Microbiology Lab Practical 2

what is a mesophile?

a species that is capable of growth at medium (comfortable for humans) temperatures

what is an obligate halophile?

require high salt concentration for growth

what is a facultative halophile?

can thrive in both high and low salt environments

What are we going to use fluid thioglycollate medium to indicate?

presence or absence of oxygen

How will we make a jar anaerobic?

Place a hydrogen generator packet in the jar along with catalyst. The hydrogen reacts with the oxygen in the jar.

What is an obligate anaerobe and where does its growth appear in fluid thioglycollate medium?

obligate anaerobes cannot grow with oxygen - growth will appear at the bottom of the FT medium.

What is an obligate aerobe and where does its growth appear in fluid thioglycollate medium?

obligate aerobe can only grow with oxygen - growth will appear at the top of the FT medium.

What is an facultative anaerobe and where does its growth appear in fluid thioglycollate medium?

facultative anaerobes can grow without oxygen with grow really well with it - growth will appear all throughout the FT tube but there will be a cluster of growth at the top of the tube.

What is an aerotolerant anaerobe and where does its growth appear in fluid thioglycollate medium?

aerotolerant anaerobes grow well with or without oxygen - growth will be seen all throughtout the FT tube.

What is an microaerophile and where does its growth appear in fluid thioglycollate medium?

microaerophiles can only grow with small amounts of oxygen - growth will be seen near the top of the tube just below where obligate aerobes would lie.

What does the oxidase test tell us about bacteria? Describe what a positive and negative result looks like.

The oxidase test tells us whether the bacteria makes oxidase or not (an enzyme essential for aerobic respiration) - a positive result will turn the solution blue/purple after 15 seconds - a negative result with turn the solution yellow or have no color change at the 15 second mark.

Catalase breaks down ________________, producing __________ and ___________.

Catalase breaks down ____hydrogen___, producing __oxygen__ and ___water___.

How can one tell if catalase is acting on its substrate?

bubbles will appear (bubbles = positive, none = negative)

Phenol red broth is a differential media. What does a color change in a phenol red broth indicate about a bacterium that has been growing in it?

whether or not the bacteria have the enzymes necessary to ferment the sugar in the tube.

what color does phenol red broth with neutral pH?

red

what color does phenol red broth with acidic pH?

yellow

what color does phenol red broth with alkaline pH?

pink

What is a Durham tube? What does it collect?

A tiny tube that sits upside down in both phenol broth tubes and collects gas.

-What two products may be produced during fermentation of simple carbohydrates?

-can you have one without the other?

-acid and gas

-yes

How are acid and gas identified in the fermentation test?

if the solution is acidic, it will turn yellow. If its not acidic, it will remain red. Gas can be seen visibly in the durham tube.

Name 3 simple carbohydrates that can be added to media for biochemical testing.

Glucose, sucrose, lactose

-icidal

to kill

-static

to inhibit

A disinfectant must __ the pathogen while in contact with them

kill

two types of bacterial cells that are more resistant to disinfectants

endospore forming (bacillus and clostridium) and acid fast (mycobacterium)

Why is antimicrobial susceptibility testing done?

to determine which antibiotic will be most effective against a certain infection

What is the name of the antimicrobial susceptibility test also known as the filter paper disk agar diffusion method?

kirby-bauer test

in the kirby-bauer test, what kind of agar is used?

mueller-hinton agar

in the kirby-bauer test, what does the 'R' stand for?

resistant- bacteria is resistant to the antibiotic on plate

in the kirby-bauer test, what does the 'S' stand for?

susceptible- bacteria is susceptible to the antibiotic on plate

what is a narrow-spectrum antibiotic and an example of one:

Effective against a restricted number of bacteria - i.e penicillin or amoxicillin

what is a broad-spectrum antibiotic and an example of one:

Effective against a wide range of both gram positive and negative bacteria - i.e ampicillin and chloramphenicol

what kind of bacteria only grows on blood agar?

fastidious (has complex needs)

What is enriched media? Give examples.

media which allows many type of bacteria to grow including fastidious - i.e blood agar and chocolate agar

What is differential media? Give examples.

media which allows you to distinguish one group of bacteria from another once grown in the media - i.e mannitol salt agar and blood agar

What is selective media? Give examples.

media in which only specific types of bacteria can grow on while other cannot - mannitol salt agar

Describe the characteristics of Staphylococci.

gram positive spheres that grow in irregular clusters; catalase positive; salt-tolerant - normally found on our skin and our membranes

Where is S. epidermidis normally found? Does it cause any diseases? If so, what?

found on skin and mucous membranes - normally does not cause disease

Where is S. saprophyticus normally found? Does it cause any diseases? If so, what?

found infrequently on our bodies - associated with urinary tract infections in women

Where is S. aureus normally found? Does it cause any diseases? If so, what?

found on skin, internal nares, vagina - causes many diseases including TTS, food poisoning, scaled skin syndrome, pneumonia, meningitis, sinusitis, otitis media, and abscessess

What do hemolysins do?

destory red blood cells

What does coagulase do?

causes plasma to clot

what does leucocidin do?

destroys white blood cells

What does hyaluronidase do?

breaks down connective tissue

what does staphylokinase do?

dissolves fibrin clots

what does enterotoxin do?

affects the digestive system

what does gelatinase do?

liquify gelatin

what species of bacteria contains all the toxins or enzymes discussed in lab 10?

staphylococcus aureus

What does Mannitol Salt Agar (MSA) select for? What ingredient is present in MSA that causes it to be selective?

