Lecture 2 & 3: Bacteria

Question 1

Distinguishing characteristics of Bacteria (prokaryote)

Answer 1

• Cell wall: present except for mycoplasma
• Cytoplasmic membrane: no sterols except mycoplasma
• nuclear membrane: absent
• nucleolus: absent
• Hereditary info: ssDNA w/ no proteins associated
• Ribosomes: small 70s
• respiration: associated particles in cytoplasmic membrane
• repro: asexual (binary fission)

Question 2

Congenital infections

Answer 2

• Sexual disease: T. Pallidum
• GI disease: L. monocytogenes
• viral causes: rubella

Question 3

Exogenous sources of microbial disease

Answer 3

Ingress

• Inhalation: common cold (viruses), coccidioidomycosis (fungus)
• Ingestion: dysentry (bacterium)

Penetration

• Wound: surgical infections & cat scratch (bacteria)
• insect bites: encephalitis (virus)

Question 4

Cell arrangement: Coccoidal

Answer 4

• pairs: diploccoci (Strep. pneumoniae)
• chains: streptococci (Strep. pyogenes)
• clusters: staphylococci (all staphylococcus sp.)
• tetrads: Sarcina sp.

Question 5

Types of stains

Answer 5

1. Gram-stain (stain -> decolourizer -> counterstain)
2. Acid Fast stain:
   - for Gram’s resistant bacteria ex. mycobacterium & Nocardia spp.
   - Ziehl-Neelsen (hot), Kinyoun (cold), fluorochrome (auramine-rhodamine; primary stain: fluorescent dyes; counterstain: potassium permanganate)

Question 6

Flagellum arrangement

Answer 6

Polar

• monotrichous: Ex. Vibrio sp.
• Amphitrichous: Ex. Spirillum sp.
• Lophotrichous: Ex. Pseudomonas sp.

Lateral

• Peritrichous: Ex. Proteus sp.

Question 7

Cell envelope components

Answer 7

1. Glycocalyx (aka capsule, slime (s-) layer
   - External mucilaginous layer (EPS) Ex. Bacillus anthracis (polypeptide)
   - slime layer: poor organisation
   - capsule: organised, K antigen (Exceptions: M - S. pyogenes, Vi -Salmonellae)
   - Fx: adherence, antigenic activity, antiphagocytic, prevention of neutrophil killing of engulfed bacteria, prevention of PMN leukocyte migration to site of infection, toxicity to host cells, protection
2. Cell wall - Peptidoglycan backbone - alternating polysaccharide chains alternating NAM & NAG
   - Gram +ve: thick peptidoglycan; Teichoic acids (Fx: bind protons, cations, act as adhesins, virus receptor sites); Lipoteichoic acid (dermal necrosis/Schwartzman rxn, induction of cell mitosis at site of infection, immune system stimulation, complement activation)
   - Gram -ve: thin peptidoglycan; LPS; porins (nutrient transport)

Question 8

Endotoxin induction of:

Answer 8

• fever
• haemorrhagic necrosis (Schwartzman rxn)
• disseminated iv coagulation
• production of TNF
• activation of alternate complement pathway
• stimulation of bone marrow cell proliferation
• enhancement of the immune and limulus lysate rxn

Question 9

Acid-Fast Bacteria

Answer 9

• Genera Mycobacterium & Nocardia
• Peptidoglycan + arabinose & galactose polymers -> mycolic acids (waxy)

Question 10

Effect of lysozyme

Answer 10

• breaks Beta1-4 bonds b/w NAM & NAG
• spheroplast: portion of cell wall remains because lysozyme only enters in portion of cell wall with porins
• protoplast: cell wall completely removed (Gram +ve more sensitive) due to lack of 2nd layer of cell wall

Question 11

Effect of penicillin

Answer 11

• prevents cell wall formation only in growing cells (no effect on Mycoplasma)
• inhibits formation of nl cross-linkages in peptidoglycan
• binds irreversibly to penicillin-binding proteins (PBP transpeptidases)
• result: defective cell walls
• no protection from osmotic shock

Question 12

Periplasmic space contains:

Answer 12

• nutrient transport proteins
• nutrient acquisition enzymes (proteases)
• detox enzymes (Beta-lactamases)
• membrane Derived Oligosaccharides (MDO)
• Osmoprotectants

