Lec. 1-3: Microbiology Foundations, Microbiology History, Tree of Life

Human Microbiota:

The complete collection of microorganisms naturally associated with human skin, mucous membranes, and the gastrointestinal tract

Industrial Role of microorganisms in Biotechnology:

Essential for producing antibiotics, vitamins, and industrial enzymes

Why is only a minor subset of microorganisms pathogenetic?

The vast majority of microorganisms are innocuous or highly beneficial to humans (eg. microbiota); only a minor subset causes severe disease

Three biological kingdoms/domains:

1. Bacteria

2. Archaea

3. Eucarya

Approximately how many species remain undiscovered/unknown?

99.99% ; An estimated minimum of 1 trillion species exist on Earth

Fundamental Attributes of Biological Life:

1. Growth

2. Metabolism

3. Motion; self-directed motility

4. Reproduction; produce separate, identical offspring entities

5. Response to Stimuli: Sensing environmental properties and dynamically reacting to changing external conditions

Lucretius, Avicenna, Fracastoro; Early disease hypothesis

Suggested invisible organisms cause disease

Zacharias Janssen; Development of Microscopy

A Dutch spectacle maker who invented and developed the first compound microscope using two lenses, achieving 3X - 9X magnification

Robert Hooke; Development of Microscopy

Depicted microfungus Mucor in 'Micrographia'; first recorded description and detailed illustration of a microorganism

Antonie van Leeuwenhoek; Development of Microscopy

Discovered live "animalcules" (bacteria/protozoa);

The Theory of Spontaneous Generation (Proposed by Aristotle and believed by Isaac Newton)

States that living organisms spontaneously form or develop directly from inanimate matter (eg. mud was believed to generate worms)

Francesco Redi's Experiment Objective:

Tested whether fly larvae (maggots) generate spontaneously from rotting meat

Francesco Redi's Experiment Setup and Results:

• unsealed flask: flies visited the meat —> maggots developed on the meat

• sealed flask: flies could not reach the meat —> no maggots developed

• Gauze-Covered Flask: flies attracted to the odor but kept out —> maggots developing on top of the gauze

Conclusion: Redi demonstrated that maggots only develop when adult flies have physical access to lay eggs, thereby disproving spontaneous generation for macroorganisms

Why does Microbiology involve the study of organisms too small to be clearly seen by the unaided eye

Most microorganisms are unicellular

Microorganism Dilemma (debate)

Scientists debated (continued after Redi’s experiment) whether;

• decomposition produced microorganisms

• microorganisms actively cause decomposition

Needham vs. Spallanzani (spontaneous generation debate)

• John Needham: boiled mutton broth, placed into flask, sealed after boiling —> Microorganisms grew —> Suggested results supported spontaneous generation

• Lazzaro Spallanzani: broth into flask, sealed first, then boiled —> No microorganisms —> claimed he disproved spontaneous generation

Louis Pasteur's Swan-Neck Flask Experiment

• Design: boiled meat broth in swan-neck flasks, allowing air to freely enter the flask while trapping airborne dust particles and microbes in the bend

• Results: Broth remained sterile in intact flask

John Tyndall and Dust-Borne Microorganisms

Dust-Free Box Experiment: confirmed Pasteur's findings by demonstrating that airborne dust carries microorganisms

Early Evidence Linking Microorganisms to Disease:

1. Agostino Bassi: Showed fungus causes silkworm disease (Germ Theory start)

2. M.J. Berkeley: Showed fungus causes Irish Potato Blight

Joseph Lister & Antiseptic Surgery

Introduced phenol / heat sterilization in surgery

Robert Koch & Anthrax

Proved Bacillus anthracis causes anthrax (Injected healthy mice with material from diseased animals/ endospores)

Gelatin Limitations: Solidifying liquid media with gelatin failed due to two major flaws:

• Flaw 1: Degraded by microbes

• Flaw 2: Melts at 37ºC

Which solid media techniques allowed Koch to successfully isolate Mycobacterium tuberculosis?

