Methods and compositions for reducing malodor

Abstract

The present technology relates generally to the field of reducing, preventing, or ameliorating the generation of undesirable body odors emanating from the skin. In particular, the present technology discloses ΔpatB-Staphylococcus hominis bacterial strains, which have been modified to reduce or prevent the production of 3-methyl-3-sulfanylhexan-l-ol (3M3SH), compositions and pharmaceutical comprising one or more of said strains, methods of genetically engineering said strains, methods for reducing body odor using said strains, and methods for altering a microbiome using said strains.

Description

Atty. Dkt. No. 135523-0115METHODS AND COMPOSITIONS FOR REDUCING MALODORCROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of and priority to U.S. Provisional Application No. 63 / 691,755, filed September 6, 2024, the disclosure of which is incorporated by reference herein in its entirety for any and all purposes.TECHNICAL FIELD

[0002] The present technology relates generally to the field of reducing, preventing, or ameliorating the generation of undesirable body odors emanating from the skin. In particular, the present technology provides ApatB-Staphylococcus hominis bacterial strains, which have been modified to prevent or reduce the production of volatile thioalcohols, such as 3-methyl- 3-sulfanylhexan-l-ol (3M3SH), from precursor molecules commonly found in sweat.BACKGROUND

[0003] The following description is provided to assist the understanding of the reader. None of the information provided or references cited is admitted to be prior art to the compositions and methods disclosed herein.

[0004] Body odors, particularly those originating from sweat, represent a significant irritant for many humans, as evidenced by the multi-billion-dollar deodorant, body -wash, and perfume industries. Sweat, released from glands throughout the body, is a complex solution of salts, minerals, metabolites, and, potentially, pharmaceuticals or other exogenous compounds. Once secreted, sweat provides a nutrient filled solution for the skin microbiota to use as a nutrient source. Unfortunately, this metabolism, and the incumbent high levels of metabolic activity, results in the production of a number of undesirable byproducts, including volatile thioalcohols, which give off a distinct and unpleasant smell commonly referred to as body odor.

[0005] Existing technologies in the anti-body odor space possess a number of limitations. Antiperspirants, so named for their sweat-blocking properties, prevent the body from engaging in natural cooling mechanisms and can be environmentally costly to produce.Atty. Dkt. No. 135523-0115Deodorants, meanwhile, do not prevent the generation of body odor, but merely conceal it with a more potent odor and, in some cases, provide antimicrobial ingredients to reduce bacterial activity and abundance. Both antiperspirants and deodorants are environmentally costly to manufacture, produce significant amounts of plastic waste, and typically require multiple applications over a period of days to retain efficacy. Accordingly, there is a need for more effective deodorizing technologies that do not pose the same negative environmental and health impacts as current anti-body odor products.SUMMARY

[0006] In one aspect, the present disclosure provides a genetically engineered Staphylococcus hominis strain whose genome comprises a disruption of a patB gene. In some embodiments, the strain exhibits decreased expression of the patB gene as compared to a corresponding wild-type Staphylococcus hominis strain. In some embodiments, the disruption prohibits transcription of a full-length wild-type mRNA from the disrupted patB gene. In some embodiments, the disruption prohibits expression of a functional PatB enzyme, inactivates the PatB protein, modifies the PatB protein, or has a negative effect on the expression of the patB gene. In some embodiments, the genetically engineered strain has decreased PatB enzyme activity as compared to a corresponding wild-type Staphylococcus hominis strain. In some embodiments, the modified PatB protein is a truncated protein. In some embodiments, the truncated protein has decreased catalytic activity compared to a corresponding wild-type Staphylococcus hominis strain. In some embodiments, the disrupted patB gene comprises a deletion of all or at least a portion of the gene, optionally wherein the deletion comprises deletion of the patB promoter. In some embodiments, the disrupted patB gene comprises an insertion of a gene cassette. In some embodiments, the disrupted patB gene comprises a deletion of at least a portion of the gene. In some embodiments, the disruption comprises a knockout of the patB gene. In some embodiments, the disruption comprises an allelic replacement knockout. In some embodiments, the disruption comprises a nucleotide substitution. In some embodiments, the nucleotide substitution is a cytosine (C) to a thymine (T). In some embodiments, the nucleotide substitution is at the W190 tryptophan amino acid site in the patB gene. In some embodiments, the nucleotide substitution results in a premature stop-codon. In some embodiments, the Staphylococcus hominis strain comprisesAtty. Dkt. No. 135523-0115SEQ ID NO: 15. In some embodiments, the premature stop codon in the patB gene results in a truncated PatB protein lacking one or more enzymatic activities. In some embodiments, the premature stop codon in the expressed mRNA of the patB gene has the sequence UAG, UGA, or UAA. In some embodiments, the disruption is introduced by homologous recombination, site-directed mutagenesis, a Transcription activator-like effector nuclease (TALEN), a meganuclease, a zinc finger nuclease, a CRISPR / Cas9 system, a CRISPR base editing system, a CRISPR prime editing system, a CRISPR / Cpfl system, a CRISPR / Csml system, or any combination thereof. In some embodiments, the disruption is introduced by a CRISPR base editing system. In some embodiments, the CRISPR based editing system is a cytosine base editor (CBE) system. In some embodiments, the fitness of the strain comprising the disruption is not negatively affected. In some embodiments, the strain has a comparable growth rate as compared to a corresponding wild-type Staphylococcus hominis strain. In some embodiments, the strain comprises Staphylococcus hominis clade A or Staphylococcus hominis clade B. In some embodiments, the strain comprises Staphylococcus hominis deposited with the International Depositary Authority of Canada (ID AC) under IDAC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20 or a variant strain thereof. In some embodiments, the strain has a reduced capacity to produce 3- methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain. In some embodiments, the disruption is a deletion or nucleotide substitution in the DNA sequence of the patB gene encoding a PatB protein, wherein the deletion inactivates the PatB protein, truncates the PatB protein, or has a negative effect on expression of the patB gene. In some embodiments, the genetically engineered Staphylococcus hominis strain has a 16S rRNA gene sequence that is at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to a sequence selected from the group consisting of SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, and SEQ ID NO: 37. In some embodiments, the strain has the 16S rRNA gene sequence represented by a sequence selected from the group consisting of SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ IDAtty. Dkt. No. 135523-0115NO: 36, and SEQ ID NO: 37. In some embodiments, the genetically engineered Staphylococcus hominis strain is derived from a strain deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-03, 030924-04, 300725-02, 300725-01, 300725-05, 300725-07, 300725-08, 300725-09, 300725- 11, 300725-12, 300725-13, or 300725-10 or a variant strain derived therefrom, wherein the genome has been engineered to comprise a disruption of a patB gene.

[0007] In another aspect, the present disclosure provides a genetically engineered Staphylococcus hominis strain deposited with the International Depositary Authority of Canada (IDAC) under IDAC Accession Number 030924-01, 030924-02, 300725-03, 300725- 04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20 or a variant strain derived therefrom, wherein the variant strain is obtained using the deposited strain as starting material, and wherein a reduced capacity of the deposited strain to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain is retained or improved in the variant strain as compared to the deposited strain.

[0008] In a different aspect, the present disclosure provides a Staphylococcus hominis strain, wherein the strain comprises a mutation in at least one endogenous polynucleotide having about 80% to about 100% sequence identity to SEQ ID NO: 14, such that the strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding strain that does not comprise the mutation. In some embodiments, the strain comprises the mutation in at least one endogenous polynucleotide having about 97% to about 100% sequence identity to SEQ ID NO: 14. In some embodiments, the strain grows at an equivalent or improved rate as compared to a Staphylococcus hominis strain lacking the mutation. In some embodiments, the strain is incapable of converting Cys-Gly-3M3SH to 3M3SH.

[0009] In another aspect, the present disclosure provides a composition comprising an effective amount of one or more of the genetically engineered Staphylococcus hominis strains of any one of the preceding embodiments. In some embodiments, the one or more Staphylococcus hominis strains are lyophilized. In some embodiments, the composition further comprises a lyoprotectant. In some embodiments, the lyoprotectant is selected fromAtty. Dkt. No. 135523-0115 one or more of sucrose, micellar casein, trehalose, or inulin. In some embodiments, the one or more Staphylococcus hominis strains are viable. In some embodiments, the one or more Staphylococcus hominis strains are nonproliferative. In some embodiments, the composition further comprises Staphylococcus epidermidis. In some embodiments, the composition further comprises an excipient. In some embodiments, the excipient is selected from one or more of oleic acid, stearic acid, almond butter, avocado butter, babassu butter, beeswax, cocoa butter, coconut butter, coconut oil, cupuacu butter, hemp seed butter, kokum butter, macadamia butter, mango butter, mowrah butter, olive butter, sal seed butter, shea butter, propylene glycol, glycerin, stearyl alcohol, myristyl alcohol, capric triglyceride, polyethylene glycol (PEG), Isopropyl Myristate, or sorbitol. In some embodiments, the Staphylococcus hominis strain is present at a concentration of about 106to about 1013bacteria per gram. In some embodiments, the Staphylococcus hominis strain is present at a concentration of about 107to about 1011of bacteria per gram. In some embodiments, the composition comprises about 90 wt. % to about 99.99 wt. % of the excipient. In some embodiments, the lyophilized Staphylococcus hominis bacteria is about 10 wt. % to about 0.01 wt. %. In some embodiments, the composition comprises about 95 wt. % to about 99.75 wt. % of the excipient. In some embodiments, the lyophilized Staphylococcus hominis bacteria is about 5 wt. % to about 0.25 wt. %. In some embodiments, the composition further comprises a fragrance selected from one or more of elettaria cardamomum seed oil, eucalyptus globulus seed oil, citrus aurantium bergamia fruit oil, mentha piperita oil, salvia sclarea oil, juniperus virginiana oil, abies sibirica oil, citrus aurantium amara, citrus aurantium dulcis lower oil, citrus nobilis oil, rosmarinus officinalis oil, melaleuca alternifolia oil, citrus limon tree oil, zingiber officinale root oil, Mangifera indica oil, Lavandula angustifolia oil, and Santalum album. In some embodiments, the composition is formulated for topical administration. In some embodiments, the composition is formulated for use as a topical deodorant. In some embodiments, the composition further comprises an antiperspirant. In some embodiments, the antiperspirant comprises one or more of aluminum salt, charcoal, and witch hazel. In some embodiments, the topical deodorant is formulated as a deodorant stick, a roll-on, a cream, a lotion, a gel, a powder, a water activated cream, a water activated powder, an injectable, a patch, or a spray. In some embodiments, the composition is a pharmaceuticalAtty. Dkt. No. 135523-0115 composition and the effective amount of one or more of the genetically engineered Staphylococcus hominis strains is a therapeutically effective amount.

[0010] In another aspect, the present disclosure provides a pharmaceutical composition comprising a therapeutically effective amount of one or more genetically engineered Staphylococcus hominis strains whose genome comprises a disruption of a patB gene, and pharmaceutically acceptable carrier or excipient, and optionally, a lyoprotectant. In some embodiments, the disruption of the patB gene comprises a nucleotide substitution. In some embodiments, the nucleotide substitution comprises a cytosine (C) to thymine (T) substitution as compared to the wild-type Staphylococcus hominis strains. In some embodiments, the substitution is at the W190 tryptophan amino acid site in the patB gene. In some embodiments, the substitution introduces a premature stop codon. In some embodiments, the one or more genetically engineered Staphylococcus hominis strains are lyophilized. In some embodiments, the pharmaceutically acceptable carrier or excipient is present at about 90 wt. % to about 99.99 wt. % and the one or more lyophilized genetically engineered Staphylococcus hominis strains are present at about 10 wt. % to about 0.01 wt. %. In some embodiments, the pharmaceutically acceptable carrier or excipient is present at about 95 wt. % to about 99.75 wt. % and the one or more lyophilized genetically engineered Staphylococcus hominis strains are present at about 5 wt. % to about 0.25 wt. %. In some embodiments, the one or more genetically engineered Staphylococcus hominis strains are present at in the composition at a concentration of about 106to about 1013bacteria per gram. In some embodiments, the one or more genetically engineered Staphylococcus hominis strains have a concentration of about 107to about 1011bacteria per gram. In some embodiments, the one or more genetically engineered Staphylococcus hominis strains are selected from the strains deposited with the International Depositary Authority of Canada (IDAC) under IDAC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the International Depositary Authority of Canada (IDAC) under IDAC Accession Number 030924-01. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with IDAC under IDAC Accession Number 030924-02. In some embodiments, theAtty. Dkt. No. 135523-0115 composition comprises the genetically engineered Staphylococcus hominis strain deposited with the ID AC under ID AC Accession Number 300725-03. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the ID AC under ID AC Accession Number 300725-04. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the ID AC under ID AC Accession Number 300725-06. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the IDAC under IDAC Accession Number 300725-14. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the IDAC under IDAC Accession Number 300725-15. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the IDAC under IDAC Accession Number 300725-16. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the IDAC under IDAC Accession Number 300725-17. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the IDAC under IDAC Accession Number 300725-18. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the IDAC under IDAC Accession Number 300725-19. In some embodiments, the composition comprises the genetically engineered Staphylococcus hominis strain deposited with the IDAC under IDAC Accession Number 300725-20. In some embodiments, the composition comprises two or more of the genetically engineered Staphylococcus hominis strains deposited with IDAC under IDAC Accession Numbers 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725- 18, 300725-19, and 300725-20.

[0011] In another aspect, the present disclosure provides a method of reducing 3-methyl-3- sulfanylhexan-l-ol (3M3SH) production from a skin microbiota of a subject, the method comprising applying: (i) a composition or pharmaceutical composition comprising the genetically engineered Staphylococcus hominis strain of any one of the preceding embodiments to the skin of the subject; (ii) the composition of any one of the preceding embodiments; or (iii) the pharmaceutical composition of any one of the preceding embodiments to the skin of the subject.Atty. Dkt. No. 135523-0115

[0012] In another aspect, the present disclosure provides a method for treating, reducing, or preventing malodor on the skin of a subject, comprising treating the skin of the subject with a therapeutically effective amount of the composition of any one of the preceding embodiments or the pharmaceutical composition of any one of the preceding embodiments. In some embodiments, the composition or pharmaceutical composition is applied topically to the skin of the subject. In some embodiments, the composition or pharmaceutical composition is applied to a sweat-gland-containing area of skin of the subject. In some embodiments, the composition or the pharmaceutical composition is administered to the subject about 1 to about 2 times per day for about 1 to about 7 days. In some embodiments, the treated skin of the subject comprises a reduction in the amount of malodorous bacteria as compared to a corresponding untreated portion of skin. In some embodiments, the malodorous bacteria comprises at least wild-type Staphylococcus hominis.

[0013] In another aspect, the present disclosure provides a method of producing a genetically engineered Staphylococcus hominis strain, the method comprising targeting the patB gene for disruption. In some embodiments, the disruption is introduced into the patB gene by homologous recombination, site-directed mutagenesis, a Transcription activator-like effector nuclease (TALEN), a meganuclease, a zinc finger nuclease, a CRISPR / Cas9 system, a CRISPR / Cpfl system, or a CRISPR / Csml system, and any combination thereof. In some embodiments, the disruption is introduced into the patB gene by the CRISPR base editing system. In some embodiments, the CRISPR based editing system is a cytosine base editor (CBE) system. In some embodiments, the CRISPR based editing system comprises transforming a wild-type Staphylococcus hominis strain with a 6-part plasmid. In some embodiments, the 6-part plasmid comprises one or more sequences selected from SEQ ID NOs: 2-7. In some embodiments, the 6 part plasmid comprises the sequence of SEQ ID NO: 1. In some embodiments, the CRISPR based editing system comprises transforming a wildtype Staphylococcus hominis strain with an 8-part plasmid. In some embodiments, the 8-part plasmid comprises one or more sequences selected from SEQ ID NOs: 17-24. In some embodiments, the 8-part plasmid comprises the sequence of SEQ ID NO: 16. In some embodiments, the strain exhibits decreased expression of the patB gene as compared to a corresponding wild-type Staphylococcus hominis strain. In some embodiments, the disruption prohibits transcription of a full-length wild-type mRNA from the disrupted patBAtty. Dkt. No. 135523-0115 gene. In some embodiments, the disruption prohibits expression of a functional PatB enzyme. In some embodiments, the genetically engineered strain has decreased PatB enzyme or catalytic activity and the PatB protein is truncated as compared to a corresponding wildtype Staphylococcus hominis strain. In some embodiments, the disrupted patB gene comprises a nucleotide substitution, wherein the substitution is a cytosine (C) to a thymine (T). In some embodiments, the strain is selected from Staphylococcus hominis strains deposited with the IDAC under IDAC Accession Number 030924-01, 030924-02, 300725- 03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20, or a variant strain thereof. In some embodiments, the genetically engineered Staphylococcus hominis strain has a reduced capacity to produce 3-methyl-3- sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain.

[0014] In another aspect, the present disclosure provides an altered microbiota comprising a genetically engineered Staphylococcus hominis strain, or a variant strain thereof, whose genome comprises a disruption of a patB gene. In some embodiments, the microbiota has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat as compared to a corresponding wild-type microbiota.

[0015] In a different aspect, the present disclosure provides a deodorant composition comprising: (i) a therapeutically effective amount of a viable Staphylococcus hominis strain whose genome comprises a disruption of a patB gene; and (ii) a therapeutically acceptable carrier or excipient; wherein the deodorant composition is formulated as a water activated formulation for topical administration. In some embodiments, the deodorant composition comprises about 90 wt. % to about 99.99 wt. % therapeutically acceptable carrier or excipient. In some embodiments, the Staphylococcus hominis strain is lyophilized and the deodorant composition comprises about 10 wt. % to about 0.01 wt. % lyophilized Staphylococcus hominis bacteria. In some embodiments, the deodorant composition comprises about 95 wt. % to about 99.75 wt. % therapeutically acceptable carrier or excipient. In some embodiments, the deodorant composition comprises about 5 wt. % to about 0.25 wt. % lyophilized Staphylococcus hominis bacteria. In some embodiments, the Staphylococcus hominis strain is present at a concentration of about 106to about 1016bacteriaAtty. Dkt. No. 135523-0115 per gram. In some embodiments, the Staphylococcus hominis strain is present at a concentration of about 107to about 1011bacteria per gram. In some embodiments, the deodorant composition further comprises: (iii) a lyoprotectant. In some embodiments, the Staphylococcus hominis strain comprises one of the strains deposited with the ID AC under IDAC Accession Numbers 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20.

[0016] In another aspect, the present disclosure provides a deodorant stick comprising the deodorant composition of any one of the preceding embodiments.

[0017] In one aspect, the present disclosure provides a pharmaceutical composition comprising a therapeutically effective amount of one or more of the genetically engineered Staphylococcus hominis strains of any one of the preceding embodiments, and a therapeutically acceptable carrier or excipient, for use in the treatment of malodor on the skin of a subject. In some embodiments, the pharmaceutical composition further comprises a lyoprotectant.

[0018] In another aspect, the present disclosure provides use of the one or more genetically engineered Staphylococcus hominis strains of any one of the preceding embodiments in the manufacture of a medicament for the treatment of malodor on the skin of a subject.

[0019] In some embodiments of any aspect of the present disclosure, the Staphylococcus hominis strain is gene edited to comprise a disruption of a patB gene. Accordingly, the disclosure provides a gene edited Staphylococcus hominis strain whose genome comprises a disruption of a patB gene. In some embodiments, the strain does not comprise any heterologous DNA.BRIEF DESCRIPTION OF THE DRAWINGS

[0020] FIGs. 1A-1L show for each of the 12 wild-type (WT) and mutant ( patB) S. hominis strains the patB allele of the wild-type parent strain aligned to the sequence of its corresponding mutant strain. Each mutant strain contains a premature stop codon at codon 190 in the patB gene (W190*), denoted by a black box. The sequence of each strain was confirmed with Sanger sequencing. FIGs. 1A-1B are diagrams showing results of ampliconAtty. Dkt. No. 135523-0115 sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00037 as compared to the WT parent Sn00016 and in strain Sn00048 as compared to the WT parent Sn00040. FIG. 1C shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00079 as compared to the WT parent Sn00042. FIG. ID shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W 190 in strain Sn00082 as compared to the WT parent Sn00015. FIG. IE shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TGA) at W 190 in strain Sn00086 as compared to the WT parent Sn00085. FIG. IF shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00325 as compared to the WT parent Sn00201. FIG. 1G shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00327 as compared to the WT parent Sn00202. FIG. 1H shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00330 as compared to the WT parent Sn00203. FIG. II shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00337 as compared to the WT parent Sn00213. FIG. 1J shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00339 as compared to the WT parent Sn00215. FIG. IK shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00343 as compared to the WT parent Sn00239. FIG. IL shows the results of amplicon sequencing to confirm insertion of a premature stop codon (TAA) at W190 in strain Sn00377 as compared to the WT parent Sn00210.

[0021] FIG. 2 is a graph of the growth curves for strain Sn00016 (also known as Staphylococcus hominis Clade A or AH5009, and designated in the graph with a “1”), strain Sn00037 (designated in the graph with a “2”), strain Sn00040 (also known as Staphylococcus hominis Clade B, and designated in the graph with a “3”), and strain Sn00048 (designated in the graph with a “4”) grown separately in triplicate, demonstrating that the disruption of patB in the strains did not significantly impact bacterial growth rate.Atty. Dkt. No. 135523-0115

[0022] FIG. 3A is a bar graph of the production of 3M3SH by (from left to right along the x- axis) strain Sn00016, strain Sn00037, strain Sn00040, strain Sn00048, S. epidermidis control, and E. coli control. FIG. 3B is a bar graph showing the ability of each of 12 wild-type S. hominis strains and their corresponding patB W190* mutants to produce 3M3SH (wild-type and engineered strains are paired from left to right along the x-axis, see the table in Example 4). When cultures were incubated with precursor molecule Cys-Gly-3M3SH, all wild-type strains exhibited production of 3M3SH and each engineered strain did not, demonstrating that patB W190* mutants ablate the production of 3M3SH in vivo.

[0023] FIG. 4 shows the phylogenetic diversity of S. hominis patB W190* mutants. A maximum-likelihood phylogeny of core genome positions from 12 engineered S. hominis strains, their wild-type counterparts, and S. hominis NCBI reference genomes demonstrates the high diversity of engineered S. hominis strains. The inset phylogeny of 5 engineered strains and their wild-type counterparts (lower right) is included to demonstrate their genetic diversity, which is not visible on the scale of the species-level phylogeny. The distance to the most recent common ancestor (dMRCA) of the presented strains and of the S. hominis species is shown to be identical. Trees are rooted at the midpoint.

[0024] FIG. 5 shows that the engineered S. hominis patB W190* mutants of the present disclosure are as diverse as S. hominis NCBI strains. The pairwise evolutionary distances amongst the genomes of NCBI strains (black) and between NCBI strains and engineered strains (gray) are not significantly different (2-sample Kolmogorov-Smirnov test), demonstrating the high phylogenetic diversity of engineered strains, which is as broad as the phylogenetic diversity of the known S. hominis species-level phylogeny.

[0025] FIG. 6 shows that the engineered S. hominis patB W190* mutants of the present disclosure engraft onto the human axilla for at least one week and reduce malodor. A single topical application was applied to both axillae of six adult volunteers. Engraftment was assessed at days 3 and 7 post-application by amplicon PCR targeting a conserved patB region flanking codon 190. Engraftment of S. hominis patB W190* mutants was detected for all six subjects at both time points. Boxes indicate per-subject, per-timepoint outcomes; “Y’7“N” within boxes denote participants’ self-reported odor reduction on questionnaires administered before sampling.Atty. Dkt. No. 135523-0115

[0026] FIG. 7 shows that the lyophilized S. hominis patB W190* mutants remain viable for at least 7 months. Viability was assessed on Day 0 (lyophilization day) and at 32, 115, and 209 days post-lyophilization. Lyophilized powder containing S. hominis patB W190* was reconstituted in PBS and plated on TSA, showing high viable counts at all time points with no discernible decrease from 32 to 209 days. Plotted points are means across replicate CFU plating experiments; error bars represent 2 standard deviations.DETAILED DESCRIPTION

[0027] It is to be appreciated that certain aspects, modes, embodiments, variations and features of the present technology are described below in various levels of detail in order to provide a substantial understanding of the present technology. The definitions of certain terms as used in this specification are provided below. Unless defined otherwise, all technical and scientific terms used herein generally have the same meaning as commonly understood by one of ordinary skill in the art to which this present technology belongs.I. Definitions

[0028] The following terms are used herein, the definitions of which are provided for guidance.

[0029] As used herein, the singular forms “a,” “an,” and “the” designate both the singular and the plural, unless expressly stated to designate the singular only.

[0030] The term “about” and the use of ranges in general, whether or not qualified by the term about, means that the number comprehended is not limited to the exact number set forth herein, and is intended to refer to ranges substantially within the quoted range while not departing from the scope of the present technology. As used herein, “about” will be understood by persons of ordinary skill in the art and will vary to some extent on the context in which it is used. If there are uses of the term which are not clear to persons of ordinary skill in the art given the context in which it is used, “about” will mean up to plus or minus 10% of the particular term.

[0031] As used herein, “administration” of an agent, drug, bacterial strain(s) or composition of the present technology to a subject includes any route of introducing or delivering to aAtty. Dkt. No. 135523-0115 subject a compound to perform its intended function. Administration can be carried out by any suitable route, including topically. In some embodiments, the compositions of the present technology are formulated for topical administration. In some embodiments, the compositions are formulated as a stick, a roll-on, a cream, a lotion, a gel, a powder, a water activated formulation (e.g., a water activated cream or a water activated powder), an injectable, a patch, or a spray. In some embodiments, the compositions are formulated for use as a probiotic. As used herein, administration includes self-administration and administration by another.

[0032] As used herein, “Strain Sn00016” or “Sn00016” or “AH5009” or “Horswill Lab Strain AH5009,” refers to a bacterial strain having been deposited under ID AC Accession No. 030924-03, or compositions comprising the strain. Strain Sn00016 is a wild-type strain and considered a member of Staphylococcus hominis and is from Clade A. In some embodiments, “ Staphylococcus hominis Clade A” refers to a strain of the species Staphylococcus hominis that belongs to Clade A, having a common ancestor.

[0033] As used herein, “Strain Sn00040” or “Sn00040” refers to a bacterial strain having been deposited under ID AC Accession No. 030924-04, or compositions comprising the strain. Strain Sn00040 is a wild-type strain and considered a member of Staphylococcus hominis from Clade B. In some embodiments, “ Staphylococcus hominis Clade B” refers to a strain of the species Staphylococcus hominis that belongs to Clade B, having a common ancestor.

[0034] As used herein, “Strain Sn00037” or “Sn00037” refers to a bacterial strain having been deposited under IDAC Accession No. 030924-01, or compositions comprising the strain. Strain Sn00037 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00016. Strain Sn00016 is deposited under IDAC Accession No. 030924-03.

[0035] As used herein, “Strain Sn00048” or “Sn00048” refers to a bacterial strain having been deposited under IDAC Accession No. 030924-02, or compositions comprising the strain. Strain Sn00048 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00040. Strain Sn00040 is deposited under IDAC Accession No. 030924-04.Atty. Dkt. No. 135523-0115

[0036] As used herein, “Strain Sn00079” or “Sn00079” refers to a bacterial strain having been deposited under ID AC Accession No. 300725-03, or compositions comprising the strain. Strain Sn00079 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00042. Strain Sn00042 is deposited under ID AC Accession No. 300725-02.

[0037] As used herein, “Strain Sn00082” or “Sn00082” refers to a bacterial strain having been deposited under ID AC Accession No. 300725-04, or compositions comprising the strain. Strain Sn00082 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00015. Strain Sn00015 is deposited under IDAC Accession No. 300725-01.

[0038] As used herein, “Strain Sn00086” or “Sn00086” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-06, or compositions comprising the strain. Strain Sn00086 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00085. Strain Sn00085 is deposited under IDAC Accession No. 300725-05.

[0039] As used herein, “Strain Sn00325” or “Sn00325” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-14, or compositions comprising the strain. Strain Sn00325 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00201. Strain Sn00201 is deposited under IDAC Accession No. 300725-07.

[0040] As used herein, “Strain Sn00327” or “Sn00327” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-15, or compositions comprising the strain. Strain Sn00327 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00202. Strain Sn00202 is deposited under IDAC Accession No. 300725-08.

[0041] As used herein, “Strain Sn00330” or “Sn00330” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-16, or compositions comprising the strain. Strain Sn00330 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00203. Strain Sn00203 is deposited under IDAC Accession No. 300725-09.

[0042] As used herein, “Strain Sn00337” or “Sn00337” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-17, or compositions comprising the strain. Strain Sn00337 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00213. Strain Sn00213 is deposited under IDAC Accession No. 300725-11.Atty. Dkt. No. 135523-0115

[0043] As used herein, “Strain Sn00339” or “Sn00339” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-18, or compositions comprising the strain. Strain Sn00339 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00215. Strain Sn00215 is deposited under IDAC Accession No. 300725-12.

[0044] As used herein, “Strain Sn00343” or “Sn00343” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-19, or compositions comprising the strain. Strain Sn00343 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00239. Strain Sn00239 is deposited under IDAC Accession No. 300725-13.

[0045] As used herein, “Strain Sn00377” or “Sn00377” refers to a bacterial strain having been deposited under IDAC Accession No. 300725-20, or compositions comprising the strain. Strain Sn00377 is a ApatB-S. hominis strain derived from the wild-type S. hominis strain Sn00210. Strain Sn00210 is deposited under IDAC Accession No. 300725-10.

[0046] As used herein, “ ApatB-Staphylococcus hominis" or “ ApatB-S. hominis" refers to a Staphylococcus hominis whose genome comprises a disruption of a patB gene or compositions comprising ApatB-Staphylococcus hominis. As used herein, “Strain Sn00037” or “Sn00037” or “ApatB of Sn00016” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00016. As used herein, “Strain Sn00048” or “Sn00048” or “ApatB of Sn00040” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00040. As used herein, “Strain Sn00079” or “Sn00079” or “ApatB of Sn00042” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00042. As used herein, “Strain Sn00082” or “Sn00082” or “ApatB of Sn00015” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00015. As used herein, “Strain Sn00086” or “Sn00086” or “ApatB of Sn00085” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00085. As used herein, “Strain Sn00325” or “Sn00325” or “ApatB of Sn00201” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00201. As used herein, “Strain Sn00327” or “Sn00327” or “ApatB of Sn00202” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00202. As used herein, “Strain Sn00330” or “Sn00330” or “ApatB of Sn00203” refers to a strain that comprises a disruption of the patBAtty. Dkt. No. 135523-0115 gene where the wild-type strain is Strain Sn00203. As used herein, “Strain SnOO337” or “Sn00337” or “ApatB of Sn00213” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00213. As used herein, “Strain Sn00339” or “Sn00339” or “ApatB of Sn00215” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00215. As used herein, “Strain Sn00343” or “Sn00343” or “ApatB of Sn00239” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00239. As used herein, “Strain Sn00377” or “Sn00377” or “ApatB of Sn00210” refers to a strain that comprises a disruption of the patB gene where the wild-type strain is Strain Sn00210.

[0047] In some embodiments, the ApatB -Staphylococcus hominis comprises a variant of strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377. In some embodiments, the ApatB-Staphylococcus hominis is from a Clade selected from Clade A or Clade B. In some embodiments, the ApatB-Staphylococcus hominis is from Clade A. In some embodiments, the ApatB- Staphylococcus hominis is from Clade B. In some embodiments, the ApatB-Staphylococcus hominis comprises a single base pair point mutation. In some embodiments, the ApatB- Staphylococcus hominis comprises a cytosine (C) to a thymine (T) point mutation. In some embodiments, the point mutation is in the patB coding region. In some embodiments, the point mutation is in the W190 tryptophan amino acid site in patB. In some embodiments, the ApatB-Staphylococcus hominis exhibits a reduced PatB activity and / or patB expression. In some embodiments, the ApatB-Staphylococcus hominis has a reduced capacity to produce thiol and / or 3M3SH when contacted with an appropriate substrate and / or precursor. In some embodiments, ApatB-Staphylococcus hominis comprises Strain Sn0037 having been deposited under ID AC Accession No. 030924-01, Strain Sn00048 having been deposited under ID AC Accession No. 030924-02, Strain Sn00079 having been deposited under ID AC Accession No. 300725-03, Strain Sn00082 having been deposited under ID AC Accession No. 300725-04, Strain Sn00086 having been deposited under IDAC Accession No. 300725-06, Strain Sn00325 having been deposited under IDAC Accession No. 300725-14, Strain Sn00327 having been deposited under IDAC Accession No. 300725-15, Strain Sn00330 having been deposited under IDAC Accession No. 300725-16, Strain Sn00337 having been deposited under IDAC Accession No. 300725-17, Strain Sn00339 having been depositedAtty. Dkt. No. 135523-0115 under IDAC Accession No. 300725-18, Strain Sn00343 having been deposited under IDAC Accession No. 300725-19, Strain Sn00377 having been deposited under IDAC Accession No. 300725-20, or a variant strain thereof, or compositions comprising one or more of the strains and / or one or more variant strains thereof. In some embodiments, ApatB-S. hominis comprises a S. hominis strain that expresses a truncated PatB protein, i.e. not the full-length PatB protein. In some embodiments, ApatB-S. hominis comprises a S. hominis strain with phenotypic loss-of-function for the patB gene (e.g.,patB~, which can be used interchangeably herein with ApatB-S. hominis). In some embodiments, the disruption is a non-synonymous (missense) substitution that yields a full-length PatB polypeptide having reduced or abolished P-lyase enzymatic activity sufficient to decrease 3M3SH production as compared to a corresponding wild-type strain. In certain embodiments, the non-synonymous substitution alters a catalytic, substrate-binding, cofactor-binding, folding, or stability determinant of PatB without truncating the protein. In some embodiments, ApatB-S. hominis comprises a single amino acid substitution such that 3M3SH production is eliminated.

