Potential applications of engineered bacteria in disease diagnosis and treatment

Abstract

Probiotics are live microorganisms that confer health benefits to the host when administered in appropriate quantities. This beneficial effect has spurred extensive research in the medical and health fields. With rapid advancements in synthetic biology, the genetic and biological characteristics of a broad array of probiotics have been elucidated. Utilizing these insights, genetic editing technologies now enable the precise modification of probiotics, leading to the development of engineered bacteria. Emerging evidence underscores the significant potential of these engineered bacteria in disease management. This review explores the methodologies for creating engineered bacteria, their preliminary applications in healthcare, and the mechanisms underlying their functions. Engineered bacteria are being developed for roles such as in vivo drug delivery systems, biosensors, and mucosal vaccines, thereby contributing to the treatment, diagnosis, and prevention of conditions including inflammatory bowel disease (IBD), metabolic disorders, cancer, and neurodegenerative diseases. The review concludes by assessing the advantages and limitations of engineered bacteria in the context of disease management.

Keywords

Engineered bacteria / probiotics / synthetic biology / IBD / cancer / biosensors / mucosal vaccines

References

[1] Society of The Chinese Institute of Food Science A, Technology. Scientific consensus on probiotics (2020).J Chin Inst Food Sci Technol2020;20:303-7

[2] Suez J,Segal E.The pros, cons, and many unknowns of probiotics.Nat Med2019;25:716-29

[3] Chowdhury S,Coker C,Arpaia N.Programmable bacteria induce durable tumor regression and systemic antitumor immunity.Nat Med2019;25:1057-63 PMCID:PMC6688650

[4] Ma J,Liu X.Engineered probiotics.Microb Cell Fact2022;21:72 PMCID:PMC9044805

[5] Gurbatri CR,Vincent R.Engineered probiotics for local tumor delivery of checkpoint blockade nanobodies.Sci Transl Med2020;12:eaax0876 PMCID:PMC7685004

[6] Li M,Zhu J.Engineered probiotics with sustained release of interleukin-2 for the treatment of inflammatory bowel disease after oral delivery.Biomaterials2024;309:122584

[7] Hu H,Liu Y.Improvement effect of a next-generation probiotic L. plantarum-pMG36e-GLP-1 on type 2 diabetes mellitus via the gut-pancreas-liver axis.Food Funct2023;14:3179-95

[8] Fang X,Miao Y,Wei J.Therapeutic effect of GLP-1 engineered strain on mice model of Alzheimer’s disease and Parkinson’s disease.AMB Express2020;10:80 PMCID:PMC7182653

[9] Gurbatri CR,Vrbanac L.Engineering tumor-colonizing E. coli Nissle 1917 for detection and treatment of colorectal neoplasia.Nat Commun2024;15:646 PMCID:PMC10799955

[10] Britton RA,Khoruts A.Probiotics and the microbiome-how can we help patients make sense of probiotics?.Gastroenterology2021;160:614-23

[11] Yadav AK,Kumar A.Adhesion of Lactobacilli and their anti-infectivity potential.Crit Rev Food Sci Nutr2017;57:2042-56

[12] Breukink E,van Kraaij C,Sahl HG.Use of the cell wall precursor lipid II by a pore-forming peptide antibiotic.Science1999;286:2361-4

[13] Hols P,Gabant P.Mobilization of microbiota commensals and their bacteriocins for therapeutics.Trends Microbiol2019;27:690-702

[14] Tiwari SK. Bacteriocin-producing probiotic lactic acid bacteria in controlling dysbiosis of the gut microbiota.Front Cell Infect Microbiol2022;12:851140 PMCID:PMC9149203

[15] Bender MJ,Phelps CM.Dietary tryptophan metabolite released by intratumoral Lactobacillus reuteri facilitates immune checkpoint inhibitor treatment.Cell2023;186:1846-62.e26 PMCID:PMC10148916

[16] Martin FP,Berger B.Metabolome-associated psychological comorbidities improvement in irritable bowel syndrome patients receiving a probiotic.Gut Microbes2024;16:2347715 PMCID:PMC11085950

[17] Srivastava S,Naghibi M.A randomized double-blind, placebo-controlled trial to evaluate the safety and efficacy of live Bifidobacterium longum CECT 7347 (ES1) and heat-treated Bifidobacterium longum CECT 7347 (HT-ES1) in participants with diarrhea-predominant irritable bowel syndrome.Gut Microbes2024;16:2338322 PMCID:PMC11028008

[18] Zhang Q,Zhao W.Efficacy of Bifidobacterium animalis subsp. lactis BL-99 in the treatment of functional dyspepsia: a randomized placebo-controlled clinical trial.Nat Commun2024;15:227 PMCID:PMC10764899

[19] Han Y,Xu X.Improvement of post-surgery constipation in patients with fractures by Lactobacillus rhamnosus JYLR-127: a single-blind randomized controlled trial.Nutrients2024;16:1505 PMCID:PMC11123980

[20] Koyama S, Fujita H, Shimosato T, et al; Yokohama Cooperative Study Group for Hematology (YACHT). Septicemia from Lactobacillus rhamnosus GG, from a probiotic enriched yogurt, in a patient with autologous stem cell transplantation. Probiotics Antimicro Prot 2019;11:295-8.

