Determination of colistin resistance by simple disk diffusion test using modified Mueller-Hinton agar

Jean Damascene Uwizeyimana; Daewon Kim; Hyunsook Lee; Jung Hyun Byun; Dongeun Yong

doi:10.3343/alm.2020.40.4.306

Determination of colistin resistance by simple disk diffusion test using modified Mueller-Hinton agar

Jean Damascene Uwizeyimana, Daewon Kim, Hyunsook Lee, Jung Hyun Byun, Dongeun Yong

Department of Laboratory Medicine

Research output: Contribution to journal › Article › peer-review

4 Citations (Scopus)

Abstract

Background: Colistin has become a last-resort antibiotic for the management of multidrug-resistant gram-negative bacteria. The disk diffusion test is cheap and easy to perform but may be unreliable for colistin susceptibility testing due to poor diffusion of the large colistin molecule. An improved agar diffusion test would increase the reliability of colistin susceptibility testing. This study aimed to modify Muller-Hinton agar (MHA) to improve colistin diffusion in agar. Methods: MHA was modified by reducing the agar concentration from 100% to 30% and supplementing with protamine. We tested 60 gram-negative clinical isolates of Pseudomonas aeruginosa (N = 27) and Acinetobacter calcoaceticus-baumannii complex (N = 33). Disk diffusion test results were interpreted based on minimum inhibitory concentrations determined by broth microdilution. Results: The modified MHA yielded the best performance metrics, including 94.7% sensitivity, 100% specificity, and an area under the curve of 0.995 (95% confidence interval, 0.982-1.000), P < 0.001, at a cut-off point of 13 mm. Conclusions: A reduction of the agar concentration from 100% to 30% and the addition of protamine improved colistin diffusion in agar and allowed routine colistin susceptibility testing in a clinical microbiology laboratory, but should be handled with caution.

Original language	English
Pages (from-to)	306-312
Number of pages	7
Journal	Annals of laboratory medicine
Volume	40
Issue number	4
DOIs	https://doi.org/10.3343/alm.2020.40.4.306
Publication status	Published - 2020

Bibliographical note

Funding Information:
This work was supported by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute, funded by the Ministry of Health & Welfare, Korea (grant number: HI14C1324); by the Research Program funded by the Korea Centers for Disease Control and Prevention, Ministry of Health and Welfare, Korea (2019-ER5403-00); by Korea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheries through Agricultural Microbiome R&D Program, funded by Ministry of Agriculture, Food and Rural Affairs (918003-4). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Publisher Copyright:
© Korean Society for Laboratory Medicine.

All Science Journal Classification (ASJC) codes

Medicine(all)

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10.3343/alm.2020.40.4.306

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title = "Determination of colistin resistance by simple disk diffusion test using modified Mueller-Hinton agar",

abstract = "Background: Colistin has become a last-resort antibiotic for the management of multidrug-resistant gram-negative bacteria. The disk diffusion test is cheap and easy to perform but may be unreliable for colistin susceptibility testing due to poor diffusion of the large colistin molecule. An improved agar diffusion test would increase the reliability of colistin susceptibility testing. This study aimed to modify Muller-Hinton agar (MHA) to improve colistin diffusion in agar. Methods: MHA was modified by reducing the agar concentration from 100% to 30% and supplementing with protamine. We tested 60 gram-negative clinical isolates of Pseudomonas aeruginosa (N = 27) and Acinetobacter calcoaceticus-baumannii complex (N = 33). Disk diffusion test results were interpreted based on minimum inhibitory concentrations determined by broth microdilution. Results: The modified MHA yielded the best performance metrics, including 94.7% sensitivity, 100% specificity, and an area under the curve of 0.995 (95% confidence interval, 0.982-1.000), P < 0.001, at a cut-off point of 13 mm. Conclusions: A reduction of the agar concentration from 100% to 30% and the addition of protamine improved colistin diffusion in agar and allowed routine colistin susceptibility testing in a clinical microbiology laboratory, but should be handled with caution.",

author = "Uwizeyimana, {Jean Damascene} and Daewon Kim and Hyunsook Lee and Byun, {Jung Hyun} and Dongeun Yong",

