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Bacterial Pathogenesis > Lecture 1 > Flashcards

Lecture 1 Flashcards

1
Q

What is a Disease?

A

Clinical signs and symptoms of damage that occur in a host as a result of its interaction with an infectious agent

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2
Q

What factors effect a host’s likelihood of infection?

A

Immune status (age, stress, diet and previous illness)
Prior Exposure
Genetic Predisposition

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3
Q

What effects a bacteria’s ability to initiate infection?

A

Site of initial infection
Specific traits of bacterial isolate (virulence factors, metabolism, growth characteristics)
Route of Inoculum
Size of inoculum

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4
Q

What does LD50 stand for?

A

Lethal dose 50 of inoculum: dosage that causes 50% mortality in animal model

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5
Q

What was the aim of Koch’s Postulates?

A

Proving that a specific microorganism causes a specific disease

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6
Q

What are Koch’s four postulates?

A

1) The microorganism must be found in abundance in all organisms suffering from the disease, but should not be found in healthy organisms
2) The microorganism must be isolated from diseased organism and grown in pure culture
3) The cultures organism should cause disease when introduced into a healthy organism
4) The microorganism must be re-isolated from the inoculated, diseased experimental host and identified as being identical to the original specific causative agent.

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7
Q

What are the implicit assumptions of Koch’s postulates?

A

1) The pathogen is culturable (in the lab)
2) The right condition are being used
- Nutrients
- O2 levels

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8
Q

Give an example of a disease which could not be cultured in the lab and therefore appeared to go against Koch’s postulates
And describe how the microorganism was eventually found

A

Cat Scratch Disease: Required 45 days to Culture
Discovered using 16S rRNA
-DNA extracted from tissue shown to have visble clusters when stained
-DNA was amplified using 16S rRNA eubacterial primer for PCR
-The DNA was cloned and sequenced

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9
Q

What are the three main types of bacterial pathogens?

A

Extracellular Invasive Pathogens
Extracellular Toxin Producing Pathogens
Intracellular Pathogens (non-obligates)

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10
Q

In order the establish and infection and cause disease pathogens must have strategies to facilitate what 6 processes?

A

1) Attachment and entry in the body
2) Local or general spread in the body
3) Multiplication
4) Evasion of host defences
5) Shedding from body
6) Causing damage to the host

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11
Q

What is used to facilitate all of those processes?

A

Virulence Factors

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12
Q

Give seven examples of virulence factors

A 
Adhesins
Flagella 
Proteins to help obtain rare/essential nutrients 
Toxins 
Capsule 
Immune Modulatory Proteins
Type III Secretory System
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13
Q

What was the first virulence factor cloned? What was it’s use?

A

Invasin - cloned in 1984, from Yersis pseudotuberculosis
Gave ability to avoid immune system
-Highlighted by conferring ability onto non-pathogenic Escherichia coli

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14
Q

What is an Endotoxin?

A 
A lipopolysaccharide from Gram -ve bacteria
Cell bound
Heat Stable
Causes fever, diarrhoea and vomiting 
Weakly toxic
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15
Q

What is an exotoxin?

A

Protein Released extracellulary by certain Gram +ve and -ve bacteria
Heat Labile
Highly toxic
Very Specific

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16
Q

What is an enterotoxin?

Give examples

A

Group of exotoxins that act specifically on the small intestine , causing changes in intestinal permeability leading to diarrhoea e.g. Clodtridium difficile toxin A, cholera toxin and Escherichia coli toxins

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17
Q

What do you call a toxin inactivated for use as a vaccine?

A

Toxoid

18
Q

Name a bacterium species which produces the most potent neurotoxin
Describe its action and how it is now used in clinical settings

A

Clostridium botulinum - a Gram +ve, anaerobic spore former
(1ng can kill 1 million guinea pigs)
Blocks acetylcholine release in synapses causing flaccid paralysis and respiratory arrest
Used for relief of spasticity, chronic migraine, excessive sweating, overactive bladder and botox

19
Q

How was Invasin confirmed as a single gene?

A

Large Y. tuberculosis genome fragments cloned in E.coli, clones enriched to allow E.coli to invade host cells, gene pinpointed using transposon mutagenesis; loss of invasion capability due to loss of single gene

20
Q

What three additional postulates were added for Molecular Koch’s postulates?

A

1) Phenotype associated with pathogenic strains of species
2) Inactivation of gene should lead to measurable reduction in virulence or if the is gene isolated will be able to confer trait
3) Replacing mutant gene with wild type should restore virulence

21
Q

What bacteria releases the cholera toxin?
How is it transmitted?
Where does it colonize?
What effect does it have?
What additional virulence factors does this bacterium employ?

