Methods and systems of characterizing and counting microbiological colonies
Described herein are methods, systems, and non-transitory computer-readable media to non-destructively acquire three-dimensional profiles of cellular microbiological samples growing on the surface of a solid growth medium. Acquisitions can be performed by an optical microscope that includes a vertical scanning interferometer. The three-dimensional profiles can enable measurement of sample parameters of microcolonies, which can be made of microbial colony forming units. The methods and systems enable early and rapid detection and quantification of microbes.
1. A method comprising:
non-destructively acquiring by a vertical scanning interferometer a three-dimensional (3D) profile that provides a height measurement of a microcolony on a surface, the microcolony comprising a microbial colony forming unit (CFU);
computing a sample parameter based on the 3D profile with the height measurement; and
enumerating, identifying, and differentiating a species among a plurality of species of the microbial CFU based on the sample parameter.
2. The method of claim 1 , wherein the surface comprises that of a solid growth medium.
3. The method of claim 1 , wherein a vertical resolution of the vertical scanning interferometer is less than or equal to 5 nm.
4. The method of claim 3 , wherein said acquiring comprises laterally sampling an area of the growth medium that is greater than or equal to 10 mm, 8 mm, 5 mm, 3 mm, 1 mm, 0.5 mm or 0.05 mm across.
5. The method of claim 1 , wherein the vertical scanning interferometer is a white light interferometer.
6. The method of claim 1 , wherein the sample parameter comprises a value based on microcolony height, aspect ratio, volume, number of microcolonies, microcolony morphology, or a combination thereof.
7. The method of claim 1 , further comprising estimating a microbial CFU population value based on the sample parameter.
8. The method of claim 1 , wherein said computing occurs at a non-zero time value that is less than 1 hour, 2 hours, 4 hours, 6 hours, or 8 hours after the microbial CFU is introduced to a solid growth medium.
9. The method of claim 8 , further comprising estimating a growth rate value of the microbial CFU based on the sample parameter and on changes over the period of time.
10. The method of claim 1 , further comprising tracking changes in the sample parameter over a period of time.
11. The method of claim 1 , further comprising screening a plurality of samples by repeating said acquiring and said computing at each of a plurality of sample wells, each sample well comprising a solid growth medium.
12. The method of claim 11 , wherein the plurality of sample wells have different solid growth media selected from the group comprising culture media, minimal media, selective media, differential media, transport media, and combinations thereof.
13. The method of claim 11 , wherein the plurality of samples wells each have a non-zero dimension that is less than or equal to 50 mm, 30 mm, 15 mm, 10 mm, or 5 mm.
14. The method of claim 1 , further comprising covering the surface of a solid growth medium with a lid comprising an optically transparent window through which light of the vertical scanning interferometer passes.
15. The method of claim 1 , wherein the surface of a solid growth medium has a non-zero root mean squared roughness value less than or equal to 200 nm, 125 nm, 100 nm, or 50 nm.
16. A non-transitory computer readable storage medium storing one or more programs, the one or more programs comprising instructions, which when executed by one or more processors of an electronic device comprising, or operably connected to, a vertical scanning interferometer, cause the device:
to acquire a 3D profile with a height measurement of a microcolony grown on a surface, the microcolony comprising a microbial colony forming unit (CFU); and
to compute a sample parameter based on the 3D profile with the height measurement,
enumerate, identify, and differentiate a species among a plurality of species of the microbial CFU based on the sample parameter, and determine a microbial CFU population value in a microcolony.
17. The non-transitory computer readable storage medium of claim 16 , wherein the sample parameter is one of microcolony height, aspect ratio, volume, number of microcolonies, microcolony morphology, microcolony count, or a combination thereof.
18. The non-transitory computer readable storage medium of claim 16 , which when executed by one or more processors of an electronic device comprising, or operably connected to, a vertical scanning interferometer, further cause the device to estimate a growth rate value of the microbial CFU, provide a positive/negative outcome of a diagnostic test, indicate a biofilm state, or perform a combination thereof based on one or more of the sample parameters.