[0048] As used herein, “variant” or “variant strain” refers to a strain derived from a strain of the present technology by any means, such as, but not limited to, genetic engineering, radiation and / or chemical treatment, and / or selection, adaptation, screening, etc. In some embodiments, the variant is naturally occurring. In some embodiments, the variant is selected for or engineered. In some embodiments, the variant is a functionally equivalent variant, e.g., a variant that has the same, or improved, properties with respect to its capacity to produce a thiol or 3M3SH when contacted with an appropriate substrate and / or precursor as the mother or wild-type strain. Such variants are a part of the present technology. In some embodiments, the term “variant” refers to a strain obtained by subjecting a strain of the present technology to any conventionally used mutagenization treatment including treatment with a chemical mutagen such as ethane methane sulphonate (EMPS) or / ' -methyl -N’-nitro- N-nitroguanidine (NTG), UV light, or to a spontaneously occurring variant. In embodiments in which variants are subjected to mutagenization, one of skill in the art may refer to the variant as a mutant. A variant may have been subjected to several mutagenization treatments (a single treatment should be understood as one mutagenization step followed by a screening / selection step). In some embodiments, less than 1%, less than 0.1%, less than 0.01%, less than 0.001%, or less than 0.0001% of nucleotides in the bacterial genome haveAtty. Dkt. No. 135523-0115 been changed (such as by replacement, insertion, deletion, or a combination thereof) compared to the mother strain. Such variants, which may be identified by using appropriate screening techniques, are a part of the present technology.

[0049] As used herein, the term “effective amount” refers to a quantity sufficient to achieve a desired therapeutic and / or prophylactic effect, e.g., an amount which results in the prevention or treatment of a condition, and / or symptom(s) thereof. In the context of therapeutic or prophylactic applications, the amount of a composition or formulation administered to the subject will depend on the type and severity of the condition and on the characteristics of the subject, such as general health, age, sex, body weight, and lifestyle, for example. It will also depend on the degree, severity, and type of condition, for example. The skilled artisan will be able to determine appropriate dosages depending on these and other factors. In some embodiments, a single dose is administered. In some embodiments, multiple doses are administered. Additionally, or alternatively, in some embodiments, multiple therapeutic compositions or compounds or formulations (e.g., pharmaceutical compositions comprising multiple bacterial strains alone or in combination with additional active agents, such as deodorizing or anti-perspiring agents). In the methods described herein, compositions comprising the bacterial strains of the present technology may be administered to a subject having, or at risk of having, body odor. In some embodiments, the compositions comprising the bacterial strains of the present technology may be administered to a subject unknown to have, or unknown to be at risk of having, body odor. In some embodiments, the compositions comprising the bacterial strains of the present technology may be administered to multiple locations on the body of a subject. For example, an “effective amount” of the compositions of the present technology, includes levels at which the presence, frequency, or severity of body odor is at a minimum reduced or ameliorated. In some embodiments, an effective amount reduces, prevents, or ameliorates the presence of body odor, and / or the likelihood of developing body odor. In some embodiments, an effective amount is achieved by multiple administrations. In some embodiments, an effective amount is achieved with a single administration.Atty. Dkt. No. 135523-0115

[0050] As used herein, the terms “freeze-dried” or “freeze-drying” and “lyophilized” or “lyophilization” are used interchangeably and refer to a process that removes water from a product after it is frozen and placed under a vacuum and the products produced therefrom.

[0051] As used herein, the term “spray drying” refers to a process in which a liquid product is atomized into a hot drying chamber causing the liquid to evaporate rapidly resulting in a fine powder. In some embodiments, the process of spray drying does not require the use of lyoprotectants.

[0052] As used herein, “desiccated” refers to a process in which liquid content of a product is significantly reduced through drying. Non-limiting examples of processes that yield a desiccated product include freeze-drying or spray drying. In some embodiments, desiccated cells can be the Staphylococcus hominis bacterial cells as disclosed herein.

[0053] “Desiccated’ refers to cells that have had their moisture content significantly reduced through drying, without specifying the exact drying method. This term effectively covers cells that have been dehydrated through various processes, including both freeze-drying and spray-drying, while still implying that the cells may retain some level of viability and can potentially be rehydrated to resume their metabolic functions.

[0054] As used herein, the term “lyoprotectant(s)” refers to an agent that prevents or ameliorates any damage caused to a product during the lyophilization process. In some embodiments, the lyoprotectant is a protein, a carbohydrate, or a combination thereof. In some embodiments, the lyoprotectant is a milk protein, and in particular a micellar casein. In some embodiments, the carbohydrate is sucrose. In some embodiments, the lypoprotectant is trehalose, inulin, or a combination thereof. In some embodiments, the lyoprotectant prevents or ameliorates any damage caused to a bacterial strain of the present technology during the lyophilization process.

[0055] As used herein, “acceptable carrier and / or diluent” or “acceptable excipient” includes but is not limited to any combination of solvents, dispersion media, coatings, antifungal agents, isotonic and absorption delaying agents, and the like. In some embodiments, the acceptable carrier comprises oleic acid, stearic acid, almond butter, avocado butter, babassu butter, beeswax, cocoa butter, coconut butter, coconut oil, cupuacu butter, hemp seed butter,Atty. Dkt. No. 135523-0115 kokum butter, macadamia butter, mango butter, mowrah butter, olive butter, sal seed butter, shea butter, propylene glycol, glycerin, stearyl alcohol, myristyl alcohol, capric triglyceride, polyethylene glycol (PEG), Isopropyl Myristate, sorbitol, or any combination thereof. The use of such agents with biologically active substances is well known in the art. Further details of excipients are provided below. Supplementary active ingredients, such as antifungal agents, antiperspirants, fragrances, and deodorants, can also be incorporated into the compositions.

[0056] As used herein, “acceptable excipient” or “excipient” refers to substances and compositions that do not produce an adverse, allergic, or other untoward reaction when administered to an animal or a human. As used herein, the term includes all inert, non-toxic, liquid or solid fillers, or diluents that do not react with the strains of the present technology in an inappropriate negative manner, including solvents, dispersion media, coatings, antifungal agents, isotonic and absorption delaying agents, preservatives and the like, for example liquid pharmaceutical carriers e.g., sterile water, saline, sugar solutions, Tris buffer, ethanol and / or certain oils. In some embodiments, the excipient is polyethylene glycol (PEG), Isopropyl Myristate, or sorbitol. In some embodiments, the excipient is polyethylene glycol (PEG). In some embodiments, the excipient is Isopropyl Myristate. In some embodiments, the excipient is sorbitol.

[0057] As used herein, “deodorant” refers to a substance which removes and / or conceals an unpleasant smell, such as body odor. In some embodiments, the strains of the present technology are formulated as deodorant compositions. In some embodiments, the deodorant compositions further comprise a fragrance. In some embodiments, the fragrance comprises one or more of elettaria cardamomum seed oil, eucalyptus globulus seed oil, citrus aurantium bergamia fruit oil, mentha piperita oil, salvia sclarea oil, juniperus virginiana oil, abies sibirica oil, citrus aurantium amara, citrus aurantium dulcis lower oil, citrus nobilis oil, rosmarinus officinalis oil, melaleuca altemifolia oil, citrus limon tree oil, zingiber officinale root oil, Mangifera indica oil, Lavandula angustifolia oil, and Santalum album. In some embodiments, the strains and compositions of the present technology are administered with a fragrance, either simultaneously or sequentially and either as separate formulations or a single formulation.Atty. Dkt. No. 135523-0115

[0058] As used herein “antiperspirant” refers to a substance that inhibits, reduces, or prevents the release of sweat from glands. In some embodiments, the strains and compositions of the present technology are administered with an antiperspirant. In some embodiments, the strains of the present technology are formulated as a deodorant that further comprises an antiperspirant. In some embodiments, the antiperspirant comprises one or more of aluminum salt, charcoal, and witch hazel.

[0059] The terms “disruption” and “mutation” are used interchangeably herein to refer to a detectable and heritable change in the genetic material. Disruptions or mutations may include insertions, deletions, substitutions (e.g., transitions, transversion, homologous recombination), transpositions, inversions, knockouts and combinations thereof. For example, in some embodiments, a disruption or mutation may include: (a) the introduction, substitution, or removal of one or more nucleotides in a Staphylococcus hominis patB gene (or an open reading frame (ORF) thereof), or the introduction, substitution, or removal of one or more nucleotides in a regulatory element required for the transcription or translation of the patB gene or ORF thereof; (b) a patB gene conversion; (c) a patB gene deletion; (d) the down-regulation of the patB gene; (e) specific mutagenesis of the patB gene; and / or (f) random mutagenesis of the patB gene. Disruptions or mutations of the patB gene, as used herein, include any genetic modification that prevents or reduces the capability of the S. hominis strains of the present technology from producing a functional patB gene product. Such disruptions or mutations may include complete or partial deletion of any portion of patB, including a polypeptide-coding sequence, a promoter, an enhancer, a ribosome-binding site (RBS), or other regulatory elements, or mutagenesis of the same, where mutagenesis encompasses substitutions, insertions, deletions, inversions, and any combinations and variations thereof that inactivate patB and prevent or reduce the production of a patB functional gene product. Mutations or disruptions may involve only a single nucleotide (e.g., a point mutation, nucleotide substitution, or a single nucleotide polymorphism) or multiple nucleotides. In some embodiments, the mutation or disruption may be a cytosine (C) to thymine (T) substitution. In some embodiments, the conversion may occur by a cytosine- based editor (CBE) tool. In some embodiments, the CBE precisely converts a cytosine (C) to thymine (T) without causing a double-stand break. In some embodiments, the cytosine (C) to thymine (T) occurs at the W 190 tryptophan amino acid site in PatB. In some embodiments,Atty. Dkt. No. 135523-0115 the cytosine (C) to thymine (T) introduces a premature stop code in the expressed mRNA (z.e., UAG, UGA, or UAA). In some embodiments, the premature stop codon introduces an early termination signal in the mRNA sequence. In some embodiments, the early termination signal leads to production of a truncated protein. In some embodiments, the truncated protein has a loss of catalytic activity. In some embodiments, the mutation or disruption causes a phenotypic change, for example, the expression level of the encoded product is altered, or the encoded product itself is altered. In some embodiments, a disruption or mutation may result in a disrupted gene with decreased levels of expression of a gene product (e.g., protein or RNA) as compared to the wild-type strain. In other embodiments, a disruption or mutation may result in an expressed protein with activity that is lower as compared to the activity of the expressed protein from the wild-type strain. In some embodiments, a gene disruption or mutation results in the removal of the gene from the genome via a substitution, such as homologous recombination. In some embodiments, a functional patB gene product enables the metabolic conversion of Cys-Gly-3M3SH to 3M3SH. Accordingly, in some embodiments, a disruption or mutation of the patB gene, as used herein, includes any genetic modification that prevents or reduces the capability of the S. hominis strains of the present technology from converting Cys-Gly-3M3SH to 3M3SH. This includes, but is not limited to, disruption of one or more PatB enzymatic activities or alteration of PatB binding with one or more metabolites or co-factors or the decreases to the relative enzymatic activity with respect to alternative substrates. Disruptions or mutations outside of the patB coding region that accomplish the effect of disrupted PatB activity are also encompassed within this disclosure, including disruptions or mutations in enhancer, repressor, promoter or other regulatory elements that result in decreased transcription or translation of the patB gene. In some embodiments, the nucleotides encoding a single amino acid are altered (e.g., replaced or deleted) which results in an elimination of 3M3SH production in the strain. Other methods may also be used to disrupt PatB activity, including, but not limited to engineered transcription factors, including Transcription Activator-Like Effectors (TALEs) that repress patB expression, antisense RNAs or small regulatory RNAs that repress patB translation, CRISPR-dCasl3 constructs that target patB RNA and prevent translation, and protein-based or small molecule constructs that post translationally inactivate or repress PatB protein activity. For example, the present disclosure explicitly contemplates constructs (e.g.,Atty. Dkt. No. 135523-0115 recombinant proteins) that post-translationally modify (e.g., phosphorylate) and / or alter the enzymatic properties of the PatB protein to reduce or inhibit enzymatic activity or metabolite binding by the PatB protein. Any construct, and method of use thereof, that downregulates, prevents, or inhibits in any way the transcription, translation, or protein activity of PatB is encompassed within the present disclosure. As used herein, “A,” for example as used in “ApatB,” is intended to encompass any disruption(s) or mutation(s) to the patB gene described herein.

[0060] The term “engineered” or “genetically engineered” is used herein to refer to an organism that has been manipulated to be genetically altered, modified, or changed, e.g., by disruption / editing of the genome. For example, an “engineered” or “genetically engineered” bacterial strain encompasses a bacterial strain that has been manipulated to be genetically altered, modified, or changed.

[0061] As used herein, the term “fitness” or “bacterial fitness” refers to the capacity of a bacterium, bacterial strain, or bacterial species to survive and reproduce in a given environment.

[0062] A “knocked out gene” or a “gene deletion” refers to a gene including a null mutation (e.g., the wild-type product encoded by the gene is not expressed, expressed at levels so low as to have no effect, or is non-functional). In some embodiments, the knocked out gene includes heterologous sequences or genetically engineered non-functional sequences of the gene itself, which renders the gene non-functional. In other embodiments, the knocked out gene is lacking a portion of the wild-type gene. For example, in some embodiments, at least about 10%, at least about 20%, at least about 30%, at least about 40% or at least about 60% of the wild-type gene sequence is deleted. In other embodiments, the knocked out gene is lacking at least about 70%, at least about 75%, at least about 80%, at least about 90%, at least about 95% or at least about 100% of the wild-type gene sequence. In other embodiments, the knocked out gene may include up to 100% of the wild-type gene sequence (e.g., some portion of the wild-type gene sequence may be deleted) but also include one or more heterologous and / or non-functional nucleic acid sequences inserted therein.Atty. Dkt. No. 135523-0115

[0063] The term “gene cassette” is used herein to refer to a DNA sequence encoding and capable of expressing one or more genes of interest (e.g., a metabolic gene, a selectable marker, or a combination thereof) that can be inserted between one or more selected restriction sites of a DNA sequence. In some embodiments, insertion of a gene cassette results in a disrupted gene. In some embodiments, disruption of the gene involves replacement of at least a portion of the gene with a gene cassette, which includes a nucleotide sequence encoding a gene of interest (e.g., a metabolic gene, a selectable marker, or a combination thereof).

[0064] As used herein, “probiotic” refers to bacteria that supplements or replaces elements of endogenous flora of a subject, which when administered to the subject confers a beneficial prophylactic and / or therapeutic effect on the subject. In some embodiments, the beneficial effect is a reduction or prevention in malodor on the skin of a subject.

[0065] As used herein, “prevention,” “prevent,” or “preventing” of a condition refers to, in a statistical sample, reduction in the occurrence or recurrence of the condition in treated subjects / samples relative to an untreated controls, or refers to delays in the onset of one or more symptoms of the condition relative to the untreated controls. In some embodiments, the condition is malodor (body odor) on the skin of the subject.

[0066] As used herein, to “reduce” or “reducing” a condition refers to results that, in a sample or specific subject, make the occurrence of the condition better or more tolerable in a sample or subject administered a therapeutic agent (e.g., a ApatB-Staphylococcus hominis strain or compositions comprising the strain) relative to a control sample or subject. In some embodiments, the condition is malodor (body odor) on the skin of the subject.

[0067] As used herein “subject” and “patient” are used interchangeably. In some embodiments, the subject is an animal subject. In some embodiments, the animal subject is a mammal. In some embodiments, the mammalian subject is a human.

[0068] As used herein, the term “simultaneous” administration refers to the administration of at least two agents by the same route and at the same time or at substantially the same time.Atty. Dkt. No. 135523-0115

[0069] As used herein, the term “separate” administration refers to an administration of at least two agents at the same time or at substantially the same time by different routes.

[0070] As used herein, the term “sequential” administration refers to administration of at least two agents at different times, the administration route being identical or different. More particularly, sequential use refers to the whole administration of one agent before administration of the other agent(s) commences. It is thus possible to administer one of the agents over several minutes, hours, or days before administering another.

[0071] A “synergistic effect” refers to a greater-than-additive effect which is produced by a combination of at least two agents, and which exceeds that which would otherwise result from the individual administration of the agents. For example, use of bacterial strain(s) of the present technology in conjunction with other agents for deodorizing or anti-perspiring may result in a greater than additive effect. In some embodiments, the synergistic effect may permit the use of lower doses of bacterial strain(s) of the present technology and / or other agents than would be required if each were used alone.

[0072] “Treating,” “treat,” “treated,” or “treatment” of a condition includes: (i) inhibiting the condition, z.e., arresting its development; (ii) relieving the condition, z.e., causing its regression; (iii) slowing progression of the condition; and / or (iv) inhibiting, relieving, or slowing progression of one or more symptoms of the condition. In some embodiments, the condition is malodor (body odor) on the skin of the subject.

[0073] As used herein, “3-methyl-3-sulfanylhexan-l-ol” and “3M3SH” are interchangeably used to refer to the alcohol compound 3-methyl-3-sulfanylhexan-l-ol having a formula of C7H16OS (CAS Number 307964-23-4). In some embodiments, a 3M3SH precursor is a compound comprising 3M3SH with additional chemical structural components. In some embodiments, Cys-Gly-3M3SH is a 3M3SH precursor.

[0074] It is to be appreciated that the various modes of treatment or prevention of medical diseases and conditions as described are intended to mean “substantial,” which includes total but also less than total treatment or prevention, and wherein some biologically or medically relevant result is achieved.Atty. Dkt. No. 135523-0115II. Sweat and Body Odor

[0075] The armpit offers two microbial habitats: the skin surface and the skin subsurface. The skin surface is the smaller of the two skin microbial habitats, with about 2 m2of surface area. It is also a harsher environment for bacteria as it is frequently disinfected through washing and deodorant use. The skin subsurface has a much larger surface area of about 24 m2and includes the epidermis, dermis, hair follicles, and sweat glands, all of which harbor skin bacteria (Gallo RL. Human Skin Is the Largest Epithelial Surface for Interaction with Microbes. J Invest Dermatol. 2017 Jun; 137(6): 1213-1214. doi: 10.1016 / j jid.2016.11.045. Epub 2017 Apr 8. PMID: 28395897; PMCID: PMC5814118; Nakatsuji T, Chiang HI, Jiang SB, Nagarajan H, Zengler K, Gallo RL. The microbiome extends to subepidermal compartments of normal skin. Nat Commun. 2013;4: 1431. doi: 10.1038 / ncomms2441. PMID: 23385576; PMCID: PMC3655727). The hair follicle provides a stable, moist, well- perfused, and UV light-protected environment. Sweat glands interface with both of these habitats and produce sweat, which bacteria feed on. Sweat glands can be categorized into apocrine glands and eccrine glands, although several additional types exist.

[0076] Eccrine glands open onto the skin surface and produce eccrine sweat, a saline solution that contains trace chemicals originating from interstitial fluid, such as potassium, bicarbonate, lactate, urea, amino acids, and calcium. This sweat causes perspiration (Groscurth P. Anatomy of sweat glands. Curr Probl Dermatol. 2002;30: 1-9. Doi:10.1159 / 000060678. PMID: 12471693). Apocrine glands open into the very upper hair follicle instead of the skin surface and produce apocrine sweat, a thick, sterile, lipid-rich, odorless fluid containing proteins, sugars, ammonia, steroids, and notably, the chemical precursors for body odor (Baker LB. Physiology of sweat gland function: The roles of sweating and sweat composition in human health. Temperature (Austin). 2019 Jul 17;6(3):211-259. doi: 10.1080 / 23328940.2019.1632145. PMID: 31608304; PMCID: PMC6773238; SanMiguel A, Grice EA. Interactions between host factors and the skin microbiome. Cell Mol Life Sci. 2015 Apr;72(8): 1499-515. doi: 10.1007 / s00018-014-1812-z. Epub 2014 Dec 30. PMID: 25548803; PMCID: PMC4376244). Under emotional stress, hormones trigger apocrine glands to release this fluid, resulting in the formation of strong body odor (Barzantny H, Brune I, Tauch A. Molecular basis of human body odour formation: insights deduced from corynebacterial genome sequences. Int J Cosmet Sci. 2012Atty. Dkt. No. 135523-0115Feb;34(l):2-11. doi: 10.1111 / j.1468-2494.2011.00669.x. Epub 2011 Jul 25. PMID: 21790661).

[0077] Staphylococcus hominis (S. hominis) is a species of gram-positive bacteria known to reside on humans. It is typically considered a non-pathogenic commensal bacterium that produces thioalcohol compounds, thus contributing to malodor in humans. S. hominis is most frequently observed in areas of the skin with high numbers of apocrine glands, such as the axilla (underarm). Wild-type S. hominis expresses the gene, thereby producing the PatB enzyme which allows the bacteria to metabolize Cys-Gly-3M3SH into 3M3SH. Apocrine sweat is typically rich in Cys-Gly-3M3SH, thereby providing a nutrient rich environment for patB expressing S. hominis and allowing for microbiome colonization of specific areas of skin with high numbers of apocrine glands. The patB gene is required for the cleavage of an amino acid from the precursor Cys-Gly-3M3SH. Amino acids are often a limiting nutrient in microbial communities, and patB expression thus creates a unique metabolic niche on the human axilla which has allowed S. hominis to dominate this microbiome site across humans. Given the essentiality of this gene to the metabolic niche of S. hominis, it is surprising that patB loss-of-function would not preclude S. hominis from surviving on the axilla, let alone persist over time.III. ApatB -Staphylococcus hominis

[0078] The technology of the present disclosure relates to the use of ApatB -Staphylococcus hominis (whose genome comprises a disruption of a patB gene), such as strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377 or variant strains thereof, to treat or prevent body odor, and methods of making the same. In some embodiments, the wild-type S. hominis strain is any one of Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210. In some embodiments, the ApatB -Staphylococcus hominis strain is any one of Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377. In some embodiments, the wild-type S. hominis strain is Sn00016 and the ApatB-Staphylococcus hominis strain is Sn00037. In some embodiments, the wild-type S. hominis strain is Sn00040 and the ApatB-Staphylococcus hominis strain is Sn00048. In some embodiments, the wild-type S. hominis strain is Sn00042Atty. Dkt. No. 135523-0115 and the ApatB-Staphylococcus hominis strain is Sn00079. In some embodiments, the wildtype S. hominis strain is Sn00015 and the ApatB-Staphylococcus hominis strain is Sn00082. In some embodiments, the wild-type S. hominis strain is Sn00085 and AIQ ApatB- Staphylococcus hominis strain is Sn00086. In some embodiments, the wild-type S. hominis strain is Sn00201 and the ApatB-Staphylococcus hominis strain is Sn00325. In some embodiments, the wild-type S. hominis strain is Sn00202 and the ApatB-Staphylococcus hominis strain is Sn00327. In some embodiments, the wild-type S. hominis strain is Sn00203 and the ApatB-Staphylococcus hominis strain is Sn00330. In some embodiments, the wildtype S. hominis strain is Sn00213 and the ApatB-Staphylococcus hominis strain is Sn00337. In some embodiments, the wild-type S. hominis strain is Sn00215 and WIQ ApatB- Staphylococcus hominis strain is Sn00339. In some embodiments, the wild-type S. hominis strain is Sn00239 and the ApatB-Staphylococcus hominis strain is Sn00343. In some embodiments, the wild-type S. hominis strain is Sn00210 and the ApatB-Staphylococcus hominis strain is Sn00377. In some embodiments, the ApatB-Staphylococcus hominis mutant comprises a TAA or a TGA mutant at the W190* position, wherein the mutation encodes a premature stop codon.

[0079] Methods for disruption of the patB gene include targeted approaches for introducing a disruption by, e.g., homologous recombination, site-directed mutagenesis, a Transcription activator-like effector nuclease (TALEN), a meganuclease, a zinc finger nuclease, a CRISPR / Cas9 system, a CRISPR base editing system, a CRISPR prime editing system, a CRISPR / Cpfl system, or a CRISPR / Csml system. Additionally or alternatively, such methods may also include random mutagenesis methods for introducing a disruption by, e.g., irradiation, chemical mutagens, passing cloned genes through mutator strains, “error-prone” PCR mutagenesis, rolling circle error-prone PCR, or saturation mutagenesis. For illustrative purposes, and for proof of concept, a patB disruption in Staphylococcus hominis (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377) was created via CRISPR base editing system as described in the Examples section. Staphylococcus hominis strain Sn00037 refers to a bacterial strain, or variant thereof, having been deposited under ID AC Accession No. 030924-01, Staphylococcus hominis strain Sn00048 refers to a bacterial strain, or variant thereof, having been deposited under ID AC Accession No. 030924-02, Staphylococcus hominis StrainAtty. Dkt. No. 135523-0115Sn00079 refers to a bacterial strain, or variant thereof, having been deposited under ID AC Accession No. 300725-03, Staphylococcus hominis Strain Sn00082 refers to a bacterial strain, or variant thereof, having been deposited under ID AC Accession No. 300725-04, Staphylococcus hominis Strain Sn00086 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-06, Staphylococcus hominis Strain Sn00325 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-14, Staphylococcus hominis Strain Sn00327 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-15, Staphylococcus hominis Strain Sn00330 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-16, Staphylococcus hominis Strain Sn00337 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-17, Staphylococcus hominis Strain Sn00339 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-18, Staphylococcus hominis Strain Sn00343 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-19, Staphylococcus hominis Strain Sn00377 refers to a bacterial strain, or variant thereof, having been deposited under IDAC Accession No. 300725-20, or compositions comprising the strain.

[0080] In some embodiments, bacterial strains, such as the genetically engineered Staphylococcus hominis strains e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377) of the present technology are used in methods, compositions, and pharmaceutical compositions for treating or preventing body odor (malodor). In some embodiments, the composition or pharmaceutical composition comprises the genetically engineered Staphylococcus hominis strains and a probiotic for preventing or controlling body odor. In some embodiments, compositions or pharmaceutical compositions of the present technology comprise vegetative bacterial cells. In some embodiments, the compositions or pharmaceutical compositions of the present technology comprise desiccated bacterial cells. In some embodiments, the compositions or pharmaceutical compositions of the present technology comprise freeze- dried bacterial cells. In some embodiments, the compositions or pharmaceutical compositions of the present technology comprise spray-dried bacterial cells. In some embodiments, the ApatB-S. hominis strains (e.g., strains Sn00037, Sn00048, Sn00079,Atty. Dkt. No. 135523-0115Sn00082, Sn00086, Sn00325, Sn00327, SnOO33O, SnOO337, SnOO339, Sn00343, or Sn00377) of the present technology do not grow significantly slower than the corresponding wild-type S. hominis strains (e.g., strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210). In some embodiments, the disruption of patB does not significantly negatively affect the fitness of the S. hominis strains whose genome comprises the disruption. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 50% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 55% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 60% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 65% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 70% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 75% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 80% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 85% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 90% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 95% when contacted with a thiol substrate compared to aAtty. Dkt. No. 135523-0115 corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decreases the amount of thiol production by at least about 96%, 97%, 98%, or 99% when contacted with a thiol substrate compared to a corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains as disclosed herein have a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decrease the amount of 3M3SH production by at least about 50% when contacted with apocrine sweat or a 3M3SH precursor as compared to corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decrease the amount of 3M3SH production by at least about 60% when contacted with apocrine sweat or a 3M3SH precursor as compared to corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decrease the amount of 3M3SH production by at least about 70% when contacted with apocrine sweat or a 3M3SH precursor as compared to corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decrease the amount of 3M3SH production by at least about 80% when contacted with apocrine sweat or a 3M3SH precursor as compared to corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decrease the amount of 3M3SH production by at least about 90% when contacted with apocrine sweat or a 3M3SH precursor as compared to corresponding wild-type strains. In some embodiments, the genetically engineered Staphylococcus hominis strains decrease the amount of 3M3SH production by at least about 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% when contacted with apocrine sweat or a 3M3SH precursor as compared to corresponding wild-type strains.

[0081] In some embodiments, bacterial strains, such as the genetically engineered Staphylococcus hominis strains (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377) of the present technology are used in methods, compositions, and pharmaceutical compositions for altering a microbiota. In some embodiments, the bacterial strains comprise a probiotic that is capable of engraftment to an existing microbiota, such that the bacterial strains persist for aAtty. Dkt. No. 135523-0115 period of time in the microbiota community. In some embodiments, the microbiota is a skin microbiota. In some embodiments, an agent is applied to disrupt the native microbiota, and then the bacterial strains and compositions of the present technology are applied, such that the disruption of the native microbiota increases engraftment of the bacterial strains.

[0082] In some embodiments, the genetically engineered Staphylococcus hominis strain whose genome comprises a disruption of a patB gene has a 16S rRNA gene sequence that is at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to any one of SEQ ID NOs: 26-37. In some embodiments, the bacterial strain for use in the present technology has the 16S rRNA gene sequence represented by any one of SEQ ID NOs: 26-37.

[0083] In some embodiments, the strain for use in the present technology is derived from a strain deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-03, 030924-04, 300725-02, 300725-01, 300725-05, 300725-07, 300725-08, 300725-09, 300725-11, 300725-12, 300725-13, or 300725-10, wherein the genome has been edited to comprise a disruption of a patB gene. In some embodiments, the strain for use in the present technology is derived from a strain that has a 16S rRNA gene sequence that is at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to any one of SEQ ID NOs: 26-37, wherein the genome has been engineered to comprise a disruption of a patB gene. Within a single genome, there are often multiple copies of 16S that are not always identical. SEQ ID NOs: 26-37 are consensus sequences of all of the 16S sequences into a single sequence. Sequences below are provided using standard IUPAC nomenclature, wherein A indicates Adenine, C indicates Cytosine, G indicates Guanine, T (or U) indicates Thymine (or Uracil), R indicates A or G, Y indicates C or T, S indicates G or C, W indicates A or T, K indicates G or T, M indicates A or C, B indicates C or G or T, D indicates A or G or T, H indicates A or C or T, V indicates A or C or G, N indicates any base, and . or - indicates a gap-Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115

[0084] In addition to strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377, other strains of Staphylococcus hominis that have been genetically engineered to comprise a disruption of a patB gene are also expected to be effective for reducing 3-methyl-3-sulfanylhexan-l-ol (3M3SH) production from a skin microbiota and / or for treating, reducing, or preventing malodor on the skin of a subject. In particular, strains that are biotypes of strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377 are also expected to be effective for reducing 3-methyl-3- sulfanylhexan-l-ol (3M3SH) production from a skin microbiota and / or for treating, reducing, or preventing malodor on the skin of a subject. As used herein, a “biotype” refers to a strain that is a closely related strain that has the same or very similar physiological and biochemical characteristics.

[0085] Similarly, in addition to strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210, other strains of Staphylococcus hominis are also expected to be suitable for genetically engineering disruption of a patB gene in order to generate a strain effective for reducing 3-methyl-3- sulfanylhexan-l-ol (3M3SH) production from a skin microbiota and / or treating, reducing, orAtty. Dkt. No. 135523-0115 preventing malodor on the skin of a subject. In particular, strains that are biotypes of strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210 are expected to be suitable for engineering to disrupt a patB gene.

[0086] Strains that are biotypes of strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377 and other Staphylococcus hominis strains that are suitable for use in the present technology may be identified by sequencing other nucleotide sequences for strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377. Similarly, strains that are biotypes of strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210 and other Staphylococcus hominis strains that are suitable for genetically engineering to disrupt a patB gene to provide a strain of the present technology may be identified by sequencing other nucleotide sequences for strains.

[0087] For example, substantially the whole genome may be sequenced and a strain for use in the present technology or for use in generating a strain for use in the present technology may have at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% sequence identity across at least 80% of its whole genome (e.g., across at least 85%, 90%, 95% or 99%, or across its whole genome). Other suitable sequences for use in identifying suitable strains may include hsp60 or repetitive sequences such as BOX, ERIC, (GTG)s or REP (Masco et al. (2003) Systematic and Applied Microbiology, 26:557-563). Suitable strains, such as biotype strains, may have sequences with at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% sequence identity to the corresponding sequence of the strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, Sn00210, Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377.

[0088] Alternatively, strains that are biotypes of strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377 and other Staphylococcus hominis strains that are suitable for use in the present technology, or strains that are biotypes of strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085,Atty. Dkt. No. 135523-0115Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210 and other Staphylococcus hominis strains that are suitable for genetically engineering to disrupt a patB gene to provide a strain of the present technology may be identified by using strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, Sn00210, Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377 and restriction fragment analysis and / or PCR analysis, for example by using fluorescent amplified fragment length polymorphism (FAFLP) and repetitive DNA element (rep)-PCR fingerprinting, or protein profiling, or partial 16S or 23 S rDNA sequencing. In some embodiments, such techniques may be used to identify other Staphylococcus hominis strains for use in the present technology, following genetic engineering to disrupt patB.

[0089] In certain embodiments, strains that are biotypes of strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377 and other Staphylococcus hominis strains that are suitable for use in the present technology, or strains that are biotypes of strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210 and other Staphylococcus hominis strains that are suitable for genetically engineering to disrupt a patB gene to provide a strain of the present technology, may be strains that provide the same pattern as strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, Sn00210, Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377 when analyzed by amplified ribosomal DNA restriction analysis (ARDRA), for example when using Sau3 Al restriction enzyme (for exemplary methods and guidance see, for example, Sriitkova et al. (2011) J Microbiol Methods, 87(1): 10-6). Alternatively, biotype strains are identified as strains that have the same carbohydrate fermentation patterns as strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, Sn00210, Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377.

[0090] Bacterial strains that have similar growth patterns, metabolic type and / or surface antigens to strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327,Atty. Dkt. No. 135523-0115SnOO33O, SnOO337, SnOO339, Sn00343, or Sn00377 may be useful in the present technology. A useful strain will have comparable PatB catalytic activity and / or 3M3SH production activity to strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377. In particular, a biotype strain will elicit comparable effects on the models shown in the Examples, which may be identified by using the culturing and assay protocols described in the Examples.

