Chapter 16 : Genetic Tools for the Enhancement of Probiotic Properties

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With renewed interest in the human microbiome and its role in human health, unique opportunities for using microbes as therapeutics have recently emerged. These opportunities range from the traditional probiotics to the engineering of intestinal mutualistic bacteria to deliver therapeutic proteins ( ). However, a more in-depth understanding of how intestinal microbes impact human health and how they function in the complex, dynamic environment of the gastrointestinal tract (GIT) requires advancements in genetic tools for nonmodel strains.

Citation: Ortiz-velez L, Britton R. 2018. Genetic Tools for the Enhancement of Probiotic Properties, p 371-387. In Britton R, Cani P (ed), Bugs as Drugs. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.BAD-0018-2016
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Figure 1

Schematic representation of dsDNA recombineering in . (A) A piece of dsDNA harboring the -- cassette ( and sites, red; Cat/CM marker, yellow) and regions with homology to the genomic insertion site (H1, green; H2, blue) is transformed into expressing an exonuclease (Lp0642), a possible host nuclease inhibitor (Lp0640), and an ssDNA binding protein (Lp0641). (B) Once the dsDNA fragments are integrated into the genome, which renders the cells resistant to CM (CM), bacteria are transformed with a plasmid that induces the expression of the recombinase Cre to recombine the sites. This recombination deletes the CM marker contained inside the sites, rendering the cells CM-sensitive and leaving a modified site (72).

Citation: Ortiz-velez L, Britton R. 2018. Genetic Tools for the Enhancement of Probiotic Properties, p 371-387. In Britton R, Cani P (ed), Bugs as Drugs. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.BAD-0018-2016
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Figure 2

Schematic representation of ssDNA recombineering in . (A) An oligomer harboring the mutation (red) and regions with homology to the genomic insertion site (blue) is transformed into cells expressing RecT. (B) The oligomer is incorporated into the lagging strand being synthesized at the DNA replication fork, generating a mutant with a mixed genotype (GT). (C) Screening of the mutant is done by MAMA-PCR, using two oligomers that amplify from a WT sequence (black) and a third oligomer (red-blue) that only amplifies when the mutation is incorporated. Thus, two amplicons are generated in the case of the mutant (lane 1), whereas only one amplicon is generated in the case of the WT (lanes 2 to 5). After the mixed genotype is identified, single colony purification is done to isolate cells containing only the mutation (pure genotype).

Citation: Ortiz-velez L, Britton R. 2018. Genetic Tools for the Enhancement of Probiotic Properties, p 371-387. In Britton R, Cani P (ed), Bugs as Drugs. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.BAD-0018-2016
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Figure 3

Overview of recombineering/CRISPR-Cas9 genome engineering in . (A) An oligomer harboring the mutation (red) and regions with homology to the genomic insertion site (blue) is transformed into expressing RecT, Cas9, and its tracrRNA. (B) The oligomer targets the lagging strains and is incorporated into one of the DNA strands at the replication fork. When cells divide, a mixed population of WT and mutant cells is generated. (C) A plasmid expressing guide RNA (pgRNA) that targets the WT sequence is introduced into the cell. Cas9 is directed by the gRNA to cleave DNA of cells that did not incorporate the mutations (WT), whereas cells that incorporate the mutation are void of cleavage (mutant).

