OBJECTIVETo investigate the resistance genes and antibiotic resistance patterns against beta-lactams in Pseudomonas aeruginosa prevalent in burn ward.

METHODSK-B method was performed to test bacterial resistance patterns against 9 species of beta-lactams in Pseudomonas aeruginosa isolated from wounds and dressings of the patient in burn wards. Seven species of resistance genes against beta-lactams were detected with PCR. Tazobactam-inhibited piperacillin resistance test was performed to study whether the above strains produce extended spectrum beta-lactams.

RESULTSAll 12 strains of bacteria with resistance genes detected were resistant to penicillin and cephalosporins (100%), among them 11 were resistant to all antibiotics. Tazobactam-inhibited piperacillin resistance test demonstrated that all strains with resistance genes were ESBLs.

CONCLUSIONHigh incidence of beta-lactams resistance genes is found in Pseudomonas aeruginosa isolated from burn ward, and they have close relationship with the occurrence of multiple drug-resistance.

Burn Units ; Burns ; microbiology ; Genes, Bacterial ; Humans ; Pseudomonas aeruginosa ; drug effects ; genetics ; isolation & purification ; beta-Lactam Resistance ; genetics

OBJECTIVETo investigate drug-resistant genes associated with beta-lactams and aminoglycosides in clinically isolated Pseudomonas aeruginosa.

METHODSTwenty strains of Pseudomonas aeruginosa were isolated from wound excretion of hospitalized burn patients. The strains resistant to 14 antibiotics were selected for detection of 16 kind of drug-resistant genes (TEM, SHV, OXA-10 cluster, PER, VEB, GES, CARB, CTX-M- I, IMP, VIM, SPM, GIM, DHA, MOX, FOX, oprD2) and 6 kind of aminoglycoside modification genes (aac(3)- I, aac(3)-II, aac(6')-I, aac(6')-II, ant (3")- I , ant(2")- I) in them by PCR.

RESULTSAmong the 20 strains resistant to beta-lactam , all of them were TEM and GES positive (100%), oprD2 gene depletion in 5 strains (25%). All other genes were negative. Among aminoglycoside resistant genes, 20 strains were aac (6') - I positive (100%), 7 were ant (2") - I positive (35%), and negative for other stains.

CONCLUSIONThere were very high existence rates of TEM, GES and aac (6')- I genes in Pseudomonas aeruginosa isolated from clinical burn patients. The fact that GES-5 gene has also been detected in Pseudomonas aeruginosa, suggesting this organism is highly drug resistant in our burn unit.

Aminoglycosides ; pharmacology ; Drug Resistance, Multiple, Bacterial ; genetics ; Humans ; Pseudomonas aeruginosa ; drug effects ; genetics ; isolation & purification ; beta-Lactam Resistance ; genetics ; beta-Lactams ; pharmacology

The gene and the amino acid sequence of the structural and regulatory region of the Pseudomonas aeruginosa with different resistance patterns were analyzed. Six strains with different resistance patterns were selected and the AmpC beta-lactamase was identified. The objective gene fragment was amplified by colonies PCR. The sequences of the PCR-products were analyzed. The DNA sequence of the structural gene ampC and the regulatory genes ampR, ampD and ampE was detected. The 6 strains and the wild-type Pseudomonas aeruginosa are highly homogeneous in structural and regulatory region. Some new mutant points were found.
Bacterial Proteins/*genetics ; DNA, Bacterial/genetics ; Point Mutation/*genetics ; Pseudomonas aeruginosa/enzymology ; Pseudomonas aeruginosa/*genetics ; Sequence Analysis, DNA ; beta-Lactam Resistance/*genetics ; beta-Lactamases/*genetics

OBJECTIVETo investigate the molecular mechanism of carbapenem resistance in the clinical isolates of Acinetobacter baumannii from Xi'an and their profile of carbapenemase production.

METHODSA total of 146 Acinetobacter baumannii strains were isolated from 6 general hospitals in Xi'an. Antimicrobial susceptibility test was performed for all the strains, followed by detection of imipenem resistance using E-test for metallo-beta-lactamase (MBL) and NaCl inhibition test for OXA type carbapenemase. Bla(OXA-23)and bla(OXA-58) were amplified by PCR, and the positive products were sequenced.

