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IRSC MCB2010L Dr. Kay

Toilet paper experiment

to assess the efficiency of toilet paper as a barrier

why did we use E.coli (TP exp)

it is the most frequent cause of enteric infections

why did we use MacConkey agar (TP exp)

selective and differential for enterobacteriaceae which are most commonly associated with urinary tract infections

what did the TP experiment tell you

TP is not a good barrier


Most commonly associated with urinary tract infections
Gram neg rod
oxidase negative
glucose positive

Know the volue of urine we plated out and how we got that volume

we plated 1/1000mll or 10^-3, which is the amount in a calibrated loop

What is the significant finding of colonies in urine test

10^5 or 100,000

Snyder Caries susceptibility test

2% dextrose in the medium - as a fermentable carbohydrate -makes acid and drops the pH
Predicts whether we have decay causing bacteria in our mouth

What organism is a major cause of dental caries

Streptococcus mutans

Interpret results of properly run Snyder Susceptibility test

Green is negative (pH of 4.8 and up)
Yellow is positive (pH of 4.4 and down) if read within 18-24 hrs

what factors influence the size of the zone of inhibition for an antibiotic (Kirby-Bauer)

Diffusibility of the agent
size of the inoculum
type of medium
resistance mechanisms of the test organism

Two antibiotics with same zone of inhibition - how can one be resistant to A but sensitive to B

resistance determination is dtermined by a cutoff number which varies between antibiotics

Kirby-Bauer uses standardized elements:

Mueller-Hinton agar poured to 4mm
pH of 7.2-7.4
swabbed with inoculating broth then incubated for 16-18 hrs at 35+/- 2 degrees C

Resistant zone

no zone

Sensitive zone

large zone of inhibition

Advantages of Enterotube II

ease of use
single inoculation
numerous tests
little media preparation
rapid results
simple interpretation

Disadvantages of Enterotube II

having to confirm questionable test results and multitest systems which are designed for a particular medically important group of pathogens and are not available for a wide variety of microbes

Which test must be performed to confirm the identity of your unknown as a member of Enterobacteriaceae? What is the expected result?

Oxidase test
Enterobacteriaceae are negative for oxidase

5 digit Enterotube II code result beginning with '0'

suggests that speciment is not a glucose fermentor and therefore not enterobacteriaceae

Coagulase positive

Virtually all strains of Staph aureus are coagulase positive

What is the role of coagulase in the pathogenesis of Staph aureus?

Surrounds itself in a coat of clotted blood protein which protects it against host defenses as the bacterium multiplies in the host

What is the role of alpha-toxin in the pathogenesis of Staph aureus?

Alpha toxin is an exotoxin that destroys RBCs to release nutrients and growth factors that increase S.aureus multiplication in the host blood stream

Nosocomial infections

Infections acquired in a hospital or other healthcare setting.

Why are staphylococci among the leading causes of nosocomial infections?

It is an opportunistic pathogen that causes infections of compromised hosts.

Why are staphylococcal infections becoming increasingly difficult to treat?

numerous strains have developed multiple resistances to common antibiotics.

Staphylococcus characteristics

Cluster of cocci
Gram +
Catalase +

Staph aureus hemolytic reaction and coagulase reaction

Alpha hemolytic
Coagulase +

Staph epidermidis hemolytic reaction and coagulase reaction

Non-alpha hemolytic
Coagulase -

Strep pyogenes & Staph aureus
what characteristics do they share?

gram + cocci

Which tests could be used to differentiate between Strep pyogenes & Staph aureus?

Catalase test
(Staph is catalase +)

Which streptococcal species includes cells that are arranged predominantly in pairs rather than chains?

Streptococcus pneumoniae

Vaginal swabs are taken from pregnant women in their third trimester. Which streptococcal species is the focus of the investigation?

Streptococcus agalactiae

Which streptococci and implicated in the development of dental caries?

Streptococcus mutans

Know how to differentiate staph from strep microscopically and biochemically.

Staph: gram +, clusters, catalase +
Strep: gram +, chain, catalase -

What does catalase do?

Catalase breaks down Hydrogen peroxide into water and oxygen

Lancefield Group A, Beta-hemolytic Streptococcus pyogenes

Microscopy: cocci
Catalase: negative
Hemolysis: beta-hemolytic
Bacitracin: postive


visible clumping

what types of compounds in bacterial cells can serve as antigens?


what are immunoglobulins?

Antibodies which are specific for a certain bacterial antigen

Slide agglutination for S.aureus
What two S.aureus antigens are being detected with this test kit?

Coagulase and protein A

What definitive test for S.aureus is highly correlated with this agglutination test?

S. aureus is the only staph species positive for the coagulase tube test whichis demonstrated by the clotting plasma. There is a 97% correlation with the agglutination test.

What advantages does the agglutination test have over the definitive S.aureus test?

agglutination test is more rapid and also tests for a second antigen of S.aureus.

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