Extenta Jan 2023

Question 1

Dysbiosis and Diversity are two terms that are often used when discussing the microbiota.
Explain the terms in detail and describe the importance of them in human health.

Answer 1

Dysbiosis is an imbalance in the microbiome

Diversity is a measure of how many different species of bacteria are present

Question 2

The bacterial taxonomy uses hierarchical levels for classification. Choose three species (of
relevance within the topics intestinal microbiota and probiotics) belonging to different classes
and name them for all levels accordingly (from phyla to species), in the correct order and
starting with the highest level. Also use upper- or lower-case letters and italics in the correct
context.

Answer 2

Firmicutes
Bacilli
Lactobacillales
/Lactobacillacea
/Lactobacillus
/L. acidophilus

Proteobacteria
Gammaproteobacteria
Enterobacteriales
/Enterobacteriacea
/Eschericia
/E. coli

Actinobacteria
Actinobacteriia
Bifidobacterilales
/Bifidobacteriacea
/Bifidobacterium
/B. bifidum

Question 3

You are going to design a clinical trial to prove that probiotics can prevent diseases. Choose
one disease or syndrome that have been discussed during the course or is mentioned in the
compendium Probiotics (Molin G., 2022).
a) Describe the study design you have chosen and suggest suitable endpoints for the trial.
Motivate your answer.
b) Suggest one suitable probiotic species for the trial and explain how it can prevent the disease
that you have chosen.
c) Describe in detail the method you would have chosen for analysing the presence of the
probiotic species in the gastrointestinal tract.
d) Describe whether there is a risk of side effects from the probiotics you have chosen for the
individuals you want to give it to, as a preventive measure to reduce the risk of disease
development.

Answer 3

IBD would be an interesting disease from a probiotics viewpoint.
a) Randomised controlled trial. Two groups. One receives probiotics and the other a placebo. Endpoints for the trial would be a change in pain level/discomfort/bowel movement/flatulence
b) Lactobacillus plantarum 299V
c) Sanger sequencing using specific primers
d) There shouldn’t really be any risks, only that the first few days may feel a little different

Question 4

Which are the typical dominating Gram-negative, and Gram-positive phyla that can be found
frequently in the intestinal tract of healthy human individuals? Name two Gram-negative and
one Gram-positive phyla.

Answer 4

Bacteroidetes (gram negative)
Proteobacteria (gram negative)
Firmicutes (gram positive)

Question 5

All Gram-negative bacteria have pro-inflammatory compounds associated to the cell wall,
what is it called?

Answer 5

LPS lipopolysaccharides contain lipid A which is an endotoxin

Question 6

Describe shortly (processes and involved cells and organs) how an inflammatory process can
be initiated in the small intestine by the microbiota.

Answer 6

Peyer’s patches present in the small intestine and contain specialised epithelial cells called M cells that sample the intestine for antigens. Dendritic cells are also present in the peyers patches and can also sample. These antigens are presented to T cells. Turn T cells into regulatory T cells. M cells can transport antigens via transcytosis. Dendritic cells can stick their dendrites through epithelial cells to sample.

Question 7

Long-term inflammation is a risk factor for several diseases, name three diseases.

Answer 7

Inflammatory bowel’s disease. Autoimmune diseases. Metabolic diseases such as type 2 diabetes.

Question 8

T/F: Cytokines are always pro-inflammatory mediators.

Answer 8

False. Can also be anti-inflammatory

Question 9

T/F: All strains from the same species will act the same way

Answer 9

False
Example
HEAL9 and 299V have different properties

Question 10

T/F: Effects of probiotic treatments are dose-specific.

Answer 10

Need to be an adequate dose

Question 11

T/F: Next generation sequencing is only detecting live cells.

Answer 11

False, it is about genetic material

Question 12

T/F: Translocation is the passage of viable cells through the intestinal wall.

Answer 12

True

Question 13

T/F: The genus Enterococcus but not Bifidobacterium are sometimes wrongly regarded as
LAB.