RESULTSFrom the collected strains, 15 non-repetitive imipenem-resistant Acinetobacter baumannii strains were identified, among which 14 yielded negative results in E-test for MBL production. All the resistant strains showed increased sensitivity to imipenem after NaCl inhibition, suggesting the presence of carbapenemase production. Eleven of the strains harbored OXA -23 type gene and 1 harbored OXA -58 type gene. The concordance rate of the results by NaCl inhibition test and PCR was 85.7%.

CONCLUSIONSProduction of OXA-type carbapenemase is the most important reason for carbapenem resistance in Acinetobacter baumannii in Xi'an. The OXA-58 type gene is a novel carbapenemase genotype in China. NaCl inhibition test is a convenient and cost-effective method for detecting carbapenemase in Acinetobacter baumannii.

Acinetobacter baumannii ; drug effects ; genetics ; isolation & purification ; Anti-Bacterial Agents ; pharmacology ; Bacterial Proteins ; genetics ; China ; Humans ; Imipenem ; pharmacology ; beta-Lactam Resistance ; beta-Lactamases ; genetics

To study the resistant mechanism and clinical significance of pseudomonas aeruginosa to beta-lactam antibiotics, the outer membrane permeability rate of 30 P. aeruginosa strains to 5 beta-lactam antibiotics was measured and their production of beta-lactamase and the beta-lactamase genes they carried detected. Furthermore, the relationship between the permeability, beta-lactamase and the clinical effects of beta-lactam antibiotics was observed. By using 14C-penicillin and liquid-scintillant isotope assay, the affinity of penicillin binding proteins (PBPS) was measured and their roles in the resistant mechanism studied. It was revealed that the permeability rate was higher in sensitive strains than in resistant ones (P < 0.05). All strains harbored 1-4 beta-lactamase genes and produced beta-lactamase. Higher permeability rate and higher degree of stability to beta-lactamase indicated better clinical therapeutic effects. The affinity of PBPs changed little without regard to the permeability and beta-lactamase. These results suggested that the permeability of outer membrane and beta-lactamase, but not PBPs, played important roles in the resistant mechanism of P. aeruginosa to beta-lactam antibiotics and affected the clinical therapeutic effectiveness of some patients.
Anti-Bacterial Agents/*pharmacology ; Bacterial Outer Membrane Proteins/metabolism ; Microbial Sensitivity Tests ; Permeability ; Pseudomonas aeruginosa/*drug effects ; beta-Lactam Resistance/*genetics ; beta-Lactamases/metabolism ; beta-Lactams/*pharmacology

To study the resistant mechanism and clinical significance of pseudomonas aeruginosa to beta-lactam antibiotics, the outer membrane permeability rate of 30 P. aeruginosa strains to 5 beta-lactam antibiotics was measured and their production of beta-lactamase and the beta-lactamase genes they carried detected. Furthermore, the relationship between the permeability, beta-lactamase and the clinical effects of beta-lactam antibiotics was observed. By using 14C-penicillin and liquid-scintillant isotope assay, the affinity of penicillin binding proteins (PBPS) was measured and their roles in the resistant mechanism studied. It was revealed that the permeability rate was higher in sensitive strains than in resistant ones (P < 0.05). All strains harbored 1-4 beta-lactamase genes and produced beta-lactamase. Higher permeability rate and higher degree of stability to beta-lactamase indicated better clinical therapeutic effects. The affinity of PBPs changed little without regard to the permeability and beta-lactamase. These results suggested that the permeability of outer membrane and beta-lactamase, but not PBPs, played important roles in the resistant mechanism of P. aeruginosa to beta-lactam antibiotics and affected the clinical therapeutic effectiveness of some patients.
Anti-Bacterial Agents ; pharmacology ; Bacterial Outer Membrane Proteins ; metabolism ; Humans ; Microbial Sensitivity Tests ; Permeability ; Pseudomonas aeruginosa ; drug effects ; beta-Lactam Resistance ; genetics ; beta-Lactamases ; metabolism ; beta-Lactams ; pharmacology

OBJECTIVETo investigate the role of AdeABC efflux pump in carbapenems resistance of Acinetobacter baumannii in light of the phenotype and genetype of the efflux pump.

