Altering microbial populations & modifying microbiota

Abstract

The invention relates to methods, uses, systems, arrays, engineered nucleotide sequences and vectors for inhibiting bacterial population growth or for altering the relative ratio of sub-populations of first and second bacteria in a mixed population of bacteria. The invention is particularly useful, for example, for treatment of microbes such as for environmental, medical, food and beverage use. The invention relates inter alia to methods of controlling microbiologically influenced corrosion (MIC) or biofouling of a substrate or fluid in an industrial or domestic system.

1 . A method of treating or preventing obesity, diabetes or a gastrointestinal condition in a subject, wherein the subject comprises a mixed gut bacterial population comprising a first bacterial sub-population and a second bacterial sub-population, wherein the first bacterial sub-population comprises  Bacteroides  bacteria and the second bacterial sub-population comprises host cells of a target bacterial species that is different from the  Bacteroides  bacteria, the method comprising increasing the relative ratio of the  Bacteroides  bacteria versus the host cells in the mixed gut bacterial population. 
     
     
         2 . The method of  claim 1 , wherein the method comprises killing the host cells or reducing growth of the host cells. 
     
     
         3 . The method of  claim 1 , wherein the method comprising:
 (a) contacting the mixed gut bacterial population with an engineered nucleic acid for producing a host modifying crRNA (HM-crRNA), and   (b) producing the HM-crRNA in the host cells;   wherein:   (i) the HM-crRNA is operable with a Cas in the host cells, wherein the engineered nucleic acid and the Cas are comprised by a HM-CRISPR/Cas system in the host cells; and   (ii) the HM-crRNA comprises a nucleotide sequence that is capable of hybridizing to a target sequence in the host cells to guide the Cas to modify the target sequence in the host cells,   wherein the host cells are killed or growth of the host cells is reduced, thereby increasing the relative ratio of the  Bacteroides  bacteria versus the second bacterial sub-population in the mixed gut bacterial population.   
     
     
         4 . The method of  claim 3 , wherein the method comprises administering to the subject an antimicrobial composition comprising the engineered nucleic acid for producing the HM-crRNA, and wherein the mixed gut bacterial population is contacted with the engineered nucleic acid for producing the HM-crRNA in vivo. 
     
     
         5 . The method of  claim 3 , wherein the mixed gut bacterial population is contacted with the engineered nucleic acid for producing the HM-crRNA ex vivo to provide a bacterial composition, and wherein the method comprises administering the bacterial composition to the subject. 
     
     
         6 . The method of  claim 1 , wherein the  Bacteroides  bacteria comprise  B. thetaiotamicron  or  B. fragilis.    
     
     
         7 . The method of  claim 1 , wherein the  Bacteroides  bacteria comprise  B. thetaiotamicron  and  B. fragilis.    
     
     
         8 . The method of  claim 1 , wherein:
 (i) the method stimulates Paneth cells in the gut of the subject;   (ii) the method induces Paneth cells in the gut of the subject to produce RegIII;   (iii) the method induces an immune response in the subject;   (iv) the method limits pathogenic bacterial colonization in the gut of the subject; or   (v) the method limits proliferation of gram-positive bacteria in the gut of the subject.   
     
     
         9 . The method of  claim 1 , wherein the target bacterial species is a  Firmicutes  species. 
     
     
         10 . The method of  claim 9 , wherein the  Firmicutes  species is selected from the group consisting of  Anaerotruncus, Acetanaerobacterium, Acetitomaculum, Acetivibrio, Anaerococcus, Anaerofilum, Annaerosinus, Anaerosaipes, Anaerovorax, Butyrivibrio, Clostridium, Capracoccus, Dehalobacter, Dialister, Dorea, Enterococcus, Ethanoligenens, Faecalibacterium, Fusobacterium, Gracilibacter, Guggenheimella, Hespellia, Lachnobacterium, Lachnospira, Lactobacillus, Leuconostoc, Megamonas, Morvella, Mitsuokella, Oribacterium, Oxobacter, Papillibacter, Proprionispira, Pseudoburvrivibrio, Pseudoramibacter, Roseburia, Ruminococcus, Sarcina, Seinonella, Shuttleworthia, Sporobacter, Sporobacterium, Streptococcus, Subdoligranulum, Syntrophococcus, Thermobacillus, Turibacter  and  Weisella  species. 
     
     
         11 . The method of  claim 1 , wherein the target bacterial species is selected from the group consisting of  Clostridium, Enterococcus, Ruminococcus, Streptococcus  and  Faecalibacterium  species. 
     
     
         12 . The method of  claim 1 , wherein the target bacterial species is  Clostridium dificile.    
     
     
         13 . The method of  claim 1 , wherein the target bacterial species is a gram-positive bacterial species. 
     
