Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
  • A61K35/66—Microorganisms or materials therefrom
  • A61K35/74—Bacteria
  • A61K35/741—Probiotics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
  • A61K35/66—Microorganisms or materials therefrom
  • A61K35/74—Bacteria
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/0012—Galenical forms characterised by the site of application
  • A61K9/0014—Skin, i.e. galenical aspects of topical compositions
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
  • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
  • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
  • A61K31/51—Thiamines, e.g. vitamin B1
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
  • A61K35/66—Microorganisms or materials therefrom
  • A61K35/74—Bacteria
  • A61K35/741—Probiotics
  • A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P17/00—Drugs for dermatological disorders
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P17/00—Drugs for dermatological disorders
  • A61P17/04—Antipruritics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/04—Antibacterial agents
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
  • C12N1/00—Microorganisms; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
  • C12N1/20—Bacteria; Culture media therefor
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K9/00—Medicinal preparations characterised by special physical form
  • A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
  • C12R2001/00—Microorganisms ; Processes using microorganisms
  • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
  • C12R2001/07—Bacillus

Definitions

• the second strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 1.
• the third strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 2.
• the first strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 13.
• the first strain further comprises having a sequence of SEQ ID NO: 13.
• the second strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 15.
• the second strain further comprises having a sequence of SEQ ID NO: 15.
• the first strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 11, SEQ ID NO: 12, or both. In some embodiments, the first strain further comprises having a sequence of SEQ ID NO: 11, SEQ ID NO: 12, or both. In some embodiments, the second strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 15. In some embodiments, the second strain further having a sequence of SEQ ID NO: 15. In some embodiments, the third strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 7, SEQ ID NO: 8, or both. In some embodiments, the third strain further comprises having a sequence of SEQ ID NO: 7, SEQ ID NO: 8, or both.
• the fourth strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 4.
• the fifth strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 1.
• the sixth strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 2.
• the bacterial strains comprise bacterial strains the first strain and the second strain.
• the bacterial strains comprise the fourth strain and the second strain.
• the bacterial strains comprise the fourth strain and the sixth strain.
• the bacterial strains comprise the first strain and the third strain.
• a composition comprises: a bacterial strain that is purified, wherein the bacterial strain comprises: a 16s rRNA sequence with at least 97% sequence identity to SEQ ID NO: 4; and wherein (a) the bacterial strain is lyophilized; or (b) the composition is formulated for delivery to a skin.
• the bacterial strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 4.
• the bacterial strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 11, SEQ ID NO: 12, or both.
• the bacterial strain further comprises having a sequence of SEQ ID NO: 11, SEQ ID NO: 12, or both.
• the composition is formulated as: a suspension, an emulsion, a cream, a lotion, a tincture, a gel, a foam, a powder, an ointment, a paste, or an oil.
• the composition further comprises a second bacterial strain.
• the composition comprises at least 10 A 3 colony forming units (cfu) per gram of bacteria.
• the composition comprises 10 A 3 to 10 A l 2 colony forming units (cfu) per gram of bacteria.
• a bacterial strain is grown in aerobic conditions. In some embodiments, a bacterial strain is grown without animal products.
• a composition comprises: a bacterial strain that is purified, wherein the bacterial strain comprises: a 16s rRNA sequence with at least 97% sequence identity to SEQ ID NO: 1; and wherein (a) the bacterial strain is lyophilized; or (b) the composition is formulated for delivery to a skin.
• the bacterial strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 1.
• the bacterial strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 15.
• the bacterial strain further comprises having a sequence of SEQ ID NO: 15.
• a bacterial strain is grown in Tryptic Soy Broth (TSB).
• TTB Tryptic Soy Broth
• the composition when stored in a sealed container placed at 20 °C retains at least about: 10 A 4 cfu after 6 months, as measured by cfu counts.
• the composition further comprises an excipient.
• the composition further comprises a lyoprotectant.
• the composition further comprises an emollient.
• the composition further comprises or a salt thereof.