METHODSThe phenotype of the efflux pump was detected in 138 clinical isolates of A.baumannii using the efflux pump inhibitor carbonyl cyanide 3-chlorophenylhydrazone (CCCP). The mRNA expression of pump-encoding gene adeB in the strains was detected using quantitative real-time RT-PCR.

RESULTSOf the 138 strains, 28 showed positivities for AdeABC efflux pump identified by Mueller-Hinton Broth with CCCP. Of the 39 strains resistant to meropenem, 15 (38.4%) showed positive results in CCCP assay, a rate significantly higher than that among the 99 sensitive strains (13.1%, 13/99) (X(2)=12.477(b), P=0.01). The mRNA expression of efflux pump-encoding gene adeB was detected by real-time RT-PCR at a level of 0.899∓∓1.172 in meropenem-sensitive strains, significantly lower than the level of 21.101∓∓21.443 in meropenem-resistant strains (t=4.403, P=0.000).

CONCLUSIONSEfflux plays a role in carbapenems resistance in the clinical isolates of A. baumannii. The AdeABC efflux pump may be an important factor in reducing carbapenems sensitivity in A. baumannii.

Acinetobacter baumannii ; drug effects ; isolation & purification ; Bacterial Proteins ; genetics ; metabolism ; Carbapenems ; pharmacology ; Humans ; Membrane Transport Proteins ; genetics ; metabolism ; beta-Lactam Resistance ; genetics

OBJECTIVETo investigate the beta-lactamase TEM gene of isolated Streptococcus pneumoniae (Sp) in Suzhou area.

METHODSTwenty-three strains of Sp were collected from respiratory tract secretions of children with respiratory diseases in Nov 2002 to Apr 2003 at Children's Hospital of Suzhou University (reference strain ATCC49619) to build TEM polymerase chain reaction (PCR) system (reference strain E. coli. 9-j53R1 with TEM gene) TEM gene of 23 strains was detected to comparo the sequences with published TEM gene sequences in GenBank for analyzing TEM gene model.

RESULTSTwenty-one strains had TEM gene with a positive rate of 91.3% (21/23). TEM-129 gene were confirmed from No.17 (SR017, penicillin resistance) TEM sequence. New discovered TEM-129 sequence had a modification (ATG[M]-->ATA[I]) at No.182 code and published (GenBank: www.ncbi.nlm.nih.gov/nucleotide, AY452662). TEM-1 genes were confirmed from other TEM sequences. New discovered TEM-1 gene of isolated Sp had been published (GenBank: www.ncbi.nlm.nih.gov/nucleotide, AY392531) too.

CONCLUSIONIsolated Sp had TEM gene (TEM-129, EM-1 genotype) with a positive rate of 91.3%. The result enriched the understanding of isolated Sp with penicillin resistance.

Base Sequence ; China ; epidemiology ; Drug Resistance, Bacterial ; genetics ; Female ; Genes, Bacterial ; genetics ; Humans ; Male ; Molecular Epidemiology ; Molecular Sequence Data ; Pneumonia, Bacterial ; epidemiology ; genetics ; microbiology ; Point Mutation ; Streptococcus pneumoniae ; enzymology ; genetics ; beta-Lactam Resistance ; beta-Lactamases ; genetics

OBJECTIVETo analyze the distribution of Enterobacteriaceae isolated from West China Hospital, investigate the antibiotic resistance profile of Enterobacteriaceae with decreased susceptibility to carbapenems and explore the molecular mechanism.

METHODSForty-five Enterobacteriaceae strains resistant or with reduced susceptibility to carbapenems were isolated from patients in West China Hospital. The antimicrobial susceptibility and carbapenemase-producing phenotypes of the bacteria were examined and specific PCR were performed to determine the molecular mechanism.

RESULTSOf the 45 isolates, 17, 21 and 36 were resistant or intermediate strains to imipenem, meropenem and ertapenem, respectively. The majority of these isolates showed resistance to cephalosporins. The modified Hodge test resulted in the highest positivity rate (77.8%), followed by EDTA disc test (57.8%) and PBA disc test (22.2%). BlaTEM, blaSHV and blaCTX-M were detected in 60.0%, 53.3% and 15.6% of these strains with reduced susceptibility. The rate of strains carrying 2 or more genes was 44.4%, and the detection rate of blaIMP was 48.9%. BlaKPC was identified in 4 (8.9%) high-level resistant strains and confirmed to locate on the plasmid.

