Profiling of 16S rRNA methylase genes and the flanking sequences in extensively drug resistant Pseudomonas aeruginosa / 中国感染与化疗杂志

Objective To profile 16S rRNA methylase genes and the flanking sequences in extensively drug resistant (XDR) Pseudomonas aeruginosa. Methods A total of 59 strains of XDR P. aeruginosa were collected. MICs of antimicrobial agents against these strains were determined by agar dilution method. The genes encoding 16S rRNA methylase (armA, rmtA, rmtB, rmtC, rmtD, rmtE, and npmA) were analyzed by PCR. The homology of strains was studied by enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR). The structure of armA and rmtB flanking regions were characterized. Results The overall prevalence of armA or rmtB genes was 62.7％ (37/59) in XDR P. aeruginosa isolates, specifically armA positive in 17 strains and rmtB positive in 22 strains. The rmtA, rmtC, rmtD, or npmA gene was not identified in any strain. ERICPCR generated 19 types (or clones) from the 59 strains. The 17 armA-positive strains were distributed in 3 clones, and 88.2％ of the armA-positive strains (15) belonged to clone D. The rmtB gene was dispersed in 9 clones. Flanking sequence analysis demonstrated that armA gene was located in a mobile element carrying multiple transposases. The sequence of the mobile element showed 99％ similarity with the sequence of armA-positive plasmid carried by E. coli and A. baumannii. The rmtB gene was also located in a mobile element containing multiple transposases. The sequence of the mobile element was highly consistent with that of rmtB-positive plasmid carried by E. coli and K. pneumoniae. Conclusions The genes encoding 16S rRNA methylase are prevalent in XDR P. aeruginosa strains. All the genes were identified in high-level gentamicin-resistant strains. Clonal dissemination may explain the spread of armA among P. aeruginosa isolates.

Antimicrobial Susceptibility and Clonal Distribution of the Blood Isolates of Pseudomonas aeruginosa from Two Korean Hospitals

An increasing prevalence of infections caused by multidrug-resistant (MDR) Pseudomonas aeruginosa (P. aeruginosa) causes a serious therapeutic problem in clinical setting. This study investigated the antimicrobial susceptibility, resistance mechanisms against aminoglycosides, and molecular epidemiology of 76 blood isolates of P. aeruginosa from two Korean hospitals. Thirty-four isolates were susceptible to all 13 antimicrobial agents tested, whereas 28 isolates showed a MDR or extensively drug-resistant phenotype. There was a significant difference in resistance rates of P. aeruginosa isolates against aztreonam, piperacillin-tazobactam, imipenem, meropenem, ciprofloxacin, and norfloxacin between two hospitals. Genes for aminoglycoside-modifying enzymes (AMEs), including aphA6 (n = 14), aadB (n = 11), aacA4 (n = 8), and aphA1 (n = 1), and 16S rRNA methylase armA (n = 6) were detected in 26 P. aeruginosa isolates resistant to aminoglycosides. There was no significant difference in carriage of genes for AME and 16S rRNA methylase between two hospitals, but aacA4 and aphA1 were specifically detected in P. aeruginosa isolates from one hospital. Seventy-six P. aeruginosa isolates were classified into 55 pulsotypes at similarity value of 0.85, and 31 and 24 pulsotypes were specifically detected in each hospital. This study demonstrates that differences in antimicrobial susceptibility of P. aeruginosa isolates between two hospitals are possibly due to the presence of diverse clones specific in each hospital.

Detection and epidemiology of drug resistance related blaOXA-23 and armA genes of Acinetobacter baumannii / 中华传染病杂志