• a composition comprises: a bacterial strain that is purified, wherein the bacterial strain comprises: a 16s rRNA sequence with at least 97% sequence identity to SEQ ID NO: 5; and wherein (a) the bacterial strain is lyophilized; or (b) the composition is formulated for delivery to a skin.
• the bacterial strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 5.
• the bacterial strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 13.
• the bacterial strain further comprises having a sequence of SEQ ID NO: 13.
• the composition is formulated as: a suspension, an emulsion, a cream, a lotion, a tincture, a gel, a foam, a powder, an ointment, a paste, or an oil.
• the composition further comprises a second bacterial strain.
• the composition comprises at least 10 A 3 colony forming units (cfu) per gram of bacteria.
• the composition comprises 10 A 3 to 10 A l 2 colony forming units (cfu) per gram of bacteria.
• a bacterial strain is grown in aerobic conditions. In some embodiments, a bacterial strain is grown without animal products.
• a composition comprises: a bacterial strain that is purified, wherein the bacterial strain comprises: a 16s rRNA sequence with at least 97% sequence identity to SEQ ID NO: 6; and wherein (a) the bacterial strain is lyophilized; or (b) the composition is formulated for delivery to a skin.
• the bacterial strain comprises the 16s rRNA sequence with at least 97% sequence identity over at least 1000 bases to SEQ ID NO: 6.
• the bacterial strain further comprises a sequence with at least 95% sequence identity to SEQ ID NO: 14.
• the bacterial strain further comprises having a sequence of SEQ ID NO: 14.
• the composition is formulated as: a suspension, an emulsion, a cream, a lotion, a tincture, a gel, a foam, a powder, an ointment, a paste, or an oil.
• the composition further comprises a second bacterial strain.
• the composition comprises at least 10 A 3 colony forming units (cfu) per gram of bacteria.
• the composition comprises 10 A 3 to 10 A l 2 colony forming units (cfu) per gram of bacteria.
• a bacterial strain is grown in aerobic conditions. In some embodiments, a bacterial strain is grown without animal products.
• aureus cause a reduction in expression of at least one of the following genes: gmk, agr,psma, saeR, ccpA, and SigB in S. aureus, wherein the reduction in expression is measured by a fluorescence reporter assay.
• the bacterial strains are present in an amount effective to suppress virulence of 5. aureus as measured by a reduction in expression of at least one of the following genes: gmk, agr, psma, saeR, ccpA, and SigB in S. aureus wherein the reduction in expression is measured by a fluorescence reporter assay.
• aureus cause a reduction in expression of at least one of the following genes: gmk, agr, psma, saeR, ccpA, and SigB in S. aureus, wherein the reduction in expression is measured by a fluorescence reporter assay.
• the bacterial strains are present in an amount effective to suppress virulence of 5. aureus as measured by a reduction in expression of at least one of the following genes: gmk, agr, psma, saeR, ccpA, and SigB in 5. aureus wherein the reduction in expression is measured by a fluorescence reporter assay.
• a method comprises administering an amount sufficient to treat a disease selected from the group consisting of: atopic dermatitis, seborrheic dermatitis, inflammation, eczema, psoriasis, rosacea, mycoses, dermatophytosis, folliculitis, acne, alopecia, vitiligo, dandruff, chronic wound, skin ulcer, Netherton syndrome, hidradenitis suppurativa, sycosis vulgaris, staphylococcal scalded skin syndrome, impetigo, ecthyma, cellulitis, carbuncle, furuncle, and abscess.
• a disease selected from the group consisting of: atopic dermatitis, seborrheic dermatitis, inflammation, eczema, psoriasis, rosacea, mycoses, dermatophytosis, folliculitis, acne, alopecia,
• the method comprises administering an amount sufficient for a reduction of incidence of a condition associated with inflammation and wherein the condition associated with inflammation comprises an itch, a rash, a redness, a pain, a swelling, a blistering, or a scaling.