     
         14 . The method of  claim 1 , wherein the target bacterial species is an antibiotic resistant bacterial species. 
     
     
         15 . The method of  claim 14 , wherein the target bacterial species is selected from (i)  Staphylococcus aureus  that is resistant to an antibiotic selected from methicillin, vancomycin, linezolid, daptomycin, quinupristin, dalfopristin and teicoplanin; (ii)  Pseudomonas aeuroginosa  that is resistant to an antibiotic selected from cephalosporins, carbapenems, fluoroquinolones, aminoglycosides and colistin; (iii)  Klebsiella  species that is resistant to carbapenem; (iv)  Streptococcus  species that is resistant to an antibiotic selected from erythromycin, clindamycin, beta-lactam, macrolide, amoxicillin, azithromycin and penicillin; (v)  Salmonella  species that is resistant to an antibiotic selected from ceftriaxone, azithromycin and ciprofloxacin; (vi)  Shigella  species that is resistant to ciprofloxacin or azithromycin; (vii)  mycobacterium tuberculosis  that is resistant to an antibiotic selected from Resistance to isoniazid (INH), rifampicin (RMP), fluoroquinolone, amikacin, kanamycin, capreomycin and azithromycin; (viii)  Enterococcus  species that is resistant to vancomycin; (ix) Enterobacteriaceae species that is resistant to an antibiotic selected from cephalosporin and carbapenem; (x)  E. coli  that is resistant to an antibiotic selected from trimethoprim, itrofurantoin, cefalexin and amoxicillin; (xi)  Clostridium  species that is resistant to metronidazole (MTZ), fluoroquinolone or carbapenem; (xii)  Neisseria gonnorrhoea  species that is resistant to an antibiotic selected from cefixime, ceftriaxone, azithromycin and tetracycline; (xiii)  Acinetoebacter baumannii  that is resistant to an antibiotic selected from beta-lactam, meropenem and carbapenem; and (xiv)  Campylobacter  species that is resistant to ciprofloxacin or azithromycin. 
     
     
         16 . The method of  claim 3 , wherein the Type II HM-CRISPR/Cas system comprises an endogenous tracrRNA of the host cells. 
     
     
         17 . The method of  claim 3 , wherein the HM-CRISPR/Cas system comprises a tracrRNA, and wherein the tracrRNA is encoded by an engineered nucleic acid. 
     
     
         18 . The method of  claim 17 , wherein the engineered nucleic acid for producing the HM-crRNA encodes a single guide RNA comprising the tracrRNA and the HM-crRNA. 
     
     
         19 . The method of  claim 3 , wherein the engineered nucleic acid for producing the HM-crRNA is present in a plasmid, phage or phagemid. 
     
     
         20 . The method of  claim 19 , wherein the plasmid, phage or phagemid is present in a carrier bacteria. 
     
     
         21 . The method of  claim 3 , wherein the Cas is a Type I Cas or a Type III Cas. 
     
     
         22 . The method of  claim 3 , wherein the Cas is encoded by an engineered nucleic acid. 
     
     
         23 . The method of  claim 3 , wherein the Cas is an endogenous Cas of the host cells. 
     
     
         24 . The method of  claim 3 , wherein the target sequence is a sequence in an essential gene of the second bacterial species. 
     
     
         25 . The method of  claim 3 , wherein the engineered nucleic acid for producing the HM-crRNA encodes a R1-S1-R1′ CRISPR unit, wherein R1 is a first CRISPR repeat, R1′ is a second CRISPR repeat, and S1 is a CRISPR spacer comprising a nucleotide sequence that is 80% or more identical to a target sequence of the host cells, and wherein S1 is not comprised by the  Bacteroides  bacteria. 
     
     
         26 . The method of  claim 25 , wherein the second bacterial species is  C. dificile , and wherein R1 and R1′ each comprises SEQ ID NO: 118 or SEQ ID NO: 119. 
     
     
         27 . The method of  claim 1 , wherein the method is for treating a gastrointestinal condition selected from the group consisting of Crohn's disease, IBD, IBS and ulcerative colitis. 
     
     
         28 . The method of  claim 1 , wherein the subject is a human. 
     
     
         29 . An engineered nucleic acid encoding a R1-S1-R1′ CRISPR unit, wherein R1 is a first CRISPR repeat, R1′ is a second CRISPR repeat, and S1 is a first CRISPR spacer comprising a nucleotide sequence that is 80% or more identical to a target sequence of a  Firmicutes  host cell, wherein R1 and R1′ do not comprise any one of SEQ ID NOs: 107-111, wherein the first and second CRISPR repeats are operable with an endogenous Cas of the  Firmicutes  host cell, and wherein S1 is not comprised by  Bacteroides  bacteria. 
     
     
         30 . A pharmaceutical composition comprising a vector comprising the engineered nucleic acid of  claim 29 .