Determining physiological role(s) of Resistance-Nodulation-Division efflux pumps of Acinetobacter baumannii

确定鲍曼不动杆菌抵抗结瘤分裂外排泵的生理作用

• 批准号: RGPIN-2021-02902
• 负责人: Kumar, Ayush
• 金额: $ 4.23万
• 依托单位: University of Manitoba
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=760962
• 关键词: Determining; physiological; role; Resistance; Nodulation

Acinetobacter baumannii is a Gram-negative pathogen and an organism of significant interest due to its high resistance to antibiotics. Although A. baumannii's resistance to antibiotics is a result of a combination of various acquired and intrinsic factors, proton-gradient-dependent pumps belonging to the Resistance-Nodulation-division (RND) family are considered the most important factors contributing to the intrinsic antibiotic resistance of A. baumannii. RND pumps are an ancient family of proteins and their homologs have been reported across all three kingdoms. RND pumps have been most studied in Gram-negative bacteria primarily due to their ability to pump out a variety of antimicrobials across the two membranes. Due to their ancient nature, there is a consensus that the efflux of antibiotics is not their primary role, however physiological function(s) of RND pumps remain unknown. In this application, we propose to study two RND pumps of A. baumannii, AdeIJK and AdeFGH to determine their natural functions. Preliminary data from our laboratory suggest that fatty acid perturbations can result in the upregulation of AdeIJK pump. Further, we have also observed that the deletion of AdeN, the transcriptional repressor of AdeIJK pump, results in an altered expression of almost 250 different genes. This suggests that perhaps AdeIJK pump is part of a response in A. baumannii that results from the loss of AdeN. Identifying and characterizing genes that are co-regulated with AdeIJK will provide important clues about the natural function(s) of AdeIJK pump. Similarly, we have isolated mutants of A. baumannii that overexpress AdeFGH pump. These mutants contain a mutation in csrA that encodes for the Carbon Storage Regulator protein. CsrA, an mRNA binding protein, represses several stationary phase processes and activates exponential-phase functions in E. coli. It also modulates the regulation of genes involved in central carbon metabolism, secondary metabolite metabolism, motility, biofilm formation, as well as virulence. We are therefore proposing to study the link between central carbon as well nitrogen metabolism and the expression of AdeFGH in A. baumannii. We will employ a unique approach and carry out the proposed studies in an antibiotic susceptible environmental isolate of A. baumannii. Our approach of carrying out these studies in an environmental isolate of A. baumannii is likely to provide more relevant findings on the physiological functions of AdeIJK and AdeFGH pumps. This work will lead to a better understanding of how the expression of RND efflux pumps may be influenced by metabolic pathways in A. baumannii.

鲍曼尼杆菌是一种革兰氏阴性的病原体，并且由于其对抗生素的耐药性高而引起了浓厚兴趣。尽管鲍曼尼曲霉对抗生素的耐药性是各种可获得和内在因素的组合，但属于抗药性 - 节点 - 族（RND）家族的质子梯度依赖性泵被认为是导致内在性抗生素耐药性的最重要因素A. Baumannii。 RND泵是古老的蛋白质家族，在所有三个王国中都有报道。 RND泵在革兰氏阴性细菌中的研究主要是由于它们能够在两个膜上抽出各种抗菌剂的能力。由于其古老的性质，人们达成共识，抗生素的排出不是其主要作用，但是RND泵的生理功能仍然未知。在此应用中，我们建议研究鲍曼尼（A. Baumannii），adeijk和adefgh的两个RND泵，以确定它们的自然功能。我们实验室的初步数据表明，脂肪酸扰动可能导致adeijk泵的上调。此外，我们还观察到，Adeijk泵的转录阻遏物Aden的缺失导致近250种不同基因的表达改变。这表明，Adeijk泵也许是A. baumannii响应的一部分，这是由于Aden的损失而导致的。识别和表征与Adeijk共同调节的基因将提供有关Adeijk泵的自然功能的重要线索。同样，我们具有过表达Adefgh泵的鲍曼尼曲霉的孤立突变体。这些突变体包含CSRA中的突变，该突变编码为碳储存器蛋白。 CSRA是一种mRNA结合蛋白，抑制了几个固定相过程，并激活大肠杆菌中的指数型函数。它还调节了中央碳代谢，继发代谢物代谢，运动，生物膜形成以及毒力的调节。因此，我们提议研究中央碳以及氮代谢与鲍曼尼曲霉中ADEFGH的表达之间的联系。我们将采用一种独特的方法，并在鲍曼尼（A. baumannii）的抗生素易感环境分离株中进行拟议的研究。我们在鲍曼尼a。A.baumannii的环境分离中进行这些研究的方法可能会提供有关Adeijk和Adefgh泵的生理功能的更相关的发现。这项工作将使您更好地了解RND外排泵的表达如何受鲍曼尼a。代谢途径的影响。