Chapter 1 Part 2: Introduction to Microbiology — Page-by-Page Notes
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Document title: Chapter 1 Part 2 Introduction to Microbiology: Classifying and Growing Microbes
Course: Microbiology (University of the Incarnate Word)
Instructor: Dr. Christopher Pierce
Copyright: © 2023 Pearson Education, Inc. All Rights Reserved
This page sets the context for the module on classifying microbes and growing them in the lab.
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1.2 Learning Objectives — Classifying Microbes
After studying this section, you should be able to:
Summarize the taxonomic hierarchy of domains and kingdoms.
Describe the binomial nomenclature system and the information it provides about an organism.
Define the terms species and strain as they relate to prokaryotes.
Compare and contrast parasitism, mutualism, and commensalism.
Define the term normal microbiota and discuss its roles.
Describe how a biofilm forms and discuss the healthcare implications of biofilms.
Provide examples of how microbes impact industry and the environment.
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Naming and Classifying Microorganisms
Carl Linnaeus (1707–1778):
Father of taxonomy.
Established criteria for classifying organisms.
Taxonomy: the science of grouping organisms by shared features (character traits) and relationships.
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Taxonomic Hierarchy: Domains — The Broadest Grouping
Domain is the broadest grouping of organisms.
The three domains are:
Bacteria — unicellular, prokaryotic organisms.
Archaea — some live in extreme environments; no known pathogens among the classic broad groups.
Eukarya — unicellular and multicellular eukaryotic organisms.
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Taxonomic Hierarchy: Domains, Kingdoms, and Major Groups
Domain Bacteria: unicellular prokaryotes.
Domain Archaea: prokaryotes often in extreme environments; no known pathogens in the classic sense.
Domain Eukarya: includes all eukaryotic organisms.
The hierarchical sequence (least specific to most specific): Domain → Kingdom → Phylum → Class → Order → Family → Genus → Species.
Example relationships (from the diagram):
Domain Bacteria → Phylum Firmicutes → Class Clostridia → Order Clostridiales → Family Clostridiaceae → Genus Clostridium → Species Clostridium tetani (causative agent of tetanus).
Notes:
“Life” sits at the broadest level, while species is among the most specific groupings.
UNICELLULAR PROKARYOTES and UNICELLULAR & MULTICELLULAR EUKARYOTES are high-level generalizations used to differentiate organisms.
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Taxonomic Hierarchy: Six-Kingdom Classification System
The six-kingdom system typically includes:
Archaea
Bacteria
Fungi
Plantae
Animalia
Protists* (not a true kingdom; a catchall category for lifeforms formerly grouped in Kingdom Protista)
Visual cues in the slide show (examples):
Sulfolobus (Archaea) — irregular, doughnut-shaped organelle; an example micrograph.
Staphylococcus aureus (Bacteria) — densely clustered, spherical cells.
Candida albicans (Fungi) — multiple ovoid bodies connected in a chain.
A flowering plant (Plantae) — blue flowers, two petal layers.
A tree frog (Animalia) — lemon-green with brown speckles.
Paramecium (Protista) — single cell with cilia; bright-field appearance.
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Taxonomic Hierarchy: Species
Eukaryotic species:
A group of similar organisms that can sexually reproduce with each other.
Prokaryotic species:
Cells that share physical characteristics and have at least 70% DNA similarity.
At least 97% identical 16S rRNA sequence similarity.
Notes:
The 70% DNA similarity threshold is a practical criterion for prokaryotic species delineation.
The 97% 16S rRNA similarity is a molecular genetic criterion frequently used to define species boundaries among prokaryotes.
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Scientific Names — Binomial Nomenclature
Carl Linnaeus established the binomial nomenclature system:
Two-name system: Genus (capitalized) + species (lowercase).
Scientific names are italicized (or underlined if handwritten).
Names can be abbreviated after first use (e.g., Escherichia coli → E. coli).
Examples and explanations:
Escherichia coli → E. coli: honors the discoverer, Theodor Escherich.
Staphylococcus aureus → S. aureus: describes clustered spherical cells and the habitat/appearance (aureus = gold-colored colonies).
Interpretations:
The genus name gives a broader grouping; the species name specifies a particular organism within that genus.
The binomial name provides essential information about relationships and characteristics.
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Taxonomic Hierarchy: Strain
Strain: a genetic variant within the same species.
Mutations and horizontal gene transfer can lead to new strains.
Strain names typically include numbers and/or letters after the species name (e.g., E. coli K-12: a laboratory strain of Escherichia coli).
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Microbes May Be Friends or Foes
Microbes constitute a massive portion of Earth's biomass.
It’s estimated there are several million microbial species in the world; more specifically:
Over 7{,}000 microbes have been characterized.
General takeaway:
Most microbes are helpful or neutral to human health; only a small minority are human pathogens.
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Host–Microbe Interactions — Symbiosis in Microbes
A symbiotic relationship exists when two or more organisms are closely connected.
Types of relationships:
Commensalism: no perceived benefit or cost to the host.
Mutualism: benefits the host.
