MBIO 1220

• Prokaryotes

• Eukaryotes

• Acellular Infectious Agents

• Carl Linnaeus (1700s)

• Genus Species

  • Ex. Homo sapien

Essential component of a healthy human

• Group of microorganisms living in/on your body & don’t usually cause disease

• Prevent disease by competing with pathogens

• Aid with digestion

• Promote immune system development

Disease-causing microbes

• Used crude microscope to view individual cells

  • Bread mold (“Microscopial Mushroom”)

  • Beginning of Cell Theory

All living organisms are composed of cells

• Amateur lens grinder

• Built microscopes that could view living microorganisms

  • Called microorganisms “little animacules”

Spontaneous Generation: Forms of life can arise from non-living matter.

Biogenesis: Living matter only arises from pre-existing life forms.

• Used swan neck flasks to disprove spontaneous generation.

• Showed that microscopic yeast (fungi) converts sugar to alcohol using fermentation.

  • Absence of oxygen - anaerobic

• Souring occurs when bacteria turns the alcohol into vinegar.

  • Presence of oxygen - aerobic

• Solution: Heat beer or wine after fermentation to kill bacteria and prevent spoilage - pasteurization.

Microorganism cause certain diseases.

• Used Phenol to clean surgical instruments and treat surgical wounds.

• Drastically reduces incidents surrounding surgical wound infections.

  • Led to development of disinfectants and antiseptics.

• first proof that bacteria cause disease.

• Investigated cause of anthrax.

• Establish a sequence of experiment steps that could be used to find the causative agent of other diseases

• Isolated bacteria

• Showed that particular bacterium was present in all cases of the disease

• Injected bacterium into healthy specimens

• Specimen contracted anthrax and died

• Re-isolated bacteria from injected specimens and showed they were identical to him first sample

• Showed that specific microbe was the cause of particular disease

• Developed smallpox vaccine

• Observed that people previously sick from cowpox did not contract smallpox.

• Purposefully inoculated a young boy with cowpox, fell ill, recovered and became immune to smallpox.

• Noticed that certain dyes stain bacteria differently than they stained a animal cells

• Proposed that a chemical might be found that would harm disease causing microbes without harming the host.

• Discovered Salvarsan, an arsenic derivative that could be used to treat syphilis.

Ability to drug target sites relative to the microorganism responsible fro infection. Some sites are unique to microorganism or more essential to the survival of the microorganism than the host.

• Noticed mold inhibited bacterial growth on contaminated plates.

• Produced first natural compound, penicillin.

  • First antibiotic

    • Compound naturally produced by molds or bacteria, inhibiting he growth of or kills other microorganisms.

• Light Microscope that uses visible light to observe a specimen

• Compound Light Microscope: Uses two lenses to magnify image

  • Objective Lens: Lens closest to specimen & can magnify between 4x - 100x

  • Ocular Lens: The eyepiece & magnifies by 10x

• Calculating magnification:

  Total Magnification = Objective lens x Ocular Lens

• Ability to distinguish fine detail & structure

• Ability to distinguish 2 points a certain distance apart

• Light must pass between 2 objects for them to be seen as two separate things.

• Need light of a short enough wavelength to fit between them, otherwise will appear as one object.

• Resolution General Principle: the shorter the wavelength, the better the resolution.

• Electrons instead of light, electrons travel in much shorter waves.

• Resolving power is greater

• Allows for greater magnification

• Allows us to view viruses and internal cell structures

• See internal structures

• Very thin slices can be cut from sample

• Samples generally stained with a metal to make structure opaque to electrons

• See surfaces, less powerful

• See’s molecules

  • Uses thin metal probe to scan a specimen revealing bumps and depressions

• Microorganisms are almost colourless when seen though a microscope, stains make them easier to see.

• Stains are composed of positively and negatively charges ions, one of which is coloured (chromophore).

• One dye is used to highlight the entire microorganism

• Steps:

  • Smear sample on slide

  • Fix with heat

  • Add stain

  • Was, dry, and view

Bacteria have a net negative charge on their outer surface, this charge attracts stains with positively charged chromophores and repels stains with negatively charged chromophores.

• Stain binds to bacterium

• Bacterium appears coloured

• Background appears clear

• Will not bind to bacterium

• Bacterium appears clear

• Background is coloured

• React different with bacteria, thus can be used to distinguish between them.

