疟疾病媒中华按蚊（双翅目疟蚊科）中与拟除虫菊酯抗性有关的 P450 基因的鉴定和功能验证。

IF 2.1 3区医学 Q2 PARASITOLOGY

Acta tropica Pub Date : 2024-09-27 DOI:10.1016/j.actatropica.2024.107413

Ying-Ao Guo, Feng-Ling Si, Bao-Zhu Han, Liang Qiao, Bin Chen

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引用次数: 0

摘要

细胞色素 P450 单氧化酶（P450s）是一个多功能蛋白质超家族，是三大类解毒酶之一。然而，对拟除虫菊酯抗性的中华按蚊种群中的 P450 基因的多样性和功能尚未进行充分的研究。本研究利用RNA-seq技术系统地筛选了三个野外除虫菊酯抗性种群（AH-FR、CQ-FR、YN-FR）和一个实验室抗性株系（WX-LR）在发育阶段、组织和血餐后与实验室易感株系（WX-LS）相比与除虫菊酯抗性相关的P450基因。重要的是，利用 RT-qPCR 验证了明显上调的 P450s 的表达，并利用四个实验室除虫菊酯抗性菌株（WX-LR、AH-LR、CQ-LR、YN-LR），通过 RNA 干扰确定了所选 P450s 在除虫菊酯解毒中的功能。在至少一个田间抗性种群中，有 16 个 P450 基因明显上调，在不同发育阶段、组织或血餐后，有 44 个基因明显上调。利用 RT-qPCR 技术，共选择了 19 个 P450 基因来验证它们与拟除虫菊酯抗性的关系，其中四个基因（AsCYP6P3v1、AsCYP6P3v2、AsCYP9J10 和 AsCYP9K1）在实验室拟除虫菊酯抗性菌株中表现出明显的上调。通过RNAi方法，敲除这四个基因均能显著降低拟除虫菊酯抗性，并使中华鳖的死亡率增加57.19%（AsCYP6P3v1和AsCYP6P3v2基因敲除组）、38.39%（AsCYP9K1基因敲除组）和48.87%（AsCYP9J10基因敲除组），从而确定了这四个基因的拟除虫菊酯解毒功能。该研究首次揭示了P450基因的多样性，并提供了4个P450在中华蚁除虫菊酯解毒过程中的功能证据，加深了我们对除虫菊酯抗性机理的认识，对除虫菊酯抗性检测和监控具有潜在价值。

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Identification and functional validation of P450 genes associated with pyrethroid resistance in the malaria vector Anopheles sinensis (Diptera Culicidae)

Cytochrome P450 monooxygenases (P450s), a multifunctional protein superfamily, are one of three major classes of detoxification enzymes. However, the diversity and functions of P450 genes from pyrethroid-resistant populations of Anopheles sinensis have not been fully explored. In this study, P450 genes associated with pyrethroid resistance were systematically screened using RNA-seq in three field pyrethroid-resistant populations (AH-FR, CQ-FR, YN-FR) and one laboratory resistant strain (WX-LR) at developmental stages, tissues, and post blood-meal in comparison to the laboratory susceptible strain (WX-LS) in An. sinensis. Importantly, the expression of significantly upregulated P450s was verified using RT-qPCR, and the function of selected P450s in pyrethroid detoxification was determined with RNA interference using four laboratory pyrethroid-resistant strains (WX-LR, AH-LR, CQ-LR, YN-LR). Sixteen P450 genes were significantly upregulated in at least one field-resistant population, and 44 were significantly upregulated in different developmental stages, tissues or post blood-meal. A total of 19 P450s were selected to verify their association with pyrethroid resistance, and four of them (AsCYP6P3v1, AsCYP6P3v2, AsCYP9J10, and AsCYP9K1) demonstrated significant upregulation in laboratory pyrethroid-resistant strains using RT-qPCR. Knockdown of these four genes all significantly reduced pyrethroid resistance and increased the mortality by 57.19% (AsCYP6P3v1 and AsCYP6P3v2 knockdown group), 38.39% (AsCYP9K1 knockdown group) and 48.87% (AsCYP9J10 knockdown group) in An. sinensis by RNAi, which determined the pyrethroid detoxification function of these four genes. This study revealed the diversity of P450 genes and provided functional evidence for four P450s in pyrethroid detoxification in An. sinensis for the first time, which increases our understanding of the pyrethroid resistance mechanism, and is of potential value for pyrethroid resistance detection and surveillance.

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来源期刊

Acta tropica 医学-寄生虫学

CiteScore

5.40

自引率

11.10%

发文量

383

审稿时长

37 days

期刊介绍： Acta Tropica, is an international journal on infectious diseases that covers public health sciences and biomedical research with particular emphasis on topics relevant to human and animal health in the tropics and the subtropics.