[0091] In another aspect, the present disclosure provides a Staphylococcus hominis strain, wherein the strain comprises a mutation in at least one endogenous polynucleotide having about 80% to about 100% sequence identity to SEQ ID NO: 14, such that the strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding strain that does not comprise the mutation. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 80% to about 100% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 85% to about 100% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 90% to about 95% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 95% to about 99% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises the mutation in at least one endogenous polynucleotide having about 97% to about 100% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 80% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 85% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 86% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 87% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 88% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotideAtty. Dkt. No. 135523-0115 having about 89% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 90% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 91% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 92% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 93% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 94% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 95% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 96% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 97% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 98% sequence identity to SEQ ID NO: 14. In some embodiments, the strain comprises a mutation in at least one endogenous polynucleotide having about 99% sequence identity to SEQ ID NO: 14. In some embodiments, the strain grows at an equivalent or improved rate as compared to a Staphylococcus hominis strain lacking the mutation. In some embodiments, the strain is incapable of converting Cys-Gly-3M3SH to 3M3SH. In some embodiments, the mutation comprises a nucleotide substitution. In some embodiments, the nucleotide substitution results in a premature stop codon in the endogenous polynucleotide having about 80% to about 100% sequence identity to the nucleotide sequence as set forth in SEQ ID NO: 14. In some embodiments, the premature stop codon is introduced at the codon that, in the wild-type sequence, encodes for tryptophan at position 190 of the PatB amino acid sequence (W190), which is set forth in SEQ ID NO: 41, such that the mutation introducing the stop codon at W190 results in the formation of a truncated PatB protein having, such as, for example, the amino acid sequence as set forth in SEQ ID NO: 42. It is to be understood that the nucleic acid sequences provided in SEQ ID NOs: 14 and 15 and the PatB amino acid sequences provided in SEQ ID NOs: 41 and 42 are illustrative, non-limiting Staphylococcus hominisAtty. Dkt. No. 135523-0115 patB gene and PatB polypeptide sequences, and one of skill in the art would understand that there is a certain degree of nucleotide and amino acid diversity across strains of the same species that is encompassed by the disclosure of the present technology.

[0092] The disclosure of the present technology encompasses genetic engineering of any Staphylococcus hominis nucleotide sequence encoding a PatB polypeptide that is at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.9%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 41, to introduce a disrupt! on / mutati on to the patB gene such that the strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding strain that does not comprise the disrupt! on / mutati on. Thus, in some embodiments, the technology of the present disclosure provides a Staphylococcus hominis strain, wherein the strain comprises a mutation in at least one endogenous nucleotide sequence selected from the group consisting of: (a) a nucleotide sequence having about 80% to about 100% sequence identity to SEQ ID NO: 14; and (b) a nucleotide sequence encoding a polypeptide comprising an amino acid sequence having about 85% to about 100% sequence identity to SEQ ID NO: 41, such that the strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding strain that does not comprise the mutation. The mutation may include any of those described herein, including, but not limited to one or more nucleotide substitutions that introduce a premature stop codon into the patB gene sequence. In some embodiments, the wildtype allele of the nucleotide sequence comprising the mutation encodes a PatB polypeptide that is at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.9%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 41.Genome Engineering and Other Methods Encompassed by the Present Disclosure

[0093] The present disclosure contemplates any and all methods for disrupting PatB metabolic enzyme activity and is not limited to any particular method. For example, the present disclosure contemplates any methods for achieving disruptions or mutations including insertions, deletions, substitutions (e.g., transitions, transversion, homologous recombination), transpositions, inversions, knockouts and combinations thereof in the patBAtty. Dkt. No. 135523-0115 gene, in any regulatory elements of the patB gene, or in any other gene or region of interest to achieve the desired result. For example, in some embodiments, a disruption or mutation may include: (a) the introduction, substitution, or removal of one or more nucleotides in a Staphylococcus hominis patB gene (or an open reading frame (ORF) thereof), or the introduction, substitution, or removal of one or more nucleotides in a regulatory element required for the transcription or translation of the patB gene or ORF thereof; (b) a patB gene conversion; (c) a patB gene deletion; (d) the down-regulation of the patB gene; (e) specific mutagenesis of the patB gene; and / or (f) random mutagenesis of the patB gene. Disruptions or mutations of the patB gene, as used herein, include any genetic modification that prevents or reduces the capability of the S. hominis strains of the present technology from producing a functional patB gene product. Such disruptions or mutations may include complete or partial deletion of any portion of patB, including a polypeptide-coding sequence, a promoter, an enhancer, a ribosome-binding site (RBS), or other regulatory elements, or mutagenesis of the same, where mutagenesis encompasses substitutions, insertions, deletions, inversions, and any combinations and variations thereof that inactivate patB and prevent or reduce the production of a patB functional gene product. Mutations or disruptions may involve only a single nucleotide (e.g., a point mutation, nucleotide substitution, or a single nucleotide polymorphism) or multiple nucleotides. In some embodiments, the mutation or disruption may be a cytosine (C) to thymine (T) substitution. In some embodiments, the conversion may occur by a cytosine-based editor (CBE) tool. In some embodiments, the CBE precisely converts a cytosine (C) to thymine (T) without causing a double-stand break. In some embodiments, the cytosine (C) to thymine (T) occurs at the W190 tryptophan amino acid site in patB. In some embodiments, the cytosine (C) to thymine (T) substitution introduces a premature stop codon in the expressed mRNA (z.e., UAG, UGA, or UAA). In some embodiments, the premature stop codon introduces an early termination signal in the mRNA sequence. In some embodiments, the early termination signal leads to production of a truncated protein. In some embodiments, the truncated protein has a loss of catalytic activity. In some embodiments, the mutation or disruption causes a phenotypic change, for example, the expression level of the encoded product is altered, or the encoded product itself is altered. In some embodiments, a disruption or mutation may result in a disrupted gene with decreased levels of expression of a gene product (e.g., protein or RNA) as compared to the wild-typeAtty. Dkt. No. 135523-0115 strain. In other embodiments, a disruption or mutation may result in an expressed protein with activity that is lower as compared to the activity of the expressed protein from the wildtype strain. In some embodiments, a gene disruption or mutation results in the removal of the gene from the genome via a substitution, such as homologous recombination.

[0094] In some embodiments, a disruption or mutation of the patB gene, as used herein, includes any genetic modification that prevents or reduces the capability of the S. hominis strains of the present technology from converting Cys-Gly-3M3SH to 3M3SH. This includes, but is not limited to, disruption of one or more PatB protein enzymatic activities or alteration of PatB protein binding with one or more metabolites or co-factors. The skilled artisan would understand that any appropriate method may be used to prevent or reduce the capability of the S. hominis strains of the present technology from converting Cys-Gly- 3M3SH to 3M3SH.

[0095] Disruptions or mutations outside of the patB coding region that accomplish the effect of disrupted PatB activity are also encompassed within this disclosure, including disruptions or mutations in enhancer, repressor, promoter, or other regulatory elements that result in decreased transcription or translation of the patB gene. Other methods may also be used to disrupt PatB activity, including, but not limited to engineered transcription factors, including Transcription Activator-Like Effectors (TALEs) that repress patB expression, antisense RNAs or small regulatory RNAs that repress patB translation, CRISPR-dCasl3 constructs that target patB RNA and prevent translation, and protein-based or small molecule constructs that post translationally inactivate or repress PatB protein activity. For example, the present disclosure explicitly contemplates constructs (e.g., recombinant proteins) that post- translationally modify the PatB protein (e.g., phosphorylate) to reduce or inhibit enzymatic activity or metabolite binding by the PatB protein. Any construct, and method of use thereof, that downregulates, prevents, or inhibits in any way the transcription, translation, or protein activity of patB is encompassed within the present disclosure.

[0096] Provided herein are methods and compositions for modifying a target genomic locus in a cell to modulate the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein. Targeted genome engineering techniques described herein include the CRISPR (clustered regularly interspaced short palindromic repeats) / CasAtty. Dkt. No. 135523-0115(CRISPR-associated) system, meganucleases, zinc finger nucleases (ZFNs), and TAL effector nucleases (TALENs). Such techniques may be employed to bind to and / or cleave a genomic region of interest. In some embodiments, the genome editing techniques described herein generate a specific sequence change or gene edit (e.g., insertion, deletion, or substitution) in the 5’-UTR of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of patB protein, such as generating a single nucleotide gene edit to form an out-of-frame start codon upstream of the gene’s ORF, thereby suppressing expression of the gene. In some embodiments, the gene edit (e.g., deletion, insertion, or substitution) results in production of an upstream, out-of- frame start codon that may result in the elimination of protein production or a nonfunctional protein. In some embodiments, the genome editing techniques described herein generate a specific sequence change or gene edit (e.g., insertion, deletion, or substitution) in the coding region or a non-coding region of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, such as generating a large deletion to form (1) an out-of-frame start codon upstream of the gene’s ORF, thereby suppressing expression of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, or (2) a non-functional protein product resulting from a frame shift downstream of the gene edit. In some embodiments, the large deletion is greater than 50 bases, greater than 100 bases, greater than 200 bases, greater than 500 bases, greater than 1000 bases, greater than 2000 bases, greater than 5000 bases, or greater than 10000 bases. In some embodiments, the large deletion is generated in the patB gene. In some embodiments, a large deletion is a complete deletion of a gene or genetic element. For example, in some embodiments, an ORF is deleted.CRISPR / Cas Systems

[0097] In some embodiments, the methods of the present technology relate to the use of a CRISPR / Cas system that binds to a target site in a region of interest in a genome, wherein the CRISPR / Cas system comprises a CRISPR / Cas nuclease and an engineered crRNA / tracrRNA (or single guide RNA (sgRNA) or guide RNA (gRNA)). In some embodiments, the CRISPRAtty. Dkt. No. 135523-0115 system generally comprises (i) a polynucleotide encoding a Cas protein, and (ii) at least one sgRNA for RNA-guided genome engineering in S. hominis cells.

[0098] Non-limiting examples of Cas proteins include Casl, CaslB, Cas2, Cas3, Cas4, Cas5, Cas6, Cas7, Cas8, Cas9 (also known as Csnl and Csxl2), CaslO, Casl2a (also known as Cpfl), Csyl, Csy2, Cys3, Csel, Cse2, Cscl, Csc2, Csa5, Csn2, Csml, Csm2, Csm3, Csm4, Csm5, Csm6, Smrl, Cmr3, Cmr4, Cmr6, Csbl, Csb2, Csb3, Csxl7, Csxl4, CsxlO, Csxl6, CsaX, Csx3, Csxl, Csxl5, Csfl, Csf2, Csf3, Csf4, homologs thereof, or modified versions thereof. In some embodiments, the Cas protein is a Streptococcus pyogenes Cas9 protein. In some embodiments, the Cas protein is a Casl2a (Cpfl) protein. In some embodiments, the Cas protein is a Csml protein. These enzymes are known. For example, the amino acid sequence of S. pyogenes Cas9 protein may be found in the SwissProt database under accession number Q99ZW2. The amino acid sequence of Francisella tularensis subsp. Novicida Cpfl protein may be found in the UniProt database under accession number A0Q7Q2. The amino acid sequence of Thermococcus onnurineus Csml protein may be found in the UniProt database under accession number B6YWB8.

[0099] The sgRNA molecules comprise a crRNA-tacrRNA scaffold polynucleotide and a targeting sequence corresponding to a genomic target of interest. In some embodiments, the CRISPR / Cas system recognizes a target site in the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein. In some embodiments, the CRISPR / Cas system recognizes a target in the patB gene or a regulatory region thereof. The CRISPR / Cas system as described herein may bind to and / or cleave the region of interest in a region upstream of the coding region of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein. In some embodiments, the CRISPR / Cas system generates a specific sequence change in the 5’-UTR of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, such as generating a single nucleotide gene edit to form an out-of-frame start codon upstream of the gene’s ORF. In some embodiments, the gene edit (e.g., deletion, insertion, or substitution) results in production of an upstream, out-of-frame start codon that may result in the elimination of protein production or a nonfunctional protein. In some embodiments, theAtty. Dkt. No. 135523-0115CRISPR / Cas system generates a specific sequence change or gene edit (e.g., insertion, deletion, or substitution) in the coding region or a non-coding region of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, such as generating a large deletion to form (1) an out-of- frame start codon upstream of the gene’s ORF, thereby suppressing expression of the gene, or (2) a non-functional protein product resulting from a frame shift downstream of the gene edit. In some embodiments, the large deletion is greater than 50 bases, greater than 100 bases, greater than 200 bases, greater than 500 bases, greater than 1000 bases, greater than 2000 bases, greater than 5000 bases, or greater than 10000 bases.

[0100] The CRISPR / Cas system can be based on the Cas9 nuclease and an engineered single guide RNA (sgRNA) that specifies the targeted nucleic acid sequence. Cas9 is a large monomeric DNA nuclease guided to a DNA target sequence adjacent to the PAM (protospacer adjacent motif) sequence motif by a complex of two non-coding RNAs: CRISPR RNA (crRNA) and trans-activating crRNA (tacrRNA). The Cas9 protein contains two nuclease domains homologous to RuvC and HNH nucleases. The HNH nuclease domain cleaves the complementary DNA strand whereas the RuvC-like domain cleaves the non- complementary strand and, as a result, a blunt cut is introduced in the target DNA.Heterologous expression of Cas9 together with an sgRNA can induce site-specific double strand breaks (DSBs) into genomic DNA of live cells. See, e.g., Mussolino, Nat.Biothechnol., 31:208-209 (2013). In some embodiments, the Cas9 protein is expressed in a S. hominis cell as a fusion protein. In some embodiments, the Cas9 protein is tagged (e.g., FLAG- or GFP-tagged). In some embodiments, promoters (e.g., PXyl tet, pCap, BBa_J23100-BBa_B0030, pHla, pBla, or pHelp) may be used to drive Cas9 expression in a S. hominis cell. In some embodiments, the Cas9 enzyme is S. pneumoniae, S. pyogenes, or S. thermophiles Cas9, and may include mutated Cas9 derived from these organisms. The enzyme may be a Cas9 homolog or ortholog. In some embodiments, the CRISPR enzyme e.g., Cas9 enzyme) is codon-optimized for expression in a S. hominis cell.

[0101] The CRISPR / Cas system can be based on the Cpfl nuclease and an engineered single guide RNA (sgRNA) that specifies the targeted nucleic acid sequence.Atty. Dkt. No. 135523-0115

[0102] Cpfl is distinguished from Cas9 by a its single RuvC endonuclease active site, its 5' protospacer adjacent motif preference, and for creating sticky rather than blunt ends at the cut site. The Cpfl protein has a RuvC-like endonuclease domain that is similar to the RuvC domain of Cas9. Cpfl does not have a HNH endonuclease domain, and the N-terminal of Cpfl does not have an alpha-helical recognition lobe, unlike Cas9. In some embodiments, the Cpfl protein is tagged (e.g., FLAG- or GFP-tagged). In some embodiments, promoters (e.g., PXyl tet, pCap, BBa_J23100-BBa_B0030, pHla, pBla, or pHelp) may be used to drive Cpfl expression in a S. hominis cell. In some embodiments, the Cpfl enzyme is Francisella tularensis subsp. Novicida Cpfl, and may include mutated Cpfl derived from these organisms. The enzyme may be a Cpfl homolog or ortholog. In some embodiments, the CRISPR enzyme (e.g., Cpfl enzyme) is codon-optimized for expression in a S. hominis cell.

[0103] The CRISPR / Cas system can be based on the Csml nuclease and an engineered single guide RNA (sgRNA) that specifies the targeted nucleic acid sequence.

[0104] Csml belongs to the CaslO family of endonucleases. Csml is the largest subunit of the Csm interference complex in the type III-A CRISPR system. Csml exhibits ssDNA- specific endo- and exonuclease activity. In some embodiments, promoters (e.g., PXyl tet, pCap, BBa_J23100-BBa_B0030, pHla, pBla, or pHelp) may be used to drive Csml expression in a S. hominis cell. In some embodiments, the Csml enzyme is Thermococcus onnurineus Csml, and may include mutated Csml derived from these organisms. The enzyme may be a Csml homolog or ortholog. In some embodiments, the CRISPR enzyme (e.g., Csml enzyme) is codon-optimized for expression in a S. hominis cell.

[0105] The single guide RNA (sgRNA) is the second component of the CRISPR / Cas system that forms a complex with a Cas nuclease. The sgRNA is created by fusing crRNA with tacrRNA. The sgRNA guide sequence located at the 5’ end confers DNA target specificity. By modifying the guide sequence, sgRNAs with different target specificities can be designed to target any desired endogenous gene. In some embodiments, the target sequence is about 1,000, about 975, about 950, about 925, about 900, about 875, about 850, about 825, about 800, about 775, about 750, about 725, about 700, about 675, about 650, about 625, about 600, about 575, about 550, about 525, about 500, about 475, about 450, about 425, about 400, about 375, about 350, about 325, about 300, about 275, about 250, about 225, about 200,Atty. Dkt. No. 135523-0115 about 175, about 150, about 125, about 100, about 90, about 80, about 70, about 60, about 50, about 40, about 30, about 20, or about 15 base pairs upstream of the transcription start site, or the target sequence may be any number of base pairs in-between these values upstream of the transcription start site. In some embodiments, the target sequence is about 1 to about 10 base pairs upstream of the transcription start site (e.g., positions -10, -9, -8, -7, -6, -5, -4, -3, -2, or -1). In some embodiments, the target sequence is located within the open reading frame of the gene of interest. In some embodiments, the target sequence is located within a coding region of the gene of interest. In some embodiments, the CRISPR / Cas system comprises at least one sgRNA. In some embodiments, a target sequence of the at least one sgRNA is about 1,000, about 975, about 950, about 925, about 900, about 875, about 850, about 825, about 800, about 775, about 750, about 725, about 700, about 675, about 650, about 625, about 600, about 575, about 550, about 525, about 500, about 475, about 450, about 425, about 400, about 375, about 350, about 325, about 300, about 275, about 250, about 225, about 200, about 175, about 150, about 125, about 100, about 90, about 80, about 70, about60, about 50, about 40, about 30, about 20, or about 15 base pairs upstream of the transcription start site, or the target sequence may be any number of base pairs in-between these values upstream of the transcription start site. In some embodiments, the target sequence of the at least one sgRNA is about 1 to about 10 base pairs upstream of the transcription start site (e.g., positions -10, -9, -8, -7, -6, -5, -4, -3, -2, or -1). In some embodiments, the target sequence of the at least one sgRNA is located within the open reading frame of the gene of interest. In some embodiments, the target sequence of the at least one sgRNA is located within a coding region of the gene of interest. In some embodiments, the target sequences of the at least one sgRNA is located within the open reading frame of the gene of interest. In some embodiments, the target sequences of the at least one sgRNA is located within a coding region of the gene of interest. In some embodiments, the CRISPR / Cas system comprises one sgRNA, wherein the one sgRNA targets two regions of a gene having the same sequence, such as two inverted terminal repeats (ITRs). In some embodiments, the target sequences of the sgRNA are separated by at least 50 bases, at least 100 bases, at least 200 bases, at least 500 bases, at least 1000 bases, at least 2000 bases, at least 5000 bases, or at least 10000 bases.Atty. Dkt. No. 135523-0115

[0106] In some embodiments, the CRISPR / Cas system comprises at least two sgRNAs. In some embodiments, a target sequence of at least one of the at least two sgRNAs is about 1,000, about 975, about 950, about 925, about 900, about 875, about 850, about 825, about 800, about 775, about 750, about 725, about 700, about 675, about 650, about 625, about 600, about 575, about 550, about 525, about 500, about 475, about 450, about 425, about 400, about 375, about 350, about 325, about 300, about 275, about 250, about 225, about 200, about 175, about 150, about 125, about 100, about 90, about 80, about 70, about 60, about 50, about 40, about 30, about 20, or about 15 base pairs upstream of the transcription start site, or the target sequence may be any number of base pairs in-between these values upstream of the transcription start site. In some embodiments, the target sequence of at least one of the at least two sgRNAs is about 1 to about 10 base pairs upstream of the transcription start site (e.g., positions -10, -9, -8, -7, -6, -5, -4, -3, -2, or -1). In some embodiments, the target sequence of at least one of the at least two sgRNAs is located within the open reading frame of the gene of interest. In some embodiments, the target sequence of at least one of the at least two sgRNAs is located within a coding region of the gene of interest. In some embodiments, the target sequences of at least two of the at least two sgRNAs are located within the open reading frame of the gene of interest. In some embodiments, the target sequences of at least two of the at least two sgRNAs are located within a coding region of the gene of interest. In some embodiments, the CRISPR / Cas system comprises two sgRNAs, wherein the two sgRNAs have non-overlapping target sequences. In some embodiments, the target sequences of the two sgRNAs are separated by at least 50 bases, at least 100 bases, at least 200 bases, at least 500 bases, at least 1000 bases, at least 2000 bases, at least 5000 bases, or at least 10000 bases.

[0107] It is not intended that the present technology be limited to any particular distance restraint with regard to the location of the guide RNA target sequence from the gene transcription start site. In some embodiments, the target sequence lies “in proximity to” a gene of interest, where “in proximity to” refers to any distance from the gene of interest, wherein the Cas-regulatory domain fusion is able to exert an effect on gene expression. In some embodiments, the target sequence lies upstream of the ORF of the gene of interest.Atty. Dkt. No. 135523-0115

[0108] The canonical length of the guide sequence is about 20 bp and the DNA target sequence is about 20 bp followed by a PAM sequence having the consensus NGG sequence. In some embodiments, sgRNAs are expressed in a S. hominis cell using S. hominis promoters.

[0109] When the DSBs are repaired by either non-homologous end joining (NHEJ) or homology directed repair (HDR), the sequence at the repair site can be modified or new genetic information can be inserted (e.g., donor DNA comprising a desired gene edit can be inserted into the target gene at the break site). Although HDR typically occurs at lower and more variable frequencies than NHEJ, it can be leveraged to generate precise, defined modifications at a target locus in the presence of an exogenously introduced repair template. Accordingly, exogenous repair templates, designed by methods known in the art, can also be delivered into a cell, most often in the form of a synthetic, single-stranded DNA donor oligo or DNA donor plasmid, to generate a precise change in the genome. Single-stranded DNA donor oligos are delivered into a cell to insert or change short sequences (SNPs, amino acid substitutions, epitope tags, etc.) of DNA in the endogenous genomic target region. The benefits of using a synthetic DNA donor oligo are that no cloning is required to generate the donor template and DNA modifications can be added during synthesis for different applications, such as increased resistance to nucleases. Traditionally, the maximum insert length recommended for use with a DNA donor oligo is about 50 nucleotides.

[0110] In some embodiments, the present technology provides an engineered, programmable, non-naturally occurring CRISPR / Cas system comprising a Cas9 protein and one or more single guide RNAs (sgRNAs) that target the genomic loci of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, and the Cas9 protein cleaves the genomic loci of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, whereby expression of the one or more gene products is altered. In some embodiments, Cas9 introduces multiple DSBs in the same cell (ie., multiplexes) via expression of one or more distinct guide RNAs.[OHl] In some embodiments, the present technology provides a method for targeted genomic modification of S. hominis cells to alter the expression of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity ofAtty. Dkt. No. 135523-0115PatB protein, the method comprising introducing into a S. hominis cell containing and expressing a DNA molecule having a target sequence and encoding the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein an engineered CRISPR / Cas system comprising (a) an expression construct comprising a first polynucleotide encoding a Cas9 protein, or a variant thereof or a fusion protein therewith, and a second polynucleotide encoding a guide RNA comprising: (i) a crRNA-tracrRNA scaffold polynucleotide, and (ii) a targeting sequence operably linked to the crRNA-tracrRNA scaffold polynucleotide, where the targeting sequence corresponds to a genomic locus of interest, and (b) delivering the expression construct into the S. hominis cell, where the first and second polynucleotides are expressed (transcribed) within the S. hominis cell. This method can optionally further include visualizing, identifying, or selecting for S. hominis cells having a genomic modification at the genomic locus of interest that is induced by the delivering the expression construct into the S. hominis cells.

[0112] In some embodiments of the methods of the present technology, the Cas9 polypeptide and one or more guide RNA are encoded on a single vector. In some embodiments, the single vector is a plasmid. In some embodiments of the methods of the present technology, the Cas9 polypeptide and the one or more guide RNA are encoded on two separate vectors. In these methods, the steps generally follow the sequence of introducing into a S. hominis cell containing and expressing a DNA molecule having a target sequence and encoding the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein an engineered CRISPR / Cas system comprising (a) a Cas9 polynucleotide or a conservative variant thereof, and a guide RNA comprising (i) a crRNA-tracrRNA scaffold polynucleotide, and (ii) a targeting sequence operably linked to the crRNA-tracrRNA scaffold polynucleotide, with the targeting sequence corresponding to a genomic locus of interest, and (b) delivering the two polynucleotides into the S. hominis cell. In variations of this method, a donor polynucleotide having homology to the genomic target of interest is included in a co-transfection. In some variations of these methods, the transfected material can be either plasmid DNA or RNA generated by in vitro transcription. In still other variations, the methods for targeted genomic modification are multiplexed, meaning that more than one genomic locus is targeted for modification. In still other variations of these methods, the transformation of the S. hominis cells can be followed byAtty. Dkt. No. 135523-0115 visualizing, identifying, or selecting for S. hominis cells having a genomic modification at the genomic locus of interest.Meganucleases

[0113] In some embodiments, the compositions and methods described herein employ a meganuclease DNA binding domain for binding to a region of interest in the genome of a S. hominis cell. Meganucleases are engineered versions of naturally occurring restriction enzymes that typically have extended DNA recognition sequences (e.g., about 14 to about 40 base pairs in length). Meganucleases (also known as homing endonucleases) are commonly grouped into five families based on sequence and structure motifs: the LAGLID ADG family (“LAGLID ADG” is disclosed as SEQ ID NO: 43), the GIY-YIG family, the His-Cyst box family, the PD-(DZE)XK family, and the HNH family. In some embodiments, the meganuclease comprises an engineered homing endonuclease. The recognition sequences of homing endonucleases and meganucleases such as I-Sce, I-Ceul, PI-PspI, Pl-Sce, I-SceIV, I- Csml, I-PanI, I-A'ccII, I- ol, I-A'ccIII, I-Crel, LTevI, I-PevII, and LTevIII are known.

[0114] In some embodiments, the meganuclease is tailored to recognize a target in one or more of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein. The meganucleases as described herein may bind to and / or cleave the region of interest in a region upstream of the coding region of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein. Gene insertion or correction can be achieved by the introduction of a DNA repair matrix containing sequences homologous to the endogenous sequence surrounding the DNA break. Gene edits can be created either at or distal to the break. In some embodiments, the meganuclease generates a specific sequence change in the 5’-UTR of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, such as generating a single nucleotide gene edit to form an out-of-frame start codon upstream of the gene’s ORF.TALENs

[0115] In some embodiments, the compositions and methods described herein employ transcription activator-like effector nucleases (TALENs) to edit S. hominis genomes byAtty. Dkt. No. 135523-0115 inducing double-strand breaks (DSBs). TALENs are restriction enzymes that can be engineered to cleave specific sequences of DNA. TALENs are constructed by fusing a TAL effector DNA-binding domain to a DNA cleavage domain (e.g., a nuclease domain such as that derived from the FokI endonuclease). Transcription activator-like effectors (TALEs) can be engineered according to methods known in the art to bind to a desired DNA sequence, and when combined with a nuclease, provide a technique for cutting DNA at specific locations. For example, after a target sequence in the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein is identified, a corresponding TALEN sequence is engineered and inserted into a plasmid. The plasmid is inserted into a target cell where it is translated to produce a functional TALEN, which then binds to and cleaves its target sequence. Such an approach can be employed to introduce an exogenous DNA sequence into the target gene as the DSB is being repaired through either homology-directed repair or non-homologous end-joining. For example, in some embodiments, the use of TALEN technology generates a specific sequence change (e.g., insertion, deletion, or substitution) in the 5’-UTR of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, resulting in the production of an out-of-frame start codon upstream of the gene’s ORF.ZFNs

[0116] In some embodiments, the compositions and methods described herein employ zinc finger nucleases (ZFNs) to edit S. hominis genomes by inducing double-strand breaks (DSBs). ZFNs are artificial restriction enzymes generated by fusing a zinc finder DNA- binding domain to a DNA cleavage domain (e.g., a nuclease domain such as that derived from the FokI endonuclease). ZFNs can be engineered to bind and cleave DNA at specific locations. ZFNs contain two protein domains. The first domain is the DNA-binding domain, which contains eukaryotic transcription factors and the zinc finger. The second domain is a nuclease domain that contains the FokI restriction enzyme responsible for cleaving DNA. ZFNs can be engineered according to methods known in the art to bind to a desired DNA sequence and cleave DNA at specific locations. For example, after a target sequence in the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA,Atty. Dkt. No. 135523-0115 or the enzymatic activity of PatB protein is identified, a corresponding ZFN sequence is engineered and inserted into a plasmid. The plasmid is inserted into a target cell where it is translated to produce a functional ZFN, which then binds to and cleaves its target sequence introducing a double strand break (DSB). Such an approach can be employed to introduce an exogenous DNA sequence into the target gene as the DSB is being repaired through either homology-directed repair or non-homologous end-joining. For example, in some embodiments, the use of ZFN technology generates a specific sequence change in the 5’-UTR of the patB gene or a gene involved in the expression of the patB gene, the translation of patB RNA, or the enzymatic activity of PatB protein, such as the insertion of an out-of-frame start codon upstream of the gene’s ORF.Additional Methodologies for Suppressing patB activity

[0117] The expression and activity of patB may be reduced by introducing mutations / disruptions as described herein for patB, or may be reduced by other well-known means of reducing expression of polynucleotides and activity of polypeptides including but not limited to the use of interfering RNAs developed from nucleic acids encoding patB. As known in the art, “interfering RNA” is RNA capable of causing gene silencing. Interfering RNA (RNAi), as used herein, includes any type of RNA molecule capable of downregulating or silencing expression of a target patB nucleic acid, including but not limited to sense RNA, antisense RNA, short interfering RNA (siRNA), micro RNA (miRNA), doublestranded RNA (dsRNA), hairpin RNA, and the like. In some embodiments, methods and constructs are provided for suppressing patB metabolic enzyme activity, such as, but not limited to, antisense, sense co-suppression, RNAi, and artificial microRNA approaches.

[0118] RNAi techniques involve stable transformation using RNAi plasmid constructs (Helliwell and Waterhouse, Methods EnzymoL 392:24-35 (2005)). Such plasmids are composed of a fragment of the target gene to be silenced in an inverted repeat structure. The inverted repeats are separated by a spacer, often an intron. The RNAi construct may be driven by a suitable promoter, for example, the PXyl tet, pCap, BBa_J23100-BBa_B0030, pHla, pBla, or pHelp promoter, and integrated into the S. hominis genome and subsequent transcription of the transgene leads to an RNA molecule that folds back on itself to form a double-stranded hairpin RNA (hpRNA).Atty. Dkt. No. 135523-0115

[0119] Artificial microRNA (amiRNA) techniques exploit the microRNA (miRNA) pathway that functions to silence endogenous genes. In this method, 21 nucleotide long fragments of the gene to be silenced are introduced into a pre-miRNA gene to form a pre-amiRNA construct. The pre-miRNA construct is transferred into the genome using transformation methods apparent to one skilled in the art. After transcription of the pre-amiRNA, processing yields amiRNAs that target genes, which share nucleotide identity with the 21 nucleotide amiRNA sequence.

[0120] In RNAi silencing techniques, two factors can influence the choice of length of the fragment. The shorter the fragment the less frequently effective silencing will be achieved, but very long hairpins increase the chance of recombination in bacterial host strains. The effectiveness of silencing also appears to be gene dependent and could reflect accessibility of target mRNA or the relative abundances of the target mRNA and the hpRNA in cells in which the gene is active. A fragment length of between 100 and 800 bp, preferably between 300 and 600 bp, is generally suitable to maximize the efficiency of silencing obtained. The other consideration is the part of the gene to be targeted. 5' UTR, coding region, and 3' UTR fragments can be used with equally good results. As the mechanism of silencing depends on sequence homology there is potential for cross-silencing of related mRNA sequences. Where this is not desirable a region with low sequence similarity to other sequences, such as a 5' or 3' UTR, should be chosen. The rule for avoiding cross-homology silencing appears to be to use sequences that do not have blocks of sequence identity of over 20 bases between the construct and the non-target gene sequences. Many of these same principles apply to selection of target regions for designing amiRNAs.

[0121] Antisense techniques involve introducing an antisense oligonucleotide that will bind to the messenger RNA (mRNA) produced by the gene of interest. The “antisense” oligonucleotide has a base sequence complementary to the gene’s messenger RNA (mRNA), which is called the “sense” sequence. Activity of the sense segment of the mRNA is blocked by the anti-sense mRNA segment, thereby effectively inactivating gene expression.

[0122] Sense co-suppression techniques involve introducing a highly expressed sense transgene resulting in reduced expression of both the transgene and the endogenous geneAtty. Dkt. No. 135523-0115(Depicker and van Montagu, Curr. Opin. Cell Biol. 9: 373-82 (1997)). The effect depends on sequence identity between transgene and endogenous gene.IV. Deodorant Compositions: Methods and Modes of Administration and Application

[0123] The following discussion is presented by way of example only, and is not intended to be limiting.

[0124] One aspect of the present technology includes methods of treating or preventing body odor in a subject. In deodorizing applications, compositions comprising one or more ApatB- Staphylococcus hominis strains of the present technology (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, and / or Sn00377), or variants thereof, are administered to a subject suspected of, or already suffering from body odor, in an amount sufficient to reduce or eliminate the odor. Without wishing to be bound by theory, the one or more ApatB -Staphylococcus hominis strains of the present technology may persist on the skin of a subject for a period of time, whereupon they do not engage in 3M3SH production or have reduced 3M3SH production and can reduce overall rates of 3M3SH production or the total amount thereof, possibly by reducing the amount of Gly-Cys-3M3SH available to other Staphylococcus bacteria present on the skin. In some embodiments, the genetically engineered Staphylococcus hominis strain of the compositions persists on the human skin for a period of time (e.g., at least about 1 hour, at least about 2 hours, at least about 3 hours, at least about 4 hours, at least about 5 hours, at least about 6 hours, at least about 7 hours, at least about 8 hours, at least about 9 hours, at least about 10 hours, at least about 11 hours, at least about 12 hours, at least about 24 hours, at least about 48 hours, at least about 72 hours, at least about 96 hours, at least about 120 hours, at least about 144 hours, at least about 1 week).