CONCLUSIONProduction of carbapenemase contributes to reduced susceptibility of carbapenems in Enterobacteriaceae. The presence of blaKPC, MBL and ESBL, and their possible combinations can be the main factor contributing to carbapenem resistance or reduced susceptibility in Enterobacteriaceae. The KPC-2 carbapenemase gene located on the plasmids we found in this study can cause potential horizontal transmission across strains.

Anti-Bacterial Agents ; pharmacology ; Bacterial Proteins ; genetics ; metabolism ; Carbapenems ; pharmacology ; Cephalosporins ; pharmacology ; Enterobacteriaceae ; drug effects ; enzymology ; genetics ; Gene Amplification ; Imipenem ; pharmacology ; Microbial Sensitivity Tests ; Polymerase Chain Reaction ; Thienamycins ; pharmacology ; beta-Lactam Resistance ; beta-Lactamases ; genetics ; metabolism ; beta-Lactams ; pharmacology

OBJECTIVETo observe the effect of amino acid substitution in conserved sequence of penicillin-binding protein (PBP) 1A, 2B, 2X on antimicrobial activity of beta-lactams against Streptococcus pneumoniae (SP).

METHODMinimal inhibitory concentration (MIC) of 6 beta-lactams was determined by the E-test in 59 SP strains. The penicillin-binding protein genes pbp1a, 2b, 2x in every SP strain were amplified by nested-polymerase chain reaction (nPCR), then the PCR products were sequenced using automatic genetic analyzer directly. To analyze the amino acid substitutions, the DNA sequences were converted to protein sequences and aligned by Clustalx software. According to amino acid substitution in conserved sequence of PBP2B, 3 phenotypes were observed, including: PBP2B phenotype I (no amino acid substitution); PBP2B phenotype II (Glutamine 432-->Leucine and/or Threonine 445/451-->Alanine/Serine, Glutamic 481-->Glycine, 1 strain had proline insertion between residues 431/432); PBP2B phenotype III (Alanine 624-->Glycine with the addition of phenotype II). According to amino acid substitution in conserved sequence of PBP1A, 3 phenotypes were observed, including: PBP1A phenotype I (no amino acid substitution); PBP1A phenotype II (Threonine 574-->Asparagine, Serine 575-->Threonine, Glutamine 576-->Glycine, Phenylalanine 577-->Tyrosine, 574TSQF-->NTGY); PBP1A III (Threonine 371-->Alanine/Serine, Proline 432-->Threonine with the addition of 574TSQF-->NTGY). According to amino acid substitution in conserved sequence of PBP2X, 4 phenotypes were observed, including: PBP2X phenotype I (no amino acid substitution); PBP2X phenotype II (Histidine 394-->Leucine or Threonine 338-->Alanine); PBP2X phenotype III (Threonine 338-->Alanine, Isoleucine 371-->Threonine, Arginine 384-->Glycine and Leucine 546-->Valine); PBP2X phenotype IV (Methionine 339-->Phenylalanine, Methionine 400-->Threonine with the addition of PBP2X phenotype III).

RESULTAmong 59 SP strains antibacterial activities distribution (sensitive strains, intermediate strains and resistant strains) of 6 beta-lactams were penicillin (12, 29, 18); amoxicillin(49, 9, 1); cefuroxime (16, 16, 27); ceftriaxone (47, 1, 11); cefotaxime (47, 3, 9); imipenem (49, 10, 0). beta-lactam antibiotics insensitive strains (intermediate + resistant strain) in PBP2B phenotype III, PBP1A phenotype III, PBP2X phenotype III and IV were significantly increased, the MIC(50) of these strains were significantly higher than that of the others.

CONCLUSIONThe amino acid substitutions in or vicinal conserved sequence of PBP of SP increase MIC for beta-lactam antibiotics.

Amino Acid Substitution ; Aminoacyltransferases ; genetics ; Anti-Bacterial Agents ; pharmacology ; Bacterial Proteins ; genetics ; Microbial Sensitivity Tests ; Penicillin-Binding Proteins ; genetics ; Peptidyl Transferases ; genetics ; Streptococcus pneumoniae ; drug effects ; beta-Lactam Resistance ; genetics ; beta-Lactams ; pharmacology