Lab Exam 1 Micro - All Topics

If you are sick.....

You must complete the Lab Make-up Request form on Canvas within 48 hours of you being absent
With covid exam make sure your ID is visible

If you have an upcoming absence due to an interview etc....

The make up request form must be submitted at least 72 hours in advance of the lab going to be missed.

Are you allowed to use your phone in lab?

No

Gloves go in the trash or biohazard bin?

Biohazard bin

Glass tubes with media go where?

Biohazard bin

Paper towels go where

The trash

Questions about chemicals can be answered by referring to the appropiate....

Material Safety Data Sheet (MSDS)

Microorganisms are

Ubiquitous in nature
Found everywhere that other lifeforms exist
Have a long rich history on Earth
And have successfully adapted to a wide range of habitats

Microorganisms can be isolated from

Soil (all kinds)
Water (over a large range of salinity)
Plants
Animals (even humans)
Even seemingly uninhabitable places such as the hot acid pools in Yellowstone
(DOES NOT APPLY TO EVERY SPECIES)

Saprophytes

perform important role of decomposition in the ecosystem

Many microorganism do not reside on or in a specific plant or animal host and are not known to cause disease

Free-Living and nonpathogenic

Pathogens

cause damage to host

mutualism

both host and microbe benefit

commensals

microbe benefits but no effect on host

Many of the commensal/ mutualistic strains are

opportunistic pathogens
Inhabit our bodies and are capable of producing a disease state if introduced into a suitable part of the body

Reservoir

is any area where a microbe with the potential to cause infection resides

Blood agar

used to differentiate bacteria based on their hemolytic characteristics
includes 5% sheep blood in a Tryptic Soy Agar base

Several types of Gram Positive cocci produce exotoxins called

Hemolysins ( streptococci produce streptolysins) which are able to destroy RBC's and hemoglobin
destruction of red blood cells

beta hemolysis

Complete destruction

results in clearing of the medium

alpha hemolysis

partial destruction

results in greenish discoloration of the medium

gamma hemolysis

no hemolysis

all cells that are able to grow on the plate will divide and produce visible

colonies

each source will likely have multiple species present representing a

Mixed Culture

If a colony is not contracting any other colonies

it is said to be

A colony appears

Color

Growth characteristics in colonies are influences by

Nutrient availability
Temperature
Incubation time

Shape

Round
Irregular
Punctiform (tiny pinpoint dots)

Margin

Entire (smooth

Elevation

flat
raised
convex
pulvinate
umbonate

Texture

Moist
Mucoid
Butyrous
Dry
Shiny
Dull

Optical properties

Opaque and translucent

Agar slants are primarily used for

Cultivation

Filiform

dense and opaque with a smooth edge

Friable

Crusty

Spreading edge

produced by motile organisms

pellicle

growth floats on top of the medium

sediment

growth sinks to the bottom

uniform fine turbidity

evenly cloudy throughout

Flocculent

clumped growth

germicides

refers to substances or systems

Decontamination

Lowest level of control
"reduction pathogenic microorganisms to a level at which items are safe to handle without protective attire"
Usually includes physical cleaning with soaps or detergents

Disinfection

next level of control
Divided into three sub levels(low

Chemical sterilants

high level disinfectants that have the ability to kill all vegetative cell and some spores

Antispetics

Disinfectants designed to reduce or eliminate pathogens on or in living tissue

Sterilization

Highest level of pathogen control
Complete elimination of viable organisms including spores
can be achieved by some chemicals and gases

steam sterilization

most effective and most common method

Autoclave

device most commonly used for stem sterilization use superheated steam under pressure to kill heat-resistant organisms In the microbiology laboratory:

• sterilization temperature is set between 121 and 127 degrees celcius

• Must reach optimum temperature for at least 15 minutes

• Time varies according to the size and consistency of the material Color coded autoclave tape biological indicators

indicator vial includes

Small ampule containing fermentation broth with pH indicator
strip of filter paper containing bacterial spores

