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Microbial Cell Structure,
Function, and Diversity
The Microbial World
What are microorganisms?
Organisms and acellular (virus-like) entities too small to be clearly seen by the unaided eye, generally less than 1mm, and often unicellular. Cellular microorganisms include fungi, protists, bacteria, and archaea. Acellular microorganisms include viruses, viroids, satellites, and prions.
Types of Microbial Cells
Prokaryotic cells lack a true membrane-bound nucleus, including bacteria and archaea. Eukaryotic cells have a membrane-enclosed nucleus and other membrane-bound organelles, are more complex morphologically, and are usually larger than prokaryotic cells.
Properties of all cells
Metabolism: Cells take up nutrients and transform them, expelling waste. Growth: Nutrients from the environment are converted into new cell materials. Evolution: Cells evolve to display new properties.
Properties of some cells
Differentiation: Some cells can form new cell structures such as a spore. Communication: Cells interact with each other by chemical messengers. Genetics exchange: Cells can exchange genes by several mechanisms. Motility: Some cells are capable of self-propulsion.
Evolution and Diversity of Microbial cells
Mutation of genetic material leads to new genotypes and advantageous phenotypes, which are then subject to natural selection. Bacteria and archaea are haploid, increasing genetic diversity through horizontal gene transfer within the same generation. The three main types of gene exchange are conjugation (sexual mating), transformation (picking up naked DNA), and transduction (virus-mediated).
The three domains of life (bacteria, archaea, and eukarya) evolved over billions of years, with bacteria being the first prokaryotes.
The importance of Microorganisms
Microorganisms are the oldest and most populous form of life, found everywhere on the planet, including underground. They play a major role in recycling essential elements and some carry out photosynthesis. Microorganisms influence all other living things, with most being beneficial or benign, and some being detrimental. Microorganisms are essential tools for study and have a significant impact on human health, agriculture, and industry.
Classification (universal phylogenetic tree)
The three-domain system, developed by Carl Woese, is based on a comparison of the DNA encoding small subunit ribosomal RNA. The three domains are Bacteria (prokaryotes), Archaea (prokaryotes), and Eukarya (eukaryotes). The genetic sequencing process for phylogenetic tree construction involves isolating DNA, making copies of the rRNA gene, sequencing the DNA, and generating the tree based on evolutionary distance.
Microorganisms of Domain Eukarya
Protists include algae, protozoa, slime molds, and water molds. Fungi include yeast and mold. Approximately 75% of the Earth's oxygen is produced by green algae and cyanobacteria.
Microbial Species
Bacteria and archaea do not reproduce sexually, so the term "species" has a different meaning, referring to a collection of strains that share many stable properties and differ significantly from other groups of strains. A microbial strain is a subset of a microbial species, consisting of the descendants of a single, pure microbial culture.
Naming Microbes
Microbes are named using the binomial nomenclature system, which includes the genus and species. The names can be derived from Greek, Latin, or the name of the discoverer. The name often reflects the shape or origin of the microbe.
Staining
Stains have two common features: chromophore groups and auxochrome groups. Chromophore groups are chemical groups with conjugated double bonds, while auxochrome groups enhance the staining ability of the chromophore.
Staining Techniques
Also gives stain its color
Stains can bind to cells and give them color, allowing for visualization under a microscope.
Ability to bind cells
Stains can bind to various cellular structures and components, enabling the identification and differentiation of microorganisms.
Types of stains
Basic stains (dyes)
Dyes with positive charges Bind to negatively charged structures
Acidic stains (dyes)
Dyes with negative charges Bind to positively charged structures
Basic stain procedure
Preparing a smear by spreading a culture and letting it air dry. Heat fixing and staining the smear. Microscopy by placing a drop of oil on the slide.
Types of staining techniques
Simple staining
Use of a single stain For visualization of morphological shape and arrangement Examples: Crystal violet, Methylene blue
Differential staining
Use of two contrasting stains separated by a decolorizing agent (e.g., water) Identifies microorganisms into groups based on their staining properties Examples: Gram stain, Acid-fast stain
Gram Staining
Created by Christian Gram Most widely used differential staining procedure Divides bacteria into two groups based on differences in cell wall structure: Gram-positive (purple) Gram-negative (pink)
Gram Staining Procedure
Flood the heat-fixed smear with crystal violet. Add iodine (mordant), which makes the crystal violet molecules larger. Decolorize briefly with alcohol. This makes the gram-positive peptidoglycan constrict and hold onto the crystal violet harder. Counterstain with safranin.
Acid-Fast Staining
Useful for staining members of the genus Mycobacterium High lipid (mycolic acid) content in the cell wall is responsible for their staining characteristics Acid-fast bacteria stain pink
Staining Specific Structures
Endospore staining
Heated, double staining technique Bacterial endospore is one color, and the vegetative cell is a different color
Capsule staining
Negative stain: capsules appear colorless against a stained background
Flagella staining
Mordant applied to increase the thickness of the flagella
Some common complex media components: Peptones: Protein hydrolysates Extracts: Aqueous extracts, usually of beef or yeast Agar: Sulfated polysaccharide used to solidify liquid media
Functional types of media
General purpose media (supportive)
Support the growth of many microorganisms
Enriched media
General purpose media supplemented with highly nutritious substances, such as blood For "picky" microorganisms
Minimal media
Contains the minimal necessities for growth of the wild-type Only inorganic salts, a simple carbon source, and water
Selective media
Favor the growth of some microorganisms and inhibit the growth of others Example: Media that selects for gram-negative bacteria
Differential media
Distinguish between different groups of microorganisms based on their biological characteristics Example: Blood agar, which differentiates between hemolytic and non- hemolytic bacteria
Isolation of Pure Cultures
Obtaining pure cultures
Pure culture: A population of cells arising from a single cell Allows for the study of a single type of microorganism Methods: Streak plate Spread plate Pour plate
Aseptic transfer
Flame loop
Sterilize the tube tip by flaming Only the sterilized portion of the loop enters the tube Tube is re-flamed and recapped; loop is resterilized
Streak plating
Sterilize the loop and remove a loopful of inoculum from the tube. Make the initial streak in one corner of the agar plate, then subsequent streaks at angles to the first. Incubation shows colonies of the bacterium on the plate.