Citation: Ortiz-velez L, Britton R. 2018. Genetic Tools for the Enhancement of Probiotic Properties, p 371-387. In Britton R, Cani P (ed), Bugs as Drugs. ASM Press, Washington, DC. doi: 10.1128/microbiolspec.BAD-0018-2016
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1. Steidler L,, Hans W,, Schotte L,, Neirynck S,, Obermeier F,, Falk W,, Fiers W,, Remaut E . 2000. Treatment of murine colitis by Lactococcus lactis secreting interleukin-10. Science 289 : 13521355.[PubMed] [CrossRef]
2. Motta J-P,, Bermudez-Humaran LG,, Deraison C,, Martin L,, Rolland C,, Rousset P,, Boue J,, Dietrich G,, Chapman K,, Kharrat P,, Vinel J-P,, Alric L,, Mas E,, Sallenave J-M,, Langella P,, Vergnolle N . 2012. Food-grade bacteria expressing elafin protect against inflammation and restore colon homeostasis. Sci Transl Med 4 : 158ra144158ra144.[CrossRef]
3. Vanderhoof JA,, Whitney DB,, Antonson DL,, Hanner TL,, Lupo JV,, Young RJ . 1999. Lactobacillus GG in the prevention of antibiotic-associated diarrhea in children. J Pediatr 135 : 564568.[PubMed] [CrossRef]
4. Stiles ME,, Holzapfel WH . 1997. Lactic acid bacteria of foods and their current taxonomy. Int J Food Microbiol 36 : 129.[PubMed] [CrossRef]
5. Steidler L,, Neirynck S,, Huyghebaert N,, Snoeck V,, Vermeire A,, Goddeeris B,, Cox E,, Remon JP,, Remaut E . 2003. Biological containment of genetically modified Lactococcus lactis for intestinal delivery of human interleukin 10. Nat Biotechnol 21 : 785789.[PubMed] [CrossRef]
6. de Moreno de LeBlanc A,, Del Carmen S,, Chatel JM,, Miyoshi A,, Azevedo V,, Langella P,, Bermúdez-Humarán LG,, LeBlanc JG . 2015. Current review of genetically modified lactic acid bacteria for the prevention and treatment of colitis using murine models. Gastroenterol Res Pract 2015 : 146972.[PubMed] [CrossRef]
7. Liu X,, Lagenaur LA,, Simpson DA,, Essenmacher KP,, Frazier-Parker CL,, Liu Y,, Tsai D,, Rao SS,, Hamer DH,, Parks TP,, Lee PP,, Xu Q . 2006. Engineered vaginal lactobacillus strain for mucosal delivery of the human immunodeficiency virus inhibitor cyanovirin-N. Antimicrob Agents Chemother 50 : 32503259.[PubMed] [CrossRef]
8. Seegers JFML . 2002. Lactobacilli as live vaccine delivery vectors: progress and prospects. Trends Biotechnol 20 : 508515.[PubMed] [CrossRef]
9. Wells JM . 2011. Immunomodulatory mechanisms of lactobacilli. Microb Cell Fact 10(Suppl 1): S17.[PubMed] [CrossRef]
10. Whitehead K,, Versalovic J,, Roos S,, Britton RA . 2008. Genomic and genetic characterization of the bile stress response of probiotic Lactobacillus reuteri ATCC 55730. Appl Environ Microbiol 74 : 18121819.[PubMed] [CrossRef]
11. Daniel C,, Roussel Y,, Kleerebezem M,, Pot B . 2011. Recombinant lactic acid bacteria as mucosal biotherapeutic agents. Trends Biotechnol 29 : 499508.[PubMed] [CrossRef]
12. Bermúdez-Humarán LG,, Aubry C,, Motta JP,, Deraison C,, Steidler L,, Vergnolle N,, Chatel JM,, Langella P . 2013. Engineering lactococci and lactobacilli for human health. Curr Opin Microbiol 16 : 278283.[PubMed] [CrossRef]
13. Kok J,, van der Vossen JMBM,, Venema G . 1984. Construction of plasmid cloning vectors for lactic streptococci which also replicate in Bacillus subtilis and Escherichia coli . Appl Environ Microbiol 48 : 726731.[PubMed]
14. Weisblum B,, Graham MY,, Gryczan T,, Dubnau D . 1979. Plasmid copy number control: isolation and characterization of high-copy-number mutants of plasmid pE194. J Bacteriol 137 : 635643.[PubMed]