Indicator Vial test

vial is autoclaved at 121 degrees celsius for 15 minutes
ampule is crushed
- Allows fermentation broth to come into contact with bacterial spores
Vials are incubated at 55 degrees celsius for 48 hours

Biological indicators are

the only way with certainty to determine that sterilization has been achieved

While typing organism names

italicized the whole name but only capitalized the first letter or word

While writing organism names

underlined the whole name but only capitalize the first word or letter

Aseptic transfer

Transfer of living microbes from one place to another without contamination of

• the culture

• the sterile medium

• or the surroundings

To prevent contamination

insulating instruments must be sterilized prior to use

• wire loops are sterilized immediately before use in an incinerator

• cool loop in agar before transfer

• mouths of tubes containing culture or media are also incinerated at the time of transfer

• instruments that can not be incinerated are sterilized inside wrappers or containers by autoclaving prior to use

Culture

A medium that contains living microbes

Pure culture

is one that contains a single species

A Mixed Culture

is one that contains multiple species
All cultures are assumed to be mixed prior to obtaining isolation

all cultures are assumed to be _______ prior to obtaining isolation

mixed

Microbial cultures are grown and maintained on or in solid and liquid substances called

media

Media comes on many forms like

Broths
Agar Slants
Plated Media

Broths

are used to grow microbes when fresh cultures or large numbers of cells are required

Agar slants

are generally used to grow stock cultures that can be refrigerated after incubation and maintained for several weeks

Plated Media

are typically used for obtaining isolation of species

what are the methods of isolation

spread plate
pour plate
streak plate

In 1850's Louis Pasteur developed concept of

Pure culture
Allowed microbiologists to study individual species
His methods of isolation were terribly inefficient
- tried to dilute cultures until he thought he had a single cell

In the 1880's Robert Koch developed a procedure for

"streaking for isolation"

The primary purpose of streaking for isolation is to

obtain isolated colonies
An isolated colony is composed of millions of cells- all identical and all descendants from a single cell or group of cells

A single colony

(composed of identical cells) is then used to inoculate another culture

In the quadrant streak method:

a bacterial sample is streaked over the surface of a plated agar medium
During Streaking

commonly used technique

QUADRANT STREAK
used with samples suspected of high cell density

Label the _______ of plates

bottom

Incinerators

Reach temperatures over 800 degrees celsius

Micropipettes

are used for the transfer of small amounts of liquids

The larger blue tips are for the

p1000

The smaller yellow tips are for the

p200
p20

what is the minimum amount you need to transfer to use a p1000 pipette?

200 uL or 0.2mL

what is the maximum amount you need to transfer to use a p1000 pipette?

1000uL or 1mL

what is the minimum amount you need to transfer to use a p200 pipette?

50uL or 0.05mL

what is the maximum amount you need to transfer to use a p200 pipette?

200uL or 0.2mL

what is the minimum amount you need to transfer to use a p20 pipette?

2uL or 0.002mL

what is the maximum amount you need to transfer to use a p20 pipette?

20uL or 0.02mL

Spread Plate method

A method of isolation in which a diluted microbial sample is deposited on an agar plate and spread uniformly across the surface

If properly diluted

Cell density is calculated by

of colonies/ (volume plated x dilution)

What is considered a countable number of colonies?

30-300

Bright- filed microscopy

Produces an image made from light transmitted through specimen

Specimen

restricts light transmission and appears dark against a light background

Contrast

is improved with he application pf stains to the specimen
Staining process usually kills cells

Image formation

Begins with light from an internal or external light source

Light passes through the condenser

Total magnification=

Magnification ocular lens x magnification objective lens

Resolution

the clarity of an image

Limit of resolution

An actual measurement of how apart two points must be for the microscope view them as separate

Resolution improves as limit of resolution decreases

magnitude/ (NA condenser + NA objective)

NA= Numerical aperture

Numerical aperture

measure of a lens ability to "capture" light coming from the specimen and use it to make the image

Determining cell morphology is an important first step in identifying a

Bacterial species

Cocci

singular coccus

Bacilli

singular bacilli

Spirilla (singular: spirillum)

spiral shaped cells

vibrios

curved rods

coccobacilli

short rods