Question 13

Pathogenicity Islands (PAI)

Answer 13

Distinct genetic elements on chromosome:

• virulence genes
• present only in pathogen genome
• occupy large genomic areas (10-200 kb)
• differ from core genome (base composition & operon useage)
• frequently located next to tRNA genes
• frequently associated with mobile genetics elements (transposons)

Question 14

Ex. of pathogens with PAI

Answer 14

Grav +ve

• Listeria spp.
• S. aureus
• Streptococcus spp.
• Enterococcus faecalis
• C. diff

Gram -ve

• H. pylori
• E. coli
• Salmonella spp.
• Shigella
• Yersinia spp.
• L. pneumophilia
• P. auruginosa
• V. cholerae
• Bacteroides fragilis

Question 15

Endospores

Answer 15

• 2 bacterial genus produce spores: Botulinum and Bacillus
• Specialised structures: produced in env. stress for spore to survive harsh conditions
• Resistant: UV, irradiation, chemical disinfection, drying
• require specialised stains (light microscopy)
• Coat: keratin-like, impermeable layer (resistance to antibacterials)
• Cortex: type of peptidoglycan (fewer cross-links)
• Spore wall: peptidoglycan layer; cell wall germinating vegetative cell
• Core: contains complete genetic material

Question 16

Bacterial shapes

Answer 16

• Spherical: Cocci
• Cylindical: Bacilli; Ex. B. perfringens (short & stubby)
• Curved: Vibrio (do not confuse with Vibrio cholera)

Question 17

Flagellum made of 3 parts

Answer 17

1. Helical filament
   - Flagellin: highly antigenic (H antigen)
   - inserted into hook
2. Hook: anchors filament into basal body
3. Basal body:
   - contains rod and 1 or 2 sets of double plates (rings)

• located in cytoplasmic membrane & cell wall
• Gram -ve flagellum have 4 rings (L, P, S & M rings ) &

Gram +ve have 2 rings (S & M rings are for both)

Question 18

Peptidoglycan layer

Answer 18

• Tetrapeptide on NAM; D-alanine always the 4th a.a.
• Cross-linkage occurs on 3rd a.a. of one tetrapeptide to the 4th a.a. of an adjacement tetrapeptide
• Gram +ve cross-linkage is via
  Penta-Glycine bridge; more than 90% is cross-linked; 3rd a.a. is always L-Lysine
• Gram-ve crosslinkage is a lipoprotein bridge b/w Diaminopimelic acid of one tetrapeptide to the D-alanine of another tetrapeptide; <60% cross-linked

Question 19

Examples of bacteria with Lipooligosaccharide (LOS)

Answer 19

• Bordetalla pertussis
• N. meningitidis
• C. jejuni
• Some endotoxins have LOS instead of LPS subunits; both stimulate immune system

Question 20

LPS structure

Answer 20

• only in Gram -ve bacteria
• endotxin
• O-antigen
• core polysaccharide
• Lipd A (fatty acids): toxic

Question 21

Comparison of Gram +ve & -ve cell walls

Answer 21

Gram +ve

• Peptidoglycan
• Teichoic & Lipoteichoic acid
• Activation of both classic and alternate complement pathway
• Activation of both specific and non-specific immune response
• Dermal necrosis
• more sensitive to penicillin b/c cell wall are thicker w/ more cross-linkages.

Gram -ve

• LPS
• Lipoprotein
• Peptidoglycan
• Porins
• Endotoxin induction of: alternate complement pathway, etc… (see other card)
• Haemorrhagic necrosis

Question 22

Pilus features

Answer 22

• hollow & helical, thinner than flagella
• protein composition is Pilin (classification & iD)
• F-Pilus (sex pilus) Gram -ve bacteria only

Question 23

Fimbriae

Answer 23

• Type I pili
• adhesion to surfaces
• predominantly Gram -ve, some Gram +ve (Corynebacterium renale, Actinomyces naeslundii)

Question 24

Axial filaments aka Endoflagellum

Answer 24

• Spirochetes - Leptospira lack flagella but are motile
• possess axial filaments instead: long thin microfibril inserted into a hook
• entire structure enclosed in periplasmic space (not exposed to external environment)