Biol 3333 Module 2 Chp 2 and 3

Standard measurement for microorganisms

Measured in micrometers (μm) and nanometers (nm)

stain used for capsules

india ink or nigrosin is commonly used to visualize the presence of capsules around bacteria.

what are eukaryotic cell walls made up of

polysaccharides

2 types of glycocalyx

capsules and slime layer

function of glycocalyx

protects against dehydration and nutrient loss as well as prevents phagocytosis

Magnification

An increase in the apparent size of an image to resolve smaller separations between objects.

Resolution

The smallest distance by which two objects can be separated and still be distinguished.

Limit of microscopy

Resolution, not magnification, limits the ability of what we can see

Refractive Index

A measure of the light-bending ability of a medium

Purpose of Immersion Oil

Used with 100X objectives to keep light from refracting/scattering, which maintains high resolution.

Brightfield Microscopy

Most widely used; specimen is darker than the surrounding bright field; used for live and preserved stained specimens

Darkfield Microscopy

Brightly illuminated specimens surrounded by a dark field; used for live and unstained specimens

Phase-contrast Microscopy

Transforms subtle changes in light waves into differences in intensity; best for observing intracellular structures

Differential Interference Contrast (DIC)

Uses two light beams and prisms to provide higher contrast, color, and a 3-D appearance to live specimens.

Fluorescence Microscopy

Uses UV radiation to excite specimens stained with fluorochromes, which then emit visible light; useful for diagnosing infections

Confocal Microscopy

Uses a laser to scan multiple "z-planes" to construct a 3-D image; useful for thick specimens like biofilms.

Electron waves vs. Light waves

Electron waves are 100,000 times shorter than visible light, providing much greater resolution

Transmission Electron Microscope (TEM)

Transmits electrons through a specimen to reveal detailed internal structures; magnification up to 1,000,000X.

Scanning Electron Microscope (SEM)

Bombards the surface of a metal-coated specimen with electrons to provide a detailed 3-D view of external structures

Purpose of Wet Mounts and Hanging Drop

Used to examine live cells to observe size, motility, shape, and arrangement.

Fixed Mounts (Smears)

Created by drying and heating a film of specimen; kills the microbes and attaches them to the slide.

Heat Fixation

Passing a slide through a Bunsen burner flame to attach the specimen and preserve it.

Chemical Fixation

Using fixatives like 10% formalin or methanol to preserve tissue structure.

Basic Dyes

Cationic, positively charged dyes attracted to negatively charged microbial surfaces.

Acidic Dyes

Anionic, negatively charged dyes repelled by microbes, resulting in a stained background

Positive Staining

The surface of the microbe is colored by the dye while the background remains white/clear.

Negative Staining

The microbe remains clear while the background is stained dark; useful for viewing capsules.

Simple Stain

Uses a single dye to reveal cell shape, size, and arrangement.

Differential Stain

Uses a primary stain and a counterstain to distinguish between cell types or parts.

Gram-Positive Bacteria

Retain crystal violet due to a thicker cell wall and appear purple

Gram-Negative Bacteria

Lose crystal violet during alcohol wash; take up safranin counterstain and appear red/pink.

Gram Stain Steps

1. Crystal violet (Primary), 2. Iodine (Mordant), 3. Alcohol (Decolorizer), 4. Safranin (Counterstain) .

Acid-Fast Stain use and what species does it identify

binds to waxy mycolic acids in cell walls; used to identify Mycobacterium and Nocardia

Endospore Stain

Uses malachite green (primary) and safranin (counterstain); spores appear green within red/pink vegetative cells

Flagellar Stain

Uses a mordant (tannic acid) to increase the diameter of flagella so they can be seen via light microscopy.

Two main categories of cells

Prokaryotes and Eukaryotes.

Distinguishing characteristics of Prokaryotes

DNA is not enclosed in a membrane, lacks membrane-enclosed organelles, and cell walls usually contain peptidoglycan .

Distinguishing characteristics of Eukaryotes

DNA is found in a nucleus (enclosed by a nuclear membrane), contains membrane-enclosed organelles, and cell walls (if present) are chemically simple .

Three main types of Cell shapes:

Coccus (spherical), Bacillus (rod-shaped), and Spiral.

Pleomorphic vs. Monomorphic

Monomorphic bacteria species maintain a single shape; pleomorphic bacteria species can have many shapes.

Glycocalyx definition

A sugar coat (polysaccharide, polypeptide, or both) secreted on the surface of many prokaryotes.

Capsule vs. Slime Layer

A capsule is an organized glycocalyx firmly attached to the cell wall; a slime layer is unorganized and loosely attached.

Functions of the Capsule

Contributes to virulence by protecting bacteria from phagocytosis by host cells.

Flagella function

Long filamentous appendages used for locomotion (motility).

