抗生素治疗及Mex B外排转运蛋白基因表达对铜绿假单胞菌生物膜耐药的影响

Evan Kello, Rochelle Greenberg, Weiqi Li, Shaya Polansky, Roberto Maldonado, Yakov Peter, P. Basu
{"title":"抗生素治疗及Mex B外排转运蛋白基因表达对铜绿假单胞菌生物膜耐药的影响","authors":"Evan Kello, Rochelle Greenberg, Weiqi Li, Shaya Polansky, Roberto Maldonado, Yakov Peter, P. Basu","doi":"10.3390/applmicrobiol3030049","DOIUrl":null,"url":null,"abstract":"Pseudomonas aeruginosa, a Gram-negative, rod-shaped bacterium, holds a prominent position as an antibiotic-resistant priority pathogen, according to the World Health Organization. Particularly prevalent in healthcare settings, this bacterium acts as an opportunistic pathogen, causing nosocomial infections. The significant antibiotic resistance observed in P. aeruginosa is multifactorial, encompassing intrinsic, acquired, and adaptive resistance mechanisms. The present study aims to explore specific RND-type efflux pump genes implicated in the acquisition of antibiotic resistances during the transition of P. aeruginosa PAO1 from its planktonic state to the more formidable and resistant biofilm form. This investigation is centered on MexB, a prominent RND-type efflux pump in P. aeruginosa. Our research is focused on MexB, a highly significant component characterized by its broad substrate specificity, primary function as the primary efflux pump, substantial expression levels, and notable clinical implications. Considering MexB’s critical role in expelling various clinically relevant antimicrobial agents and its significant contribution to multidrug resistance, our study aims to evaluate the comparative efficacy of three distinct antibiotic categories, namely, Ofloxacin (OFX), Tobramycin (TOB), and Ceftazidime (CAZ), in regulating the expression levels of identified multidrug efflux pump genes associated with the biofilm’s ability to remove antibiotics from bacterial cells. Expression analysis of efflux transporter genes in P. aeruginosa was performed by isolating total RNA from both planktonic and biofilm samples, both untreated and treated with Tobramycin (TOB), Ofloxacin (OFX), and Ceftazidime (CAZ). Real-time quantitative reverse transcriptase PCR was employed to investigate changes in the expression levels of MexA, MexB, MexX, MexY, OprM, and RPSL genes in the collected samples. In the absence of antibiotic treatment, the MexB efflux pump gene exhibited higher expression compared to other efflux pump genes in the biofilm’s state, supporting its involvement in multidrug resistance when active. To further explore the role of the MexB gene in antibiotic resistance, P. aeruginosa was cultured in both planktonic and biofilm forms while simultaneously treating them with TOB, OFX, and CAZ. Among the three antibiotics employed, OFX demonstrated superior efficacy in inhibiting the growth of biofilms by downregulating the expression of the Mex B efflux pump gene in P. aeruginosa, thereby enhancing its susceptibility to OFX. TOB yielded comparable outcomes to OFX, albeit with a slightly lesser extent of Mex B expression reduction. Conversely, CAZ exhibited ineffectiveness in reducing MexB gene expression in both biofilm and planktonic forms of the organism, rendering it incapable of eradicating the pathogen.","PeriodicalId":8080,"journal":{"name":"Applied microbiology","volume":"58 1","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2023-07-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in Pseudomonas Aeruginosa Biofilms\",\"authors\":\"Evan Kello, Rochelle Greenberg, Weiqi Li, Shaya Polansky, Roberto Maldonado, Yakov Peter, P. Basu\",\"doi\":\"10.3390/applmicrobiol3030049\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"Pseudomonas