Rapid optical determination of β-lactamase and antibiotic activity

BMC Microbiology

Table 2 Comparison of different methods of β-lactamase detection and cefazolin antibiotic susceptibility/activity determination

*S. aureus*isolate #	β-LACTAMSE GENOTYPE (‘blaZ’PCR)	β-LACTAMASE PHENOTYPE			CEFAZOLIN SUSCEPTIBILITY/ACTIVITY
		β-LEAF assay*	Nitrocefin disk test	Zone edge test	Disk diffusion	Antibiotic activity – β-LEAF assay**
	‘+’ = positive PCR		Uniform orange color = ‘+’ (positive)	Sharp zone edge = ‘+’ (positive)	S = susceptible	LA = less active
	$: contained stop codon or deletion				(!) = sharp zone edge	A = active
1	+	+	+	+	S (!)	LA
2	-	-	-	-	S	A
3	+	-	-	-	S	A
4	-	-	-	-	S	A
5	+	-	-	-	S	A
6	+	+	+	+	S (!)	LA
7	+	-	-	-	S	A
8	+	-	-	-	S	A
9	+	-	-	-	S	A
10	+^$	-	-	-	S	A
11	+	-	-	-	S	A
12	+	-	-	-	S	A
13	+	-	-	-	S	A
14	+	-	-	-	S	A
15	+	-	-	-	S	A
16	+^$	-	-	-	S	A
17	+^$	-	-	-	S	A
18	+	+	+	+	S (!)	LA
19	+	+	+	+	S (!)	LA
20	+	+	+	+	S (!)	LA
21	-	-	-	-	S	A
22	+	(Weak) +	-	-	S	A
23	-	-	-	-	S	A
24	Unknown	-	-	-	S	A
25	-	-	-	-	S	A
26	+	-	-	-	S	A
27	+	-	-	-	S	A
	Col. 1	Col. 2	Col. 3	Col. 4	Col. 5	Col. 6

^$Special comment – blaZ contained Stop codon or deletion (so non-functional) (Robert L. Skov, unpublished results).
*Classification into positive and negative is based on proposed cut-off depicted in Figure 2 (upper limit of mean ± 3X Std. deviation for strain #2, β-LEAF probe reaction) to demarcate β-lactamase production. Isolates showing cleavage rates of β-LEAF (black bars in Figure 2) lower than or equal to the cut-off were designated ‘negative’, while isolates with higher cleavage rates were designated ‘positive’.
**Classification of cefazolin as ‘active’ or ‘less active’: When difference in cleavage rates (fluorescence change) in the absence and presence of cefazolin was minimal, antibiotic predicted to be ‘active’. Drastically lowered cleavage rate in presence of cefazolin compared to when probe assayed alone led to prediction of cefazolin as ‘less active’ respectively (also see Figure 2).
Details of Disk Diffusion results are presented in Table 3.

ISSN: 1471-2180