MBIO 1220 Microscopy/Cell Structure (1)

What type of microscope describes any kind of microscope that uses visible light to observe a specimen?

Light microscope

What type of microscope uses 2 lenses to magnify the image?

Compound light microscope

Where is the objective lens?

Closest to the specimen (4x - 100x)

Where is the ocular lens?

The eye piece (10x)

How would you calculate the total magnification for a compound microscope?

Objective lens x ocular lens

What 2 things do we have the ability to do through resolution?

1. Distinguish fine detail and structure

2. Distinguish 2 points a certain distance apart

Two points can be distinguish if there are at least _nm apart

“4”

Do you need light of a shorter or longer wavelength to fit between 2 points, so that they don’t appear as 1 object?

Short-enough wavelength

What is an electron microscope?

Uses electrons instead of light

Do electrons travel in much shorter or longer waves than light?

Shorter waves than light

Is the resolving power greater or weaker with an electron microscope?

Greater

What type of microscope allows greater magnification (some up to 50 million times!)

Electron microscope

What 2 things do electron microscopes allow us to view?

Viruses + internal cell structures

What are the 2 types of electron microscopes?

Transmission (TEM) + scanning (SEM)

What is a transmission EM used to see?

Internal structures

Very ____ slices can be cut from sample using a TEM

“thin”

Why are samples under a TEM generally stained with a metal?

To make structures opaque to electrons

What is a scanning EM used to see?

To see surfaces

Which electron microscope is less powerful than the other?

Scanning EM

What is atomic force used to see?

Molecules

How does atomic force scan a specimen?

Uses thin metal probe, revealing bumps + depressions

Human eye can see an object about __mm

“0.2”

Compound light microscope can view an object about __μm

“2”

Electron microscope can see an object about __nm

“2”

Microorganisms are almost colourless when seen through a microscope. How can we make them easier to see ?

Stains

In what case can we still see microorganisms without a stain?

Wet mount

What are stains composed of?

Positively + negatively charged ions (one of which is coloured) = chromophore

What is a simple stain?

When only 1 dye is used to highlight the entire microorganism

What are the 4 steps to create a simple stain?

1. Smear sample on slide

2. Heat

3. Stain

4. Wash, dry, view

Bacteria have a net _______ charge on their outer surface, attracting stains with ________ charged chromophores

“negative,” "positively”

A positive stain will/will not bind to a bacterium?

Will

What is an example of a positive stain?

Crystal violet

A negative stain will/will not bind to a bacterium?

Will not

What is an example of a negative stain?

Nigrosin

What characterizes differential stains?

React differently with different bacteria (can be distinguished)

An example of a differential stain is a gram stain, which differentiates bacteria based on what?

Structure of the cell wall

Gram positive using the gram stain appears purple, meaning?

Thick cell wall

Does Staphylococcus aureus have a thick or thin cell wall?

Thick

Gram negative using the gram stain appears pink, meaning?

Thin cell wall

Does Escherichia coli have a thick or thin cell wall?

Thin

Bacteria with a thin cell wall lose crystal violet during staining, why does it turn pink?

Takes on colour of counterstain (safranin)

What type of stain stains the internal structure of some bacteria?

Endospore stain

Primary stain of the endospore stain colours what structure malachite green?

Endospores

The counterstain of the endospore stain is _______, which colours the rest of the cell ____

“safranin,” “red”

What is an example of a bacterium that has endospores?

Bacillus anthacis

What type of stain stains the external structure of some bacteria?

Flagella stain

What does mordant do in order to observe something under light microscope?

Thickens the flagella (= external structure helping specimen to move e.g. sperm flagella)

What’s an example of a bacterium that has peritrichous flagella?

(peritrichous = all around its surface)

Bordetella bronchisepticum

What does an acid-fast stain detect the presence of?

Waxy compound in cell wall

What type of stain is used to identify the genus Mycobacterium?

Acid-fast stain

What dye does a Mycobacterium cell wall retain?

Carbol fuchsin (darker)

Counterstain with methylene blue stains non _______ bacteria + ________ blue

“acid-fast,” “tissues”

What does a capsule stain detect a thick layer of?

Polysaccharide outside the cell (hence capsule)

In a capsule stain, a negative stain colours the?

Background

In a capsule stain, a positive stain colours the?

Cell

The capsule does not take up most dyes, remaining?

Colourless

What’s an example of a bacterium we would use a capsule stain on?

Streptococcus pneumoniae

Breaking down Pro-karyote, what does either word mean?

Pro = before

Karyon = nucleus

In a prokaryote, where is the DNA enclosed?

Trick question - DNA is not enclosed within a nucleus in a prokaryote

Where is DNA usually arranged in a prokaryote?

One circular chromosome

Breaking down Eu-karyote, what does either word mean?

Eu = true

Karyon = nucleus

What is DNA arranged as within the nucleus of a eukaryote?

Multiple chromosomes

What are the 5 morphology (shapes) of bacteria?

Coccus, bacillus, vibrio, spirillum, spirochete

What shape is coccus/cocci?

Spherical

What shape is bacillus/bacilli?

Rod-shaped

What shape is vibrio?

Curved

What shape is spirillum/spirilla?

Spiral shaped

What shape is spirochete?

Corkscrew shaped

Flip to see bacteria cell structure (for reference)

Some bacteria have capsules + slime layers, which are?

Sticky gelatinous layer external to the cell

What are capsules + slime layers composed of (3)?

Polysaccharide, protein or both

How can you differentiate a capsule from a slime layer?

Layer organized + firmly attached = CAPSULE

Layer unorganized + loosely attached = SLIME LAYER

Some bacteria capsules play a role in virulence (severity of disease), which protects against?

Phagocytosis

What’s an example of a bacterium that plays a role in virulence?

Streptococcus pneumoniae

Is disease caused with or without a presenting capsule?

With a capsule

Slime layers often allow bacteria to attach to surfaces. What are 3 examples?

Medical implants, water pipes, teeth

What’s an example of a bacterium that has a slime layer?

Streptococcus mutans

Streptococcus mutans makes polysaccharide slime from?

Sucrose

Streptococcus mutans attaches to teeth, which causes?

Cavities

What are the 4 typical arrangements of flagella in bacterial cells?

Monotrichous, lophotrichous, amphitrichous, peritrichous

Monotrichous meaning?

Single polar flagellum

Lophotrichous meaning?

2+ flagella originating from one pole

Amphitrichous meaning?

Tufts of flagella originating from opposite poles

Peritrichous meaning?

Flagella distributed all over the cell

Flagella turn causing cell to move in one direction “__”

“run”

Periodically flagella reverse direction causing a random change in direction “______”

“tumble”

Flagella allow chemotaxis, which means?

Movement toward/away from a stimulant

Towards a nutrient, chemotaxis causes?

Attraction (nutrient = attractant)

Away from a toxin, chemotaxis causes?

Repelling (toxin = repellent)

For example, E. coli will move towards ______ (attractant)

“glucose”

Flagellar _______ can be used to distinguish among strains of a species.

“protein”

e.g. E. coli O157:H7

What are fimbriae + pili?

Short, hollow, hair-like appendages (NOT flagella)

What type of bacterial appendage allows the cell to adhere to surfaces?

Fimbriae

What does fimbriae contribute too?

Pathogenicity (ability to cause disease in a host)

Some strains of E. coli have fimbriae that allow them to attach to what?

The intestinal wall

What type of bacterial appendage allows the attachment of 2 bacteria to each other?

Pili

Pili is involved in the transfer of what between bacteria?

Genetic material

What is an example of pili?

E. coli’s sex pilus