Objective To investigate the drug resistant genes against carbapenems,aminoglycosides and quinolones and the molecular epidemiology of clinical isolates of Acinetobacter baumannii.Methods Forty non-duplicate strains of Acinetobacter baumannii were collected from clinical specimens in First Affiliated Hospital of Wenzhou Medical University.The identification of strains was conducted by Vitek 2 Compact system.The susceptibilities to antimicrobials commonly used were determined by agar plate dilution method and broth microdilution method.The presence of class B metalloenzyme-encoding genes (blaIMP,blaVIM,blaNDM,blaSIM,blaGIM),class D cabapenemase-encoding genes (blaOXA-23,blaOXA-48,blaOXA-58),16S rRNA methylase genes (armA,rmtB) and quinolone resistance-determining regions (QRDR) in gyrA and parC were detected by polymerase chain reaction (PCR) and sequenced.Chromosomal or plasmid location of blaOXA-23 and armA genes were assessed by Southern blot.Multiple loci sequence classification (MLST) was performed to analyze the molecular epidemiology of these strains.Results All of the 40 isolates were multi-drug resistant Acinetobacterbaumannii (MDR-AB) and showed high level resistance to all of the tested antimicrobial agents excluding colistin and tigecycline.The positive rates of blaOXA-23 and armA were 90％ and 95％,respectively.All of the 40 isolates carried QRDR mutations in gyrA and parC genes,leading to the Ser83→ Leu and the Ser80→ Leu amino-acid substitutions,respectively.Southern blot showed the chromosomal location of blaOXA-23 and armA genes.Six different ST (ST191,ST381,ST373,ST426,ST208 and ST207) were assigned for these isolates by MLST and the most dominant clones were ST191 (23/40) and ST381 (10/40).Conclusions The predominant cabapenemase-encoding gene and 16S rRNA methylase gene of Acinetobacter baumannii isolates in First Affiliated Hospital of Wenzhou Medical University are blaOXA-23 and armA,respectively,which may be located on the chromosome and vertically transmit the drug resistance.ST191 MDR-AB with blaOXa-23 and armA gene clonally spread in this hospital.

Genetic basis of high level aminoglycoside resistance in Acinetobacter baumannii from Beijing, China

The objective of this study was to investigate the genetic basis of high level aminoglycoside resistance in Acinetobacter baumannii clinical isolates from Beijing, China. 173 A. baumannii clinical isolates from hospitals in Beijing from 2006 to 2009 were first subjected to high level aminoglycoside resistance (HLAR, MIC to gentamicin and amikacin>512 µg/mL) phenotype selection by broth microdilution method. The strains were then subjected to genetic basis analysis by PCR detection of the aminoglycoside modifying enzyme genes (aac(3)-I, aac(3)-IIc, aac(6')-Ib, aac(6')-II, aph(4)-Ia, aph(3')-I, aph(3')-IIb, aph(3')-IIIa, aph(3')-VIa, aph(2″)-Ib, aph(2″)-Ic, aph(2″)-Id, ant(2″)-Ia, ant(3″)-I and ant(4')-Ia) and the 16S rRNA methylase genes (armA, rmtB and rmtC). Correlation analysis between the presence of aminoglycoside resistance gene and HLAR phenotype were performed by SPSS. Totally 102 (58.96%) HLAR isolates were selected. The HLAR rates for year 2006, 2007, 2008 and 2009 were 52.63%, 65.22%, 51.11% and 70.83%, respectively. Five modifying enzyme genes (aac(3)-I, detection rate of 65.69%; aac(6')-Ib, detection rate of 45.10%; aph(3')-I, detection rate of 47.06%; aph(3')-IIb, detection rate of 0.98%; ant(3″)-I, detection rate of 95.10%) and one methylase gene (armA, detection rate of 98.04%) were detected in the 102 A. baumannii with aac(3)-I+aac(6')-Ib+ant(3″)-I+armA (detection rate of 25.49%), aac(3)-I+aph(3')-I+ant(3″)-I+armA (detection rate of 21.57%) and ant(3″)-I+armA (detection rate of 12.75%) being the most prevalent gene profiles. The values of chi-square tests showed correlation of armA, ant(3″)-I, aac(3)-I, aph(3')-I and aac(6')-Ib with HLAR. armA had significant correlation (contingency coefficient 0.685) and good contingency with HLAR (kappa 0.940). The high rates of HLAR may cause a serious problem for combination therapy of aminoglycoside with β-lactams against A. baumannii infections. As armA was reported to be able to cause high level aminoglycoside resistance to most of the clinical important aminoglycosides (gentamicin, amikacin, tobramycin, etc), the function of aminoglycoside modifying enzyme gene(s) in A. baumannii carrying armA deserves further investigation.