[0125] Compositions or pharmaceutical compositions of the present technology (e.g., deodorant compositions) can be formulated for topical administration, including for administration to a subject as a topical deodorant. In some embodiments, the deodorant compositions or pharmaceutical compositions is a topical deodorant. In some embodiments, the deodorant compositions or pharmaceutical compositions are formulated into a topical deodorant. In some embodiments, the subject is a mammal. In some embodiments, theAtty. Dkt. No. 135523-0115 mammalian subject is human. The deodorant compositions or pharmaceutical compositions comprising the bacterial strains or the genetically engineered Staphylococcus hominis strains (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377, or variant strains thereof) of the present technology can be applied to any dermal location on a subject, including but not limited to the armpit, groin, nipples, and / or anywhere that glands, such as apocrine sweat glands, are located. Additionally, the deodorant compositions or pharmaceutical compositions as disclosed herein can be formulated using any appropriate means, including as a deodorant stick, a roll-on, a cream, a lotion, a gel, a powder, a water activated cream, a water activated powder, an injectable, a patch, or a spray. In some embodiments, the compositions or pharmaceutical compositions of the present technology comprise one or more lyophilized or desiccated bacterial strains of the present technology (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377, or a variant thereof), optionally in a powdered form, which can be mixed with water or a water-containing composition and subsequently applied to a subject as a topical deodorant, optionally to the armpits. In some embodiments, the water or a water-containing composition can be naturally occurring sweat from the subject. In some embodiments, the deodorant compositions or pharmaceutical compositions as disclosed herein comprise lyophilized or desiccated strains of the present technology. In some embodiments, the topical compositions or pharmaceutical compositions comprise lyophilized or desiccated strains of the present technology at an amount of about 0.0001% to about 35% by weight of the total composition (wt. %). In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.001 wt. % to about 15 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.01 wt. % to about 10 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.1 wt. % to about 10 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.15 wt. % to about 10 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.2 wt. %Atty. Dkt. No. 135523-0115 to about 10 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 10 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 9 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 8 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 7 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 6 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 5 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 4 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 3 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 2 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized or desiccated strains at an amount of about 0.25 wt. % to about 1 wt. %. In some embodiments, the deodorant compositions or pharmaceutical compositions comprise the lyophilized strains or desiccated at an amount of about 0.25 wt. % to about 0.50 wt. %. In some embodiments, the amount of acceptable carrier or excipient in the deodorant compositions or pharmaceutical compositions ranges from about 65 wt. % to about 99.9999 wt. %. In some embodiments, the amount of acceptable carrier or excipient in the deodorant compositions or pharmaceutical compositions ranges from about 90 wt. % to about 99.9999 wt. %. In some embodiments, the topical deodorant compositions and or pharmaceutical compositions comprise about 0.25 wt. % to about 5 wt. % lyophilized strains of the present technology and about 95 wt. % to about 99.75 wt. % acceptable carrier or excipient. Herein, any of the upper limits of the amounts of the components of the deodorant compositions or pharmaceutical compositions disclosed can be combined with any of theAtty. Dkt. No. 135523-0115 lower limits of the amounts of the components of the deodorant compositions or pharmaceutical compositions. In some embodiments, the excipient is polyethylene glycol (PEG), Isopropyl Myristate, or sorbitol, or a combination of excipients. In some embodiments, the topical deodorant comprises about 105to about 1014viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 106to about 1013viable bacteria per gram of the deodorant compositions or pharmaceutical composition. In some embodiments, the topical deodorant comprises about 107to about 1011viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 105viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 106viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 107viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 108viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 109viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about IO10viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1011viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1012viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1013viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1014viable bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the amount of viable bacteria as disclosed herein is a colony-forming unit (CFU). For example, in some embodiments, the topical deodorant comprises about 105to about 1014viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 106to about 1013viable CFUs per gram of the deodorant compositions or pharmaceutical composition. InAtty. Dkt. No. 135523-0115 some embodiments, the topical deodorant comprises about 107to about 1011viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 105viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 106viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 107viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 108viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 109viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about IO10CFUs bacteria per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1011viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1012viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1013viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, the topical deodorant comprises about 1014viable CFUs per gram of the deodorant compositions or pharmaceutical compositions. In some embodiments, topical deodorant application of the strains, deodorant compositions, and pharmaceutical compositions of the present technology deposits about 0.1 grams to about 2 grams of the product at the application location. In some embodiments, about 0.1 grams, about 0.2 grams, about 0.4 grams, about 0.6 grams, about 0.8 grams, about 1 gram, about 1.2 grams, about 1.4 grams, about 1.6 grams, about 1.8 grams, or about 2 grams or more of the present technology is deposited at the application location.

[0126] In some embodiments, the compositions of the present technology include an acceptable carrier. In some embodiments, the acceptable carrier comprises oleic acid, stearic acid, almond butter, avocado butter, babassu butter, beeswax, cocoa butter, coconut butter, coconut oil, cupuacu butter, hemp seed butter, kokum butter, macadamia butter, mango butter, mowrah butter, olive butter, sal seed butter, shea butter, propylene glycol, glycerin, stearyl alcohol, myristyl alcohol, sunflower extract, capric triglyceride, or any combinationAtty. Dkt. No. 135523-0115 thereof. Additional components of the compositions of the present technology may include a preservative selected from the group consisting of sucrose, sodium ascorbate, and glutathione. In some embodiments the preservative is a cryoprotectant selected from the group consisting of a nucleotide, a disaccharide, a polyol, and a polysaccharide. In some embodiments, the cryoprotectant is selected from the group consisting of inosine-5’- monophosphate (IMP), guanosine-5 ’-monophosphate (GMP), adenosine-5’ -monophosphate (AMP), uranosine-5’ -monophosphate (UMP), cytidine-5’ -monophosphate (CMP), adenine, guanine, uracil, cytosine, guanosine, uridine, cytidine, hypoxanthine, xanthine, orotidine, thymidine, inosine, trehalose, maltose, lactose, sucrose, sorbitol, mannitol, dextrin, inulin, sodium ascorbate, glutathione, and skim milk.

[0127] In some embodiments, the composition comprises a lyoprotectant. In some embodiments, the lyoprotectant is a protein, a carbohydrate, or a combination thereof. In some embodiments, the lyoprotectant is a milk protein, and in particular a micellar casein. In some embodiments, the carbohydrate is sucrose. In some embodiments, the lyoprotectant is present in an amount of about 0.1% by weight of the composition to about 15% by weight of the composition.

[0128] In some embodiments, the compositions of the present technology comprise one or more ApatB-Staphylococcus hominis strains of the present technology and an additional probiotic agent (wild-type Staphylococcus epidermidis). that may improve engraftment efficacy for the ApatB-Staphylococcus hominis strains of the present technology.

[0129] In some embodiments, the disclosure of the present technology relates to a method of manufacturing the deodorant compositions, comprising homogeneously dispersing the ApatB- Staphylococcus hominis strains (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, and / or Sn00377) of the present technology in an acceptable carrier or excipient. In some embodiments, the excipient is polyethylene glycol (PEG), Isopropyl Myristate, or sorbitol. In some embodiments, the excipient is polyethylene glycol (PEG). In some embodiments, the excipient is Isopropyl Myristate. In some embodiments, the excipient is sorbitol.Atty. Dkt. No. 135523-0115

[0130] In another aspect, the present disclosure provides an article of manufacture comprising a composition of any of the preceding embodiments. In some embodiments, the article of manufacture is a deodorant stick. In some embodiments, the article of manufacture comprises one or more chambers for housing one or more lyophilized strains of the present technology. In some embodiments, the article of manufacture comprises one or more lyophilized strains of the present technology and a carrier in separate chambers, wherein the article of manufacture can be used by inducing the mixture of the one or more lyophilized strains and the carrier prior to application of the mixture to the skin of a subject.

[0131] The strains, compositions, and pharmaceutical compositions of the present technology can be administered to a subject one or more times per day. In some embodiments, the strains and compositions are administered more than once per day, including, for example, twice per day or three times per day, etc. In some embodiments, the strains and compositions are administered once per day. In some embodiments, the strains, compositions and pharmaceutical compositions are administered once every 2 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 3 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 4 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 5 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 6 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 7 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 8 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 9 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 10 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 11 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 12 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 13 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 14 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 15Atty. Dkt. No. 135523-0115 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 16 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 17 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 18 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 19 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 20 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 21 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 22 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 23 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 24 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 25 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 26 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 27 days. In some embodiments, the strains, compositions, and pharmaceutical compositions are administered once every 28 days.

[0132] In some embodiments, administering the strains and compositions of the present technology results in a reduction of malodorous bacteria, including, for example, wild-type Staphylococcus hominis. at the application location.V. Combination Treatment with ApatB-Staphylococcus hominis

[0133] In some embodiments, the ApatB-Staphylococcus hominis strains (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, and / or Sn00377) of the present technology may be combined with one or more additional agents for the prevention or treatment of body odor. Additional agents include, but are not limited to, deodorants, antiperspirants, and / or fragrances.

[0134] In some embodiments, an additional agent is administered to a subject in combination with the ApatB-Staphylococcus hominis strains (e.g strains Sn00037, Sn00048, Sn00079,Atty. Dkt. No. 135523-0115Sn00082, Sn00086, Sn00325, Sn00327, SnOO33O, SnOO337, SnOO339, Sn00343, and / or Sn003778) of the present technology such that a synergistic effect is produced. For example, administration of ApatB Staphylococcus hominis (e.g., strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, and / or Sn00377) with one or more additional agents for the prevention or treatment of body odor will have greater than additive effects in the prevention or treatment of the body odor.

[0135] In any case, the multiple agents may be administered in any order or even simultaneously. If simultaneously, the multiple agents may be provided in a single, unified form, or in multiple forms (by way of example only, either as a single deodorant stick or as two deodorant sticks). One of the agents may be given in multiple doses, or both may be given as multiple doses. In addition, the combination methods, compositions and formulations are not to be limited to the use of only two agents.EXAMPLES

[0136] The present technology is further illustrated by the following examples, which should not be construed as limiting in any way.Materials and Methods

[0137] Bacterial Strains. A strain of Staphylococcus hominis Sn00016, as described herein, Severn et al. (The Ubiquitous Human Skin Commensal Staphylococcus hominis Protects against Opportunistic Pathogens, Bacteriology, Vol. 13, 1. 3 (2022)) was obtained from Dr. Alexander Horswill (strain referred to AH5009) (IDA deposit ID 030924-03). A strain of Staphylococcus hominis Sn000040 (IDA deposit ID 030924-04) was isolated from the human axilla. Puritan swabs (Puritan Medical, 25-1506 1PF 100 INNER) pre-wetted in PBS (Gibco™ 10010031) were used to swab the human axilla for 30 seconds. Swabs were then resuspended in PBS and vortexed for one minute. A 1 : 10,000 dilution was performed, and 100 uL of the resulting dilution was plated on a TSA plate and grown overnight at 37 °C.

[0138] Genotyping Bacterial Strains. A single colony of Staphylococcus hominis Sn000040 was picked, inoculated in 5 ml B2 low broth, and incubated at 250 rpm and 37°C overnight (B2 low: 25 g yeast extract, 10 g tryptone, 10 g NaCl, 5 g glucose, and 1 g K2HPO4 in 1 L.Atty. Dkt. No. 135523-0115 wt / vol, pH 7.5, adjusted with NaOH Filter sterilized). A single colony of Staphylococcus hominis strain Sn00016 was also inoculated in 5 ml B2 low broth and incubated at 250 rpm and 37°C overnight. ImL of each 5 mL overnight cultures was centrifuged for 1 minute at 12,000 x g, and the supernatant was discarded. The resulting cell pellet was resuspended in 90 pL of cold PBS, followed by the addition of 10 pL of lysostaphin solution (10 mg / mL, Lysostaphin Recombinant ENZ2691MG, Supplier: Carbosynth LLC). The mixture was briefly vortexed and then incubated at 37°C for 40 minutes. Both samples were run through a Monarch® Genomic DNA Purification Kit #T3010S according to the manufacturer's protocol (New England Biolabs, 240 County Road, Ipswich, MA 01938), and purified gDNA was analyzed by Plasmidsaurus (Plasmidsaurus, 1850 Millrace Drive, Suite 200 Eugene, OR 97403) for Big Bacterial Genome sequencing, according to the company’s internal procedures. To identify informative core-genome positions among the S. hominis strains, a pangenome-based multiple-sequence alignment that included the study strains and publicly available S. hominis references was generated. A pangenome alignment of study strains and NCBI references was constructed with Panaroo 1.5.0, which was then filtered to retain only positions present and single-copy in all genomes (i.e., universally conserved sites), yielding >1.6 x 106nucleotide positions for downstream analyses. Positions containing gaps or ambiguous bases were excluded. A maximum-likelihood (ML) tree was inferred with RAxML using the GTR+T substitution model and both random and parsimony starting trees (10 each). All trees were midpoint-rooted. To clarify relationships among closely related strains in the highlighted clade in FIG. 4, a separate maximum-parsimony tree was constructed using only members of that clade; default settings were used, and the tree was midpoint-rooted. To assess how the presented strains span known Staphylococcus hominis diversity, pairwise evolutionary distances were computed on the same filtered core-genome alignment used for FIG. 4 (>1.6 x 106nucleotide positions; universally conserved sites). Pairwise Jukes-Cantor distances were calculated between: (i) all NCBI S. hominis reference genomes (diverse representatives of the species), and (ii) each NCBI reference versus each presented strain. Empirical cumulative distribution functions (CDFs) of these two distance sets were compared. The distributions did not differ significantly (two-sample Kolmogorov- Smirnov test, p > 0.05), supporting that the presented strains span the species-level diversity observed in public references. Based on the known membership of references in clade A andAtty. Dkt. No. 135523-0115 clade B, it was observed that the S. hominis strains of the present disclosure are well distributed throughout Clade A and Clade B, as shown in FIG. 4.

[0139] 3-methyl-3-sulfanylhexan-l-ol (3M3SH) Assay. To measure the capability of the bacterial strains of the present technology to produce 3M3SH, the following assay was performed. A single inoculated colony (e.g., WT S. hominis strain Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, or Sn00210 or ApatB-S. hominis strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, or Sn00377) was grown in 5 mL of B2 low media in a 50 mL Falcon tube at 250 RPM overnight at 37°C. The cells were pelleted at 4000xg for 10 minutes (Thermo Fisher Sorvall X4R Pro-MD), and resuspended in 2 mL of IX M9 buffer (MP Biomedicals™ 113037022). A 200 pL reaction assay mixture was prepared in a 96-well assay plate (Falcon, 96-well tissue culture plate, Ref: 353072) as follows: 150 pL resuspended cells normalized to an Optical Density (OD) 600 nm of 5.0, 50 pL of 10 mM Cys-Gly-3M3SH (Dalton Pharma services, DC-003684) dissolved in IX M9 buffer and allowed to grow for five hours at 250 RPM 37°C. After the five hour incubation, samples were pelleted at 4000xg for 10 minutes. After centrifugation, 50 pL of supernatant was transferred to a clear bottom 96 well plate and mixed with 3.2 pL of 25 mM DTNB (CAS: 68-78-3, REF: 22582, Thermo Scientific) dissolved in lx M9 buffer. Each well was worked up to a total volume of 150 pL by adding IX M9 buffer with 50mM Tris-HCl, pH 8.0. For the reagent-free negative control, the same process was carried out with cell free media. The reaction assay mixture was incubated at room temperature for 5 minutes. Photometric reactions were measured using a Promega Glomax Discovery (REF: GM3000, SN: 9700100744) at A405 nm. This experiment was carried out with three biological replicates per sample. The average and standard deviation were calculated based on biological replicates. The average absorbance for the cell free control was subtracted from all samples to remove background signal. All plotting was performed using MATLAB.

[0140] Cytosine Base Editor. A cytosine base editor plasmid as described below was utilized to catalytically obliterate the function of PatB in Staphylococcus hominis Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, and Sn00210. Specifically, the cytosine based editor (CBE) genome tool was designed toAtty. Dkt. No. 135523-0115 convert cytosine (C) to a thymine (T) in DNA without causing double strand brakes using single guide RNA (sgRNA) targeting the W 190 tryptophan amino acid site in patB. Conversion of at least one cytosine (C) on the complementary stand at the W 190 site of PatB (5’-TGG-3’ is the W190 nucleic acid sequence of the sense strand, while 3’-ACC-5’ is the nucleic acid sequence of the complementary antisense strand) introduces a premature stop codon in the expressed mRNA as UAA or UGA or UAG. The CBE system is based on a modified form of the CRISPR / Cas9 system, where the Cas9 protein is rendered “nickase” by mutation of the RuvC nuclease domain. The nickase form of Cas9 (Cas9n) can only cut one strand of DNA, reducing the likelihood of introduction double-strand breaks. The CBE system comprises a cytidine deaminase enzyme (i.e., APOBEC1 or AID). The enzyme is fused to the Cas9n and is responsible for converting cytosine (C) to uracil (U) in a targeted manner. Uracil is then interpreted as thymine (T) during DNA replication or repair, resulting in a C»G to T»A base pair substitution.

[0141] Microbiome Profiling Protocol. The following microbiome profiling protocol was carried out to determine whether the compositions comprising the bacterial strains of the present technology are present on the skin of a subject to whom the compositions are applied post-application and whether such application alters self-reported odor levels.

[0142] Product application'. A single ~1 g dose of a topical formulation containing 0.5% (w / v) lyophilized material containing Staphylococcus hominis \N\ 9* patB mutant cells was applied to both axillae of six adult volunteers using a silicone applicator. Participants completed brief questionnaires on perceived body-odor reduction immediately before sampling on study days 3 and 7 after the single application.

[0143] Sample collection'. On days 3 and 7, axillary samples were collected by gently pressing foam-tipped swabs (Puritan Medical, 25-1506 1PF 100) pre-wetted in PBS with 0.005% Tween-20 against the epidermis for ~30 s in a zig-zag pattern (one swab per axilla). Swab heads were placed into cryotubes containing 750 pL PBS with 20% glycerol (v / v), vortexed ~1 min, and stored at -80 °C until processing.

[0144] DNA extraction'. From each primary sample, 200 pL was incubated with lysozyme (Millipore Sigma Cat # 6876, final concentration of 1 mg / mL) at 37 °C for ~24 h, thenAtty. Dkt. No. 135523-0115 treated with Proteinase K (NEB Cat # 8107, final 10 U / mL) and SDS (final 0.1% w / v) at 55 °C for ~3 h. DNA was purified using PureLink Pro 96 Genomic DNA Purification Kits (Thermo Fisher Cat # 182104) following the manufacturer’s protocol, with an added 100% ethanol wash and a brief 37 °C incubation prior to elution to improve yield.

[0145] Amplicon library preparation and sequencing'. Amplicons spanning patB around codon 190 were generated by a two-step PCR: (i) gene-specific amplification with primers flanked by Nextera-style (Illumina-compatible) overhangs targeting conserved regions upstream / downstream of codon 190 (SEQ ID Nos. 37 and 38); (ii) limited-cycle indexing to add flow-cell adapters and sample indices. Libraries were purified, normalized to equal mass, pooled, and sequenced with paired-end 2^300 bp reads. SEQ ID Nos. 37 and 37 (Forward and reverse Nextera-overhang primers) are as follows:Forward: 5’-TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGCGTATGTGGGTAGCAGAT-3’(SEQ ID NO: 38)Reverse: 5’-GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGCTGCGTTATGAACAAATTGCT CTG-3 (SEQ ID NO: 39)

[0146] Bioinformatic processing and engraftment metrics'. Reads were demultiplexed and adapter / primer-trimmed (cutadapt) and processed into AS Vs (USEARCH vl 1) with chimera removal. Because codon 190 in S. hominis patB is conserved as TGG (Trp) in wild-type genomes, AS Vs encoding premature stop codons at position 190 were attributed to the applied engineered strains. For each sample, engraftment (%) was defined as the proportion of filtered patB reads which could be positively attributed to an applied engineered strain.

[0147] Quantification of Viable Cells in Lyophilized Powder Suspended in PBS: At each viability timepoint, lyophilized S. hominis powder was reconstituted in PBS (pH 7.4) and quantified by plate count. A sterile 15 mL polypropylene conical tube was pre-weighed to the nearest 0.001 g, the balance was tared, a small amount of lyophilized powder was added, the tube was re-weighed, and the powder mass was recorded by difference. PBS (1.0 mL) was added and the tube vortexed, followed by an additional 3.0 mL PBS and vortexing to homogeneity. Serial 10-fold dilutions were prepared in PBS, and 100 pL from the 105- 107Atty. Dkt. No. 135523-0115 dilutions were spread-plated in duplicate onto TSA plates. Plates were incubated at 30 °C for 24-48 h until colonies were countable. CFU mL1of the stock suspension was calculated from plate counts and dilution factors, then normalized to the recorded powder mass to report viable CFU g1of lyophilized powder.Example 1 : Creation of Cytosine Base Editor Plasmids pXS0034 and pAB00038.

[0148] pXS0034: A plasmid containing six-part inserts was constructed using Golden Gate assembly which utilizes Type IIS restriction enzymes for multi-part DNA cloning (SEQ ID NO: 1) Each DNA fragment was synthesized and amplified with Bsal sites. The fragments were cloned into a universal entry vector via Bsal assembly creating “part” plasmids. The plasmid was transformed into E. coli cells and colonies were screened and verified through restriction mapping and sequencing to confirm correct assembly in line with Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) procedures.

[0149] Each part plasmid was categorized by function (Types 1-6) with unique BsmBI- generated overhangs to ensure correct assembly. In a one-pot reaction, the six-part plasmids were combined and digested with Bsal and T4 DNA ligase. Part 1 is a 2701 bp region (SEQ ID NO: 2) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes parts of a pIMAY vector (Addgene Plasmid #68939). Specifically, part 1 encodes the repBCAD temperature sensitive Staph, origin, the constitutive E. coli! Staph, promoter, Phelp (a strong constitutive Staph, promoter) and a chloramphenicol resistance marker (cat). Part 2 is a 547 bp region (SEQ ID NO: 3) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes the low copy E. coli origin, pl5A ori. Part 3 is a 152 bp region (SEQ ID NO: 4) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes a leaky inducible Staph, promoter. Part 4 is a 5167 bp region (SEQ ID NO: 5) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes the open reading frame (ORF) for the cytosine base editor. Part 5 is a 128 bp region (SEQ ID NO: 6) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes a low strength constitutive Staph, promoter. Part 6 is a 121 bp region (SEQ ID NO: 7) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110,Atty. Dkt. No. 135523-0115Hayward, CA 94545) and encodes a sgRNA (sgW190) that introduces a premature stop codon at W 190 of patB.

[0150] pAB00038'. A plasmid containing eight-part inserts was constructed using Golden Gate assembly which utilizes Type IIS restriction enzymes for multi-part DNA cloning (SEQ ID NO: 16) Each DNA fragment was synthesized and amplified with Bsal sites. The fragments were cloned into a universal entry vector via Bsal assembly creating “part” plasmids. The plasmid was transformed into E. coli cells and colonies were screened and verified through restriction mapping and sequencing to confirm correct assembly in line with Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) procedures.

[0151] Each part plasmid was categorized by function (Types 1-8) with unique BsmBI- generated overhangs to ensure correct assembly. In a one-pot reaction, the eight-part plasmids were combined and digested with BsmBI and T4 DNA ligase. Part 1 is a 2197 bp region (SEQ ID NO: 17) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes parts of a pIMAY vector (Addgene Plasmid #68939) and half of the coding sequence for a chloramphenicol resistance marker. Specifically, part 1 encodes the repBCAD temperature sensitive Staph, origin. Part 2 is a 508 bp region (SEQ ID NO: 18) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd Suite 110, Hayward, CA 94545) that encodes the rest of the chloramphenicol resistance marker coding sequence and pHelp, a strong promoter which originates from Listeria monocytogenes and is functional in both E. coli and Staphylococcus. Part 3 is a 1343 bp region (SEQ ID NO: 19) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes the E. coli pSClOl ori for plasmid replication and propagation of this plasmid in E. coli. Part 4 is a 155 bp region (SEQ ID NO: 20) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes a leaky inducible Staph, promoter. Part 5, Part 6, and Part 7 are 1852, 1781, 1614 bp regions respectively (SEQ ID NOs: 21-23) that were synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encode the cytosine base editor machinery segmented into three separate parts. Parts 5-7 encode a fusion protein consisting of a cytidine deaminase (APOBEC1), nickase version ofAtty. Dkt. No. 135523-0115Cas9 (Cas9n), and uracil glycosylase inhibitor (UGI). Part 8 is a 170 bp region (SEQ ID NO: 24) that was synthesized by Clutch Biotechnologies, Inc. (23595 Cabot Blvd, Suite 110, Hayward, CA 94545) and encodes a sgRNA (sgW190) that introduces a premature stop codon at W 190 of patB.Example 2: Transformation of plasmids pXS0034 and pAB00038 into Sdcm E. coli BL21 strain to produce unmethylated DNA.

[0152] The pXS0034 and pAB00038 plasmids were transformed into Edcm E. coli BL21. Briefly, transformation into Sdcm E. coli BL21 strain helps evade Type IV restriction systems in Staphylococcus species because the strain is deficient in the dem gene, which encodes DNA cytosine methyltransferase, an enzyme that methylates cytosines in the sequence CCWGG (where W is A or T). Type IV restriction enzymes in Staphylococcus species recognize and cleave methylated DNA, particularly at methylated cytosines. By using a Sdcm strain such as BL21, the DNA produced is unmethylated, making it less recognizable to Type IV restriction systems in Staphylococcus. This increases the likelihood that the DNA will remain intact and functional after introduction into Staphylococcus cells.

[0153] Next, high concentration DNA (1000 ng / ul) was extracted and prepared using the ZymoPure II Plasmid Midiprep Kit (Zymo Research Corporation, 17062 Murphy Avenue, Irvine, CA 92614, USA) according to the manufacture’s protocol. High concentration DNA significantly enhances transformation efficiency in Staphylococcus strains for at least the following reasons. First, higher DNA concentration increases the likelihood that Staphylococcus cells will encounter and take up the DNA during transformation, crucial for species with naturally lower transformation efficiency. Additionally, high purity DNA reduces the addition of contaminants such as proteins, salts, and other impurities that can inhibit the downstream electroporation process by interfering with the electrical field.Example 3 : Plasmid Electroporation into S. hominis Strains.

[0154] The 5. hominis strains Sn00016, Sn00040, Sn00042, Sn00015, Sn00085, Sn00201, Sn00202, Sn00203, Sn00213, Sn00215, Sn00239, and Sn00210 were streaked onto TSA plates and then incubated overnight at 37°C. A single colony per strain was inoculated into 50 mL of BHIS (BD Difco™ 237500) and incubated overnight at 37°C in a shakingAtty. Dkt. No. 135523-0115 incubator at 250 RPM. A 1 : 10 dilution of the overnight culture in 1 mL volume was performed and the OD measured using a spectrophotometer at OD 578 nm (Thermo Scientific Spectronic Genesys 20 Model 4001 / 4). The overnight culture was then subcultured to an OD of 0.2 in 50 mL BHIS and incubated at 37°C in a shaking incubator at 250 RPM until an OD of 0.7 was achieved. Next, the subculture was placed on ice for 10 minutes followed by centrifugation at 3500 x g for 10 minutes at 4°C using a pre chilled centrifuge. The supernatant was discarded and the pellet was resuspended in an equal volume of 10% ice-cold glycerol. The centrifugation and resuspension process was repeated five times. At completion of the fifth cycle, the pellet was resuspended in 100 pL of 10% ice-cold glycerol per 50 mL of starting culture and transferred to a sterile microcentrifuge tube ready for electroporation. If electroporation was not to be performed on the same day, the solution was flash frozen using liquid nitrogen or an ethanol dry ice bath. The frozen electrocompetent cells were later prepared for electroporation by thawing on ice for 5 minutes and then incubating at room temperature for 5 minutes. Electrocompetent cells were centrifuged at 6,000xg for 1 min, and the supernatant was discarded. Electrocompetent cells were resuspended in 10% Glycerol with 0.5M Sucrose.

[0155] Next, 50 pl of electrocompetent cells and 500 fmol of plasmid DNA from Example 2 (suspended in water) was transferred to an electroporation cuvette with a 0.1 cm gap, incubated for 5 minutes at room temperature, and electroporated using a BioRad Micropulser II for 2.5 milliseconds according to the following parameters: Voltage: 1.8 kV, Resistance: 100 Q, and Capacitance: 25 pF. The cell suspension was then transferred to 1 mL of room temperature BHIS and incubated while shaking at 250 RPM for 3 hours at 30°C. The cell suspension was then centrifuged at 3500 x g for 10 minutes, the cell pellet was resuspended in 100 pL BHIS, and plated on pre-warmed 30°C TSA CmlO plates (10 pg / ml chloramphenicol) and incubated at 30°C for 48 hours.Example 4: Validation of ApatB-S. hominis Strains.

[0156] 16S sequencing was performed by Quintara Biosciences, Inc. according to their internal procedure (3563 Investment Blvd, Suite 2, Hayward, CA 94545) on a single transformant. The 16S was amplified out of the transformant using the 27F forward 16S primer (SEQ ID NO.: 8) and the 1492R reverse 16S primers (SEQ ID NOs: 9-11). TheAtty. Dkt. No. 135523-0115 amplicon was sequenced using Sanger sequencing and then aligned to the S. hominis reference genome. The results confirmed that the transformant was from the S. hominis species.

[0157] Next, using the patB004 forward patB primer (SEQ ID NO: 12) and patB002 reverse patB primer (SEQ ID NO: 13), PCR was performed by Quintara Biosciences, Inc. (3563 Investment Blvd, Suite 2, Hayward, CA 94545) to verify that the transformant had a premature stop codon introduced at the W190 tryptophan amino acid site in the patB gene.

[0158] These results confirmed that a single patB-S. hominis Sn00037 strain has a premature stop codon of TAA at W190 compared to the wild-type strain (FIG. 1A) and a single patB-S. hominis Sn00048 strain has a premature stop codon of TAA at W 190 compared to the wild-type strain (FIG. IB). Other disclosed strains also possess a premature stop codon of TAA / TGA at W 190 compared to the wild-type strain (FIGs. 1C-1L). A summary of the strains of the present disclosure is provided in the Table below:

[0159] The genetic diversity of the engineered S. hominis strains was compared against all known S. hominis strains in the NCBI database as described in the materials and methods. As shown in FIG. 5, the pairwise distance amongst NCBI S. hominis strains (black) and between NCBI S. hominis strains and the engineered S. hominis strains (gray) are not significantly different (2-sample Kolmogorov-Smirnov test), demonstrating the high phylogenetic diversity of engineered S. hominis strains, which is similar to the phylogenetic diversity of the known S. hominis species-level phylogenyAtty. Dkt. No. 135523-0115

[0160] Accordingly, the engineered S. hominis strains of the present technology were validated for the insertion of a premature stop codon at the *W190 position and were shown to possess a broad range of genetic diversity.Example 5: patB-S. hominis Strains Exhibit Wild-Type Growth Rates.

[0161] Triplicate biological replicates of wild-type (WT) S. hominis Clade A strain Sn00016, knockout patB S. hominis Clade A strain Sn00037, WT S. hominis Clade B strain Sn00040, and knockout patB S. hominis Clade B strain Sn00048,were grown in a 96-well plate, in 200 pL TSB liquid media for 10 hours at 37°C and 220 RPM, in a BioTek Epoch 2 Microplate Spectrophotometer with A600nm recorded at 10 minute intervals. It was anticipated that patB-S. hominis strains (Sn00037 and Sn00048) would possess a growth disadvantage compared to wild-type, given that patB-S. hominis is more metabolically restricted and disruption of metabolic pathways can have unpredictable side effects.

[0162] Surprisingly, the resulting growth curve, shown in FIG. 2, demonstrated that neither patB-S. hominis cell line (Sn00037 and Sn00048; knockout-patB; represented by the triangles) exhibited a growth disadvantage compared to the wild-type (Sn00016 and Sn00040; WT patB; represented by the circles) bacteria, and therefore ^patB-S. hominis does not possess any competitive growth disadvantage or advantage. These results are surprising because a highly conserved gene such as patB should be essential for cell fitness. Moreover, as described above, the patB gene operates at a critical juncture in a key metabolic pathway for S. hominis, which would suggest that disruption of patB would affect growth. Surprisingly, however, knocking out patB did not reduce cell fitness at any of the observed growth stages.

[0163] Accordingly, these results demonstrate that the ^patB-S. hominis cell lines (strain Sn00037 and strain Sn00048) of the present disclosure possess wildtype growth capabilities. Accordingly, the patB-S. hominis cell lines of the present disclosure are useful in multiple forms, including as strain isolates and compositions, and have a sufficient growth rate to persist within a microbiota, and are useful in methods and compositions for inhibiting or reducing malodor on the skin of a subject (body odor).Atty. Dkt. No. 135523-0115Example 6: SpalB-S. hominis Strains Engraft on Subjects and Reduce Body Odor.

[0164] The ability of the patB-S. hominis strains of the present disclosure to treat, prevent, or reduce body odor (malodor) in a subject was determined using the odor studies as described in the materials and methods sections on Microbiome Profiling Protocol, Product application, Sample Collection, DNA extraction, Amplicon library preparation and sequencing, and Bioinformatic processing and engraftment metrics.

[0165] Briefly, a single dose of the Sn00079 S. hominis bacteria was applied to both axillae of six adult volunteers using a silicone applicator. Odor assessments and microbiome profiling were performed at 3- and 7-days post treatment. As shown in FIG. 6, a surprising 100% of participants showed engraftment of Sn00079 at both 3- and 7-days post treatment with a threshold of 10% or more strain prevalence. This sustained result was unexpected, as the patB-S. hominis Sn00079 strain was predicted to possess a fitness defect relative to the native microbiota in the axillae environment due to the metabolic deficiency induced by the patB W190* mutation. Furthermore, treatment with the SpatB-S. hominis Sn00079 strain was shown to reduce odor in 6 / 6 participants at 3-days post treatment and 5 / 6 participants at 7-days post treatment, demonstrating that a single application of the strains of the present technology is effective for reducing malodor over an extended duration.

[0166] Accordingly, these results demonstrate that the compositions of the present technology are useful for methods of treating, reducing, or preventing malodor on the skin of a subject.Example 7: SpatB-S. hominis Strains have Reduced 3M3SH Production.

[0167] The ability of the SpatB-Staphylococcus hominis strains (strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, and Sn00377) of the present disclosure to produce 3M3SH was assayed as described above in the 3-methyl-3-sulfanylhexan-l-ol (3M3SH) Assay section, using a comparable wild-type Staphylococcus hominis strain as a control.

