐药性相关的Leu-164突变，作为与Asn-108、Arg-59和（或）Ile-51的多重突变，也未出现。
本研究表明马查周边地区恶性疟原虫对叶酸拮抗剂的耐药水平在不断上升。建议持续监测，以确保在广泛耐药造成严重公共卫生损失之前及时修订政策。
