Figueiredo Luiza Almeida, Rebouças Thais Fuscaldi, Ferreira Sebastião Rodrigo, Rodrigues-Luiz Gabriela Flavia, Miranda Rodrigo Cambraia, Araujo Ricardo Nascimento, Fujiwara Ricardo Toshio
Department of Parasitology, Institute of Biological Sciences, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Center of Agrarian and Biological Sciences, University of Vale do Acaraú, Sobral, Ceará, Brazil.
PLoS One. 2018 Feb 23;13(2):e0192995. doi: 10.1371/journal.pone.0192995. eCollection 2018.
While diseases caused by nematodes remains a considerable drawback for the livestock, agriculture and public health, anthelmintics drug resistance has been observed over the past years and is a major concern for parasite control. Ivermectin, initially considered as a highly potent drug, currently presents a reduced anti-helminthic efficacy, which is influenced by expression of several ATP-binding cassette transporters (ABC), among them the P-glycoproteins (Pgps). Here we present some evidences of Pgps dominance during Ivermectin resistance/susceptibility using Pgps double silencing in C. elegans and the phylogenetic relationship of Pgps among nematodes, which strengthen the use of this model for study of drug resistance in nematodes. Firstly, we evaluated the quantitative gene expression of 12 out the 15 known Pgps from resistant and WT strains of C. elegans, we demonstrated the upregulation of Pgps 12 and 13 and downregulation of all remaining Pgps in ivermectin resistant strain. By using an RNAi loss-of-function approach we observed that Pgp 12 gene silencing reverts the resistance phenotype to ivermectin, while Pgp 4 gene silencing does not alter the resistance phenotype but induces a resistance in wild type strain. Interestingly, the dual silencing of Pgp 12 and Pgp 4 expression demonstrates the dominance of phenotype promoted by Pgp 12 silencing. Finally, in silico analysis reveals a close relationship between Pgps from C. elegans and several nematodes parasites. Taken together, our results indicate that Pgp 12 is crucial for the resistance to ivermectin and thus a good candidate for further studies aiming to develop specific inhibitors to this transporter, allowing the continuous use of ivermectin to control the burden on animal and human health inflicted by nematode parasites globally.
虽然线虫引起的疾病仍然是畜牧业、农业和公共卫生的一个重大缺陷，但在过去几年中已观察到驱虫药耐药性，这是寄生虫控制的一个主要问题。伊维菌素最初被认为是一种高效药物，目前其抗蠕虫功效降低，这受到几种ATP结合盒转运蛋白（ABC）的表达影响，其中包括P-糖蛋白（Pgps）。在这里，我们通过在秀丽隐杆线虫中对Pgps进行双重沉默以及线虫中Pgps的系统发育关系，展示了Pgps在伊维菌素耐药性/敏感性中的主导地位的一些证据，这加强了该模型在研究线虫耐药性方面的应用。首先，我们评估了秀丽隐杆线虫耐药菌株和野生型菌株中15种已知Pgps中的12种的定量基因表达，我们证明了伊维菌素耐药菌株中Pgps 12和13的上调以及所有其余Pgps的下调。通过使用RNAi功能丧失方法，我们观察到Pgp 12基因沉默使对伊维菌素的耐药表型恢复，而Pgp 4基因沉默不会改变耐药表型，但会在野生型菌株中诱导耐药性。有趣的是，Pgp 12和Pgp 4表达的双重沉默证明了Pgp 12沉默促进的表型的主导地位。最后，计算机分析揭示了秀丽隐杆线虫的Pgps与几种线虫寄生虫之间的密切关系。综上所述，我们的结果表明Pgp 12对伊维菌素耐药性至关重要，因此是进一步研究开发该转运蛋白特异性抑制剂的良好候选者，从而能够持续使用伊维菌素控制全球线虫寄生虫对动物和人类健康造成的负担。
