lecture 2- med micro

Prokaryotic Cell Structures

• Cell envelope: Includes the cytoplasmic membrane, cell wall, and glycocalyx/capsule.

• Cytoplasmic membrane: Acts as a barrier between the cell's interior and exterior environment.

• Cell wall: A rigid structure that prevents the cell from bursting, maintaining cell contents.

• Glycocalyx/capsule: A layer outside the cell wall, offering protection and attachment.

• Appendages: Include flagella (for motility) and pili (for adherence).

Cytoplasmic Membrane

• The boundary of the cell, made of a phospholipid bilayer.

• Embedded membrane proteins move constantly (fluid mosaic model).

• Regulates the movement of proteins and other molecules (ions) in and out of the cell.

• Selectively permeable, allowing only certain molecules to pass.

• Movement across the membrane is regulated by simple diffusion and osmosis (no energy required) or active transport (uses energy).

Cell Wall

• A rigid structure that maintains cell integrity and determines the shape of the organism.

• Cell wall architecture distinguishes Gram-positive and Gram-negative bacteria.

• Contains unique structures and molecules, some recognized by the immune system as signs of invaders.

• Antimicrobial agents target some of these wall structures.

Peptidoglycan

• Found in bacterial cell walls.

• A polymer of NAM and NAG.

• Its structure is a key difference between Gram-positive and Gram-negative bacteria.

• A major target for antibiotics.

Gram-positive Cell Wall

• Thick peptidoglycan layer (around 30 layers).

• Permeable to many substances (sugars and amino acids).

• Composed of peptidoglycan and teichoic acid.

• Some also contain lipoteichoic acids attached to the cytoplasmic membrane.

• The teichoic acids give the cell a negative charge.

Gram-negative Cell Wall

• Thin peptidoglycan layer.

• Outer membrane, a unique lipid bilayer with many proteins, including lipoproteins.

• The outside leaflet is composed of lipopolysaccharides (LPS).

• This layer acts as a protective barrier, making Gram-negative bacteria less sensitive to many medications.

• Contains porin proteins.

Lipopolysaccharide (LPS)

• Extremely important medically.

• Also known as endotoxin due to its toxic activity.

• Elicits symptoms associated with Gram-negative bacterial infections.

• Two notable parts: Lipid A (anchors LPS in the lipid bilayer; immune system recognizes it as an invader which is associated with endotoxic activity) and the O-specific polysaccharide side chain (O antigen).

Antibacterial Compounds Targeting Peptidoglycan

• Differences in prokaryotic and eukaryotic cells allow for development of antibacterial agents that target prokaryotic life processes.

• Some compounds interfere with peptidoglycan synthesis or alter its structural integrity, weakening the rigid molecule and causing cell bursting.

Penicillin

• Interferes with peptidoglycan synthesis.

• Binds to proteins/enzymes, preventing cross-linking of adjacent glycan chains (peptide interbridge formation).

• More effective against Gram-positive cells due to outer membrane preventing penicillin reaching peptidoglycan layer.

Lysozyme

• An enzyme found in bodily secretions (tears, saliva).

• Breaks bonds linking NAM and NAG in peptidoglycans.

• Destroys the structural integrity of glycans.

• More effective against Gram-positive bacteria due to outer membrane of Gram-negative bacteria

Bacteria Lacking a Cell Wall

• Example: Mycoplasma

• Lack a rigid cell wall, leading to variable shapes.

• Some cause mild pneumonia.

• Not affected by penicillin or lysozyme.

• Cell membrane is stronger, containing sterols.

Comparison of Gram-positive and Gram-negative Bacteria

• Summary table of key differences and structural components (see table 3.2 in the document).

Structures Outside the Bacterial Cell Wall: Capsules and Slime Layers

• Many bacteria have external layers.

• Distinct, gelatinous layer is a capsule.

• Diffuse, irregular layer is a slime layer.

• Capsules and slime layers allow attachment and protection, and can lead to biofilm formation.

• Important in adherence, pathogenesis, and biofilm formation (e.g. streptococcus mutans and dental plaque).

Other Surface Layers/Structures

• Flagella: Long protein structures for motility. May be peritrichous (all over the cell) or polar (at one or both ends). The filament is composed of flagellin.

• Pili/Fimbriae: Shorter protein appendages for attachment to surfaces, important in pathogenesis (e.g., enterotoxigenic E. coli). "Sex pili" transfer DNA.

Internal Components of Bacterial Cells: Nuclear Material

• Chromosome: A single, circular, supercoiled DNA molecule containing all genetic information. It resides in the nucleoid.

• Plasmids: Extra-chromosomal, double-stranded DNA molecules with additional genetic material.

• Plasmids carry genes for additional traits such as antibiotic resistance.

Ribosomes

• Organelles involved in protein synthesis.

• Composed of large and small subunits (prokaryotic are 70S; composed of 30S and 50S subunits; eukaryotic are 80S).

• Differences in ribosome structure are used as targets for certain antibiotics.

Endospores

• Dormant bacterial cells formed during unfavorable conditions.

• Extremely resistant to heat, desiccation, chemicals, and radiation.

• Germinate into vegetative cells under favorable conditions.

• Cause some medically important diseases, such as tetanus, botulism, gas gangrene, and anthrax.

Methods to Observe Bacterial Cells

• Often need to be immobilized and stained for observation due to rapid movement and transparency.

• Staining provides contrast between bacteria and surrounding medium.

• Simple staining uses a single dye and differential staining distinguishes cell types(Gram stain, Acid-fast stain).

Dyes and Staining Techniques

• Basic dyes: Positively charged, used to stain negatively charged cell components (nucleic acids, proteins). Examples include methylene blue, crystal violet, safranin, and malachite green.

• Acidic dyes: Negatively charged, used for negative staining (stain background).

• Differential staining: Distinguishes cell types, such as Gram staining.

Differential Staining

• Gram Staining: Distinguishes Gram-positive and Gram-negative bacteria based on their cell wall structure.

• Acid-fast staining: Distinguishes acid-fast bacteria (e.g., Mycobacterium) that have waxy cell walls from non-acid-fast bacteria.

Fluorescent Dyes and Tags

• Fluorescence: Used to observe total cells, subsets of cells (live or dead), or cells with specific proteins.

• Various dyes bind to certain compounds in cells.

• Examples: Acridine orange binds DNA; CTC dye binds to respiration proteins in live cells; Bac light in live vs dead cells; Calcofluor white binds to fungi and some bacterial cell wall components.