Diameter of the disk = 8mm
Isolate A
CAZ = 20mm
CAZ-CLV = 30mm
Isolate B
CAZ = no inhibition (0mm????)
CAZ-CLV = 10mm
We would know that isolate A tested positive for ESBL
since the diameter is greater than 5mm with clv
However, for isolate B, this is harder for me to determine
since I can’t make a judgement on whether the diameter
with Caz-clv should be taken to be more than 5mm,
even though the observable increase in diameter is only 2mm
owing to the disk being 8mm itself
(10mm – 8mm = 2mm; ESBL-ve)
OR (10mm – 0mm = 10; ESBL+ve)
(should Isolate B be counted as an ESBL producer???)
Isolate B is obviously ESBLs producer, since there is a enhancment from 0 to 10mm. However, you can confirm by examining the blaCTX or blaTEM in them.
For isolate A, you have demostrated ESBL. For isolate B, the result is indeterminate as you need to take into consideration the diameter of the disc (usually 6 mm). It seems likely that isolate B may have ESBL activity as there is some increased susceptibility with clavulanate - however if the isolate also produces AmpC enzyme this can lead to the type of result you are seeing (i.e. the clavulanate cannot completely nullify the combination of resistance mechanisms). To confirm ESBL using PCR requires screening for a variety of genes (including those encoding CTX-M and variants of SHV, TEM, etc..). It is easier to investigate such isolates further using phenotypic methods e.g. by including an AmpC inhibitor in the disc (e.g. a boronic acid derivative) or by selecting a cephalosporin that is relatively stable to hydrolysis by AmpC ß-lactamase (e.g. cefepime). Such discs are commercially available (e.g.
http://www.mastgrp.com/Identification%20strips%20discs%20and%20rings/Glossies/ESBL_6pp.pdf)
I think the discrepancy cannot be fully resolved with just that test. You could equally use another indicator cephalosporin such as cefotaxime in the combined disc diffusion method by placing amoxicillin-clavulanate disc in-between ceftazidime and cefotaxime at appropriate distance (30mm) after seeding the sensitivity plate with the test organism. This method is far more reliable and detects ESBL production in which several genes have been implicated viz SHV, TEM and CTX-M. Ultimately genotypic tests are most reliable but considering the array of genes you would have to screen for with newer genes/variants increasingly being implicated, I would recommend this simple phenotypic test. Other antibiotics such as aztreonam and ceftriaxone could also be included at the same distance (30mm) from the amoxicillin-clavulanate to increase the sensitivity of the test.
I guess that isolate B does possess ESBL activity. You can not consider the difference in ZOI to be 10 mm. It is actually 4 mm because usually the disc diameter is 6 mm and not 8 mm. Only a difference of 5 mm or more than 5 mm is considered to be postive but still a difference an error of 1 mm is possible during naked eye estimation. Even I had such a issue while screening for ESBL in Acinetobacter baumannii. You can encounter a similar issue when your isolate produces both ESBL and AmpC. In such cases, clavulunate will not be able to withstand the hydrolysing activity of both ESBL and AmpC. How can you proceed is:
A) Measure the ZOI again to confirm.
B) Screening for ESBL secretion genotype by PCR which would be more accurate.
C) If you don't have access to sequencing, you can phenotypically demonstrate by using an AmpC stable cephalosporin like Cefepime.
D) To just rule out AmpC production, you can also use combination discs which also contain an AmpC inhibitor.
- Badges
- Science topic
- Similar topics
- Medicine
- Disease
- Infectious Diseases