15. Morello E,, Bermúdez-Humarán LG,, Llull D,, Solé V,, Miraglio N,, Langella P,, Poquet I . 2008. Lactococcus lactis, an efficient cell factory for recombinant protein production and secretion. J Mol Microbiol Biotechnol 14 : 4858.[PubMed] [CrossRef]
16. Le Loir Y,, Azevedo V,, Oliveira SC,, Freitas DA,, Miyoshi A,, Bermúdez-Humarán LG,, Nouaille S,, Ribeiro LA,, Leclercq S,, Gabriel JE,, Guimaraes VD,, Oliveira MN,, Charlier C,, Gautier M,, Langella P . 2005. Protein secretion in Lactococcus lactis: an efficient way to increase the overall heterologous protein production. Microb Cell Fact 4 : 2.[PubMed] [CrossRef]
17. Bermúdez-Humarán LG . 2009. Lactococcus lactis as a live vector for mucosal delivery of therapeutic proteins. Hum Vaccin 5 : 264267.[PubMed] [CrossRef]
18. Wyszyńska A,, Kobierecka P,, Bardowski J,, Jagusztyn-Krynicka EK . 2015. Lactic acid bacteria: 20 years exploring their potential as live vectors for mucosal vaccination. Appl Microbiol Biotechnol 99 : 29672977. (Erratum, 99 : 4531. doi:10.1007/s00253-015-6569-2.)[PubMed] [CrossRef]
19. Cano-Garrido O,, Seras-Franzoso J,, Garcia-Fruitós E . 2015. Lactic acid bacteria: reviewing the potential of a promising delivery live vector for biomedical purposes. Microb Cell Fact 14 : 137.[PubMed] [CrossRef]
20. LeBlanc JG,, Aubry C,, Cortes-Perez NG,, de Moreno de LeBlanc A,, Vergnolle N,, Langella P,, Azevedo V,, Chatel JM,, Miyoshi A,, Bermúdez-Humarán LG . 2013. Mucosal targeting of therapeutic molecules using genetically modified lactic acid bacteria: an update. FEMS Microbiol Lett 344 : 19.[PubMed] [CrossRef]
21. Wells JM,, Mercenier A . 2008. Mucosal delivery of therapeutic and prophylactic molecules using lactic acid bacteria. Nat Rev Microbiol 6 : 349362.[PubMed] [CrossRef]
22. Walker DC,, Klaenhammer TR . 1994. Isolation of a novel IS3 group insertion element and construction of an integration vector for Lactobacillus spp. J Bacteriol 176 : 53305340.[PubMed] [CrossRef]
23. Raya RR,, Fremaux C,, De Antoni GL,, Klaenhammer TR . 1992. Site-specific integration of the temperate bacteriophage phi adh into the Lactobacillus gasseri chromosome and molecular characterization of the phage (attP) and bacterial (attB) attachment sites. J Bacteriol 174 : 55845592.[PubMed] [CrossRef]
24. Dupont L,, Boizet-Bonhoure B,, Coddeville M,, Auvray F,, Ritzenthaler P . 1995. Characterization of genetic elements required for site-specific integration of Lactobacillus delbrueckii subsp. bulgaricus bacteriophage mv4 and construction of an integration-proficient vector for Lactobacillus plantarum . J Bacteriol 177 : 586595.[PubMed] [CrossRef]
25. Douglas GL,, Klaenhammer TR . 2011. Directed chromosomal integration and expression of the reporter gene gusA3 in Lactobacillus acidophilus NCFM. Appl Environ Microbiol 77 : 73657371.[PubMed] [CrossRef]
26. Lambert JM,, Bongers RS,, Kleerebezem M . 2007. Cre-lox-based system for multiple gene deletions and selectable-marker removal in Lactobacillus plantarum . Appl Environ Microbiol 73 : 11261135.[PubMed] [CrossRef]
27. Vos M,, Didelot X . 2009. A comparison of homologous recombination rates in bacteria and archaea. ISME J 3 : 199208.[PubMed] [CrossRef]
28. Klaenhammer TR . 1995. Genetics of intestinal lactobacilli. Int Dairy J 5 : 10191058.[CrossRef]
29. Hols P,, Ferain T,, Garmyn D,, Bernard N,, Delcour J . 1994. Use of homologous expression-secretion signals and vector-free stable chromosomal integration in engineering of Lactobacillus plantarum for alpha-amylase and levanase expression. Appl Environ Microbiol 60 : 14011413.[PubMed]
