WORKSHEET: Identification of Staphylococci

Answer the following questions: 

1. Why is it necessary to avoid getting colonies from Blood Agar petri plate when
performing catalase test? Explain.
ANSWER: In performing catalase test, getting colonies from Blood agar plate
should be avoided because BAP contains red blood cells and those cells have large
amounts of catalase. Therefore, testing colonies from a blood agar plate for a
catalase test would possibly give a false positive reaction.

2. Enumerate possible explanations if you observe weak catalase positive reaction.

ANSWER: Possibly, the hydrogen peroxide being used isn’t stable anymore, since
the chemical is very unstable. Another possible explanation is because other
bacteria produce peroxidase which cause the

3. Explain why Staphylococcus aureus can gives a negative coagulase reaction after 3
to 4hrs of incubation? Why is it necessary to incubate overnight? 
ANSWER: It is possible for bacteria which utilize citrate to contaminate
Staphylococcus cultures on which the coagulase test is being performed and they
may, upon prolonged incubation, give false-positive results due to utilization of the
citrate. The presence of EDTA in the coagulase plasma should overcome this
problem. 3. When checking results of the Coagulase Test, tubes should be observed
every 30 minutes during the first four hours of incubation. Some strains of
Staphylococcus aureus produce fibrinolysin which may lyse clots formed earlier. If
the tubes are not read until 24 hours of incubation, reversion to a false-negative
may occur.
 An incubation time of 24 h at 35 to 38°C is recommended for the optimal
performance of MRSASelect (Bio-Rad) chromogenic screening agar

Reference:
https://www.pro-lab.com/wp-content/uploads/2017/01/PL850_Rabbit-Coagulase
-Plasma_English.pdf
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2650924/

4. Explain the principle behind Mannitol Salt agar test.

ANSWER: Mannitol Salt Agar has beef extracts and peptones that supply vitamins,
minerals, amino acids and nitrogen that is needed for the growth of the colony. A
sodium chloride concentration of 7.5% affects the partial or complete inhibition of
other bacterial contaminants other than staphylococci.

5. What is the purpose of performing the Novobiocin test?

ANSWER: The purpose of doing a Novobiocin test is to differentiate and
distinguish coagulase-negative staphylocci (CONS). Urine is the commonly used
specimen for this test. Novobiocin testing identifies the isolate as S. saprophyticus
which is resistant to novobiocin.