[21] Rossi F,Gasperi M.Lactobacilli infection case reports in the last three years and safety implications.Nutrients2022;14:1178 PMCID:PMC8954171

[22] Kassam Z,Ramakrishna B.Donor screening for fecal microbiota transplantation.N Engl J Med2019;381:2070-2

[23] Nasrollahzadeh A,Khomeiri M.Antifungal preservation of food by lactic acid bacteria.Foods2022;11:395 PMCID:PMC8834354

[24] Zuo F,Hammarström L.Inducible plasmid self-destruction (IPSD) assisted genome engineering in lactobacilli and bifidobacteria.ACS Synth Biol2019;8:1723-9

[25] Bober JR,Nair NU.Synthetic biology approaches to engineer probiotics and members of the human microbiota for biomedical applications.Annu Rev Biomed Eng2018;20:277-300 PMCID:PMC6100750

[26] Wu J,Kong J.Genetic tools for the development of recombinant lactic acid bacteria.Microb Cell Fact2021;20:118 PMCID:PMC8214781

[27] Seyedian SS,Malamir MD.A review of the diagnosis, prevention, and treatment methods of inflammatory bowel disease.J Med Life2019;12:113-22 PMCID:PMC6685307

[28] Kaplan GG.The global burden of IBD: from 2015 to 2025.Nat Rev Gastroenterol Hepatol2015;12:720-7

[29] Alipour M,Valcheva R.Mucosal barrier depletion and loss of bacterial diversity are primary abnormalities in paediatric ulcerative colitis.J Crohns Colitis2016;10:462-71 PMCID:PMC4946763

[30] Danne C,Marteyn B.Neutrophils: from IBD to the gut microbiota.Nat Rev Gastroenterol Hepatol2024;21:184-97

[31] Smith PM,Panikov N.The microbial metabolites, short-chain fatty acids, regulate colonic Treg cell homeostasis.Science2013;341:569-73 PMCID:PMC3807819

[32] Lautenschläger C,Fischer D.Drug delivery strategies in the therapy of inflammatory bowel disease.Adv Drug Deliv Rev2014;71:58-76

[33] Yasmin F,Shaikh S.Novel drug delivery systems for inflammatory bowel disease.World J Gastroenterol2022;28:1922-33 PMCID:PMC9150062

[34] Steidler L,Schotte L.Treatment of murine colitis by Lactococcus lactis secreting interleukin-10.Science2000;289:1352-5

[35] Steidler L,Huyghebaert N.Biological containment of genetically modified Lactococcus lactis for intestinal delivery of human interleukin 10.Nat Biotechnol2003;21:785-9

[36] Braat H,Hommes DW.A phase I trial with transgenic bacteria expressing interleukin-10 in Crohn’s disease.Clin Gastroenterol Hepatol2006;4:754-9

[37] Jiang R,Zhou Y.Strategies to overcome the challenges of low or no expression of heterologous proteins in Escherichia coli.Biotechnol Adv2024;75:108417

[38] Plotkin JB.Synonymous but not the same: the causes and consequences of codon bias.Nat Rev Genet2011;12:32-42 PMCID:PMC3074964

[39] Hanson ML,Li W.Oral delivery of IL-27 recombinant bacteria attenuates immune colitis in mice.Gastroenterology2014;146:210-21.e13 PMCID:PMC3920828

[40] Zhou J,Chen Q.Programmable probiotics modulate inflammation and gut microbiota for inflammatory bowel disease treatment after effective oral delivery.Nat Commun2022;13:3432 PMCID:PMC9198027

[41] Chen H,Ji H.Escherichia coli Nissle 1917 ghosts alleviate inflammatory bowel disease in zebrafish.Life Sci2023;329:121956

[42] Qin J,Cai Z.A metagenome-wide association study of gut microbiota in type 2 diabetes.Nature2012;490:55-60

[43] Zhou W,Contrepois K.Longitudinal multi-omics of host-microbe dynamics in prediabetes.Nature2019;569:663-71 PMCID:PMC6666404

[44] Tilg H.Microbiota and diabetes: an evolving relationship.Gut2014;63:1513-21

[45] Rittiphairoj T,Janchot K,Li T.Probiotics contribute to glycemic control in patients with type 2 diabetes mellitus: a systematic review and meta-analysis.Adv Nutr2021;12:722-34 PMCID:PMC8166562

[46] Wang Y,Wu Y,Sun X.Composite probiotics alleviate type 2 diabetes by regulating intestinal microbiota and inducing GLP-1 secretion in db/db mice.Biomed Pharmacother2020;125:109914

[47] Holst JJ.Glucagonlike peptide 1: a newly discovered gastrointestinal hormone.Gastroenterology1994;107:1848-55

[48] Sandoval DA.Physiology of proglucagon peptides: role of glucagon and GLP-1 in health and disease.Physiol Rev2015;95:513-48

[49] Grandl G,Dimarchi R,Müller TD.Gut peptide agonism in the treatment of obesity and diabetes. Compr Physiol 2019;10:99-124.