30. Scheirlinck T,, Mahillon J,, Joos H,, Dhaese P,, Michiels F . 1989. Integration and expression of alpha-amylase and endoglucanase genes in the Lactobacillus plantarum chromosome. Appl Environ Microbiol 55 : 21302137.[PubMed]
31. Fitzsimons A,, Hols P,, Jore J,, Leer RJ,, O’Connell M,, Delcour J . 1994. Development of an amylolytic Lactobacillus plantarum silage strain expressing the Lactobacillus amylovorus alpha-amylase gene. Appl Environ Microbiol 60 : 35293535.[PubMed]
32. Russell WM,, Klaenhammer TR . 2001. Efficient system for directed integration into the Lactobacillus acidophilus and Lactobacillus gasseri chromosomes via homologous recombination. Appl Environ Microbiol 67 : 43614364.[CrossRef]
33. Law J,, Buist G,, Haandrikman A,, Kok J,, Venema G,, Leenhouts K . 1995. A system to generate chromosomal mutations in Lactococcus lactis which allows fast analysis of targeted genes. J Bacteriol 177 : 70117018.[PubMed] [CrossRef]
34. Sauer B . 1987. Functional expression of the cre-lox site-specific recombination system in the yeast Saccharomyces cerevisiae . Mol Cell Biol 7 : 20872096.[PubMed] [CrossRef]
35. Sauer B,, Henderson N . 1988. Site-specific DNA recombination in mammalian cells by the Cre recombinase of bacteriophage P1. Proc Natl Acad Sci USA 85 : 51665170.[PubMed] [CrossRef]
36. Albert H,, Dale EC,, Lee E,, Ow DW . 1995. Site-specific integration of DNA into wild-type and mutant lox sites placed in the plant genome. Plant J 7 : 649659.[PubMed] [CrossRef]
37. Nguyen TT,, Mathiesen G,, Fredriksen L,, Kittl R,, Nguyen TH,, Eijsink VGH,, Haltrich D,, Peterbauer CK . 2011. A food-grade system for inducible gene expression in Lactobacillus plantarum using an alanine racemase-encoding selection marker. J Agric Food Chem 59 : 56175624.[PubMed] [CrossRef]
38. Stevens MJA,, Vollenweider S,, Meile L,, Lacroix C . 2011. 1,3-Propanediol dehydrogenases in Lactobacillus reuteri: impact on central metabolism and 3-hydroxypropionaldehyde production. Microb Cell Fact 10 : 61.[PubMed] [CrossRef]
39. Croux C,, Nguyen N-P-T,, Lee J,, Raynaud C,, Saint-Prix F,, Gonzalez-Pajuelo M,, Meynial-Salles I,, Soucaille P . 2016. Construction of a restriction-less, marker-less mutant useful for functional genomic and metabolic engineering of the biofuel producer Clostridium acetobutylicum . Biotechnol Biofuels 9 : 23.[PubMed] [CrossRef]
40. Wang Y,, Zhang C,, Gong T,, Zuo Z,, Zhao F,, Fan X,, Yang C,, Song C . 2015. An upp-based markerless gene replacement method for genome reduction and metabolic pathway engineering in Pseudomonas mendocina NK-01 and Pseudomonas putida KT2440. J Microbiol Methods 113 : 2733.[PubMed] [CrossRef]
41. Shi T,, Wang G,, Wang Z,, Fu J,, Chen T,, Zhao X . 2013. Establishment of a markerless mutation delivery system in Bacillus subtilis stimulated by a double-strand break in the chromosome. PLoS One 8 : e81370.[PubMed] [CrossRef]
42. Goh YJ,, Azcárate-Peril MA,, O’Flaherty S,, Durmaz E,, Valence F,, Jardin J,, Lortal S,, Klaenhammer TR . 2009. Development and application of a upp-based counterselective gene replacement system for the study of the S-layer protein SlpX of Lactobacillus acidophilus NCFM. Appl Environ Microbiol 75 : 30933105.[PubMed] [CrossRef]
43. Selle K,, Goh YJ,, O’Flaherty S,, Klaenhammer TR . 2014. Development of an integration mutagenesis system in Lactobacillus gasseri . Gut Microbes 5 : 326332.[PubMed] [CrossRef]
44. Bron PA,, Benchimol MG,, Lambert J,, Palumbo E,, Deghorain M,, Delcour J,, De Vos WM,, Kleerebezem M,, Hols P . 2002. Use of the alr gene as a food-grade selection marker in lactic acid bacteria. Appl Environ Microbiol 68 : 56635670.[PubMed] [CrossRef]