[0168] The PatB enzyme in S. hominis is known to cleave Gly-3M3SH during thiol production. As shown in FIG. 3A, the wild-type (WT) strains from both major clades of S.Atty. Dkt. No. 135523-0115 hominis (WT S. hominis Clade A strain Sn00016 and WT S. hominis Clade B strain Sn00040) successfully metabolized Cys-Gly-3M3SH and produced 3M3SH, as indicated by the significant increase in absorbance at 405nm wavelength over time compared to the negative control species of E. coli and S. epidermidis. In contrast, ApatB-S. hominis (strain Sn00037 and strain Sn00048) is largely incapable of generating 3M3SH, as shown by the lack of significant change in absorbance at the 405nm wavelength. These results indicate that the respective ApatB-S. hominis strains Sn00037 and Sn00048 lack a functional patB gene and cannot produce 3M3SH. Moreover, FIG. 3B shows that for each mutant (strains Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325, Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, and Sn00377) and wild-type strain of the present disclosure, the WT S. hominis successfully produced 3M3SH, whereas the ApatB-S. hominis strains were largely incapable of producing 3M3SH. Accordingly, these results demonstrate that a probiotic deodorant comprising one or more ApatB-S. hominis strains of the present technology is effective for reducing, preventing, or ameliorating body odor.Example 8: Lyophilized SpatB-S. hominis Strains Retain Viability.

[0169] The viability of a lyophilized patB-S. hominis strain of the present disclosure was tested. Briefly, 50mL of a patB-S. hominis strain prepared by the methods described herein was grown up overnight in TSB at 37°C. The overnight culture was centrifuged (4,000 g, 10 min) and resuspended (lOx dilution) in PBS with 10% (m / v) trehalose and 10% (m / v) inulin. ImL of this suspension was poured into 1.5mL microfuge tubes. The mixture was thoroughly vortexed and stored at -80 °C before freeze-drying or spray drying.

[0170] The samples were lyophilized in open microfuge tubes using a VirTis Advantage Pro per the manufacturer’s protocol. During lyophilization, samples were first kept at -40 °C with a condenser temperature of -55 °C and chamber pressure of 10 Pa for 18 hours, and then the condenser was turned off, and the samples remained at room temperature and 10 Pa for 5 hours. After lyophilization, samples were kept in sealed 1.5mL microfuge tubes at room temperature and kept under pressure. Lyophilized samples of the patB-S. hominis strain were assessed for viability as described in the Materials and Methods.Atty. Dkt. No. 135523-0115

[0171] As shown in FIG. 7, lyophilized samples retained viability pose resuspension as late as 200 days post lyophilization. Accordingly, these results demonstrate that the patB-S. hominis strains of the present disclosure retain viability in lyophilized form for an extended duration, and are therefore useful in compositions, articles of manufacture (e.g., deodorants), and methods for the treatment or prevention of malodor.

[0172] Accordingly, these results demonstrate that the patB-S. hominis strains of the present technology retains viability in long-term storage, and therefore a probiotic deodorant or pharmaceutical composition comprising the patB-S. hominis strains can be used to treat, reduce, or prevent body odor.Example 9: patB-S. hominis Strains Reduces Body Odor.

[0173] The ability of patB-S. hominis to reduce body odor will be tested. Briefly, a mixture of the patB-S. hominis strains of the present technology (e.g., strain Sn00037, strain Sn00048, and / or any of the strains described herein or variants thereof) alone or in combination in an excipient will be applied to one armpit of a subject while excipient without the bacterial strains will be applied to the other armpit of the subject. One week and one month later the subject will engage in exercise and an odor test will be performed by smelling the subject’s armpits. It is anticipated that the armpit receiving the ^patB-S. hominis strains either alone or in combination in excipient will be identified as substantially less odorous as compared to the armpit receiving excipient alone. Accordingly, it is anticipated that these results will demonstrate that a probiotic deodorant or pharmaceutical composition comprising the ^patB-S. hominis strains of the present disclosure will be effective for treating, reducing, or preventing body odor.EQUIVALENTS

[0174] The present technology is not to be limited in terms of the particular embodiments described in this application, which are intended as single illustrations of individual aspects of the present technology. Many modifications and variations of this present technology can be made without departing from its spirit and scope, as will be apparent to those skilled in the art. Functionally equivalent methods and apparatuses within the scope of the present technology, in addition to those enumerated herein, will be apparent to those skilled in the artAtty. Dkt. No. 135523-0115 from the foregoing descriptions. Such modifications and variations are intended to fall within the scope of the appended claims. The present technology is to be limited only by the terms of the appended claims, along with the full scope of equivalents to which such claims are entitled. It is to be understood that this present technology is not limited to particular methods, reagents, compounds compositions or biological systems, which can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting.

[0175] Each and every publication and patent mentioned in the above specification is herein incorporated by reference in its entirety for all purposes. Various modifications and variations of the described methods and system of the present technology will be apparent to those skilled in the art without departing from the scope and spirit of the present technology. Although the present technology has been described in connection with specific embodiments, the present technology as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the present technology which are obvious to those skilled in the art and in fields related thereto are intended to be within the scope of the following claims.Atty. Dkt. No. 135523-0115BIOLOGICAL DEPOSITS

[0176] The Applicant requests that a sample of the deposited microorganisms should be made available only to an expert approved by the Applicant.

[0177] Staphylococcus hominis strain identified as Sn00016 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on September 3, 2024, under IDAC Accession Number 030924-03.

[0178] Staphylococcus hominis strain identified as Sn00040 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on September 3, 2024, under IDAC Accession Number 030924-04.

[0179] Staphylococcus hominis strain identified as Sn00037 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on September 3, 2024, under IDAC Accession Number 030924-01.

[0180] Staphylococcus hominis strain identified as Sn00048 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on September 3, 2024, under IDAC Accession Number 030924-02.

[0181] Staphylococcus hominis strain identified as Sn00015 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-01.

[0182] Staphylococcus hominis strain identified as Sn00042 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-02.Atty. Dkt. No. 135523-0115

[0183] Staphylococcus hominis strain identified as Sn00079 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-03.

[0184] Staphylococcus hominis strain identified as Sn00082 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-04.

[0185] Staphylococcus hominis strain identified as Sn00085 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-05.

[0186] Staphylococcus hominis strain identified as Sn00086 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-06.

[0187] Staphylococcus hominis strain identified as Sn00201 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-07.

[0188] Staphylococcus hominis strain identified as Sn00202 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-08.

[0189] Staphylococcus hominis strain identified as Sn00203 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-09.Atty. Dkt. No. 135523-0115

[0190] Staphylococcus hominis strain identified as Sn00210 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-10.

[0191] Staphylococcus hominis strain identified as Sn00213 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-11.

[0192] Staphylococcus hominis strain identified as Sn00215 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-12.

[0193] Staphylococcus hominis strain identified as Sn00239 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-13.

[0194] Staphylococcus hominis strain identified as Sn00325 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-14.

[0195] Staphylococcus hominis strain identified as Sn00327 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-15.

[0196] Staphylococcus hominis strain identified as Sn00330 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-16.Atty. Dkt. No. 135523-0115

[0197] Staphylococcus hominis strain identified as Sn00337 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-17.

[0198] Staphylococcus hominis strain identified as Sn00339 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-18.

[0199] Staphylococcus hominis strain identified as Sn00343 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-19.

[0200] Staphylococcus hominis strain identified as Sn00377 deposited with the International Depositary Authority of Canada (IDAC), National Microbiology Laboratory, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, Manitoba Canada R3E 3R2, on July 30, 2025, under IDAC Accession Number 300725-20.

[0201] The deposit was made according to the Budapest treaty on the international recognition of the deposit of microorganisms for the purposes of patent procedure.

[0202] Name of Depositor: Taxa Technologies, Inc. Address of Depositor: 135 Mississippi St., San Francisco, CA, 94107.

[0203] In respect of all designated states to which such action is possible and to the extent that it is legally permissible under the law of the designated state, it is requested that a sample of the deposited biological material be made available only by the issue thereof to an independent expert, in accordance with the relevant Patent legislation, for example EPC Rule 32, UK Patent Rules 2007 (as amended) Rule 13(1) and Schedule 1 paragraphs 6 and 7, and generally similar provisions mutatis mutandis for any other designated state.Atty. Dkt. No. 135523-0115Embodiments

[0204] Certain numbered embodiments of the present disclosure are contemplated as described below.

[0205] Embodiment 1. A genetically engineered Staphylococcus hominis strain whose genome comprises a disruption of a patB gene.

[0206] Embodiment 2. The genetically engineered Staphylococcus hominis strain of embodiment 1, wherein the strain exhibits decreased expression of the patB gene as compared to a corresponding wild-type Staphylococcus hominis strain.

[0207] Embodiment 3. The genetically engineered Staphylococcus hominis strain of embodiment 1 or embodiment 2, wherein the disruption prohibits transcription of a full- length wild-type mRNA from the disrupted patB gene.

[0208] Embodiment 4. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-3, wherein the disruption prohibits expression of a functional PatB enzyme, inactivates the PatB protein, modifies the PatB protein, or has a negative effect on the expression of the patB gene.

[0209] Embodiment 5. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-4, wherein the genetically engineered strain has decreased PatB enzyme activity as compared to a corresponding wild-type Staphylococcus hominis strain.

[0210] Embodiment 6. The genetically engineered Staphylococcus hominis strain of embodiment 4, wherein the modified PatB protein is a truncated protein.

[0211] Embodiment 7. The genetically engineered Staphylococcus hominis strain of embodiment 6, wherein the truncated protein has decreased catalytic activity compared to a corresponding wild-type Staphylococcus hominis strain.

[0212] Embodiment 8. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-7, wherein the disrupted patB gene comprises a deletion of all or at least a portion of the gene, optionally wherein the deletion comprises deletion of the patB promoter.Atty. Dkt. No. 135523-0115

[0213] Embodiment 9. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-7, wherein the disrupted patB gene comprises an insertion of a gene cassette.

[0214] Embodiment 10. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-7, wherein the disrupted patB gene comprises a deletion of at least a portion of the gene.

[0215] Embodiment 11. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-7, wherein the disruption comprises a knockout of the patB gene.

[0216] Embodiment 12. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-7, wherein the disruption comprises an allelic replacement knockout.

[0217] Embodiment 13. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-7, wherein the disruption comprises a nucleotide substitution.

[0218] Embodiment 14. The genetically engineered Staphylococcus hominis strain of embodiment 13, wherein the nucleotide substitution is a cytosine (C) to a thymine (T).

[0219] Embodiment 15. The genetically engineered Staphylococcus hominis strain of embodiment 14, wherein the nucleotide substitution is at the W190 tryptophan amino acid site in the patB gene.

[0220] Embodiment 16. The genetically engineered Staphylococcus hominis strain of embodiment 15, wherein the nucleotide substitution results in a premature stop-codon.

[0221] Embodiment 17. The genetically engineered Staphylococcus hominis strain of embodiment 16, wherein the Staphylococcus hominis strain comprises a disrupted patB gene having about 80% to about 100% sequence identity to SEQ ID NO: 15.

[0222] Embodiment 18. The genetically engineered Staphylococcus hominis strain of embodiment 16, wherein the premature stop codon in the patB gene results in a truncated PatB protein lacking one or more enzymatic activities.Atty. Dkt. No. 135523-0115

[0223] Embodiment 19. The genetically engineered Staphylococcus hominis strain of embodiment 16, wherein the premature stop codon in the expressed mRNA of the patB gene has the sequence UAG, UGA, or UAA.

[0224] Embodiment 20. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-19, wherein the disruption is introduced by homologous recombination, site-directed mutagenesis, a Transcription activator-like effector nuclease (TALEN), a meganuclease, a zinc finger nuclease, a CRISPR / Cas9 system, a CRISPR base editing system, a CRISPR prime editing system, a CRISPR / Cpfl system, a CRISPR / Csml system, or any combination thereof.

[0225] Embodiment 21. The genetically engineered Staphylococcus hominis strain of embodiment 20, wherein the disruption is introduced by a CRISPR base editing system.

[0226] Embodiment 22. The genetically engineered Staphylococcus hominis strain of embodiment 21, wherein the CRISPR based editing system is a cytosine base editor (CBE) system.

[0227] Embodiment 23. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-22, wherein the fitness of the strain comprising the disruption is not negatively affected.

[0228] Embodiment 24. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-23, wherein the strain has a comparable growth rate as compared to a corresponding wild-type Staphylococcus hominis strain.

[0229] Embodiment 25. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-24, wherein the strain comprises a Staphylococcus hominis of clade A or a Staphylococcus hominis of clade B.

[0230] Embodiment 26. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-25, wherein the strain comprises Staphylococcus hominis deposited with the International Depositary Authority of Canada (ID AC) under ID AC AccessionAtty. Dkt. No. 135523-0115Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20, or a variant strain thereof.

[0231] Embodiment 27. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-26, wherein the strain has a reduced capacity to produce 3-methyl-3- sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain.

[0232] Embodiment 28. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-27, wherein the disruption is a deletion or nucleotide substitution in the DNA sequence of the patB gene encoding a PatB protein, wherein the deletion or nucleotide substitution inactivates the PatB protein, truncates the PatB protein, or has a negative effect on expression of the patB gene.

[0233] Embodiment 29. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-28, wherein the genetically engineered Staphylococcus hominis strain has a 16S rRNA gene sequence that is at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to a sequence selected from the group consisting of SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, and SEQ ID NO: 37.

[0234] Embodiment 30. The genetically engineered Staphylococcus hominis strain of embodiment 29, wherein the strain has the 16S rRNAgene sequence represented by a sequence selected from the group consisting of SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, and SEQ ID NO: 37.

[0235] Embodiment 31. The genetically engineered Staphylococcus hominis strain of any one of embodiments 1-30, wherein the genetically engineered Staphylococcus hominis strain is derived from a strain deposited with the International Depositary Authority of Canada (IDAC) under IDAC Accession Number 030924-03, 030924-04, 300725-02, 300725-01, 300725-05, 300725-07, 300725-08, 300725-09, 300725-11, 300725-12, 300725-13, or 300725-10, or a variant strain derived therefrom, wherein the genome has been engineered to comprise a disruption of a patB gene.Atty. Dkt. No. 135523-0115

[0236] Embodiment 32. A genetically engineered Staphylococcus hominis strain deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20, or a variant strain derived therefrom, wherein the variant strain is obtained using the deposited strain as starting material, and wherein a reduced capacity of the deposited strain to produce 3-methyl-3- sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain is retained or improved in the variant strain as compared to the deposited strain.

[0237] Embodiment 33. A Staphylococcus hominis strain, wherein the strain comprises a mutation in at least one endogenous polynucleotide selected from the group consisting of:(a) a nucleotide sequence having about 80% to about 100% sequence identity to SEQ ID NO: 14; and(b) a nucleotide sequence encoding a polypeptide comprising an amino acid sequence having about 80% to about 100% sequence identity to SEQ ID NO: 41 , such that the strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding strain that does not comprise the mutation.

[0238] Embodiment 34. The strain of embodiment 33, wherein the nucleotide sequence of (a) has about 97% to about 100% sequence identity to SEQ ID NO: 14, and / or the nucleotide sequence of (b) encodes a polypeptide comprising an amino acid sequence having about 97% to about 100% sequence identity to SEQ ID NO: 41

[0239] Embodiment 35. The strain of embodiment 33 or 34, wherein the strain grows at an equivalent or improved rate as compared to a Staphylococcus hominis strain lacking the mutation.

[0240] Embodiment 36. The strain of any one of embodiments 33-35, wherein the strain is incapable of converting Cys-Gly-3M3SH to 3M3SH.

[0241] Embodiment 37. A composition comprising an effective amount of one or more of the genetically engineered Staphylococcus hominis strains of any one of embodiments 1-36.Atty. Dkt. No. 135523-0115

[0242] Embodiment 38. The composition of embodiment 37, wherein the one or more Staphylococcus hominis strains are lyophilized.

[0243] Embodiment 39. The composition of embodiment 37 or embodiment 38 further comprising a lyoprotectant.

[0244] Embodiment 40. The composition of embodiment 39, wherein the lyoprotectant is selected from one or more of sucrose, micellar casein, trehalose, or inulin.

[0245] Embodiment 41. The composition of any one of embodiments 37-40, wherein the one or more Staphylococcus hominis strains are viable.

[0246] Embodiment 42. The composition of any one of embodiments 37-41, wherein the one or more Staphylococcus hominis strains are nonproliferative.

[0247] Embodiment 43. The composition of any one of embodiments 37-42 further comprising Staphylococcus epidermidis.

[0248] Embodiment 44. The composition of any one of embodiments 37-43 further comprising an excipient.

[0249] Embodiment 45. The composition of embodiment 44, wherein the excipient is selected from one or more of oleic acid, stearic acid, almond butter, avocado butter, babassu butter, beeswax, cocoa butter, coconut butter, coconut oil, cupuacu butter, hemp seed butter, kokum butter, macadamia butter, mango butter, mowrah butter, olive butter, sal seed butter, shea butter, propylene glycol, glycerin, stearyl alcohol, myristyl alcohol, capric triglyceride, polyethylene glycol (PEG), Isopropyl Myristate, or sorbitol.

[0250] Embodiment 46. The composition of any one of embodiments 37-45, wherein the Staphylococcus hominis strain is present at a concentration of about 106to about 1013bacteria per gram.

[0251] Embodiment 47. The composition of embodiment 46, wherein the Staphylococcus hominis strain is present at a concentration of about 107to about 1011bacteria per gram.Atty. Dkt. No. 135523-0115

[0252] Embodiment 48. The composition of any one of embodiments 44-47, comprising about 90 wt. % to about 99.99 wt. % excipient.

[0253] Embodiment 49. The composition of embodiment 48, wherein the lyophilized Staphylococcus hominis bacteria is present at about 10 wt. % to about 0.01 wt. %.

[0254] Embodiment 50. The composition of embodiment 48, comprising about 95 wt. % to about 99.75 wt. % excipient.

[0255] Embodiment 51. The composition of embodiment 50, wherein the lyophilized Staphylococcus hominis bacteria is present at about 5 wt. % to about 0.25 wt. %.

[0256] Embodiment 52. The composition of any one of embodiments 37-51, further comprising a fragrance selected from one or more of elettaria cardamomum seed oil, eucalyptus globulus seed oil, citrus aurantium bergamia fruit oil, mentha piperita oil, salvia sclarea oil, juniperus virginiana oil, abies sibirica oil, citrus aurantium amara, citrus aurantium dulcis lower oil, citrus nobilis oil, rosmarinus officinalis oil, melaleuca alternifolia oil, citrus limon tree oil, zingiber officinale root oil, Mangifera indica oil, Lavandula angustifolia oil, and Santalum album.

[0257] Embodiment 53. The composition of any one of embodiments 37-52, wherein the composition is formulated for topical administration.

[0258] Embodiment 54. The composition of embodiment 53, wherein the composition is formulated for use as a topical deodorant.

[0259] Embodiment 55. The composition of any one of embodiments 37-54, further comprising an antiperspirant.

[0260] Embodiment 56. The composition of embodiment 55, wherein the antiperspirant comprises one or more of aluminum salt, charcoal, and witch hazel.

[0261] Embodiment 57. The composition of any one of embodiments 37-56, wherein the topical deodorant is formulated as a deodorant stick, a roll-on, a cream, a lotion, a gel, a powder, a water activated cream, a water activated powder, an injectable, a patch, or a spray.Atty. Dkt. No. 135523-0115

[0262] Embodiment 58. The composition of any one of embodiments 37-57, wherein the composition is a pharmaceutical composition and the effective amount of one or more of the genetically engineered Staphylococcus hominis strains is a therapeutically effective amount.

[0263] Embodiment 59. A pharmaceutical composition comprising a therapeutically effective amount of one or more genetically engineered Staphylococcus hominis strains whose genome comprises a disruption of a patB gene, and pharmaceutically acceptable carrier or excipient, and optionally, a lyoprotectant.

[0264] Embodiment 60. The pharmaceutical composition of embodiment 59, wherein the disruption of the patB gene comprises a nucleotide substitution, optionally wherein the nucleotide substitution comprises a cytosine (C) to thymine (T) substitution as compared to a corresponding wild-type Staphylococcus hominis strain.

[0265] Embodiment 61. The pharmaceutical composition of embodiment 60, wherein the substitution is at the W190 tryptophan amino acid site in the patB gene.

[0266] Embodiment 62. The pharmaceutical composition of embodiment 60 or 61, wherein the substitution introduces a premature stop codon.

[0267] Embodiment 63. The pharmaceutical composition of any one of embodiments 59-62, wherein the one or more genetically engineered Staphylococcus hominis strains are lyophilized.

[0268] Embodiment 64. The pharmaceutical composition of embodiment 63, wherein the pharmaceutically acceptable carrier or excipient is present at about 90 wt. % to about 99.99 wt. % and the one or more lyophilized genetically engineered Staphylococcus hominis strains is present at about 10 wt. % to about 0.01 wt. %.

[0269] Embodiment 65. The pharmaceutical composition of embodiment 64, wherein the pharmaceutically acceptable carrier or excipient is present at about 95 wt. % to about 99.75 wt. % and the one or more lyophilized genetically engineered Staphylococcus hominis strains is present at about 5 wt. % to about 0.25 wt. %.Atty. Dkt. No. 135523-0115

[0270] Embodiment 66. The pharmaceutical composition of any one of embodiments 59-65, wherein the one or more genetically engineered Staphylococcus hominis strains are present at a concentration of about 106to about 1013bacteria per gram.

[0271] Embodiment 67. The pharmaceutical composition of embodiment 66, wherein the one or more genetically engineered Staphylococcus hominis strains are present at a concentration of about 107to about 10nbacteria per gram.

[0272] Embodiment 68. The pharmaceutical composition of any one of embodiments 59-67, wherein the one or more genetically engineered Staphylococcus hominis strains are selected from the strains deposited with the International Depositary Authority of Canada (ID AC) under IDAC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20.

[0273] Embodiment 69. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the International Depositary Authority of Canada (IDAC) under IDAC Accession Number 030924-01.

[0274] Embodiment 70. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with IDAC under IDAC Accession Number 030924-02.

[0275] Embodiment 71. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-03.

[0276] Embodiment 72. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-04.

[0277] Embodiment 73. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-06.Atty. Dkt. No. 135523-0115

[0278] Embodiment 74. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-14.

[0279] Embodiment 75. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-15.

[0280] Embodiment 76. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-16.

[0281] Embodiment 77. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-17.

[0282] Embodiment 78. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-18.

[0283] Embodiment 79. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-19.

[0284] Embodiment 80. The pharmaceutical composition of embodiment 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the IDAC under IDAC Accession Number 300725-20.

[0285] Embodiment 81. The pharmaceutical composition of any one of embodiments 59-67, comprising two or more of the genetically engineered Staphylococcus hominis strains deposited with IDAC under IDAC Accession Numbers 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725- 19, and 300725-20.Atty. Dkt. No. 135523-0115

[0286] Embodiment 82. A method of reducing 3-methyl-3-sulfanylhexan-l-ol (3M3SH) production from a skin microbiota of a subject, the method comprising applying: (i) a composition or a pharmaceutical composition comprising the Staphylococcus hominis strain of any one of embodiments 1-36 to the skin of the subject; (ii) the composition of any one of embodiments 37-58; or (iii) the pharmaceutical composition of any one of embodiments 59- 81 to the skin of the subject.

[0287] Embodiment 83. A method for treating, reducing, or preventing malodor on the skin of a subject, comprising treating the skin of the subject with a therapeutically effective amount of the composition of any one of embodiments 37-58 or the pharmaceutical composition of any one of embodiments 59-81.

[0288] Embodiment 84. The method of embodiment 83, wherein the composition or pharmaceutical composition is applied topically to the skin of the subject.

[0289] Embodiment 85. The method of embodiment 83 or 84, wherein the composition or pharmaceutical composition is applied to a sweat-gland-containing area of skin of the subject.

[0290] Embodiment 86. The method of any one of embodiments 83-85, wherein the composition or the pharmaceutical composition is administered to the subject about 1 to about 2 times per day for about 1 to about 7 days.

[0291] Embodiment 87. The method of any one of embodiments 83-86, wherein the treated skin of the subject comprises a reduction in the amount of malodorous bacteria as compared to a corresponding untreated portion of skin.

[0292] Embodiment 88. The method of embodiment 87, wherein the malodorous bacteria comprises at least wild-type Staphylococcus hominis.

[0293] Embodiment 89. A method of producing a genetically engineered Staphylococcus hominis strain, the method comprising targeting the patB gene for disruption.

[0294] Embodiment 90. The method of embodiment 89, wherein the disruption is introduced into the patB gene by homologous recombination, site-directed mutagenesis, a Transcription activator-like effector nuclease (TALEN), a meganuclease, a zinc finger nuclease, aAtty. Dkt. No. 135523-0115CRISPR / Cas9 system, a CRISPR / Cpfl system, or a CRISPR / Csml system, and any combination thereof.

[0295] Embodiment 91. The method of embodiment 90, wherein the disruption is introduced into the patB gene by the CRISPR base editing system.

[0296] Embodiment 92. The method of embodiment 91, wherein the CRISPR based editing system is a cytosine base editor (CBE) system.

[0297] Embodiment 93. The method of embodiment 91, wherein the CRISPR based editing system comprises transforming a wild-type Staphylococcus hominis strain with a 6-part plasmid.

[0298] Embodiment 94. The method of embodiment 93, wherein the 6-part plasmid comprises one or more sequences selected from SEQ ID NOs: 2-7.

[0299] Embodiment 95. The method of embodiment 94, wherein the 6-part plasmid comprises the sequence of SEQ ID NO: 1.

[0300] Embodiment 96. The method of embodiment 91, wherein the CRISPR based editing system comprises transforming a wild-type Staphylococcus hominis strain with an 8-part plasmid.

[0301] Embodiment 97. The method of embodiment 96, wherein the 8-part plasmid comprises one or more sequences selected from SEQ ID NOs: 17-24.

[0302] Embodiment 98. The method of embodiment 97, wherein the 8-part plasmid comprises the sequence of SEQ ID NO: 16.

[0303] Embodiment 99. The method of any one of embodiments 89-98, wherein the strain exhibits decreased expression of the patB gene as compared to a corresponding wild-type Staphylococcus hominis strain.

[0304] Embodiment 100. The method of any one of embodiments 89-99, wherein the disruption prohibits transcription of a full-length wild-type mRNA from the disrupted patB gene.Atty. Dkt. No. 135523-0115

[0305] Embodiment 101. The method of any one of embodiments 89-100, wherein the disruption prohibits expression of a functional PatB enzyme.

[0306] Embodiment 102. The method of any one of embodiments 89-101, wherein the genetically engineered strain has decreased PatB enzyme or catalytic activity and the PatB protein is truncated as compared to a corresponding wild-type Staphylococcus hominis strain.

[0307] Embodiment 103. The method of any one of embodiments 89-102, wherein the disrupted patB gene comprises a nucleotide substitution, wherein the substitution is a cytosine (C) to a thymine (T).

[0308] Embodiment 104. The method of any one of embodiments 89-103, wherein the strain is selected from Staphylococcus hominis strains deposited with the ID AC under ID AC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20, or a variant strain thereof.

[0309] Embodiment 105. The method of any one of embodiments 89-104, wherein the genetically engineered Staphylococcus hominis strain has a reduced capacity to produce 3- methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain.

[0310] Embodiment 106. An altered microbiota comprising a genetically engineered Staphylococcus hominis strain, or a variant strain thereof, whose genome comprises a disruption of a patB gene.

[0311] Embodiment 107. The altered microbiota of embodiment 106, wherein the microbiota has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat as compared to a corresponding wild-type microbiota.

[0312] Embodiment 108. A deodorant composition comprising:(i) a therapeutically effective amount of a viable Staphylococcus hominis strain of any one of embodiments 1-36; and(ii) a therapeutically acceptable carrier or excipient;Atty. Dkt. No. 135523-0115 wherein the deodorant composition is formulated as a water activated formulation for topical administration.

[0313] - Embodiment 109. The deodorant composition of embodiment 108, comprising about 90 wt. % to about 99.99 wt. % therapeutically acceptable carrier or excipient.

[0314] Embodiment 110. The deodorant composition of embodiment 109, wherein the Staphylococcus hominis strain is lyophilized.

[0315] Embodiment 111. The deodorant composition of embodiment 110, wherein the deodorant composition comprises about 10 wt. % to about 0.01 wt. % lyophilized Staphylococcus hominis bacteria.

[0316] Embodiment 112. The deodorant composition of embodiment 109, comprising about 95 wt. % to about 99.75 wt. % of the therapeutically acceptable carrier or excipient.

[0317] Embodiment 113. The deodorant composition of embodiment 112, wherein the deodorant composition comprises about 5 wt. % to about 0.25 wt. % lyophilized Staphylococcus hominis bacteria.

[0318] Embodiment 114. The deodorant composition of any one of embodiments 108-113, wherein the Staphylococcus hominis strain is present at a concentration of about 106to about 1016bacteria per gram.

[0319] Embodiment 115. The deodorant composition of embodiment 114, wherein the Staphylococcus hominis strain is present at a concentration of about 107to about 1011bacteria per gram.

[0320] Embodiment 116. The deodorant composition of any one of embodiments 108-115 further comprising: (iii) a lyoprotectant.

[0321] Embodiment 117. The deodorant composition of any one of embodiments 108-116, wherein the Staphylococcus hominis strain comprises one or more of the strains deposited with the IDAC under IDAC Accession Numbers 030924-01, 030924-02, 300725-03, 300725-Atty. Dkt. No. 135523-011504, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20.

[0322] Embodiment 118. A deodorant stick comprising the deodorant composition of any one of embodiments 108-117.

[0323] Embodiment 119. A pharmaceutical composition comprising a therapeutically effective amount of one or more of the Staphylococcus hominis strains of any one of embodiments 1-36, and a therapeutically acceptable carrier or excipient, for use in the treatment of malodor on the skin of a subject.

[0324] Embodiment 120. The pharmaceutical composition of embodiment 119, wherein the one or more Staphylococcus hominis strains are lyophilized.

[0325] Embodiment 121. The pharmaceutical composition of embodiment 120 further comprising a lyoprotectant.

