Microbiology: Microbes, Prokaryotes, & Eukaryotes

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Humans and the Microbial World

• Spontaneous generation is the hypothesis that life arises from nonliving matter, with a “vital force” necessary for life, but was disproven after over 200 years of experiments.
• The microbiome refers to a group of microbes that live stably on/in the human body.
• Microbes¹ are classified into Bacteria (prokaryote), Archaea (prokaryote), and Eukarya (eukaryote).

Prokaryotes vs. Eukaryotes

• Prokaryotes are single-celled organisms without a membrane-bound nucleus, instead having a nucleoid or organelles.
• Eukaryotes can be single-celled or multicellular and have a membrane-bound nucleus and organelles like mitochondria, endoplasmic reticulum, and the Golgi apparatus.
• Both prokaryotes and eukaryotes contain a plasma membrane, cytoplasm, and ribosomes.

Types of Living Microorganisms

• Bacteria are single-celled prokaryotes with specific shapes (cylindrical, spherical, spiral) and cell walls consisting of peptidoglycan.
• Archaea are single-celled prokaryotes, similar in size and shape to bacteria.
• Many archaea can live in harsh environments and do not have peptidoglycan in their cell walls.
• Fungi can be single-celled yeasts or multicellular molds/mushrooms that spread via spores and absorb nutrients from organic materials for energy.
• Algae can be single-celled or multicellular organisms found in freshwater, saltwater, and moist soil, using photosynthesis to generate energy.
• Protozoa are single-celled eukaryotes that can be motile and ingest organic material.
• Helminths include roundworms, tapeworms, and flukes, with microscopic eggs and larvae, but adults can be seen with the naked eye.

Types of Acellular Microorganisms

• Viruses are obligate intracellular parasites using the machinery and nutrients of host cells to replicate, consisting of either DNA or RNA surrounded by a protein coat.
• Viroids are obligate intracellular parasites that consist of only RNA and have no protein coat.
• Prions are misfolded versions of normal cellular proteins that cause the normal versions to misfold, consisting of only protein and have no DNA or RNA.

Scientific Names

• Scientific names consist of a binomial system of nomenclature reflecting a characteristic of the organism or honoring a scientist.
• Each scientific name is made up of a genus and a specific epithet, the genus should be capitalized, while the species shouldn’t be capitalized.
• Both the genus and species should be italicized or underlined.

Cells and Methods to Observe Them

• Prokaryotes have no nucleus (only a nucleoid region), circular DNA, no organelles (but can have granules or vesicles), peptidoglycan cell walls, flagella, axial filaments, pili, 70s ribosomes and reproduces via binary fission.
• Eukaryotes have a membrane-bound nucleus, linear DNA wrapped around histones, a multitude of organelles, cellulose or chitin cell walls, flagella and cilia, 80s ribosomes and reproduce via mitosis or meiosis.
• Pleomorphism refers to variations in cell structure caused by slight genetic or nutritional differences, resulting in an undefined shape.

Shapes Observed in Bacteria

• Coccus: spherical
• Cocobacillis: short but slightly elongated
• Bacillus: rod-like
• Spiral: Includes vibrio (comma-shaped bacterium), spirillum (a rigid, wavy-shaped bacterium), and spirochete (a corkscrew-shaped bacterium).

Arrangements Observed in Bacteria

• Diplo-: division in one plane produces cells in pairs.
• Strepto-: division in one plane produces cells in chains.
• Tetra-: division in two planes produces cells in tetrads-four cells arranged in a cube.
• Sarcinae: division in three planes-eight cells arranged in a cube.
• Staphylo-: random division producing grapelike clusters.
• Palisades: chains of cells attached by a small hinge region at the ends.
• Rosette: bacteria attached together via stalks.

Selectively Permeable Plasma Membrane

• The plasma membrane is a selectively permeable lipid bilayer with proteins, defining the boundaries of a cell.
• Passive transport involves substances moving from a higher to lower concentration, requiring no energy.
• Simple Diffusion: Nonpolar substances and small polar substances (water and small ions), pass through pores.
• Facilitated Diffusion: Large molecules and ions require transport proteins to aid in their entry into the cell.
• Osmosis: Movement of water from low to high solute concentrations, or from high to low water concentrations.
• Active Transport: Requires ATP to transport substances against a concentration gradient

Peptidoglycan Layer

• Peptidoglycan is a polymer consisting of sugars and amino acids, forming a mesh-like layer outside the plasma membrane of bacteria and serves as the most important component of the cell wall
• Gram-positive bacteria have a thicker peptidoglycan layer (60-90%), lack an outer membrane, and contain teichoic acid and lipoteichoic acid, making them highly susceptible to penicillin.
• Gram-negative bacteria have a thinner peptidoglycan layer (10-20%), have an outer membrane containing lipopolysaccharides (LPS), contain periplasmic spaces, and are less sensitive to penicillin.