note = "Funding Information: This work was supported by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute, funded by the Ministry of Health & Welfare, Korea (grant number: HI14C1324); by the Research Program funded by the Korea Centers for Disease Control and Prevention, Ministry of Health and Welfare, Korea (2019-ER5403-00); by Korea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheries through Agricultural Microbiome R&D Program, funded by Ministry of Agriculture, Food and Rural Affairs (918003-4). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Publisher Copyright: {\textcopyright} Korean Society for Laboratory Medicine.",

year = "2020",

doi = "10.3343/alm.2020.40.4.306",

language = "English",

volume = "40",

pages = "306--312",

journal = "Annals of Laboratory Medicine",

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publisher = "Seoul National University",

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T1 - Determination of colistin resistance by simple disk diffusion test using modified Mueller-Hinton agar

AU - Uwizeyimana, Jean Damascene

AU - Kim, Daewon

AU - Lee, Hyunsook

AU - Byun, Jung Hyun

AU - Yong, Dongeun

N1 - Funding Information: This work was supported by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute, funded by the Ministry of Health & Welfare, Korea (grant number: HI14C1324); by the Research Program funded by the Korea Centers for Disease Control and Prevention, Ministry of Health and Welfare, Korea (2019-ER5403-00); by Korea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheries through Agricultural Microbiome R&D Program, funded by Ministry of Agriculture, Food and Rural Affairs (918003-4). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Publisher Copyright: © Korean Society for Laboratory Medicine.

PY - 2020

Y1 - 2020

N2 - Background: Colistin has become a last-resort antibiotic for the management of multidrug-resistant gram-negative bacteria. The disk diffusion test is cheap and easy to perform but may be unreliable for colistin susceptibility testing due to poor diffusion of the large colistin molecule. An improved agar diffusion test would increase the reliability of colistin susceptibility testing. This study aimed to modify Muller-Hinton agar (MHA) to improve colistin diffusion in agar. Methods: MHA was modified by reducing the agar concentration from 100% to 30% and supplementing with protamine. We tested 60 gram-negative clinical isolates of Pseudomonas aeruginosa (N = 27) and Acinetobacter calcoaceticus-baumannii complex (N = 33). Disk diffusion test results were interpreted based on minimum inhibitory concentrations determined by broth microdilution. Results: The modified MHA yielded the best performance metrics, including 94.7% sensitivity, 100% specificity, and an area under the curve of 0.995 (95% confidence interval, 0.982-1.000), P < 0.001, at a cut-off point of 13 mm. Conclusions: A reduction of the agar concentration from 100% to 30% and the addition of protamine improved colistin diffusion in agar and allowed routine colistin susceptibility testing in a clinical microbiology laboratory, but should be handled with caution.

AB - Background: Colistin has become a last-resort antibiotic for the management of multidrug-resistant gram-negative bacteria. The disk diffusion test is cheap and easy to perform but may be unreliable for colistin susceptibility testing due to poor diffusion of the large colistin molecule. An improved agar diffusion test would increase the reliability of colistin susceptibility testing. This study aimed to modify Muller-Hinton agar (MHA) to improve colistin diffusion in agar. Methods: MHA was modified by reducing the agar concentration from 100% to 30% and supplementing with protamine. We tested 60 gram-negative clinical isolates of Pseudomonas aeruginosa (N = 27) and Acinetobacter calcoaceticus-baumannii complex (N = 33). Disk diffusion test results were interpreted based on minimum inhibitory concentrations determined by broth microdilution. Results: The modified MHA yielded the best performance metrics, including 94.7% sensitivity, 100% specificity, and an area under the curve of 0.995 (95% confidence interval, 0.982-1.000), P < 0.001, at a cut-off point of 13 mm. Conclusions: A reduction of the agar concentration from 100% to 30% and the addition of protamine improved colistin diffusion in agar and allowed routine colistin susceptibility testing in a clinical microbiology laboratory, but should be handled with caution.

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Determination of colistin resistance by simple disk diffusion test using modified Mueller-Hinton agar

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