• a method comprises administering an amount sufficient to suppress virulence of 5. aureus as measured by a reduction in expression of at least one of the following genes: gmk, agr, psma, saeR, ccpA, and SigB in S. aureus, wherein the reduction in expression is measured by a fluorescence reporter assay.
• the method of administering is topical administration.
• Also disclosed herein are methods for reducing an incidence of a condition associated with inflammation comprising: topically administering a composition comprising: a first bacterial strain, which comprises a 16s rRNA sequence with at least 97% sequence identity to SEQ ID NO: 1; and a second bacterial strain, which comprises a 16s rRNA sequence with at least 97% sequence identity to SEQ ID NO: 6, to a subject in need thereof, wherein the bacterial strains are purified, wherein the bacterial strains are present in an amount sufficient for a reduction of the incidence of a condition associated with inflammation and wherein the condition associated with inflammation comprises an itch, a rash, a redness, a pain, a swelling, a blistering, or a scaling.
• FIGURE 5 is a bar graph that plots percent change of 5. aureus gene expression by a fluorescent reporter in the three-strain mixture of Strain jl.83, Strain jl.27 and Strain jl.77 as compared to S. aureus monoculture in different environmental conditions.
• the Y-axis shows the percent change as compared to the 5. aureus monoculture.
• the X-axis shows the environmental condition that was tested.
• FIG. 5 shows inhibition of different 5. aureus promoter-reporter constructs (agr, psmA, GMK) when the reporter strains of S.
• FIGURE 11 is a bar graph that plots percent growth of mixed cultures as compared to 5. aureus monoculture.
• the Y-axis shows the effect of growth inhibition (%) by the strain combination.
• the X-axis shows the growth of different S. aureus strains in the presence of an ensemble (e.g., the three-strain combination) or the S. aureus strain alone.
• FIG. 11 shows inhibition of different S. aureus strains when grown with the strains Strain jl.21, Strain jl.68, and Strain jl.121. This figure shows the combination of the isolates are able to inhibit growth of numerous S. aureus isolates.
• FIGURE 12 is a bar graph that plots percent growth of mixed cultures as compared to 5. aureus monoculture.
• the Y-axis shows the effect of growth inhibition (%) by the strain combination.
• the X-axis shows the growth of different S. aureus strains in the presence of an ensemble (e.g., the three-strain combination) or the S. aureus strain alone.
• FIG. 12 shows inhibition of different S. aureus strains when grown with the strains Strain jl.27, Strain jl.68, and Strain jl.121. This figure shows the combination of the isolates are able to inhibit growth of numerous S. aureus isolates.
• the level of identity in relation to a nucleotide sequence is determined for the entire sequence searched.
• Percent identity may be at least 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% to a reference bacterial 16S rRNA sequence, 16S rRNA V4 region sequence, or whole genome sequence.
• Percent identity may be at least 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% to a reference bacteria 16S rRNA: VI region, V2 region, V3 region, V5 region, V6 region, V7 region, V8 region or V9 region sequence.
• Percent identity may be at least 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% to a reference bacterial sequence.
• the skin of a human provides many environments for the microbiome.
• both the microbiome of the skin and the host environment of the skin are important for maintaining skin health.
• dysbiosis of the skin microbiome and replacement of commensal bacterial species with pathogenic bacterial species can cause disease.
• perturbation of a damaged skin microbiome with bacterial isolates from healthy individuals can restore function and diversity of a skin microbiome.
• restoration and/or modification of a damaged microbiome can be used to treat a disease.
• Compositions described herein are used for treatment of a damaged skin microbiome and contain bacterial species that are purified.
• Strain jl.68 comprises a 16s rRNA sequence with at least: 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 5.
• Strain jl.121 comprises the 16s rRNA sequence of SEQ ID NO: 6.
• Strain jl.121 comprises a 16s rRNA sequence with at least: 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 6.
• Strain jl.21 comprises the 16s rRNA sequence of SEQ ID NO: 3.
• Strain jl.21 comprises a 16s rRNA sequence with at least: 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 3.
