AIMNineteen compounds related to salicylic acid were evaluated for their in vitro activity of inhibiting beta-lactamase isolated from a resistant strain of Pseudomonas aeruginosa, and their structure-activity relationships were examined.

METHODSNitrocefin method was used.

RESULTSThe 50% inhibitory concentration (IC50) of salicylic acid inhibiting beta-lactamase was 22 mmol x L(-1); four analogs had IC50 lower than that of salicylic acid; fifteen analogs had IC50 higher than that of salicylic acid.

CONCLUSIONExamination of the structure-activity relationships of the compounds revealed that carboxyl group and adjoining hydroxyl group were active group, and replacement of adjoining hydroxyl by carboxyl increased activity nearly 4-fold. Moreover, addition of a sulfonic group at C-5 and nitro group at C-3, 5 of benzenoic ring of salicylic acid resulted in a 2-fold to 3-fold increase in activity, addition of a amino group at C-5 of benzenoic ring of salicylic acid decreased activity, add addition of -Cl or -F at C-2,4 position of benzenoic ring of benzoic acid did not show activity.

Anti-Bacterial Agents ; chemical synthesis ; chemistry ; pharmacology ; Cephalosporins ; metabolism ; Inhibitory Concentration 50 ; Pseudomonas aeruginosa ; enzymology ; Salicylates ; chemical synthesis ; chemistry ; pharmacology ; Structure-Activity Relationship ; beta-Lactamases ; isolation & purification ; metabolism

BACKGROUND: We evaluated the combined use of the modified Hodge test (MHT) and carbapenemase inhibition test (CIT) using phenylboronic acid (PBA) and EDTA to detect carbapenemase-producing Enterobacteriaceae (CPE) and metallo-beta-lactamase (MBL)-producing Pseudomonas spp. METHODS: A total of 49 isolates of CPE (15 Klebsiella pneumoniae carbapenemase [KPC], 5 Guiana extended-spectrum beta-lactamase [GES]-5, 9 New Delhi metallo-beta-lactamase [NDM]-1, 5 Verona integron-encoded metallo-beta-lactamase [VIM]-2, 3 imipenem-hydrolyzing beta-lactamase [IMP], and 12 oxacillinase [OXA]-48-like), 25 isolates of MBL-producing Pseudomonas spp. (14 VIM-2 and 11 IMP), and 35 carbapenemase-negative controls were included. The MHT was performed for all isolates as recommended by the Clinical and Laboratory Standards Institute. Enhanced growth of the indicator strain was measured in mm with a ruler. The CIT was performed by directly dripping PBA and EDTA solutions onto carbapenem disks that were placed on Mueller-Hinton agar plates seeded with the test strain. RESULTS: Considering the results of the MHT with the ertapenem disk in Enterobacteriaceae and Pseudomonas spp., the CIT with the meropenem disk in Enterobacteriaceae, and the imipenem disk in Pseudomonas spp., three combined disk tests, namely MHT-positive plus PBA-positive, EDTA-positive, and MHT-positive plus PBA-negative plus EDTA-negative, had excellent sensitivity and specificity for the detection of KPC- (100% sensitivity and 100% specificity), MBL- (94% sensitivity and 100% specificity), and OXA-48-like-producing isolates (100% sensitivity and 100% specificity), respectively. CONCLUSIONS: Combined use of the MHT and CIT with PBA and EDTA, for the detection of CPE and MBL-producing Pseudomonas spp., is effective in detecting and characterizing carbapenemases in routine laboratories.
Bacterial Proteins/antagonists & inhibitors/*metabolism ; Boronic Acids/chemistry/pharmacology ; Disk Diffusion Antimicrobial Tests/*methods ; Edetic Acid/chemistry/pharmacology ; Enterobacteriaceae/drug effects/*enzymology ; Enterobacteriaceae Infections/diagnosis ; Humans ; Pseudomonas/drug effects/*enzymology ; Pseudomonas Infections/diagnosis ; Sensitivity and Specificity ; beta-Lactamases/chemistry/*metabolism

