Microbiology : Lecture 2

culture

introducing microorganisms to a medium that supports their growth and multiplication

classification

the process of arranging microbes to groups based on similar characteristics

Inoculation → Incubation → Isolation → Inspection → Identification

The Five I’s (in order)

Inoculation

adding a microbial sample to a medium

Incubation

giving the culture the right conditions to grow

Isolation

separating species, if necessary

Inspection

perform tests and observe

Identification

use this information to determine the organism

medium (inoculation); (media)

provides nutrients to microorganisms which allows them to grow outside their natural habitat

colony

A discrete mount of cells

Isolated Colony

A colony that was formed from a single cell and contains just one species

sterile (aseptic) technique

prevents contamination; limits exposure to environment, disinfect materials

agar

solid physical state of media

defined media

compositions are precisely chemically defined (salts, amino acids, water)

complex media

one or more component of the medium is not chemically definable (ex : extracts from animals, blood, serum, ground up cells, milk, TSA)

general purpose media

allow the growth of as broad of a spectrum of microbes as possible (complex media, TSA, nutrient agar and broth)

enriched media

contain complex organic substances usch as blood, serum, hemoglobin, or growth factors that certain species must have in order to grow (sheep blood agar, chocolate agar)

fastidious

bacteria that require enriched media

selective media

contains one or more agents that inhibit the growth of certain types of microbes but not others (MacConkey agar, tomato juice agar, mannitol salt agar (MSA))

differential media

do not inhibit growth, but do display differences in how microbes grow (MSA, MacConkey agar)

MacConkey Agar

• Selective and Differential

• contains bile and crystal violet, which select for intestinal bacteria and gram-negative bacteria

• contains lactose and neutral red (pH indicator), which differentiate bacteria based on their ability to ferment lactose

Mannitol Salt Agar (MSA)

• selective and differential

• contains 7.5% NaCl, which selects for bacteria that can tolerate high salt concentrations

• contains mannitol, which differentiates bacteria based on their ability to ferment this agar.

Human Body (The Taj Mahal)

• bathed in nutrients

• waste washed away

• moist and comfy

Petri Plate (The Sahara Desert)

• limited nutrients

• build up of waste

• very dry

millimeters > micrometer > nanometers

nanometers, millimeters, and micrometers in order from biggest to smallest

Resolution (microscope)

the capacity of an optical system to distinguish two adjacent objects or points from one another.

oil immersion lens (100x)

• helps capture light that would be lost to scatter

• increases resolution to .2 um, meaning objects .2 um in diameter and objects .2 um apart can be differentiated.

contract (microscopes)

how well an object is differentiated from its background and surroundings

dyes (stains) and microscopy techniques

we use ___ and _____ _____ to increase contrast

fluorescence microscopy

• uses UV light

• uses fluorescent proteins and dyes

• UV rays excite the fluorescent molecules and they emit light

Transmission Electron Microscopy (TEM)

• used to see detailed structures

• has .5 nm resolution

• transmits electrons through specimens

• must be thin samples

• cross-sections

  “T = through”

Scanning Electron Microscopy (SEM)

• used for a detailed, 3D view of all kinds of objects

• 10 nm resolution

• shoots electrons at objects and detects their shape by how the electrons bounce

• “S=surface'“

Fresh, living preparations

“wet mounts” - microbe is suspended in liquid (water, saline, broth) that temporarily maintains the viability of the organism

fixed specimens

• organisms are killed and stuck in place

staining

the process of adding colored dyes to specimens

simple stain

only a single dye

differential stains

use two differently colored dyes that distinguish between cell types or parts

The Gram Stain

universal diagnostic technique for bacteria

purple

gram-positive stain color

red (pink)

gram-negative stain color