scholarly journals Altered permeability and beta-lactam resistance in a mutant of Mycobacterium smegmatis.

1997 ◽  
Vol 41 (8) ◽  
pp. 1721-1724 ◽  
Author(s):  
S Mukhopadhyay ◽  
P Chakrabarti
Keyword(s):  
Mycobacterium Smegmatis ◽  
Parent Strain ◽  
Beta Lactamase ◽  
Wild Type ◽  
Beta Lactam ◽  
Beta Lactams ◽  
Intact Cells ◽  
Glycine Uptake ◽  
Penicillin Binding Proteins ◽  
High Level

Beta-lactam resistance in mycobacteria results from an interplay between the following: (i) beta-lactamase production, (ii) affinity of the penicillin-binding proteins (PBPs) for the drugs, and (iii) permeation of the drugs. A laboratory mutant of Mycobacterium smegmatis was studied in order to evaluate the roles of these factors in beta-lactam resistance. Mutant M13 was between 7- and 78-fold more resistant than the wild type to cephaloridine, cefoxitin, cefazolin, cefamandole, and cephalothin. Increased beta-lactamase activity toward these antibiotics was not observed in the mutant. The PBP profiles of the wild type and M13 were comparable. However, the affinities of PBP 1 for the beta-lactams tested were lower for the mutant than for the wild type. The permeation of the drugs measured in intact cells was lower for M13 than for the parent strain. The liposome swelling technique, which could be used for cephaloridine, also supported this view. Reduced permeation was not restricted to the beta-lactams alone. Glycine uptake was also lower in M13. Taken together, the results suggest that decreased affinities of PBP 1 for beta-lactams, combined with the decreased permeability of the cell wall of the mutant, lead to the development of high-level acquired beta-lactam resistance.

scholarly journals Mechanism of action of dd-peptidases: role of asparagine-161 in the Streptomyces R61 dd-peptidase

1993 ◽  
Vol 293 (1) ◽  
pp. 195-201 ◽  
Author(s):  
J M Wilkin ◽  
M Jamin ◽  
B Joris ◽  
J M Frere
Keyword(s):  
Site Directed Mutagenesis ◽  
Beta Lactamase ◽  
Wild Type ◽  
Peptide Substrate ◽  
Beta Lactam ◽  
Beta Lactams ◽  
Wild Type Enzyme ◽  
Class A ◽  
First Case

The role of residue Asn-161 in the interaction between the Streptomyces R61 DD-peptidase and various substrates or beta-lactam inactivators was probed by site-directed mutagenesis. The residue was successively replaced by serine and alanine. In the first case, acylation rates were mainly affected with the peptide and ester substrates but not with the thiol-ester substrates and beta-lactams. However, the deacylation rates were decreased 10-30-fold with the substrates yielding benzoylglycyl and benzoylalanyl adducts. The Asn161Ala mutant was more generally affected, although the acylation rates with cefuroxime and cefotaxime remained similar to those observed with the wild-type enzyme. Surprisingly, the deacylation rates of the benzoylglycyl and benzoylalanyl adducts were very close to those observed with the wild-type enzyme. The results also indicate that the interaction with the peptide substrate and the transpeptidation reaction were more sensitive to the mutations than the other reactions studied. The results are discussed and compared with those obtained with the Asn-132 mutants of a class A beta-lactamase.

scholarly journals Mechanisms of Resistance to Imipenem and Ampicillin in Enterococcus faecalis

2005 ◽  
Vol 49 (7) ◽  
pp. 2954-2958 ◽  
Author(s):  
Seiji Ono ◽  
Tetsuro Muratani ◽  
Tetsuro Matsumoto
Keyword(s):  
Amino Acid ◽  
Enterococcus Faecalis ◽  
Point Mutations ◽  
Beta Lactam ◽  
Beta Lactams ◽  
Beta Lactamases ◽  
Penicillin Binding Proteins ◽  
Resistant Strains ◽  
High Level ◽  
Level Resistance