• Eg. Gram Stain

  • Differentiates bacteria based on the structure of the cell wall

• Gram Positive: Bacteria with a thick cell wall retain the primary stain crystal violet and appear purple.

• Gram Negative: Bacteria with a thin cell wall lose crystal violet during destaining, take on the colour of counterstain safranin and appear pink.

• Stains internal structures of some bacteria

• Primary stain colours endospores green

• Counterstain colours the rest of cell red

• Stains external structures

• Mordant (dye fixative) thickens flagella so they can be observed under light microscope

• Detects presence of waxy compound in cell wall

• Used to identify the genus Mycobacterium

• Mycobacterium cell wall retains dye carbol fuschin

• Counterstain with methylene blue stains non acid-fast bacteria and tissues blue

• Detects thick layer of polysaccharide outside the cell.

  • Negative stain colours background

  • Positive stain colours the cell

  • Does not take up most dyes and remains colourless.

• DNA not enclosed within a nucleus

• Usually arranged as one circular chromosome

• Lack membrane bound organelles

• Single celled organisms: Bacteria, Archaea

• DNA found in nucleus, surrounded by nuclear membrane

• DNA arranged as multiple chromosomes

• Membrane bound organelles

• Single celled or multicellular organisms

  • Algae, Protozoa, Fungi, Plants, Animals

Morphology

• Coccus - Spherical

• Bacillus - Rod

• Vibrio - Curved

• Spirillum - Spiral Shaped

• Spirochete - Corkscrew Shaped

• Sticky, gelatinous layer external to the cell

• Composed of polysaccharides, protein, or both

• If layer is organized and firmly attached to cell wall, capsule

  • In some bacteria capsules a play a role in virulence

  • Protection against phagocytosis

  • Streptococcus pneumoniae

    • With capsule: causes disease

    • Without capsule: no disease

• Sticky, gelatinous layer external to the cell

• Composed of polysaccharides, protein, or both

• If layer is unorganized and loosely attached to the cell wall, slime layer

  • Often allow bacteria to attach to surfaces

  • Medical Implants, water pipes, teeth

    • Streptococcus mutans

      • Makes polysacccharide slime from sucrose

      • Attaches to teeth, leading to cavities

• Long protein appendages

• Used in motility

• Semi rigid helical, turns like a propeller

Monotrichous: Single polar flagellum

Lophotrichous: Two or more flagella originating from one pole

Amphitrichous: Tufts of flagella originating from opposite poles

Peritrichous: Flagella distributed all over the cell

• Flagella turn causing cell to move in one direction - “run”

• Periodically flagella reverse direction causing a random change in direction - “tumble”

• Flagella allow chemotaxis

  • Movement toward or away from a stimulant

  • Toward nutrients (attractant)

  • Away from toxins (repellent)

  • Flagella protein can be used to distinguish among strains of species

• Short, hair like appendages

• Hollow

  • Allow cell to adhere to surfaces

  • Contribute to pathogenicity

• Short, hair like appendages

• Hollow

  • Allows attachment of two bacteria to each other

  • Involved in transfer of genetic material between bacteria

1. Pilus Formation

   The donor cells (F+ cells) form a sex pilus and begin contact with an F- recipient cell.

2. Physical Contact between Donor and Recipient Cell

   The pilus forms a conjugation tube and enables direct contact between the donor and the recipient cells.

3. Transfer of F-Plasmid

   The F-factor opens at the origin of replication. One strand is cut at the origin of replication, and the 5’ end enters the recipient cell.

4. Synthesis of Complementary Strand

   The donor and the recipient strand both contain a single strand of the F-plasmid. Thus, a complementary strand is synthesized in both the recipient and the donor. The recipient cell now contains a copy of F plasmid and becomes a donor cell.

