JeffState Microbiology Ch #3

What is the Microsphere Nanoscope?

A specialized microscopic technique utilizing microspheres to achieve super-resolution imaging beyond the diffraction limit of light.

How does the Microsphere Nanoscope work?

It places a transparent microsphere on or near the specimen to collect near-field evanescent waves and converts them into far-field virtual images.

What distinguishes the Microsphere Nanoscope from a regular light microscope?

It breaks the diffraction limit of light without needing complex fluorophore tagging or ultra-high-power laser setups.

What can be observed using the Microsphere Nanoscope?

Sub-diffraction cellular structures, viruses, fine cell wall details, live cell organelles, and molecular assemblies in real time.

What are the Five I's in microbiology?

Inoculation, Incubation, Isolation, Inspection, Identification.

What does Inoculation entail?

Introduction of microbes into or upon media for culture.

What is Incubation in microbiology?

Creating the proper growth conditions such as temperature and gas requirements.

What is Isolation in microbiology?

Separating individual species to obtain discrete, pure isolated colonies.

What does Inspection involve?

Observing cultures macroscopically and microscopically.

What is Identification in microbiology?

Determining the identity of the isolated microbe to the species level.

What is a culture in microbiology?

Propagation of microorganisms with various media; growth of microbes in or on a nutrient medium.

What is a medium in microbiology?

A nutrient-containing environment in which microbes can multiply.

What does sterile mean in microbiology?

Completely free of all life forms including endospores and viruses.

What are the standard laboratory temperatures for incubation?

Typically range from 20 °C to 45 °C.

What is the growth indication in liquid media?

Cloudiness (turbidity), sediment formation, scum/pellicle, or color changes.

What indicates growth on solid media?

Formation of colonies, which are visible masses of piled-up cells.

What is the optimum growth temperature for Listeria monocytogenes?

25 to 30 °C.

How long does it take to culture Pseudomonas fluorescens?

1-2 days.

What is the optimum growth temperature for Streptococcus pyogenes?

37 °C.

How long does it take to culture Mycobacterium tuberculosis?

28 days (4 weeks).

What are the three fundamental properties of media classification?

Physical State, Chemical Composition, Functional Type.

What are the physical states of media?

Liquid, Semisolid, Solid.

What is Liquid Media?

Water-based solutions that flow freely and do not solidify above freezing.

What is Semisolid Media used for?

Testing bacterial motility and localizing reactions.

What is Solid Media used for?

Firm surfaces for discrete colony formation; essential for isolating bacteria and fungi.

What is Agar?

A complex polysaccharide extracted from red alga that liquefies at 100 °C and solidifies at 42 °C.

What characterizes Defined (Synthetic) Media?

Media with exact known chemical structure, formulated with pure organic/inorganic compounds.

What is Complex Media?

Media that contains at least one ingredient that is not chemically definable.

What is General-Purpose Media?

Designed to grow a wide variety of microbes.

What is Enriched Media?

Contains complex organic substances required by fastidious organisms.

What is Selective Media?

Contains agents that inhibit certain microbes while encouraging growth of desired species.

What is Differential Media?

Allows multiple species to grow but displays visible differences based on metabolic reactions.

What does Blood Agar differentiate?

Species via hemolysins (Beta = complete lysis, Alpha = partial lysis, Gamma = no lysis).

What is Reducing Media used for?

Contains chemicals that absorb oxygen; used for obligate anaerobes.

What does carbohydrate fermentation media contain?

It contains sugars and a pH indicator to detect acid production and gas formation.

What is the purpose of transport media?

It maintains and preserves clinical specimens during transit.

What is assay media used for?

It is used to test antimicrobial drugs, antiseptics, and cosmetics.

What is enumeration media used for?

It is used in industrial and environmental testing to count microbial numbers in food, water, or soil.

What is a colony in microbiology?

A discrete mound of cells formed on solid agar originating from a single cell.