ABSTRACT We found ampicillin- and imipenem-resistant isolates of vanA-possessing Enterococcus faecalis with MICs of 8 to 16 μg/ml and 4 to 32 μg/ml, respectively. There have been few reports about penicillin- and imipenem-resistant E. faecalis. Two mechanisms of beta-lactam resistance in E. faecalis, the production of beta-lactamase and the overproduction of penicillin-binding proteins (PBPs), have been reported. The resistant isolates in the current study did not produce any beta-lactamases and analysis of the PBPs showed no overproduction. However, the affinities of PBP4 for beta-lactams in the resistant strains were lower than those of susceptible strains but the affinities of other PBPs for beta-lactams did not change. Accordingly, whole pbp4 fragments from these resistant isolates were sequenced. Two amino acid substitutions at positions 520 and 605 were observed in the highly resistant strains compared to the susceptible ones, Pro520Ser and Tyr605His, and a single Tyr605His amino acid substitution was found in the low-resistance strains. These two point mutations exist in the region between the active-site-defining motifs SDN and KTG of the penicillin-binding domain, the main target of beta-lactams. A strong correlation was seen between these substitutions and decreasing affinities of PBP4 to beta-lactams. In E. faecalis, resistance due to mutations in PBPs has not been reported, though it has in Enterococcus faecium. Our results suggest that development of high-level resistance to penicillins and imipenem depends on point mutations of PBP4 at positions 520 and 605.

scholarly journals In Vivo Activity of QPX7728, an Ultrabroad-Spectrum Beta-Lactamase Inhibitor, in Combination with Beta-Lactams against Carbapenem-Resistant Klebsiella pneumoniae

2020 ◽  
Vol 64 (11) ◽  
Author(s):  
Mojgan Sabet ◽  
Ziad Tarazi ◽  
David C. Griffith
Keyword(s):  
Klebsiella Pneumoniae ◽  
Beta Lactamase ◽  
Beta Lactam ◽  
Beta Lactams ◽  
Bacterial Killing ◽  
Clinical Issue ◽  
Content Type ◽  
Beta Lactam Antibiotics ◽  
Carbapenem Resistant ◽  
Lactamase Inhibitor

ABSTRACT Resistance to beta-lactams has created a major clinical issue. QPX7728 is a novel ultrabroad-spectrum cyclic boronic acid beta-lactamase inhibitor with activity against both serine and metallo-beta-lactamases developed to address this resistance for use in combination with beta-lactam antibiotics. The objective of these studies was to evaluate the activity of QPX7728 in combination with multiple beta-lactams against carbapenem-resistant Klebsiella pneumoniae isolates in a neutropenic mouse thigh infection model. Neutropenic mice were infected with strains with potentiated beta-lactam MICs of ≤2 mg/liter in the presence of 8 mg/liter QPX7728. Two strains of carbapenem-resistant K. pneumoniae were tested with aztreonam, biapenem, cefepime, ceftazidime, ceftolozane, and meropenem alone or in combination with 12.5, 25, or 50 mg/kg of body weight of QPX7728 every 2 hours for 24 hours. Treatment with all beta-lactams alone either was bacteriostatic or allowed for bacterial growth. The combination of QPX7728 plus each of these beta-lactams produced bacterial killing at all QPX7728 doses tested. Overall, these data suggest that QPX7728 administered in combination with different partner beta-lactam antibiotics may have utility in the treatment of bacterial infections due to carbapenem-resistant K. pneumoniae.

scholarly journals A TEM-derived extended-spectrum beta-lactamase in Pseudomonas aeruginosa.

1996 ◽  
Vol 40 (11) ◽  
pp. 2488-2493 ◽  
Author(s):  
P Mugnier ◽  
P Dubrous ◽  
I Casin ◽  
G Arlet ◽  
E Collatz
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Clinical Strain ◽  
Beta Lactamase ◽  
Beta Lactam ◽  
Aminoglycoside Resistance ◽  
Double Mutation ◽  
Extended Spectrum Beta Lactamase ◽  
Beta Lactam Antibiotics ◽  
Extended Spectrum ◽  
High Level

A clinical strain of Pseudomonas aeruginosa, PAe1100, was found to be resistant to all antipseudomonal beta-lactam antibiotics and to aminoglycosides, including gentamicin, amikacin, and isepamicin. PAe1100 produced two beta-lactamases, TEM-2 (pI 5.6) and a novel, TEM-derived extended-spectrum beta-lactamase called TEM-42 (pI 5.8), susceptible to inhibition by clavulanate, sulbactam, and tazobactam. Both enzymes, as well as the aminoglycoside resistance which resulted from AAC(3)-IIa and AAC(6')-I production, were encoded by an 18-kb nonconjugative plasmid, pLRM1, that could be transferred to Escherichia coli by transformation. The gene coding for TEM-42 had four mutations that led to as many amino acid substitutions with respect to TEM-2: Val for Ala at position 42 (Ala42), Ser for Gly238, Lys for Glu240, and Met for Thr265 (Ambler numbering). The double mutation Ser for Gly238 and Lys for Glu240, which has so far only been described in SHV-type but not TEM-type enzymes, conferred concomitant high-level resistance to cefotaxime and ceftazidime. The novel, TEM-derived extended-spectrum beta-lactamase appears to be the first of its class to be described in P. aeruginosa.

