Introduction to Microbiology

Question 1

What are the sizes of Viruses and Bacteria, and what type of microscope is used to view them?

Answer 1

Viruses (0.03-0.3 microns; Electron-light microscope)

Bacteria (0.1-10 microns; Light microscope)

Question 2

Unique Features: Viruses

Answer 2

Smallest infectious particles

Require host for replication

Question 3

Unique Features: Bacteria

Answer 3

Simple unicellular organisms

Prokaryotes

Question 4

Unique Features: Fungi

Answer 4

More complex
Eukaryotes
Unicellular (yeast) or filamentous (mold)

Question 5

Unique Features: Parasites

Answer 5

Most complex
Eukaryotes
Unicellular to multicellular

Question 6

What is unique about prokaryotes compared to eukaryotes?

Answer 6

1. No nucleus
2. Single, double-stranded, circular DNA chromosome (Haploid)
3. Smaller ribosome (70s)
4. Mesh-like peptidoglycan cell wall

Question 7

Classification of Bacteria (4 things)

Answer 7

1. Visible features (shape, spore formation, gram reaction)
2. Nutrition (growth media, aerobic vs anaerobic, temperature for optimal growth)
3. End products (production of specific enzymes/toxins)
4. Surface molecules (antigen composition)

***Currently developing a more rigid classification system by nucleic acid analysis (PCR, nucleic acid homology, and gene probes)

Question 8

What are the shapes of Cocci, Bacilli, and Spirochetes?

Answer 8

Cocci (spherical)
Bacilli (rod; straight or curved)
Spirochetes (spiral)

Question 9

Are gram positive or gram negative cell walls more simple?

Answer 9

Gram Positive

Question 10

Do gram positive or negative cells stain purple?

Answer 10

Positive (Purple = Positive)

Question 11

Where is the peptidoglycan layer found?

Answer 11

Internal to the capsule (if present) and external to the cytoplasmic membrane (not found in Mycoplasma and Chlamydia)

Question 12

Functions of peptidoglycan

Answer 12

Provide protection
Determines shape

Unique to bacteria; both gram positive and gram negative have it

Question 13

Structure of Peptidoglycan

Answer 13

• Linear chain of alternating N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) with tetra peptide crosslinks
• Linked via B-1,4 linkages

Question 14

Where does lysozyme from endothelial cells attack bacteria?

Answer 14

At the B-1,4 linkages of the peptidoglycan layer

Question 15

Proinflammatory effects of Peptidoglycan

Answer 15

Fixes Complement
Binds pattern recognition receptors (e.g. Toll-like receptors)
Triggers TNF production

Question 16

Unique features of gram-positive envelope

Answer 16

1. Thick layer of peptidoglycan (many layers w/ extensive crosslinking)
2. Teichoic Acid (PG associated; PM associated (Lipoteichoic Acid, LTA))
3. Various proteins and lipids which differ between bacteria

Question 17

What are the functions of Teichoic Acid and LTA?

Answer 17

Promote attachment of bacteria to other bacteria or host cells (virulence factors)

Question 18

Unique features of gram-negative envelope

Answer 18

1. Greatly reduced peptidoglycan
2. PG (peptidoglycan) is within the periplasmic space
3. Surrounded by an OUTER MEMBRANE rich in porins and endotoxin/LPS (lipopolysaccharide; important pathogen-associated molecular pattern)

Question 19

Structure of outer membrane of gram-negative envelope

Answer 19

• “Stiff canvas sack”
• Maintains structure/permeability barrier
• Rich in porins (allow diffusino of hydrophilic molecules less than 700 Da through)
• Structural and transport/secretory proteins
• Asymmetric phospholipid bilayer

Question 20

Inner leaflet vs outer leaflet of Outer Membrane of gram-negative envelope

Answer 20

Inner leaflet contains phospholipids

Outer leaflet contains LPS (creates asymmetry of membrane)

Question 21

What are the 3 subunits of LPS/Endotoxin?

Answer 21

Lipid A, Core Polysaccharide, O Antigen

Question 22

Lipid A

Answer 22

• Responsible for endotoxin activity of LPS*

- Fatty acids anchor it into outer membrane

Question 23

Core Polysaccharide

Answer 23

• Branched polysaccharide of 9 to 12 sugars

- Required for bacterial structure and viability

Question 24

O Antigen

Answer 24

• Long, linear polysaccharide of repeating saccharide units of 4 to 7 sugars/unit
• **Helps to classify specific serotypes bacteria due to VARIATION
• Missing in Neisseria (LOS)

Question 25

How does LPS/Endotoxin cause inflammation/septic shock?

