MICR20010 Lecture 4 2024 PDF

MICR20010 Lecture 4 Basic microbial morphology Dr. Jennifer Mitchell Microbiology School of Biomolecular and Biomedical Science  Lecture 3 Light Microscopy Preparing bacterial cells for microscopy Light microscope resolution Electron Microsco...

MICR20010 Lecture 4 Basic microbial morphology Dr. Jennifer Mitchell Microbiology School of Biomolecular and Biomedical Science  Lecture 3 Light Microscopy Preparing bacterial cells for microscopy Light microscope resolution Electron Microscope – Scanning Electron Microscope – Transmission Electron Microscope Domains of Life Types of microorganism Eukaryotic Cell Structure Eukaryotic Versus Prokaryotic Cells  Learning Outcomes Prokaryotic cell morphology Bacterial cell structure The gram stain – Gram stain mechanism Bacterial shapes – Different morphological shapes Bacterial cell structure – G+ve G-ve Archaea – Cell membrane – Cell wall – Outer membrane – Cell appendages  Prokaryotic cells Bacteria Archaea Gram positive Gram negative Bacterial Cell Structure  In the lab! YOU MUST PRINT OUT THE LAB MANUAL BEFOREHAND!!! YOU MUST BRING A NOTEBOOK TO RECORD OBSERVATIONS!!! YOU MUST READ THE INSTRUCTIONS AND VIEW ONLINE MATERIAL BEFORE THE LAB!!!!!!! Bunsen Burner  The Gram Stain Most important differential staining method in Microbiology Gram-positive Gram-negative (staphylococci) (Escherichia coli) 1. Crystal Violet 2. Iodine 3. Alcohol 4. Neutral Red Gram Stain Use tongs to fix smear!!  The microscope Oil must be removed from 100x lens immediately after use using lens tissue.  Gram Stain Mechanism Differential lipid content of G+ and G- cell envelopes Crystal violet-iodine complex forms within the cells (Blue colour) Alcohol treatment G+ cell envelope has low lipid G- cell envelope has content and is dehydrated by high lipid content which alcohol - making it impermeable is extracted by alcohol to permeabilise the membrane Crystal violet-iodine complex diffuses out and neutral red is taken up Streaking out a mixed culture Microbes are everywhere a. Air, outside and inside; b. fingertips, before or after washing or after touching these to your hair; c. Soil; d. Water, a drop from a tap; e. Blade of grass; f. A drop of milk; g. Leaf of a plant Incubate at 27°C  LAB WRITE UP Instructions on page 2 of practical manual!!! Bring sharpie to write on petri dishes. You have one week to write up and submit on Brightspace. Ask all questions to your demonstrator in the lab. Do not leave until all your questions are answered! Some bacteria don’t stain using the Gram method Mycobacteria have a high wax content in their cell envelope and suspected mycobacteria are stained using the Ziehl- Nielson stain Mycoplasmas, the smallest known bacteria, have no cell wall to stain  Bacterial Shapes Cocci (spherical) Bacilli (rod shaped) Curved or spiral shaped Morphological Shapes of Different Bacteria Thiomargarita magnifica  Bacterial Cell Structure Chromosome The bacterial chromosome contains the bacterial genetic information. Plasmids may also be present. Cytoplasmic Membrane The cytoplasmic membrane surrounds the cytoplasm Cell Wall Rigid layer surrounding the cytoplasmic membrane Outer Membrane of Gram-negative bacteria Covers the cell wall and acts as a molecular sieve  Typical Gram-negative and Gram-positive Cell Envelopes  Cytoplasmic Membrane Composed primarily of lipids and phospholipids Osmotic barrier – Only molecules smaller than glycerol diffuse into the cytoplasm Site of energy production oxidative phosphorylation Transport of important molecules via Permeases Facilitated diffusion (passive) and Active transport Synthesis of new cell wall Anchor the chromosome  Cell Wall Domains of Life Cell wall composed 1. Eukaryotes primarily of peptidoglycan 2. Bacteria Prokaryotes 3. Archaea Lack peptidoglycan - wall composed of other polysaccharides or proteins  Function of the cell wall Bacterial cells contain high concentrations of dissolved solutes (salts, sugars etc). Generates a high pressure within the cell caused by the cytoplasm pressing against the cell envelope (similar to pressure in car tyre) Cell wall allows cell to withstand turgor pressure Gives the cell shape and rigidity  Bacterial Cell Wall Peptidoglycan = the principal component of the cell wall, is a unique polysaccharide which gives the cell its characteristic shape and prevents osmotic lysis Gram-positive Gram-negative Many peptidoglycan layers One peptidoglycan layer (90% of cell envelope material) (2-20% of cell envelope material) Penicillin disrupts peptidoglycan synthesis Many antigens are presented on cell wall surface  Peptidoglycan NAG = N-acetylglucosamine NAM = N-acetylmuramic acid  Basic structure of the peptidoglycan disaccharide unit (left) and multiple peptidoglycan units liked to give the cell wall structure (right Amino acids G = N-acetylglucosamine G-M: b 1,4 glycosidic bond M = N-acetylmuramic acid Gram-positive cell envelope  Cell walls of Archaea No peptidoglycan S-layer composed of a ordered layer of protein or glycoprotein – Examples: Many thermophiles, halophiles, methanogens Few Archaea contain pseudopeptidoglycan – (Repeated sugar units, however, ab1,3-linked) – Example: Methanogens Polysaccharides  Gram-negative Cell Envelope: Outer Membrane Phospholipid-Lipopolysaccharide (LPS) Bilayer (extra lipid layer - mechanism of the Gram stain) Bacterial cell adhesion Resistance to phagocytosis Molecular sieve - access of some molecules to cell wall and cytoplasmic membrane LPS vs Phospholipid Gram-negative cell envelope  Gram positive cell surface Note different surface textures Gram negative cell surface  Cell Appendages and other Cell Structures Flagella and Pili extend from the cell surface Flagellae rotate and are required for motility (chemotaxis) Bacteria swim towards chemoattractants and away from chemorepellents Flagella Bacteria use flagella to swim. Changing the direction of the flagellar rotation can cause the cell to tumble and change direction.  Pili (from latin for hair) Common Pili - adherence UTI’s Conjugative Pili - plasmid transfer  Further Reading Microbiology an Introduction, Tortora, Funke and Case 12th Ed. Chapter 4 “Functional Anatomy of Prokaryotic and Eukaryotic Cells”