aeruginosa, a Gram-negative, rod-shaped bacterium, holds a prominent position as an antibiotic-resistant priority pathogen, according to the World Health Organization. Particularly prevalent in healthcare settings, this bacterium acts as an opportunistic pathogen, causing nosocomial infections. The significant antibiotic resistance observed in P. aeruginosa is multifactorial, encompassing intrinsic, acquired, and adaptive resistance mechanisms. The present study aims to explore specific RND-type efflux pump genes implicated in the acquisition of antibiotic resistances during the transition of P. aeruginosa PAO1 from its planktonic state to the more formidable and resistant biofilm form. This investigation is centered on MexB, a prominent RND-type efflux pump in P. aeruginosa. Our research is focused on MexB, a highly significant component characterized by its broad substrate specificity, primary function as the primary efflux pump, substantial expression levels, and notable clinical implications. Considering MexB’s critical role in expelling various clinically relevant antimicrobial agents and its significant contribution to multidrug resistance, our study aims to evaluate the comparative efficacy of three distinct antibiotic categories, namely, Ofloxacin (OFX), Tobramycin (TOB), and Ceftazidime (CAZ), in regulating the expression levels of identified multidrug efflux pump genes associated with the biofilm’s ability to remove antibiotics from bacterial cells. Expression analysis of efflux transporter genes in P. aeruginosa was performed by isolating total RNA from both planktonic and biofilm samples, both untreated and treated with Tobramycin (TOB), Ofloxacin (OFX), and Ceftazidime (CAZ). Real-time quantitative reverse transcriptase PCR was employed to investigate changes in the expression levels of MexA, MexB, MexX, MexY, OprM, and RPSL genes in the collected samples. In the absence of antibiotic treatment, the MexB efflux pump gene exhibited higher expression compared to other efflux pump genes in the biofilm’s state, supporting its involvement in multidrug resistance when active. To further explore the role of the MexB gene in antibiotic resistance, P. aeruginosa was cultured in both planktonic and biofilm forms while simultaneously treating them with TOB, OFX, and CAZ. Among the three antibiotics employed, OFX demonstrated superior efficacy in inhibiting the growth of biofilms by downregulating the expression of the Mex B efflux pump gene in P. aeruginosa, thereby enhancing its susceptibility to OFX. TOB yielded comparable outcomes to OFX, albeit with a slightly lesser extent of Mex B expression reduction. Conversely, CAZ exhibited ineffectiveness in reducing MexB gene expression in both biofilm and planktonic forms of the organism, rendering it incapable of eradicating the pathogen.\",\"PeriodicalId\":8080,\"journal\":{\"name\":\"Applied microbiology\",\"volume\":\"58 1\",\"pages\":\"\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2023-07-06\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Applied microbiology\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.3390/applmicrobiol3030049\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Applied microbiology","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.3390/applmicrobiol3030049","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0

摘要