Parasitism: harms the host.
These relationships underpin the concept of normal microbiota (see next page).
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Normal Microbiota — Roles and Scope
Normal microbiota includes bacteria, archaea, and eukaryotic microbes found on or in our bodies.
Functions:
Train our immune system.
Produce vitamins for us.
Help digest foods.
Potentially influence mood and brain function.
Note: Our normal microbiota can include opportunistic pathogens; they may cause disease under certain conditions.
Statistic example:
Approximately 27\% of adults asymptomatically carry Staphylococcus aureus on their skin.
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Normal Microbiota and the Human Microbiome — Site-Specific Populations
Skin:
Populations vary by skin region; at least 1{,}000 species identified.
Examples: Candida albicans, Corynebacterium species, Pityrosporum ovale, Staphylococcus species, Streptococcus species, Trichosporon cutaneum.
Stomach:
Transient populations mainly from swallowed materials; up to 25 species, including Bacteroides species, Helicobacter pylori, Lactobacillus species, Streptococcus species.
Intestines:
Over 40{,}000 species, including Bacteroides species, Clostridioides difficile, Escherichia coli, Lactobacillus species.
Mouth, pharynx, and upper respiratory system:
At least 6{,}000 species, including Candida albicans, Neisseria sicca, Staphylococcus species, Streptococcus species.
Urogenital tract:
About 60 species, including Corynebacterium species, Lactobacillus species (in vagina), Streptococcus species, Ureaplasma species.
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Disruptions in Normal Microbiota
Disruptions to normal microbiota increase risk of infections.
Antibiotic therapy can disrupt normal microbiota:
Kills resident bacteria and can reduce competition against pathogens.
Reduction of normal microbiota allows opportunistic pathogens to establish infections.
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Biofilms — Formation and Healthcare Implications
Biofilm lifecycle:
Attachment: Cells attach to a surface and begin to replicate.
Growth: A sticky matrix produced by biofilm residents promotes adhesion and protects the community; hard to penetrate.
Detachment: Planktonic (free-floating) cells can be released to seed new sites.
Visual note: SEM image example of biofilm on dental floss/tooth surfaces.
Target surface: Biofilms can form on nearly any surface.
Key statistics:
60\%–80\% of infectious diseases in humans are due to biofilm-forming microbes.
Clinical significance:
Internal biofilms are more resistant to antibiotics and immune system defenses than planktonic cells.
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1.3 Learning Objectives — Growing, Staining, and Viewing Microbes
After this section, you should be able to:
Discuss the main formats of culture media used in the laboratory.
Describe the goal of aseptic culture technique and identify central elements.
Explain the goal of the streak plate technique and why it is important in microbiology.
Summarize simple versus structural staining techniques and what information they provide.
Describe the Gram stain procedure and why it works.
Correctly label the parts of the compound light microscope.
Define the term resolution.
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We Culture Microbes So We Can Study Them
The first step in studying a microbe is to grow it in the laboratory.
Growth is not trivial; many known species require complex growth environments.
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Growth Media
Growth media (culture media) are mixtures of nutrients that support microbial growth.
Types of media:
Broths (Liquid)
Plates (Solid and contains Agar)
Slants (Solid and contains Agar)
Deeps (Solid and contains Agar)
Agar is often added as a solidifying agent to allow isolation of colonies.
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Aseptic Culture Techniques
Nature often yields mixed cultures rather than single-species cultures.
Pure culture: a culture consisting of a single microbial species isolated from a diverse sample.
Aseptic culture techniques are practices that limit contamination:
Use sterile media and sterile instruments.
Decontaminate work surfaces.
Wear gloves and other protective clothing.
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Aseptic Culture Techniques — Streak Plate Method
Streak plate technique is used to isolate colonies of a single microbe for study.
Concept: Dilute sample on solid medium to obtain discrete colonies.
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Aseptic Culture Techniques — Colony and Cultures
Colony: a grouping of cells (clones) that developed from a single parent cell.
Mixed culture: contains more than one type of organism, resulting in multiple colony morphologies.
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Staining Specimens — Basic Dyes and Positive Staining
Stains (dyes) increase contrast to visualize cells under the microscope.
Basic dyes:
Dyes are positively charged and are attracted to negatively charged cell surfaces.
Result: cells take on the color of the dye.
Common basic dyes:
Methylene blue
Crystal violet
Safranin
Malachite green
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Staining Specimens — Acidic Dyes and Negative Staining
Acidic dyes are used in negative staining:
Dyes are negatively charged and are repelled by the negatively charged cell surface.
Result: stains the background rather than the cell.
Common acidic dyes:
Nigrosin
India ink
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Mordants
Mordants are chemicals that interact with a dye to fix or trap it on or inside a specimen.
Example: Iodine is a mordant used in certain staining procedures to intensify or fix the stain.
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Simple Stains
Simple staining techniques use a single dye.
Purpose: determine cell size, shape, and/or cellular arrangement.
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Structural Stains — Capsule Staining
Capsule staining aims to visualize capsules: sticky carbohydrate-based outer layers produced by some bacteria.
