Test : Penicillinase Test
Indications : The test is ordered to detect antibiotic resistance in certain infection- causing bacterial strains. No preparation is required prior to the test.
Samples are collected using cotton swabs from parts of the body, based on the type of infection. It is transferred to a plate containing appropriate media to enable the growth of these organisms.
There are two methods to carry out the acidometric tests.
(i)
Tube method
For this method, 2 mL of 0.5% (w/v) aqueous phenol red solution is made dilute with 16.6 mL of distilled water. Then 1.2 g of
benzylpenicillin is added and the pH is adjusted to 8.5 using 1 M NaOH. This results in a violet colored solution which can be stored at -20°C. 100 µl of this solution is then transferred into tubes or microtitre wells and inoculated with a sample of cultured bacteria to form dense suspensions. Experiments using controls must run parallel to the ones using samples.
(ii)
Paper strip method
For this method, seven numbers of Whatman No. 1 filter paper is cut into 5 x 1 cm strips and allowed to soak in a freshly-prepared solution containing 125 g/L benzylpenicillin, 0.1% (w/v) bromocresol purple and 1.25 mM NaOH.
These strips are later allowed to dry, after which they are stored at 4°C for 6 months. They must be moistened with distilled water before use. A small portion of the bacterial isolates from the culture plates are smeared on the moistened strips.
Physiology : The Penicillinase test is used for rapid detection of beta lactamase production from isolated colonies of
Haemophilus influenzae,
Neisseria gonorrhoeae, Moraxella catarrhalis, S. aureus, and enterococci sp.
Beta lactamase enzyme, also known as penicillin lactamase, renders a few bacterial species resistant to some antibiotics, such as beta lactams. This acidimetric test is based on the fact that hydrolysis of the beta lactam ring generates a carboxyl ring that acidifies unbuffered systems.
Interpretation : Tube method - A yellow color develops within 5 min of inoculation, indicating β-lactamase activity.
Paper strip method - A yellow color develops within five minutes of smearing the sample, indicating β-lactamase activity.
Sample : Haemophilus infeluenzae, Moraxella catarrhalis, Neisseria gonorrhoeae, enterococci, Sttaphylococcus aureus, gram negative anaerobic rods.
Test Method : Acidimetric method
Related Tests : Neisseria gonorrhoeae culture, Susceptibility testing.
References :
1. http://bsac.org.uk/wp-content/uploads/2012/02/Chapter_6.pdf
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