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Probiotics > Module 12 - Genetic Tools > Flashcards

Module 12 - Genetic Tools Flashcards

1
Q

LAB and Bifidobacterium as Delivery Sytems (3)

A

GRAS status + live mucosal vectors for production and delivery of therapeutic molecules + survive stress

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2
Q

LAB and Bifidobacterium survive

A

Low pH of stomach and bile salts

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3
Q

Survive harsh condtions up to 48 hours

A

L. lactis

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4
Q

Persist 4 and 7 days under harsh conditions

A

Lactobacillus and Bifidobacterium

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5
Q

Pioneer bacterium

A

L. lactis

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6
Q

Best studied from physiological aspect

A

L. lactis

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7
Q

Why poineer (8)

A

Several sequences available + GRAS + model for mucosal delivery + high transformation via electroporation + does not colonize GI tract + several genetic tools + simple secretion system + best studies from physiological aspect

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8
Q

No intestinal colonization

A

Non-invasive

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9
Q

L. lactis genetic tools

A

Plasmids and Expression Systems

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10
Q

Lactobacillus as delivery vector

A

Persist longer than Lactococcus in GI tract + might be part of native microbiota and have interacted with host IS

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11
Q

Very difficult to transform and genetically manipulate

A

Lactobacillus and Bifidobacterium

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12
Q

Bifidobacterium as new delivery vector

A

Similar advantages as Lactobacillus

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13
Q

Nisin-Controlled Gene Expression (NICE) Expression Vector

A

Nisin –> binds NisK on membrane –> ST to NisR –> regulates gene expression of PnisZ –> makes device protein

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14
Q

Based on expression of heterologous proteins from nisZ promoter

A

Nisin-Controlled Gene Expression (NICE)

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15
Q

NisK and NisR are

A

Regulatory Proteins

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16
Q

NICE mostly in…

A

Bacillus subtilis

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17
Q

Increasing NICE in

A

L. lactic and L. plantarum

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18
Q

Zinc Inducible Expression Systems

A

PZNZitR-driven Heterologous Expression and Xirex

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19
Q

Xirex

A

Expression highly inducible but protein quality low

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20
Q

Xylose-inducible expression vector (XIES)

A

Cytoplasmic and secreted proteins + high amounts

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21
Q

pSIP

A

Similar to NICE for expression of cytoplasmic, cell wall anchoring, and secretion proteins

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22
Q

SICE and P170

A

Self-regulated promoters for environmental conditions

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23
Q

Controlled by pGroEL Chaperon promoter

A

Stress-inducible Controlled Expression (SICE)

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24
Q

SICE

A

pGroEL chaperon promoter controlled where a variety of conditions produce highly inducible proteins via constitutive expression

