Introduction to Bacteriology

Question 1

How do prokaryotes differ from eukaryotes?

Answer 1

Prokaryotes have closed, circular DNA, no histones, and no introns. Can contain up to 34% imported DNA.

Question 2

Name the main shapes used to classify bacteria.

Answer 2

• Rods (bacilli): e.g., E. coli
• Spheres (Cocci): e.g., Staphylococcus aureus
• Spirals
• Threads/Filaments: e.g., Treponema pallidum.

Question 3

What is the main structural difference between Gram-positive and Gram-negative bacteria?

Answer 3

• Gram-positive: thick peptidoglycan layer and one membrane.
• Gram-negative: thin peptidoglycan layer between two membranes

Question 4

What is peptidoglycan made of?

Answer 4

• Carbohydrate part: NAG and NAM linked by β-1,4 glycosidic bonds.
• Peptide part: amino acids in L- and D-stereochemistry attached to NAM

Question 5

How does peptidoglycan polymerisation differ between Gram-positive and Gram-negative bacteria?

Answer 5

• Gram-negative: fewer peptide cross-links, looser organisation.
• Gram-positive: more peptide cross-links for structural rigidity.

Question 6

What is teichoic acid, what type of bacteria is it found in and what are its functions?

Answer 6

Polymer of alcohol molecules joined by phosphate groups, found in Gram-Positive Bacteria, Functions: structural integrity, contributes to surface composition, role in inflammatory response.

Question 7

Describe the function of porins in Gram-negative bacteria.

Answer 7

Control entry and exit of molecules (e.g., antibiotics), Size limit: molecules >700 Daltons cannot pass into cell.

Question 8

What are the three components of lipopolysaccharides (LPS) in Gram-negative bacteria?

Answer 8

• Lipid A: pro-inflammatory, sits in the outer membrane.
• Core oligosaccharide: bridge to oligosaccharide chain.
• Oligosaccharide chain: highly variable, aids in identification.

Question 9

What are the functions of bacterial capsules and slime layers?

Answer 9

Protect bacteria and prevent dehydration, aids immune evasion by hiding recognition molecules.

Question 10

What generates the energy for flagellar motion in bacteria?

Answer 10

Proton gradients across membranes.

Question 11

Describe the polar flagellar arrangement.

Answer 11

Single flagellum at one end.

Question 12

Describe the amphitrichous flagellar arrangement.

Answer 12

Two flagella, one at each end.

Question 13

Describe the lopotrichous flagellar arrangement.

Answer 13

Bunch of flagella at one end.

Question 14

Describe the peritrichous flagellar arrangement.

Answer 14

Flagella around the entire cell like a halo.

Question 15

What are the roles of pili (fimbriae) in bacteria?

Answer 15

Adhesion to surfaces, subsequent retraction allows movement toward a surface.

Question 16

Describe the composition of pili (fimbriae) in bacteria.

Answer 16

Protein filaments with specialised adhesion subunits at the tip.

Question 17

What is the signficance of peptidoglycan containing both L and D steroisomers of amino acids when proteins only contain the L form?

Answer 17

Aids function of peptidoglycan to provide mechanical strength: D amino acids will not be broken down by proteolytic enzymes.

Question 18

What is special about DAP (amino acid found in peptidoglycan)?

Answer 18

It is not one of the 20 amino acids that makes up proteins.

Question 19

In which type of bacteria does its peptidoglycan contain an interpeptide chain attached to its DAP?

Answer 19

Gram-Positive peptidoglycan.

Question 20

What is the order of the 5 amino acids attached to NAM in peptidoglycan, starting from the NAM component?

Answer 20

L-alanine, D-glutamate, DAG, D-alanine, D-alanine.

Question 21

4 reasons

Why is it important to identify pathogens?

Answer 21

• Predict symptoms.
• Suggest treatment.
• Identify the source of infection for better transmission management.
• Predict when infections might occur.

