Experimental Induction of Paromomycin Resistance in Antimony-Resistant Strains of L. donovani: Outcome Dependent on In Vitro Selection Protocol
Experimental Induction of Paromomycin Resistance in Antimony-Resistant Strains of L. donovani: Outcome Dependent on In Vitro Selection Protocol
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DOI：
10.1371/journal.pntd.0001664
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发表时间：
2012-05-01
影响因子：
3.8
通讯作者：
Maes, Louis
中科院分区：
文献类型：
--
作者：
Hendrickx, Sarah;da Luz, Raquel Andrea Inocencio;Maes, Louis
巴龙霉素（PMM）最近被引入印度用于治疗内脏利什曼病。虽然尚未报告临床耐药性，但应保持积极警惕。本体外研究比较了前鞭毛体和细胞内无鞭毛体实验性PMM抗性诱导的结果和稳定性。克隆抗锑L.来自印度和尼泊尔的donovani现场分离株暴露于逐步增加浓度的PMM（高达500 μ M），无论是作为前鞭毛体或细胞内无鞭毛体。克隆得到的一种耐药菌株，并通过作为前鞭毛体在体外无药物传代20周或在金黄仓鼠体内单次传代来检查耐药性的稳定性。前鞭毛体中的抗性选择需要大约25周才能达到不影响正常生长的最大97 μ M包涵体水平。亲本和所选菌株之间的IC 50值的比较显示，印度菌株的抗性为9至11倍，尼泊尔菌株的抗性为3至5倍，由此抗性表型也保持在无鞭毛体水平。与亲本菌株（IC 50 = 45 μ M）相比，对细胞内无鞭毛体施加PMM压力仅在两个选择周期后产生抗性（IC 50 = 199 μ M）。在无鞭毛体诱导的菌株/克隆中，仅在无鞭毛体中观察到较低的PMM亲和性，而在前鞭毛体中根本没有。这种抗性表型在作为前鞭毛体连续体外传代20周和在仓鼠体内单次传代后保持稳定。这项研究清楚地表明，一个不同的PMM抗性表型，获得药物选择是否适用于前鞭毛体或细胞内无鞭毛体。这些发现可能具有重要的相关性，耐药机制的调查和耐药发展的可能性和检测领域。
Paromomycin (PMM) has recently been introduced for treatment of visceral leishmaniasis in India. Although no clinical resistance has yet been reported, proactive vigilance should be warranted. The present in vitro study compared the outcome and stability of experimental PMM-resistance induction on promastigotes and intracellular amastigotes. Cloned antimony-resistant L. donovani field isolates from India and Nepal were exposed to stepwise increasing concentrations of PMM (up to 500 mu M), either as promastigotes or intracellular amastigotes. One resulting resistant strain was cloned and checked for stability of resistance by drug-free in vitro passage as promastigotes for 20 weeks or a single in vivo passage in the golden hamster. Resistance selection in promastigotes took about 25 weeks to reach the maximal 97 mu M inclusion level that did not affect normal growth. Comparison of the IC50 values between the parent and the selected strains revealed a 9 to 11-fold resistance for the Indian and 3 to 5-fold for the Nepalese strains whereby the resistant phenotype was also maintained at the level of the amastigote. Applying PMM pressure to intracellular amastigotes produced resistance after just two selection cycles (IC50 = 199 mu M) compared to the parent strain (IC50= 45 mu M). In the amastigote-induced strains/clones, lower PMM susceptibilities were seen only in amastigotes and not at all in promastigotes. This resistance phenotype remained stable after serial in vitro passage as promastigote for 20 weeks and after a single in vivo passage in the hamster. This study clearly demonstrates that a different PMM-resistance phenotype is obtained whether drug selection is applied to promastigotes or intracellular amastigotes. These findings may have important relevance to resistance mechanism investigations and the likelihood of resistance development and detection in the field.