[0326] Embodiment 122. Use of the one or more Staphylococcus hominis strains of any one of embodiments 1-36 in the manufacture of a medicament for the treatment of malodor on the skin of a subject.Atty. Dkt. No. 135523-0115SEQUENCE LISTINGStaphylococcus hominis pXS0034 plasmid: Six part cytosine base editor plasmid nucleotide sequence (SEQ ID NO: 1):CGTGATGGTAACTTCAGCATGCGTTTTAGCGTTTATTTCGTTTAGTTATCGGCATA ATCGTTAAAACAGGCGTTATCGTAGCGTAAAAGCCCTTGAGCGTAGCGTGGCTTT GCAGCGAAGATGTTGTCTGTTAGATTATGAAAGCCGATGACTGAATGAAATAAT AAGCGCAGCGCCCTTCTATTTCGGTTGGAGGAGGCTCAAGGGAGTATGAGGGAA TGAAATTCCCTCATGGGTTTGATTTTAAAAATTGCTTGCAATTTTGCCGAGCGGTA GCGCTGGAAAATTTTTGAAAAAAATTTGGAATTTGGAAAAAAATGGGGGGAAAG GAAGCGAATTTTGCTTCCGTACTACGACCCCCCATTAAGTGCCGAGTGCCAATTT TTGTGCCAAAAACGCTCTATCCCAACTGGCTCAAGGGTTTAAGGGGTTTTTCAAT CGCCAACGAATCGCCAACGTTTTCGCCAACGTTTTTTATAAATCTATATTTAAGT AGCTTTATTGTTGTTTTTATGATTACAAAGTGATACACTAACTTTATAAAATTATT TGATTGGAGTTTTTTAAATGGTGATTTCAGAATCGAAAAAAAGAGTTATGATTTC TCTGACAAAAGAGCAAGATAAAAAATTAACAGATATGGCGAAACAAAAAGGTTT TTCAAAATCTGCGGTTGCGGCGTTAGCTATAGAAGAATATGCAAGAAAGGAATC AGAACAAAAAAAATAAGCGAAAGCTCGCGTTTTTAGAAGGATACGAGTTTTCGC TACTTGTTTTTGATAAGGTAATTATATCATGGCTATTAAAAATACTAAAGCTAGA AATTTTGGATTTTTATTATATCCTGACTCAATTCCTAATGATTGGAAAGAAAAATT AGAGAGTTTGGGCGTATCTATGGCTGTCAGTCCTTTACACGATATGGACGAAAAA AAAGATAAAGATACATGGAATAATAGTAATATTATACAAAATGGAAAGCACTAT AAAAAACCACACTATCACGTTATATATATTGCACGAAATCCTGTAACAATAGAA AGCGTTAGGAACAAGATTAAGCGAAAATTGGGGAATAGTTCAGTTGCTCATGTT GAGATACTTGATTATATCAAAGGTTCATATGAATATTTGACTCATGAATCAAAGG ACGCTATTGCTAAGAATAAACATATATACGACAAAAAAGATATTTTGAACATTA ATGATTTTGATATTGACCGCTATATAACACTTGATGAAAGCCAAAAAAGAGAATT GAAGAATTTACTTTTAGATATAGTGGATGACTATAATTTGGTAAATACAAAAGAT TTAATGGCTTTTATTCGCCTTAGGGGAGCGGAGTTTGGAATTTTAAATACGAATG ATGTAAAAGATATTGTTTCAACAAACTCTAGCGCCTTTAGATTATGGTTTGAGGG CAATTATCAGTGTGGATATAGAGCAAGTTATGCAAAGGTTCTTGATGCTGAAACG GGGGAAATAAAATGACAAACAAAGAAAAAGAGTTATTTGCTGAAAATGAGGAA TTAAAAAAAGAAATTAAGGACTTAAAAGAGCGTATTGAAAGATACAGAGAAATG GAAGTTGAATTAAGTACAACAATAGATTTATTGAGAGGAGGGATTATTGAATAA ATAAAAGCCCCCTGACGAAAGTCGAAGGGGGTTTTTATTTTGGTTTGATGTTGCG ATTAATAGCAATACATTCTATAATAGAAGGTATGGAGGATGTTATATAATGAGAC AGAATTATGATGATCATATGTCAACTAACGGGGCAGGTTAGTGACATTAGAAAA CCGACTGTAAAAAGTACAGTCGGCATTATCTCATATTATAAAAGCCAGTCATTAG GCCTATCTGACAATTCCTGAATAGAGTTCATAAACAATCCTGCATGATAACCATC ACAAACAGAATGATGTACCTGTAAAGATAGCGGTAAATATATTGAATTACCTTTA TTAATGAATTTTCCTGCTGTAATAATGGGTAGAAGGTAATTACTATTATTATTGAT ATTTAAGTTAAACCCAGTAAATGAAGTCCATGGAATAATAGAAAGAGAAAAAGC ATTTTCAGGTATAGGTGTTTTGGGAAACAATTTCCCCGAACCATTATATTTCTCTA CATCAGAAAGGTATAAATCATAAAACTCTTTGAAGTCATTCTTTACAGGAGTCCA AATGCCAGAGAATGTTTTAGATACACCATCAAAAATTGTATAAAGTGGCTCTAAC TTATCCCAATAACCTAACTCTCCGTCGCTATTGTAACCAGTTCTAAAAGCTGTATTAtty. Dkt. No. 135523-0115TGAGTTTATCACCCTTGTCACTAAGAAAATAAATGCAGGGTAAAATTTATATCCT TCTTGTTTTATGTTTCGGTATAAAACACTAATATCAATTTCTGTGGTTATACTAAA AGTCGTTTGTTGGTTCAAATAATGATTAAATATCTCTTTTCTCTTCCAATTGTCTA AATCAATTTTATTAAAGTTCATGGGTTTCACTCTCCTTCTACATTTTTTAACCTAA TAATGCCAAATACCGTTTGCCACCCCTCTCTTTGATAATTATAATATTGGCGAAA TTCGCTTCTAAAGATGAAACGCAATATTATATGCTTGCTTTATCGGCCGTATGTG ATTATACCAGCCCCCTCACTACATGTCAAGAATAAACTGCCAAAGCATAATGGG ATAATTAACCCTCTTTTCCATAGGCTCCGCCCCCCTGACAAGCATCACGAAATCT GACGCTCAAATCAGTGGTGGCGAAACCCGACAGGACTATAAAGATACCAGGCGT TTCCCCCTGGCGGCTCCCTCGTGCGCTCTCCTGTTCCTGCCTTTCGGTTTACCGGT GTCATTCCGCTGTTATGGCCGCGTTTGTCTCATTCCACGCCTGACACTCAGTTCCG GGTAGGCAGTTCGCTCCAAGCTGGACTGTATGCACGAACCCCCCGTTCAGTCCGA CCGCTGCGCCTTATCCGGTAACTATCGTCTTGAGTCCAACCCGGAAAGACATGCA AAAGCACCACTGGCAGCAGCCACTGGTAATTGATTTAGAGGAGTTAGTCTTGAA GTCATGCGCCGGTTAAGGCTAAACTGAAAGGACAAGTTTTGGTGACTGCGCTCCT CCAAGCCAGTTACCTCGGTTCAAAGAGTTGGTAGCTCAGAGAACCTTCGAAAAA CCGCCCTGCAAGGCGGTTTTTTCGTTTTCAGAGCAAGAGATTACGCGCAGACCAA AACGATCTCAAGTAGTCTCCTTTTTCGCTTCTTTATTCCAATTGCTTTATTGACGTT GAGCCTCGGAACCGTCGACTTTATTTGGATCCCCTCGAGTTCATGAAAAACTAAA AAAAATATTGAAACTCTATCATTGATAGAGTATAATTAACAAGGAGGAATAAAA AATGAGCTCAGAGACTGGCCCAGTGGCTGTGGACCCCACATTGAGGCGGCGGAT CGAGCCCCATGAGTTTGAGGTATTCTTCGATCCGAGAGAGCTCCGCAAGGAGAC TTGCCTGCTTTACGAAATTAATTGGGGGGGCCGGCACTCCATTTGGCGACATACA TCACAGAACACTAACAAGCACGTCGAAGTCAACTTCATCGAGAAGTTCACGACA GAAAGATATTTCTGTCCGAACACAAGGTGCAGCATTACCTGGTTTCTCAGCTGGA GCCCATGCGGCGAATGTAGTAGGGCCATCACTGAATTCCTGTCAAGGTATCCCCA CGTCACTCTGTTTATTTACATCGCAAGGCTGTACCACCACGCTGACCCCCGCAAT CGACAAGGCCTGCGGGATTTGATCTCTTCAGGTGTGACTATCCAAATTATGACTG AGCAGGAGTCAGGATACTGCTGGAGAAACTTTGTGAATTATAGCCCGAGTAATG AAGCCCACTGGCCTAGGTATCCCCATCTGTGGGTACGACTGTACGTTCTTGAACT GTACTGCATCATACTGGGCCTGCCTCCTTGTCTCAACATTCTGAGAAGGAAGCAG CCACAGCTGACATTCTTTACCATCGCTCTTCAGTCTTGTCATTACCAGCGACTGCCCCCACACATTCTCTGGGCCACCGGGTTGAAATCTGGTGGTTCTTCTGGTGGTTCT AGCGGCAGCGAGACTCCCGGGACCTCAGAGTCCGCCACACCCGAAAGTTCTGGT GGTTCTTCTGGTGGTTCTGATAAAAAGTATTCTATTGGTTTAGCCATCGGCACTAA TTCCGTTGGATGGGCTGTCATAACCGATGAATACAAAGTACCTTCAAAGAAATTT AAGGTGTTGGGGAACACAGACCGTCATTCGATTAAAAAGAATCTTATCGGTGCC CTCCTATTCGATAGTGGCGAAACGGCAGAGGCGACTCGCCTGAAACGAACCGCT CGGAGAAGGTATACACGTCGCAAGAACCGAATATGTTACTTACAAGAAATTTTT AGCAATGAGATGGCCAAAGTTGACGATTCTTTCTTTCACCGTTTGGAAGAGTCCT TCCTTGTCGAAGAGGACAAGAAACATGAACGGCACCCCATCTTTGGAAACATAG TAGATGAGGTGGCATATCATGAAAAGTACCCAACGATTTATCACCTCAGAAAAA AGCTAGTTGACTCAACTGATAAAGCGGACCTGAGGTTAATCTACTTGGCTCTTGC CCATATGATAAAGTTCCGTGGGCACTTTCTCATTGAGGGTGATCTAAATCCGGAC AACTCGGATGTCGACAAACTGTTCATCCAGTTAGTACAAACCTATAATCAGTTGT TTGAAGAGAACCCTATAAATGCAAGTGGCGTGGATGCGAAGGCTATTCTTAGCG CCCGCCTCTCTAAATCCCGACGGCTAGAAAACCTGATCGCACAATTACCCGGAGAtty. Dkt. No. 135523-0115AGAAGAAAAATGGGTTGTTCGGTAACCTTATAGCGCTCTCACTAGGCCTGACACCAAATTTTAAGTCGAACTTCGACTTAGCTGAAGATGCCAAATTGCAGCTTAGTAAGGACACGTACGATGACGATCTCGACAATCTACTGGCACAAATTGGAGATCAGTATGCGGACTTATTTTTGGCTGCCAAAAACCTTAGCGATGCAATCCTCCTATCTGACATACTGAGAGTTAATACTGAGATTACCAAGGCGCCGTTATCCGCTTCAATGATCAAAAGGTACGATGAACATCACCAAGACTTGACACTTCTCAAGGCCCTAGTCCGTCAGCAACTGCCTGAGAAATATAAGGAAATATTCTTTGATCAGTCGAAAAACGGGTACGCAGGTTATATTGACGGCGGAGCGAGTCAAGAGGAATTCTACAAGTTTATCAAACCCATATTAGAGAAGATGGATGGGACGGAAGAGTTGCTTGTAAAACTCAATCGCGAAGATCTACTGCGAAAGCAGCGGACTTTCGACAACGGTAGCATTCCACATCAAATCCACTTAGGCGAATTGCATGCTATACTTAGAAGGCAGGAGGATTTTTATCCGTTCCTCAAAGACAATCGTGAAAAGATTGAGAAAATCCTAACCTTTCGCATACCTTACTATGTGGGACCCCTGGCCCGAGGGAACTCTCGGTTCGCATGGATGACAAGAAAGTCCGAAGAAACGATTACTCCATGGAATTTTGAGGAAGTTGTCGATAAAGGTGCGTCAGCTCAATCGTTCATCGAGAGGATGACCAACTTTGACAAGAATTTACCGAACGAAAAAGTATTGCCTAAGCACAGTTTACTTTACGAGTATTTCACAGTGTACAATGAACTCACGAAAGTTAAGTATGTCACTGAGGGCATGCGTAAACCCGCCTTTCTAAGCGGAGAACAGAAGAAAGCAATAGTAGATCTGTTATTCAAGACCAACCGCAAAGTGACAGTTAAGCAATTGAAAGAGGACTACTTTAAGAAAATTGAATGCTTCGATTCTGTCGAGATCTCCGGGGTAGAAGATCGATTTAATGCGTCACTTGGTACGTATCATGACCTCCTAAAGATAATTAAAGATAAGGACTTCCTGGATAACGAAGAGAATGAAGATATCTTAGAAGATATAGTGTTGACTCTTACCCTCTTTGAAGATCGGGAAATGATTGAGGAAAGACTAAAAACATACGCTCACCTGTTCGACGATAAGGTTATGAAACAGTTAAAGAGGCGTCGCTATACGGGCTGGGGACGATTGTCGCGGAAACTTATCAACGGGATAAGAGACAAGCAAAGTGGTAAAACTATTCTCGATTTTCTAAAGAGCGACGGCTTCGCCAATAGGAACTTTATGCAGCTGATCCATGATGACTCTTTAACCTTCAAAGAGGATATACAAAAGGCACAGGTTTCCGGACAAGGGGACTCATTGCACGAACATATTGCGAATCTTGCTGGTTCGCCAGCCATCAAAAAGGGCATACTCCAGACAGTCAAAGTAGTGGATGAGCTAGTTAAGGTCATGGGACGTCACAAACCGGAAAACATTGTAATCGAGATGGCACGCGAAAATCAAACGACTCAGAAGGGGCAAAAAAACAGTCGAGAGCGGATGAAGAGAATAGAAGAGGGTATTAAAGAACTGGGCAGCCAGATCTTAAAGGAGCATCCTGTGGAAAATACCCAATTGCAGAACGAGAAACTTTACCTCTATTACCTACAAAATGGAAGGGACATGTATGTTGATCAGGAACTGGACATAAACCGTTTATCTGATTACGACGTCGATCACATTGTACCCCAATCCTTTTTGAAGGACGATTCAATCGACAATAAAGTGCTTACACGCTCGGATAAGAACCGAGGGAAAAGTGACAATGTTCCAAGCGAGGAAGTCGTAAAGAAAATGAAGAACTATTGGCGGCAGCTCCTAAATGCGAAACTGATAACGCAAAGAAAGTTCGATAACTTAACTAAAGCTGAGAGGGGTGGCTTGTCTGAACTTGACAAGGCCGGATTTATTAAACGTCAGCTCGTGGAAACCCGCCAAATCACAAAGCATGTTGCACAGATACTAGATTCCCGAATGAATACGAAATACGACGAGAACGATAAGCTGATTCGGGAAGTCAAAGTAATCACTTTAAAGTCAAAATTGGTGTCGGACTTCAGAAAGGATTTTCAATTCTATAAAGTTAGGGAGATAAATAACTACCACCATGCGCACGACGCTTATCTTAATGCCGTCGTAGGGACCGCACTCATTAAGAAATACCCGAAGCTAGAAAGTGAGTTTGTGTATGGTGATTACAAAGTTTATGACGTCCGTAAGATGATCGCGAAAAGCGAACAGGAGATAGGCAAGGCTACAGCCAAATACTTCTTTTATTCTAACATTATGAATTTCTTTAAGACGGAAATCACTCTGGCAAACGGAGAGATACGCAAACGACCTTTAATTGAAACCAATGGGGAGACAGGTGAAATCGTATGGGATAAGGGCCGGGACTTCGCGACGGTGAtty. Dkt. No. 135523-0115AGAAAAGTTTTGTCCATGCCCCAAGTCAACATAGTAAAGAAAACTGAGGTGCAG ACCGGAGGGTTTTCAAAGGAATCGATTCTTCCAAAAAGGAATAGTGATAAGCTC ATCGCTCGTAAAAAGGACTGGGACCCGAAAAAGTACGGTGGCTTCGATAGCCCT ACAGTTGCCTATTCTGTCCTAGTAGTGGCAAAAGTTGAGAAGGGAAAATCCAAG AAACTGAAGTCAGTCAAAGAATTATTGGGGATAACGATTATGGAGCGCTCGTCTT TTGAAAAGAACCCCATCGACTTCCTTGAGGCGAAAGGTTACAAGGAAGTAAAAA AGGATCTCATAATTAAACTACCAAAGTATAGTCTGTTTGAGTTAGAAAATGGCCG AAAACGGATGTTGGCTAGCGCCGGAGAGCTTCAAAAGGGGAACGAACTCGCACT ACCGTCTAAATACGTGAATTTCCTGTATTTAGCGTCCCATTACGAGAAGTTGAAA GGTTCACCTGAAGATAACGAACAGAAGCAACTTTTTGTTGAGCAGCACAAACAT TATCTCGACGAAATCATAGAGCAAATTTCGGAATTCAGTAAGAGAGTCATCCTA GCTGATGCCAATCTGGACAAAGTATTAAGCGCATACAACAAGCACAGGGATAAA CCCATACGTGAGCAGGCGGAAAATATTATCCATTTGTTTACTCTTACCAACCTCG GCGCTCCAGCCGCATTCAAGTATTTTGACACAACGATAGATCGCAAACGATACA CTTCTACCAAGGAGGTGCTAGACGCGACACTGATTCACCAATCCATCACGGGATT ATATGAAACTCGGATAGATTTGTCACAGCTTGGGGGTGACTCTGGTGGTTCTGGA GGATCTGGTGGTTCTACTAATCTGTCAGATATTATTGAAAAGGAGACTGGTAAGC AACTGGTTATCCAGGAATCCATCCTCATGCTCCCAGAGGAGGTGGAAGAAGTCATTGGGAACAAGCCGGAAAGCGATATACTCGTGCACACCGCCTACGACGAGAGCA CCGACGAGAATGTCATGCTTCTGACTAGCGACGCCCCTGAATACAAGCCTTGGGC TCTGGTCATACAGGATAGCAACGGTGAGAACAAGATTAAGATGCTCTAATGGGT CCAGGCATCAAATAAAACGAAAGGCTCAGTCGAAAGACTGGGCCTTTCGTTTTA TCTGTTGTTTGTCGGTGAACGCTCTCTTCCTTGCAAAATATACAGGGGATTATATA TAATGGAAAACAAGTCTGACCAGACACGTCCCGTGTTTTAGAGCTAGAAATAGC AAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGTCGGT GCTTTTTTTGCGGCCGCGTGATCTAATAAACPart 1 of Staphylococcus hominis pXS0034 plasmid: (SEQ ID NO: 2):CGTGATGGTAACTTCAGCATGCGTTTTAGCGTTTATTTCGTTTAGTTATCGGCAT AATCGTTAAAACAGGCGTTATCGTAGCGTAAAAGCCCTTGAGCGTAGCGTGGCT TTGCAGCGAAGATGTTGTCTGTTAGATTATGAAAGCCGATGACTGAATGAAATA ATAAGCGCAGCGCCCTTCTATTTCGGTTGGAGGAGGCTCAAGGGAGTATGAGG GAATGAAATTCCCTCATGGGTTTGATTTTAAAAATTGCTTGCAATTTTGCCGAGC GGTAGCGCTGGAAAATTTTTGAAAAAAATTTGGAATTTGGAAAAAAATGGGGG GAAAGGAAGCGAATTTTGCTTCCGTACTACGACCCCCCATTAAGTGCCGAGTGC CAATTTTTGTGCCAAAAACGCTCTATCCCAACTGGCTCAAGGGTTTAAGGGGTT TTTCAATCGCCAACGAATCGCCAACGTTTTCGCCAACGTTTTTTATAAATCTATA TTTAAGTAGCTTTATTGTTGTTTTTATGATTACAAAGTGATACACTAACTTTATA AAATTATTTGATTGGAGTTTTTTAAATGGTGATTTCAGAATCGAAAAAAAGAGT TATGATTTCTCTGACAAAAGAGCAAGATAAAAAATTAACAGATATGGCGAAAC AAAAAGGTTTTTCAAAATCTGCGGTTGCGGCGTTAGCTATAGAAGAATATGCAA GAAAGGAATCAGAACAAAAAAAATAAGCGAAAGCTCGCGTTTTTAGAAGGATA CGAGTTTTCGCTACTTGTTTTTGATAAGGTAATTATATCATGGCTATTAAAAATA CTAAAGCTAGAAATTTTGGATTTTTATTATATCCTGACTCAATTCCTAATGATTG GAAAGAAAAATTAGAGAGTTTGGGCGTATCTATGGCTGTCAGTCCTTTACACGA TATGGACGAAAAAAAAGATAAAGATACATGGAATAATAGTAATATTATACAAAAtty. Dkt. No. 135523-0115ATGGAAAGCACTATAAAAAACCACACTATCACGTTATATATATTGCACGAAATC CTGTAACAATAGAAAGCGTTAGGAACAAGATTAAGCGAAAATTGGGGAATAGT TCAGTTGCTCATGTTGAGATACTTGATTATATCAAAGGTTCATATGAATATTTGA CTCATGAATCAAAGGACGCTATTGCTAAGAATAAACATATATACGACAAAAAA GATATTTTGAACATTAATGATTTTGATATTGACCGCTATATAACACTTGATGAAA GCCAAAAAAGAGAATTGAAGAATTTACTTTTAGATATAGTGGATGACTATAATT TGGTAAATACAAAAGATTTAATGGCTTTTATTCGCCTTAGGGGAGCGGAGTTTG GAATTTTAAATACGAATGATGTAAAAGATATTGTTTCAACAAACTCTAGCGCCTTTAGATTATGGTTTGAGGGCAATTATCAGTGTGGATATAGAGCAAGTTATGCAA AGGTTCTTGATGCTGAAACGGGGGAAATAAAATGACAAACAAAGAAAAAGAGT TATTTGCTGAAAATGAGGAATTAAAAAAAGAAATTAAGGACTTAAAAGAGCGT ATTGAAAGATACAGAGAAATGGAAGTTGAATTAAGTACAACAATAGATTTATT GAGAGGAGGGATTATTGAATAAATAAAAGCCCCCTGACGAAAGTCGAAGGGGG TTTTTATTTTGGTTTGATGTTGCGATTAATAGCAATACATTCTATAATAGAAGGT ATGGAGGATGTTATATAATGAGACAGAATTATGATGATCATATGTCAACTAACG GGGCAGGTTAGTGACATTAGAAAACCGACTGTAAAAAGTACAGTCGGCATTATCTCATATTATAAAAGCCAGTCATTAGGCCTATCTGACAATTCCTGAATAGAGTT CATAAACAATCCTGCATGATAACCATCACAAACAGAATGATGTACCTGTAAAG ATAGCGGTAAATATATTGAATTACCTTTATTAATGAATTTTCCTGCTGTAATAATGGGTAGAAGGTAATTACTATTATTATTGATATTTAAGTTAAACCCAGTAAATGA AGTCCATGGAATAATAGAAAGAGAAAAAGCATTTTCAGGTATAGGTGTTTTGG GAAACAATTTCCCCGAACCATTATATTTCTCTACATCAGAAAGGTATAAATCAT AAAACTCTTTGAAGTCATTCTTTACAGGAGTCCAAATGCCAGAGAATGTTTTAG ATACACCATCAAAAATTGTATAAAGTGGCTCTAACTTATCCCAATAACCTAACT CTCCGTCGCTATTGTAACCAGTTCTAAAAGCTGTATTTGAGTTTATCACCCTTGT CACTAAGAAAATAAATGCAGGGTAAAATTTATATCCTTCTTGTTTTATGTTTCGG TATAAAACACTAATATCAATTTCTGTGGTTATACTAAAAGTCGTTTGTTGGTTCAAATAATGATTAAATATCTCTTTTCTCTTCCAATTGTCTAAATCAATTTTATTAAA GTTCATGGGTTTCACTCTCCTTCTACATTTTTTAACCTAATAATGCCAAATACCG TTTGCCACCCCTCTCTTTGATAATTATAATATTGGCGAAATTCGCTTCTAAAGAT GAAACGCAATATTATATGCTTGCTTTATCGGCCGTATGTGATTATACCAGCCCCC TCACTACATGTCAAGAATAAACTGCCAAAGCATAATGGGATAATTAACCCTCPart 2 of Staphylococcus hominis pXS0034 plasmid: (SEQ ID NO: 3):TTTTCCATAGGCTCCGCCCCCCTGACAAGCATCACGAAATCTGACGCTCAAATCA GTGGTGGCGAAACCCGACAGGACTATAAAGATACCAGGCGTTTCCCCCTGGCGG CTCCCTCGTGCGCTCTCCTGTTCCTGCCTTTCGGTTTACCGGTGTCATTCCGCTGTT ATGGCCGCGTTTGTCTCATTCCACGCCTGACACTCAGTTCCGGGTAGGCAGTTCG CTCCAAGCTGGACTGTATGCACGAACCCCCCGTTCAGTCCGACCGCTGCGCCTTA TCCGGTAACTATCGTCTTGAGTCCAACCCGGAAAGACATGCAAAAGCACCACTG GCAGCAGCCACTGGTAATTGATTTAGAGGAGTTAGTCTTGAAGTCATGCGCCGGT TAAGGCTAAACTGAAAGGACAAGTTTTGGTGACTGCGCTCCTCCAAGCCAGTTACCTCGGTTCAAAGAGTTGGTAGCTCAGAGAACCTTCGAAAAACCGCCCTGCAAGG CGGTTTTTTCGTTTTCAGAGCAAGAGATTACGCGCAGACCAAAACGATCTCAAAtty. Dkt. No. 135523-0115Part 3 of Staphylococcus hominis pXS0034 plasmid: (SEQ ID NO: 4):TCTCCTTTTTCGCTTCTTTATTCCAATTGCTTTATTGACGTTGAGCCTCGGAACCGTCGACTTTATTTGGATCCCCTCGAGTTCATGAAAAACTAAAAAAAATATTGAAACTCTATCATTGATAGAGTATAATTAACAAGGAGGAATAAAAAPart 4 of Staphylococcus hominis pXS0034 plasmid: (SEQ ID NO: 5):ATGAGCTCAGAGACTGGCCCAGTGGCTGTGGACCCCACATTGAGGCGGCGGATCGAGCCCCATGAGTTTGAGGTATTCTTCGATCCGAGAGAGCTCCGCAAGGAGACTTGCCTGCTTTACGAAATTAATTGGGGGGGCCGGCACTCCATTTGGCGACATACATCACAGAACACTAACAAGCACGTCGAAGTCAACTTCATCGAGAAGTTCACGACAGAAAGATATTTCTGTCCGAACACAAGGTGCAGCATTACCTGGTTTCTCAGCTGGAGCCCATGCGGCGAATGTAGTAGGGCCATCACTGAATTCCTGTCAAGGTATCCCCACGTCACTCTGTTTATTTACATCGCAAGGCTGTACCACCACGCTGACCCCCGCAATCGACAAGGCCTGCGGGATTTGATCTCTTCAGGTGTGACTATCCAAATTATGACTGAGCAGGAGTCAGGATACTGCTGGAGAAACTTTGTGAATTATAGCCCGAGTAATGAAGCCCACTGGCCTAGGTATCCCCATCTGTGGGTACGACTGTACGTTCTTGAACTGTACTGCATCATACTGGGCCTGCCTCCTTGTCTCAACATTCTGAGAAGGAAGCAGCCACAGCTGACATTCTTTACCATCGCTCTTCAGTCTTGTCATTACCAGCGACTGCCCCCACACATTCTCTGGGCCACCGGGTTGAAATCTGGTGGTTCTTCTGGTGGTTCTAGCGGCAGCGAGACTCCCGGGACCTCAGAGTCCGCCACACCCGAAAGTTCTGGTGGTTCTTCTGGTGGTTCTGATAAAAAGTATTCTATTGGTTTAGCCATCGGCACTAATTCCGTTGGATGGGCTGTCATAACCGATGAATACAAAGTACCTTCAAAGAAATTTAAGGTGTTGGGGAACACAGACCGTCATTCGATTAAAAAGAATCTTATCGGTGCCCTCCTATTCGATAGTGGCGAAACGGCAGAGGCGACTCGCCTGAAACGAACCGCTCGGAGAAGGTATACACGTCGCAAGAACCGAATATGTTACTTACAAGAAATTTTTAGCAATGAGATGGCCAAAGTTGACGATTCTTTCTTTCACCGTTTGGAAGAGTCCTTCCTTGTCGAAGAGGACAAGAAACATGAACGGCACCCCATCTTTGGAAACATAGTAGATGAGGTGGCATATCATGAAAAGTACCCAACGATTTATCACCTCAGAAAAAAGCTAGTTGACTCAACTGATAAAGCGGACCTGAGGTTAATCTACTTGGCTCTTGCCCATATGATAAAGTTCCGTGGGCACTTTCTCATTGAGGGTGATCTAAATCCGGACAACTCGGATGTCGACAAACTGTTCATCCAGTTAGTACAAACCTATAATCAGTTGTTTGAAGAGAACCCTATAAATGCAAGTGGCGTGGATGCGAAGGCTATTCTTAGCGCCCGCCTCTCTAAATCCCGACGGCTAGAAAACCTGATCGCACAATTACCCGGAGAGAAGAAAAATGGGTTGTTCGGTAACCTTATAGCGCTCTCACTAGGCCTGACACCAAATTTTAAGTCGAACTTCGACTTAGCTGAAGATGCCAAATTGCAGCTTAGTAAGGACACGTACGATGACGATCTCGACAATCTACTGGCACAAATTGGAGATCAGTATGCGGACTTATTTTTGGCTGCCAAAAACCTTAGCGATGCAATCCTCCTATCTGACATACTGAGAGTTAATACTGAGATTACCAAGGCGCCGTTATCCGCTTCAATGATCAAAAGGTACGATGAACATCACCAAGACTTGACACTTCTCAAGGCCCTAGTCCGTCAGCAACTGCCTGAGAAATATAAGGAAATATTCTTTGATCAGTCGAAAAACGGGTACGCAGGTTATATTGACGGCGGAGCGAGTCAAGAGGAATTCTACAAGTTTATCAAACCCATATTAGAGAAGATGGATGGGACGGAAGAGTTGCTTGTAAAACTCAATCGCGAAGATCTACTGCGAAAGCAGCGGACTTTCGACAACGGTAGCATTCCACATCAAATCCACTTAGGCGAATTGCATGCTATACTTAGAAGGCAGGAGGATTTTTATCCGTTCAtty. Dkt. No. 135523-0115CTCAAAGACAATCGTGAAAAGATTGAGAAAATCCTAACCTTTCGCATACCTTACTATGTGGGACCCCTGGCCCGAGGGAACTCTCGGTTCGCATGGATGACAAGAAAGTCCGAAGAAACGATTACTCCATGGAATTTTGAGGAAGTTGTCGATAAAGGTGCGTCAGCTCAATCGTTCATCGAGAGGATGACCAACTTTGACAAGAATTTACCGAACGAAAAAGTATTGCCTAAGCACAGTTTACTTTACGAGTATTTCACAGTGTACAATGAACTCACGAAAGTTAAGTATGTCACTGAGGGCATGCGTAAACCCGCCTTTCTAAGCGGAGAACAGAAGAAAGCAATAGTAGATCTGTTATTCAAGACCAACCGCAAAGTGACAGTTAAGCAATTGAAAGAGGACTACTTTAAGAAAATTGAATGCTTCGATTCTGTCGAGATCTCCGGGGTAGAAGATCGATTTAATGCGTCACTTGGTACGTATCATGACCTCCTAAAGATAATTAAAGATAAGGACTTCCTGGATAACGAAGAGAATGAAGATATCTTAGAAGATATAGTGTTGACTCTTACCCTCTTTGAAGATCGGGAAATGATTGAGGAAAGACTAAAAACATACGCTCACCTGTTCGACGATAAGGTTATGAAACAGTTAAAGAGGCGTCGCTATACGGGCTGGGGACGATTGTCGCGGAAACTTATCAACGGGATAAGAGACAAGCAAAGTGGTAAAACTATTCTCGATTTTCTAAAGAGCGACGGCTTCGCCAATAGGAACTTTATGCAGCTGATCCATGATGACTCTTTAACCTTCAAAGAGGATATACAAAAGGCACAGGTTTCCGGACAAGGGGACTCATTGCACGAACATATTGCGAATCTTGCTGGTTCGCCAGCCATCAAAAAGGGCATACTCCAGACAGTCAAAGTAGTGGATGAGCTAGTTAAGGTCATGGGACGTCACAAACCGGAAAACATTGTAATCGAGATGGCACGCGAAAATCAAACGACTCAGAAGGGGCAAAAAAACAGTCGAGAGCGGATGAAGAGAATAGAAGAGGGTATTAAAGAACTGGGCAGCCAGATCTTAAAGGAGCATCCTGTGGAAAATACCCAATTGCAGAACGAGAAACTTTACCTCTATTACCTACAAAATGGAAGGGACATGTATGTTGATCAGGAACTGGACATAAACCGTTTATCTGATTACGACGTCGATCACATTGTACCCCAATCCTTTTTGAAGGACGATTCAATCGACAATAAAGTGCTTACACGCTCGGATAAGAACCGAGGGAAAAGTGACAATGTTCCAAGCGAGGAAGTCGTAAAGAAAATGAAGAACTATTGGCGGCAGCTCCTAAATGCGAAACTGATAACGCAAAGAAAGTTCGATAACTTAACTAAAGCTGAGAGGGGTGGCTTGTCTGAACTTGACAAGGCCGGATTTATTAAACGTCAGCTCGTGGAAACCCGCCAAATCACAAAGCATGTTGCACAGATACTAGATTCCCGAATGAATACGAAATACGACGAGAACGATAAGCTGATTCGGGAAGTCAAAGTAATCACTTTAAAGTCAAAATTGGTGTCGGACTTCAGAAAGGATTTTCAATTCTATAAAGTTAGGGAGATAAATAACTACCACCATGCGCACGACGCTTATCTTAATGCCGTCGTAGGGACCGCACTCATTAAGAAATACCCGAAGCTAGAAAGTGAGTTTGTGTATGGTGATTACAAAGTTTATGACGTCCGTAAGATGATCGCGAAAAGCGAACAGGAGATAGGCAAGGCTACAGCCAAATACTTCTTTTATTCTAACATTATGAATTTCTTTAAGACGGAAATCACTCTGGCAAACGGAGAGATACGCAAACGACCTTTAATTGAAACCAATGGGGAGACAGGTGAAATCGTATGGGATAAGGGCCGGGACTTCGCGACGGTGAGAAAAGTTTTGTCCATGCCCCAAGTCAACATAGTAAAGAAAACTGAGGTGCAGACCGGAGGGTTTTCAAAGGAATCGATTCTTCCAAAAAGGAATAGTGATAAGCTCATCGCTCGTAAAAAGGACTGGGACCCGAAAAAGTACGGTGGCTTCGATAGCCCTACAGTTGCCTATTCTGTCCTAGTAGTGGCAAAAGTTGAGAAGGGAAAATCCAAGAAACTGAAGTCAGTCAAAGAATTATTGGGGATAACGATTATGGAGCGCTCGTCTTTTGAAAAGAACCCCATCGACTTCCTTGAGGCGAAAGGTTACAAGGAAGTAAAAAAGGATCTCATAATTAAACTACCAAAGTATAGTCTGTTTGAGTTAGAAAATGGCCGAAAACGGATGTTGGCTAGCGCCGGAGAGCTTCAAAAGGGGAACGAACTCGCACTACCGTCTAAATACGTGAATTTCCTGTATTTAGCGTCCCATTACGAGAAGTTGAAAGGTTCACCTGAAGATAACGAACAGAAGCAACTTTTTGTTGAGCAGCACAAACATTATCTCGACGAAATCATAGAGCAAATTTCGGAATTCAGTAAGAGAGTCATCCTAGCTAtty. Dkt. No. 135523-0115GATGCCAATCTGGACAAAGTATTAAGCGCATACAACAAGCACAGGGATAAACCCATACGTGAGCAGGCGGAAAATATTATCCATTTGTTTACTCTTACCAACCTCGGCG CTCCAGCCGCATTCAAGTATTTTGACACAACGATAGATCGCAAACGATACACTTC TACCAAGGAGGTGCTAGACGCGACACTGATTCACCAATCCATCACGGGATTATATGAAACTCGGATAGATTTGTCACAGCTTGGGGGTGACTCTGGTGGTTCTGGAGGATCTGGTGGTTCTACTAATCTGTCAGATATTATTGAAAAGGAGACTGGTAAGCAACTGGTTATCCAGGAATCCATCCTCATGCTCCCAGAGGAGGTGGAAGAAGTCATTGGGAACAAGCCGGAAAGCGATATACTCGTGCACACCGCCTACGACGAGAGCACCG ACGAGAATGTCATGCTTCTGACTAGCGACGCCCCTGAATACAAGCCTTGGGCTCT GGTCATACAGGATAGCAACGGTGAGAACAAGATTAAGATGCTCTAATPart 5 of Staphylococcus hominis pXS0034 plasmid: (SEQ ID NO: 6):GGGTCCAGGCATCAAATAAAACGAAAGGCTCAGTCGAAAGACTGGGCCTTTCGT TTTATCTGTTGTTTGTCGGTGAACGCTCTCTTCCTTGCAAAATATACAGGGGATTA TATATAATGGAAAACAAGPart 6 of Staphylococcus hominis pXS0034 plasmid: (SEQ ID NO: 7):TCTGACCAGACACGTCCCGTGTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGG CTAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGTCGGTGCTTTTTTTGCGGCC GCGTGATCTAAT27F (forward primer) for amplifying the 16S of the transformant (SEQ ID NO: 8):GAGTTTGATYMTGGCTCAG1492R (reverse primer) for amplifying the 16S of the transformant (SEQ ID NO: 9):GYTACCTTGTTACGACTT1492R (reverse primer) for amplifying the 16S of the transformant (SEQ ID NO: 10):GCTACCTTGTTACGACTT1492R (reverse primer) for amplifying the 16S of the transformant (SEQ ID NO: 11):GTTACCTTGTTACGACTT patB004 (path forward) primer to verify introduction of premature stop codon at W 190 tryptophan amino acid site of patB (SEQ ID NO: 12):GGTGTGACATACGTTATTGATGGAtty. Dkt. No. 135523-0115 patB002 (patB reverse) primer to verify introduction of premature stop codon at W 190 tryptophan amino acid site of patB (SEQ ID NO: 13):GGTGTATGACCTATGTGTGNucleic acid sequence of patB (SEQ ID NO: 14):GTGAATTACAATTTTGATGAAATCATTGATCGTCGTTATACAAATGCAATGAAT GTGGAAGGCTATAAAGGCTATCTTTTTGGTGATGCAGATACGTCAGATTTAAAA GATAATGATGAACTGATTCGTATGTGGGTAGCAGATATGGACTTTGGTACACCA GAAGTCGTATTAAATGCGATTCGTGAACGTTTAAATAAGAAAATTTTAGGTTAC ACAAACGTCTTCGGTTCTGAATATTATGAAGCTTTTGTATCTTGGACAAAGAAA CGTTATGGTTTCACTTTCTCTCAAGAGCACCTCGTATTCTCTCACGGTATTGTAG CTGGGTTAATTGAACTTGTAGGTTATATTTGTGATAAGGATGATAAAGCATTAA TTGTTACGCCGAGTTATGGACCATTTAAAATGGCATGTGATAAAAATCATATTT CAACAGTATATTCTCCACTAATCAATCATCATGGATATTATGAGATTGATTTTGA TGATGTACGTAAAAAAGTAGAAACAGAAAATATTAAATTATGCATTTTTGCGAA TCCACATAATCCAACGGGACGTGTCTGGTCAGAAGAAGAGTTAGCCACATTAG GTCAAATTATGAAAGAGAATGATGTATGGCTCATTTCAGATGAAATCCATTGTG ATATCAAACGTTCAGGTCAATCACATATCCCATTTGCCAAAGCTGTCCCGGATT ACGACAAAATCATTACAACTATGTCTCAAAGTAAAGCCTTCAACATTGCAGGTT TAATGTTTTCAAATATCATTATTCAAAATGAATCACTTTTAAAAACTTGGAACAC ACATCACTTTGGTACGGAAAATCCTTTAAGTGTGGTTGCCACTCAAGCTGCTTAT GAAAAAGGCGAAGGTTGGTTACAAGCAATGAATCATTATTTAGATGATAACTTT AACTATTTAGCAGATTTCCTAGAAAAAGAACTACCACATGCAGAGTTTAAAATT CCAGAGGCAACGTATTTAGCTTGGGTAGATTTAAGCTATTATATTAAAGAAAAA GATATTGATGAATCAATGGCGAAATTCTTTATTAAAAACGCTGGTGTTATCATT GAAGGCGCAGAGCAATTTGTTCATAACGCAGAGGGGCATATTAGAATTAATATT GCAGTTCCTAGAGAAGTAATGAAAAAGGGACTACAAAAAATAAAAGCGGCTTT AGTCTAANucleic acid sequence of mutated patB with introduction of an exemplary stop codon at the W190 codon shown in bold and underlined text (SEQ ID NO: 15):GTGAATTACAATTTTGATGAAATCATTGATCGTCGTTATACAAATGCAATGAAT GTGGAAGGCTATAAAGGCTATCTTTTTGGTGATGCAGATACGTCAGATTTAAAA GATAATGATGAACTGATTCGTATGTGGGTAGCAGATATGGACTTTGGTACACCA GAAGTCGTATTAAATGCGATTCGTGAACGTTTAAATAAGAAAATTTTAGGTTAC ACAAACGTCTTCGGTTCTGAATATTATGAAGCTTTTGTATCTTGGACAAAGAAA CGTTATGGTTTCACTTTCTCTCAAGAGCACCTCGTATTCTCTCACGGTATTGTAG CTGGGTTAATTGAACTTGTAGGTTATATTTGTGATAAGGATGATAAAGCATTAA TTGTTACGCCGAGTTATGGACCATTTAAAATGGCATGTGATAAAAATCATATTT CAACAGTATATTCTCCACTAATCAATCATCATGGATATTATGAGATTGATTTTGA TGATGTACGTAAAAAAGTAGAAACAGAAAATATTAAATTATGCATTTTTGCGAA TCCACATAATCCAACGGGACGTGTCTAATCAGAAGAAGAGTTAGCCACATTAGAtty. Dkt. No. 135523-0115GTCAAATTATGAAAGAGAATGATGTATGGCTCATTTCAGATGAAATCCATTGTGATATCAAACGTTCAGGTCAATCACATATCCCATTTGCCAAAGCTGTCCCGGATTACGACAAAATCATTACAACTATGTCTCAAAGTAAAGCCTTCAACATTGCAGGTTTAATGTTTTCAAATATCATTATTCAAAATGAATCACTTTTAAAAACTTGGAACACACATCACTTTGGTACGGAAAATCCTTTAAGTGTGGTTGCCACTCAAGCTGCTTATGAAAAAGGCGAAGGTTGGTTACAAGCAATGAATCATTATTTAGATGATAACTTTAACTATTTAGCAGATTTCCTAGAAAAAGAACTACCACATGCAGAGTTTAAAATTCCAGAGGCAACGTATTTAGCTTGGGTAGATTTAAGCTATTATATTAAAGAAAAAGATATTGATGAATCAATGGCGAAATTCTTTATTAAAAACGCTGGTGTTATCATTGAAGGCGCAGAGCAATTTGTTCATAACGCAGAGGGGCATATTAGAATTAATATTGCAGTTCCTAGAGAAGTAATGAAAAAGGGACTACAAAAAATAAAAGCGGCTTTAGTCTAAStaphylococcus