Lecture 11: Reverse Vaccinology

Question 1

What are 3 types of vaccines?

Answer 1

Killed microorganisms
• detoxified

Attenuated living microorganisms
• Viruses
- Recombinant viruses (Adenovirus)
• Bacteria
− related species (BCG  M. bovis)

Component vaccines -> Adjuvant needed
• Toxoids (tetanus)
• Immunogenic components (a-cellular vaccines)
− Sub-structures and surface structures (capsule/vesicles)
− Proteins, polysaccharides, RNA/DNA

Question 2

Vaccin Development: classical vs. reverse genomics approach

Answer 2

Classical: identification/selection of vaccine components by: knowledge of disease.
(− Virulence factors
− Serology
− Immune response)

Reverse genomics approach:
Identification/Selection of vaccine candidates
• Genomics
− Surface exposed
− Homology to known Virulence factors
− Antigen structure recognized by immune system

Question 3

Clinical development of classical vs reverse genomics?

Answer 3

Clinical development:

• Purification, testing immunogenicity and testing protection
• Proof safety

Question 4

Reverse Vaccinology and Genomics: differences between Classical, psn-genomic and subtractive?

Answer 4

Classical: single genome based > Identify surface exposed antigens’

1.Pan-genomic (S. pneumoniae)
> Multiple genomes
> Select shared antigens

2.Subtractive (Pathogenic E. coli):
> Multiple genomes
> Pathogenic and non-pathogenic
> Select pathogen-specific antigens

Question 5

What starts the disease of Neisseria meningitidis?

Answer 5

Entry into the bloodstream starts disease (rare cases)
- within 10 days of colonization
- only encapsulated forms (to survive in the bloodstream)
- serogroups A, B, C, Y and W (types of polysaccharides)
• Damage caused by bacterial components (LPS/endotoxin) and the response of the host defense system (shock)

Question 6

Why are Invasive strains are encapsulated ?

Answer 6

survival of complement/blood stream

Question 7

Neisseria Capsule is Major virulence factor

 why not use just purified Capsule Polysacharides?

Answer 7

c• Poorly immunogenic in children

• No immunogenic memory induced

Question 8

What can be done to get an immune reaction?

Answer 8

Capsular Polysaccharide is chemically linked to an immunogenic “carrier” protein. Then triggers immune response.

Question 9

 conjugate vaccines available for serogroups ACWY, but not B. Why?

Answer 9

Problem Serogroup B:
• capsular polysaccharide = homopolymer of sialic acid  
• also present on human neuronal cells
− poorly immunogenic
− response also leads to auto-immunity

Question 10

PorA protein = ?

Answer 10

Porin in OM of Neisseria and acts as a channel for nutrients. Exposed on surface. Raises bactericidal antibodies.

Question 11

Neisseria serogroup B vaccines: What was used?

Answer 11

OMV (de-toxified: LPS removal) + PorA types

Question 12

Why is N. . meningitidis Model for reverse vaccinology.?

Answer 12

more >190 available genomes.

Question 13

What can you search for for vaccines when looking at genome?

Answer 13

ORFs (proteins) -> functional annotation
• Homology-based
• Structure-based

Promoter sequences
Regulatory sequences

For vaccines selected: surface structures

• OMPs
• Proteins extending from cell surface

Question 14

Vaccine candidates are predicted in genome sequence of neisseria. And then? Filter for?

Answer 14

1. Surface Exposure
2. Bactericidal activity
3. Presence in disease isolates
4. Antigenic variation

Question 15

What is immunogold labeling?

Answer 15

Bining of antibodies to surface

Question 16

There is variation of antigens that are present in the vaccine. Hoewver..

Answer 16

-> However: one component enough effectivity
-> Long range protection seems possible; but has not been
proven yet

Question 17

2 methods to select antigens for vaccines. One is IVET. Explain?

Answer 17

IVET:Used to signal induction of virulence genes.

Clone library of genes in a plasmid -> let it integrate into the genome of the pathogen -> genes on plasmid can have certain markers: e.g. survival in presence of certain nutrients/antibiotic/fluorescence.
Selective marker is under control of promoter of genome of bacterium, infect the model. Add the antibiotic or nutrient: promoters that are active during this infection are thus important for infection.
Sequence pool of recovered plasmids
-> genes expressed during infection
-> select: surface exposed/immunogenic

Question 18

The other: Tn-seq in infection models. Explain?

Answer 18

• Transposon that you let jump into genes of pathogen -> library
  Bacteria that survive -> inject in model ->if gene is needed for experimental condition it needs to be expressed. If transposon is there, gene will not be (correctly) expressed, bacterium will die.
  Recover library -> sequence -> compare to as you had it before. No transposons where gene is needed.