BB5MOP - Week 18

What are the shapes of bacteria?

Cocci (spherical), bacilli (rod), spirillum (spiral)

What technique is used to distinguish between gram negative and gram positive bacteria and what does it test?

Gram staining, tests the thickness of the peptidoglycan layer.

Describe gram positive bacteria.

Purple stain, thick peptidoglycan layer.

Describe gram negative bacteria.

Pink stain, thin peptidoglycan layer.

What is the order of gram staining?

Crystal violet, iodine, ethanol, carbol fuschin

What is blood agar?

Enriched as it has all the nutrients for a wide range of bacteria to grow and is differential.

Describe the types of haemolysis on blood agar.

1. Alpha haemolysis is a reduction of haemoglobin, green greyish colour.

2. Beta haemolysis is complete lysis of blood, clear area appears yellow.

3. Gamma haemolysis where there is no lysis of blood.

What are biochemical tests?

Insight into bacteria's metabolic process.

Describe the catalase test.

1. Whether bacteria can break down hydrogen peroxide into water and oxygen.

2. Positive shows cloudy formation, negative no changes.

Describe the oxidase test.

1. Presence of cytochrome C.

2. Positive shows blue colour, negative no colour change.

Describe the indoles test.

1. Whether bacteria can convert tryptophan into indole using enzyme tryptophanase.

2. Positive colour change to purple, negative no colour change remains yellow.

Describe the urease test.

1. Whether bacteria can convert urea into ammonia, carbon dioxide and water using enzyme urease.

2. Positive shows pink, negative remains yellow no colour change.

Describe the coagulase test.

1. If enzyme is present and can colt plasma, by converting fibrinogen to fibrin.

2. Positive cloud, negative no colour change.

Where is Staphylococcus epidermidis found and what can it cause?

1. Found all over the skin.

2. Can cause infections in people with weakened immune systems or medical devices like catheters.

Describe Staphylococcus aureus.

A true pathogen that can naturally cause disease by colonising different parts of the body - an opportunistic pathogen.

What is the three step strategy that S. aureus uses to cause infection?

Colonise, Evade and Damage

Describe the first stage, colonisation, of S. aureus infection.

Has MSCRAMMS that grab, hook and adhere to tissues.

What does MSCRAMMS stand for?

Microbial surface components recognizing adhesive matrix molecules

What are the three types of MSCRAMMS?

1. Clumping factor A (ClfA)

2. Fibronectin binding proteins (FnbpA)

3. Collagen and elastin binding proteins (EbpS)

What is clumping factor A (ClfA)?

Binds to fibrinogen to produce fibrin, hijacking plasma clotting system.

What are fibronectin binding proteins (FnbpA)?

Binds to fibronectin.

What are collagen and elastin binding proteins (EbpS)?

Promote adhesion to severely damaged tissue.

Describe the second stage, evasion, of S. aureus infection.

Has a microcapsule that hides it from immune cells, it acts as a bound coagulase forming clots around S. aureus to protect it form neutrophils - phagocytosis minimised.

Apart from the bound coagulase, how else does S. aureus evade the immune system?

Makes protein A binding to antibodies, blocking the immune system from targeting the pathogen.

Describe the third stage, damage, of S. aureus infection.

S. aureus produces invasins, enzymes that can break down tissues to get nutrients.

What are invasins?

A type of free coagulase released to surrounding area that form fibrin clots.

What are staphylokinases?

Breaks down fibrin clots if a pathogen gets trapped.

What are hyaluronidases?

Break down hyaluronic acid, the substance that holds human tissues together.

What are DNases?

Break down DNA to give nutrients to the pathogen.

What are fatty acid modifying enzymes (FAME)?

Convert bactericidal fatty acids in infected tissues to alcohol or cholesterol.

How does a-haemolysin work?

Causes cell death by binding to ADAM10 and punching holes in cell membranes, cells leak out and die.

Apart from invasins, how else does S. aureus cause damage in the third stage of infection?

Produces exotoxins

What are the three types of exotoxins that S. aureus produces?

1. Enterotoxins, from contaminated foods.

2. Toxic shock syndrome toxin 1 (TSST-1), common in tampon users, organ failure from systemic shock.

3. Exfoliative toxins, produced systemically, scaled skin syndrome.

What are superantigens?

Trigger a massive uncontrolled immune response, leading to tissue damage, bind non-complementary to avoid cytokine inflammation.

What are the characteristics of S. aureus?

1. Gram positive cocci

2. Beta haemolytic, golden colonies on blood agar.

3. Coagulase positive

4. DNase positive

5. Protein A positive

What are the characteristics of S. epidermidis?

1. Gram positive cocci

2. Gamma haemolytic, white colonies on blood agar.

3. Coagulase negative

4. DNase negative

5. Protein A negative

Which agar is selective for S. aureus?

Bird Parker Agar (BPA)

What do S. aureus and S. epidermidis look like on BPA?

1. S. aureus forms black colonies with halo, due to lecithinase activity.

2. S. epidermidis forms black colonies with no halo, no lecithinase activity.

What do S. aureus and S. epidermidis look like in a DNase test?

1. S. aureus forms yellow colonies.

2. S. epidermidis remains green.

What does the side coagulase and test coagulase tests test for?

1. SCT tests for bound coagulase (clumping factor A).

2. TCT tests for free coagulase.

How is Protein A tested for and what does a positive result look like?

Latex agglutination test - coated with IgG specific to Protein A, positive result shows clumping of Protein A.