hominis pAB00038 plasmid: Eight part cytosine base editor plasmid nucleotide sequence (SEQ ID NO: 16):CGTGATGGTAACTTCAGCTTGCGTTTTAGCGTTTATTTCGTTTAGTTATCGGCATAATCGTTAAAACAGGCGTTATCGTAGCGTAAAAGCCCTTGAGCGTAGCGTGGCTTTGCAGCGAAGATGTTGTCTGTTAGATTATGAAAGCCGATGACTGAATGAAATAATAAGCGCAGCGCCCTTCTATTTCGGTTGGAGGAGGCTCAAGGGAGTATGAGGGAATGAAATTCCCTCATGGGTTTGATTTTAAAAATTGCTTGCAATTTTGCCGAGCGGTAGCGCTGGAAAATTTTTGAAAAAAATTTGGAATTTGGAAAAAAATGGGGGGAAAGGAAGCGAATTTTGCTTCCGTACTACGACCCCCCATTAAGTGCCGAGTGCCAATTTTTGTGCCAAAAACGCTCTATCCCAACTGGCTCAAGGGTTTAAGGGGTTTTTCAATCGCCAACGAATCGCCAACGTTTTCGCCAACGTTTTTTATAAATCTATATTTAAGTAGCTTTATTGTTGTTTTTATGATTACAAAGTGATACACTAACTTTATAAAATTATTTGATTGGAGTTTTTTAAATGGTGATTTCAGAATCGAAAAAAAGAGTTATGATTTCTCTGACAAAAGAGCAAGATAAAAAATTAACAGATATGGCGAAACAAAAAGGTTTTTCAAAATCTGCGGTTGCGGCGTTAGCTATAGAAGAATATGCAAGAAAGGAATCAGAACAAAAAAAATAAGCGAAAGCTCGCGTTTTTAGAAGGATACGAGTTTTCGCTACTTGTTTTTGATAAGGTAATTATATCATGGCTATTAAAAATACTAAAGCTAGAAATTTTGGATTTTTATTATATCCTGACTCAATTCCTAATGATTGGAAAGAAAAATTAGAGAGTTTGGGCGTATCTATGGCTGTCAGTCCTTTACACGATATGGACGAAAAAAAAGATAAAGATACATGGAATAATAGTAATATTATACAAAATGGAAAGCACTATAAAAAACCACACTATCACGTTATATATATTGCACGAAATCCTGTAACAATAGAAAGCGTTAGGAACAAGATTAAGCGAAAATTGGGGAATAGTTCAGTTGCTCATGTTGAGATACTTGATTATATCAAAGGTTCATATGAATATTTGACTCATGAATCAAAGGACGCTATTGCTAAGAATAAACATATATACGACAAAAAAGATATTTTGAACATTAATGATTTTGATATTGACCGCTATATAACACTTGATGAAAGCCAAAAAAGAGAATTGAAGAATTTACTTTTAGATATAGTGGACGACTATAATTTGGTAAATACAAAAGATTTAATGGCTTTTATTCGCCTTAGGGGAGCGGAGTTTGGAATTTTAAATACGAATGATGTAAAAGATATTGTTTCAACAAACTCTAGCGCCTTTAGATTATGGTTTGAGGGCAATTATCAGTGTGGATATAGAGCAAGTTATGCAAAGGTTCTTGATGCTGAAACGGGGGAAATAAAATGACAAACAAAGAAAAAGAGTTATTAtty. Dkt. No. 135523-0115TGCTGAAAATGAGGAATTAAAAAAAGAAATTAAGGACTTAAAAGAGCGTATTGAAAGATACAGAGAAATGGAAGTTGAATTAAGTACAACAATAGATTTATTGAGAGGAGGGATTATTGAATAAATAAAAGCCCCCTGACGAAAGTCGAAGGGGGTTTTTATTTTGGTTTGATGTTGCGATTAATAGCAATACATTCTATAATAGAAGGTATGGACGATGTTATATAATGAGACAGAATTATGATGAGCATATGTCAACTAACGGGGCAGGTTAGTGACATTAGAAAACCGACTGTAAAAAGTACAGTCGGCATTATCTCATATTATAAAAGCCAGTCATTAGGCCTATCTGACAATTCCTGAATAGAGTTCATAAACAATCCTGCATGATAACCATCACAAACAGAATGATGTACCTGTAAAGATAGCGGTAAATATATTGAATTACCTTTATTAATGAATTTTCCTGCTGTAATAATGGGTAGAAGGTAATTACTATTATTATTGATATTTAAGTTAAACCCAGTAAATGAAGTCCATGGAATAATAGAAAGAGAAAAAGCATTTTCAGGTATAGGTGTTTTGGGAAACAATTTCCCCGAACCATTATATTTCTCTACATCAGAAAGGTATAAATCATAAAACTCTTTGAAGTCATTCTTTACAGGAGTCCAAATGCCAGAGAATGTTTTAGATACACCATCAAAAATTGTATAAAGTGGCTCTAACTTATCCCAATAACCTAACTCTCCGTCGCTATTGTAACCAGTTCTAAAAGCTGTATTTGAGTTTATCACCCTTGTCACTAAGAAAATAAATGCAGGGTAAAATTTATATCCTTCTTGTTTTATGTTTCGGTATAAAACACTAATATCAATTTCTGTGGTTATACTAAAAGTCGTTTGTTGGTTCAAATAATGATTAAATATCTCTTTTCTCTTCCAATTGTCTAAATCAATTTTATTAAAGTTCATGGGTTTCACTCTCCTTCTACATTTTTTAACCTAATAATGCCAAATACCGTTTGCCACCCCTCTCTTTGATAATTATAATATTGGCGAAATTCGCTTCTAAAGATGAAACGCAATATTATATGCTTGCTTTATCGGCCGTATGTGATTATACCAGCCCCCTCACTACATGTCAAGAATAAACTGCCAAAGCATAATGGGATAATTAACCCTCACGACTGCGGAACTGACTAAAGTAGTGAGTTATACACAGGGCTGGGATCTATTCTTTTTATCTTTTTTTATTCTTTCTTTATTCTATAAATTATAACCACTTGAATATAAACAAAAAAAACACACAAAGGTCTAGCGGAATTTACAGAGGGTCTAGCAGAATTTACAAGTTTTCCAGCAAAGGTCTAGCAGAATTTACAGATACCCACAACTCAAAGGAAAAGGACTAGTAATTATCATTGACTAGCCCATCTCAATTGGTATAGTGATTAAAATCACCTAGACCAATTGAGATGTATGTCTGAATTAGTTGTTTTCAAAGCAAATGAACTAGCGATTAGTCGCTATGACTTAACGGAGCATGAAACCAAGCTAATTTTATGCTGTGTGGCACTACTCAACCCCACGATTGAAAACCCTACAAGGAAAGAACGGACGGTATCGTTCACTTATAACCAATACGCTCAGATGATGAACATCAGTAGGGAAAATGCTTATGGTGTATTAGCTAAAGCAACCAGAGAGCTGATGACGAGAACTGTGGAAATCAGGAATCCTTTGGTTAAAGGCTTTGAGATTTTCCAGTGGACAAACTATGCCAAGTTCTCAAGCGAAAAATTAGAATTAGTTTTTAGTGAAGAGATATTGCCTTATCTTTTCCAGTTAAAAAAATTCATAAAATATAATCTGGAACATGTTAAGTCTTTTGAAAACAAATACTCTATGAGGATTTATGAGTGGTTATTAAAAGAACTAACACAAAAGAAAACTCACAAGGCAAATATAGAGATTAGCCTTGATGAATTTAAGTTCATGTTAATGCTTGAAAATAACTACCATGAGTTTAAAAGGCTTAACCAATGGGTTTTGAAACCAATAAGTAAAGATTTAAACACTTACAGCAATATGAAATTGGTGGTTGATAAGCGAGGCCGCCCGACTGATACGTTGATTTTCCAAGTTGAACTAGATAGACAAATGGACCTCGTAACCGAACTTGAGAACAACCAGATAAAAATGAATGGTGACAAAATACCAACAACCATTACATCAGATTCCTACCTACATAACGGACTAAGAAAAACACTACACGATGCTTTAACTGCAAAAATTCAGCTCACCAGTTTTGAAtty. Dkt. No. 135523-0115GGCAAAATTTTTGAGTGACATGCAAAGTAAGTATGACCTCAATGGTTCGTTCTCATGGCTCACGCAAAAACAACGAACCACACTAGAGAACATACTGGCTAAATACGGAAGGATCTGAGGTTCTTATGGCTCTTGTATCTATCAGTGAAGCATCAAGACTAACAAACAAAAGTAGAACAACTGTTCACCGTTACATATCAAAGGGAAAACTGTCGTAGTCTCCTTTTTCGCTTCTTTATTCCAATTGCTTTATTGACGTTGAGCCTCGGAACCGTCGACTTTGTTTGGATCCCCTCGAGTTCATGAAAAACTAAAAAAAATATTGAAACTCTATCATTGATAGAGTATAATTAACAAGGAGGAATAAAAAATGAGCTCAGAGACTGGCCCAGTGGCTGTGGACCCCACATTGAGGCGGCGGATCGAGCCCCATGAGTTTGAGGTATTCTTCGACCCGAGAGAGCTCCGCAAGGAGACTTGCCTGCTTTACGAAATTAATTGGGGGGGCCGGCACTCCATTTGGCGACATACATCACAGAACACTAACAAACACGTCGAAGTCAACTTCATCGAGAAGTTCACGACAGAAAGATATTTCTGTCCGAACACAAGGTGTAGCATTACCTGGTTTCTCAGCTGGAGCCCATGTGGCGAATGTAGTAGGGCCATCACTGAATTCCTGTCAAGGTATCCCCACGTCACTCTGTTTATTTACATCGCAAGGCTGTACCACCACGCTGACCCCCGCAATCGACAAGGCCTGCGGGATTTGATCTCTTCAGGTGTGACTATCCAAATTATGACTGAGCAGGAGTCAGGATACTGCTGGAGAAACTTTGTGAATTATAGCCCGAGTAATGAAGCCCACTGGCCTAGGTATCCCCATCTGTGGGTACGACTGTACGTTCTTGAACTGTACTGCATCATACTGGGCCTGCCTCCTTGTCTCAACATTCTGAGAAGGAAGCAGCCACAGCTGACATTCTTTACCATCGCTCTTCAGTCTTGTCATTACCAGCGACTGCCCCCACACATTCTCTGGGCCACCGGGTTGAAATCTGGTGGTTCTTCTGGTGGTTCTTCAGGCAGCGAGACTCCCGGGACCTCAGAGTCCGCCACACCCGAAAGTAGTGGTGGTTCTTCTGGTGGTTCTGATAAAAAGTATTCTATTGGTTTAGCCATCGGCACTAATTCCGTTGGTTGGGCTGTCATAACCGATGAATACAAAGTACCTTCAAAGAAATTTAAGGTGTTGGGGAACACAGACCGTCATTCGATTAAAAAGAATCTTATCGGTGCCCTCCTATTCGATAGTGGCGAAACGGCAGAGGCGACTCGCCTGAAACGAACCGCTCGGAGAAGGTATACACGTCGCAAGAACCGAATATGTTACTTACAAGAAATTTTTAGCAATGAGATGGCCAAAGTTGACGATTCTTTCTTTCACCGTTTGGAAGAGTCCTTCCTTGTCGAAGAGGACAAGAAACATGAACGGCACCCCATCTTTGGAAACATAGTAGATGAGGTGGCATATCATGAAAAGTACCCAACGATTTATCACCTCAGAAAAAAGCTAGTTGACTCAACTGATAAAGCGGACCTGAGGTTAATCTACTTGGCTCTTGCCCATATGATAAAGTTCCGTGGGCACTTTCTCATTGAGGGTGACCTAAATCCGGACAACTCGGACGTCGACAAACTGTTCATTCAGTTAGTACAAACCTATAATCAGTTGTTTGAAGAGAACCCTATAAATGCAAGTGGCGTGGACGCGAAGGCTATTCTTAGCGCCCGCCTCTCTAAATCCCGACGGCTAGAAAACCTGATCGCACAATTACCCGGAGAGAAGAAAAATGGGTTGTTCGGTAACCTTATAGCGCTCTCACTAGGCCTGACACCAAATTTTAAGTCGAACTTCGACTTAGCTGAAGATGCCAAACTTCAGCTTAGTAAGGACACGTACGATGACGACCTCGACAATCTACTGGCACAAATTGGAGATCAGTACGCGGACTTATTTTTGGCTGCCAAAAACCTTAGCGATGCAATCCTCCTATCTGACATACTGAGAGTTAATACTGAGATTACCAAGGCGCCGTTATCCGCTTCAATGATAAAAAGGTACGATGAACATCACCAAGACTTGACACTTCTCAAGGCCCTAGTCCGTCAGCAACTGCCTGAGAAATATAAGGAAATATTCTTTGACCAGTCGAAAAACGGGTACGCAGGTTATATTGACGGCGGAGCGAGTCAAGAGGAATTCTACAAGTTTATCAtty. Dkt. No. 135523-0115AAACCCATATTAGAGAAGATGGACGGGACGGAAGAGTTGCTTGTAAAACTCAATCGCGAAGATCTACTGCGAAAGCAGCGGACTTTCGACAACGGTAGCATTCCACATCAAATCCACTTAGGCGAATTGCATGCTATACTTAGAAGGCAGGAGGATTTTTATCCGTTCCTCAAAGACAATCGTGAAAAGATTGAGAAAATCCTAACCTTTCGTATACCTTACTATGTGGGACCCCTGGCCCGAGGGAACTCTCGGTTCGCTTGGATGACAAGAAAGTCCGAGGAAACGATTACTCCATGGAATTTTGAGGAAGTTGTCGATAAAGGTGCGTCAGCTCAATCGTTCATCGAGAGAATGACCAACTTTGACAAGAATTTACCGAACGAAAAAGTATTGCCTAAGCACAGTTTACTTTACGAGTATTTCACAGTGTACAATGAACTCACGAAAGTTAAGTATGTCACTGAGGGCATGAGAAAACCCGCCTTTCTAAGCGGAGAACAGAAGAAAGCAATAGTAGACCTGTTATTCAAGACCAACCGCAAAGTGACAGTTAAGCAATTGAAAGAGGACTACTTTAAGAAAATTGAATGCTTCGATTCTGTCGAGATCTCCGGGGTAGAAGACCGATTTAATGCTTCACTTGGTACGTATCATGACCTCCTAAAGATAATTAAAGATAAGGACTTCCTGGATAACGAAGAGAATGAAGATATCTTAGAAGATATAGTGTTGACTCTTACCCTCTTTGAAGACCGGGAAATGATTGAGGAAAGACTAAAAACATACGCTCACCTGTTCGACGATAAGGTTATGAAACAGTTAAAGAGGCGTCGCTATACGGGCTGGGGACGATTGTCGCGGAAACTTATCAACGGGATAAGAGACAAGCAAAGTGGTAAAACTATTCTCGATTTTCTAAAGAGCGACGGCTTCGCCAATAGGAACTTTATGCAACTGATTCATGATGACTCTTTAACCTTCAAAGAGGATATACAAAAGGCACAGGTTTCCGGACAAGGGGACTCATTGCACGAACATATTGCGAATCTTGCTGGTTCGCCAGCCATCAAAAAGGGCATACTCCAGACAGTCAAAGTAGTGGACGAGCTAGTTAAGGTCATGGGACGTCACAAACCGGAAAACATTGTAATCGAGATGGCACGCGAAAATCAAACGACTCAGAAGGGGCAAAAAAACAGTCGAGAGAGAATGAAGAGAATAGAGGAGGGTATTAAAGAACTGGGCAGCCAGATATTAAAGGAGCACCCTGTGGAAAATACCCAATTGCAGAACGAGAAACTTTACCTCTATTACCTACAAAATGGAAGGGACATGTATGTTGACCAGGAACTGGACATAAACCGTTTATCTGATTACGACGTCGACCACATTGTACCCCAATCCTTTTTGAAGGACGATTCAATCGACAATAAAGTGCTTACACGCTCGGATAAGAACCGAGGGAAAAGTGACAATGTTCCAAGCGAGGAAGTCGTAAAGAAAATGAAGAACTATTGGCGGCAGCTCCTAAACGCGAAACTGATAACGCAAAGAAAGTTCGATAACTTAACTAAAGCTGAGAGGGGTGGCTTGTCTGAACTTGACAAGGCCGGATTTATTAAACGTCAGCTCGTGGAAACCCGCCAAATCACAAAGCATGTTGCACAGATACTAGATTCCCGAATGAATACGAAATACGACGAAAACGATAAGCTGATTCGTGAAGTCAAAGTAATCACTTTAAAGTCAAAATTGGTGTCGGACTTCAGAAAGGATTTTCAATTCTATAAAGTTAGGGAGATAAATAACTACCACCACGCGCACGACGCTTATCTTAATGCCGTCGTAGGGACCGCACTCATTAAGAAATACCCGAAGCTAGAAAGTGAGTTTGTGTATGGTGATTACAAAGTTTATGACGTCCGTAAAATGATAGCGAAAAGCGAACAGGAGATAGGCAAGGCTACAGCCAAATACTTCTTTTATTCTAACATTATGAATTTCTTTAAGACGGAAATCACTCTGGCAAACGGAGAGATACGCAAACGACCTTTAATTGAAACCAATGGGGAGACAGGTGAAATCGTATGGGATAAGGGCCGGGACTTCGCGACGGTGAGAAAAGTTTTGTCCATGCCCCAAGTCAACATAGTAAAGAAAACTGAGGTGCAGACCGGAGGGTTTTCAAAGGAATCGATTCTTCCAAAAAGGAATAGTGATAAGCTCATCGCTCGTAAAAAGGACTGGGACCCGAAAAAGTACGGTGAtty. Dkt. No. 135523-0115GCTTCGATAGCCCTACAGTTGCCTATTCTGTCCTAGTAGTGGCAAAAGTTGAGAAG GGAAAATCCAAGAAACTGAAGTCAGTCAAAGAATTATTGGGCATTACGATTATGG AGCGCTCGTCTTTTGAAAAGAACCCCATCGACTTCCTTGAGGCGAAAGGTTACAA GGAAGTAAAAAAGGACCTCATAATTAAACTACCAAAGTATAGTCTGTTTGAGTTAG AAAATGGCCGAAAAAGAATGTTGGCTAGCGCCGGAGAGCTTCAAAAGGGGAACG AACTCGCACTACCGTCTAAATACGTGAATTTCCTGTATTTAGCGTCCCATTACGAGAAGTTGAAAGGTTCACCTGAAGATAACGAACAGAAGCAACTTTTTGTTGAACAACA CAAACATTATCTCGACGAAATCATAGAGCAAATTTCGGAATTCAGTAAGAGAGTCA TACTAGCTGATGCCAATCTGGACAAAGTATTAAGCGCTTACAACAAGCACAGGGAT AAACCCATACGTGAGCAGGCGGAAAATATTATCCATTTGTTTACTCTTACCAACCTC GGCGCTCCGGCCGCGTTCAAGTATTTTGACACAACGATAGATCGCAAACGATACA CTTCTACCAAGGAGGTGCTAGACGCGACACTGATTCACCAATCCATCACGGGATTATATGAAACTCGGATAGATTTGTCACAGCTTGGGGGTGACTCTGGGGGGTCTGGGG GTTCTGGGGGTTCTACTAATCTGTCAGATATTATTGAAAAGGAGACTGGTAAGCAA CTGGTTATCCAGGAATCCATACTCATGCTCCCAGAGGAGGTGGAAGAAGTCATTG GGAACAAGCCGGAAAGCGATATACTCGTGCACACCGCCTACGACGAGAGCACCG ACGAGAATGTCATGCTTCTGACTAGCGACGCCCCTGAATACAAGCCTTGGGCTCT GGTCATACAGGATAGCAACGGTGAGAACAAGATTAAGATGCTCTAACCAGGCATCAAATAAAACGAAAGGCTCAGTCGAAAGACTGGGCCTTTCGTTTTATCTGTTGTTTG TCGGTGAACGCTCTCTTCCTTGATTTAATTCTAAGATATTTGTATTTTTATATTGTAAT CGATTTCTGACCAGACACGTCCCGTGTTTTAGAGCTAGAAATAGCAAGTTAAAATA AGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGTCGGTGCTTTTTTTGCG GCCGCGTGATTTAATAAACPart 1 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 17):AAACCGTGATGGTAACTTCAGCTTGCGTTTTAGCGTTTATTTCGTTTAGTTATCGGC ATAATCGTTAAAACAGGCGTTATCGTAGCGTAAAAGCCCTTGAGCGTAGCGTGGCT TTGCAGCGAAGATGTTGTCTGTTAGATTATGAAAGCCGATGACTGAATGAAATAAT AAGCGCAGCGCCCTTCTATTTCGGTTGGAGGAGGCTCAAGGGAGTATGAGGGAAT GAAATTCCCTCATGGGTTTGATTTTAAAAATTGCTTGCAATTTTGCCGAGCGGTAG CGCTGGAAAATTTTTGAAAAAAATTTGGAATTTGGAAAAAAATGGGGGGAAAGGAAGCGAATTTTGCTTCCGTACTACGACCCCCCATTAAGTGCCGAGTGCCAATTTTT GTGCCAAAAACGCTCTATCCCAACTGGCTCAAGGGTTTAAGGGGTTTTTCAATCG CCAACGAATCGCCAACGTTTTCGCCAACGTTTTTTATAAATCTATATTTAAGTAGCT TTATTGTTGTTTTTATGATTACAAAGTGATACACTAACTTTATAAAATTATTTGATTG GAGTTTTTTAAATGGTGATTTCAGAATCGAAAAAAAGAGTTATGATTTCTCTGACA AAAGAGCAAGATAAAAAATTAACAGATATGGCGAAACAAAAAGGTTTTTCAAAATCTGCGGTTGCGGCGTTAGCTATAGAAGAATATGCAAGAAAGGAATCAGAACAAAA AAAATAAGCGAAAGCTCGCGTTTTTAGAAGGATACGAGTTTTCGCTACTTGTTTTT GATAAGGTAATTATATCATGGCTATTAAAAATACTAAAGCTAGAAATTTTGGATTTTTAtty. Dkt. No. 135523-0115ATTATATCCTGACTCAATTCCTAATGATTGGAAAGAAAAATTAGAGAGTTTGGGCGTATCTATGGCTGTCAGTCCTTTACACGATATGGACGAAAAAAAAGATAAAGATACATGGAATAATAGTAATATTATACAAAATGGAAAGCACTATAAAAAACCACACTATCACGTTATATATATTGCACGAAATCCTGTAACAATAGAAAGCGTTAGGAACAAGATTAAGCGAAAATTGGGGAATAGTTCAGTTGCTCATGTTGAGATACTTGATTATATCAAAGGTTCATATGAATATTTGACTCATGAATCAAAGGACGCTATTGCTAAGAATAAACATATATACGACAAAAAAGATATTTTGAACATTAATGATTTTGATATTGACCGCTATATAACACTTGATGAAAGCCAAAAAAGAGAATTGAAGAATTTACTTTTAGATATAGTGGACGACTATAATTTGGTAAATACAAAAGATTTAATGGCTTTTATTCGCCTTAGGGGAGCGGAGTTTGGAATTTTAAATACGAATGATGTAAAAGATATTGTTTCAACAAACTCTAGCGCCTTTAGATTATGGTTTGAGGGCAATTATCAGTGTGGATATAGAGCAAGTTATGCAAAGGTTCTTGATGCTGAAACGGGGGAAATAAAATGACAAACAAAGAAAAAGAGTTATTTGCTGAAAATGAGGAATTAAAAAAAGAAATTAAGGACTTAAAAGAGCGTATTGAAAGATACAGAGAAATGGAAGTTGAATTAAGTACAACAATAGATTTATTGAGAGGAGGGATTATTGAATAAATAAAAGCCCCCTGACGAAAGTCGAAGGGGGTTTTTATTTTGGTTTGATGTTGCGATTAATAGCAATACATTCTATAATAGAAGGTATGGACGATGTTATATAATGAGACAGAATTATGATGAGCATATGTCAACTAACGGGGCAGGTTAGTGACATTAGAAAACCGACTGTAAAAAGTACAGTCGGCATTATCTCATATTATAAAAGCCAGTCATTAGGCCTATCTGACAATTCCTGAATAGAGTTCATAAACAATCCTGCATGATAACCATCACAAACAGAATGATGTACCTGTAAAGATAGCGGTAAATATATTGAATTACCTTTATTAATGAATTTTCCTGCTGTAATAATGGGTAGAAGGTAATTACTATTATTATTGATATTTAAGTTAAACCCAGTAAATGAAGTCCATGGAATAATAGAAAGAGAAAAAGCATTTTCAGGTATAGGTGTTTTGGGAAACAATTTCCCCGAACCATTATATTTCTCTACATCAGAAAGGTATAAATCATAAAACTCTTTGAAGTCATTCTTTACAGGAGTCC AAATPart 2 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 18):GCCAGAGAATGTTTTAGATACACCATCAAAAATTGTATAAAGTGGCTCTAACTTATCCCAATAACCTAACTCTCCGTCGCTATTGTAACCAGTTCTAAAAGCTGTATTTGAGTTTATCACCCTTGTCACTAAGAAAATAAATGCAGGGTAAAATTTATATCCTTCTTGTTTTATGTTTCGGTATAAAACACTAATATCAATTTCTGTGGTTATACTAAAAGTCGTTTGTTGGTTCAAATAATGATTAAATATCTCTTTTCTCTTCCAATTGTCTAAATCAATTTTATTAAAGTTCATGGGTTTCACTCTCCTTCTACATTTTTTAACCTAATAATGCCAAATACCGTTTGCCACCCCTCTCTTTGATAATTATAATATTGGCGAAATTCGCTTCTAAAGATGAAACGCAATATTATATGCTTGCTTTATCGGCCGTATGTGATTATACCAGCCCCCTCACTACATGTCAAGAATAAACTGCCAAAGCATAATGGGATAATTAACCCTCPart 3 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 19):ACGACTGCGGAACTGACTAAAGTAGTGAGTTATACACAGGGCTGGGATCTATTCTTTTTATCTTTTTTTATTCTTTCTTTATTCTATAAATTATAACCACTTGAATATAAACAAAAtty. Dkt. No. 135523-0115AAAAACACACAAAGGTCTAGCGGAATTTACAGAGGGTCTAGCAGAATTTACAAGTTTTCCAGCAAAGGTCTAGCAGAATTTACAGATACCCACAACTCAAAGGAAAAGGACTAGTAATTATCATTGACTAGCCCATCTCAATTGGTATAGTGATTAAAATCACCTAGACCAATTGAGATGTATGTCTGAATTAGTTGTTTTCAAAGCAAATGAACTAGCGATTAGTCGCTATGACTTAACGGAGCATGAAACCAAGCTAATTTTATGCTGTGTGGCACTACTCAACCCCACGATTGAAAACCCTACAAGGAAAGAACGGACGGTATCGTTCACTTATAACCAATACGCTCAGATGATGAACATCAGTAGGGAAAATGCTTATGGTGTATTAGCTAAAGCAACCAGAGAGCTGATGACGAGAACTGTGGAAATCAGGAATCCTTTGGTTAAAGGCTTTGAGATTTTCCAGTGGACAAACTATGCCAAGTTCTCAAGCGAAAAATTAGAATTAGTTTTTAGTGAAGAGATATTGCCTTATCTTTTCCAGTTAAAAAAATTCATAAAATATAATCTGGAACATGTTAAGTCTTTTGAAAACAAATACTCTATGAGGATTTATGAGTGGTTATTAAAAGAACTAACACAAAAGAAAACTCACAAGGCAAATATAGAGATTAGCCTTGATGAATTTAAGTTCATGTTAATGCTTGAAAATAACTACCATGAGTTTAAAAGGCTTAACCAATGGGTTTTGAAACCAATAAGTAAAGATTTAAACACTTACAGCAATATGAAATTGGTGGTTGATAAGCGAGGCCGCCCGACTGATACGTTGATTTTCCAAGTTGAACTAGATAGACAAATGGACCTCGTAACCGAACTTGAGAACAACCAGATAAAAATGAATGGTGACAAAATACCAACAACCATTACATCAGATTCCTACCTACATAACGGACTAAGAAAAACACTACACGATGCTTTAACTGCAAAAATTCAGCTCACCAGTTTTGAGGCAAAATTTTTGAGTGACATGCAAAGTAAGTATGACCTCAATGGTTCGTTCTCATGGCTCACGCAAAAACAACGAACCACACTAGAGAACATACTGGCTAAATACGGAAGGATCTGAGGTTCTTATGGCTCTTGTATCTATCAGTGAAGCATCAAGACTAACAAACAAAAGTAGAACAACTGTTCACCGTTACATATCAAAGGGAAAACTG TCPart 4 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 20):GTAGTCTCCTTTTTCGCTTCTTTATTCCAATTGCTTTATTGACGTTGAGCCTCGGAACCGTCGACTTTGTTTGGATCCCCTCGAGTTCATGAAAAACTAAAAAAAATATTGAAACTCTATCATTGATAGAGTATAATTAACAAGGAGGAATAAAAPart 5 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 21):AATGAGCTCAGAGACTGGCCCAGTGGCTGTGGACCCCACATTGAGGCGGCGGATCGAGCCCCATGAGTTTGAGGTATTCTTCGACCCGAGAGAGCTCCGCAAGGAGACTTGCCTGCTTTACGAAATTAATTGGGGGGGCCGGCACTCCATTTGGCGACATACATCACAGAACACTAACAAACACGTCGAAGTCAACTTCATCGAGAAGTTCACGACAGAAAGATATTTCTGTCCGAACACAAGGTGTAGCATTACCTGGTTTCTCAGCTGGAGCCCATGTGGCGAATGTAGTAGGGCCATCACTGAATTCCTGTCAAGGTATCCCCACGTCACTCTGTTTATTTACATCGCAAGGCTGTACCACCACGCTGACCCCCGCAATCGACAAGGCCTGCGGGATTTGATCTCTTCAGGTGTGACTATCCAAATTATGACTGAGCAGGAGTCAGGATACTGCTGGAGAAACTTTGTGAATTATAGCCCGAGTAATGAAGCCCACTGGCCTAGGTATCCCCATCTGTGGGTACGACTGTACGTTCTTGAACTGTACTGCATCAtty. Dkt. No. 135523-0115ATACTGGGCCTGCCTCCTTGTCTCAACATTCTGAGAAGGAAGCAGCCACAGCTGA CATTCTTTACCATCGCTCTTCAGTCTTGTCATTACCAGCGACTGCCCCCACACATTC TCTGGGCCACCGGGTTGAAATCTGGTGGTTCTTCTGGTGGTTCTTCAGGCAGCGA GACTCCCGGGACCTCAGAGTCCGCCACACCCGAAAGTAGTGGTGGTTCTTCTGGT GGTTCTGATAAAAAGTATTCTATTGGTTTAGCCATCGGCACTAATTCCGTTGGTTGG GCTGTCATAACCGATGAATACAAAGTACCTTCAAAGAAATTTAAGGTGTTGGGGA ACACAGACCGTCATTCGATTAAAAAGAATCTTATCGGTGCCCTCCTATTCGATAGT GGCGAAACGGCAGAGGCGACTCGCCTGAAACGAACCGCTCGGAGAAGGTATACA CGTCGCAAGAACCGAATATGTTACTTACAAGAAATTTTTAGCAATGAGATGGCCAA AGTTGACGATTCTTTCTTTCACCGTTTGGAAGAGTCCTTCCTTGTCGAAGAGGACA AGAAACATGAACGGCACCCCATCTTTGGAAACATAGTAGATGAGGTGGCATATCAT GAAAAGTACCCAACGATTTATCACCTCAGAAAAAAGCTAGTTGACTCAACTGATA AAGCGGACCTGAGGTTAATCTACTTGGCTCTTGCCCATATGATAAAGTTCCGTGGG CACTTTCTCATTGAGGGTGACCTAAATCCGGACAACTCGGACGTCGACAAACTGT TCATTCAGTTAGTACAAACCTATAATCAGTTGTTTGAAGAGAACCCTATAAATGCA AGTGGCGTGGACGCGAAGGCTATTCTTAGCGCCCGCCTCTCTAAATCCCGACGGC TAGAAAACCTGATCGCACAATTACCCGGAGAGAAGAAAAATGGGTTGTTCGGTAA CCTTATAGCGCTCTCACTAGGCCTGACACCAAATTTTAAGTCGAACTTCGACTTAG CTGAAGATGCCAAACTTCAGCTTAGTAAGGACACGTACGATGACGACCTCGACAA TCTACTGGCACAAATTGGAGATCAGTACGCGGACTTATTTTTGGCTGCCAAAAACC TTAGCGATGCAATCCTCCTATCTGACATACTGAGAGTTAATACTGAGATTACCAAGG CGCCGTTATCCGCTTCAATGATAAAAAGGTACGATGAACATCACCAAGACTTGACA CTTCTCAAGGCCCTAGTCCGTCAGCAACTGCCTGAGAAATATAAGGAAATATTCTTTGACCAGTCGAAAAACPart 6 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 22):GGGTACGCAGGTTATATTGACGGCGGAGCGAGTCAAGAGGAATTCTACAAGTTTAT CAAACCCATATTAGAGAAGATGGACGGGACGGAAGAGTTGCTTGTAAAACTCAAT CGCGAAGATCTACTGCGAAAGCAGCGGACTTTCGACAACGGTAGCATTCCACATC AAATCCACTTAGGCGAATTGCATGCTATACTTAGAAGGCAGGAGGATTTTTATCCG TTCCTCAAAGACAATCGTGAAAAGATTGAGAAAATCCTAACCTTTCGTATACCTTA CTATGTGGGACCCCTGGCCCGAGGGAACTCTCGGTTCGCTTGGATGACAAGAAAG TCCGAGGAAACGATTACTCCATGGAATTTTGAGGAAGTTGTCGATAAAGGTGCGT CAGCTCAATCGTTCATCGAGAGAATGACCAACTTTGACAAGAATTTACCGAACGA AAAAGTATTGCCTAAGCACAGTTTACTTTACGAGTATTTCACAGTGTACAATGAAC TCACGAAAGTTAAGTATGTCACTGAGGGCATGAGAAAACCCGCCTTTCTAAGCGG AGAACAGAAGAAAGCAATAGTAGACCTGTTATTCAAGACCAACCGCAAAGTGAC AGTTAAGCAATTGAAAGAGGACTACTTTAAGAAAATTGAATGCTTCGATTCTGTCG AGATCTCCGGGGTAGAAGACCGATTTAATGCTTCACTTGGTACGTATCATGACCTC CTAAAGATAATTAAAGATAAGGACTTCCTGGATAACGAAGAGAATGAAGATATCTTAtty. Dkt. No. 135523-0115AGAAGATATAGTGTTGACTCTTACCCTCTTTGAAGACCGGGAAATGATTGAGGAA AGACTAAAAACATACGCTCACCTGTTCGACGATAAGGTTATGAAACAGTTAAAGAGGCGTCGCTATACGGGCTGGGGACGATTGTCGCGGAAACTTATCAACGGGATAAG AGACAAGCAAAGTGGTAAAACTATTCTCGATTTTCTAAAGAGCGACGGCTTCGCC AATAGGAACTTTATGCAACTGATTCATGATGACTCTTTAACCTTCAAAGAGGATATACAAAAGGCACAGGTTTCCGGACAAGGGGACTCATTGCACGAACATATTGCGAATC TTGCTGGTTCGCCAGCCATCAAAAAGGGCATACTCCAGACAGTCAAAGTAGTGGA CGAGCTAGTTAAGGTCATGGGACGTCACAAACCGGAAAACATTGTAATCGAGATG GCACGCGAAAATCAAACGACTCAGAAGGGGCAAAAAAACAGTCGAGAGAGAAT GAAGAGAATAGAGGAGGGTATTAAAGAACTGGGCAGCCAGATATTAAAGGAGCACCCTGTGGAAAATACCCAATTGCAGAACGAGAAACTTTACCTCTATTACCTACAAA ATGGAAGGGACATGTATGTTGACCAGGAACTGGACATAAACCGTTTATCTGATTACGACGTCGACCACATTGTACCCCAATCCTTTTTGAAGGACGATTCAATCGACAATAA AGTGCTTACACGCTCGGATAAGAACCGAGGGAAAAGTGACAATGTTCCAAGCGAGGAAGTCGTAAAGAAAATGAAGAACTATTGGCGGCAGCTCCTAAACGCGAAACT GATAACGCAAAGAAAGTTCGATAACTTAACTAAAGCTGAGAGGGGTGGCTTGTCTGAACTTGACAAGGCCGGATTTATTAAACGTCAGCTCGTGGAAACCCGCCAAATCA CAAAGCATGTTGCACAGATACTAGATTCCCGAATGAATACGAAATACGACGAAAA CGATAAGCTGATTCGPart 7 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 23):TGAAGTCAAAGTAATCACTTTAAAGTCAAAATTGGTGTCGGACTTCAGAAAGGAT TTTCAATTCTATAAAGTTAGGGAGATAAATAACTACCACCACGCGCACGACGCTTATCTTAATGCCGTCGTAGGGACCGCACTCATTAAGAAATACCCGAAGCTAGAAAGT GAGTTTGTGTATGGTGATTACAAAGTTTATGACGTCCGTAAAATGATAGCGAAAAGCGAACAGGAGATAGGCAAGGCTACAGCCAAATACTTCTTTTATTCTAACATTATGA ATTTCTTTAAGACGGAAATCACTCTGGCAAACGGAGAGATACGCAAACGACCTTT AATTGAAACCAATGGGGAGACAGGTGAAATCGTATGGGATAAGGGCCGGGACTTCGCGACGGTGAGAAAAGTTTTGTCCATGCCCCAAGTCAACATAGTAAAGAAAACTG AGGTGCAGACCGGAGGGTTTTCAAAGGAATCGATTCTTCCAAAAAGGAATAGTGA TAAGCTCATCGCTCGTAAAAAGGACTGGGACCCGAAAAAGTACGGTGGCTTCGAT AGCCCTACAGTTGCCTATTCTGTCCTAGTAGTGGCAAAAGTTGAGAAGGGAAAATCCAAGAAACTGAAGTCAGTCAAAGAATTATTGGGCATTACGATTATGGAGCGCTC GTCTTTTGAAAAGAACCCCATCGACTTCCTTGAGGCGAAAGGTTACAAGGAAGTAAAAAAGGACCTCATAATTAAACTACCAAAGTATAGTCTGTTTGAGTTAGAAAATGG CCGAAAAAGAATGTTGGCTAGCGCCGGAGAGCTTCAAAAGGGGAACGAACTCGC ACTACCGTCTAAATACGTGAATTTCCTGTATTTAGCGTCCCATTACGAGAAGTTGAA AGGTTCACCTGAAGATAACGAACAGAAGCAACTTTTTGTTGAACAACACAAACAT TATCTCGACGAAATCATAGAGCAAATTTCGGAATTCAGTAAGAGAGTCATACTAGCTGATGCCAATCTGGACAAAGTATTAAGCGCTTACAACAAGCACAGGGATAAACCCAtty. Dkt. No. 135523-0115ATACGTGAGCAGGCGGAAAATATTATCCATTTGTTTACTCTTACCAACCTCGGCGCTCCGGCCGCGTTCAAGTATTTTGACACAACGATAGATCGCAAACGATACACTTCTACCAAGGAGGTGCTAGACGCGACACTGATTCACCAATCCATCACGGGATTATATGAAACTCGGATAGATTTGTCACAGCTTGGGGGTGACTCTGGGGGGTCTGGGGGTTCTGGGGGTTCTACTAATCTGTCAGATATTATTGAAAAGGAGACTGGTAAGCAACTGGTTATCCAGGAATCCATACTCATGCTCCCAGAGGAGGTGGAAGAAGTCATTGGGAACAAGCCGGAAAGCGATATACTCGTGCACACCGCCTACGACGAGAGCACCGACGAGAATGTCATGCTTCTGACTAGCGACGCCCCTGAATACAAGCCTTGGGCTCTGGTCATACAGGATAGCAACGGTGAGAACAAGATTAAGATGCTCTAACCAGGCATCAAATAAAACGAAAGGCTCAGTCGAAAGACTGGGCCTTTCGTTTTATCTGTTGTTTGTCGGTGAAC GCTCTCPart 8 of Staphylococcus hominis pAB00038 plasmid (SEQ ID NO: 24):TTCCTTGATTTAATTCTAAGATATTTGTATTTTTATATTGTAATCGATTTCTGACCAGACACGTCCCGTGTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAGTGGCACCGAGTCGGTGCTTTTTTTGCGGCCGCGTGATTTA AT gRNAused to engineer Sn00037, Sn00048, Sn00079, Sn00082, Sn00086, Sn00325,Sn00327, Sn00330, Sn00337, Sn00339, Sn00343, and Sn00377 (SEQ ID NO: 25):TCTGACCAGACACGTCCCGTAtty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Atty. Dkt. No. 135523-0115Forward Nextera-overhang primer: 5’-TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGCGTATGTGGGTAGCAGAT-3’(SEQ ID NO: 38)Reverse Nextera-overhang primer: 5’-GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGCTGCGTTATGAACAAATTGCTCTG-3 (SEQ ID NO: 39)PXyl tet promoter tctcctttttcgcttctttattccaattgctttattgacgttgagcctcggaaccgtcgactttatttggatcccctcgagttcatgaaaaacta aaaaaaatattgaaactctatcattgatagagtataattaacaaggaggaataaaaa (SEQ ID NO: 40)Staphylococcus hominis PatB amino acid sequence (SEQ ID NO: 41):1 mnynfdeiid rrytnamnve gykgylfgda dtsdlkdnde lirmwvadmd fgtpevvlna61 irerlnkkil gytnvfgsey yeafvswtkk rygfifpqeh Ivfshgivag lielvgyicd121 kddkvlivtp sygpfkmacd knhistvysp linhhgyyei dfddvrkkve teniklcifa181 nphnptgrvw seeelatlgq imkendvwli sdeihcdikr sgqshipfak avpdydkiit241 tmsqskafni aglmfsniii qnesllktwn thhfgtenpl svvatqaaye kgedwlqamn301 hylddnfnyl adflekelph aefkipeaty lawvdlsyyi kekdidesma kffiknagvi361 iegaeqfvhn aeghirinia vprevmkkgl qkikaalvStaphylococus homins truncated PatB amino acid sequence resulting from the introduction of a stop codon at W190 (SEQ ID NO: 42):VNYNFDEIIDRRYTNAMNVEGYKGYLFGDADTSDLKDNDELIRMWVADMDFG TPEVVLNAIRERLNKKILGYTNVFGSEYYEAF VSWTKKRYGFTF SQEHLVF SHGI VAGLIELVGYICDKDDKALIVTPSYGPFKMACDKNHISTVYSPLINHHGYYEIDF DDVRKKVETENIKLCIFANPHNPTGRV*

