Detection, isolation and identification of microorganisms
A method of selectively identifying live microorganisms in a sample involves contacting a sample suspected of containing a live microorganism of interest with a biomolecular precursor labeled with a reactive chemical label. The live microorganism of interest is grown under conditions that promote selective growth of the microorganism to permit the microorganism to utilize the labeled biomolecular precursor to synthesize labeled biomolecules on a cell surface of the live microorganism. Labeled live microorganisms are contacted with a reporter and/or capture element bearing a functional group that reacts with the reactive chemical label. The labeled live microorganism is analyzed to identify it. The method can specifically isolate live microorganisms from dead ones thereby reducing the possibility of false identification of contaminating microorganisms and is able to detect specific strains using tailored metabolites that are specific for a given microorganism. The present method is considerably faster than currently used cell culture-based methods.
1. A method of selectively identifying a live microorganism in a soil, food or water sample comprising:
a) contacting a sample suspected of containing the live microorganism with one or more biomolecular precursors labeled with a reactive chemical label, wherein each of said one or more biomolecular precursors is labeled with the same reactive chemical label;
b) growing the live microorganism under conditions that promote selective growth of the live microorganism to permit the live microorganism to utilize the one or more biomolecular precursors labeled with the same reactive chemical label to synthesize labeled biomolecules on a cell surface of the live microorganism to produce a labeled live microorganism;
c) contacting the labeled live microorganism with a reporter and/or capture element bearing a functional group that reacts with the same reactive chemical label; and,
d) analyzing the labeled live microorganism to identify it.
2. The method according to claim 1 further comprising separating the labeled live microorganism from unlabeled microorganisms in the sample.
3. The method according to claim 1 , wherein the reactive chemical label and the functional group comprise paired chemical species, the paired chemical species selected from the group consisting of an alkyne/azide pair, a nitrone/alkyne pair, an azide/phosphine pair, an azide/strained alkyne pair, a strained alkene/tetrazine pair, an aldehyde/aminiooxy pair, an aldehyde/hydrazide pair, a ketone/aminooxy pair, a ketone/hydrazide pair, a thiol/alkene pair and an alkene/diaryl tetrazole pair.
4. The method according to claim 1 , wherein the functional group of the reporter and/or capture element comprises an azide group.
5. The method according to claim 1 , wherein the same reactive chemical label comprises an alkyne group.
6. The method according to claim 1 , wherein the reporter and/or capture element is bound to a surface of a physical substrate.
7. The method according to claim 6 , wherein the physical substrate comprises a magnetic particle, a magnetic bead or a microfluidic device.
8. The method according to claim 1 , wherein each of said one or more biomolecular precursors is specific for the live microorganism.
9. The method according to claim 1 , wherein the one or more biomolecular precursors is an amino acid, a nucleic acid, a sugar or a fatty acid, or a mixture of two or more thereof.
10. The method according to claim 9 , wherein the one or more biomolecular precursors is a 3-deoxy-D-manno-oct-2-ulosonic acid analogue.
11. The method according to claim 9 , wherein the one or more biomolecular precursors is methionine or a methionine analogue.
12. The method according to claim 1 , wherein step a) comprises contacting the sample with at least two different biomolecular precursors, each labeled with the same reactive chemical label.
13. The method according to claim 1 , wherein the one or more biomolecular precursors comprises a mixture of methionine or a methionine analogue and a 3-deoxy-D-manno-oct-2-ulosonic acid analogue.
14. The method according to claim 1 , wherein the microorganism is a species of bacteria.
15. The method according to claim 1 , wherein an immobilized strain-specific antibody is used to capture the labeled live microorganism.
16. The method according to claim 15 , wherein the strain-specific antibody is immobilized by a magnetic bead.
17. A kit for selectively identifying a live microorganism in a soil, food or water sample comprising:
a) a biomolecular precursor labeled with a reactive chemical label,
b) a reporter and/or capture element bearing a functional group that reacts with the reactive chemical label,
c) a vessel for growing the live microorganism and conducting a reaction between the labeled biomolecular precursor and said reporter and/or capture element bearing a functional group that reacts with the reactive chemical label,
d) growth medium for the live microorganism,
e) nutritional factors necessary for growing the live microorganism, and
f) instructions for using the kit to identify the live microorganism.
18. A method of selectively identifying a live microorganism in a sample comprising:
a) treating the sample with a medium containing amino acid biosynthesis inhibitory amino acids,
b) contacting the sample with an amino acid labeled with a reactive chemical label;
c) growing the live microorganism under conditions that promote selective growth of the live microorganism to permit the live microorganism to utilize the amino acid labeled with the reactive chemical label to synthesize labeled biomolecules on a cell surface of the live microorganism to produce a labeled live microorganism;
d) contacting the labeled live microorganism with a reporter and/or capture element bearing a functional group that reacts with the reactive chemical label; and,
e) analyzing the labeled live microorganism to identify it.
19. The method of claim 18 , wherein the labeled amino acid labeled with the reactive chemical label is methionine or a methionine analogue.