2: Which pathogen strain to use?

Question 1

What is Cystic Fibrosis?

Answer 1

In Cystic Fibrosis (CF) mucus clogs the lungs and leads to chronic respiratory infections. It can be caused by infection with Pseudomonas aeruginosa.

Question 2

What are the pro’s of using a sequenced strain?

Answer 2

The sequenced strain is known so the genetics and proteomics is easier, and we can exchange data/materials with other labs.

Question 3

What are the con’s of using a sequenced strain?

Answer 3

You have to be careful for attenuation by in vitro growth. This could induce mutations. Also keep in mind genome plasticity, each strain is different and has its own specifications and perhaps different virulence factors (max 10% difference).

Question 4

What is attenuation by in vitro growth?

Answer 4

most ‘workhorses’ have been around for a long time and have been frequently subcultured. This could induce mutations (in virulence factors). Attenuated strains can be used for vaccines (because they can be less virulent after attenuation).

Question 5

When are bacteria called a species?

Answer 5

When there is 98% 16S rDNA identity. Except when there are imporant metabolic/pathogenic differences!

Question 6

What is horizontal gene transfer?

Answer 6

It is exchanging parts of genomes by bacteria (not mixing genomes!), by replacing genes or introducing new genes.

Question 7

What are the causes of horizontal gene transfer?

Answer 7

Transduction, transformation and conjugation.

Question 8

What is transduction?

Answer 8

Bacteriophages mediate transfer of DNA between bacteria, whereby DNA from a donor bacterium is packages into a virus particle and transferred into a recipient bacterium during infection. Is only extremely efficient for transferring phage DNA, not chromosomal DNA.

Question 9

What is transformation?

Answer 9

Some bacteria are able to take up free DNA from the environment and incorporate it into their chromosome. This is extremely efficient.

Question 10

What is conjugation?

Answer 10

The mechanism of gene transfer responsible for the most concerning aspects of antimicrobial resistance. A sex pilus forms between two bacterial cells through which a plasmid is transferred from one to the other. This is extremely efficient and the transfer of chromosomal DNA is possible.

Question 11

When is recombination required and what is it dependent on?

Answer 11

Recombination is required after transformation and transduction. It is dependent on the RecA protein and DNA homology (high homology, increased recombination efficiency).

Question 12

How can you determine horizontal gene transfer?

Answer 12

By G-C nucleotide content, codon usage and gene order/syntheny.

Question 13

Why is horizontal gene transfer more frequent between closely related strains/species?

Answer 13

Because of high homology, so there is no difference between codon usage and G-C nucleotide content (threfore probably underestimated).

Question 14

What is an example of a virulene plasmid?

Answer 14

Virulence of Mycobacterium ulcerans is dependent on production of mycolactone. Enzymes for mycolactone production are encoded by a plasmid.

Question 15

What are integrated bacteriophages?

Answer 15

Integrated bacteriophages are also responsible for virulence, like with Vibrio cholera where the causative agent of cholera is cholera toxin on bacteriophages (so it’s easily transferred).

Question 16

What are pathogenicity islands?

Answer 16

Pathogenicity islands are clusters of contiguous genes that are present in some related strains but not in others. They are acquired by horizontal gene transfer. They code for distinctive virulence characteristics and are often associated with transfer RNA loci. Some are excisable, so it can be transferred somewhere else.

Question 17

What is the example of pathogenicity islands in Salmonella?

Answer 17

In Salmonella the type III secretion systems, which are high precision virulence factors ,are endcoded on pathogenicity islands.

Question 18

What is the core genome?

Answer 18

The common gene pool of a bacteria species.

Question 19

What is the pan-genome?

Answer 19

All genes that are present in at least one of the strains of a species.

Question 20

What is the difference between an open pangenome and a closed pangenome?

Answer 20

Some bacterial species are considered to have an unlimited large gene repertoire (open pangenome) and other species seem to be limited by a maximum number of genes in their gene pool (closed pangenome).

Question 21

What are the limits to horizontal gene transfer?

Answer 21

Gene acquisition will also lead to gene loss, not much of basal genes but mostly of newly acquired genes.

Question 22

What is genetic headroom?

Answer 22

The amount of dispensable information in the genome. This determines genetic flexibilty and evolutionary potential.

Question 23

What should you be aware of when using the sequenced strain?

Answer 23

You can use the standard strain but be aware of differences and check also other clones in your assays. Determine the genome sequence of your optimal virulent strain.

Question 24

Why do RNA viruses have a higher error rate?

Answer 24

because they have no proofreading activity of RNA-dependent RNA polymerase.

Question 25

What are the pro’s and con’s of DNA viruses?

Answer 25

Consequences for DNA viruses are lower variability in sequence, but more possibilities to incorporate new genes, including host genes.

Question 26

What are quasispecies?

Answer 26

A viral quasispecies is a population structure of RNA viruses with a large numbers of variant genomes (mutant cloud). Quasispecies result from high mutation rates as mutants arise continually and change in relative frequency as viral replication and selection proceeds.

Question 27

What is the quasispecies effect?

Answer 27

Quasispecies infection leads to ways for RNA viruses to escape the immune system because they have so many different forms.

Question 28

What are the pro’s and con’s of RNA viruses?

Answer 28

Consequences for RNA viruses are that RNA viruses evolve very quickly, however, they hardly acquire new genes and therefore also limited new (virulence) traits. Genetic headroom is ery small/absent: no room for gene acquisition. Despite this, all recently emerged viruses are RNA viruses.