Gram Positive
Gram Positive cocci
Staphylococcus epidermis / aureus
- 1/2 - 2/3 all infections
Streptococcus viridans
- 25%
Other
- Enterococci
- Strept. pyogenes / pneumoniae
- Neiserria
Gram positive bacilli (all anaerobes)
Clostridiae (bacilli)
Listerius
Bacillus
Gram Negative
Gram negative bacilli
10-20%
Coliforms
- E Coli, Proteus, Salmonella, enterobacter, acinetobacter
Pseudomonas
Hemophilus
Gram Negative Cocci
Neiserria
Gram negative Anaerobes
Bacteroides
Resistant microbes
MRSA, MRSE & VRE
Cell Structure
Cytoplasmic Membrane
Innermost layer
- present in both Gram positive and negative
- functions as a permeability barrier and transport system
Cell Wall
Function
- maintains cell shape
- protects high internal osmotic pressure
- cell wall injury = Lysis
Gram positive
- thick Peptidoglycan Layer (x5)
Gram negative
- thin Peptidoglycan Layer
- second outer membrane of lipopolysaccharide (endotoxin)
Capsule & Glycocalyx
Extracellular Polymer
- if forms condensed, well-defined layer, called Capsule
- if forms loose meshwork of fibrils, called Glycocalyx
Slime
- polysaccharide Glycocalyx
- envelopes bugs infecting prosthesis
- S aureus & epidermis
- Pseudomonas
Function
- adhere to and survive on synthetic surfaces
- protects from host-defense factors / Complement fixation / Neutrophil ingestion
- 500 x more resistant
"Window of Opportunity"
- theoretical period when can kill microbe before biofilm forms
- basis behind Acute Early TJR infection regimes
- time frame unknown ? 2/52
Spores
- gram positive rods
- Bacillus & Clostridium
- occurs in unfavourable conditions
- resistant to drying / antiseptic
- only killed by Betadine
- killing of spores is the difference between Sterilization & Disinfection
Staining
1. Gram Stain
Difference is in cell wall
- reason unclear
Technique
- flood with Crystal Violet and wash
- flood with Iodine applied and wash
- all microbes are blue at this point
- carefully decolourise with ethanol
- counterstain with safranin
Gram positive
- retain crystal Violet - Iodine complex
- purple
- don't take up counterstain
Gram negative
- cells completely decolourised
- take up safranin
- take on contrasting Red
2. Acid - Fast Stain / Ziehl-Neilsen
- AFB retain Carbol-Fuschin stain even when decolourised with acid-alcohol
- most commonly Mycobacterium
Process
- red Carbol-fuschin applied
- heated on steam bath
- decolourised with hydrochloric acid in alcohol
- contrasting blue counterstain applied
Acid-fast bacteria appear red
- others are blue
Antimicrobial Actions
Four basic methods of action
1. Cell Wall Synthesis
- ß lactam drugs
- Vancomycin
2. Inhibition of Cell Membrane Permeability
- Polymyxins
- Amphotericin B & Fungi
3. Inhibition of Protein Synthesis
- Gentamicin
- Erythromycin
- Tetracyclines
4. Inhibition of Nucleic Acid Synthesis
- Quinolones (Ciprofloxacin)
- Rifampicin
- Sulfonamides
- Trimethoprim
Antibiotic Resistance
1. Genetic Exchange
- plasmids
- Entire chromosomes
2. mutation
Mechanisms
- degrade antibiotics / B Lactamases
- modify receptor sites or target
- alter 30s binding site of ribosome
- decrease bacteria's permeability to the antibiotic
- protective glycocalyx
- produce cell membrane antibiotic pumps