[50] Agarwal P,Billack B,Shao J.Oral delivery of glucagon like peptide-1 by a recombinant Lactococcus lactis.Pharm Res2014;31:3404-14

[51] Luo J,Lu J,Chen T.Antidiabetic effect of an engineered bacterium Lactobacillus plantarum-pMG36e -GLP-1 in monkey model.Synth Syst Biotechnol2021;6:272-82 PMCID:PMC8455315

[52] Wang L,Wang H.Engineered bacteria of MG1363-pMG36e-GLP-1 attenuated obesity-induced by high fat diet in mice.Front Cell Infect Microbiol2021;11:595575 PMCID:PMC7959774

[53] Takiishi T,Van Belle TL.Reversal of autoimmune diabetes by restoration of antigen-specific tolerance using genetically modified Lactococcus lactis in mice.J Clin Invest2012;122:1717-25 PMCID:PMC3336982

[54] Chavkin TA,Kostic A.E. coli Nissle 1917 modulates host glucose metabolism without directly acting on glucose.Sci Rep2021;11:23230 PMCID:PMC8636602

[55] Bazi Z,Hekmatdoost A.The long term oral regulation of blood glucose in diabetic patients by using of Escherichia coli Nissle 1917 expressing CTB-IGF-1 hybrid protein.Med Hypotheses2013;81:961-2

[56] Tu P,Wang H.Expression of CTB-10×rolGLP-1 in E. coli and its therapeutic effect on type 2 diabetes.Curr Pharm Biotechnol2015;16:564-72

[57] Qin Q,Li Y.Intestinal microbiota play an important role in the treatment of type I diabetes in mice with BefA protein.Front Cell Infect Microbiol2021;11:719542 PMCID:PMC8485065

[58] Wang H,Hu H.Oral administration of bacterial β cell expansion factor A (BefA) alleviates diabetes in mice with type 1 and type 2 diabetes.Oxid Med Cell Longev2022;2022:9206039 PMCID:PMC8853770

[59] Blau N,Langenbeck U.Diagnosis, classification, and genetics of phenylketonuria and tetrahydrobiopterin (BH4) deficiencies.Mol Genet Metab2011;104 Suppl:S2-9

[60] Spronsen FJ, Blau N, Harding C, Burlina A, Longo N, Bosch AM. Phenylketonuria.Nat Rev Dis Primers2021;7:36 PMCID:PMC8591558

[61] Lichter-Konecki U.Phenylketonuria: current treatments and future developments.Drugs2019;79:495-500

[62] van Spronsen FJ,Ahring K.Key European guidelines for the diagnosis and management of patients with phenylketonuria.Lancet Diabetes Endocrinol2017;5:743-56

[63] Martinez M,Schwank G.State-of-the-art 2023 on gene therapy for phenylketonuria.J Inherit Metab Dis2024;47:80-92 PMCID:PMC10764640

[64] Kim W,Surendran S.Trends in enzyme therapy for phenylketonuria.Mol Ther2004;10:220-4

[65] Levy HL,Scriver CR.Phenylalanine ammonia lyase (PAL): from discovery to enzyme substitution therapy for phenylketonuria.Mol Genet Metab2018;124:223-9

[66] Durrer KE,Hunt von Herbing I.Genetically engineered probiotic for the treatment of phenylketonuria (PKU); assessment of a novel treatment in vitro and in the PAHenu2 mouse model of PKU.PLoS One2017;12:e0176286 PMCID:PMC5435137

[67] Ramírez AM,Ardila A.Production of human recombinant phenylalanine hydroxylase in Lactobacillus plantarum for gastrointestinal delivery.Eur J Pharm Sci2017;109:48-55

[68] Phumkhachorn P.A broad host range food-grade cloning vector for lactic acid bacteria.Biologia2016;71:457-63

[69] Kaur T,Kaur B.Correction to: construction of a shuttle expression vector for lactic acid bacteria.J Genet Eng Biotechnol2020;18:38 PMCID:PMC7403355

[70] Platteeuw C,van Schalkwijk S.Food-grade cloning and expression system for Lactococcus lactis.Appl Environ Microbiol1996;62:1008-13 PMCID:PMC167865