A 
Vibrio cholerae 
Water-oral
Small intestine
The exotoxin causes increased adenylate cyclase activity; increasing cAMP levels and effects sodium/chloride flux in/out of cell (see Microbiology module for more). This leads to loss of fluid and electrolytes (aka diarrhoea)
Adhesin, Mucinase, Flagella
22
Q

Where did the cholera toxin come from?

A

Part of genome of lysogenic phage

23
Q

What is TCP?

What is its role?

A

Toxin co-regulated pilus

Required for colonization and acts as receptor for other phage

24
Q

What is a pathogen?

A

An organism capable of causing disease

25
Q

Name and describe the role of 6 virulence factors seen in some Streptococcus pneumoniae

A

Capsule: Prevents Phagocytosis
Surface Adhesions: Attach to respiratory lining
Secretory IgA protease: Cleaves IgA and promotes spread of infection
Neuroaminidase: Cleaves terminal acetylated neuraminic acids from sugar residues
Pneumolysins: Pore forming; tissue damage

26
Q

What is the microbiome?

A

The ecologocial community of commensals which live in the human body; provide benefits to us.

27
Q

What are the benefits of a healthy microbiome?

A

Stimulate the immune system
Prevent colonization of pathogens; by out competing for nutrients
Produce beneficial nutrients for the host

28
Q

What is metagenomics? What are the benefits?

A

The study of genetic material recovered directly from environmental samples
No need to cultivate organisms

28
Q

Why will defining the healthy micro biome help in the understanding of disease?

A

Easier to notice differences in micro biome and correlate these to diseases

29
Q

Which areas of the body should be free of microbes?

A 
Blood
Cerebrospinal fluid 
Urine(not so much now)
Muscles 
Brain
30
Q

How many bacterial species exist in a healthy micro biome in the mouth?
And how many individual bacterial cells are in the healthy micro biome of each hand?

A

400 Species

10,000 - 10 million

31
Q

Name the five areas in the human body which make up the micro biome

A 
Anterior Nares
Buccal Mucosa 
Supragingival Plaque
Distal Gut
Posterior Fornix
32
Q

Explain the mouse experiment example

A

When the microbiota from the gut of an obese twin and the microbiota from the gut of a lean twin are transplanted into mice the mouse receiving the obese biota will have increased adiposity; even on same diet.
Possible for the lean microbiota to invade the mice with obese biiota and stop increased adiposity; but not the reverse

33
Q

How can commensals cause disease?

A

Enter parts of the body which should be sterile

overgrow if normal balance is disturbed (eg. by pH imbalanced mucus and immune deficiency)

34
Q

What effect has antibiotic use had on Clostridium difficile?

A

A normal gut flora member which flourishes under antibiotic selective pressure and leads to antibiotic-associated diarrhoea

35
Q

Would these commensals that cause disease be classed as pathogenic?

A

Yes - if they fulfil Koch’s postulates

36
Q

What is the modern ‘supplement’ to Koch’s postulates?

A

In some cases, human disease is more apt to reflect a ‘disturbed microbial community structure’ instead of the singular mischief of a single pathogenic species

37
Q

What is a overt or strict pathogen?
What is a opportunistic pathogen?
What is a facultative pathogen?

A

Only associated with human disease, and are not found as members of normal, healthy microbial flora
Members of the normal flora that only cause disease when introduces into unprotected sites
Can grow and survive in environment as well as in host (only become pathogens due to accidental entry into host)

38
Q

What three ways can be used to identify genes needed to survive* during infection?

A

1) Differential fluorescence induction
- Clone random promoters to express GFP when active
- screen for promoters active only during infection
2) Random mutagenesis
- 1000’s of mutants made: challenge to track and achieve
3) Signature Tagged Mutagenesis
- Create unique DNA tags with RE sites for insertion into plasmid and removal of invariant arms
- Incorporate each each into transposon plasmid
- Pool transposons (input pool) and inject into animal model
- Mutants that survive used for output pool
- 96 well plate of input and output pools compared
- Mutants that did not survive animal model screen had important survival gene

39
Q

What did STM show about Vibrio cholera?

A

Experiment 1: 1100 mutants, in pools of 48, showed 51 attenuated strains
Experiment 2: 9600 mutants, in pools of 96, showed 251 attenuated strains, many metabolism genes affected
These 251 were screened for acid tolerance - 9 genes essential for acid survival and colonisation

40
Q

What is the competitive index?

A

Ratio of mutant to wild type
When below 1 = competitive disadvantage to mutant
When above 1 = competitive advantage to mutant

41
Q

What is TraDis?

A

Transposon Directed Insertion site Sequencing
Deep sequencing of the unique transposon flanking sequences
High coverage and quantitative information

Bacterial Pathogenesis (9 decks)

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