Cell Wall

• The cell wall maintains the characteristic shape of the cell and protects it from bursting due to osmosis.
• Cell walls are found in nearly all prokaryotes EXCEPT the bacteria Mycoplasma, which instead have sterols in the plasma membrane.
• Atypical cell walls are described in acid-fast bacteria mycobacterium and nocardia. Due to lipids impeding entry of nutrients into the bacterium, growing slowly, and expending large amounts of energy to synthesize lipids.

Cell Walls

• Cell walls are thick, composed of 60% waxy lipid, also known as mycolic acid.
• Cell walls contain less peptidoglycan than gram (+) bacteria
• Penicillin interferes with peptidoglycan synthesis.
• Lysozyme breaks bonds linking glycan chains found in human excretions and destroys the structural integrity of peptidoglycan molecules

Bacterial Flagella

• A flagellum, singular, is made of protein subunits called flagellin that include a filament, hook, and basal body.
• Atrichous: bacteria without flagella.
• Monotrichous: single flagellum at one end.
• Amphitrichous: single flagellum at both ends.
• Lophotrichous: tuft of flagella at one or both ends.
• Peritrichous: flagella distributed all over the bacterium.
• Rotate in a twirling like manner using their L-shaped hooks, including Runs and Tumbles. Move due to chemotaxis.

Axial Filaments on Bacteria

• Periplasmic flagella that wrap around the bacterium, causing it to take on a helical/spirochete shape and is found between the cell wall and cytoplasmic membrane of bacteria.

Bacterial Pili

• Conjugation Pili (sex pili) attach two cells to transfer DNA via conjugation. Furnish genetic variety for bacteria and can pass antibiotic resistance.
• Fimbriae (attachment pili) help bacteria adhere to cell surfaces and enhance colonization on the surfaces of other organisms.

Glycocalyx

• A gel-like layer outside the cell wall that protects or allows for attachment to a surface and is made of repeating polysaccharide or glycoprotein units.
• Capsules: are thick, neatly organized, and firmly attached to the cell, formed by pathogenic bacteria and have greater disease-causing capabilities.
• Slime Layer: is thin, unorganized, loosely attached to the cell and protect against loss of water and nutrients.

Using Microscopes to Observe Bacteria

• Used to magnify small objects of the Brightfield objects visible against a bright background
• Darkfield: light objects visible against a bright background.
• Phase-Contrast: makes cells appear darker.
• Differential Interference Contrast: forms a 3D appearance.
• Fluorescence: observe cells/materials that are naturally fluorescent or tagged with fluorescent dyes.
• Electron: include transmission (TEM) and scanning (SEM) and can magnify objects 100,000X.
• Image is black and white but can be artificially colored.
• Lenses and specimens must be used in a vacuum.

Staining Bacteria

• Staining is a molecule that binds to a cellular structure to give it color. Used to make microorganisms stand out against their backgrounds, to group major categories of microorganisms, and to examine structural/chemical differences in cellular structures.
• Cationic (basic) Dyes are attracted to negatively charged cell elements, while Anionic (acidic) Dyes: attracted to positively charged cell elements.
• Simple stains are single dyes, that reveals basic cell shapes and arrangements, with Negative Stain allows visualization of organisms with structures that WILL NOT accept most stains and reveals reveal cell morphology and size
• Differential stains uses two or more dyes to distinguishes between two kinds of organisms OR between two different parts of an organism
• Gram Stain distinguishes Gram (+), Gram (-), is gram variable, and gram-nonreactive organisms. Is a cell wall retains crystal violet stain on Gram Positive, cell wall does NOT retain crystal violet stain on Gram Negative, cell wall does not stain/stain properly, with cell wall stains unevenly on Gram Variable
• Acid-Fast Stain distinguishes members of Mycobacterium and Nocardia from other bacteria Special stains are used to identify various specialized structures
• Flagellar Stain Indicates the presence of flagella by building up layers of stain on their surface Spore Stain allows visualization of hard-to-stain bacterial endospores-I.e. members of genera Clostridium and Bacillus