• Semi Rigid structure giving shape to the cell

• Major function is to prevent rupture of the cell - protects against environmental changes

• Useful in identification of bacteria - ie. Gram Stain

• Composed of peptidoglycan

• Mesh-like structure composed of polysaccharide and amino acids

• Polysaccharide portion composed of 2 alternating monosaccharide covalently joined

  • N-acetyl glucosamine (NAG)

  • N-acetyl muramic acid (NAM)

• Peptide portion composed of short changing of amino acid

• Polysaccharide chains run parallel

• Peptide chains link polysaccharides together

• Forms a mesh-like net surrounding cell

• Completely different from anything found in animal cells

• Many antibiotics have been discovered that act against peptidoglycan

  • Penicillin: inhibits production of peptidoglycan

  • Lysozyme: Degrades and is found in tears, saliva, mucous

• Made of thick layers of peptidoglycan outside of plasma membrane

  • Contains teichoic acids

    • Wall teichoic acids: Attached to peptidoglycan

    • Lipoteichoic acids: Attached to plasma membrane and extend through peptidoglycan

• Have only one membrane - cytoplasmic membrane

• Thin peptidoglycan layer sandwiched between two membranes

• Outer membrane made of phospholipid, proteins, and lipopolysaccharide (LPS)

• Polysaccharide portion of LPS is composed of O-sugars

  • Useful for distinguishing negative bacteria

• Lipid portion of LPS is toxic

  • Referred to as endotoxin

• Thick peptidoglycan traps crystal violet - stain purple

• Thin peptidoglycan does not trap crystal violet, and outer membrane gets disrupted by alcohol

• Crystal violet is washed away

• Safranin counterstain stains cells pink

• Composed of phospholipid bilayer

• Separates interior from outside environment

  • Serves as a semi-permeable barrier

  • Selectively allowed inflow and outflow of materials

• Exists in a semi-fluid state

• Alcohols disrupt the membrane

• Can be used as disinfectant

• Substance inside plasma membrane

  • 80% water

  • Contains most of the stuff needed for life

    • Sugars, amino acids, nucleotide, etc.

    • Enzymes

    • Some functional structures

• Contains bacterial chromosome (DNA)

  • All genetic information required for cell’s structures and functions

• Not surrounded by nuclear membrane

• May contain plasmids

  • Smaller double stranded DNA molecules

  • Contain non-essential genes - eg. Genes for antibiotic resistance

• Site of protein synthesis (translation)

• Made of protein and ribosomal RNA (rRNA)

• Two Parts:

  • 30s Subunit

  • 50s Subunit

    • Together form the complete 70s ribosome

• Ribosomes of bacteria differ from eukaryotic ribosomes

  • Eukaryotes have 80s ribosomes

  • Several antibiotics target bacterial ribosomes

  • Prevent bacteria from making new proteins

• Usually deposits or granules of nutrients, stored for late use

• Examples:

  • Sulfur granules

  • Polysaccharide (Glycogen)

  • Lipid inclusions

  • Enzymes

  • Magnetite

• Variety of inclusion bodies occur in different bacterial species - can serve as a basis for identification

• Formed only by some Gram-positive bacteria

• Special resisting structure - allows bacteria to enter dormant state

• Extremely durable

  • Resist heat, desiccation chemicals, radiation

  • Some endospores can survive in boiling water for hours

• Remains dormant until good growth conditions occur

  • Can for new population

1. Cell replicates its DNA

2. Septum forms, dividing cell into unequal compartments

3. Larger compartment engulfs the smaller

4. Peptidoglycan and other protective material forms around the foreshore - spore coat

5. Finished spore is freed from the mother cell as the mother cell dies

• Includes microorganisms algae, fungi, protozoa and higher organisms, plants, animals.

  • Larger and more complex than prokaryotes

  • Genetic material housed in nucleus

  • Membrane bound organelles

• Composed of phospholipid bilayer

• Separates interior from outside environment

  • Serves as a semi-permeable barrier

  • Selectively allowed inflow and outflow of materials

• Exists in a semi-fluid state

• Contains phospholipids, proteins, and sterols

  • Sterols make membrane rigid compared to bacteria

• Not all eukaryotes have one

  • Allows endocytosis

• Simple structure compared to bacteria

• Made of:

  • Cellulose

  • Chitin

• Substance inside plasma membrane but outside nuclear membrane

• Has complex internal structure - Cytoskeleton

  • Protein filaments on the inside of plasma membrane

  • Provides support and shape

  • Transports substance through the cell

• Larger and heavier than bacterial ribosomes

  • 80s

• Several antibiotics target bacterial ribosomes

• Prevent bacteria from making new proteins

• Structures with specialized functions

• Not present in bacteria

• Example:

