Antisense locked nucleic acids targeting agrA inhibit quorum sensing and pathogenesis of community-associated methicillin-resistant Staphylococcus aureus
Antisense locked nucleic acids targeting agrA inhibit quorum sensing and pathogenesis of community-associated methicillin-resistant Staphylococcus aureus
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DOI：
10.1111/jam.13321
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发表时间：
2017-01-01
影响因子：
4
通讯作者：
Luo, X.
中科院分区：
文献类型：
--
作者：
Da, F.;Yao, L.;Luo, X.
目的：社区相关耐甲氧西林金黄色葡萄球菌(CA-MRSA)毒力大，常与非院内皮肤软组织感染相关，主要受辅助基因调控(AGR)群体感应系统(QS)控制。本研究(Kff)(3)针对agra基因设计了K肽共轭锁定核酸(PLNAs)，用于抑制agr活性和阻断CA-MRSA的致病性。方法与结果：在预测了agra基因二级结构后，设计并合成了两个PLNAs。采用生长曲线法测定其对细菌生长的影响。用RT-qPCR、溶血实验、乳酸脱氢酶释放实验和趋化实验评价PLNAs对AGR-QS的抑制作用。采用小鼠皮肤感染模型，检测PLNAs的体内保护作用。这些PLNAs在体外均未发现抑菌或杀菌作用。然而，PLNA34在USA300LAC株中表现出很强的抑制AGRA和效应分子RNAIII表达的能力。此外，PLNA34还抑制了AGR上调的毒力基因的表达，包括Hla、Psmα、Psmβ和Pvl。PLNA34处理的细菌上清液的溶血活性以及裂解和招募中性粒细胞的能力也显著降低。在CA-MRSA小鼠皮肤感染模型中，PLNA34显示出较高的保护作用。结论：抗AgA PLNA34能有效抑制AgR QS，抑制CA-MRSA的致病作用。
Aim: Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is commonly associated with nonnosocomial skin and soft tissue infections due to its virulence, which is mainly controlled by the accessory gene regulator (agr) quorum sensing (QS) system. In this study (KFF)(3)K peptide-conjugated locked nucleic acids (PLNAs) targeting agrA mRNA were developed to inhibit agr activity and arrest the pathogenicity of CA-MRSA.Methods and Results: Two PLNAs were designed, and synthesized, after predicting the secondary structure of agrA mRNA. The influence on bacterial growth was tested using a growth curve assay. RT-qPCR, haemolysis assay, lactate dehydrogenase release assay and chemotaxis assay were used to evaluate the effects of the PLNAs on inhibiting agr QS. A mouse skin infection model was employed to test the protective effect of the PLNAs in vivo. None of the PLNAs were found to be bacteriostatic or bactericidal in vitro. However, one PLNA, PLNA34, showed strong ability to suppress expression of agrA and the effector molecule RNAIII in USA300 LAC strain. Furthermore, PLNA34 inhibited the expression of virulence genes that are upregulated by agr, including hla, psm alpha, psm beta and pvl. The haemolytic activity of the supernatants from PLNA34-treated bacteria was also dramatically reduced, as well as the capacity to lyse and recruit neutrophils. Moreover, PLNA34 showed high levels of protection in the CA-MRSA mouse skin infection model.Conclusions: The anti-agrA PLNA34 can effectively inhibit the agr QS and suppress CA-MRSA pathogenicity.Significance and Impact of the Study: agrA is a promising target for the development of antisense oligonucleotides to block agr QS.