The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa Biofilms
<i>Pseudomonas aeruginosa</i>, 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, caus...
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MDPI AG
2023-07-01
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| Series: | Applied Microbiology |
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| Online Access: | https://www.mdpi.com/2673-8007/3/3/49 |
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| author | Evan Kello Rochelle Greenberg Weiqi Li Shaya Polansky Roberto Maldonado Yakov Peter Paramita Basu |
| author_facet | Evan Kello Rochelle Greenberg Weiqi Li Shaya Polansky Roberto Maldonado Yakov Peter Paramita Basu |
| author_sort | Evan Kello |
| collection | DOAJ |
| description | <i>Pseudomonas aeruginosa</i>, 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 <i>P. aeruginosa</i> 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 <i>P. aeruginosa</i> 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. <i>aeruginosa</i>. 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 <i>P. aeruginosa</i> 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, <i>P. aeruginosa</i> 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 <i>P. aeruginosa</i>, 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. |
| first_indexed | 2024-03-10T23:05:05Z |
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| spelling | doaj.art-01cde6ecee4e4676b5d3718fe677b2c62023-11-19T09:21:05ZengMDPI AGApplied Microbiology2673-80072023-07-013370972110.3390/applmicrobiol3030049The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa BiofilmsEvan Kello0Rochelle Greenberg1Weiqi Li2Shaya Polansky3Roberto Maldonado4Yakov Peter5Paramita Basu6New York College of Podiatric Medicine, 53 E 124 Street, New York, NY 10035, USANew York College of Podiatric Medicine, 53 E 124 Street, New York, NY 10035, USATouro College of Pharmacy, 230 W 125 Street, New York, NY 10027, USALander College for Men, Touro University, 75-31 150th Street, Flushing, NY 11367, USATouro College of Pharmacy, 230 W 125 Street, New York, NY 10027, USALander College for Men, Touro University, 75-31 150th Street, Flushing, NY 11367, USANew York College of Podiatric Medicine, 53 E 124 Street, New York, NY 10035, USA<i>Pseudomonas aeruginosa</i>, 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 <i>P. aeruginosa</i> 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 <i>P. aeruginosa</i> 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. <i>aeruginosa</i>. 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 <i>P. aeruginosa</i> 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, <i>P. aeruginosa</i> 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 <i>P. aeruginosa</i>, 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.https://www.mdpi.com/2673-8007/3/3/49antibiotic resistancebiofilmpseudomonas aeruginosaofloxacintobramycinceftazidime |
| spellingShingle | Evan Kello Rochelle Greenberg Weiqi Li Shaya Polansky Roberto Maldonado Yakov Peter Paramita Basu The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa Biofilms Applied Microbiology antibiotic resistance biofilm pseudomonas aeruginosa ofloxacin tobramycin ceftazidime |
| title | The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa Biofilms |
| title_full | The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa Biofilms |
| title_fullStr | The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa Biofilms |
| title_full_unstemmed | The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa Biofilms |
| title_short | The Effect of Antibiotic Treatment and Gene Expression of Mex B Efflux Transporters on the Resistance in <i>Pseudomonas</i> Aeruginosa Biofilms |
| title_sort | effect of antibiotic treatment and gene expression of mex b efflux transporters on the resistance in i pseudomonas i aeruginosa biofilms |
| topic | antibiotic resistance biofilm pseudomonas aeruginosa ofloxacin tobramycin ceftazidime |
| url | https://www.mdpi.com/2673-8007/3/3/49 |
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