• a bacterial strain comprises Strain jl. 19, Strain jl.39, Strain jl.26, Strain jl.116, Strain jl. 119, or Strain jl.45.
• Strain jl.19 comprises the 16s rRNA sequence of SEQ ID NO: 16.
• Strain jl.19 comprises a 16s rRNA sequence with at least: 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 16.
• Strain jl.39 comprises the 16s rRNA sequence of SEQ ID NO: 17.
• Strain jl.39 comprises a 16s rRNA sequence with at least: 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 17.
• Strain jl.26 comprises the 16s rRNA sequence of SEQ ID NO: 18.
• Strain jl.26 comprises a 16s rRNA sequence with at least: 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 18.
• Strain jl. 116 comprises the 16s rRNA sequence of SEQ ID NO: 19.
• Strain jl.21 comprises an identifier sequence with at least: 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 9. In some cases, Strain jl.21 comprises an identifier sequence with at least: 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to SEQ ID NO: 10.
• a composition comprises three or more bacterial species or strains.
• a composition comprises at least three of the following bacterial species: Staphylococcus cohnii, Staphylococcus capitis, Staphylococcus caprae, Bacillus tropicus, Bacillus mycoides, Bacillus wiedmannii, Bacillus mediterraneensis, Bacillus amyloliquefaciens, Bacillus velezensis, Bacillus cereus, Bacillus cecembensis, Kocuria marina, and Lysinibacillus macroides.
• a composition comprises a bacterial species Bacillus amyloliquefaciens, Bacillus tropicus, and Staphylococcus caprae. In some cases, a composition comprises bacterial strains Strain jl.68, Strain jl.21, and Strain jl.121. In some embodiments, a composition comprises a bacterial species combination from Table 3. In some embodiments, a composition is a bacterial species combination from Table 3. In some embodiments, a composition comprises a bacterial strain combination from Table 4. In some embodiments, a composition is a bacterial strain combination from Table 4.
• a composition comprises Bacillus mediterraneensis. In some cases, a composition comprises Bacillus mediterraneensis jl.44. In some embodiments, a composition comprises Bacillus cereus. In some cases, a composition comprises Strain jl.116. In some embodiments, a composition comprises Bacillus cecembensis. In some cases, a composition comprises Strainjl.119. In some embodiments, a composition comprises Kocuria marina. In some cases, a composition comprises Strain jl.45.
• Bacillus tropicus is Bacillus tropicus ATCC 4342 (Bacillus tropicus NRS 731), which can be obtained from the ATCC (American Type Culture Collection) depository.
• a bacterial species or a strain described herein when administered to a subject, can reduce or eliminate colonization of the skin of pathogenic: bacteria, fungi or viruses.
• a composition described herein is used to kill a pathogenic bacteria, fungi or virus.
• a composition described herein is used to suppress growth of a pathogenic bacteria, fungi or virus.
• a composition described herein is used to control growth of a pathogenic bacteria, fungi or virus.
• Skin pathogenic bacteria include, without limitation, Staphylococcus sp., Micrococcus sp., Corynebacterium sp., Staphylococcus aureus, Streptococcus pyogenes, Pseudomonas aeruginosa, Pasteurella multocida, Capnocytophaga canimorsus, Bartonella sp.., Klebsiella rhinoscleromatis, and Vibrio vulnificus.
• a pathogenic bacteria comprises Gardnerella vaginalis.
