Sequence specific antimicrobials
AssigneeThe Rockefeller University
InventorsDavid Bikard, Luciano Marraffini
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Abstract
Provided are compositions and methods for selectively reducing the amount of antibiotic resistant and/or virulent bacteria in a mixed bacteria population, or for reducing any other type of unwanted bacteria in a mixed bacteria population. The compositions and methods involve targeting bacteria that are differentiated from other members of the population by at least one unique clustered regularly interspaced short palindromic repeats (CRISPR) targeted DNA sequence. The compositions and methods can be readily adapted to target any bacteria or any bacteria plasmid, or both.
Claims15
A method for killing targeted bacteria in a mixed bacterial population,
- wherein the mixed bacterial population comprises at least two bacterial species selected from Staphylococcus, Clostridium, Bacillus, Salmonella, Helicobacter pylori, Neisseria gonorrhoeae, Neisseria meningitidis, and Escherichia coli, comprising:
- providing a pharmaceutical composition comprising a pharmaceutically acceptable carrier and packaged, recombinant phagemids that are packaged in phage capsids,
- wherein the packaged phagemids comprise a clustered regularly interspaced short palindromic repeats (CRISPR) system,
- wherein the CRISPR system comprises DNA encoding:
- i) a type II CRISPR-associated enzyme;
- and ii) a targeting RNA that targets at least one bacterial chromosome at a target site;
- and
- contacting the bacterial population with the pharmaceutical composition,
- wherein the contacting with the pharmaceutical composition introduces at least some of the phagemids into at least some of the bacteria in the bacterial population,
- wherein subsequent to the introduction of the phagemids, the bacteria into which the phagemid is introduced expresses the targeting RNA and the type II CRISPR-associated enzyme,
- wherein the expressed type II CRISPR-associated enzyme cleaves the bacterial chromosome at the target site of the targeting RNA, and
- wherein the cleavage of the bacterial chromosome at the target site kills the bacteria.
A method for killing targeted bacteria in a mixed bacterial population, wherein the mixed bacterial population comprises at least two different bacterial species, comprising:
- providing a pharmaceutical composition comprising a pharmaceutically acceptable carrier and packaged, recombinant phagemids that are packaged in phage capsids,
- wherein the packaged phagemids comprise a clustered regularly interspaced short palindromic repeats (CRISPR) system,
- wherein the CRISPR system comprises DNA encoding:
- i) a type II CRISPR-associated enzyme;
- and ii) multiple targeting RNAs, one of which targets a virulence gene on a chromosome or on a plasmid at a target site within the chromosome or within the plasmid and one of which targets an antibiotic resistance gene on a plasmid at a target site within the plasmid;
- and
- contacting the bacterial population with the pharmaceutical composition,
- wherein the contacting with the pharmaceutical composition introduces at least some of the phagemids into at least some of the bacteria in the bacterial population that contain the virulence gene on a chromosome or on a plasmid and the antibiotic resistance gene on a plasmid,
- wherein subsequent to the introduction of the phagemids, the bacteria into which the phagemid is introduced expresses the targeting RNAs and the type II CRISPR-associated enzyme,
- wherein the expressed type II CRISPR-associated enzyme cleaves the virulence gene on a chromosome or on a plasmid at the target site within the chromosome or within the plasmid and the antibiotic resistance gene on a plasmid at the target site within the plasmid, and
- wherein the cleavage of the bacterial plasmid at the target site of the antibiotic resistance gene within the plasmid kills the bacteria in the presence of the antibiotic.
Dependencies are inferred from the claim text.