Question 22

What factors need to be considered when using bacterial culture conditions to identify bacteria?

Answer 22

Bacterial response at different temperature, different levels of oxygen availability, different carbon sources (e.g., glucose, lactose), interaction with indicators, blood, bile salts, or antibiotics.

Question 23

What are the four main types of culture media?

Answer 23

• Rich media grows everything.
• Selective media promotes growth of specific species.
• Differential media distinguishes species based on metabolic processes (e.g., colour changes).
• Enrichment media supports bacteria with specific growth requirements.

Question 24

Give an example of enrichment media and its use.

Answer 24

Blood agar supports bacteria causing bloodstream infections and distinguishes Gram-positive bacteria via haemolytic activity.

Question 25

What does the catalase test differentiate between?

Answer 25

Staphylococcus (catalase-positive, causes skin infections), Streptococcus (catalase-negative, causes respiratory infections).

Question 26

What does the coagulase test identify?

Answer 26

Different strains of Staphlococcus bacteria: Staphylococcus aureus (positive: clumping observed), Staphylococcus epidermidis (negative).

Question 27

What is the IMViC test used for?

Answer 27

Discriminating between coliform species by testing for tryptophanase activity, acid production, acetoin production, and citrate utilisation.

Question 28

What are the three main types of antigens used in serotyping gut bacteria?

Answer 28

- **O-antigens:** sugar side chains of Gram-negative LPS. - **K-antigens:** capsular monomers. - **H-antigens:** flagellar proteins.

Question 29

What is CHROMagar, and when is it effective?

Answer 29

A medium that identifies bacteria (e.g., UTI pathogens) by producing distinct colours, works well with high bacterial concentrations such as in urine.

Question 30

Why does the BactAlert blood culture system take time to identify bacteria?

Answer 30

Requires bacterial growth to detect CO₂ production and identify bacteria, needs 36–72 hours for results and subsequent identification.

Question 31

How does PCR help in bacterial identification?

Answer 31

- RAPD-PCR uses primers (random sequences) to create genetic fingerprints for tracking transmission. Sequences are unknown so will not enable species identification. - MLST identifies strain-specific alleles by targeting essential genes. - Rapid identification of MRSA (by showing presence of mecA gene).

Question 32

What is the advantage of Whole Genome Sequencing (WGS) in outbreak detection?

Answer 32

Identifies similarities and differences between bacterial genomes, provides precise identification of transmission events.

Question 33

What is a coliform?

Answer 33

a type of Gram-negative, rod-shaped bacterium.

Question 34

When grown on blood agar, what is observed in α-haemolytic gram-positive bacteria?

Answer 34

Biliverdin is produced, observations are dark green cloudiness.

Question 35

When grown on blood agar, what is observed in β-haemolytic Group A gram-positive bacteria?

Answer 35

Completely clear areas of blood agar.

Question 36

When grown on blood agar, what is observed in β-haemolytic Group B gram-positive bacteria?

Answer 36

Slight orange haze in areas of partial clearing in the blood agar.

Question 37

What is meant by the term 'microbiota' and what did it used to be called?

Answer 37

It is the community of commensal, mutualistic and pathogenic microorganisms that share our body space. Used to be called flora.

Question 38

What is meant by the term 'microbiome'?

Answer 38

It is the totality of organisms and their collective genetic material.

Question 39

Why is it important to study microorganisms?

Answer 39

They are everywhere! Humans are constantly in contact with microorganisms. Even our genome contains viral genetic material. As humans, we are the hosts of many microorganisms.

Question 40

What are the 3 main types of Host-Microbe Interactions?

Answer 40

**Mutualism:** Both host and microbes benefit from the symbiotic relationship (e.g. human gut microbiota) this is the most common interaction. **Commensalism:** Microbes benefit from the interaction but host does not (e.g. skin, nose and oral cavity microbiota). **Parasitism:** Microbes benefits from interaction whilst harming the host. In parasitism, the microbe is referred to as pathogenic.