[71] Kim JH.Improvement of a nisin-inducible expression vector for use in lactic acid bacteria.Plasmid2007;58:275-83

[72] Chae JP,Hwang IC.Construction of a Bile-responsive expression system in Lactobacillus plantarum.Food Sci Anim Resour2019;39:13-22 PMCID:PMC6413156

[73] Guan C,Ma Y.Development of a novel expression system in lactic acid bacteria controlled by a broad-host-range promoter P_srfA.Microb Cell Fact2022;21:23 PMCID:PMC8845276

[74] Isabella VM,Castillo MJ.Development of a synthetic live bacterial therapeutic for the human metabolic disease phenylketonuria.Nat Biotechnol2018;36:857-64

[75] Jiang Y,Qian F.Expression of phenylalanine ammonia lyase as an intracellularly free and extracellularly cell surface-immobilized enzyme on a gut microbe as a live biotherapeutic for phenylketonuria.Sci China Life Sci2023;66:127-36 PMCID:PMC9362719

[76] Puurunen MK,Searle SL.Safety and pharmacodynamics of an engineered E. coli Nissle for the treatment of phenylketonuria: a first-in-human phase 1/2a study.Nat Metab2021;3:1125-32

[77] Adolfsen KJ,Monahan CE.Improvement of a synthetic live bacterial therapeutic for phenylketonuria with biosensor-enabled enzyme engineering.Nat Commun2021;12:6215 PMCID:PMC8553829

[78] Triassi AJ,Monahan CE.Redesign of an Escherichia coli Nissle treatment for phenylketonuria using insulated genomic landing pads and genetic circuits to reduce burden.Cell Syst2023;14:512-24.e12

[79] Heineman HS,Cooper WM.Hodgkin’s disease and salmonella typhimurium infection.JAMA1964;188:632-4

[80] Stritzker J,Hill PJ,Goebel W.Tumor-specific colonization, tissue distribution, and gene induction by probiotic Escherichia coli Nissle 1917 in live mice.Int J Med Microbiol2007;297:151-62

[81] Toso JF,Hwu P.Phase I study of the intravenous administration of attenuated Salmonella typhimurium to patients with metastatic melanoma.J Clin Oncol2002;20:142-52

[82] Chondrou P,Kiousi DE.Lactobacillus paracasei K5 displays adhesion, anti-proliferative activity and apoptotic effects in human colon cancer cells.Benef Microbes2018;9:975-83

[83] Leschner S,Dietrich N.Tumor invasion of Salmonella enterica serovar Typhimurium is accompanied by strong hemorrhage promoted by TNF-alpha.PLoS One2009;4:e6692 PMCID:PMC2724709

[84] Kasinskas RW.Salmonella typhimurium lacking ribose chemoreceptors localize in tumor quiescence and induce apoptosis.Cancer Res2007;67:3201-9

[85] Kwon SY,Son J,Min JJ.Exploiting bacteria for cancer immunotherapy.Nat Rev Clin Oncol2024;21:569-89

[86] Saito S,Maekawa T,Okuno A.Lactococcus lactis subsp. cremoris C60 upregulates macrophage function by modifying metabolic preference in enhanced anti-tumor immunity.Cancers2024;16:1928 PMCID:PMC11120145

[87] Herrero-Fresno A.Salmonella Typhimurium metabolism affects virulence in the host - a mini-review.Food Microbiol2018;71:98-110

[88] Low KB,Le T.Lipid A mutant Salmonella with suppressed virulence and TNFalpha induction retain tumor-targeting in vivo.Nat Biotechnol1999;17:37-41

[89] Clairmont C,Pike J.Biodistribution and genetic stability of the novel antitumor agent VNP20009, a genetically modified strain of Salmonella typhimurium.J Infect Dis2000;181:1996-2002

[90] Song M,Kim EY.ppGpp-dependent stationary phase induction of genes on Salmonella pathogenicity island 1.J Biol Chem2004;279:34183-90

[91] Na HS,Lee HC,Rhee JH.Immune response induced by Salmonella typhimurium defective in ppGpp synthesis.Vaccine2006;24:2027-34

[92] Sieow BF,Yong WP,Chang MW.Tweak to treat: reprograming bacteria for cancer treatment.Trends Cancer2021;7:447-64

[93] Park SH,Nguyen VH.RGD peptide cell-surface display enhances the targeting and therapeutic efficacy of attenuated salmonella-mediated cancer therapy.Theranostics2016;6:1672-82 PMCID:PMC4955065

[94] Massa PE,Monegal A,Rescigno M.Salmonella engineered to express CD20-targeting antibodies and a drug-converting enzyme can eradicate human lymphomas.Blood2013;122:705-14

[95] Wu D,Liu H.Escherichia coli Nissle 1917-driven microrobots for effective tumor targeted drug delivery and tumor regression.Acta Biomater2023;169:477-88