Drug Resistance Mechanism of Patients Infected with Aminoglycoside-resistant Acinetobacter Baumannii in Emergency Intensive Care Unit / 医药导报

Objective To investigate drug resistance mechanism of aminoglycoside-resistant Acinetobacter baumannii by detecting 16S rRNA methylase gene and three common genes of aminoglycoside-modifying enzymes in Acinetobacter baumannii infected patients at EICU. Methods The 48 Acinetobacter baumannii strains were collected,and antimicrobial susceptibility tests were performed by VITEK automicroscan. The MIC was detected by 2-fold agar dilution method,and genes were analyzed by polymerase chain reaction( PCR) . Results Among 48 strains,28 were highly resistant to aminoglycosides and 20 showed lower resistances. The 16S rRNA armA,APH(3')-I,ANT(3'')-Ia,AAC(6')-Ib genes were detected in 71. 43%,60. 71%,82. 14%, and 53. 57%of the 28 highly resistant strains,but only present in 0. 00%,0. 05%,0. 05%,and 0. 05%of the low-resistant isolates(P<0. 01). Conclusion The aminoglycoside-modifying enzymes and 16S rRNA methylase were frequently found in Acinetobacter baumannii clinical isolates,which is closely related to the high-level resistance to aminoglycoside antibiotics.

Drug resistance genes of 16S rRNA methylase in Klebsiella pneumoniae / 中国感染控制杂志

Objective To investigate antimicrobial susceptibility of Klebsiella pneumoniae (K .pneumoniae )to aminoglycosides and detection of 16S rRNA methylase genes in K .pneumoniae .Methods Ninety-six non-repetitive clinical K .pneumoniae isolates were collected from Xiangya hospital of Central South University from January to Ju-ly 2009,minimal inhibitory concentrations (MICs)of gentamycin,amikacin and tobramycin were determined by agar dilution method ;genotype of 16S rRNA methylase genes (armA,rmtA,rmtB ,rmtC,rmtD ,npmA)were detec-ted by polymerase chain reaction(PCR).Results MIC50 of amikacin,gentamycin and tobramycin was 256μg/mL, 512μg/mL and 512μg/mL respectively;and MIC90 were all>512μg/mL;antimicrobial resistance rate was 21 .88%, 63.54%,and 41 ,67% respectively.68 isolates (70.83%)were resistant to at least one kind of antimicrobial agent, 21 isolates(21 .88%)were resistant to three kinds of antimicrobial agents.22 isolates(22.92%)carried armA,but rmtA,rmtB ,rmtC,rmtD and npmA were not detected;of 22 isolates harboring armA 16S rRNA methylase genes, 17(77.27%)were highly resistant to gentamicin,amikacin and tobramycin,the homology of armA positive isolate and armA (FJ410928.1 )was 100%.Conclusion armA 16S rRNA methylase gene harbored in K .pneumoniae plays an important role in aminoglycoside resistance.

Analysis of the genetic context of high level aminoglycosides resistance gene 16S rRNA methylase gene armA / 国际检验医学杂志

Objective To analysis the genetic context of armA gene identified in different species .Methods BLASTN searches were used to analysis sequences harboring armA submitted to the GenBank database .Results armA gene was mostly found down-stream of ISCR1 element and was most commonly detected in Enterobacter spp ..Its flanking sequences were highly conserved in Enterobacter spp ..Conclusion The genetic context of armA gene reported in different species scattered in different geometrical lo-cations share some common characteristics .

The Genetic Characteristics of Multidrug-resistant Acinetobacter baumannii Coproducing 16S rRNA Methylase armA and Carbapenemase OXA-23