MSA is selective for salt-tolerant bacteria (like staph) - MSA contains 9.5% salt

How is MSA differential? What does it differentiate between? What ingredients are present in MSA that cause it to be differential?

MSA distinguishes from mannitol fermenters from NON fermenters - MSA contains mannitol and an acid-base indicator

what color does s. aureus present on blood agar and what type of hemolysis does this indicate?

yellow - beta

what color does s. epidermis present on blood agar and what type of hemolysis does this indicate?

white - gamma

what color does s. saprophyticus present on blood agar and what type of hemolysis does this indicate?

white or yellow - gamma

what is the coagulase test result for s. aureus?

postive

what is the coagulase test result for s. epidermis?

negative

what is the coagulase test result for s. saprophyticus?

negative

what is the mannitol fermentation result for s. aureus?

positive

what is the mannitol fermentation result for s. epidermis?

negative

what is the mannitol fermentation result for s. saprophyticus?

varies (+,-)

Is S. aureus susceptible or resistant to novobiocin?

susceptible

is S. epidermis susceptible or resistant to novobiocin?

susceptible

Is S. saprophyticus resistant or susceptible to novobiocin?

resistant

Describe the members of the genus Streptococcus.

Gram-positive spheres arranged in chains; catalase negative

What does fastidious mean?

bacteria that require growth factors and complex nutrients

what type of bacteria does strep need to grow on?

enriched - blood agar

substances that destroy red blood cells are called ____

hemolysins

what color does alpha hemolytic strep produce on blood agar plate?

green - because they do not completely break down hemoglobin

what color does beta hemolytic strep produce on blood agar plate?

yellow - because they completely break down hemoglobin

what color does gamma hemolytic strep produce on blood agar plate?

no change "white"

Which group causes disease? List several of the diseases that are caused by a member of this group.

beta hemolytic (streptococcus pyogenes) - TTS, strep throat, scarlet fever, and rheumatic fever

What is an antigen?

any substances that causes antibodies to be produced in the body - only reacts with specific antibodies

what group do human strep pathogens belong to?

group a

what does a positive coagulase test look like?

coagulase positive is indicated by clot formation

what does a negative coagulase test look like?

no clot formation

what does a positive gelatinase test look like?

solution is liquid after incubation

what does a negative gelatinase test look like?

solution becomes solid after incubation

what does a positive oxidase test look like?

purple/blue color appears after 15 seconds

what does a negative oxidase test look like?

no color change/yellow after 15 seconds

what does a positive phenol broth result look like?

turns yellow

Describe the Enterobacteriaceae

gram-negative nonsporing rods, facultative anaerobes

Enterobacteriaceae that causes pneumonia

Klebsiella pneumoniae

Enterobacteriaceae that causes shigellosis

Shigella dysenteriae

Enterobacteriaceae that causes typhoid fever

Salmonella typhi

How do the pathogenic enterobacteriacaeae differ from the non-pathogenic forms?

the pathogenic forms DO NOT ferment lactose - the nonpathogenic forms DO ferment lactose.

do all e. coli normally causes disease?

no

What are MacConkey and EMB agar selective for?

gram negative rods

What do MacConkey and EMB agar differentiate?

lactose fermenters from non-lactose fermenters

Phenylalanine is broken down into _________ when bacteria that produce

phenylalanine deaminase are grown in culture.

phenylpyruvic acid

In the phenylalanine deaminase test, a reagent called ferric chloride is added after incubation. What does a

positive reaction look like (color)?

green

Name a genus in Enterobacteriaceae that is phenylalanine deaminase positive.

proteus

Urea is hydrolyzed into ________ and _________

carbon dioxide and ammonia

Name the exoenzyme studied in the urea hydrolysis exercise

urease

Name one genus of bacteria that can hydrolyze urea.

proteus

After incubation, how can you tell if a urea slant was inoculated with a urease-positive organism or a

urease-negative organism?

a positive reaction yields a pink color change after incubation - a negative reaction turns yellow/orange

Name the indicator that was added to the urea slant before inoculation

phenol red

the indicator added to the urea slant before inoculation changes solution _______ in a solution below pH 8.4 and ________ above pH 8.4

below pH 8.4 -> yellow/orange

above pH 8.4 -> red/pink

What does the "SIM" acronym stand for?

Sulfide, Indole, Motility

Name the enzyme responsible for sulfide production

cysteine desulfurase or thiosulfate reductase

After incubation, how can you tell if a SIM tube had been inoculated with an organism positive for sulfide production?

a black precipitate formed

Name a genus in Enterobacteriaceae that is positive for sulfide production.

salmonella, shigella, proteus