Distribution of Conjugative-Plasmid-Mediated 16S rRNA Methylase Genes among Amikacin-Resistant Enterobacteriaceae Isolates Collected in 1995 to 1998 and 2001 to 2006 at a University Hospital in South Korea and Identification of Conjugative Plasmids Mediating Dissemination of 16S rRNA Methylase

Hee Young Kang; Ki Young Kim; Jungmin Kim; Je Chul Lee; Yoo Chul Lee; Dong Taek Cho; Sung Yong Seol

doi:10.1128/JCM.01677-07

DOI: 10.1128/JCM.01677-07

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Tables

TABLE 1.

Numbers of isolates studied and rates of resistance to amikacin among Enterobacteriaceae that were collected from a university hospital in South Korea between 1995 and 2006

Species	No. of isolates studied (no. of isolates resistant to amikacin) in:									Total no. of isolates (total no. resistant to amikacin)	Rate of resistance to amikacin (%)
Species	1995	1996	1997	1998	2001 to ∼2002	2003	2004	2005	2006	Total no. of isolates (total no. resistant to amikacin)	Rate of resistance to amikacin (%)
E. coli	132 (0)	197 (5)	157 (1)	6 (2)	492 (13)	156 (9)	418 (14)	908 (31)	860 (25)	3,326 (100)	3.0
K. pneumoniae	60 (2)	61 (3)	141 (3)	42 (3)	304 (8)	83 (5)	226 (26)	605 (111)	537 (108)	2,059 (269)	13.1
Klebsiella spp.	2 (0)	9 (0)	24 (2)	4 (0)	39 (0)	12 (0)	18 (0)	36 (2)	5 (1)	149 (5)	3.4
E. cloacae	27 (4)	25 (3)	29 (9)	5 (2)	86 (5)	13 (2)	65 (6)	109 (6)	125 (5)	484 (42)	8.7
Enterobacter spp.	5 (0)	7 (1)	29 (0)	4 (2)	45 (0)	5 (0)	44 (0)	107 (2)	108 (5)	354 (10)	2.8
S. marcescens	16 (0)	9 (0)	43 (2)	7 (0)	75 (5)	45 (7)	23 (2)	38 (1)	51 (6)	307 (23)	7.5
Serratia spp.								2 (0)	2 (0)	4 (0)	0
C. freundii	4 (0)	2 (0)	8 (1)	1 (1)	15 (0)	14 (2)	17 (0)	47 (3)	53 (1)	161 (8)	5.0
Citrobacter spp.							3 (0)	6 (0)	4 (1)	13 (1)	2.6
Morganella morganii	4 (0)	2 (0)	4 (0)		10 (0)	4 (0)	25 (1)	2 (1)	25 (0)	76 (2)	3.2
P. mirabilis	6 (1)	3 (0)	6 (0)	3 (0)	18 (0)	10 (0)	15 (0)	39 (1)	40 (1)	140 (3)	2.1
Proteus spp.	2 (0)	1 (0)	2 (0)		5 (0)	2 (0)	4 (0)	11 (0)	16 (0)	43 (0)	0
Providencia spp.	3 (0)				3 (0)		2 (0)	3 (0)		11 (0)	0
Total no. of isolates (total no. resistant to amikacin)	261 (7)	316 (12)	443 (18)	72 (10)	1,092 (31)	344 (25)	860 (49)	1,913 (158)	1,826 (153)	7,127 (463)
Rate of resistance to amikacin (%)	2.7	3.8	4.1	13.9	2.8	7.3	5.7	8.3	8.4	6.5

TABLE 2.

Distribution of aacA4 and 16S rRNA methylase genes among the 218 amikacin-resistant isolates of Enterobacteriaceae that were collected from a university hospital in South Korea

Yr of isolation	No. of strains carrying the following gene(s):						Total
Yr of isolation	aacA4 alone	armA alone	armA and aacA4	rmtB alone	rmtB and aacA4	None^a	Total
1995	2						2
1996	9						9
1997	1	7	2				10
1998	1	3	1				5
2001		4		2	5		11
2002		1		5			6
2003			9				9
2004		19	6	1			26
2005		45	8	20	1	1	75
2006		24	24	17			65
Total no. of strains	13	103	50	45	6	1	218

↵ a No gene was detected.

TABLE 3.

Distribution of aacA4 and 16S rRNA methylase genes among the 218 amikacin-resistant isolates of Enterobacteriaceae collected from a university hospital in South Korea between 1995 and 2006

Species	No. of strains carrying the following amikacin resistance gene(s):						Total no. of strains
Species	aacA4 alone	armA alone	armA and aacA4	rmtB alone	rmtB and aacA4	None^a	Total no. of strains
E. coli	5	14	14	17	1	1	52
K. pneumoniae	3	65	23	27	5		123
Klebsiella spp.		1	1				2
E. cloacae	4	17	2				23
Enterobacter spp.	1	5	1				7
S. marcescens			6				6
C. freundii			3	1			4
M. morganii		1					1
Total no. of strains	13	103	50^b	45	6^c	1	218

↵ a No gene was detected.
↵ b Thirteen of 50 strains transferred the armA gene alone to the recipient E. coli strain, as determined by the conjugation assay.
↵ c All six strains transferred the rmtB gene alone to the recipient E. coli strain, as determined by the conjugation assay.