Gram-negative Enterobacteriaceae with resistance to carbapenem conferred by New Delhi metallo-β-lactamase 1 (NDM-1) are a type of newly discovered antibioticresistant bacteria. The rapid pandemic spread of NDM-1 bacteria worldwide (spreading to India, Pakistan, Europe, America, and Chinese Taiwan) in less than 2 months characterizes these microbes as a potentially major global health problem. The drug resistance of NDM-1 bacteria is largely due to plasmids containing the blaNDM-1 gene shuttling through bacterial populations. The NDM-1 enzyme encoded by the blaNDM-1 gene hydrolyzes β-lactam antibiotics, allowing the bacteria to escape the action of antibiotics. Although the biological functions and structural features of NDM-1 have been proposed according to results from functional and structural investigation of its homologues, the precise molecular characteristics and mechanism of action of NDM-1 have not been clarified. Here, we report the three-dimensional structure of NDM-1 with two catalytic zinc ions in its active site. Biological and mass spectroscopy results revealed that D-captopril can effectively inhibit the enzymatic activity of NDM-1 by binding to its active site with high binding affinity. The unique features concerning the primary sequence and structural conformation of the active site distinguish NDM-1 from other reported metallo-β-lactamases (MBLs) and implicate its role in wide spectrum drug resistance. We also discuss the molecular mechanism of NDM-1 action and its essential role in the pandemic of drug-resistant NDM-1 bacteria. Our results will provide helpful information for future drug discovery targeting drug resistance caused by NDM-1 and related metallo-β-lactamases.
Amino Acid Sequence ; Anti-Bacterial Agents ; metabolism ; Binding Sites ; Captopril ; chemistry ; pharmacology ; Catalytic Domain ; Crystallography, X-Ray ; Drug Resistance, Bacterial ; Enterobacteriaceae ; enzymology ; Molecular Sequence Data ; Sequence Alignment ; Sequence Homology, Amino Acid ; beta-Lactamases ; chemistry ; metabolism

BACKGROUND: Rapid detection of carbapenemase-producing gram-negative bacilli (GNB) is required for optimal treatment of infected patients. We developed and assessed a new disk carbapenemase test (DCT). METHODS: Paper disks containing 0.3 mg of imipenem and bromothymol blue indicator were developed, and the performance of the DCT were evaluated by using 742 strains of GNB with or without carbapenemases. RESULTS: The paper disks were simple to prepare, and the dried disks were stable at -20℃ and at 4℃. The DCT detected 212 of 215 strains (98.6% sensitivity with 95% confidence interval [CI] 96.0-99.5%) of GNB with known class A (KPC and Sme) and class B (NDM, IMP, VIM, and SIM) carbapenemases within 60 min, but failed to detect GES-5 carbapenemase. The DCT also detected all two Escherichia coli isolates with OXA-48, but failed to detect GNB with OXA-232, and other OXA carbapenemases. The DCT showed 100% specificity (95% CI, 99.2-100%) in the test of 448 imipenem-nonsusceptible, but carbapenemase genes not tested, clinical isolates of GNB. CONCLUSIONS: The DCT is simple and can be easily performed, even in small laboratories, for the rapid detection of GNB with KPC, NDM and the majority of IMP, VIM, and SIM carbapenemases.
Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/*metabolism ; Bromthymol Blue/chemistry ; Drug Resistance, Bacterial ; Gram-Negative Bacteria/drug effects/*enzymology ; Imipenem/pharmacology ; Microbial Sensitivity Tests/*methods ; Paper ; beta-Lactamases/*metabolism

To investigate the distribution of the genes of two major metallo-beta-lactamases (MBL; i.e., IMP and VIM) and class 1 integrons (intI) in the clinical imipenem-resistant Pseudomonas aeruginosa, a total of 65 isolates, from a university hospital in Sichuan between December 2004 and April 2005 were screened for MBL genes by PCR using primers specific for bla ( IMP-1 ), bla ( VIM ) and bla ( VIM-2 ) genes. The MBL-positive isolates were further assessed for class 1 integrons by PCR using specific primers. The nucleotide sequences of several PCR products were also determined. The results revealed that the bla ( VIM ) gene was found in 81.5% (53/65) of all isolates, bla ( VIM-2 ) gene was found in only 1 isolate and the intI gene was observed in 45.3% (24/53) of bla ( VIM )-positive isolates. One isolate carried simultaneously both bla ( IMP-1 ) and intI genes, and to the best of our knowledge this is the first report of such isolate in southwest China. These observations highlight that the genes for VIM beta-lactamase and class 1 integrons were predominantly present among the imipenem-resistant P. aeruginosa tested, confirming the current widespread threat of imipenem-resistant, integron-borne P. aeruginosa.
Anti-Bacterial Agents/pharmacology ; China ; DNA Primers/chemistry ; Drug Resistance, Bacterial ; Gene Expression Regulation, Bacterial ; Imipenem/*pharmacology ; Integrons ; Microbial Sensitivity Tests ; Models, Genetic ; Pseudomonas Infections/genetics ; Pseudomonas Infections/*microbiology ; Pseudomonas aeruginosa/*metabolism ; Sequence Analysis, DNA ; beta-Lactamases/*metabolism