Acinetobacter baumannii is a gram-negative organism reported worldwide as a cause of health-care associated infections. Due to its increasing drug resistance, several studies on coproduction of armA and carbapenemase in South Korea and other parts of the world were reported, which can pose significant therapeutic threat. The aim of this study was to investigate genetic characteristics of multidrug-resistant A. baumannii coproducing armA and carbapenemase and its epidemiological relatedness. Forty-five multidrug resistant (MDR) A. baumannii clinical isolates were collected. Antimicrobial susceptibility was determined by agar dilution, Etest and VITEK 2 system. The presence of 16S rRNA methylase and carbapenemase were analyzed by polymerase chain reaction (PCR) and sequencing. Repetitive element palindromic (REP)-PCR was also performed for epidemiologic investigation. All of A. baumannii isolates harbored blaOXA-51 -like gene and 10 isolates showed an upstream ISAba1. 36 isolates (80%) showed amplification of OXA-23, all of which except one had an upstream ISAba1. 16S rRNA methylase armA was found in 44 isolates with high level resistance to aminoglycosides. The rate of coproduction was found in 36 isolates (80%). All isolates showed dominant two patterns in REP-PCR profile. The prevalence of MDR A. baumannii coproducing OXA-23 and armA was high, which the rate of blaOXA-23 coproduction was also high.

Distribution and resistance mechanism of 16S rRNA methylase in ESBL-producing Klebsiella pneumoniae / 中华微生物学和免疫学杂志

ObjectiveTo investigate the prevalence and distribution of 16S rRNA methylase gene and research the relationship with drug resistant spectrum.And preliminary explore its role in molecular epidemiology analysis.MethodsCollected 69 clinical isolates of non repetitive ESBL-producing Klebsiella pneumoniae in our hospital from Mar to Sep 2010.Detection 16S rRNA methylation enzyme gene by PCR,and analyze ESBL genetype and integron gene of the positive strains.All PCR products were sequenced for determination.Plasmid conjugation test and plasmid elimination method to determine dissemination of 16S rRNA methylase gene.Then we used ERIC-PCR genotyping technology for the establishment of DNA fingerprinting.ResultsIn sixty-nine strains,twenty isolates were rmtB positive (28.9％),two isolates were armA positive,and two strains coproduce rmtB and armA.All positive isolates carried the CTX-M gene,detemined by sequencing,14 strains of CTX-M-14 gene,6 strains of CTX-M-15 gene,14 strains carried TEM1 gene,8 strains carried SHY gene,sequencing showed that 5 strains of SHV-12 gene,3 strains of SHV-11 gene,3 strains carried OXA-10 gene,3 strains carried VBE-1 gene.In addition,the intl gene was found in 12 isolates of 20 rmtB positive strains.All the intl gene positive strains were divided into five kinds gene cassettes,which contained drfA25,drfA1,drfA12,aadA1,aadA2,sat and blaVEB-1 genes.Respectivily,16S rRNA methylase gene positive strains were divided into five genetypes using ERIC-PCR technology.A genetype was the advantage popular clones.Conjugative plasmid and elimination test found that rmtB gene was located in a plasmid in KP5 and KP16 isolates with A genetype,and can disseminate by conjugation.ConclusionA high prevalence of 16S rRNA methylase gene-rmtB was found among clinical ESBL-producing K.pneumoniae isolates in our hospital,which could lead to resistant to almost all aminoglycoside at a high level.Both horizontal gene transfer and clonal spread were responsible for the dissemination of the rmtB gene.In addition,K.pneumoniae co-producing ESBLs,16S rRNA methylation enzymes and class Ⅰ integron existed and were spreading.