TABLE 4.

Incompatibility groups of conjugative plasmids which confer a high level of resistance to amikacin due to the presence of the 16S rRNA methylase gene and/or aacA4 gene

Incompatibility group(s)	No. of transconjugants carrying the following gene(s):					Total	No. (%) of transconjugants with:		P value^c
Incompatibility group(s)	aacA4 alone	armA alone	armA and aacA4	rmtB alone	None^a	Total	armA	rmtB	P value^c
FIIAs							20 (13.1)		0.007
HI2		4				4	4 (2.6)
L/M		8	29			37	37 (24.2)		<0.001
L/M, F			1			1	1 (0.7)
A/C	2	4	3	41	1	51	7 (4.6)	41 (80.3)	<0.001
A/C, I1-Iγ		1	1			2	2 (1.3)
A/C, F				1		1		1 (2.0)
A/C, F, FIA				1		1		1 (2.0)
F		7		6	1	14	7 (4.6)	6 (11.7)	0.069
F, Y				1		1		1 (2.0)
I1-Iγ				1		1		1 (2.0)
F, FIB	1					1
B/O	4					4
NI^b	4	72	3		1	80	75 (49.0)		<0.001
Total no. of transconjugants	11	116	37	51	3	218	153	51

↵ a No gene was detected.
↵ b NI, not identified.
↵ c The chi-square test was performed.

TABLE 5.

Antimicrobial susceptibilities of transconjugants grouped by presence of the armA or rmtB gene and by plasmid incompatibility group^a

Antimicrobial agent	MIC₅₀ for isolates with the indicated gene and Inc plasmid^b:
	armA (n = 143)					rmtB (n = 47)
	FIIAs (n = 20)	L/M (n = 37)	A/C (n = 7)	F (n = 7)	NI (n = 75)	A/C (n = 41)	F (n = 6)
Ampicillin	512	>512	>512	>512	>512	>512	>512
Aztreonam	<1	8	<1	<1	32	8	<1
Ceftazidime	<1	2	<1	<1	32	2	<1
Cefotaxime	<1	64	<1	<1	4	64	<1
Cefepime	<1	8	<1	<1	<1	8	<1
Cefoxitin	32	4	1	2	32	4	4
Ciprofloxacin	0.25	0.03	0.015	0.03	0.25	0.015	0.25
Chloramphenicol	4	4	64	4	4	128	4
Streptomycin	256	4	512	256	256	256	4
Sulfamethoxazole	16	16	>512	16	16	>512	16
Trimethoprim	<1	<1	<1	8	<1	>512	<1
Tetracycline	<1	<1	1	<1	<1	32	<1

↵ a Groups with fewer than five strains were excluded from calculations of MIC₅₀s.
↵ b Numbers of strains included in each group are shown in parentheses. NI, not identified.

TABLE 6.

Years of isolation and species of Enterobacteriaceae isolates which were able to transfer the plasmids of various incompatibility groups carrying armA or rmtB

Yr of isolation and species^a	No. of strains that transfer a conjugative plasmid of the indicated Inc group carrying gene:
	armA						rmtB
	FIIAs	HI2	L/M	A/C	F	NI^b	A/C	F	I1-Iγ
1997-1998
E. coli				3^d	1
K. pneumoniae
Others		4		3		2
2001
E. coli								1
K. pneumoniae						1	6
Others	3
2002
E. coli							4^e	1^g
K. pneumoniae
Others	1
2003
E. coli			2
K. pneumoniae			1
Others			6
2004
E. coli	1		1	1		2	1
K. pneumoniae	1		1		2	10
Others					1	5
2005
E. coli	1		1		1	5	4^f	4	1
K. pneumoniae	6		2	1	1	30	10	1
Others	2		1		1	1	1
2006
E. coli	1		6			2	2
K. pneumoniae	4		13^c	1		14	15
Others			4			3
Total no. of strains	20	4	38^c	9^d	7	75	43^e, ^f	7^g	1

↵ a Others, Enterobactericeae isolates other than E. coli and K. pneumoniae, including 19 E. cloacae isolates, 6 Enterobacter species isolates, 6 S. marcescens isolates, 4 C. freundii isolates, 2 K. oxytoca isolates, and 1 M. morganii isolate.
↵ b NI, not identified.
↵ c One isolate was positive for L/M and F.
↵ d Two isolates were positive for A/C and I1-Iγ.
↵ e One isolate was positive for A/C and F.
↵ f One isolate was positive for A/C, F, and FIA.
↵ g One isolate was positive for F and Y.