Microbiology Lab

• Either inhibits the growth of a certain group of microbes or supports the growth of very few microbes
• Antibiotics
• Salts
• Toxic Dyes
• Limit carbon/energy source
• Either Gram + OR Gram -

• Distinguish closely related organisms from one another (usually biochemical abilities)
• Typically a visual difference (pH indicator)

• A type of selective medium
• Contains NaCL and mannitol
• Lacks phenol red
• Favors colony pigmentation by different strains of S. aureus
• No color change as mannitol is fermented

A type of differential medium

• A partial breakdown of hemoglobin to methemoglobin
• Red to green

Capable of completely hydrolyzing hemoglobin

Possess no hemolytic actions whatsoever

• Indicator: Neutral red, pH
• Inhibitor: Crystal violet and bile salts are toxic to gram +
• Results:
• -Gram += inhibited growth
• -Gram - fermenting lactose= pink colonies
• -Gram - not fermenting lactose= colorless colonies

• Indicator: Methylene blue and eosin Y
• Inhibitor: Methylene blue is toxic to gram +
• Results:
• -Gram += inhibited growth
• -Gram - fermenting lactose= pink or purple colonies with a metallic green sheen
• -Gram - not fermenting lactose= colorless colonies

• Indicator: Neutral red for lactose fermentation, FeS for H₂S production
• Inhibitor: Sodium citrate and brilliant green dye are toxic to most bacteria except these
• Results:
• -Gram += inhibited growth
• -Gram - fermenting lactose= pink colonies; if H₂S is produced, the center of the colony will be black
• -Gram - not fermenting lactose= colorless colonies

• Indicator: Phenol red, pH
• Inhibitor: 7.5% NaCl
• Results:
• -Gram + staphylococci fermenting mannitol= yellow media, neutral pH
• -Gram + staphylococci not fermenting mannitol= red media, basic pH
• -Gram + streptococci= inhibited growth
• -Gram -= inhibited growth
• NaCl inhibits other organisms than staphylococci

How to Remove Immersion Oil

• Ensures most light will travel straight into the objective lens
• Prevents light from being lost to the surrounding air

• Two types of dye- acidic and basic
• Used to see the morphology and shape of bacteria

Colors the cell, leaving the background colorless

Cell remains colorless, the background retains the stain

• Any organism that takes up the red safranin in gram staining
• Red
• Have a higher lipid content in the cell wall
• Lipids are dissolved by the alcohol causing the crystal violet to leak from the cell

• Any organism that retains the primary dye in gram staining
• Purple
• Have a thick layer of peptidoglycan
• Pores in the cell wall are closed and prevent the loss of the crystal violet

A method to distinguish between different types of bacteria

Common species that form endospores

• Composed of designated amounts of specific compounds
• Exact contents are known
• Amounts are known

• Contain nutrient-rich substances and precise chemical constituents are unknown
• Amounts of chemicals are not known
• Yeast extract, peptone, and tryptone are some substances are used

Why can't all bacteria be cultured?

• Useful in isolating colonies
• Can be made from broth and/or plate cultures

• Useful in quantifying bacteria
• Also isolate colonies (if proper dilution is used)
• Must be done with a liquid culture

• A function of molecular structure weight, charge, solubility, and diffusion within the medium
• Indicates some chemical activity occurred in vitro

When microbes grow within or on the surface of foods

A potential major health hazard

Types of fermenting

Why store food at low temperatures

• Live in very hot conditions (above 45C)
• Compost heaps, hot springs

• Live in conditions like body temperature (~37C)
• Most human pathogens

• Live in cooler climates
• Refrigerators (4C)
• Causes spoilage of refrigerated foods

What changes the osmotic conditions of food environments?

Live in high salt conditions

Live in high sugar concentrations

Live in neutral pH environments

Live in more acidic pH's

• Live in higher pH environments
• Degrade proteins for their energy
• Produce amines and ammonia

What can prevent food spoilage?

Babies are sterile until...

• Live with humans in a harmonious relationship
• Normal flora

Live off the nutrients of dead cells and tissues

Some bacteria produce...

What changes the structure of the normal flora community in a host?

• Normal flora for the skins
• Deadly pathogen if introduced in the bloodstream

• Normal flora:
• Staphylococcus
• Corynebacterium
• Proprionibacterium
• Streptococcus

• Normal flora:
• Streptococcus
• Corynebacterium
• Neisseria
• Branhamella

• Normal flora:
• Escherichia
• Bacteriodes
• Lactobacillus
• Candida
• Clostridium
• Enterococcus

• Normal flora:
• Escherichia
• Lactobacillus
• Enterococcus
• Bacteriodes
• Streptococcus

• Normal flora:
• Lactobacillus
• Candida
• Stretococcus

• Dry environment
• Acidic pH
• Inhospitable for many microbes
• Microflora are gram-positive
• Anaerobic bacteria living in pores

• Metabolizes sebum
• Releases fatty acids
• Enhance anti-microbial environment provided by the skin

Two medias used for primary isolation on skin

Gram stains help differentiate between what two bacterias?