Aminoglycoside-resistant genes in Klebsiella pneumoniae / 中华临床感染病杂志

ObjectiveTo investigate the prevalence of 16S rRNA methylase genes, aminoglycoside modifying enzymes (AMEs) genes and small multidrug resistance efflux pump gene smr-2 in Klebsiella pneumoniae. MethodsTotally 138 Klebsiella pneumoniae isolates were collected in the First Affiliated Hospital, college of medicine, Zhejiang University from January 2007 to December 2009. Polymerase chain reaction (PCR) and DNA sequencing were performed to screen the presence of six 16S rRNA methylase genes ( rmtA, rmtB, rmtC, rmtD, armA and npmA), seven AMEs genes[aac ( 3 )- Ⅰ , aac ( 3 )- Ⅱ,aac(6′)- Ⅰ b, aac(6′)-Ⅱ, ant(2″)- Ⅰ , ant(3″)- Ⅰ , aph(3′)-Ⅵa]and small multidrug resistance efflux pumps gene (smr-2).Results Thirteen (9. 4％) isolates were found to carry rmtB gene, whereas 87 (63.0％) isolates were found to carry at least one kind of AMEs genes but no smr-2 was detected. The positive rates of aac(3)-Ⅱ, aac(6′)- Ⅰ b, ant (3″)- Ⅰ and ant(2″)- Ⅰ were 40.6％ (56/138), 31.9％ (44/138), 28.3％ (39/138) and 2.2％ (3/138), respectively. All strains harboring rmtB gene carried one to three AMEs genes. Among 44 aac(6′)- Ⅰ b positive strains, 37 (84. 1％ ) were confirmed to carryaac(6′)- Ⅰ b-cr. ConclusionFor Klebsiella pneumoniae, rmtB is the predominant subtype in 16S rRNA methylase genes, accompanying with several AMEs genes.

Co-production of carbapenem-hydrolyzing enzyme KPC-2 and ArmA 16S rRNA methylase in pandrugs resistant Enterobacter cloacae / 中华微生物学和免疫学杂志

Objective To investigate the production of carbapenemase and 16S rRNA methylase in five isolates of pan-drugs resistant E.cloacae recovered in Ruijin hospital.Methods MICs of the five isolates to 10 antibiotics were determined by E test.Six kinds of 16S rRNA methylase genes and a series of β- lactamase genes were amplified by PCR.Shotgun cloning was performed to detect carbapenem resistance determinant.The conjugal transfer of carbapenemase gene and 16S rRNA methylase gene was performed in broth culture with E.coli J53 as the recipient.Pulsed-field gel electrophoresis (PFGE) was carried out to analyse the genotyping.IEF was performed to detect β-1actamases.Southern blot was performed to determine the location of carbapenem resistance determinant.Results The MICs of 10 antibiotics were ＞32 mg/L.Four β-1actamases with pIs of 5.4 ( TEM-1 ),6.7 ( KPC-2 ),8.2 ( SHV-12 ),8.4 (CTX-M-14) were determined.The insertion sequence in the recombinant plasmid was blaKPC-2 flanked by a transposon.blaKPC-2 was located on a large non-conjugative plasmid whereas armA was located on an other conjugative plasmid.PFGE patterns of 5 isolates were identical.Conclusion KPC-2 was responsible for carbapenem resistance in pandrugs resistant Enterobacter cloacae.There was no relationship between blaKPC-2 and armA.Although pandrug resistant Enterobacteriaceae remain rare,the emergence of this group of organism merits monitoring.

aac(6')-Ⅰ b gene in drug-resistant strains of Pseudomonas aeruginosa / 中华临床感染病杂志

Objective To investigate the distribution of aminoglycoside modifying enzyme genes (AMEs) and 16S rRNA methylase genes in drug-resistant strains of Pseudomonas aeruginosa. Methods Twenty strains of drug-resistant Pseudomonas aeruginosa were isolated from sputum and wound secretion samples collected from the First People's Hospital of Huai' an in Jiangsu province. Eight AMEs [aac(3)-Ⅰ , aac(3)-Ⅱ, aac(6')-Ⅰb,aac(6')-Ⅱ, ant{2)-Ⅰ ,ant(3)-Ⅰ , ant(4')-Ⅰ , aph(3')-Ⅱb] and 6 16S rRNA methylase genes (armA, rmtA, rmtB, rmtC, rmtD, npmA) were analyzed by PCR and verified by DNA sequencing. Results Out of 20 strains of Pseudomonas aeruginosa, aac(6')-Ⅱ was positive in 8 strains (40.0% ) , ant2- Ⅰ in 8 strains (40.0% ) , aac(3)-Ⅱ in 5 strains (25.0% ) , aac(6')- Ⅰ b in 2 strains (10.0% ) and rmtB in 1 strains (5.0% ) , respectively. The rest 9 genes were not detected. Among 2 strains harboring aac(6')- Ⅰ b, DNA sequencing confirmed that 1 was aac(6')- Ⅰ b (the clssical type) and another was aac(6')- Ⅰ b-cr. Conclusion Gene aac(6')- Ⅰ b-cr exists in drug-resistant Pseudomonas aeruginosa, and it has modifying effect on both aminoglycosides and quinolones.

