巴西亚马孙西部地区间日疟原虫分离株的体外氯喹抗药性。
In vitro chloroquine resistance for Plasmodium vivax isolates from the Western Brazilian Amazon.
机构信息
Fundação de Medicina Tropical Dr, Heitor Vieira Dourado, Av, Pedro Teixeira, 25, Dom Pedro, Manaus, AM 69040-000, Brazil.
出版信息
Malar J. 2013 Jul 3;12:226. doi: 10.1186/1475-2875-12-226.
BACKGROUND
Chloroquine (CQ) and primaquine (PQ) are still the drugs of choice to treat Plasmodium vivax malaria in many endemic areas, Brazil included. There is in vivo evidence for the P. vivax resistance to CQ in the Brazilian Amazon, where the increase in the proportion of P. vivax malaria parallels the increase of unusual clinical complications related to this species. In this study, in vitro CQ and mefloquine (MQ)-susceptibility of P. vivax isolates from the Western Brazilian Amazon was tested using the double-site enzyme-linked lactate dehydrogenase immunodetection (DELI) assay.
METHODS
A total of 112 P. vivax isolates were tested in vitro for CQ-susceptibility and out of these 47 were also tested for MQ-susceptibility. The DELI assay was used to detect P. vivax growth at 48-hour short-term culture in isolates with ring stages ranging from 50 to %. Each isolate was tested in triplicate and geometric means of IC50's was obtained. Nineteen isolates were genetically characterized for pvdhfr, pvmrp1, pvmdr1 and pvdhps candidate genes likely related to CQ resistance (10 with IC50<40 nM and 9 with IC50 >100 nM).
RESULTS
Twelve out of 112 isolates were considered resistant to CQ, resulting in 10.7% (IC95% 5.0-16.4), while 3 out of 47 (6.4%; IC95% 0.0-12.8) were resistant to MQ. A discrete correlation was observed between IC50's of CQ and MQ (Spearman=0.294; p=0.045). For pvdhps gene, a non-synonymous mutation was found at codon 382 (S→C) in 5/8 CQ-sensitive samples and 1/9 CQ-resistant samples (p=0.027). The other molecular markers were not associated to CQ-susceptibility.
CONCLUSIONS
In vitro CQ-resistance estimated in this study, estimated by the DELI test, was very similar to that observed in clinical trials, suggesting that in vitro procedures developed by capable local laboratories are useful in the surveillance of CQ-resistance in the Amazon; concurrent Amazon P. vivax strains with both CQ and MQ resistance may be common; and a non-synonymous mutation at pvdhps codon 382 (S→C) was associated to in vitro susceptibility to CQ, needing further studies to be confirmed.
背景
在许多流行地区，包括巴西在内，氯喹（CQ）和伯氨喹（PQ）仍然是治疗间日疟原虫疟疾的首选药物。在巴西亚马逊地区，已经有体内证据表明间日疟原虫对 CQ 产生了耐药性，而该地区间日疟原虫疟疾的比例增加与该物种相关的不寻常临床并发症的增加是平行的。在这项研究中，使用双位点酶联乳酸脱氢酶免疫检测（DELI）试验测试了来自巴西西部亚马逊地区的间日疟原虫分离株的体外 CQ 和甲氟喹（MQ）敏感性。
方法
对 112 株间日疟原虫分离株进行了体外 CQ 敏感性检测，其中 47 株还进行了 MQ 敏感性检测。使用 DELI 试验检测环早期为 50-70%的分离株在 48 小时短期培养中的疟原虫生长情况。每个分离株检测 3 次，并获得 IC50 的几何平均值。对 19 株可能与 CQ 耐药相关的候选基因 pvdhfr、pvmrp1、pvmdr1 和 pvdhps 进行了基因特征分析（10 株 IC50<40 nM，9 株 IC50 >100 nM）。
结果
112 株分离株中有 12 株被认为对 CQ 耐药，耐药率为 10.7%（95%CI5.0-16.4），而 47 株中有 3 株（6.4%；95%CI0.0-12.8）对 MQ 耐药。CQ 和 MQ 的 IC50 之间存在明显的相关性（Spearman=0.294；p=0.045）。在 pvdhps 基因中，5/8 株 CQ 敏感样本和 1/9 株 CQ 耐药样本在密码子 382 处发现非同义突变（S→C）（p=0.027）。其他分子标记与 CQ 敏感性无关。
结论
本研究采用 DELI 试验测定的体外 CQ 耐药性与临床试验观察到的耐药性非常相似，表明由有能力的当地实验室开发的体外程序可用于监测亚马逊地区的 CQ 耐药性；同时具有 CQ 和 MQ 耐药性的亚马逊地区间日疟原虫株可能很常见；pvdhps 密码子 382 处的非同义突变（S→C）与 CQ 的体外敏感性相关，需要进一步研究证实。
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