Flagellar arrangements

Atrichous (none), Monotrichous (single), Lophotrichous (tuft at one end), Amphitrichous (flagella at both ends), and Peritrichous (distributed over entire cell) .

Three parts of a flagellum

Filament (outermost region), Hook (attachment point), and Basal Body (anchors to the cell wall and membrane).

Taxis definition

The movement of a bacterium toward or away from a stimulus (e.g., chemotaxis for chemicals, phototaxis for light).

Axial Filaments (Endoflagella)

Bundles of fibrils found in spirochetes that wrap around the cell, enabling a corkscrew motion.

Fimbriae function

Hairlike appendages that allow microbes to adhere to surfaces and each other; involved in forming biofilms .

Pili function

Longer than fimbriae; involved in motility (twitching/gliding) and DNA transfer between cells (conjugation)

Primary functions of the cell wall

Prevents osmotic lysis (rupturing), maintains cell shape, and serves as an anchor for flagella.

Peptidoglycan (Murein)

The macromolecular network making up the bacterial cell wall, consisting of repeating disaccharides (NAG and NAM) and polypeptides.

Gram-Positive Cell Wall

Consists of many layers of peptidoglycan (thick/rigid) and contains teichoic acids.

Gram-Negative Cell Wall

Consists of one or very few layers of peptidoglycan (thin) and an outer membrane; lacks teichoic acids.

Gram-Negative Outer Membrane

Contains lipopolysaccharides (LPS), lipoproteins, and phospholipids; provides a barrier against antibiotics and enzymes.

Lipopolysaccharide (LPS) components

Lipid A (endotoxin), Core polysaccharide (stability), and O polysaccharide (antigen) .

Acid-Fast Cell Wall

Contains high concentrations (60%) of mycolic acid (a hydrophobic waxy lipid) which prevents uptake of most dyes.

Wall-less bacteria example

Mycoplasma species, which are the smallest known bacteria and have sterols in their plasma membranes.

Plasma Membrane structure

A phospholipid bilayer with a "fluid mosaic" of proteins; lacks sterols in most prokaryotes.

Selective Permeability

The property of the membrane that allows certain molecules to pass through while others are stopped.

Passive Processes

Movement of substances from high to low concentration without using energy (Simple diffusion, Facilitated diffusion, Osmosis).

Active Processes

Movement of substances against a concentration gradient using energy (ATP).

Group Translocation

A special form of active transport in prokaryotes where a substance is chemically altered as it crosses the membrane so it cannot leave.

Isotonic vs. Hypotonic vs. Hypertonic

Isotonic (equal), Hypotonic (water enters cell/bursts), Hypertonic (water leaves cell/shrinks)

Cytoplasm

The substance of the cell inside the plasma membrane; 80% water.

Nucleoid

Region containing the bacterial chromosome (a single, continuous, circularly arranged thread of DNA).

Plasmids

Small, circular, double-stranded DNA molecules that are extrachromosomal; often carry genes for antibiotic resistance.

Gas Vesicles

increase buoyancy of bacteria

Prokaryotic Ribosomes

70S ribosomes (consisting of 30S and 50S subunits); the site of protein synthesis.

Inclusions

Reserve deposits where cells accumulate nutrients (e.g., Metachromatic granules, Polysaccharide granules, Lipid inclusions) .

Endospores

Highly resistant, "resting" cells produced by certain Gram-positive bacteria (e.g., Bacillus, Clostridium) to survive extreme environments.

Sporulation vs. Germination

Sporulation is the process of endospore formation; germination is the return to a vegetative state.

Eukaryotic Flagella and Cilia

Used for locomotion; consist of microtubules in a "9 + 2" arrangement.

Eukaryotic Ribosomes

80S ribosomes (60S and 40S subunits); found free in cytoplasm or attached to Rough ER.

Nucleus

Enclosed by a double membrane (nuclear envelope); contains the cell's DNA organized into chromosomes.

Rough Endoplasmic Reticulum (ER)

Studded with ribosomes; factory for synthesizing secretory proteins and membrane molecules.

Smooth Endoplasmic Reticulum (ER)

No ribosomes; site of phospholipid, fat, and steroid synthesis.

Golgi Complex

Modifies, sorts, and packages proteins into vesicles for transport.

Lysosomes

Contain digestive enzymes to break down molecules and harmful bacteria.

Mitochondria

"Powerhouses" of the cell; site of ATP production via cellular respiration; contain 70S ribosomes and their own DNA.

Chloroplasts

Found in algae and green plants; site of photosynthesis; contain 70S ribosomes and DNA.

Endosymbiotic Theory

The theory that eukaryotic organelles (mitochondria and chloroplasts) evolved from symbiotic prokaryotes living inside host cells.

counterclockwise flagellar motion

results in smooth linear direction - run

clockwise flagellar motion

tumble motion