16S rRNA methylase genes in ESBLs-producing Enterobacteriaceae / 中华临床感染病杂志

Objective To investigate the prevalence and dissemination mechanism of 16S rRNA methylase genes in extended-spectrum beta-lactamases(ESBLs)-producing Enterobacteriaceae in China.Methods PCR amplification and DNA sequencing were used for screening and identifing 16S rRNA methylase genes and ESBLs genes.Minimal inhibitory concentrations(MICs)of the antimicrobial agents were detected by Etest.Conjugation and plasmid extract were performed to study dissemination mechanism of 16S rRNA methylase genes and ESBLs genes.Results Only one strain.Klebsiella oxytoca strain ZJ157 was screened as positive for armA gene from 447 ESBLs-producing isolates,which also contained CTX-M-15 and TEM-1 genes.It was resistant to aminoglycesides,ciprofloxacin,and most β-lactams,except carbapenems,polymyxin E and tigecyeline.Resistance to amikacin and β-lactams was transferred to a recipient Escherichia coli 600 by conjugation experiment.arntA.CTX-M-15 and TEM-1 genes were detected in the transconjugant.A plasmid about 55 kb was extracted from Klebsiella oxytoca ZJl57 and the transconjugant.Conclusions A 16S rRNA methylase gene armA was detected in an isolate of Klebsiella oxytoca.armA,CTX-M-15 and TEM-1 genes can be co-transferred in the same plasmid leading to multi-drug resistance.

Study on carbapenemase and 16S rRNA methylase of imipenem-resistant Acinetobacter baumannii / 中华流行病学杂志

Objective To investigate the prevalence of 16S rRNA methylases gene in imipenem-resistant Acinetobacter baumannii isolates from China. Methods A total of 342 imipenem-resistant A.baumannii isolates were collected between December 2004 and December 2005, from 25 hospitals of China. Agar dilution was used to determinate the minimal inhibitory concentration (MIC) of these isolates. The homology of these isolates was analyzed by pulse-field gel electrophoresis (PFGE). Several 16S rRNA methylase genes and carbapenemase genes were detected by PCR-based assays and PCR products were sequenced. Results The rates of resistance to ampicillin-sulbactam, cefoperazone-sulbactam, tobramycin, and minocycline were 68.0%, 54.2%, 87.4%, and 75.9%, respectively. The rate of resistance to polymyxin E was 10.8%, the lowest among the tested agents. The rates of resistance to all other tested antimicrobial agents were more than 90%. The A.baumannii isolates belonged to 29 distinct clones. Among them, 6 clones were dominant, consisting of 303 isolates in total. All isolates contained the blaOXA-51-1ike gene (blaOXA-66) and 322 isolates contained the blaOXA-23-1ike gene. PCR with the ISAbal-OXA-23-like primers generated a PCR product in 314 isolates, and PCR with the ISAbal-OXA-51-1ike primers generated a PCR product in 13 strains. 221 armA-positive isolates were identified. Conclusion Most of the imipenem-resistant A.baumannii contained blaOXA-23, with ISAbal upstream of the gene. 16S rRNA methylase gene armA was widely distributed in these isolates. The results suggested that the spread of clones played an important role in the outbreak of imipenem-resistant A. baumannii in China.

The prevalence of 16S rRNA methylase gene armA and drug resistance in Acinetobacter baumannii / 中华微生物学和免疫学杂志

Objective To investigate the prevalence of 16S rRNA methylase gene armA and to analyze their effect on the drug resistance in multi drug-resistant strains of Acinetobacter baumannii . Methods A total of 72 Acinetobacter baumannii isolates were collected from the Second Xiangya Hospital from Jan. 2008 to Dec. 2008. The size of inhibitory zone of these strains to gentamycin, tobramycin and amikacin were determinate using Kirby-Bauer( K-B) method. The 16S rRNA methylase genes armA were detected by PCR. PCR products were purified and sequenced. Then we used randomly amplified polymorphic DNA method (RAPD) genotyping technology for the establishment of DNA fingerprinting. In addition, we compared drug sensitivity test with RAPD technology. Results Twenty isolates of 72 strains were armA positive and the resistance rates of the strains with armA gene to gentamycin, tobramycin, amikacin were 90.0% , 90.0% and 90. 0% , respectivily. armA positive stains were divided into 7 types using RAPD technology. A genotype was the advantage type. Conclusion The study showed that 16S rRNA methylases gene armA was prevalent in Acinetobacter baumannii which could lead to resistant to almost all aminoglycosides at a high level. And the main form of armA gene prevalence in our hospital was the spread of the same clone strain inside and outside of clinic department.