  • Mitochondria: Site of energy production

  • Chloroplast: Site of photosynthesis in algae and plant cells

Flagellum & Cillia

• Long flexible projections that contain protein and cytoplasm

• Move in whip-like fashion

• Can be used for:

  • Motility

  • Sweeping material past stationary cells

• Has 9 + 2 array (9 pairs of microtubules with 2 in enter of ring)

  • Microtubules: Long, hollow tubes made of protein called tubulin

• Refers to increase in bacterial cell numbers

  • Not an increase in size of individual cells

• Most bacteria reproduce by binary fission

• Bacterial cell:

  • Elongates and makes copy of its DNA

  • Divides into two identical cells

• Binary fission → Population of cells double every generation

• Time required for population to double = generation time

• Varies greatly between different bacteria

• Inoculation: Introducing microbes into a medium to start culture

• Culture: Microbes Growing in a medium

• Closed System

• Once started, no other nutrients added

• When nutrients are used up - bacteria stomp growing

• Open system

• Nutrients are continuously added, wasted are continuously removed

  • Supports indefinite growth

• Period of adaptation

  • Cells adjust to new media and prepare to grow

• Period of maximal reproduction - cell numbers increase exponentially

  • Used to calculate generation time

• Cells have reached maximum population density

• Nutrients have been used in, or wasted have accumulated

• No increase in cell number

• Toxic waste products have accumulated

  • Cells die at uniform rate

• Sometimes a small fraction of population survived the death phase

• May consume nutrients release from dying cells

• Selects for the strongest cells in population

• Temperature

• Oxygen

• pH

• Osmotic Pressure

• Nutritional Factors

• Nutritional Diversity

• Each microbe species has its own temperature range

  • Usually spans about 30°C

    • Maximum: Lowest temperature supporting growth

    • Optimum: Temperature supporting best growth

    • Maximum: Highest temperature supporting growth

• Cold loving

• Grows between 5°C - 15°C

• Killed at 20°C

• Very broad temperature range

• Minimum: ~5°C

• Maximum: ~30°C - 45°C

• Optimum: 15°C - 30°C

• The microbes that cause food to spoil in fridge

• Moderate temperature loving

• Minimum: ~10°C

• Maximum: ~45°C

• Optimum: 25°C - 45°C

• Most bacteria

• Most pathogens have temperature optimum of 37°C

• Heat loving

• Minimum: ~40°C

• Maximum: ~80°C

• Optimum: 65°C

• Minimum: ~75°C

• Maximum: ~121°

• Restricted to very few places on earth where water reaches these temperatures

  • E.g. Deep ocean vents

• Involves use of both hot and cold temperatures

• Heat is used to kill mesophilic and psychrotrophic microbes

• Cold temperature is used to slow growth

  • Only psychrotophs will grow in a refrigerator - and slowly

• Require O2 for respiration (energy generation)

• Can use O2 for respiration but also grow in it its absence

• Cannot use O2 and are killed by it

• Require O2 in low amounts, but killed in high concentrations

• Cannot use O2, but are not killed by it

• Measurement of acidity or alkalinity

  • pH < 7 = Acidic

  • pH > 7 = Alkaline

  • pH of 7 = Neutral

• Most bacteria grow at or near neutral

• Bacteria that grow at low pH are Acidophiles

• Bacteria that grow at high pH are Alkaliphiles

• Movement of solvent molecules across a semi-permeable barrier

  • E.g. Movement of water through cytoplasmic membrane

• H2O will move from area of high concentration to low

• High solute concentration

• Ex. Salt or sugar

• Water flows out of cell

• Cell dries up - Plasmolysis

• Low solute concentration

• Water flows into cell

• Cell bursts - Osmotic Lysis

• Condition where solute concentration on outside of cell is equal to that inside the cell

• Some bacteria have adapted to life in high salt concentrations - requiring up to 30% NaCl

  • Extreme Halophiles

• Blood has a salt concentration of about 0.9%

  • Does not inhibit the growth of most microorganisms

• required for all organic molecules - backbone of living matter

• Heterotrophs - Take carbon from organic matter

• Autotrophs - Use inorganic carbon

• Required in smaller amounts for synthesis of cellular material

• E.g. Protein, nucleic acids, phospholipids, ATP

• Required in very small amounts

• E.g. Iron, zinc, molybdenum

• Essential to functions of certain enzymes