scholarly journals Plasmid-mediated dissemination of the metallo-beta-lactamase gene blaIMP among clinically isolated strains of Serratia marcescens.

1995 ◽  
Vol 39 (4) ◽  
pp. 824-829 ◽  
Author(s):  
H Ito ◽  
Y Arakawa ◽  
S Ohsuka ◽  
R Wacharotayankun ◽  
N Kato ◽  
...  
Keyword(s):  
Serratia Marcescens ◽  
The Other ◽  
Beta Lactamase ◽  
Beta Lactams ◽  
E Coli ◽  
Aichi Prefecture ◽  
Imipenem Resistance ◽  
High Level ◽  
Lactamase Gene ◽  
Moderately Resistant

The distribution of strains producing metallo-beta-lactamase among 105 strains of Serratia marcescens was investigated. All of these strains were isolated in seven general hospitals located in Aichi Prefecture, Japan, from April to May 1993. Southern hybridization analysis suggested that four S. marcescens strains, AK9373, AK9374, AK9385, and AK9391, had a metallo-beta-lactamase genes similar to the blaIMP gene found by our laboratory (E. Osano, Y. Arakawa, R. Wacharotayankun, M. Ohta, T. Horii, H. Ito, F. Yoshimura, and N. Kato, Antimicrob. Agents Chemother. 38:71-78, 1994), and these four strains showed resistance to carbapenems as well as to the other broad-spectrum beta-lactams. In particular, strains AK9373, AK9374, and AK9391 showed an extraordinarily high-level resistance to imipenem (MICs, > or = 64 micrograms/ml), whereas strain AK9385 demonstrated moderate imipenem resistance (MIC, 8 micrograms/ml). The imipenem resistance of AK9373 was transferred to Escherichia coli CSH2 by conjugation with a frequency of 10(-5). The DNA probe of the blaIMP gene hybridized to a large plasmid (approximately 120 kb) transferred into the E. coli transconjugant as well as to the large plasmids harbored by AK9373. On the other hand, although we failed in the conjugational transfer of imipenem resistance from strains AK9374, AK9385, and AK9391 to E. coli CSH2, imipenem resistance was transferred from these strains to E. coli HB101 by transformation. A plasmid (approximately 25 kb) was observed in each transformant which acquired imipenem resistance. The amino acid sequence at the N terminus of the enzyme purified from strain AK9373 was identical to that of the metallo-beta-lactamase IMP-1. In contrast, strains ES9348, AK9386, and AK93101, which were moderately resistant to imipenem (MICs, > or = 4 to < or = 8 micrograms/ml), had no detectable blaIMP gene. As a conclusion, 19% of clinically isolated S. marcescens strains in Aichi Prefecture, Japan, in 1993 were resistant to imipenem (MICs, > or = 2 micrograms/ml), and strains which showed high-level imipenem resistance because of acquisition of a plasmid-mediated blaIMP-like metallo-beta-lactamase gene had already proliferated as nosocomial infections, at least in a general hospital.

scholarly journals PBP5, PBP6 and DacD play different roles in intrinsic β-lactam resistance of Escherichia coli

Microbiology ◽  
2011 ◽  
Vol 157 (9) ◽  
pp. 2702-2707 ◽  
Author(s):  
Sujoy Kumar Sarkar ◽  
Mouparna Dutta ◽  
Chiranjit Chowdhury ◽  
Akash Kumar ◽  
Anindya S. Ghosh
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Parent Strain ◽  
Amino Acid Identity ◽  
Enzymic Activity ◽  
Exponential Phase ◽  
Wild Type ◽  
Acid Identity ◽  
Penicillin Binding Proteins ◽  
E Coli