Method involves using both a basic dye to stain the cell and an acidic dye to stain the background.
Capsule appears as a clear halo around the cell.
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Structural Stains — Endospore Staining
Endospores are dormant structures formed by some bacteria under harsh conditions.
Staining approach:
Apply heat to drive the dye (malachite green) into the spores.
Counterstain non-sporulating cells with safranin.
Endospores are visible as green structures within red/pink cells (depending on the counterstain).
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Differential Stains — Gram and Acid-Fast Stains
Differential staining highlights differences in bacterial cell walls to discriminate classes of cells.
Examples:
Gram stain
Acid-fast stain
These stains help categorize bacteria by cell wall structure and composition.
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Gram Stain — Classification by Cell Wall Structure
Bacteria are classified as Gram-positive or Gram-negative based on cell wall characteristics:
Gram-positive: thick peptidoglycan layer.
Gram-negative: thin peptidoglycan layer plus an outer lipid-containing layer (lipopolysaccharide).
Shapes:
Rod (gram-negative)
Coccus (gram-positive)
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Gram Stain Procedure — Stepwise
Steps and outcomes:
Apply crystal violet (purple dye).
Apply iodine (mordant).
Decolorize with alcohol.
Apply safranin (counterstain).
Result interpretation:
Gram-positive: retain crystal violet → appear purple.
Gram-negative: lose crystal violet and take up safranin → appear pink/red.
Key reagents in order: Crystal violet → Iodine → Alcohol → Safranin.
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Acid-Fast Staining
Purpose: distinguish cells with and without waxy cell walls.
Acid-fast bacteria have waxy walls rich in mycolic acid and retain the red primary dye, Carbol-fuchsin, after acid-alcohol wash.
Non–acid-fast cells lose the primary dye after acid wash and are counterstained (commonly with methylene blue).
Important diagnostic targets:
Mycobacterium species (e.g., M. tuberculosis)
Nocardia species
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Light Microscopy
Light microscopy uses visible light to illuminate specimens.
The compound light microscope is the most common type of optical microscope used in microbiology.
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Parts of the Compound Light Microscope
Objective lens: located near the specimen; multiple objectives with different magnifications.
Ocular lens: located at the top near the viewer's eyes.
Final magnification is the product of the magnifications of the ocular and the objective lenses.
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Additional Microscope Components
Condenser lenses: focus light into a precise cone to illuminate the specimen.
Iris diaphragm: controls the amount of light reaching the specimen to improve contrast.
Coarse focus knob: roughly focuses the image by moving the stage or objective lens to adjust distance.
Fine focus knob: precise focusing for sharp image.
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Resolution — The Ability to Distinguish Detail
Resolution definition: the ability to distinguish two distinct points as separate.
Naked eye resolution: about 0.1\ \text{mm} = 100{,}000\ \text{nm}.
Most compound light microscopes have a resolution around 200\ \text{nm}.
Note: The transcript contains a missing value for maximum magnification; the important functional point is the resolution limit, not the exact maximum magnification.
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1.2 Learning Objectives (Revisited) — Classifying Microbes
Reiteration of the objectives covered in this section:
Summarize the taxonomic hierarchy of domains and kingdoms.
Describe binomial nomenclature and the information it conveys about organisms.
Define species and strain for prokaryotes.
Compare parasitism, mutualism, and commensalism.
Define normal microbiota and discuss its roles.
Describe how a biofilm forms and its healthcare implications.
Provide examples of microbial impacts on industry and the environment.
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1.3 Learning Objectives — Growing, Staining, and Viewing Microbes
After this section, you should be able to:
Discuss the main formats of culture media used in the laboratory.
Describe the goal of aseptic culture technique and the central elements involved.
Explain the goal of the streak plate technique and why it is important in microbiology.
Summarize simple versus structural staining techniques and what information they provide about a sample.
Describe the Gram stain procedure and why it works.
Correctly label the parts of the compound light microscope.
Define the term resolution.
Key cross-topic connections and takeaways:
Taxonomy provides a framework for organizing microbial diversity and understanding evolutionary relationships. The three-domain system consolidates bacteria, archaea, and eukaryotes as fundamental groups.
Binomial nomenclature communicates universally about organisms and encodes information about genus, species, and sometimes discoverers or notable features (e.g., E. coli, S. aureus).
Species and strain concepts are practical ways to categorize organisms, especially prokaryotes, for communication and research, with specific similarity thresholds guiding definitions (DNA similarity, 16S rRNA similarity).
Normal microbiota play essential roles in health but can become opportunistic pathogens when ecosystems are disrupted (e.g., by antibiotics).
Biofilms represent a major clinical challenge due to their resistance to antibiotics and immune clearance; they account for a substantial share of infectious diseases.
Laboratory techniques (culture media, aseptic technique, streak plating, staining methods, and microscopy) are fundamental tools for identifying, characterizing, and understanding microbes.
Knowledge of staining (simple, structural, differential) and staining mechanisms (capsules, endospores, Gram, acid-fast) provides critical practical means to classify and study microbes.