45. Palumbo E,, Favier CF,, Deghorain M,, Cocconcelli PS,, Grangette C,, Mercenier A,, Vaughan EE,, Hols P . 2004. Knockout of the alanine racemase gene in Lactobacillus plantarum results in septation defects and cell wall perforation. FEMS Microbiol Lett 233 : 131138.[PubMed] [CrossRef]
46. Hols P,, Defrenne C,, Ferain T,, Derzelle S,, Delplace B,, Delcour J . 1997. The alanine racemase gene is essential for growth of Lactobacillus plantarum . J Bacteriol 179 : 38043807.[PubMed] [CrossRef]
47. Yang P,, Wang J,, Qi Q . 2015. Prophage recombinases-mediated genome engineering in Lactobacillus plantarum . Microb Cell Fact 14 : 154.[PubMed] [CrossRef]
48. van Pijkeren J-P,, Britton RA . 2012. High efficiency recombineering in lactic acid bacteria. Nucleic Acids Res 40 : e76.[PubMed] [CrossRef]
49. Oh J-H,, van Pijkeren J-P . 2014. CRISPR-Cas9-assisted recombineering in Lactobacillus reuteri . Nucleic Acids Res 42 : e131.[PubMed] [CrossRef]
50. Yang XW,, Model P,, Heintz N . 1997. Homologous recombination based modification in Escherichia coli and germline transmission in transgenic mice of a bacterial artificial chromosome. Nat Biotechnol 15 : 859865.[PubMed] [CrossRef]
51. Jiang W,, Bikard D,, Cox D,, Zhang F,, Marraffini LA . 2013. RNA-guided editing of bacterial genomes using CRISPR-Cas systems. Nat Biotechnol 31 : 233239.[PubMed] [CrossRef]
52. Court DL,, Sawitzke JA,, Thomason LC . 2002. Genetic engineering using homologous recombination. Annu Rev Genet 36 : 361388.[PubMed] [CrossRef]
53. Sharan SK,, Thomason LC,, Kuznetsov SG,, Court DL . 2009. Recombineering: a homologous recombination-based method of genetic engineering. Nat Protoc 4 : 206223.[PubMed] [CrossRef]
54. Montiel D,, Kang H-S,, Chang F-Y,, Charlop-Powers Z,, Brady SF . 2015. Yeast homologous recombination-based promoter engineering for the activation of silent natural product biosynthetic gene clusters. Proc Natl Acad Sci USA 112 : 89538958.[PubMed] [CrossRef]
55. Mosberg JA,, Lajoie MJ,, Church GM . 2010. Lambda red recombineering in Escherichia coli occurs through a fully single-stranded intermediate. Genetics 186 : 791799.[PubMed] [CrossRef]
56. Yu D,, Sawitzke JA,, Ellis H,, Court DL . 2003. Recombineering with overlapping single-stranded DNA oligonucleotides: testing a recombination intermediate. Proc Natl Acad Sci USA 100 : 72077212.[PubMed] [CrossRef]
57. Cha RS,, Zarbl H,, Keohavong P,, Thilly WG . 1992. Mismatch amplification mutation assay (MAMA): application to the c-H-ras gene. PCR Methods Appl 2 : 1420.[PubMed] [CrossRef]
58. van Pijkeren JP,, Neoh KM,, Sirias D,, Findley AS,, Britton RA . 2012. Exploring optimization parameters to increase ssDNA recombineering in Lactococcus lactis and Lactobacillus reuteri . Bioengineered 3 : 209217.[PubMed] [CrossRef]
59. al-Bar OA,, O’Connor CD,, Giles IG,, Akhtar M . 1992. d-alanine: d-alanine ligase of Escherichia coli: expression, purification and inhibitory studies on the cloned enzyme. Biochem J 282 : 747752.[PubMed] [CrossRef]
60. Kahne D,, Leimkuhler C,, Lu W,, Walsh C . 2005. Glycopeptide and lipoglycopeptide antibiotics. Chem Rev 105 : 425448.[PubMed] [CrossRef]
61. Chapot-Chartier M-P,, Kulakauskas S . 2014. Cell wall structure and function in lactic acid bacteria. Microb Cell Fact 13(Suppl 1): S9.[PubMed] [CrossRef]