Escherichia coli PBP5, PBP6 and DacD, encoded by dacA, dacC and dacD, respectively, share substantial amino acid identity and together constitute ~50 % of the total penicillin-binding proteins of E. coli. PBP5 helps maintain intrinsic β-lactam resistance within the cell. To test if PBP6 and DacD play simlar roles, we deleted dacC and dacD individually, and dacC in combination with dacA, from E. coli 2443 and compared β-lactam sensitivity of the mutants and the parent strain. β-Lactam resistance was complemented by wild-type, but not dd-carboxypeptidase-deficient PBP5, confirming that enzymic activity of PBP5 is essential for β-lactam resistance. Deletion of dacC and expression of PBP6 during exponential or stationary phase did not alter β-lactam resistance of a dacA mutant. Expression of DacD during mid-exponential phase partially restored β-lactam resistance of the dacA mutant. Therefore, PBP5 dd-carboxypeptidase activity is essential for intrinsic β-lactam resistance of E. coli and DacD can partially compensate for PBP5 in this capacity, whereas PBP6 cannot.

Occurrence of beta-lactamase producing Escherichia coli in healthy and diarrhoeic dogs in Andhra Pradesh, India

2018 ◽  
Author(s):  
N. Mohammad Sharif ◽  
B. Sreedevi ◽  
R. K. Chaitanya ◽  
Ch. Srilatha
Keyword(s):  
Escherichia Coli ◽  
Andhra Pradesh ◽  
Diffusion Method ◽  
Beta Lactamase ◽  
Beta Lactam ◽  
Beta Lactams ◽  
Extended Spectrum Beta Lactamase ◽  
E Coli ◽  
Healthy Dogs ◽  
Group 2

The present study was carried out to characterize beta-lactam resistance in Escherichia coli isolated from healthy and diarrhoeic dogs. A total of 93 E. coli were isolated from the rectal swabs of 136 dogs (60/92 of healthy dogs and 33/44 of diarrhoeic dogs). Predominant serotypes detected include rough (19 isolates), O141 (5), O9 (2), O126 (2), O128 (2), O15, O20, O35, O49, O63, O85, O101, O116, O117, O118, O119 (1 isolate each) and the rest of 52 isolates were untypable (UT). Disc diffusion method revealed resistance to cefotaxime (41.9%), ceftriaxone (34.4%), ceftazidime (30.1%) and aztreonam (18.2%). Overall frequency of extended spectrum beta-lactamase (ESBL) phenotype was found to be 29% (27/93). Beta-lactamase genes detected include blaAmpC (86.0%), blaSHV (30.1%), blaCTX-M group-1 (19.3%), blaTEM (17.2%), blaOXA (13.9%) and blaCTX-M group-2 (7.5%). The study revealed resistance to commonly prescribed beta-lactams, with ESBL phenotype in E. coli of canine origin in Andhra Pradesh, India.

scholarly journals A Statistical Approach for Determination of Disk Diffusion-Based Cutoff Values for Systematic Characterization of Wild-Type and Non-Wild-Type Bacterial Populations in Antimicrobial Susceptibility Testing

2015 ◽  
Vol 53 (6) ◽  
pp. 1812-1822 ◽  
Author(s):  
Giorgia Valsesia ◽  
Malgorzata Roos ◽  
Erik C. Böttger ◽  
Michael Hombach
Keyword(s):  
Resistance Mechanisms ◽  
Beta Lactamase ◽  
Wild Type ◽  
Beta Lactam ◽  
Bacterial Populations ◽  
Content Type ◽  
Extended Spectrum Beta Lactamase ◽  
E Coli

In this study, we introduce a new approach for determination of epidemiologic cutoffs (ECOFFs) and resistant-population cutoffs (RCOFFs) based on receiver operating characteristic (ROC) curves. As an example, the method was applied for determination of ECOFFs for seven different beta-lactam antibiotics and wild-type populations ofEscherichia coli,Klebsiella pneumoniae, andEnterobacter cloacae. In addition, RCOFFs were determined for bacterial populations with defined resistance mechanisms (“resistotypes”), i.e., extended-spectrum beta-lactamase (ESBL)-positiveE. coli, ESBL-positiveK. pneumoniae, and ESBL-positiveE. cloacae; AmpC cephalosporinase-positiveE. coliand AmpC-positiveK. pneumoniae; and broad-spectrum beta-lactamase (BSBL)-positiveE. coli. RCOFFs and ECOFFs are instrumental for a systematic characterization of associations between resistotypes and wild-type populations.

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