TABLE 1.

Sample information and results of phenotypic assay, conventional PCR, MLST, LF-qPCR, in vitro-synthesized carbapenemase activity assay, and sequencing^a

Strain or variant	Vitek 2 MIC (mg/liter) of:		PCRc	STd	Result of indicated step of PCR-P assay		Sequencing result(s)
	IPM	ETPb			LF-qPCR (CT value)	Antibiotic degradation assay (rate [−%A0/min])e
Strains
E. coli 53 (negative control)	≤1	2	Neg	—	Neg	—	—
A. baumannii 65 (positive control)	≥16	—	+	New	13.27	1.338 ± 0.135	WT
E. coli 23	≤1	2	Neg	—	Neg	—	—
E. coli 59	2	≤0.5	Neg	—	Neg	—	—
E. coli 75	≤1	2	Pos	—	12.89	1.393 ± 0.083	WT
K. pneumoniae 17	≥16	≥8	Pos	—	28.24	1.344 ± 0.092	WT
K. pneumoniae 19	2	1	Neg	—	Neg	—	—
K. pneumoniae 76	2	1	Neg	—	Neg	—	—
E. cloacae 60	≤1	4	Neg	—	Neg	—	—
E. cloacae 63	2	≤0.5	Neg	—	Neg	—	—
K. ozaenae 16	12	≥8	Pos	—	14.4	1.264 ± 0.134	WT
A. baumannii 26	≥16	—	Neg	254	Neg	—	—
A. baumannii 30	≥16	—	Neg	254	Neg	—	—
A. baumannii 31	≥16	—	Pos	254	30.96	1.284 ± 0.093	WT
A. baumannii 33	≥16	—	Neg	254	Neg	—	—
A. baumannii 39	≥16	—	Neg	254	Neg	—	—
A. baumannii 40	≥16	—	Neg	254	Neg	—	—
A. baumannii 41	≥16	—	Pos	254	28	1.301 ± 0.089	WT
A. baumannii 42	≥16	—	Neg	New	Neg	—	—
A. baumannii 59	8	—	Neg	208	Neg	—	—
A. baumannii 62	≥16	—	Pos	208	24.94	1.181 ± 0.245	WT
A. baumannii 63	≥16	—	Pos	381	26.58	1.391 ± 0.053	WT
A. baumannii 66	≥16	—	Neg	368	Neg	—	—
A. baumannii 69	≥16	—	Neg	New	Neg	—	—
A. baumannii 70	≥16	—	Pos	229	14.23	1.511 ± 0.087	WT
Variantsf
NDM-K211E	—	—	Pos	—	10.63	0.132 ± 0.004	A611G
NDM-2(P28A)	—	—	Pos	—	15.73	1.140 ± 0.069	C82G
NDM-3(D95N)	—	—	Pos	—	11.52	1.120 ± 0.047	G283A
NDM-4(M154L)	—	—	Pos	—	11.30	2.236 ± 0.060	A460C
NDM-5(V88L, M154L)	—	—	Pos	—	12.63	1.773 ± 0.017	G262T, A460C
NDM-6(A233V)	—	—	Pos	—	13.27	1.187 ± 0.105	C698T

^aIPM, imipenem; ETP, ertapenem; ST, sequence type; Pos, positive; Neg, negative; —, not tested; WT, wild type.

^bErtapenem has limited activity against A. baumannii.

^cConventional PCR to detect bla_NDM-1.

^dNew, new ST.

^e−%A₀/min is the percentage of absorbance decrease per minute. %A₀ is the percentage of absorbance relative to the absorbance at time zero (just after imipenem is added).

^fbla_NDM-1 variants were constructed by using a site-directed gene mutagenesis kit and overlap PCR. The mutations in these variants were confirmed by sequencing as shown in the last column.