Clinical microbiology

Question 1

ILOs

Answer 1

• Have an appreciation of disease mechanisms – entry, establishment, evasion, damage
• Have an understanding of the clinical significance of the anatomy of disease causing microorganisms

Question 2

Requirements for disease

Answer 2

1. Point of entry
2. Establishment
3. Avoiding host defence mechanisms
4. Damaging the host

Question 3

Portals of entry

Answer 3

Most common hospital infection is UTI

Normally a change in pH in urinary tract (normally acidic) causes UTI

Question 4

Portals of entry - mucous membranes

Answer 4

Mucous membranes need to be protected:

1.Washing with secretions, e.g. tears,
saliva, mucus (nasal passages) and urine (urethra).

2. Filter hairs in nasal passages prevents entry of large particles.
3. Cilia in respiratory tract push mucous and microbes upward.

Non-specific innate Mechanisms
Mechanical Factors
Keratinised surface of skin - tough so acts as an
effective barrier against entry i.e., mechanical barrier to microbial to invasion

Question 5

Mucous membranes

Answer 5

• Form a protective covering that resists penetration and traps many microbes
• are often bathed in antimicrobial secretions which contain a variety of antimicrobial substances
• contain mucosal-associated lymphoid tissue (MALT) (part of the secondary lymphatic system)

Question 6

Antimicrobial secretions

Answer 6

Many body fluids, e.g. blood, are not normally suitable for microbial growth due to the presence /absence of various factors. e.g.

Normally iron is not bioavailable in blood or breast milk (it is complexed in molecules, e.g. Transferrins or haemoglobin) - bacteria need iron to multiply

Long chains of fatty acids (e.g. oleic acid) occur in slightly acid secretion of the skin (pH 4-6) and these are lethal to many bacteria.

Lactenin - (nitrogenous protein) present in breast milk which are selectively bactericidal for Streptococcus pyogenes - protects against mastitis (inflammation of the mammary gland).

Question 7

Antimicrobial Secretions - Lysozyme

Answer 7

Main mediator as protection against infection.
Lysozyme hydrolyzes bond connecting sugars in peptidoglycan (cell wall)

Action of lysozyme on the cell wall of Gram+ve bacteria. Peptidoglycan backbone, the β(1-4) bonds connect alternating N-acetylglucosamine and N-acetylmuramic acid (NAM) residues.

Action of lysozyme is more effective on G+ve bacteria as the peptidoglycan cell wall is more easily accessed (phospholipid bilayer somewhat protects a G-ve bacterium)

Question 8

Gastrointestinal tract

Answer 8

• stomach
– gastric acid

• intestines
– pancreaticenzymes– bile
– intestinal enzymes– GALT (M cell patches - Peyers patch)
– peristalsis

. intestines

– shedding of columnar epithelial cells

– secretoryIgA
– normal microbiota– Paneth cells

• produce lysozyme
• produce cryptins (defensins: peptides)

Genitourinary tract

low pH of urine and vaginal epithelia
urea and other toxic metabolic end products in urine hypertonic nature of kidney medulla
flushing with urine and mucus

Question 9

Antimicrobial Peptides - Defensins and cationic peptides

Answer 9

Defensins:

1. Defend from pathogens
2. Shape Microbiota
3. Protect stem cells

peptides that are open- ended, rich in arginine and cysteine, and disulfide linked. Found in neutrophils, intestinal Paneth cells and intestinal and respiratory epithelial cells

Cationic peptides

e. g. cathelicidin, produced by a variety of cells
e. g. neutrophils and respiratory epithelial cells

LL37 (leucine leucine 37 a.a.)
- proteolytic product of a cathelicidin

• amphipathic

Question 10

Antimicrobial Peptides - Bacteriocins

Answer 10

Bacteriocins

Peptides produced by bacteria including normal microbiota

Lethal to closely related species

e. g., colicins produced by E. coli
e. g., sakacins produced by lactobacilli

Question 11

Antimicrobial Peptides - Selectivity

Answer 11

• host mammalian amphipathic antimicrobial peptides selectively bind to the differential membranes
• host secretions are amphipathic so really good at binding to bacterial membranes (cholesterol molecule in host membrane decreases charge on the membrane, bacteria do not have this, and are much more negatively charged)
• damage the integrity of the cell they bind to

Question 12

Establishment

Question 13

Adherence factors associated with establishment of infection

Answer 13

Adherence mediation: nearly always proteins

Question 14

Adherence factors associated with establishment of infection - N. meningiditis

Answer 14

• interactions are multi-adhesion, multi-molcule
• proteins are predominantly involved in attachement
• through pili and pores

Question 15

Evading host defences

Answer 15

• Leukocidins
– Kill white blood cells including neutrophils and macrophages!