据世界卫生组织(World Health Organization)称,铜绿假单胞菌(Pseudomonas aeruginosa)是一种革兰氏阴性杆状细菌,是一种具有抗生素耐药性的优先病原体。这种细菌作为一种机会性病原体,在卫生保健环境中尤为普遍,可引起医院感染。在铜绿假单胞菌中观察到的显著抗生素耐药性是多因素的,包括内在的、获得性的和适应性的耐药机制。本研究旨在探索在P. aeruginosa PAO1从浮游状态向更强大和耐药的生物膜形式转变过程中涉及获得抗生素耐药性的特定rnd型外流泵基因。本研究以铜绿假单胞菌中一个突出的rnd型外排泵MexB为中心。我们的研究重点是MexB,这是一种非常重要的成分,其特点是具有广泛的底物特异性、作为主要外排泵的主要功能、大量的表达水平和显著的临床意义。考虑到MexB在驱逐各种临床相关抗菌药物方面的关键作用及其对多药耐药的重要贡献,我们的研究旨在评估三种不同的抗生素类别,即氧氟沙星(OFX),妥布霉素(TOB)和头孢他啶(CAZ),在调节与生物膜从细菌细胞中去除抗生素能力相关的多药外排泵基因表达水平方面的比较功效。通过从浮游和生物膜样品中分离总RNA,对铜绿假单胞菌外排转运蛋白基因进行表达分析,这些样品均未经妥布霉素(TOB)、氧氟沙星(OFX)和头孢他啶(CAZ)处理。采用实时定量逆转录酶PCR检测采集样本中MexA、MexB、MexX、MexY、OprM和RPSL基因的表达水平变化。在没有抗生素治疗的情况下,与其他外排泵基因相比,MexB外排泵基因在生物膜状态下表现出更高的表达,支持其在活性时参与多药耐药。为了进一步探索MexB基因在抗生素耐药性中的作用,我们以浮游和生物膜两种形式培养铜绿假单胞菌,同时用TOB、OFX和CAZ处理它们。在三种抗生素中,OFX通过下调P. aeruginosa中Mex B外排泵基因的表达,从而增强其对OFX的易感性,从而表现出抑制生物膜生长的优越效果。TOB产生的结果与OFX相当,尽管Mex B表达减少的程度略小。相反,CAZ在生物膜和浮游形式的生物体中都表现出降低MexB基因表达的无效,使其无法根除病原体。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in Pseudomonas Aeruginosa Biofilms
Pseudomonas aeruginosa, a Gram-negative, rod-shaped bacterium, holds a prominent position as an antibiotic-resistant priority pathogen, according to the World Health Organization. Particularly prevalent in healthcare settings, this bacterium acts as an opportunistic pathogen, causing nosocomial infections. The significant antibiotic resistance observed in P. aeruginosa is multifactorial, encompassing intrinsic, acquired, and adaptive resistance mechanisms. The present study aims to explore specific RND-type efflux pump genes implicated in the acquisition of antibiotic resistances during the transition of P. aeruginosa PAO1 from its planktonic state to the more formidable and resistant biofilm form. This investigation is centered on MexB, a prominent RND-type efflux pump in P. aeruginosa. Our research is focused on MexB, a highly significant component characterized by its broad substrate specificity, primary function as the primary efflux pump, substantial expression levels, and notable clinical implications. Considering MexB’s critical role in expelling various clinically relevant antimicrobial agents and its significant contribution to multidrug resistance, our study aims to evaluate the comparative efficacy of three distinct antibiotic categories, namely, Ofloxacin (OFX), Tobramycin (TOB), and Ceftazidime (CAZ), in regulating the expression levels of identified multidrug efflux pump genes associated with the biofilm’s ability to remove antibiotics from bacterial cells. Expression analysis of efflux transporter genes in P. aeruginosa was performed by isolating total RNA from both planktonic and biofilm samples, both untreated and treated with Tobramycin (TOB), Ofloxacin (OFX), and Ceftazidime (CAZ). Real-time quantitative reverse transcriptase PCR was employed to investigate changes in the expression levels of MexA, MexB, MexX, MexY, OprM, and RPSL genes in the collected samples. In the absence of antibiotic treatment, the MexB efflux pump gene exhibited higher expression compared to other efflux pump genes in the biofilm’s state, supporting its involvement in multidrug resistance when active. To further explore the role of the MexB gene in antibiotic resistance, P. aeruginosa was cultured in both planktonic and biofilm forms while simultaneously treating them with TOB, OFX, and CAZ. Among the three antibiotics employed, OFX demonstrated superior efficacy in inhibiting the growth of biofilms by downregulating the expression of the Mex B efflux pump gene in P. aeruginosa, thereby enhancing its susceptibility to OFX. TOB yielded comparable outcomes to OFX, albeit with a slightly lesser extent of Mex B expression reduction. Conversely, CAZ exhibited ineffectiveness in reducing MexB gene expression in both biofilm and planktonic forms of the organism, rendering it incapable of eradicating the pathogen.
求助全文
通过发布文献求助,成功后即可免费获取论文全文。 去求助
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信