Aminoglycoside-modifying Enzyme and 16S rRNA Methylase Gene Expressions in Acinetobacter baumannii / 中华医院感染学杂志

OBJECTIVE To understand aminoglycoside-modifying enzyme and 16S rRNA methylase gene expressions in Acinetobacter baumannii in Xinjiang region.METHODS 20 A.baumannii strains were isolated and test the anti-bacterial drug sensitivity.PCR methods were used to test aminoglycoside-modifying enzyme and 16S rRNA methylase gene.RESULTS From thirteen A.baumannii strains detected the aminoglycoside-modifying enzyme genes,the detection rate was 65%;in which aac(3)-Ⅰ gene was positive in 4 strains(20%),aac(3)-Ⅱ gene in 8 strains(40%),aac(6′)-Ⅰad gene in 4 strain(20%),ant(3″)-Ⅰ gene in 4 strain(20%),and ant(2″)-Ⅰ gene was positive in 1 strain(5%).The aac(6′)-Ⅰ b and aac(6′)-Ⅱ gens and were not detected;the 16S rRNA gene methylation was negative.CONCLUSIONS There are aminoglycoside-modifying enzyme genes existing in A.baumannii,no 16S rRNA methylase gene was detected in Xinjiang region.

Drug-resistant Genes Associated with Aminoglycosides in Acinetobacter baumannii Isolated from ICU / 中华医院感染学杂志

OBJECTIVE To investigate the antibiotic resistance of multi-resistant Acinetobacter baumannii(ABA) and distribution of aminoglycoside-modifying enzymes and 16S rRNA methylase genes in ICU in Yinzhou People′s Hospital in Ningbo. METHODS The samples of 20 ABA isolates were collected from Oct 2007 to Jul 2008 in ICU.K-B method was used to determine the sensitivity to 32 antibacterials and the aminoglycoside-modifying enzymes and 16S rRNA methylase genes were analyzed by polymerase chain reaction(PCR). RESULTS From 20 ABA isolates,8 strains carried aminoglycoside-modifying enzymes genes aac(3)-Ⅰ,aac(3)-Ⅱ,aac(6′)-Ⅰb,and ant(3″)-Ⅰ,their positive rate was 10%,15%,30% and 25%,respectively;no strain carried 16S rRNA methylase genes. CONCLUSIONS The antibiotics resistance of A.baumannii is very serious in Yinzhou People′s Hospital in Ningbo.Aminoglycoside-modifying enzymes genes aac(3)-Ⅰ,aac(3)-Ⅱ,aac(6′)-Ⅰb and ant(3″)-Ⅰ exist in multi-resistant A.baumannii widely.They would be the main causes of high drug-resistantce to aminoglycosides.

16S rRNA Methylase Gene and Aminoglycoside-modifying Enzyme Genes in Pseudomonas aeruginosa Isolated from Burned Patients / 中华医院感染学杂志