• a compound comprises acetate, beta-alanine, bicarbonate, biotin, butyrate, caffeine, citrate, creatine, D-cellobiose, D-fructose, D-glucosamine, D-glucose, D- mannitol, D-raffinose, D-sorbitol, D-sucrose, D-trehalose, D-xylose, formate, GlcNAc, glycerol, glycine, L-alanine, L-arabinose, L-arginine, L-citrulline, L-glutamine, L-hydroxyproline, L- isoleucine, L-leucine, L-methionine, L-omithine, L-proline, L-serine, L-taurine, L-threonine, L- valine, L-ascorbate, L-lactate, nicotinamine, polysorbate 20, polysorbate 80, propionat
• Non-limiting examples of optional additives include preservatives, such as sorbate; solvents, such as isopropanol and propylene glycol; astringents, such as menthol and ethanol; emollients, such as polyalkylene methyl glucosides; humectants, such as glycerin; emulsifiers, such as glycerol stearate, PEG-100 stearate, polyglyceryl-3 hydroxyluryl ether, and polysorbate 60; sorbitol and other polyhydroxy alcohols, such as polyethylene glycol; sunscreen agents, such as methoxy octyl cinnamate (Parsol MCX) and butyl methoxy benzoylmethane (Parsol 1789); antioxidants, such as ascorbic acid (vitamin C), a-tocopherol (Vitamin E), [S-tocopherol, y- tocopherol, 6-tocopherol
• Dosing may include single or multiple administrations of pharmaceutical compositions described herein. Examples include: multiple times a day, daily, every other day, 1, 2, 3, 5, 6, or 7 times a week, weekly, or less often, a single administration, a course of treatment involving several treatments on a regular or irregular basis, or multiple administrations for a period of time a disease or condition is treated. In some cases, dosing can occur every day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 1 year, 2 years, or as needed.
• compositions described herein may comprise about: 10 2 to 10 12 cfu, 10 3 to 10 12 cfu, 10 3 to 10 11 cfu, 10 3 to IO 10 cfu, 10 3 to 10 9 cfu, 10 3 to 10 8 cfu, 10 3 to 10 7 cfu, 10 3 to 10 6 cfu, 10 3 to about 10 5 cfu, 10 3 to 10 4 cfu, 10 4 to 10 12 cfu, 10 4 to 10 11 cfu, 10 4 to IO 10 cfu, 10 4 to 10 9 cfu, 10 4 to 10 8 cfu, 10 4 to 10 7 cfu, 10 4 to 10 6 cfu, 10 5 to 10 12 cfu, 10 5 to 10 11 cfu, about 10 5 to about IO 10 cfu, 10 6 to 10 12 cfu, 10 7 to 10 12 cfu, 10 8 to 10 12 cfu, 10 9 to 10 12 cfu
• compositions comprise about 10 3 cfu, about 10 4 cfu, about 10 5 cfu, about 10 6 cfu, about 10 7 cfu, about 10 8 cfu, about 10 9 cfu, about IO 10 cfu, about 10 11 cfu, about 10 12 cfu, or about 10 13 cfu of bacteria, a bacterial species, or a bacterial strain described herein.
• Compositions, such as pharmaceutical compositions, described herein may comprise 10 2 to 10 15 colony forming units (cfu) of bacteria, a bacterial species, or a bacterial strain described herein per mL.
• compositions described herein may comprise about 10 2 to 10 12 cfu, 10 3 to 10 12 cfu, 10 3 to 10 11 cfu, 10 3 to IO 10 cfu, 10 3 to 10 9 cfu, 10 3 to 10 8 cfu, 10 3 to 10 7 cfu, 10 3 to 10 6 cfu,
• compositions described comprises isolated bacteria present in an amount sufficient for a reduction in the virulence of a pathogenic bacteria, fungi or virus.
• a disease or condition can relate to a bacterial infection.
• Sources for bacterial infections for treatment with pharmaceutical compositions described herein include, without limitation, S. aureus (methicillin-resistant S. aureus (MRSA) and methicillin-sensitive S. aureus (MSSA)), Gardnerella vaginalis, and Streptococcus pyogenes.
• a skin disease or condition comprises eczema, diaper rash, seborrheic dermatitis, chickenpox, measles, warts, acne, fifth disease, hives, ringworm, rashes from a bacterial or a fungal infections (e.g., cutaneous candidiasis), or rashes from an allergic reaction.
• compositions and methods described herein are compositions used to improve a cosmetic irregularity.
• compositions and methods described herein are compositions used to treat a condition selected from the group consisting of: a pruritis, an aesthetic condition, and a body odor.