Answer 25

Binds CD14 and TLR4 on phagocytes/APCs, which activates the immune system and causes transcription of proinflammatory cytokines (TNF, IL-1, IL-6)

Question 26

Acid-fast Bacteria Characteristics

Answer 26

- Complex cell envelope - Mycolic acids (long, branched chain fatty acids) covalently bound to the peptidoglycan via a polysaccharide - Other mycolic acid-containing compounds as well as complex lipids form a thick waxy membranous layer outside of the peptidoglycan

Question 27

Examples of Acid-Fast Bacteria

Answer 27

Mycobacteria, Nocardia, and Corynebacteria

Question 28

Weakly-stained bacteria

Answer 28

Legionella

Question 29

Cell wall too thin to be visible

Answer 29

Treponema, Leptospira

Question 30

Wall-less bacteria

Answer 30

Mycoplasma

Question 31

Gram Stain Steps

Answer 31

1. Crystal Violet 2. Gram Iodine 3. Decolorizer (Alcohol or Acetone) 4. Safranin Red

Question 32

Acid-Fast Stain Steps

Answer 32

1. Stain all bacteria red by carbol fuchsin 2. Acid alcohol added to remove stain from everything except those that are Acid-Fast 3. Methylene blue used to counter-stain and label other bacteria

Question 33

Acid Fast Cell Wall

Answer 33

- Waxy outer wall found on Mycobacteria and Nocardia | - Outermost layer is rich in mycolic acids, which causes a very SLOW GROWING bacteria

Question 34

Acid Fast Structure

Answer 34

***Similar to Gram Positive (Inner plasma membrane overlaid with peptidoglycan and no outer membrane) -Contains Lipoarabinomannan (LAM), Arabinogalactans, and Mycolic Acid

Question 35

Lipoarabinomannan (LAM)

Answer 35

Functionall related to LPS (structure, viability, activates immune system)

Question 36

Arabinogalactans

Answer 36

Branched polysaccharides that bind to mycolic acid

Question 37

Mycolic Acid

Answer 37

60-90 carbons | -WAXY

Question 38

Pili/Fimbrae (two functions)

Answer 38

- Common or somatic (involved in attachment to epithelial cells; usually called adhesins, lectins, evasins, or aggressins) - Sex - only one per cell (involved in gene transfer; usually a plasmid)

Question 39

Important antigen associated with Flagella

Answer 39

H-Antigen

Question 40

Flagella function

Answer 40

Locomotion; can be located at one or both poles | -Important VIRULENCE FACTOR (if removed, more difficult to cause infection)

Question 41

Important Qualities of Capsule

Answer 41

- Can be present on both gram positive and negative - Called slime layer or glycocalyx - Hydrophilic - Aides in protection from immune system (ANTIPHAGOCYTIC, poorly antigenic, major VIRULENCE FACTOR) - Needed to survive in host, not in culture * **Capsule + protein conjugate serves as antigen for many vaccines

Question 42

Biofilm

Answer 42

An organized community of microbial cells that has a capsule/slime layer over the entire population

Question 43

Quellung reaction

Answer 43

Used to detect encapsulated bacteria

Question 44

How are encapsulated bacteria cleared from the body?

Answer 44

Opsonized and cleared by the spleen

Question 45

Endospores

Answer 45

Only formed by Gram POSITIVE bacteria in harsh environmental conditions (e.g. low nutrition) - Bacteria convert to vegetative state - Gives rise to a single bacteria when conditions are favorable

Question 46

Endospore Characteristics

Answer 46

- Dehydrated, multishelled structure - Contains a complete copy of the chromosome - Has minimum proteins and ribosomes - High concentration of calcium bound to DIPCOLINIC ACID - Inner membrane, two peptidoglycan layers, and an outer keratin-like protein coat * **Multiple coats make it resistant to many treatments***

Question 47

Endospores: Human pathology

Answer 47

-Can be aerosolized (airborne, bioterrorism) -Difficult to manage in hospital settings, as they can withstand extreme pH, temperature (boiling), radiation, attack by enzymes and chemicals -Can exist for centuries Sterilization: Autoclave, 120 degrees C at high pressure for over 20 mins, 70% EtOH, 10% bleach and water