Spread plate and pour plate
Spread plate: Small volume of diluted culture is transferred and spread evenly over the agar surface. Pour plate: Diluted samples are mixed with liquid agar, and the mixture is poured into sterile culture dishes. Both methods provide isolated colonies and the opportunity to enumerate the bacteria in a sample.
Microbial growth on solid surfaces
Species form characteristic colonies. Differences in growth rate from the edges to the center are due to oxygen, nutrients, and toxic products. Cells on the outside are younger because of more oxygen.
Microbial Cell Structure and Function
Size, shape, and arrangement
Common shapes: cocci (spheres) and bacilli (rods) Arrangements determined by the plane of division and the degree of separation after division Size varies between microorganisms
Cocci
Diplococci: Pairs Streptococci: Chains Staphylococci: Grape-like clusters Tetrads: 4 cocci in a square Sarcinae: Cubic configuration of 8 cocci
Bacilli
Rods, coccobacilli (very short rods) Vibrios: Comma-shaped Spirilla: Rigid helices Spirochetes: Flexible helices
Membrane structure
Lipid bilayer composed of phospholipids Amphipathic: Polar ends (hydrophilic) and non-polar tails (hydrophobic) Saturation levels reflect the environment Hopanoids add structural stability
Membrane proteins
Peripheral: Loosely connected to the membrane (20-30% of membrane proteins) Integral: Embedded within the membrane (70-80% of membrane proteins), carry out important functions
Bacterial Cell Wall
Rigid structure that lies just outside the plasma membrane Contains peptidoglycan Functions: Provides shape to the cell Protects from osmotic lysis May contribute to pathogenicity Protects from toxic substances
Bacterial Cell Walls
Gram-Positive Cell Walls
Composed primarily of peptidoglycan (up to 90% of wall) May also contain teichoic acids, which are negatively charged and help maintain the structure of the cell wall, bind calcium and magnesium, and protect from harmful substances Teichoic acids can be attached to the cell membrane (lipoteichoic acids) or the peptidoglycan layer Some gram-positive bacteria have a layer of proteins on the surface of the peptidoglycan
Gram-Negative Cell Walls
Consist of a thin layer of peptidoglycan surrounded by an outer membrane Outer membrane is composed of lipids, lipoproteins, and lipopolysaccharide (LPS) Peptidoglycan makes up only about 10% of the cell wall Periplasm may constitute 20%-40% of the cell volume and contains many enzymes, including hydrolytic enzymes, binding proteins, and chemoreceptors Outer membrane has porin channels through which small, hydrophilic molecules can pass
Braun's lipoproteins connect the outer membrane to the peptidoglycan layer
Lipopolysaccharide (LPS)
LPS has three main parts: Lipid A Core polysaccharide O-specific polysaccharide (O-antigen) Lipid A is embedded in the outer membrane, while the core polysaccharide and O-side chain extend from the cell LPS contributes to the negative charge on the cell surface, helps stabilize the outer membrane structure, may contribute to attachment and biofilm formation, creates a permeability barrier, and can act as an endotoxin (lipid A)
Comparison of Gram-Positive and Gram-Negative Cell
Walls
Cell walls provide osmotic protection: In hypotonic environments, the cell wall protects the cell from lysis as water moves into the cell and it swells In hypertonic environments, the cell wall protects the cell from dehydration as water moves out of the cell Penicillin inhibits peptidoglycan synthesis, and without a cell wall, cells will lyse in hypotonic solutions Protoplasts (gram-positive without cell wall) and spheroplasts (gram- negative without peptidoglycan) can survive in isotonic environments
Bacterial Capsules, Inclusions, and Endospores
Capsules and Slime Layers
Capsules are usually composed of polysaccharides and are well- organized, not easily removed from the cell Advantages of capsules include attachment to surfaces and each other (biofilms), protection against desiccation, phagocytosis, viruses, detergents, toxins, antimicrobials, and absorption of nutrients Slime layers are similar to capsules but are diffuse, unorganized, and easily removed; they may aid in motility
Cell Inclusions
Aggregates of organic or inorganic material, such as: Storage inclusions (e.g., glycogen, poly-β-hydroxybutyrate, polyphosphate granules, cyanophycin granules) Carbonate minerals (biomineralization) Gas vesicles (to help cells float) Magnetosomes
Spread by direct contact through broken skin, mucous membranes, saliva, or brain/nervous tissue from an infected animal Symptoms include weakness, fever, headache, prickling/itching at the bite area, anxiety, confusion, agitation, delirium, and hallucinations
Pertussis
Caused by the gram-negative, encapsulated, coccobacillus bacterium Bordetella pertussis Spread by coughing or sneezing Symptoms include uncontrollable coughing, a high-pitched whooping sound, runny nose, congestion, and fever