[96] Zhu J,Liu Q.Engineered Lactococcus lactis secreting Flt3L and OX40 ligand for in situ vaccination-based cancer immunotherapy.Nat Commun2022;13:7466 PMCID:PMC9719518

[97] Zhou DX,Xu X.Anti-tumor effects of engineered VNP20009-Abvec-Igκ-mPD-1 strain in melanoma mice via combining the oncolytic therapy and immunotherapy.Pharmaceutics2022;14:2789 PMCID:PMC9781615

[98] Ciaćma K,Kędracka-Krok S.Secretion of tumoricidal human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) by recombinant Lactococcus lactis: optimization of in vitro synthesis conditions.Microb Cell Fact2018;17:177 PMCID:PMC6238363

[99] Yoon W,Chae YS,Kim BM.Therapeutic advantage of genetically engineered Salmonella typhimurium carrying short hairpin RNA against inhibin alpha subunit in cancer treatment.Ann Oncol2018;29:2010-7

[100] Zheng JH,Jiang SN.Two-step enhanced cancer immunotherapy with engineered Salmonella typhimurium secreting heterologous flagellin.Sci Transl Med2017;9:eaak9537

[101] Tan W,Zuo C.Targeting of pancreatic cancer cells and stromal cells using engineered oncolytic Salmonella typhimurium.Mol Ther2022;30:662-71 PMCID:PMC8821930

[102] Satoh Y,Iida Y,Notsu Y.Supplementation of l-arginine boosts the therapeutic efficacy of anticancer chemoimmunotherapy.Cancer Sci2020;111:2248-58 PMCID:PMC7484823

[103] Canale FP,Antonini G.Metabolic modulation of tumours with engineered bacteria for immunotherapy.Nature2021;598:662-6

[104] Reich N.The neuroprotective effects of glucagon-like peptide 1 in Alzheimer’s and Parkinson’s disease: an in-depth review.Front Neurosci2022;16:970925 PMCID:PMC9475012

[105] Diz-Chaves Y,Spuch C,Mallo F.Anti-inflammatory effects of GLP-1 receptor activation in the brain in neurodegenerative diseases.Int J Mol Sci2022;23:9583 PMCID:PMC9455625

[106] Nowell J,Edison P.Incretin and insulin signaling as novel therapeutic targets for Alzheimer’s and Parkinson’s disease.Mol Psychiatry2023;28:217-29 PMCID:PMC9812772

[107] Batista AF,De Felice FG.Neuroprotective actions of glucagon-like peptide-1 (GLP-1) analogues in Alzheimer’s and Parkinson’s diseases.CNS Drugs2019;33:209-23

[108] Wang RF,Hölscher C.Post-treatment with the GLP-1 analogue liraglutide alleviate chronic inflammation and mitochondrial stress induced by Status epilepticus.Epilepsy Res2018;142:45-52

[109] Loh JS,Tan LKS.Microbiota-gut-brain axis and its therapeutic applications in neurodegenerative diseases.Signal Transduct Target Ther2024;9:37 PMCID:PMC10869798

[110] Chen T,Huang Z.Engineered commensal bacteria prevent systemic inflammation-induced memory impairment and amyloidogenesis via producing GLP-1.Appl Microbiol Biotechnol2018;102:7565-75

[111] Yue M,Chen W,Chen T.Neurotrophic role of the next-generation probiotic strain L. lactis MG1363-pMG36e-GLP-1 on Parkinson’s disease via inhibiting ferroptosis.Nutrients2022;14:4886 PMCID:PMC9698534

[112] Fang X,Zhao X,Chen T.Neuroprotective effects of an engineered commensal bacterium in the 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine Parkinson disease mouse model via producing glucagon-like peptide-1.J Neurochem2019;150:441-52

[113] Mukherjee S.Bacterial quorum sensing in complex and dynamically changing environments.Nat Rev Microbiol2019;17:371-82 PMCID:PMC6615036

[114] Gupta RK,Lee CY.RNAIII of the Staphylococcus aureus agr system activates global regulator MgrA by stabilizing mRNA.Proc Natl Acad Sci U S A2015;112:14036-41 PMCID:PMC4653210

[115] Lubkowicz D,Hwang IY,Lee YS.Reprogramming probiotic Lactobacillus reuteri as a biosensor for staphylococcus aureus derived AIP-I detection.ACS Synth Biol2018;7:1229-37

[116] Li H,Qi Q.Engineered probiotic Lactobacillus plantarum WCSF I for monitoring and treatment of Staphylococcus aureus infection.Microbiol Spectr2023;11:e0182923 PMCID:PMC10848683

[117] Mao N,Cameron DE.Probiotic strains detect and suppress cholera in mice.Sci Transl Med2018;10:eaao2586 PMCID:PMC7821980