62. Park IS,, Lin CH,, Walsh CT . 1996. Gain of d-alanyl-d-lactate or d-lactyl-d-alanine synthetase activities in three active-site mutants of the Escherichia coli d-alanyl-d-alanine ligase B. Biochemistry 35 : 1046410471.[PubMed] [CrossRef]
63. Il-Park IS,, Walsh CT . 1997. d-Alanyl-d-lactate and d-alanyl-d-alanine synthesis by d-alanyl-d-alanine ligase from vancomycin-resistant Leuconostoc mesenteroides: effects of a phenylalanine 261 to tyrosine mutation. J Biol Chem 272 : 92109214.[CrossRef]
64. Wang HH,, Isaacs FJ,, Carr PA,, Sun ZZ,, Xu G,, Forest CR,, Church GM . 2009. Programming cells by multiplex genome engineering and accelerated evolution. Nature 460 : 894898.[PubMed] [CrossRef]
65. Terns MP,, Terns RM . 2011. CRISPR-based adaptive immune systems. Curr Opin Microbiol 14 : 321327.[PubMed] [CrossRef]
66. Doudna JA,, Charpentier E . 2014. The new frontier of genome engineering with CRISPR-Cas9. Science 346 : 1258096.[PubMed] [CrossRef]
67. Ran FA,, Hsu PD,, Wright J,, Agarwala V,, Scott DA,, Zhang F . 2013. Genome engineering using the CRISPR-Cas9 system. Nat Protoc 8 : 22812308.[PubMed] [CrossRef]
68. Hsu PD,, Lander ES,, Zhang F . 2014. Development and applications of CRISPR-Cas9 for genome engineering. Cell 157 : 12621278.[PubMed] [CrossRef]
69. Marraffini LA . 2015. CRISPR-Cas immunity in prokaryotes. Nature 526 : 5561.[PubMed] [CrossRef]
70. Mao Z,, Bozzella M,, Seluanov A,, Gorbunova V . 2008. DNA repair by nonhomologous end joining and homologous recombination during cell cycle in human cells. Cell Cycle 7 : 29022906.[PubMed] [CrossRef]
71. Mali P,, Yang L,, Esvelt KM,, Aach J,, Guell M,, DiCarlo JE,, Norville JE,, Church GM . 2013. RNA-guided human genome engineering via Cas9. Science 339 : 823826.[PubMed] [CrossRef]
72. Brissett NC,, Doherty AJ . 2009. Repairing DNA double-strand breaks by the prokaryotic non-homologous end-joining pathway. Biochem Soc Trans 37 : 539545.[PubMed] [CrossRef]
73. Reisch CR,, Prather KLJ . 2015. The no-SCAR (Scarless Cas9 Assisted Recombineering) system for genome editing in Escherichia coli . Sci Rep 5 : 15096.[PubMed] [CrossRef]
74. Jacob F,, Monod J . 1961. Genetic regulatory mechanisms in the synthesis of proteins. J Mol Biol 3 : 318356.[CrossRef]
75. Eaton TJ,, Shearman CA,, Gasson MJ . 1993. The use of bacterial luciferase genes as reporter genes in Lactococcus: regulation of the Lactococcus lactis subsp. lactis lactose genes. J Gen Microbiol 139 : 14951501.[PubMed] [CrossRef]
76. Llull D,, Poquet I . 2004. New expression system tightly controlled by zinc availability in Lactococcus lactis . Appl Environ Microbiol 70 : 53985406.[PubMed] [CrossRef]
77. Miyoshi A,, Jamet E,, Commissaire J,, Renault P,, Langella P,, Azevedo V . 2004. A xylose-inducible expression system for Lactococcus lactis . FEMS Microbiol Lett 239 : 205212.[PubMed] [CrossRef]
78. Madsen SM,, Arnau J,, Vrang A,, Givskov M,, Israelsen H . 1999. Molecular characterization of the pH-inducible and growth phase-dependent promoter P170 of Lactococcus lactis . Mol Microbiol 32 : 7587.[PubMed] [CrossRef]
79. Kok J . 1996. Inducible gene expression and environmentally regulated genes in lactic acid bacteria. Antonie van Leeuwenhoek 70 : 129145.[PubMed] [CrossRef]
80. Benbouziane B,, Ribelles P,, Aubry C,, Martin R,, Kharrat P,, Riazi A,, Langella P,, Bermúdez-Humarán LG . 2013. Development of a stress-inducible controlled expression (SICE) system in Lactococcus lactis for the production and delivery of therapeutic molecules at mucosal surfaces. J Biotechnol 168 : 120129.[PubMed] [CrossRef]