Agar + Petri dish + enriched media (resembling body fluids)

"Father of Immunology"; Immunized individuals against smallpox using fluid from cowpox lesions

Edward Jenner

Attenuated live vaccines mode of preparation:

Made by growing pure pathogen cultures and treating them with heat (42–43°C) or potassium bichromate

Daniel Elmer Salmon & Theobald Smith:

Discovered that killed microbial cells are also effective as protective vaccines

Humoral Immunity:

antibodies produced in the blood neutralize toxins

Cellular Immunity:

phagocytes (white blood cells / leukocytes) directly engulf disease-causing bacteria to confer protection

Pasteur's vaccine developments:

Developed attenuated vaccines for anthrax (Bacillus anthracis), avian cholera (Vibrio cholerae), and rabies (rabies virus)

Taxonomy:

Artificial classification system based on visible or morphological similarities

Phylogeny:

Natural classification system that reflects the true evolutionary relatedness and historical lineage between organisms, using ubiquitous gene sequences (shared across all living entities)

Carl Woese and Colleagues

Revolutionized classification by introducing a molecular approach based on 16S ribosomal RNA (rRNA) gene sequencing; established the Three-Domain/Kingdom Scheme

Koch's Postulates; A set of four criteria established to prove a causal relationship between a specific microorganism and a specific disease (define pathogen)

1. Presence: The microorganism must be present in every case of the disease, but absent from healthy organisms

2. Isolation: The suspected microorganism must be isolated and grown in a pure culture

3. Inoculation: The same disease must result when the isolated microorganism is inoculated into a healthy host

4. Re-isolation: The same microorganism must be isolated again from the experimentally diseased host

Methodology for Building a Molecular Phylogenetic Tree

1. Gene Selection: Obtain the sequence of a ubiquitous marker gene present in all target organisms

2. Sequence Alignment: Align the gene sequences across all organisms using specialized sequence alignment software

3. Algorithmic Distance Calculation: Feed the aligned sequences into a phylogenetic algorithm that identifies sequence homology and links closest evolutionary neighbors

4. Tree Generation: Branching patterns are created based on sequence similarity

What is the length of the Phylogenetic Tree branches connecting organisms directly proportional to?

The evolutionary distance between organismsWhi

Which organelles do plants/algae posses?

Both mitochondria and chloroplast (eukaryotes possess only mitochondria)

How do bacteria and eukaryotes differ in terms of organellar structures?

prokaryotes lack organellar structures; eukaryotes possess only mitochondria

prokaryotic features of mitochondria and chloroplasts:

• Physical Dimension: Measure ~1 µm in diameter (identical to typical bacteria)

• Genome Autonomy: Possess their own active, circular DNA genomes.

• Ribosomal Machinery: Contain 70S ribosomes (characteristic of prokaryotes), distinct from eukaryotic 80S cytosolic ribosomes6.

• Membrane Architecture: Possess a double-membrane system resembling the inner cytoplasmic and outer membranes of Gram-negative bacteria (though lacking a peptidoglycan cell envelope).

• Replication: Multiply independently via binary fission

Lynn Margulis Proposal

Theory: A primitive prokaryote (an ancestral archaeon) engulfed an ancient aerobic heterotrophic bacterium to generate the first eukaryotic cell

First Prokaryotes:

~3.5 billion years ago

First Eukaryotes:

~1.0 billion years ago

Endosymbiont Benefit:

Gained a sheltered, nutrient-rich environment inside the host cell

Host Cell Benefit of Endosymbiosis:

Gained an efficient internal energy source generating ATP via respiration (mitochondrion) or photosynthesis (chloroplast)

Mitochondria Descendants:

Evolved from an ancient 𝜶-proteobacterium; closest modern relative is Rickettsia prowazekii (a Gram-negative, obligate intracellular parasite)

Chloroplast Descendants:

Evolved from an ancient cyanobacterium

Direct experimental support for the endosymbiotic theory:

Obligate Interdependence: endosymbiont not viable outside the host, and the host dies if its endosymbionts are killed

Primary Goals of Classification:

1. Establish stability and predictability in biological naming (nomenclature)

2. Construct a hierarchical framework

3. Enable predictive learning: studying one representative member provides generalizable knowledge about the entire group

The 7-Rank Taxonomic Hierarchy

1. Kingdom/Domain

2. Phlyum

3. Class

4. Order

5. Family

6. Genus

7. Species (Basic Taxon)