[118] Borrero J,Dunny GM.Modified lactic acid bacteria detect and inhibit multiresistant enterococci.ACS Synth Biol2015;4:299-306 PMCID:PMC4384838

[119] Strimbu K.What are biomarkers?.Curr Opin HIV AIDS2010;5:463-6 PMCID:PMC3078627

[120] Mimee M,Hayward A.An ingestible bacterial-electronic system to monitor gastrointestinal health.Science2018;360:915-8 PMCID:PMC6430580

[121] Zou ZP,Fang TT,Ye BC.Biomarker-responsive engineered probiotic diagnoses, records, and ameliorates inflammatory bowel disease in mice.Cell Host Microbe2023;31:199-212.e5

[122] Xia JY,Tran P,Bass J.Engineered calprotectin-sensing probiotics for IBD surveillance in humans.Proc Natl Acad Sci U S A2023;120:e2221121120 PMCID:PMC10410751

[123] Meng T,Fan M,Duan X.Enhancing the contrast of tumor imaging for image-guided surgery using a tumor-targeting probiotic with the continuous expression of a biomarker.Anal Chem2022;94:10109-17

[124] Danino T,Kwong GA.Programmable probiotics for detection of cancer in urine.Sci Transl Med2015;7:289ra84 PMCID:PMC4511399

[125] Tanniche I.Engineered live bacteria as disease detection and diagnosis tools.J Biol Eng2023;17:65 PMCID:PMC10598922

[126] Perdigon G,Nader DE Macias ME,de Ruiz Holgado AP.The oral administration of lactic acid bacteria increase the mucosal intestinal immunity in response to enteropathogens.J Food Prot1990;53:404-10

[127] Hanniffy SB,Hitchin E.Mucosal delivery of a pneumococcal vaccine using Lactococcus lactis affords protection against respiratory infection.J Infect Dis2007;195:185-93

[128] Lee JS,Han DP.Mucosal immunization with surface-displayed severe acute respiratory syndrome coronavirus spike protein on Lactobacillus casei induces neutralizing antibodies in mice.J Virol2006;80:4079-87 PMCID:PMC1440448

[129] Xu YXY,Weng MM.Oral immunization of mice with recombinant Lactobacillus plantarum expressing a Trichinella spiralis galectin induces an immune protection against larval challenge.Parasit Vectors2022;15:475 PMCID:PMC9764493

[130] Zhang R,Duan G.An engineered Lactococcus lactis strain exerts significant immune responses through efficient expression and delivery of Helicobacter pylori Lpp20 antigen.Biotechnol Lett2016;38:2169-75

[131] Zhang F,Zhang Z.Recombinant L. lactis vaccine LL-plSAM-WAE targeting four virulence factors provides mucosal immunity against H. pylori infection.Microb Cell Fact2024;23:61 PMCID:PMC10893618

[132] Huynh DT,Chathuranga K,Kim CJ.Mucosal administration of Lactobacillus casei surface-displayed HA1 induces protective immune responses against avian influenza A virus in mice.J Microbiol Biotechnol2024;34:735-45 PMCID:PMC11016770

[133] Sarnelli G,Palenca I.Intranasal administration of Escherichia coli Nissle expressing the spike protein of SARS-CoV-2 induces long-term immunization and prevents spike protein-mediated lung injury in mice.Biomed Pharmacother2024;174:116441

[134] Zhao Z,Zhang D.Oral vaccination with recombinant Lactobacillus casei expressing Aeromonas hydrophila Aha1 against A. hydrophila infections in common carps.Virulence2022;13:794-807 PMCID:PMC9067532

[135] Zhang Y,Zhang J.Oral or intranasal immunization with recombinant Lactobacillus plantarum displaying head domain of Swine Influenza A virus hemagglutinin protects mice from H1N1 virus.Microb Cell Fact2022;21:185 PMCID:PMC9461438

[136] Qiao N,Zhong X.Recombinant lactic acid bacteria as promising vectors for mucosal vaccination.Exploration2021;1:20210026 PMCID:PMC10191043

[137] Guo S,McDonough SP.The recombinant Lactococcus lactis oral vaccine induces protection against C. difficile spore challenge in a mouse model.Vaccine2015;33:1586-95

[138] Sarate PJ,Poiret S.E. coli Nissle 1917 is a safe mucosal delivery vector for a birch-grass pollen chimera to prevent allergic poly-sensitization.Mucosal Immunol2019;12:132-44

[139] Rao S,McHugh L.Toward a live microbial microbicide for HIV: commensal bacteria secreting an HIV fusion inhibitor peptide.Proc Natl Acad Sci U S A2005;102:11993-8 PMCID:PMC1189328

[140] Huang L,He L.Engineered probiotic Escherichia coli elicits immediate and long-term protection against influenza A virus in mice.Nat Commun2024;15:6802 PMCID:PMC11315933