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25
P170 System
pH inducible of pH 6 or below
26
High recombinant protein production
P170 system
27
pSIP System proteins
SapIP protein triggers membrane protein sapK which triggers sapR and acts on PsapA to get POI
28
Vectors for mucosal delivery of molecules...
Requires mastering thief targeting mode
29
Protein location important for
Efficiency of treatment of therapeutic molecule and IR modulation
30
Three main expression vectors (signal peptides)
pCYT, pSEC, pCWA
31
Extra signal peptide
Fusion to cell wall
32
pCTY
Cytoplasmic expression
33
pSEC
Proteins fused to a signal peptide
34
pCWA
anchors to cell wall
35
Anti-protease Roles
Prevent inflammation and restore mucosal homeostasis
36
Elafin Secretion
Inhibits high proteolytic activity, CXCL-8, MCP-1, and IP-10 + increase intestinal integrity
37
Anti-protease
Elafin Secretion
38
Secreted SOD
Enzyme produced that prevent neutrophil and Mac ROS release
39
DNA Delivery Roles
Production of epithelial cells or POI + transfer DNA depending on capacity of LAB to do so
40
Enzyme protein expression roles
Caecal and colonic inflammation decreases
41
Plasmid transfer
DNA delivery to produce protein expression of eukaryotic cells
42
Cytokine and Antigen POI Role
Long term diabetes reversion in NOD mice + Local T-reg increase
43
Cytokines and antigens produced by recLAB
IL-10 and pro-insulin
44
Mechanism of IL-10 and Pro-insulin
Interact with Tregs to make functional after migration to intestine
45
IBD
Group of gut disorders characterised by uncontrollable and chronic inflammation of GIT
46
Two major forms of IBD
Crohn's disease and UC
47
IBD Characterised by
Microbiota disturbances + Significant production of inflammatory Mediators
48
Inflammatory Mediators of IBD
Protease, ROS, and cytokines
49
Prevent IBD via expression of
Elafin, antioxidant enzymes (SOD), and cytokines from Lactococcus and Lactobacillus
50
Bacteria used to treat IBD
Lactococcus and Lactobacillus
51
Elafin produced by LAB
Oral administration in mice --> reduced local inflammation and epithelial damage to restrole colon homeostasis
52
SOD
Antioxidant enzyme
53
SOD (superoxide dismutase)
Converts superoxide anion to H2O2
54
Present in very few LABs
SOD
55
Produce SOD to protect mice
L. plantarum and L. Lactis --> colitis protection
56
Enzyme expression that is similar to SOD
CAT
57
Production of IL-10 leads to
Reduced inflammation
58
Reduced inflammation in DSS-inducible colitis mice models
recombinant L. lactis producing IL10
59
Recombinant L. lactis producing IL-10
Reduce inflammation in colitis mouse models + decrease activity of Crohn's disease in human trial
60
IL27 production
Reduce inflammation in DSS-iduced colitis mouse models
61
IL27 produced by
L. lactis
62
IL27 mechanism
Th1 response induced --> suppress Th17 and Th2 differentiation + induce IL-10
63
Allergy to Cow's Milk
Common in young infants due to response to milk proteins such as B-lactoglobulin (BLG)
64
B-galactoglobulin (BLG)
Cow's milk protein
65
Treatment of milk allergy
Express BLG in L. lactis --> BLG-specfic IgA in mice --> protect from further sensitization
66
Asthma
Chronic inflammatory disease correlated with airway inflammation driven by Th2 cytokines
67
Drivers of asthma
Th2 cytokines IL4 and IL5 and reduced Th1 response
68
Allergic Airway Diseases
Asthma
69
Asthma reduction via
Expression of Th1 inducer IL12 by L. lactis in OVA-induced asthma --> switch from Th2 to Th1 in mice
70
T1DM
Autoimmune disease characterised by destruction of pancreatic Beta cells
71
Heterologous Protein Expression for T1DM
Produce auto-antigenic peptide in NOD model of diabetes in mice
72
Human insulin gene
Express in murine model as auto-antigenic peptide
73
Auto-antigenic peptide
Human insulin gene
74
Autoantigens that help reduce DM1 (4)
Human insulin gene, glutamic acid decarboxylase GAD-65, HSP65, P277
75
Celiac Disease
Autoimmune disease characterised by insufficient tolerance toward gliadin
76
Gliadin
Component of gluten
77
Celiac Disease Protein Expression
Deaminated gliadin epitope carried by L. lactis can suppress local and systemic responses + Elafin
78
Elafin role in celiac disease reduction
Inhibit TG-2 activity to restore intestinal barrier and reduce inflammation
79
TG-2
Involved in deamination of gliadin
80
Mucosal Delivery of Bacterial Antigens
S. mutant PAc + Tatanus toxin (TTFC) + Listeriolysin O
81
Surface protein antigen (Pac)
On S. mutant and first antigen tested for carries prevention by increasing salivary IgA and IgG
82
Tetanus toxoid (TTFC)
L. lactis expression of fragment C of TT produced by C. tetani
83
Listeriolysin O-secreted
L. lactis --> Pore-forming toxin LLO
84
Only IP and not Oral route
LLO expression
85
HPV-16 E7 Antigen
Administer L. lactis IN --> specific humoral and cellular responses
86
HPV bacteria
L. lactis, L. plantarum, L. casei
87
HPV better results when...
Protein anchored on the surface of bacteria
88
Rotavirus
Most common cause of diarrhea in infants and young children
89
Rotavirus LAB
L. lactis and L. casei
90
Rotavirus antigen delivered
NSP4 and VP7
91
H5N1
Avian Influenza Virus with a 60% mortality rate in humans
92
H5N1 Antigens
HA via L. lactis --> HA-specfic serum IgG and fecal IgA
93
Parasitic diseases and LAB
Malaria + Giardiasis + Crytosporidiosis
94
Malaria
L. lactis expressing fusion protein (MSP3 and GLIRP from parasite) --> high levels of antibodies in mice
95
Giardiasis (Giardia intestinalis)
Water borne parasite that disseminates as cysts
96
Giardiasis treatment
Use cyst form antigen expressed in L. lactis
97
Giardiasis antigen
CWP2
98
Cryptosporidosis
Waster borne disease mainly by Cryptosporidium parvum that can be lethal in immunosuppressed hosts
99
Crytosporidiosis treatment
L. casei expressing P23 ahesion protein --> IgG and IgA

Probiotics (16 decks)

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