• an aesthetic condition comprises wrinkles or appearance of aging.
• an aesthetic condition is a cosmetic irregularity.
• efficacy assessments can be taken over a period of time, for example, after each treatment, or every week, or every month. All grading assessments can be performed by the same investigator at each visit or by a computer implementing an algorithm to ensure grading consistency.
• a measurement is taken, such as the number or wrinkles, or the depth of wrinkles, and is a factor in a clinical score.
• a symptom comprises a raised bump, a discolored bump, a rash, a pain, an itch, a scaly skin, a rough skin, a peeling skin, an ulcer, an open sore, a lesion, a dry skin, a cracked skin, a discolored patch of skin, a fleshy bump, a wart, a skin growth, a mole, a change in a mole size or color, a loss of skin pigment, or an excessive flushing.
• a symptom comprises an itch, a rash, a redness, a pain, a swelling, a blistering, or a scaling.
• a composition is administered in an amount sufficient to reduce symptoms associated with dry skin.
• kits wherein the kit comprises: a first container, wherein the first container comprises a purified, and lyophilized bacteria that comprises: Strain jl.83, Strain jl.27, and Strain jl.77; and a second container, wherein the second container comprises a pharmaceutically acceptable excipient.
• the kit comprises: a first container, wherein the first container comprises a purified, and lyophilized bacteria that comprises: Strainjl.68, Strain jl.27, and Strain jl.121; and a second container, wherein the second container comprises a pharmaceutically acceptable excipient.
• kits wherein the kit comprises: a first container, wherein the first container comprises a purified, and lyophilized bacteria that comprises: Strainjl.68; Strain jl.21; and Strain jl.121; and a second container, wherein the second container comprises a pharmaceutically acceptable excipient.
• Embodiment 3 The composition of embodiment 1, further comprising a fourth bacterial species.
• Embodiment 12 A composition, wherein the composition comprises: a bacterial species that is purified, wherein the bacterial species comprises: Bacillus velezensis,' and wherein (a) the bacterial species is lyophilized; or (b) the composition is formulated for delivery to a skin.
• Embodiment 13 The composition of embodiment 12, wherein the composition is formulated as: a suspension, an emulsion, a cream, a lotion, a tincture, a gel, a foam, a powder, an ointment, a paste, or an oil.
• Embodiment 14 The composition of embodiment 12, further comprising a second bacterial species.
• Embodiment 18 A composition, wherein the composition comprises: a bacterial species that is purified, wherein the bacterial species comprises: Lysinibacillus macroides,' and wherein (a) the bacterial species is lyophilized; or (b) the composition is formulated for delivery to a skin.
• Embodiment 19 The composition of embodiment 18, wherein the composition is formulated as: a suspension, an emulsion, a cream, a lotion, a tincture, a gel, a foam, a powder, an ointment, a paste, or an oil.
• Embodiment 21 A composition, wherein the composition comprises: a bacterial species that is purified, wherein the bacterial species comprises: Bacillus tropicus,' and wherein (a) the bacterial species is lyophilized; or (b) the composition is formulated for delivery to a skin.
• Embodiment 28 The composition of any one of embodiments 1-27, wherein the bacterial species are grown in Tryptic Soy Broth (TSB).
• TLB Tryptic Soy Broth
• Embodiment 32 The composition of any one of embodiments 1-31, further comprising an emollient.
• Embodiment 33 The composition of any one of embodiments 1-32, further comprising thiamine or a salt thereof.
• Embodiment 34 The composition of any one of embodiments 1-33, wherein the bacterial species when contacted with 5. aureus cause a reduction in expression of at least one of the following genes: gmk, agr,psma, saeR, ccpA, and SigB in S. aureus, wherein the reduction in expression is measured by a fluorescence reporter assay.
