Method and device for rapid detection of bacterial antibiotic resistance/susceptibility
Inventors
Sauer-Budge, Alexis • Sharon, Andre • Kalashnikov, Maxim • Wirz, Holger
Assignees
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Publication Number
US-8785148-B2
Publication Date
2014-07-22
Expiration Date
Abstract
Described herein is a method and a device for expediting delivery of an agent to a damaged bacterial cell. In one embodiment, the methods and devices are useful for screening candidate antibiotics. In another embodiment, the methods and devices described herein are used to determine susceptibility of bacteria to an antibiotic. The methods also provide a method for determining an appropriate antibiotic to treat an individual having a bacterial infection.
Core Innovation
The invention relates to a method for determining sensitivity of bacteria to an antibiotic by immobilizing bacteria from a bacterial suspension through covalent attachment to a solid support. The immobilized bacteria are contacted with an agent comprising a reporter moiety that preferentially binds to damaged bacterial cells, and the bacteria are subjected to a stressor in the presence of an antibiotic without the need for bacterial growth phase.
After contacting the covalently immobilized bacteria with the stressor in the presence of the antibiotic, a signal from the reporter moiety in the immobilized bacteria is detected between 1 and 30 minutes. The detected signal is compared to a control comprising immobilized bacteria subjected to the stressor in the absence of the antibiotic, and an increase in the signal in the presence of the antibiotic compared to the control indicates susceptibility, while a signal comparable to the control indicates resistance.
The disclosed approach includes stressor and reporter concepts such as biochemical and physical or chemical stressors, including lysostaphin, lysozyme, endolysin, oxidative stress, and a porin, together with reporter moieties and fluorescent dyes used to label damaged cells. The partial content also describes rapid fluorescence-based detection and comparison between antibiotic and control conditions over minutes, enabling differentiation such as MRSA versus MSSA in the disclosed context.
Claims Coverage
The relevant independent claim is clm-00001, which contains the core method framework with a rapid detection window and signal comparison to infer susceptibility versus resistance without bacterial growth phase. Additional dependent claims listed in the provided material refine aspects such as detection timing, fluorescence modality, specific fluorescent dyes, specific stressors, and narrowing to pathogenic bacteria.
Covalent immobilization of bacteria on a solid support
The method immobilizes the bacteria from a bacterial suspension by covalent attachment to a solid support.
Reporter moiety preferentially binding damaged bacterial cells
The method contacts the immobilized bacteria with an agent comprising a reporter moiety, which preferentially binds to damaged bacterial cells.
Stressor exposure with antibiotic without bacterial growth phase
The method subjects the immobilized bacteria to a stressor in the presence of an antibiotic without the need for bacterial growth phase.
Rapid reporter signal detection and susceptibility/resistance interpretation
The method detects a signal from the reporter moiety in the immobilized bacteria, where the detecting is performed between 1 and 30 minutes from contacting the covalently immobilized bacteria with the stressor in the presence of the antibiotic, and compares the signal to a control comprising immobilized bacteria subjected to the stressor in the absence of the antibiotic, wherein detection of an increase in said signal in the presence of an antibiotic compared to a control indicates susceptibility and a signal comparable to the control indicates resistance.
Across the provided claim material, the inventive coverage centers on covalently immobilizing bacteria, using a reporter moiety that preferentially binds damaged cells, applying a stressor in the presence of an antibiotic without bacterial growth phase, and performing rapid time-bounded detection followed by comparison to an antibiotic-free stressor control to infer susceptibility versus resistance.
Stated Advantages
Determines sensitivity of bacteria to an antibiotic between 1 and 30 minutes from contacting the covalently immobilized bacteria with the stressor in the presence of the antibiotic.
Provides susceptibility versus resistance indication based on reporter signal change relative to a control subjected to the stressor in the absence of the antibiotic.
Does not require bacterial growth phase.
Enables signal-based detection (including fluorescence) from immobilized bacteria after stressor exposure with or without the antibiotic.
Documented Applications
Rapid antibiotic susceptibility/resistance detection using immobilized bacteria and a reporter moiety that binds damaged cells, with susceptibility/resistance inferred from signal change relative to a control.
Differentiation of resistant versus susceptible bacteria in an antibiotic exposure context, including MRSA versus MSSA in the provided partial content summary.
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