No abstract available.
Anti-Bacterial Agents/pharmacology ; DNA, Bacterial/chemistry/genetics/metabolism ; Drug Resistance, Bacterial ; Integrons/*genetics ; Microbial Sensitivity Tests ; Pseudomonas aeruginosa/drug effects/*enzymology/isolation & purification ; Sequence Analysis, DNA ; beta-Lactamases/*genetics

NDM-1 (New Delhi metallo-beta-lactamase) gene encodes a metallo-beta-lactamase (MBL) with high carbapenemase activity, which makes the host bacterial strain easily dispatch the last-resort antibiotics known as carbapenems and cause global concern. Here we present the bioinformatics data showing an unexpected similarity between NDM-1 and beta-lactamase II from Erythrobacter litoralis, a marine microbial isolate. We have further expressed these two mature proteins in E. coli cells, both of which present as a monomer with a molecular mass of 25 kDa. Antimicrobial susceptibility assay reveals that they share similar substrate specificities and are sensitive to aztreonam and tigecycline. The conformational change accompanied with the zinc binding visualized by nuclear magnetic resonance, Zn(2+)-bound NDM-1, adopts at least some stable tertiary structure in contrast to the metal-free protein. Our work implies a close evolutionary relationship between antibiotic resistance genes in environmental reservoir and in the clinic, challenging the antimicrobial resistance monitoring.
Amino Acid Sequence ; Anti-Bacterial Agents ; pharmacology ; Aztreonam ; pharmacology ; Cephalosporinase ; chemistry ; genetics ; metabolism ; Computational Biology ; methods ; Drug Resistance, Bacterial ; genetics ; Enzyme Stability ; drug effects ; Evolution, Molecular ; Minocycline ; analogs & derivatives ; pharmacology ; Molecular Sequence Data ; Phylogeny ; Protein Structure, Tertiary ; drug effects ; Sequence Homology, Nucleic Acid ; Sphingomonadaceae ; drug effects ; enzymology ; genetics ; Tigecycline ; Zinc ; pharmacology ; beta-Lactamases ; chemistry ; genetics ; metabolism

BACKGROUND: The emergence of carbapenem-resistant Klebsiella pneumoniae poses a serious problem to antibiotic management. We investigated the beta-lactamases in a group of carbapenem-resistant K. pneumoniae clinical isolates from Turkey. METHODS: Thirty-seven strains of K. pneumoniae isolated from various clinical specimens were analyzed by antimicrobial susceptibility testing, PCR for the detection of beta-lactamase genes, DNA sequencing, and repetitive extragenic palindronic (REP)-PCR analysis. RESULTS: All 37 isolates were resistant to ampicillin, ampicillin/sulbactam, piperacillin, piperacillin/tazobactam, ceftazidime, cefoperazone/sulbactam, cefepime, imipenem, and meropenem. The lowest resistance rates were observed for colistin (2.7%), tigecycline (11%), and amikacin (19%). According to PCR and sequencing results, 98% (36/37) of strains carried at least one carbapenemase gene, with 32 (86%) carrying OXA-48 and 7 (19%) carrying NDM-1. No other carbapenemase genes were identified. All strains carried a CTX-M-2-like beta-lactamase, and some carried SHV- (97%), TEM- (9%), and CTX-M-1-like (62%) beta-lactamases. Sequence analysis of bla(TEM) genes identified a bla(TEM-166) with an amino acid change at position 53 (Arg53Gly) from bla(TEM-1b), the first report of a mutation in this region. REP-PCR analysis revealed that there were seven different clonal groups, and temporo-spatial links were identified within these groups. CONCLUSIONS: Combinations of beta-lactamases were found in all strains, with the most common being OXA-48, SHV, TEM, and CTX-M-type (76% of strains). We have reported, for the first time, a high prevalence of the NDM-1 (19%) carbapenemase in carbapenem-resistant K. pneumoniae from Turkey. These enzymes often co-exist with other beta-lactamases, such as TEM, SHV, and CTX-M beta-lactamases.
Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/*genetics/metabolism ; Carbapenems/*pharmacology ; DNA, Bacterial/chemistry/genetics/metabolism ; Drug Resistance, Bacterial ; Genotype ; Humans ; Klebsiella Infections/diagnosis/microbiology ; Klebsiella pneumoniae/*drug effects/enzymology/isolation & purification ; Microbial Sensitivity Tests ; Polymerase Chain Reaction ; Sequence Analysis, DNA ; Turkey ; beta-Lactamases/*genetics/metabolism

