Compositions and methods for detecting antibiotic responsive mRNA expression signatures and uses thereof

Patent metadata

Field	Value
Publication number	US-12442035-B2
Application number	US-201917271496-A
Country	US
Kind code	B2
Filing date	Aug 26, 2019
Priority date	Aug 27, 2018
Publication date	Oct 14, 2025
Grant date	Oct 14, 2025

Abstract

The present disclosure relates to compositions, methods, and kits for rapid phenotypic detection of antibiotic resistance/susceptibility.

First claim

What is claimed is:

1. A method, comprising:
obtaining a sample including one or more bacterial cells, wherein the sample is obtained from a patient;
processing the sample to enrich the one or more bacterial cells;
contacting the sample with one or more antibiotic compounds;
lysing the sample to release messenger ribonucleic acid (mRNA) from the one or more bacterial cells;
hybridizing the released mRNA to at least one set of two nucleic acid probes, wherein each nucleic acid probe includes a unique barcode or tag and wherein the at least one set of nucleic acid probes includes one or more probes from SEQ ID NOs: 74-86 and one or more probes from SEQ ID NOs: 91-103;
detecting the hybridized nucleic acid probes;
identifying one or more genetic resistance determinants;
determining antibiotic susceptibility of the one or more bacterial cells using a machine learning algorithm trained on a susceptibility classification dataset, wherein the machine learning algorithm generates a ranked list of two or more susceptibility features, thereby determining antibiotic susceptibility of the one or more bacterial cells; and
selecting an appropriate antibiotic based on the determined antibiotic susceptibility of the one or more bacterial cells; and
administering the appropriate antibiotic to the patient.

2. The method of claim 1 , wherein the hybridizing occurs at a temperature between about 64° C. and about 69° C.

3. The method of claim 1 , wherein the hybridizing occurs at a temperature between about 65° C. and about 67° C.

4. The method of claim 1 , wherein the hybridizing occurs at about 65° C. or about 66° C. or about 67° C.

5. The method of claim 1 , wherein the at least one set of two nucleic acid probes is selected from the group consisting of: SEQ ID NO: 74 and SEQ ID NO: 91; SEQ ID NO: 75 and SEQ ID NO: 92; SEQ ID NO: 76 and SEQ ID NO: 93; SEQ ID NO: 77 and SEQ ID NO: 94; SEQ ID NO: 78 and SEQ ID NO: 95; SEQ ID NO: 79 and SEQ ID NO: 96; SEQ ID NO: 80 and SEQ ID NO: 97; SEQ ID NO: 81 and SEQ ID NO: 98; SEQ ID NO: 82 and SEQ ID NO: 99; SEQ ID NO: 83 and SEQ ID NO: 100; SEQ ID NO: 84 and SEQ ID NO: 101; SEQ ID NO: 85 and SEQ ID NO: 102; and SEQ ID NO: 86 and SEQ ID NO: 103.

6. The method of claim 5 , wherein the at least one set of two nucleic acid probes is selected from the group consisting of: SEQ ID NO: 74 and SEQ ID NO: 91; SEQ ID NO: 75 and SEQ ID NO: 92; SEQ ID NO: 76 and SEQ ID NO: 93; SEQ ID NO: 77 and SEQ ID NO: 94; SEQ ID NO: 78 and SEQ ID NO: 95; SEQ ID NO: 80 and SEQ ID NO: 97; SEQ ID NO: 82 and SEQ ID NO: 99; SEQ ID NO: 83 and SEQ ID NO: 100; and SEQ ID NO: 85 and SEQ ID NO: 102.

7. The method of claim 1 , wherein at least two sets of nucleic acid probes are selected.

8. The method of claim 1 , wherein at least three sets of nucleic acid probes are selected.

9. The method of claim 1 , wherein at least four sets of nucleic acid probes are selected.

10. A method, comprising:
obtaining a sample including one or more bacterial cells, wherein the sample is obtained from a patient;
processing the sample to enrich the one or more bacterial cells;
contacting the sample with one or more antibiotic compounds;
lysing the sample to release messenger ribonucleic acid (mRNA) from the one or more bacterial cells;
hybridizing the released mRNA to at least five nucleic acid probe sets, wherein each nucleic acid probe includes a unique barcode or tag and wherein the at least one set of nucleic acid probes includes one or more probes from SEQ ID NOs: 74-86 and one or more probes from SEQ ID NOs: 91-103;
detecting the hybridized nucleic acid probes;
identifying one or more genetic resistance determinants;
determining antibiotic susceptibility of the one or more bacterial cells using a machine learning algorithm trained on a susceptibility classification dataset, wherein the machine learning algorithm generates a ranked list of two or more susceptibility features, thereby determining antibiotic susceptibility of the one or more bacterial cells; and
selecting an appropriate antibiotic based on the determined antibiotic susceptibility of the one or more bacterial cells and administering the appropriate antibiotic to the patient.

11. The method of claim 10 , wherein the at least five nucleic acid probe sets comprise: SEQ ID NO: 74 and SEQ ID NO: 91; SEQ ID NO: 75 and SEQ ID NO: 92; SEQ ID NO: 76 and SEQ ID NO: 93; SEQ ID NO: 78 and SEQ ID NO: 95; and SEQ ID NO: 84 and SEQ ID NO: 101.

Assignees

• Broad Inst Inc
• Massachusetts Gen Hospital

Inventors

• Hung Deborah
• Bhattacharyya Roby
• Livny Jonathan
• Ma Peijun

Classifications

• C12Q1/689 Primary

  for bacteria · CPC title

• C12Q1/6876

  Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes · CPC title

• C12N1/20

  Bacteria; Culture media therefor · CPC title

• C12N1/06

  Lysis of microorganisms · CPC title

• C12Q1/6816 Primary

  characterised by the detection means (C12Q1/6804 takes precedence) · CPC title