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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
Q

P170 System

A

pH inducible of pH 6 or below

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26
Q

High recombinant protein production

A

P170 system

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27
Q

pSIP System proteins

A

SapIP protein triggers membrane protein sapK which triggers sapR and acts on PsapA to get POI

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28
Q

Vectors for mucosal delivery of molecules…

A

Requires mastering thief targeting mode

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29
Q

Protein location important for

A

Efficiency of treatment of therapeutic molecule and IR modulation

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30
Q

Three main expression vectors (signal peptides)

A

pCYT, pSEC, pCWA

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31
Q

Extra signal peptide

A

Fusion to cell wall

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32
Q

pCTY

A

Cytoplasmic expression

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33
Q

pSEC

A

Proteins fused to a signal peptide

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34
Q

pCWA

A

anchors to cell wall

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35
Q

Anti-protease Roles

A

Prevent inflammation and restore mucosal homeostasis

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36
Q

Elafin Secretion

A

Inhibits high proteolytic activity, CXCL-8, MCP-1, and IP-10 + increase intestinal integrity

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37
Q

Anti-protease

A

Elafin Secretion

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38
Q

Secreted SOD

A

Enzyme produced that prevent neutrophil and Mac ROS release

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39
Q

DNA Delivery Roles

A

Production of epithelial cells or POI + transfer DNA depending on capacity of LAB to do so

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40
Q

Enzyme protein expression roles

A

Caecal and colonic inflammation decreases

41
Q

Plasmid transfer

A

DNA delivery to produce protein expression of eukaryotic cells

42
Q

Cytokine and Antigen POI Role

A

Long term diabetes reversion in NOD mice + Local T-reg increase

43
Q

Cytokines and antigens produced by recLAB

A

IL-10 and pro-insulin

44
Q

Mechanism of IL-10 and Pro-insulin

A

Interact with Tregs to make functional after migration to intestine

45
Q

IBD

A

Group of gut disorders characterised by uncontrollable and chronic inflammation of GIT

46
Q

Two major forms of IBD

A

Crohn’s disease and UC

47
Q

IBD Characterised by

A

Microbiota disturbances + Significant production of inflammatory Mediators

48
Q

Inflammatory Mediators of IBD

A

Protease, ROS, and cytokines

49
Q

Prevent IBD via expression of

A

Elafin, antioxidant enzymes (SOD), and cytokines from Lactococcus and Lactobacillus

50
Q

Bacteria used to treat IBD

A

Lactococcus and Lactobacillus

51
Q

Elafin produced by LAB

A

Oral administration in mice –> reduced local inflammation and epithelial damage to restrole colon homeostasis

52
Q

SOD

A

Antioxidant enzyme

53
Q

SOD (superoxide dismutase)

A

Converts superoxide anion to H2O2

54
Q

Present in very few LABs

A

SOD

55
Q

Produce SOD to protect mice

A

L. plantarum and L. Lactis –> colitis protection

56
Q

Enzyme expression that is similar to SOD

A

CAT

57
Q

Production of IL-10 leads to

A

Reduced inflammation

58
Q

Reduced inflammation in DSS-inducible colitis mice models

A

recombinant L. lactis producing IL10

59
Q

Recombinant L. lactis producing IL-10

A

Reduce inflammation in colitis mouse models + decrease activity of Crohn’s disease in human trial

60
Q

IL27 production

A

Reduce inflammation in DSS-iduced colitis mouse models

61
Q

IL27 produced by

A

L. lactis

62
Q

IL27 mechanism

A

Th1 response induced –> suppress Th17 and Th2 differentiation + induce IL-10

63
Q

Allergy to Cow’s Milk

A

Common in young infants due to response to milk proteins such as B-lactoglobulin (BLG)

64
Q

B-galactoglobulin (BLG)

A

Cow’s milk protein

65
Q

Treatment of milk allergy

A

Express BLG in L. lactis –> BLG-specfic IgA in mice –> protect from further sensitization

66
Q

Asthma

A

Chronic inflammatory disease correlated with airway inflammation driven by Th2 cytokines

67
Q

Drivers of asthma

A

Th2 cytokines IL4 and IL5 and reduced Th1 response

68
Q

Allergic Airway Diseases

A

Asthma

69
Q

Asthma reduction via

A

Expression of Th1 inducer IL12 by L. lactis in OVA-induced asthma –> switch from Th2 to Th1 in mice

70
Q

T1DM

A

Autoimmune disease characterised by destruction of pancreatic Beta cells

71
Q

Heterologous Protein Expression for T1DM

A

Produce auto-antigenic peptide in NOD model of diabetes in mice

72
Q

Human insulin gene

A

Express in murine model as auto-antigenic peptide

73
Q

Auto-antigenic peptide

A

Human insulin gene

74
Q

Autoantigens that help reduce DM1 (4)

A

Human insulin gene, glutamic acid decarboxylase GAD-65, HSP65, P277

75
Q

Celiac Disease

A

Autoimmune disease characterised by insufficient tolerance toward gliadin

76
Q

Gliadin

A

Component of gluten

77
Q

Celiac Disease Protein Expression

A

Deaminated gliadin epitope carried by L. lactis can suppress local and systemic responses + Elafin

78
Q

Elafin role in celiac disease reduction

A

Inhibit TG-2 activity to restore intestinal barrier and reduce inflammation

79
Q

TG-2

A

Involved in deamination of gliadin

80
Q

Mucosal Delivery of Bacterial Antigens

A

S. mutant PAc + Tatanus toxin (TTFC) + Listeriolysin O

81
Q

Surface protein antigen (Pac)

A

On S. mutant and first antigen tested for carries prevention by increasing salivary IgA and IgG

82
Q

Tetanus toxoid (TTFC)

A

L. lactis expression of fragment C of TT produced by C. tetani

83
Q

Listeriolysin O-secreted

A

L. lactis –> Pore-forming toxin LLO

84
Q

Only IP and not Oral route

A

LLO expression

85
Q

HPV-16 E7 Antigen

A

Administer L. lactis IN –> specific humoral and cellular responses

86
Q

HPV bacteria

A

L. lactis, L. plantarum, L. casei

87
Q

HPV better results when…

A

Protein anchored on the surface of bacteria

88
Q

Rotavirus

A

Most common cause of diarrhea in infants and young children

89
Q

Rotavirus LAB

A

L. lactis and L. casei

90
Q

Rotavirus antigen delivered

A

NSP4 and VP7

91
Q

H5N1

A

Avian Influenza Virus with a 60% mortality rate in humans

92
Q

H5N1 Antigens

A

HA via L. lactis –> HA-specfic serum IgG and fecal IgA

93
Q

Parasitic diseases and LAB

A

Malaria + Giardiasis + Crytosporidiosis

94
Q

Malaria

A

L. lactis expressing fusion protein (MSP3 and GLIRP from parasite) –> high levels of antibodies in mice

95
Q

Giardiasis (Giardia intestinalis)

A

Water borne parasite that disseminates as cysts

96
Q

Giardiasis treatment

A

Use cyst form antigen expressed in L. lactis

97
Q

Giardiasis antigen

A

CWP2

98
Q

Cryptosporidosis

A

Waster borne disease mainly by Cryptosporidium parvum that can be lethal in immunosuppressed hosts

99
Q

Crytosporidiosis treatment

A

L. casei expressing P23 ahesion protein –> IgG and IgA

Probiotics (16 decks)

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