Produced by Staphylococcal and Streptococcal spp.

• Coagulase

– Causes fibrin clots to form in the blood of a host. Advantageous to the bacteria in evasion

Question 16

Damage (exotoxins)

Answer 16

• Neurotoxins – cause paralysis
• Enterotoxins – cause sickness and diarrhoea
• Cytotoxins – cause cell death

Question 17

Damage

Answer 17

• classic damage by TOXINS
• EXOTOXINS (secreted)
• EXOTOXINS (LPS, lipopolysaccharide on the SURFACE of G-VE bacteria)

TB has no obvious virulence factor (toxin that causes damage in absence of the bacterial cell)

Question 18

Damage - comparison between exotoxins and endotoxins

Answer 18

LPS = endotoxin, part of the cell

Highly toxic

Question 19

Bacterial anatomy

Answer 19

Certain bacterial structures satisfy several of the requirements for satisfactory infection.

The clinical significance of bacterial anatomy:

1. Structures outside the bacterial cell wall - cause the most disease
2. The bacterial cell wall
3. Inside the cell wall

Question 20

Bacterial anatomy - G+ve vs G-ve

Answer 20

Question 21

Bacterial anatomy: clinical significance of the bacterial cell wall

Gram Positive

Answer 21

• Gram positive:

– Teichoic acid is being considered as a vaccine target for both S. aureus and C. difficiledue to the high levels of antibiotic resistance.

– M protein from Strep. pyogenesis highly variable but important in infection- protrudes from cell wall

– Mycolic acid from Mycobacterium tuberculosis prevents the action of many antibiotics and host effects (‘myco’ = Greek for slimy, slimy because of mycolic acid) unique to actinomycetes

Question 22

Bacterial anatomy: clinical significance of the bacterial cell wall

Gram Negative

Answer 22

Gram negative:

• Lipid A anchors LPS to the outer phosholipid bilayer. Release of this leads to a heightened immune reaction. Endotoxin!
• O-antigen highly variable and recognized by the immune system. Can be used in typing (e.g. Ecoli 0157)

Question 23

Bacterial anatomy: structures outside the cell wall: Glycocalyx

Answer 23

Structures outside the bacterial cell wall - called Glycocalyx

– Polysaccharides and proteins. If loosely attached referred to as slime layer. If they form a highly organized structure more commonly known as Capsule. (poly-d-glutamic acid)

– Capsule is important in preventing phagocytosis and allowing the infection process to continue

Question 24

Bacterial anatomy: Fimbriae and Pili

Answer 24

Structures outside the bacterial cell wall.

• Fimbriae and pili

– Involved in adherence. Bundle forming pili from E. coli.

– Shorter than flagella and typically

found on Gram negative bacteria.

– Also aids in motility such as gliding or twitching motility

– Potential vaccine candidates.

– Pili also used in immune evasion through a process of antigenic variation.

Question 25

Bacterial anatomy: structures outside the cell wall: flagella and axial filaments

Answer 25

Structures outside the bacterial cell wall.

• Flagella and Axial filaments

– Flagella protrudes far beyond the cell wall and glycocalyx.

– Several configurations (see Yr1 notes)

– Aids in movement to distal tissues

– H. pylori uses flagella to penetrate through gastric mucous

– Axial filaments like flagella can produce a rotational movement of the whole organism. B. burgdorferi

Question 26

Bacterial anatomy: Structures INSIDE the bacterial cell wall

Answer 26

• Plasma membrane
– Target for therapeutic strategies,

colistin and polymyxin B.

• DNA

– Spread of antibiotic resistance on plasmids. Target of antibiotic therapy.

• Ribosomes
– Target for antibiotics

Question 27

Bacterial anatomy

Clinical relevance of bacterial cell structures - summary

Answer 27