Rapid microbial detection and antimicrobial susceptibility testing
A method for the detection of microorganisms in a sample comprising contacting said sample with a biosensor concentration module, allowing microorganisms to grow for a first period of time and detecting growth of discrete microorganisms as an indication of the presence of said microorganisms.
1. A method comprising:
concentrating a plurality of individual microorganisms on a detection surface;
incubating the microorganisms in a medium containing an antibiotic agent;
determining, by a computer-based system configured to perform image analysis and comprising a processor, a memory, an interface and a microorganism detection device, a set of values corresponding to a growth rate of each individual microorganism on at least a portion of the detection surface;
calculating, by the computer-based system, a reference growth rate for the plurality of individual microorganisms; and
scoring, by the computer-based system, a microorganism having a growth rate different than the reference growth rate as resistant to the antimicrobial agent,
wherein the reference growth rate of the plurality of individual microorganisms is determined based on
a) number of pixels associated with an image of the plurality of individual microorganisms,
b) quantitative growth information comprising integrated intensity data for a clone, or
c) both a) and b).
2. The method of claim 1 , wherein the scoring is determined within a timeframe associated with 1 to 10 doubling times of the plurality of individual microorganisms.
3. The method of claim 1 , wherein the scoring is determined within a timeframe associated with 1 to 4 doubling times of the plurality of individual microorganisms.
4. The method of claim 1 , wherein the scoring is determined within a timeframe associated with 1 to 2 doubling times of the plurality of individual microorganisms.
5. The method of claim 1 , wherein the reference growth rate of the plurality of individual microorganisms is further determined based on a doubling time of the plurality of individual microorganisms.
6. The method of claim 1 , wherein the reference growth rate of the plurality of individual microorganisms is further determined based on an amount of fluorescence associated with an image of the plurality of individual microorganisms.
7. The method of claim 1 , wherein the reference growth rate of the plurality of individual microorganisms is further determined based on an amount of light scatter associated with an image of the plurality of individual microorganisms.
8. The method of claim 1 , further comprising, determining, by the computer-based system, a growth curve for an individual clone.
9. The method of claim 1 , wherein scoring the microorganism as antimicrobial resistant comprises determining that the growth rate for the microorganism exceeds a reference rate.
10. The method of claim 9 , wherein the reference rate comprises an average growth rate of a population of microorganisms incubated at the antibiotic concentration.
11. The method of claim 9 , wherein the reference rate comprises a growth rate of a most-susceptible microorganism.
12. The method of claim 1 , further comprising determining and reporting a minimum inhibitory concentration (MIC) for the antimicrobial resistant microorganism having a growth rate different than the reference growth rate based on selection of a reference curve from experimentally determined time-kill data points for the antimicrobial resistant microorganism having a growth rate different than the reference growth rate.
13. The method of claim 1 , wherein scoring the microorganism that is resistant to the antimicrobial agent comprises scoring the microorganism that is resistant to the antimicrobial agent at least a level of sensitivity of one resistant microorganism per 100 susceptible microorganisms.
14. The method of claim 1 , wherein the computer-based system comprises an image analysis module.
15. The method of claim 1 , wherein the reference growth rate for the plurality of individual microorganisms is further determined based on a method selected from a group consisting of surface plasmon resonance, light microscopy, confocal microscopy, phase contrast microscopy, brightfield microscopy, darkfield microscopy, fluorescence microscopy, epi-fluorescence microscopy, light scattering imaging, brightfield imaging, darkfield imaging, phase imaging, fluorescence imaging, upconverting phosphor imaging, quantum dot imaging, and chemilumiscence imaging.