Abstract
Background
Recently a novel plasmid-mediated resistant mechanism that conferred high-level resistance to aminoglycoside via methylation of 16S rRNA was reported. The aims of this study were to determine the prevalence of the 16S rRNA methylase genes and to characterize the coresistance to other antibiotics in Gram-negative bacilli.
Methods
Consecutive non-duplicate Gram-negative bacilli were isolated from clinical specimens at a Korean secondary- and tertiary-care hospital from July 2006 to June 2007. The antimicrobial susceptibility was tested by the CLSI agar dilution method, and PCR was performed to detect the 16S rRNA methylase genes in the arbekacin-resistant isolates.
Results
In Gram-negative bacilli, the proportions of 16S rRNA methylase gene-positive isolates were 5% (75/1,471) in the secondary-carehospital and 4% (48/1,251) in the tertiary-care hospital, and the positive rates by species were 1% Escherichiae coli 16% (10/1,062), Klebsiella pneumoniae 16% (75/ 460), K. oxytoca 2% (1/44), Citrobacter spp. 9% (7/ 82), Enterobacter spp. 2% (4/181), Serratia marcescens 6% (6/100), Proteus miriabilis 4% (2/57), Achromobacter xylosoxidans 20% (1/5), Pseudomonas aer- uginosa <1% (1/505), Acinetobacter spp. 10% (11/ 112), and Stenotrophomonas maltophilia 2% (1/66), respectively. Among 16S rRNA methylase-positive isolates from secondary- and tertiary-care hospitals,93% (70/75) and 90% (43/48), respectively, were armA positive, and others, except one rmtA positive isolate, were positive for the rmtB gene, according to PCR results. The rates of ESBL-positive and cefoxitin-resistant K. pneumoniae were 59% and 92%, respectively. In addition, 91% of 16S rRNA methylase-producing K. pneumoniae were positive for qnrB. There were no MBL producers among 16S rRNA methylase-producing Pseudomonas and Acinetobacter species.
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Table 1.
Primers used for PCR detection of 16S rRNA methylase genes, qnr, and qepA genes
Name | Nucleotide sequence (5’→3’) | Product size (bp) | GenBank accession No. |
---|---|---|---|
armA F armA R | AGG TTG TTT CCA TTT CTG AG TCT CTT CCA TTC CCT TCT CC | 590 | AB116388 |
rmtA F rmtA R | CTA GCG TCC ATC CTT TCC TC TTT GCT TCC ATG CCC TTG CC | 635 | AB083212 |
rmtB F rmtB R | CCC AAA CAG ACC GTA GAG GC CTC AAA CTC GGC GGG CAA GC | 584 | AB103506 |
rmtC F rmtC R | CGA AGA AGT AAC AGC CAA AG ATC CCA ACA TCT CTC CCA CT | 711 | AB194779 |
rmtD F rmtD R | ATG AGC GAA CTG AAG GAA AAA CTG C GCT CCA AAA GCG GCA GCA CCT TA | 532 | DQ914960 |
qnrA F qnrA R | AGA GGA TTT CTC ACG CCA GG TGC CAG GCA CAG ATC TTG AC | 580 | qnrA1-qnrA6∗ |
qnrB F qnrB R | GGA ATT GAA ATT CGC CAC TG TTT GCC GCC CGC CAG TCG AA | 264 | qnrB1-qnrB6∗ |
qnrS F qnrS R | GCA AGT TCA TTG AAC AGG GT TCT AAA CCG TCG AGT TCG GCG | 428 | qnrS1-qnrS2∗ |
qepA F qepA R | CCG ACA GGC CCA CGA CGA GGA TGC TCG GCG GCG TGT TGC TGG AGT TCT | 549 | AB263754 |
Table 2.
The rates and distributions of 16S rRNA methylase in gram-negative bacilli isolated at a secondary- and tertiary-care hospital from 2006 to 2007
Bacterial species | Secondary-care hospital (Myongji Hospital) | Tertiary-care hospital (Severance Hospital) | ||||||||
---|---|---|---|---|---|---|---|---|---|---|
No. isolated | No. of PCR positive for | No. isolated | No. (%) of PCR positive for | |||||||
Total (%) | armA | rmtB | armA+rmtB | Total (%) | armA | rmtB | armA+rmtB | |||
E. coli | 646 | 8 (1) | 8 | 0 | 0 | 416 | 2 (<1) | 0 | 1 | 1 |
K. pneumoniae | 245 | 51 (21) | 49 | 0 | 2 | 215 | 24 (11) | 24 | 0 | 0 |
K. oxytoca | ND | 44 | 1 (2) | 1 | 0 | 0 | ||||
Citrobacter spp. | 37 | 3 (8) | 3 | 0 | 0 | 45 | 4 (10) | 2 | 2 | 0 |
Enterobacter spp. | 129 | 2 (2) | 2 | 0 | 0 | 52 | 2 (4) | 2 | 0 | 0 |
S. marcescens | 52 | 4 (8) | 4 | 0 | 0 | 48 | 2 (4) | 2 | 0 | 0 |
P. mirabilis | 57 | 2 (4) | 2 | 0 | 0 | ND | ||||
M. morganii | 27 | 1 (4) | 1 | 0 | 0 | ND | ||||
Providencia spp. | 21 | 3 (14) | 0 | 2 | 1 | ND | ||||
A. xylosoxidans∗ | ND | 5 | 1 (20) | 0 | 0 | 0 | ||||
P. aeruginosa | 257 | 1 (<1) | 1 | 0 | 0 | 248 | 0 (0) | 0 | 0 | 0 |
Acinetobacter spp. | ND | 112 | 11 (10) | 11 | 0 | 0 | ||||
S. maltophilia | ND | 66 | 1 (2) | 1 | 0 | 0 | ||||
Total | 1,471 | 75 (5) | 70 | 2 | 3 | 1,251 | 48 (4) | 44 | 3 | 1 |
Table 3.
Comparisons of MICs (μg/mL) of antibiotics for 16S rRNA methylase-positive and -negative gram-negative bacilli
Table 4.
Comparisons of the rates of β-lactam resistance and quinolone resistance in 16S rRNA methylases-positive and -negative gram-negative bacilli
Bacterial species (No. of isolates) | No. (%) of other resistance mechanisms | |||||
---|---|---|---|---|---|---|
ESBL | PABL | qnrA | qnrB | qnrS | qepA∗ | |
16S rRNA methylase-positive | ||||||
E. coli (10) | 7 (70) | 7 (70) | 0 (0) | 2 (20) | 1 (10) | 1 (10) |
K. pneumoniae (75) Other Enterobacteriaceae (23) | 44 (59) 13 (57) | 69 (92) NT | 1 (2) 2 (9) | 68 (91) 9 (39)† | 0 0 | 0 1 (4) |
Pseudomonas spp. and Acinetobacter spp. (12) | 0‡ | NT | 0 | 1 (8) | 0 | 0 |
16S rRNA methylase-negative | ||||||
E. coli (4) | 2 (50) | 1 (25) | 0 | 0 | 0 | 1 (25) |
K. pneumoniae (14) | 6 (43) | 9 (64) | 0 | 8 (57) | 1 (7) | 0 |
Other Enterobacteriaceae (4) | 1 (25) | NT | 0 | 1 (25) | 0 | 0 |
Pseudomonas spp. and Acinetobacter spp. (11) | NT | NT | 0 | 4 (36) | 0 | 0 |