Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron Microscopy
Background. Colistin (Polymyxin E) has reemerged in the treatment of MDR Gram-negative infections. Traditional Colistin AST methods have long turnaround times and are cumbersome for routine use. We present a SEM-AST technique enabling rapid detection of Colistin resistance through direct observation...
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| Format: | Article |
| Language: | English |
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Wiley
2024-01-01
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| Series: | International Journal of Microbiology |
| Online Access: | http://dx.doi.org/10.1155/2024/8917136 |
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| author | Omar Zmerli Sara Bellali Gabriel Haddad Rim Iwaza Akiko Hisada Erino Matsumoto Yusuke Ominami Didier Raoult Jacques Bou Khalil |
| author_facet | Omar Zmerli Sara Bellali Gabriel Haddad Rim Iwaza Akiko Hisada Erino Matsumoto Yusuke Ominami Didier Raoult Jacques Bou Khalil |
| author_sort | Omar Zmerli |
| collection | DOAJ |
| description | Background. Colistin (Polymyxin E) has reemerged in the treatment of MDR Gram-negative infections. Traditional Colistin AST methods have long turnaround times and are cumbersome for routine use. We present a SEM-AST technique enabling rapid detection of Colistin resistance through direct observation of morphological and quantitative changes in bacteria exposed to Colistin. Methods. Forty-four Gram-negative reference organisms were chosen based on their Colistin susceptibility profiles. Bacterial suspensions of ∼107 CFU/mL were exposed to Colistin at EUCAST-ECOFF, with controls not exposed, incubated at 37°C, and then sampled at 0, 15, 30, 60, and 120 minutes. Phosphotungstic Acid (PTA) staining was applied, followed by SEM imaging using Hitachi TM4000PlusII-Tabletop-SEM at ×2000, ×5000 and ×7000 magnifications. Bacterial viability analysis was performed for all conditions by quantifying viable and dead organisms based on PTA-staining and morphologic changes. Results. We identified a significant drop in the percentage of viable organisms starting 30 minutes after exposure in susceptible strains, as compared to nonsignificant changes in resistant strains across all tested organisms. The killing effect of Colistin was best observed after 120 minutes of incubation with the antibiotic, with significant changes in morphologic features, including bacterial inflation, fusion, and lysis, observed as early as 30 minutes. Our observation matched the results of the gold standard-based broth microdilution method. Conclusions. We provide an extended application of the proof of concept for the utilization of the SEM-AST assay for Colistin for a number of clinically relevant bacterial species, providing a rapid and reliable susceptibility profile for a critical antibiotic. |
| format | Article |
| id | doaj-art-f8e4b5a3b82c4761874ba95f5c1b73da |
| institution | Kabale University |
| issn | 1687-9198 |
| language | English |
| publishDate | 2024-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | International Journal of Microbiology |
| spelling | doaj-art-f8e4b5a3b82c4761874ba95f5c1b73da2025-02-03T05:56:55ZengWileyInternational Journal of Microbiology1687-91982024-01-01202410.1155/2024/8917136Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron MicroscopyOmar Zmerli0Sara Bellali1Gabriel Haddad2Rim Iwaza3Akiko Hisada4Erino Matsumoto5Yusuke Ominami6Didier Raoult7Jacques Bou Khalil8Institut Hospitalo-Universitaire Méditerranée-InfectionInstitut Hospitalo-Universitaire Méditerranée-InfectionInstitut Hospitalo-Universitaire Méditerranée-InfectionInstitut Hospitalo-Universitaire Méditerranée-InfectionHitachi, Ltd.Hitachi, Ltd.Hitachi High-Tech CorporationConsulting Infection MarseilleInstitut Hospitalo-Universitaire Méditerranée-InfectionBackground. Colistin (Polymyxin E) has reemerged in the treatment of MDR Gram-negative infections. Traditional Colistin AST methods have long turnaround times and are cumbersome for routine use. We present a SEM-AST technique enabling rapid detection of Colistin resistance through direct observation of morphological and quantitative changes in bacteria exposed to Colistin. Methods. Forty-four Gram-negative reference organisms were chosen based on their Colistin susceptibility profiles. Bacterial suspensions of ∼107 CFU/mL were exposed to Colistin at EUCAST-ECOFF, with controls not exposed, incubated at 37°C, and then sampled at 0, 15, 30, 60, and 120 minutes. Phosphotungstic Acid (PTA) staining was applied, followed by SEM imaging using Hitachi TM4000PlusII-Tabletop-SEM at ×2000, ×5000 and ×7000 magnifications. Bacterial viability analysis was performed for all conditions by quantifying viable and dead organisms based on PTA-staining and morphologic changes. Results. We identified a significant drop in the percentage of viable organisms starting 30 minutes after exposure in susceptible strains, as compared to nonsignificant changes in resistant strains across all tested organisms. The killing effect of Colistin was best observed after 120 minutes of incubation with the antibiotic, with significant changes in morphologic features, including bacterial inflation, fusion, and lysis, observed as early as 30 minutes. Our observation matched the results of the gold standard-based broth microdilution method. Conclusions. We provide an extended application of the proof of concept for the utilization of the SEM-AST assay for Colistin for a number of clinically relevant bacterial species, providing a rapid and reliable susceptibility profile for a critical antibiotic.http://dx.doi.org/10.1155/2024/8917136 |
| spellingShingle | Omar Zmerli Sara Bellali Gabriel Haddad Rim Iwaza Akiko Hisada Erino Matsumoto Yusuke Ominami Didier Raoult Jacques Bou Khalil Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron Microscopy International Journal of Microbiology |
| title | Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron Microscopy |
| title_full | Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron Microscopy |
| title_fullStr | Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron Microscopy |
| title_full_unstemmed | Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron Microscopy |
| title_short | Antimicrobial Susceptibility Testing for Colistin: Extended Application of Novel Quantitative and Morphologic Assay Using Scanning Electron Microscopy |
| title_sort | antimicrobial susceptibility testing for colistin extended application of novel quantitative and morphologic assay using scanning electron microscopy |
| url | http://dx.doi.org/10.1155/2024/8917136 |
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