OBJECTIVE To investigate the 16S rRNA methylase gene and aminoglycoside-modifying enzyme genes in Pseudomonas aeruginosa isolated from burned patients. METHODS GNS-448 and K-B tests were performed to detect the susceptibility to 19 kinds of antimicrobial agents against these strains. 16S rRNA methylase gene and aminoglycoside-modifying enzyme genes were amplified by polymerase chain reaction (PCR) and verified by DNA sequencing. RESULTS The 32 isolated strains were all resistant to ampicillin,cefuroxime,cefoxitin,SMZ-TMP,The sensitive rates to amikacin and gentamicin were 68% and 46.9%,respectively. The resistant rates to imipenem and meropenem were 68.8% and 59.4%,respectively. The 16S rRNA methylase gene and aminoglycoside-modifying enzyme genes including aac(6')-Ⅰb,aac(6')-Ⅱ,ant(3″)-Ⅰ,ant(2″)-Ⅰ and rmtB were found and positive rates were 9.4%,3.1%,28.1%,25.0% and 3.1%,respectively. A novel subtype of aac(6')-Ⅰb was reported firstly. CONCLUSIONS There are high positive percentage of 16S rRNA methylase gene and aminoglycoside-modifying enzyme genes in P. aeruginosa isolated from burned patients. P. aeruginosa resistance to aminoglycoside relates to the existence of 16S rRNA methylase gene and aminoglycoside-modifying enzyme genes.

Detection of 16S rRNA methylase gene in gram-negative bacteria / 中国感染与化疗杂志

Objective To investigate the prevalence of 16S rRNA methylase genes, armA and rmtB, which mediate high level aminoglycoside resistance, in gram-negative bacteria isolated from 2 hospitals in Dalian and study the mechanism of aminoglycoside resistance.Methods A total of 134 amikacin-resistant clinical isolates of gram-negative bacteria were collected. Two 16S rRNA methylase genes, armA and rmtB, were identified by PCR-based assays. PCR products were extracted for DNA sequencing analysis. armA and rmtB gene mapping were conducted by plasmid extraction, conjugation and transformation. The MICs of amikacin, gentamicin and tobramycin were determined for the positive isolates, transconjugants and the resultant strain of transformation using agar dilution technique. Results Overall armA was identified in 21 strains of Acinetobacter baumannii, rmtB in 5 strains of Escherichia coli and 5 strains of Klebsiella pneumoniae. Plasmid extraction and conjugation experiments were only successful for rmtB-positive isolates. Transconjugant and DH5a (pMDarmA) exhibited high-level resistance to aminoglycosides.Conclusions The 16S rRNA methylase genes, armA and rmtB are identified in Dalian. armA gene is identified in A. baumannii. rmtB gene is located on the plasmid of E. coli and K. pneumoniae. armA and rmtB can induce high-level resistance to aminoglycosides.

Emergence of 16S rRNA methylase gene rmtB in Klebsiella pneumoniae isolates from the inpatients in Zhejiang province, China / 中华流行病学杂志

Objective To investigate the presence and genetic background of 16S rRNA methylase gene and Aminoglycoside modifying enzymes(AMEs) genes in Klebsiella pneumoniae isolated from the People's Liberation Army 98th HospitaI,Huzhou district,Zhejiang province,China.Methods 25 strains of Klebsiella pneumoniae were isolated from the inpatienta between September,2005 and April,2006.6 kinds of 16S rRNA methylase gene (including armA,rmtA,rmtB,rmtC,rmtD and npmA ),6 kinds of AMEs genes [ including aac (3)-Ⅰ,sac (3)-Ⅱ,sac (6')-Ⅰ,aac (6')-Ⅱ,ant (3")-Ⅰand ant(2")-Ⅰ],intI1,intI2,intI3,mercuric reductase gene merA (merA gene were the collective genetic markers of transposona of Tn21 and Tn501 ) and tnpA ( tnpA gene were the collective genetic markers of transposons of Tn1,Tn2,Tn3 and Tn1000) were analyzed by PCR and verificated by DNA sequencing.Results In 25 strains of Klebsiella pneumoniae,the positive rate of genes of rmtB,sac (3)-Ⅱ,sac (6')-Ⅰ,ant(3")-Ⅰ and intI1 were 60.0%(15/25),4.0%(1/25),48.0%(12/25),60.0%(15/25) and 96.0%(24/25),respectively.The rest 12 kinds of genes were all tested negative.The total positive rate of 6 kinds of AMEs gene was 84.0%(21/25).Conclusion There were very high positive rate on both genes of rmtB and AMEs genotypes in Klebsiella pneumoniae isolated from inpatients,and this was the first report of the emergence of 16S rRNA methylase gene rmtB in Klebsiella pneumoniae identified in Zhejiang province,China.