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Publication Number
US-11427851-B2
Publication Date
2022-08-30
Expiration Date
Abstract
A fluidic device has a culture chamber configured to house a 3D culture matrix comprising a culture of microorganisms. A concentration gradient of a test substance is established over the 3D culture matrix by providing respective fluid flows at different end portions of the culture chamber and comprising different concentrations of the test substance. The response of the microorganisms to the test substance is determined based on the position of a border zone in the 3D culture matrix.
Core Innovation
The invention provides a system and fluidic device for determining a response of a microorganism to a test substance in a culture chamber configured to house a culture of the microorganism in a three dimensional (3D) culture matrix. The device includes a first fluid channel flanking a first end portion of the culture chamber and a second fluid channel flanking a second, different end portion, enabling formation of a concentration gradient of the test substance across the 3D culture matrix.
The system includes a first fluid reservoir with a first fluid comprising the test substance at a first concentration, and a second fluid reservoir with a second fluid lacking the test substance or comprising the test substance at a second concentration lower than the first concentration. The first and second reservoirs are connected to inputs of the first and second fluid channels to form the concentration gradient over at least a portion of the 3D culture matrix, based on gradient information defining concentration versus position in the 3D culture matrix.
A computer-based system takes at least one image of the 3D culture matrix and processes the image to identify a position relative to the first end portion and/or the second different end portion of any border zone between a growth zone of growing microorganism and a non-growth zone lacking growth. The computer-based system also identifies a width of the border zone and determines a minimal inhibitory concentration (MIC) by correlating the border-zone position to a concentration or concentration range based on the gradient information, and determines resistance based on the width of the border zone.
Claims Coverage
The claim set includes one independent claim that covers the core system for gradient formation, imaging-based border-zone identification, and MIC/resistance determination. The dependent claims further refine the independent claim by specifying additional fluid handling, timing relative to steady-state gradient formation, multi-test-substance configurations, additional channels for additional gradients, and an imaging feature based on light intensity.
Concentration gradient in a 3D culture matrix with imaging-based border-zone detection
A fluidic device with a culture chamber holding a culture of a microorganism in a 3D culture matrix, including a first fluid channel flanking a first end portion and a second fluid channel flanking a second, different end portion; first and second fluid reservoirs configured to form a concentration gradient of a test substance over at least a portion of the 3D culture matrix; and a computer-based system configured to take and process at least one image to identify a position and width of any border zone between a growth zone of growing microorganism and a non-growth zone lacking growth that is not inhibited by the test substance, then determine a MIC by correlating the border-zone position to a concentration or concentration range based on gradient information defining concentration versus position, and determine resistance based on the width of the border zone.
Overall claim coverage is centered on correlating an imaged border zone position and width in a 3D culture matrix to MIC and resistance, using concentration-versus-position gradient information generated by first and second fluid channels with first and second fluid reservoirs.
Stated Advantages
Continuous concentration range for test substance determination based on gradient information defining concentration versus position.
Short readout time.
No labeling generally.
Determination of MIC based on the position of a border zone between growth and non-growth regions.
Determination of resistance based on the width of a border zone.
Documented Applications
Rapid antimicrobial susceptibility testing in a 3D culture matrix using a fluidic device that establishes a steady concentration gradient, images the 3D matrix to locate a border zone, and maps the border-zone position (and optionally width) to concentration thresholds such as MIC and resistance.
Demonstrated use with a microorganism identified as E. coli K12 MG1655 and test substances including ampicillin and spectinomycin, with MIC and border-zone width differences indicating resistance behavior.
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