• Checks for the presence of the enzyme catalyse
• Splits hydrogen peroxide into hydrogen and water
• Bubbles= positive
• Lack of bubbles= negative

• Gram positive cocci
• Either micrococci or staphylococcus

• Gram positive cocci
• Streptococcus

• Belong to Lancefield groups A, B, and D
• Most important pathogens

• Uses blood agar
• BAP distinguishes between 3 different types of hemolysis

• Greenish zone around the colony
• Due to partial breakdown of red blood cells
• Most bacillus species

• Clear around the colonies
• Complete lysis of the red blood cells
• Strep pyogenes, Staph aureus, and all gram negative enteric bacteria

• No hemolysis
• Staph epidermis and Strep faecalis

Over incubation in the hemolysins' test causes

• Confirms Staphylococcus aureus
• 97% of strains are positive
• Others are negative
• Test only valid for gram-positive
• Some gram-negative rods cause false-positive reations

• A loose clot
• Solid immovable clot

Slanted agar

Split complex organic compounds into smaller units in the presence of water

Identify bacteria

• Bacteria that hydrolyze starch produce..
• Clear

The breakdown of starch produces...

Iodine and Starch

• Gelatin is a protein that can be broken down by the enzyme..
• Indicated by the liquid in the tube

Medium has a clear zone

Two medias used for primary isolation in the nose

Fussy eaters

What type of agar is used for the throat?

What type of test is designed for the throat?

Differentiates alpha-hemolytic viridans group streptococci from the pneumococci

An enlarged arrowhead-shaped hemolytic zone at the juncture where the unknown meets the S. aureus means it is positive

Enteric bacteria may be isolated by...

Two types of agar used for normal flora of the intestinal tract

Contains bile salts to inhibit non-enteric bacteria and two dyes, neutral red and crystal violet, which are taken up by lactose-fermenting bacteria

Inhibitory to gram-positive bacteria

• Appears red
• E. coli is brick red
• Enterobacter, Klebsiella and other lactose fermenters are pink to red with a mucoid texture

Isolates to confirm the presence of gram-negative rods

• E. coli- blue-black with a green metallic sheen in reflected light
• Enterobacter aerogenes- convex and mucoid with dark centers and creamy pink borders

• Valuable medium for the further differentiation of enteric bacteria
• Contains three carbohydrates: glucose, lactose, and sucrose
• pH indicator phenol red
• Anaerobic conditions
• Contains iron ions and sodium thiosulfate

Cracks in the agar of TSI means..

If the TSI medium is black, that means...

Bacteria isolated on MAC and EMB plates will be differentiated by their reactions in...

IMViC Series

• Amino acids are produced as a result of the hydrolytic degradation of proteins, peptones, and peptides
• Tryptophan

• Depends on an organism's ability to ferment glucose to acid
• Red in acid solutions
• Yellow in alkaline or neutral solutions

• Depends on the digestion of glucose to acetylmethylcarbinol
• If present it will form a red color

• Based on the ability of certain bacteria to utilize a salt of a citric acid as a sole carbon source in growth
• Indicator bromthymol blue becomes deep blue when pH of medium rises
• If utilization does not occur, then appears green

• Positive- presence of a red surface layer
• Negative- no production

• Positive- acid is produced- red
• Negative- yellow

• Positive- red
• Negative- lack of color change

• Positive- ability to utilize citrate- blue
• Negative- green

Common cause of UTIs

Two major infections of the urinary tract

Two types of agar used for primary detection in urinary tract

Range of countable colonies

Transfer of a known volume of a sample into a known volume of diluent

Dilution=

Total dilution=

Dilution Factor=

= # of bacteria in the original sample

How do you score an enterotube test?

What tests are E. coli positive and negative in IMViC?

What is the indicator for citrate and methyl red test?

What are 3 general outcomes of TSI slant?

Hospital acquired infections

What are the indicators in MAC and EMB agar?

Bacteria that grows on the outside of the body is...

Bacteria that grows on the inside of the body is...

What are the two bacteria used in IMViC?

Pair of cocci or bacilli

chain of three or more cocci or bacilli

Cluster of cocci

Packet of four cocci in a square

Packet of eight cocci in a cube

What part of your body is sterile?

What two types of bacteria are used in water?

What are two types of hydrolysis reactions?