Answer 13

Enterococcus is a LAB but not bifidobacterium. So must be false.

Question 14

T/F: Staphylococcus belongs to the phylum Firmicutes

Answer 14

True

Question 15

T/F: Through phenotypic classification organisms are grouped together based on similar
observable characteristics.

Answer 15

True

Question 16

T/F: Fermented plant-based products contain in general 10^5−7 lactic acid bacteria/ml or g

Answer 16

True

Question 17

T/F: Lactobacillus plantarum is now named Lactiplantibacillus plantarum.

Answer 17

True

Question 18

T/F: QPS in QPS list stands for Qualified Probiotic Species

Answer 18

No Qualitied Presumption of Safety

Question 19

T/F: The 18S rRNA gene is used for identification of bacterial strains.

Answer 19

False, 16S for bacteria, 18S for yeast

Question 20

T/F: Today there is non approved health claim on probiotic by EFSA.

Answer 20

No health claim specific to probiotics has been authorised in the EU

Question 21

In principle, what is the difference between bacterial identification and bacterial
classification?

Answer 21

You classify based on the different characteristics you can identify.

Question 22

Explain why the 16S rRNA gene is used for identification of bacteria?

Answer 22

Because it has both conserved and hypervariable regions

Question 23

Describe one traditional lactic acid fermented food based on plant-material (name of product,
production process, microorganisms involved, and estimated bacterial concentration in the
final product)

Answer 23

Kimchi
Sauerkraut
Lactic acid bacteria
Estimated bacterial conc is between 10^5-7 for fermented foods.

Question 24

Describe the concept transient microbiota (or variable microbiome) and reflect upon the
possibilities for the microorganisms in the fermented food mentioned in 7a, to affect the
composition of the gut microbiota after consumption.

Answer 24

The transient microbiome is passing through and not part of the core commensals but will still have an effect. Particularly if the probiotics is consumed regularly. While it may not colonise the gut it still has the potential to affect the composition of the microbiome.

Question 25

Describe in detail three different ways in which the microbiota can benefit healthy
individuals.

Answer 25

It helps to outcompete pathogens, synthesise essential vitamins and break down food that humans can’t such as dietary fibers, regulating the metabolism of host, help to balance the immune response, neurotransmitter production

Question 26

Mention four of the major factors influencing our gut microbiota composition?

Answer 26

Diet
Antibiotics use
Genetics
Lifestyle and environment

Question 27

Mention eight properties of an ideal probiotic.

Answer 27

The most important properties of strains to even be considered for probiotic use include: resistance to gastric acidity, bile acid resistance, adherence to mucus and/or human epithelial cells and cell lines, antimicrobial activity against potentially pathogenic bacteria or fungi, ability to reduce pathogen adhesion to surfaces, bile salt hydrolase activity, enhancing viability of probiotics.

Question 28

What is Next Generation Probiotics?

Answer 28

Notably, “traditional probiotic strains” were isolated from gut and traditional fermented foods. By contrast, NGPs have been recently isolated using new tools allowing isolation, identification, and modification of commensal bacterial species.

Question 29

Describe possible risks associated with the use of next gen probiotics.

Question 30

Describe schematically the different steps in Next Generation Sequencing using Illumina
MiSeq

Question 31

What can this method NGS preferably determine

Answer 31

Microbiome composition in the gut, sequences large data sets so makes sense to use it to “map out” all bacteria present

Question 32

Formulate a question within the topic of probiotics that seems relevant to you (naturally you
are not allowed to use or refer to any of the above questions), and then answer your own
question. The answer must be 500 words ± 10 %. The relevance of the question and the
quality of the answer will be evaluated and given a maximum of 10 points.

Question 33

Difference between identify and classify

Answer 33

Identification involves describing something in enough detail so that it could be recognised in a given context. Classifying is an investigative approach that involves sorting objects or events into groups or categories.

Question 34

Which bacteria dominates in sauerkraut?

Answer 34

L. plantarum

Question 35

Which bacteria dominates in kefir?

Answer 35

L. kefir