• Embodiment 36 A method of administering the composition of any one of embodiments 1-35, comprising administering an amount sufficient to treat a disease selected from the group consisting of: atopic dermatitis, seborrheic dermatitis, inflammation, eczema, psoriasis, rosacea, mycoses, dermatophytosis, folliculitis, acne, alopecia, vitiligo, dandruff, chronic wound, skin ulcer, Netherton syndrome, hidradenitis suppurativa, sycosis vulgaris, staphylococcal scalded skin syndrome, impetigo, ecthyma, cellulitis, carbuncle, furuncle, and abscess.
• Embodiment 37 A method of administering the composition of any one of embodiments 1-35, comprising administering an amount sufficient to reduce symptoms associated with atopic dermatitis.
• compositions comprising bacterial species that are purified.
• the bacterial species comprises: Bacillus velezensis,' Bacillus wiedmannii,' and Lysinibacillus macroides.
• the bacterial species are lyophilized; or (b) the composition is formulated for delivery to a skin.
• the composition is formulated as: a suspension, an emulsion, a cream, a lotion, a tincture, a gel, a foam, a powder, an ointment, a paste, or an oil.
• the composition can further comprise a fourth bacterial species.
• the method comprises administering an amount sufficient to treat a condition selected from the group consisting of: pruritis, an aesthetic condition, and body odor.
• the aesthetic condition comprises wrinkles or appearance of aging.
• the method comprises administering an amount sufficient to reduce symptoms associated with atopic dermatitis.
• the method comprises administering the composition to a subject who has an eczema prone microbiome prior to the administration.
• the method comprises administering an amount sufficient to reduce symptoms associated with dry skin.
• the S. aureus promoter/reporter strains contained a fluorescent reporter under the control of a promoter of a gene of interest.
• the promoter/reporter strains generated were as follows: Strain 1- agr promoter for quorum sensing induction, Strain 2- psmA promoter for a toxin that damages host tissue, Strain 3- saeR promoter for virulence regulation, Strain 4- sigB promoter for the stress response sigma factor, Strain 5- ccpA promoter for metabolism (e.g, carbon catabolite repression), and strain 6- GMK promoter for constitutive metabolic function.
• strains tested were Strain jl.27 (jl27), Strain jl.77 (jl77), Strain jl.83 (jl83), Strain jl.21 (jl21), Strain jl.68 (jl68), and Strain jl.121 (jll21).
• strains grown in animal-free media there was a 10-100X improvement in cultivation cfu/ml after 48 hours.
• the data shows the strains can be cultivated in a variety of media and can be manufactured in animal free media for production of a composition, such as a pharmaceutical composition.
• EXAMPLE 7 Assays of X. aureus growth inhibition with combinations of bacterial isolates
• Strain jl.27 was isolated from the skin of a healthy individual and was screened in a pair- wise combinatorial screening assay against 4 X. aureus behavior reporters. X. aureus behaviors were measured via a set of plasmid-mediated “promoter-reporter” X. aureus strains, whose fluorescence report on one specific activity. Combinations of strains were screened in an array assay. The strain mixture conditions were: 1) Strain jl.27 self-cross, 2) Strain jl.27 with random skin isolate, and 3) Strain jl.27 with another isolate that showed inhibition of X. aureus gene expression.
• the screen analyzed the expression of the reporters: agr for quorum sensing induction, psmA for toxin that damages host tissue, and GMK for constitutive metabolic function.
• Combinations of strains were screened in an array after 1 day of growth.
• the strain mixture conditions were: 1) three-wise combination with an additional set of 4 strains, 2) two-wise combination with an additional set of 5 strains, and 3) one individual strain with an additional set up 6 strains.
• jll21/jl21/jl68 + 4X shows tested communities where jl.121, jl.21 and jl.68 and up to 4 additional microbes (chosen among 89 strains) were present.
• aureus agr quorum sensing
• psmA toxin production
• GMK metabolic function
• strong performance of the strain combination (ensemble) still occurs even with other microbes present, suggesting the combination will perform well in a native microbiome; and (3) that performance is stronger than when any subset of the 3-species combination is present.