81. O’Sullivan DJ,, Walker SA,, West SG,, Klaenhammer TR . 1996. Development of an expression strategy using a lytic phage to trigger explosive plasmid amplification and gene expression. Biotechnology (N Y) 14 : 8287.[CrossRef]
82. de Ruyter PG,, Kuipers OP,, de Vos WM . 1996. Controlled gene expression systems for Lactococcus lactis with the food-grade inducer nisin. Appl Environ Microbiol 62 : 36623667.[PubMed]
83. Eichenbaum Z,, Federle MJ,, Marra D,, de Vos WM,, Kuipers OP,, Kleerebezem M,, Scott JR . 1998. Use of the lactococcal nisA promoter to regulate gene expression in Gram-positive bacteria: comparison of induction level and promoter strength. Appl Environ Microbiol 64 : 27632769.[PubMed]
84. Mierau I,, Kleerebezem M . 2005. 10 years of the nisin-controlled gene expression system (NICE) in Lactococcus lactis . Appl Microbiol Biotechnol 68 : 705717.[PubMed] [CrossRef]
85. Wu C-M,, Lin C-F,, Chang Y-C,, Chung T-C . 2006. Construction and characterization of nisin-controlled expression vectors for use in Lactobacillus reuteri . Biosci Biotechnol Biochem 70 : 757767.[PubMed] [CrossRef]
86. Piard JC,, Hautefort I,, Fischetti VA,, Ehrlich SD,, Fons M,, Gruss A . 1997. Cell wall anchoring of the Streptococcus pyogenes M6 protein in various lactic acid bacteria. J Bacteriol 179 : 30683072.[PubMed] [CrossRef]
87. Sørvig E,, Mathiesen G,, Naterstad K,, Eijsink VGH,, Axelsson L . 2005. High-level, inducible gene expression in Lactobacillus sakei and Lactobacillus plantarum using versatile expression vectors. Microbiology 151 : 24392449.[PubMed] [CrossRef]
88. Tauer C,, Heinl S,, Egger E,, Heiss S,, Grabherr R . 2014. Tuning constitutive recombinant gene expression in Lactobacillus plantarum . Microb Cell Fact 13 : 150.[PubMed] [CrossRef]
89. Karlskås IL,, Maudal K,, Axelsson L,, Rud I,, Eijsink VGH,, Mathiesen G . 2014. Heterologous protein secretion in Lactobacilli with modified pSIP vectors. PLoS One 9 : e91125.[PubMed] [CrossRef]
90. Karimi S,, Ahl D,, Vågesjö E,, Holm L,, Phillipson M,, Jonsson H,, Roos S . 2016. In vivo and in vitro detection of luminescent and fluorescent Lactobacillus reuteri and application of red fluorescent mCherry for assessing plasmid persistence. PLoS One 11 : e0151969.[PubMed] [CrossRef]
91. Berlec A,, Ravnikar M,, Štrukelj B . 2012. Lactic acid bacteria as oral delivery systems for biomolecules. Pharmazie 67 : 891898.[PubMed]
92. Braat H,, Rottiers P,, Hommes DW,, Huyghebaert N,, Remaut E,, Remon JP,, van Deventer SJH,, Neirynck S,, Peppelenbosch MP,, Steidler L . 2006. A phase I trial with transgenic bacteria expressing interleukin-10 in Crohn’s disease. Clin Gastroenterol Hepatol 4 : 754759.[PubMed] [CrossRef]
93. Bermúdez-Humarán LG,, Kharrat P,, Chatel J-M,, Langella P . 2011. Lactococci and lactobacilli as mucosal delivery vectors for therapeutic proteins and DNA vaccines. Microb Cell Fact 10(Suppl 1): S4.[PubMed] [CrossRef]
94. Oh PL,, Benson AK,, Peterson DA,, Patil PB,, Moriyama EN,, Roos S,, Walter J . 2010. Diversification of the gut symbiont Lactobacillus reuteri as a result of host-driven evolution. ISME J 4 : 377387.[PubMed] [CrossRef]
95. Mimee M,, Tucker AC,, Voigt CA,, Lu TK . 2015. Programming a human commensal bacterium, Bacteroides thetaiotaomicron, to sense and respond to stimuli in the murine gut microbiota. Cell Syst 1 : 6271.[PubMed] [CrossRef]
96. Rud I,, Jensen PR,, Naterstad K,, Axelsson L . 2006. A synthetic promoter library for constitutive gene expression in Lactobacillus plantarum . Microbiology 152 : 10111019.[PubMed] [CrossRef]

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