[141] Mohseni AH,Keyvani H,Khavari-Nejad RA.Oral immunization with recombinant Lactococcus lactis NZ9000 expressing human papillomavirus type 16 E7 antigen and evaluation of its immune effects in female C57BL/6 mice.J Med Virol2019;91:296-307

[142] Li Y,Liu H,Yang J.Intranasal immunization with recombinant Lactococci carrying human papillomavirus E7 protein and mouse interleukin-12 DNA induces E7-specific antitumor effects in C57BL/6 mice.Oncol Lett2014;7:576-82 PMCID:PMC3881950

[143] Benz I.Cloning and expression of an adhesin (AIDA-I) involved in diffuse adherence of enteropathogenic Escherichia coli.Infect Immun1989;57:1506-11 PMCID:PMC313306

[144] Buddenborg C,Liebrecht S,Humberg V.Development of a tripartite vector system for live oral immunization using a gram-negative probiotic carrier.Int J Med Microbiol2008;298:105-14

[145] Sarnelli G,Pesce M.Oral Immunization with Escherichia coli Nissle 1917 expressing SARS-CoV-2 spike protein induces mucosal and systemic antibody responses in mice.Biomolecules2023;13:569 PMCID:PMC10046078

[146] Remer KA,Roeger B,Sonnenborn U.Split immune response after oral vaccination of mice with recombinant Escherichia coli Nissle 1917 expressing fimbrial adhesin K88.Int J Med Microbiol2009;299:467-78

[147] Wen LJ,Wang GH.Immunization with recombinant Lactobacillus casei strains producing K99, K88 fimbrial protein protects mice against enterotoxigenic Escherichia coli.Vaccine2012;30:3339-49

[148] Huang Y,Yu S,Chen W.Intestinal engineered probiotics as living therapeutics: chassis selection, colonization enhancement, gene circuit design, and biocontainment.ACS Synth Biol2022;11:3134-53

[149] Li MC.IL-10 and its related cytokines for treatment of inflammatory bowel disease.World J Gastroenterol2004;10:620-5 PMCID:PMC4716896

[150] Yang M,Li L.Bile acid-gut microbiota axis in inflammatory bowel disease: from bench to bedside.Nutrients2021;13:3143 PMCID:PMC8467364

[151] Wang Y,Peng X.Biochemotaxis-oriented engineering bacteria expressing GLP-1 enhance diabetes therapy by regulating the balance of immune.Adv Healthc Mater2024;13:e2303958

[152] Hausmann O,Longo N.Pegvaliase: Immunological profile and recommendations for the clinical management of hypersensitivity reactions in patients with phenylketonuria treated with this enzyme substitution therapy.Mol Genet Metab2019;128:84-91

[153] Mater DD,Corthier G.A probiotic Lactobacillus strain can acquire vancomycin resistance during digestive transit in mice.J Mol Microbiol Biotechnol2008;14:123-7

[154] Anisimova E,Karimullina G.Alarming antibiotic resistance of Lactobacilli isolated from probiotic preparations and dietary supplements.Antibiotics2022;11:1557 PMCID:PMC9686474

[155] Castro CP, Drumond MM, Batista VL, Nunes A, Mancha-Agresti P, Azevedo V. Vector development timeline for mucosal vaccination and treatment of disease using Lactococcus lactis and design approaches of next generation food grade plasmids.Front Microbiol2018;9:1805 PMCID:PMC6102412

[156] Charbonneau MR,Li N.Developing a new class of engineered live bacterial therapeutics to treat human diseases.Nat Commun2020;11:1738 PMCID:PMC7142098

[157] Yan X,Zhao C.Applications of synthetic biology in medical and pharmaceutical fields.Signal Transduct Target Ther2023;8:199 PMCID:PMC10173249

[158] Gibson DG,Lartigue C.Creation of a bacterial cell controlled by a chemically synthesized genome.Science2010;329:52-6

[159] Sung BH,Kim SC.Construction of a minimal genome as a chassis for synthetic biology.Essays Biochem2016;60:337-46

[160] Calero P.Chasing bacterial chassis for metabolic engineering: a perspective review from classical to non-traditional microorganisms.Microb Biotechnol2019;12:98-124 PMCID:PMC6302729

[161] Zhu D,Liu F,Saris PE.Enhanced heterologous protein productivity by genome reduction in Lactococcus lactis NZ9000.Microb Cell Fact2017;16:1 PMCID:PMC5210298

[162] Hille F,Wong SP,Ressel S.The biology of CRISPR-Cas: backward and forward.Cell2018;172:1239-59

[163] Hryhorowicz M,Zeyland J.CRISPR/Cas9 immune system as a tool for genome engineering.Arch Immunol Ther Exp2017;65:233-40 PMCID:PMC5434172