We report a case of CTX-M-55-type extended-spectrum beta-lactamase (ESBL)-producing Shigella sonnei infection in a 27-year-old Korean woman who had traveled to China. The patient was admitted to the hospital due to abdominal pain, watery diarrhea, and fever (39.3degrees C). S. sonnei was isolated from her stool specimens, and the pathogen was found to be resistant to cefotaxime due to CTX-M-55-type ESBL. Insertion sequence (IS)Ecp1 was found upstream of the blaCTX-M-55 gene. The blaCTX-M-55 gene was transferred from the S. sonnei isolate to an Escherichia coli J53 recipient by conjugation. Pulsed-field gel electrophoresis and Southern blotting revealed that the blaCTX-M-55 gene was located on a plasmid of approximately 130 kb.
Adult ; Anti-Bacterial Agents/pharmacology ; Asian Continental Ancestry Group ; Cefotaxime/pharmacology ; China ; Drug Resistance, Bacterial/drug effects ; Dysentery, Bacillary/diagnosis/*microbiology ; Electrophoresis, Gel, Pulsed-Field ; Escherichia coli/metabolism ; Feces/microbiology ; Female ; Humans ; Plasmids/chemistry/genetics ; Republic of Korea ; Shigella sonnei/enzymology/*isolation & purification ; Travel ; beta-Lactamases/genetics/*metabolism

BACKGROUND: We investigated the prevalence of plasmid-mediated quinolone resistance and its association with extended-spectrum beta-lactamase (ESBL) and AmpC beta-lactamase in Enterobacteriaceae. METHODS: A total of 347 non-duplicated isolates of Enterobacteriaceae were collected between August and October 2006 from 2 hospitals. Qnr determinant screening was conducted using PCR amplification, and all positive results were confirmed by direct sequencing. Qnr-positive strains were determined on the basis of the presence of ESBL and AmpC beta-lactamase genes. RESULTS: The qnr gene was detected in 47 of 347 clinical Enterobacteriaceae isolates. Among the 47 qnr-positive strains, Klebsiella pneumoniae (N=29) was the most common, followed by Escherichia coli (N=6), Enterobacter cloacae (N=6), Citrobacter freundii (N=5), and Enterobacter aerogenes (N=1). These isolates were identified as qnrA1 (N=6), 8 qnrB subtypes (N=40), and qnrS1 (N=1). At least 1 ESBL was detected in 38 of the 47 qnr-positive strains. Qnr-positive strains also showed high positive rates of ESBL or AmpC beta-lactamase, such as TEM, SHV, CTX-M, and DHA. DHA-1 was detected in 23 of 47 qnr-positive strains, and this was co-produced with 1 qnrA1 and 22 qnrB4. Strains harboring MIR-1T and CMY were also detected among the qnr-positive strains. Antimicrobial-resistance rates of qnr-positive strains to ciprofloxacin, levofloxacin, norfloxacin, nalidixic acid, and moxifloxacin were 51.1%, 46.8%, 46.8%, 74.5%, and 53.2%, respectively. CONCLUSIONS: The qnr genes were highly prevalent in Enterobacteriaceae, primarily the qnrB subtypes. They were closely associated with EBSL and AmpC beta-lactamase.
Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/biosynthesis/*genetics ; DNA, Bacterial/chemistry/genetics ; Drug Resistance, Bacterial/*genetics ; Enterobacteriaceae/enzymology/*genetics/isolation & purification ; Enterobacteriaceae Infections/microbiology ; *Genetic Variation ; Hospitals, University ; Humans ; Microbial Sensitivity Tests ; Plasmids/genetics/*metabolism ; Quinolones/*pharmacology ; beta-Lactamases/biosynthesis/genetics