Claims

Atty. Dkt. No. 135523-0115CLAIMSWhat is claimed is:

1. A genetically engineered Staphylococcus hominis strain whose genome comprises a disruption of a patB gene.

2. The genetically engineered Staphylococcus hominis strain of claim 1, wherein the strain exhibits decreased expression of the patB gene as compared to a corresponding wildtype Staphylococcus hominis strain.

3. The genetically engineered Staphylococcus hominis strain of claim 1 or claim 2, wherein the disruption prohibits transcription of a full-length wild-type mRNA from the disrupted patB gene.

4. The genetically engineered Staphylococcus hominis strain of any one of claims 1-3, wherein the disruption prohibits expression of a functional PatB enzyme, inactivates the PatB protein, modifies the PatB protein, or has a negative effect on the expression of the patB gene.

5. The genetically engineered Staphylococcus hominis strain of any one of claims 1-4, wherein the genetically engineered strain has decreased PatB enzyme activity as compared to a corresponding wild-type Staphylococcus hominis strain.

6. The genetically engineered Staphylococcus hominis strain of claim 4, wherein the modified PatB protein is a truncated protein.

7. The genetically engineered Staphylococcus hominis strain of claim 6, wherein the truncated protein has decreased catalytic activity compared to a corresponding wild-type Staphylococcus hominis strain.

8. The genetically engineered Staphylococcus hominis strain of any one of claims 1-7, wherein the disrupted patB gene comprises a deletion of all or at least a portion of the gene, optionally wherein the deletion comprises deletion of the patB promoter.Atty. Dkt. No. 135523-01159. The genetically engineered Staphylococcus hominis strain of any one of claims 1-7, wherein the disrupted patB gene comprises an insertion of a gene cassette.

10. The genetically engineered Staphylococcus hominis strain of any one of claims 1-7, wherein the disrupted patB gene comprises a deletion of at least a portion of the gene.

11. The genetically engineered Staphylococcus hominis strain of any one of claims 1-7, wherein the disruption comprises a knockout of the patB gene.

12. The genetically engineered Staphylococcus hominis strain of any one of claims 1-7, wherein the disruption comprises an allelic replacement knockout.

13. The genetically engineered Staphylococcus hominis strain of any one of claims 1-7, wherein the disruption comprises a nucleotide substitution.

14. The genetically engineered Staphylococcus hominis strain of claim 13, wherein the nucleotide substitution is a cytosine (C) to a thymine (T).

15. The genetically engineered Staphylococcus hominis strain of claim 14, wherein the nucleotide substitution is at the W190 tryptophan amino acid site in the patB gene.

16. The genetically engineered Staphylococcus hominis strain of claim 15, wherein the nucleotide substitution results in a premature stop-codon.

17. The genetically engineered Staphylococcus hominis strain of claim 16, wherein the Staphylococcus hominis strain comprises a disrupted patB gene having about 80% to about 100% sequence identity to SEQ ID NO: 15.

18. The genetically engineered Staphylococcus hominis strain of claim 16, wherein the premature stop codon in the patB gene results in a truncated PatB protein lacking one or more enzymatic activities.

19. The genetically engineered Staphylococcus hominis strain of claim 16, wherein the premature stop codon in the expressed mRNA of the patB gene has the sequence UAG, UGA, or UAA.Atty. Dkt. No. 135523-011520. The genetically engineered Staphylococcus hominis strain of any one of claims 1-19, wherein the disruption is introduced by homologous recombination, site-directed mutagenesis, a Transcription activator-like effector nuclease (TALEN), a meganuclease, a zinc finger nuclease, a CRISPR / Cas9 system, a CRISPR base editing system, a CRISPR prime editing system, a CRISPR / Cpfl system, a CRISPR / Csml system, or any combination thereof.

21. The genetically engineered Staphylococcus hominis strain of claim 20, wherein the disruption is introduced by a CRISPR base editing system.

22. The genetically engineered Staphylococcus hominis strain of claim 21, wherein the CRISPR based editing system is a cytosine base editor (CBE) system.

23. The genetically engineered Staphylococcus hominis strain of any one of claims 1-22, wherein the fitness of the strain comprising the disruption is not negatively affected.

24. The genetically engineered Staphylococcus hominis strain of any one of claims 1-23, wherein the strain has a comparable growth rate as compared to a corresponding wild-type Staphylococcus hominis strain.

25. The genetically engineered Staphylococcus hominis strain of any one of claims 1-24, wherein the strain comprises a Staphylococcus hominis of clade A or a Staphylococcus hominis of clade B.

26. The genetically engineered Staphylococcus hominis strain of any one of claims 1-25, wherein the strain comprises Staphylococcus hominis deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725- 17, 300725-18, 300725-19, or 300725-20, or a variant strain thereof.

27. The genetically engineered Staphylococcus hominis strain of any one of claims 1-26, wherein the strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain.Atty. Dkt. No. 135523-011528. The genetically engineered Staphylococcus hominis strain of any one of claims 1-27, wherein the disruption is a deletion or nucleotide substitution in the DNA sequence of the patB gene encoding a PatB protein, wherein the deletion or nucleotide substitution inactivates the PatB protein, truncates the PatB protein, or has a negative effect on expression of the patB gene.

29. The genetically engineered Staphylococcus hominis strain of any one of claims 1-28, wherein the genetically engineered Staphylococcus hominis strain has a 16S rRNA gene sequence that is at least 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical to a sequence selected from the group consisting of SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, and SEQ ID NO: 37.

30. The genetically engineered Staphylococcus hominis strain of claim 29, wherein the strain has the 16S rRNA gene sequence represented by a sequence selected from the group consisting of SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, and SEQ ID NO: 37.

31. The genetically engineered Staphylococcus hominis strain of any one of claims 1-30, wherein the genetically engineered Staphylococcus hominis strain is derived from a strain deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-03, 030924-04, 300725-02, 300725-01, 300725-05, 300725-07, 300725-08, 300725-09, 300725-11, 300725-12, 300725-13, or 300725-10, or a variant strain derived therefrom, wherein the genome has been engineered to comprise a disruption of a patB gene.

32. A genetically engineered Staphylococcus hominis strain deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725- 16, 300725-17, 300725-18, 300725-19, or 300725-20, or a variant strain derived therefrom, wherein the variant strain is obtained using the deposited strain as starting material, and wherein a reduced capacity of the deposited strain to produce 3-methyl-3-sulfanylhexan-l-olAtty. Dkt. No. 135523-0115(3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain is retained or improved in the variant strain as compared to the deposited strain.

33. A Staphylococcus hominis strain, wherein the strain comprises a mutation in at least one endogenous polynucleotide selected from the group consisting of:(a) a nucleotide sequence having about 80% to about 100% sequence identity to SEQ ID NO: 14; and(b) a nucleotide sequence encoding a polypeptide comprising an amino acid sequence having about 80% to about 100% sequence identity to SEQ ID NO: 41 , such that the strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding strain that does not comprise the mutation.

34. The strain of claim 33, wherein the nucleotide sequence of (a) has about 97% to about 100% sequence identity to SEQ ID NO: 14, and / or the nucleotide sequence of (b) encodes a polypeptide comprising an amino acid sequence having about 97% to about 100% sequence identity to SEQ ID NO: 41.

35. The strain of claim 33 or 34, wherein the strain grows at an equivalent or improved rate as compared to a Staphylococcus hominis strain lacking the mutation.

36. The strain of any one of claims 33-35, wherein the strain is incapable of converting Cys-Gly-3M3SH to 3M3SH.

37. A composition comprising an effective amount of one or more of the genetically engineered Staphylococcus hominis strains of any one of claims 1-36.

38. The composition of claim 37, wherein the one or more Staphylococcus hominis strains are lyophilized.

39. The composition of claim 37 or claim 38 further comprising a lyoprotectant.

40. The composition of claim 39, wherein the lyoprotectant is selected from one or more of sucrose, micellar casein, trehalose, or inulin.Atty. Dkt. No. 135523-011541. The composition of any one of claims 37-40, wherein the one or more Staphylococcus hominis strains are viable.

42. The composition of any one of claims 37-41, wherein the one or more Staphylococcus hominis strains are nonproliferative.

43. The composition of any one of claims 37-42 further comprising Staphylococcus epidermidis.

44. The composition of any one of claims 37-43 further comprising an excipient.

45. The composition of claim 44, wherein the excipient is selected from one or more of oleic acid, stearic acid, almond butter, avocado butter, babassu butter, beeswax, cocoa butter, coconut butter, coconut oil, cupuacu butter, hemp seed butter, kokum butter, macadamia butter, mango butter, mowrah butter, olive butter, sal seed butter, shea butter, propylene glycol, glycerin, stearyl alcohol, myristyl alcohol, capric triglyceride, polyethylene glycol (PEG), Isopropyl Myristate, or sorbitol.

46. The composition of any one of claims 37-45, wherein the Staphylococcus hominis strain is present at a concentration of about 106to about 1013bacteria per gram.

47. The composition of claim 46, wherein the Staphylococcus hominis strain is present at a concentration of about 107to about 1011bacteria per gram.

48. The composition of any one of claims 44-47, comprising about 90 wt. % to about 99.99 wt. % excipient.

49. The composition of claim 48, wherein the lyophilized Staphylococcus hominis bacteria is present at about 10 wt. % to about 0.01 wt. %.

50. The composition of claim 48, comprising about 95 wt. % to about 99.75 wt. % excipient.

51. The composition of claim 50, wherein the lyophilized Staphylococcus hominis bacteria is present at about 5 wt. % to about 0.25 wt. %.Atty. Dkt. No. 135523-011552. The composition of any one of claims 37-51, further comprising a fragrance selected from one or more of elettaria cardamomum seed oil, eucalyptus globulus seed oil, citrus aurantium bergamia fruit oil, mentha piperita oil, salvia sclarea oil, juniperus virginiana oil, abies sibirica oil, citrus aurantium amara, citrus aurantium dulcis lower oil, citrus nobilis oil, rosmarinus officinalis oil, melaleuca altemifolia oil, citrus limon tree oil, zingiber officinale root oil, Mangifera indica oil, Lavandula angustifolia oil, and Santalum album.

53. The composition of any one of claims 37-52, wherein the composition is formulated for topical administration.

54. The composition of claim 53, wherein the composition is formulated for use as a topical deodorant.

55. The composition of any one of claims 37-54, further comprising an antiperspirant.

56. The composition of claim 55, wherein the antiperspirant comprises one or more of aluminum salt, charcoal, and witch hazel.

57. The composition of any one of claims 37-56, wherein the topical deodorant is formulated as a deodorant stick, a roll-on, a cream, a lotion, a gel, a powder, a water activated cream, a water activated powder, an injectable, a patch, or a spray.

58. The composition of any one of claims 37-57, wherein the composition is a pharmaceutical composition and the effective amount of one or more of the genetically engineered Staphylococcus hominis strains is a therapeutically effective amount.

59. A pharmaceutical composition comprising a therapeutically effective amount of one or more genetically engineered Staphylococcus hominis strains whose genome comprises a disruption of a patB gene, and pharmaceutically acceptable carrier or excipient, and optionally, a lyoprotectant.

60. The pharmaceutical composition of claim 59, wherein the disruption of the patB gene comprises a nucleotide substitution, optionally wherein the nucleotide substitution comprises a cytosine (C) to thymine (T) substitution as compared to a corresponding wild-type Staphylococcus hominis strain.Atty. Dkt. No. 135523-011561. The pharmaceutical composition of claim 60, wherein the substitution is at the W190 tryptophan amino acid site in the patB gene.

62. The pharmaceutical composition of claim 60 or 61, wherein the substitution introduces a premature stop codon.

63. The pharmaceutical composition of any one of claims 59-62, wherein the one or more genetically engineered Staphylococcus hominis strains are lyophilized.

64. The pharmaceutical composition of claim 63, wherein the pharmaceutically acceptable carrier or excipient is present at about 90 wt. % to about 99.99 wt. % and the one or more lyophilized genetically engineered Staphylococcus hominis strains is present at about 10 wt. % to about 0.01 wt. %.

65. The pharmaceutical composition of claim 64, wherein the pharmaceutically acceptable carrier or excipient is present at about 95 wt. % to about 99.75 wt. % and the one or more lyophilized genetically engineered Staphylococcus hominis strains is present at about 5 wt. % to about 0.25 wt. %.

66. The pharmaceutical composition of any one of claims 59-65, wherein the one or more genetically engineered Staphylococcus hominis strains are present at a concentration of about 106to about 1013bacteria per gram.

67. The pharmaceutical composition of claim 66, wherein the one or more genetically engineered Staphylococcus hominis strains are present at a concentration of about 107to about 101'bacteria per gram.

68. The pharmaceutical composition of any one of claims 59-67, wherein the one or more genetically engineered Staphylococcus hominis strains are selected from the strains deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20.Atty. Dkt. No. 135523-011569. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the International Depositary Authority of Canada (ID AC) under ID AC Accession Number 030924-01.

70. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with ID AC under ID AC Accession Number 030924-02.

71. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-03.

72. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-04.

73. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-06.

74. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-14.

75. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-15.

76. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-16.

77. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-17.Atty. Dkt. No. 135523-011578. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-18.

79. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-19.

80. The pharmaceutical composition of claim 68, wherein the one or more genetically engineered Staphylococcus hominis strains is the strain deposited with the ID AC under ID AC Accession Number 300725-20.

81. The pharmaceutical composition of any one of claims 59-67, comprising two or more of the genetically engineered Staphylococcus hominis strains deposited with ID AC under IDAC Accession Numbers 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, and 300725-20.

82. A method of reducing 3-methyl-3-sulfanylhexan-l-ol (3M3SH) production from a skin microbiota of a subject, the method comprising applying: (i) a composition or a pharmaceutical composition comprising the Staphylococcus hominis strain of any one of claims 1-36 to the skin of the subject; (ii) the composition of any one of claims 37-58; or (iii) the pharmaceutical composition of any one of claims 59-81 to the skin of the subject.

83. A method for treating, reducing, or preventing malodor on the skin of a subject, comprising treating the skin of the subject with a therapeutically effective amount of the composition of any one of claims 37-58 or the pharmaceutical composition of any one of claims 59-81.

84. The method of claim 83, wherein the composition or pharmaceutical composition is applied topically to the skin of the subject.

85. The method of claim 83 or 84, wherein the composition or pharmaceutical composition is applied to a sweat-gland-containing area of skin of the subject.Atty. Dkt. No. 135523-011586. The method of any one of claims 83-85, wherein the composition or the pharmaceutical composition is administered to the subject about 1 to about 2 times per day for about 1 to about 7 days.

87. The method of any one of claims 83-86, wherein the treated skin of the subject comprises a reduction in the amount of malodorous bacteria as compared to a corresponding untreated portion of skin.

88. The method of claim 87, wherein the malodorous bacteria comprises at least wildtype Staphylococcus hominis.

89. A method of producing a genetically engineered Staphylococcus hominis strain, the method comprising targeting the patB gene for disruption.

90. The method of claim 89, wherein the disruption is introduced into the patB gene by homologous recombination, site-directed mutagenesis, a Transcription activator-like effector nuclease (TALEN), a meganuclease, a zinc finger nuclease, a CRISPR / Cas9 system, a CRISPR / Cpfl system, or a CRISPR / Csml system, and any combination thereof.

91. The method of claim 90, wherein the disruption is introduced into the patB gene by the CRISPR base editing system.

92. The method of claim 91, wherein the CRISPR based editing system is a cytosine base editor (CBE) system.

93. The method of claim 91, wherein the CRISPR based editing system comprises transforming a wild-type Staphylococcus hominis strain with a 6-part plasmid.

94. The method of claim 93, wherein the 6-part plasmid comprises one or more sequences selected from SEQ ID NOs: 2-7.

95. The method of claim 94, wherein the 6-part plasmid comprises the sequence of SEQ ID NO: 1.

96. The method of claim 91, wherein the CRISPR based editing system comprises transforming a wild-type Staphylococcus hominis strain with an 8-part plasmid.Atty. Dkt. No. 135523-011597. The method of claim 96, wherein the 8-part plasmid comprises one or more sequences selected from SEQ ID NOs: 17-24.

98. The method of claim 97, wherein the 8-part plasmid comprises the sequence of SEQ ID NO: 16.

99. The method of any one of claims 89-98, wherein the strain exhibits decreased expression of the patB gene as compared to a corresponding wild-type Staphylococcus hominis strain.

100. The method of any one of claims 89-99, wherein the disruption prohibits transcription of a full-length wild-type mRNA from the disrupted patB gene.

101. The method of any one of claims 89-100, wherein the disruption prohibits expression of a functional PatB enzyme.

102. The method of any one of claims 89-101, wherein the genetically engineered strain has decreased PatB enzyme or catalytic activity and the PatB protein is truncated as compared to a corresponding wild-type Staphylococcus hominis strain.

103. The method of any one of claims 89-102, wherein the disrupted patB gene comprises a nucleotide substitution, wherein the substitution is a cytosine (C) to a thymine (T).

104. The method of any one of claims 89-103, wherein the strain is selected from Staphylococcus hominis strains deposited with the ID AC under ID AC Accession Number 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725- 16, 300725-17, 300725-18, 300725-19, or 300725-20, or a variant strain thereof.

105. The method of any one of claims 89-104, wherein the genetically engineered Staphylococcus hominis strain has a reduced capacity to produce 3-methyl-3-sulfanylhexan- l-ol (3M3SH) when contacted with apocrine sweat or a 3M3SH precursor as compared to a corresponding wild-type strain.

106. An altered microbiota comprising a genetically engineered Staphylococcus hominis strain, or a variant strain thereof, whose genome comprises a disruption of a patB gene.Atty. Dkt. No. 135523-0115107. The altered microbiota of claim 106, wherein the microbiota has a reduced capacity to produce 3-methyl-3-sulfanylhexan-l-ol (3M3SH) when contacted with apocrine sweat as compared to a corresponding wild-type microbiota.

108. A deodorant composition comprising:(i) a therapeutically effective amount of a viable Staphylococcus hominis strain of any one of claims 1-36; and(ii) a therapeutically acceptable carrier or excipient; wherein the deodorant composition is formulated as a water activated formulation for topical administration.

109. The deodorant composition of claim 108, comprising about 90 wt. % to about 99.99 wt. % therapeutically acceptable carrier or excipient.

110. The deodorant composition of claim 109, wherein the Staphylococcus hominis strain is lyophilized.

111. The deodorant composition of claim 110, wherein the deodorant composition comprises about 10 wt. % to about 0.01 wt. % lyophilized Staphylococcus hominis bacteria.

112. The deodorant composition of claim 109, comprising about 95 wt. % to about 99.75 wt. % of the therapeutically acceptable carrier or excipient.

113. The deodorant composition of claim 112, wherein the deodorant composition comprises about 5 wt. % to about 0.25 wt. % lyophilized Staphylococcus hominis bacteria.

114. The deodorant composition of any one of claims 108-113, wherein the Staphylococcus hominis strain is present at a concentration of about 106to about 1016bacteria per gram.

115. The deodorant composition of claim 114, wherein the Staphylococcus hominis strain is present at a concentration of about 107to about 1011bacteria per gram.

116. The deodorant composition of any one of claims 108-115 further comprising: (iii) a lyoprotectant.Atty. Dkt. No. 135523-0115117. The deodorant composition of any one of claims 108-116, wherein the Staphylococcus hominis strain comprises one or more of the strains deposited with the ID AC under IDAC Accession Numbers 030924-01, 030924-02, 300725-03, 300725-04, 300725-06, 300725-14, 300725-15, 300725-16, 300725-17, 300725-18, 300725-19, or 300725-20.

118. A deodorant stick comprising the deodorant composition of any one of claims 108- 117.

119. A pharmaceutical composition comprising a therapeutically effective amount of one or more of the Staphylococcus hominis strains of any one of claims 1-36, and a therapeutically acceptable carrier or excipient, for use in the treatment of malodor on the skin of a subject.

120. The pharmaceutical composition of claim 119, wherein the one or more Staphylococcus hominis strains are lyophilized.

121. The pharmaceutical composition of claim 120 further comprising a lyoprotectant.

122. Use of the one or more Staphylococcus hominis strains of any one of claims 1-36 in the manufacture of a medicament for the treatment of malodor on the skin of a subject.

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