What is the streak plate method?

A technique where a sample is spread in a thinning pattern across agar using a loop, flame-sterilizing between quadrants.

What is the pour plate method?

A method where serial dilutions are mixed into melted agar and poured into Petri dishes; colonies grow on the surface and within the agar.

What is the spread plate method?

A technique where a liquid sample is pipetted onto agar and spread evenly with a sterile glass 'hockey stick'.

What is a pure culture?

A culture that contains only a single known species.

What is a subculture?

A second-level culture made from an isolated colony.

What is a mixed culture?

A culture that holds two or more identified, easily differentiated species.

What is a contaminated culture?

A culture that was once pure or mixed but now contains unwanted contaminants or microbes.

What is the range of sizes for viruses?

Viruses range from 20 to 800 nm.

What is the size range for bacteria?

Bacteria range from 200 nm (Mycoplasma) to 750 µm (Thiomargarita).

What is the size of yeasts?

Yeasts are typically 3 to 4 µm.

What is the size range for protozoa?

Protozoa range from 100 to 300 µm.

What is magnification in microscopy?

Magnification results from the interaction of visible light waves and lens curvature.

What is refraction in optics?

Refraction is the bending or change in light ray angle as it passes from one medium to another.

What is a real image in microscopy?

An image formed by the objective lens.

What is a virtual image in microscopy?

An image formed by the ocular lens (eyepiece) as perceived by the eye.

How do you calculate total magnification?

Total magnification is calculated as Ocular Lens Power × Objective Lens Power.

What are the three elements of effective microscopy?

1. Magnification, 2. Resolution (Resolving Power), 3. Contrast.

What is the oil immersion technique?

It is applied only with the 100x objective lens to prevent light refraction/scattering into air, increasing resolution.

What types of light microscopes are there?

Standard bright-field, dark-field, phase-contrast, differential interference contrast (DIC).

What is a fluorescence microscope used for?

It uses a UV light source and fluorescent dyes.

What is a confocal microscope?

A microscope that uses focused laser beams to scan depth sections of specimens.

What is the purpose of a super-resolution microscope?

It uses dual lasers to bypass light diffraction limits.

What is the function of an electron microscope?

It uses electron beams instead of light for ultra-high magnification and nanometer resolution.

What is a wet mount?

A drop of culture on a slide with a coverslip, fast but dries quickly.

What is a hanging drop mount?

A technique using a concave slide and Vaseline seal to preserve viability and motility without crushing cells.

What is a fixed smear?

An air-dried film gently heat-fixed to kill cells and preserve structure.

What is a basic dye?

A positively charged cationic dye that binds to negative cell walls.

What is an acidic dye?

A negatively charged anionic dye that is repelled by the cell wall, staining the background.

What is a simple stain?

A stain using a single dye to reveal shape, size, and arrangement.

What is a differential stain?

A stain that uses a primary stain and counterstain to distinguish cell types.

What is the purpose of the Gram stain?

To differentiate bacteria into Gram-positive (purple) and Gram-negative (pink) categories based on cell wall structure.

What is the acid-fast stain used for?

To differentiate acid-fast (pink/red) from non-acid-fast (blue) cells.

What does the endospore stain distinguish?

It distinguishes dormant endospores (green) from vegetative cells (pink/red).

What is a special stain?

A stain that highlights specific structures like capsules or flagella.

What happens when X-Gal is present in a media?

Lactose fermenting colonies turn blue while non-fermenting colonies remain white.

What is the definition of culture in microbiology?

(a) Propagation of microorganisms with various media; (b) The growth of microorganisms in or on a nutrient medium.

Why do we use immersion oil with 100X objective lenses?

To reduce light scatter by filling the air gap with a medium having the same refractive index as glass.

What type of microscopes are commonly used in laboratories?

Compound light microscopes.

What is the total magnification of a specimen viewed with the 40× objective?

400x (40x objective × 10x ocular).