• a 3-species combination of Strain jl.121, Strain jl.21, and Strain jl.68 are robust to the larger community and more robust than their subsets as agr expression was reduced to less than about 5%, psmA expression was reduced to less than about 2%, and GMK expression was reduced to less than about 20%.
• FIG. 15 shows the percent expression of the reporter as compared to the 5. aureus monoculture (i.e., the monoculture is 100%).
• the X-axis shows the strain combinations that were tested.
• the tested strains were Strain jl.121, Strain jl.27, and Strain jl.68. These strains were tested in different combinations with a random selection of strains to form 7-strain communities.
• the data shows the strain combination containing Strain jl.121, Strain jl.27, and Strain jl.68 were (1) effective at suppressing S.
• aureus agr quorum sensing
• psmA toxin production
• GMK metabolic function
• strong performance of the strain combination (ensemble) still occurs even with other microbes are present, suggesting the combination will perform well in a native microbiome; and (3) that performance is stronger than when any subset of the 3-species combination is present.
• a 3-species combination of Strain jl.121, Strain j 1.27, and Strain j 1.68 are robust to the larger community and more robust than their subsets as agr expression was reduced to less than about 5%, psmA expression was reduced to less than about 2%, and GMK expression was reduced to less than about 40%.
• EXAMPLE 12 Sequences of isolated strains [0202] Isolated strains were subjected to whole genome sequencing and assembly of generated contigs. Additionally, 16s rRNA gene sequences were sequenced by Sanger sequencing and manually trimmed. The 16s rRNA sequences from the isolated strains are presented in Table 6. Additionally, several identifier sequences were pulled from the assembly for each isolated strain for identification. The identifier sequences from the isolated strains are presented in Table 7. To identify the identifier sequences, the genome assemblies were assessed for strain identifying sequences. The assemblies were preprocessed by: removing 1000 bp from each side of each assembly sequence to minimize edge effects and if the resulting sequence length after cropping was ⁇ 10 kbp, it was dropped from consideration.
• a series of new sub-sequences was created by applying a sliding window of some length, with a 10% overlap across windows; a 5000bp window with a 500bp overlap was used and a 1200bp window with a 120bp overlap was used.
• the sequences were searched by BLAST from a nucleotide collection database (the Nucleotide Collection (nt) database) provided by The National Center for Biotechnology Information (NCBI) downloaded on September 17, 2022.
• nt Nucleotide Collection
• NCBI National Center for Biotechnology Information
• S. aureus reporter strains containing a GMK promoter fused GFP reporter or an agr promoter fused GFP reporter were grown in liquid cultures in the following conditions: grown in monoculture, mixed with S. aureus WT (control); mixed with Strain jl.27, Strain jl.68, and Strain jl.77; mixed with Strain jl.68 alone; mixed with supernatant from Strain jl.27, Strain jl.68, and Strain jl.77; or mixed with supernatant from strain jl.68 alone.
• the expression of GMK and agr were determined after incubation for 24 hours.
• FIGS. 18C-D show in a 1:1 coculture with Strain jl.27, Strain jl.68, and Strain jl.77 or with Strain jl.68 there was reduced cell viability ( ⁇ 1% (CFU)).
• quorum sensing agr-GFP was reduced to negligible levels (near 0%) compared to the PBS control
• GMK-GFP metabolism
• EXAMPLE 16 Growth inhibition of S. aureus after mixture with supernatant of a selected strain
• aureus containing an agr promoter fused GFP reporter was plated in 10-fold dilutions on TSB agar plates containing 0% (PBS control), 5% or 10% (v/v) supernatant from Strain jl.68. The plates were incubated for 18 hours at 37 °C and colonies were counted.
• FIG. 20 shows images of the plates containing the dilutions of 5. aureus on the agar plates comprising the various percentages of supernatant from Strain jl.68. In the presence of unaltered TSB medium, individual colonies were countable around 6-7 10-fold dilutions of a saturated culture. They were bright and fluorescent, indicating quorum sensing was on.

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