Question 41

What is dysbiosis?

Answer 41

When alterations that generate an imbalance occur, pathogens may colonise, causing disease. Factors may include stress, production of toxins by pathogens, type of food ingested, medications (e.g. antibiotics), malfunction of organs. Can lead to infection of opportunistic pathogens.

Question 42

How can we use sequencing to identify bacteria of the microbiome at the species level?

Answer 42

Based on the 16S rRNA gene which is composed of 9 hypervariable regions. Amplification using PCR and primers to target 16S rRNA gene allows us to compare to a bacterial species database to identify them at the species level.

Question 43

Where in the human body is the microbiota most diverse?

Answer 43

The gut.

Question 44

Which 5 biological processes in humans might the bacteria of the microbiota aid?

Answer 44

Extracting energy from food, producing essential vitamins, regulating our immune system, regulating glucose levels and metabolism, protecting us against pathogens.

Question 45

What factors can affect the microbiota?

Answer 45

Obesity, age, malnutrition, environment, changes in diet can change microbiota composition. Studies transplanting microbiota of mice, show a relationship between obesity and microbiota. We don't know if this is cause or effect though.

Question 46

Approximately what percentage of the adult population suffer from irritable bowl syndrome (IBS)?

Answer 46

11%, they have an altered microbiome.

Question 47

What does the gut-brain axis refer to?

Answer 47

Relationship between brain and gut, bloodstream and nerve system connects them. This is a form of bi-directional communication.

Question 48

What are the key findings in relation to Gut-Brain axis research?

Answer 48

Serotonin, dopamine and other relevant neurological molecules are produced by some microbes living in out gut, such as Candida, E.coli. Animal Studies found associations between autism spectrum disorders and intestinal problems, faecal transplant from ASD individual causing ASD-like behaviours in previous neurotypical mice. Human studies found that stress increases gut permeability, allowing bacteria antigens to cross - neural mechanism unclear.

Question 49

What are some examples of bacteriotherapy?

Answer 49

Probiotics, prebiotics, synbiotics and Faecal Micriobiota Transplantation

Question 50

What are probiotics?

Answer 50

Live microorganisms which when administered in adequate amounts confer a health benefit on the host.

Question 51

What are prebiotics?

Answer 51

Non-digestible food ingredient that beneficially affects the host by selectively stimulating the growth / activity of a limited number of bacteria in the colon.

Question 52

What are synbiotics?

Answer 52

Food ingredients or dietary supplements combining probiotics and prebiotics in a form of synergism.

Question 53

When are Faecal Microbiota Transplants used and what is the process of adminstration?

Answer 53

Used in patients with recurrent C. difficile infections as a last resort. 50g-300g faeces are donated from a healthy individual. Faecal material is washed in saline and administered in pills or as a faecal slurry (tube to stomach). Approximately 1-2 days after transplantation, recipient microbiota shifts dramatically to resemble healthy gut microbiome.

Question 54

What is MALDI-TOF identification? When can it be used? How does it work?

Answer 54

- MALDI-TOF is used to identify bacterial species causing infection. - Often requires cultures on plate but can use blood cultures directly from machines that purify bacterial proteins. - Only works for single species identification (90% of the time this is the case for bloodstream infections). - Ionisation of material, causing movement towards a magnetic field: size of molecule dictates speed of movement. - Gives a series of peaks represent charge:mass of the mixture, indicative of a particular species. - Spectrum compared to database containing sprectra of bacteria. - Machine looks for closest match and gives a likelihood score for the match.

Question 55

What are the main limitations of MALDI-TOF?

Answer 55

- Where there are multiple species causing infection, it is useless. - Less accurate with less common species as there are less spectra of this species on the database. - There is little clinical relevance in knowing the species. What matters is whether bacteria are resistant or susceptible to antibiotics. Often knowing the species would not identify this.