US 12,246,061 B2
Sequence specific antimicrobials
David Bikard, Paris (FR); and Luciano Marraffini, Brooklyn, NY (US)
Assigned to The Rockefeller University, New York, NY (US)
Filed by The Rockefeller University, New York, NY (US)
Filed on Jan. 29, 2024, as Appl. No. 18/425,204.
Application 17/168,971 is a division of application No. 16/877,010, filed on May 18, 2020.
Application 17/168,971 is a division of application No. 16/877,030, filed on May 18, 2020, granted, now 11,135,273, issued on Oct. 5, 2021.
Application 17/088,302 is a division of application No. 16/877,030, filed on May 18, 2020, granted, now 11,135,273, issued on Oct. 5, 2021.
Application 17/088,297 is a division of application No. 16/877,030, filed on May 18, 2020, granted, now 11,135,273, issued on Oct. 5, 2021.
Application 16/877,030 is a division of application No. 15/159,929, filed on May 20, 2016, granted, now 11,918,631.
Application 15/159,929 is a division of application No. 14/766,675, granted, now 10,660,943, issued on May 26, 2020, previously published as PCT/US2014/015252, filed on Feb. 7, 2014.
Application 18/425,204 is a continuation of application No. 17/168,971, filed on Feb. 5, 2021, granted, now 12,168,040.
Application 18/425,204 is a continuation of application No. 17/088,297, filed on Nov. 3, 2020.
Application 18/425,204 is a continuation of application No. 17/088,302, filed on Nov. 3, 2020.
Application 18/425,204 is a continuation of application No. 16/877,010, filed on May 18, 2020.
Application 18/425,204 is a continuation of application No. 15/159,929, filed on May 20, 2016, granted, now 11,918,631.
Application 17/168,971 is a continuation of application No. 15/159,929, filed on May 20, 2016, granted, now 11,918,631.
Application 16/877,010 is a continuation of application No. 14/766,675, granted, now 10,660,943, issued on May 26, 2020, previously published as PCT/US2014/015252, filed on Feb. 7, 2014.
Claims priority of provisional application 61/761,971, filed on Feb. 7, 2013.
Prior Publication US 2024/0165210 A1, May 23, 2024
This patent is subject to a terminal disclaimer.
Int. Cl. A61K 38/46 (2006.01); A01N 63/00 (2020.01); A61K 31/7105 (2006.01); A61K 31/713 (2006.01); A61K 45/06 (2006.01); C12N 9/16 (2006.01); C12N 9/22 (2006.01); C12N 15/113 (2010.01); C12N 15/74 (2006.01)
CPC A61K 38/465 (2013.01) [A01N 63/00 (2013.01); A61K 31/7105 (2013.01); A61K 31/713 (2013.01); A61K 45/06 (2013.01); C12N 9/16 (2013.01); C12N 9/22 (2013.01); C12N 15/113 (2013.01); C12N 15/74 (2013.01); C12Y 301/00 (2013.01); C12N 2310/10 (2013.01); C12N 2310/20 (2017.05); C12N 2795/10331 (2013.01); C12N 2795/10332 (2013.01); C12N 2795/10343 (2013.01); C12N 2795/10371 (2013.01)] 24 Claims
 
1. A method for selectively modifying a mixed population of bacteria, wherein the mixed population of bacteria comprises at least a first and a second bacterial species, comprising:
(a) contacting the mixed population of bacteria with packaged, recombinant phagemids that are packaged in phage capsids, wherein the packaged phagemids comprise a clustered regularly interspaced short palindromic repeats (CRISPR) system comprising i) a type II CRISPR-associated enzyme and ii) two or more crRNA guides that target different chromosomal sequences in bacterial cells of the second bacterial species and,
(b) producing the crRNA guides in the bacterial cells of the second bacterial species, wherein the crRNA guides comprise i) a first crRNA guide complementary to a first target chromosomal sequence in the bacterial cells of the second bacterial species and ii) a second crRNA guide complementary to a second target chromosomal sequence in the bacterial cells of the second bacterial species, wherein the second target chromosomal sequence is different from the first target chromosomal sequence; and wherein:
1) the first target chromosomal sequence is comprised by a first antibiotic resistance gene and the second target chromosomal sequence is comprised by a second antibiotic resistance gene,
2) the first target chromosomal sequence is comprised by an antibiotic resistance gene and the second target chromosomal sequence is comprised by a virulence gene, or
3) the first target chromosomal sequence is comprised by a first virulence gene and the second target chromosomal sequence is comprised by a second virulence gene;
wherein the expressed type II CRISPR-associated enzyme cleaves the first and second target chromosomal sequences at the target sites of the crRNA guides in the bacterial cells of the second bacterial species,
wherein the first and second bacterial species are different and are selected from Streptococcus, Staphylococcus, Clostridium, Bacillus, Salmonella, Helicobacter pylori, Neisseria gonorrhoeae, Neisseria meningitidis, and Escherichia coli, and
wherein the growth of the bacterial cells of the first bacterial species in the mixed population is not inhibited, and wherein at least 50% of the bacterial cells of the second bacterial species are killed.