Eukaryotic vs Prokaryotic Membranes

• Prokaryotic membranes are simple ones, but eukaryotic membranes are complex ones,.
• BOTH: Possess a phospholipid bilayer with proteins ONLY EUKARYOTES: Contain sterols—to increase stability—and lipid rafts-to detect/respond to signals

Mobility in Eukaryotic and Prokaryotic Cells

• Prokaryotic Flagella move via rotation clockwise or counterclockwise and are relatively simple structures naked, and have no sheath covering the structure.
• Eukaryotic Flagella move in a whip-like fashion and are 10x thicker; made of microtubules placed in a 9+2 arrangement and is covered by an extension of the plasma membrane.

Eukaryotic Organisms

• Nucleus: holds the cell's DNA; surrounded by a nuclear envelope.
• Nucleolus: site of ribosomal RNA synthesis.
• Ribosomes: involved in protein synthesis.
• Cytoskeleton: framework of the cell; responsible for cell shape.
• Rough ER: synthesizes and modifies proteins.
• Smooth ER: synthesizes and stores nonprotein macromolecules such as lipids.
• Golgi Body: modifies, packages, and transports proteins to & out of the cell.
• Lysosomes: contain enzymes that digest food particles, debris, and foreign microorganisms.
• Mitochondria: generate ATP for the cell.
• Chloroplast: site of photosynthesis for plants and algae.
• The endosymbiotic theory is when larger bacterial cells engulfed smaller ones, leading to the development of the first eukaryotes.
• Ingested aerobic bacteria became mitochondria, and ingested photosynthetic bacteria became chloroplasts.
• Both Mitochondria and chloroplasts have 70S ribosomes and their own DNA separate from the nucleus, and it is circular like prokaryotic DNA and both replicate without input from the nucleus in a process similar to binary fission

Fungi

• Members of a group of eukaryotic organisms that include molds, yeasts, and mushrooms morphological forms Mold: MULTIcellular filamentous fungi. Yeast: SINGLE-celled fungi. Mushroom: reproductive structure of certain fungi Can obtain nutrients from dead or decaying organic material, living tissue, and even petroleum products,often have mutualistic relationships with other organisms ie: Fungi and algae can form lichens EX: Fungi and plant roots can form mycorrhizae Fungal cell walls contain chitin.

Protozoa

• Microscopic, single-celled eukaryotic organisms, that lack a cell wall, but DO have structures for movement ie, flagella or cilia, found in aquatic environments or in the soil
• One type includes intestinal protozoans which can exist as a trophozoite growing form and as a cyst infectious form Viruses
• Viruses are infectious agents with genetic information contained within a protective protein coat: Require live organisms as hosts: obligate intracellular parasites and can be enveloped surrounded by a lipid bilayer or naked, their families end with viridae and they can describe viruses that infect bacteria

Lytic Cycle

• Lytic Life involves 5 steps: attachment, genome entry, synthesis, assembly, and release Attachment: phage attaches to receptors Genome Entry: phage lysosomes degrade cell wall Synthesis: degrades host DNA, produces phage genome and its viral components instead Assembly: components are assembled within the bacterial cell Release: the cell lyses, releasing the infective viruses; those new viruses then enter and infect new hosts

Lysogenic life

• Lysogenic Life is characteristic of temperate phages where the Dna enters the hosrt
• It incorporates itself into the bacterial chromosome as a prophage and can remain in a latent state indefinitely and then replicates along with the chromosome through many cell divisions

Bacterial Transduction

• A method of transferring genetic material from one bacterium to another (horizontal gene transfer) using a bacteriophage
• Generalized: any bacterial gene can be transferred with random host bacterial DNA that can be mistakenly packaged into the head of a virus When host lyses, viruses can transfer any random bacterial gene to a recipient cell

Special Transduction

• Only specific genes are transferred when viral DNA is integrated into the host bacterial chromosome at a specific location.
• When excised, it can transfer specific bacterial genes located near the phage integration site to a recipient cell.
• Acute infections have a rapid onset, short duration—influenza.
• Bursts of viral particles are released from an infected host cell as the immune system gradually eliminates viral particles.

Persistent Infections

• Persistent Infections continue for years or throughout a lifetime with or without any apparent symptoms and can include chronic or latent infections Chronic entails constant production of low levels of viral particles. Latent: viral genome remains silent in host cell then can reactivate to cause an infection, while the Viral genome can be incorporated into a host cell chromosome as a provirus.