[164] Xiao Y,Nam KH.How type II CRISPR-Cas establish immunity through Cas1-Cas2-mediated spacer integration.Nature2017;550:137-41 PMCID:PMC5832332

[165] Kleinstiver BP,Tsai SQ.Engineered CRISPR-Cas9 nucleases with altered PAM specificities.Nature2015;523:481-5 PMCID:PMC4540238

[166] Kim H.A guide to genome engineering with programmable nucleases.Nat Rev Genet2014;15:321-34

[167] Gaj T,Barbas CF 3rd.ZFN, TALEN, and CRISPR/Cas-based methods for genome engineering.Trends Biotechnol2013;31:397-405 PMCID:PMC3694601

[168] Ren J,Na D.Recent advances in genetic engineering tools based on synthetic biology.J Microbiol2020;58:1-10

[169] Gaudelli NM,Rees HA.Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage.Nature2017;551:464-71 PMCID:PMC5726555

[170] Anzalone AV,Davis JR.Search-and-replace genome editing without double-strand breaks or donor DNA.Nature2019;576:149-57 PMCID:PMC6907074

[171] Peters JE.Tn7: smarter than we thought.Nat Rev Mol Cell Biol2001;2:806-14

[172] Crawley AB,Stout E,Barrangou R.Characterizing the activity of abundant, diverse and active CRISPR-Cas systems in lactobacilli.Sci Rep2018;8:11544 PMCID:PMC6070500

[173] Pan M,Hidalgo-Cantabrana C.Genomic and epigenetic landscapes drive CRISPR-based genome editing in Bifidobacterium.Proc Natl Acad Sci U S A2022;119:e2205068119 PMCID:PMC9335239

[174] Song X,Xiong Z,Yang S.CRISPR-Cas9^D10A Nickase-assisted genome editing in Lactobacillus casei.Appl Environ Microbiol2017;83:e01259-17 PMCID:PMC5666132

[175] Myrbråten IS,Salehian Z.CRISPR interference for rapid knockdown of essential cell cycle genes in Lactobacillus plantarum.mSphere2019;4:e00007-19 PMCID:PMC6429040

[176] Hidalgo-Cantabrana C,Pan M,Barrangou R.Genome editing using the endogenous type I CRISPR-Cas system in Lactobacillus crispatus.Proc Natl Acad Sci U S A2019;116:15774-83 PMCID:PMC6690032

[177] Han X,Chen H.Harnessing the endogenous Type I-C CRISPR-Cas system for genome editing in Bifidobacterium breve.Appl Environ Microbiol2024;90:e0207423 PMCID:PMC10952402

[178] Ninyio N,Sergon G,Andersson S.Stable expression of HIV-1 MPER extended epitope on the surface of the recombinant probiotic bacteria Escherichia Coli Nissle 1917 using CRISPR/Cas9.Microb Cell Fact2024;23:39 PMCID:PMC10840157

[179] Zhou Y,Yang H.Construction of a food-grade gene editing system based on CRISPR-Cas9 and its application in Lactococcus lactis NZ9000.Biotechnol Lett2023;45:955-66

[180] Yu J,Zhang S.Design of a self-driven probiotic-CRISPR/Cas9 nanosystem for sono-immunometabolic cancer therapy.Nat Commun2022;13:7903 PMCID:PMC9780327

[181] Tian X,Zhang Y.Metabolic engineering coupled with adaptive evolution strategies for the efficient production of high-quality L-lactic acid by Lactobacillus paracasei.Bioresour Technol2021;323:124549

[182] Oh JH.CRISPR-Cas9-assisted recombineering in Lactobacillus reuteri.Nucleic Acids Res2014;42:e131 PMCID:PMC4176153

[183] Fang M,Wang C.Engineering probiotic Escherichia coli Nissle 1917 to block transfer of multiple antibiotic resistance genes by exploiting a type I CRISPR-Cas system.Appl Environ Microbiol2024;90:e0081124

[184] Lan Y,Cheng S.Development of Escherichia coli Nissle 1917 derivative by CRISPR/Cas9 and application for gamma-aminobutyric acid (GABA) production in antibiotic-free system.Biochem Eng J2021;168:107952

[185] Luo W,Zhou D.Deep tumor penetration of CRISPR-Cas system for photothermal-sensitized immunotherapy via probiotics.Nano Lett2023;23:8081-90

[186] Yang Y,Liu Y.GEDpm-cg: genome editing automated design platform for point mutation construction in corynebacterium glutamicum.Front Bioeng Biotechnol2021;9:768289 PMCID:PMC8554027

[187] Chien T,Kepecs B.Enhancing the tropism of bacteria via genetically programmed biosensors.Nat Biomed Eng2022;6:94-104 PMCID:PMC8956018

[188] Hryhorowicz M,Zeyland J.Evolution of CRISPR/Cas systems for precise genome editing.Int J Mol Sci2023;24:14233 PMCID:PMC10532350