scholarly journals Mutation in Serratia marcescens AmpC β-Lactamase Producing High-Level Resistance to Ceftazidime and Cefpirome

2001 ◽  
Vol 45 (8) ◽  
pp. 2331-2339 ◽  
Author(s):  
Alessandro Raimondi ◽  
Francesca Sisto ◽  
Hiroshi Nikaido
Keyword(s):  
Serratia Marcescens ◽  
Catalytic Efficiency ◽  
Fold Increase ◽  
Mutant Enzyme ◽  
Kinetic Properties ◽  
Cloning And Sequencing ◽  
Mutant Strains ◽  
High Level ◽  
Outer Membrane Permeability ◽  
Level Resistance

ABSTRACT Starting from a clinical isolate of Serratia marcescens that produced a chromosomally encoded AmpC β-lactamase inducibly, we isolated by stepwise selection two laboratory mutants that showed high levels of resistance to some cephalosporins. The 98R mutant apparently overproduced the unaltered β-lactamase constitutively, but the 520R mutant produced an altered enzyme, also constitutively. Ceftazidime and cefpirome MICs for the 520R mutant were much higher (512 and 64 μg/ml, respectively) than those for the 98R mutant (16 and 16 μg/ml, respectively). Yet the MICs of cephaloridine and piperacillin for the 520R mutant were four- to eightfold lower than those for the 98R mutant. Cloning and sequencing of theampC alleles showed that in the 520R mutant enzyme, the Thr64 residue, about two turns away from the active-site serine, was mutated to isoleucine. This resulted in a >1,000-fold increase in the catalytic efficiency (k cat/K m ) of the mutated AmpC enzyme toward ceftazidime, whereas there was a >10-fold decrease in the efficiency of the mutant enzyme toward cefazolin and cephaloridine. The outer membrane permeability of the 520R strain to cephalosporins was also less than in the 98R strain, and the alteration of the kinetic properties of the AmpC enzyme together with this difference in permeability explained quantitatively the resistance levels of both mutant strains to most agents studied.

scholarly journals Phenotypic Characterization of Two Ancylostoma caninum Isolates with Different Susceptibilities to the Anthelmintic Pyrantel

2008 ◽  
Vol 52 (11) ◽  
pp. 3980-3986 ◽  
Author(s):  
Steven R. Kopp ◽  
Glen T. Coleman ◽  
James S. McCarthy ◽  
Andrew C. Kotze
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Resistance Mechanism ◽  
Fold Increase ◽  
Phenotypic Characterization ◽  
Gastrointestinal Helminths ◽  
Ancylostoma Caninum ◽  
High Level ◽  
Level Resistance

ABSTRACT The anthelmintic pyrantel plays an important role in the control of gastrointestinal helminths of humans and domestic animals. Despite the demonstration of pyrantel resistance in several helminth species over the last 20 years, the resistance mechanism remains unclear. It has been hypothesized that resistance may arise as a consequence of changes to the relative proportions of subpopulations of nicotinic acetylcholine receptors (nAchRs). To test this hypothesis, we examined the responses of two isolates of the canine hookworm Ancylostoma caninum with low-level resistance (isolate NT) and high-level resistance (isolate PR) to pyrantel to nicotinic agonist drugs reported to be selective for three nAchR subtypes. We used larval motility and conformation assays and force transduction experiments with adult worms. Pyrantel and levamisole were less potent against larvae of isolate PR than larvae of isolate NT (up to an 18-fold increase in the 50% inhibitory concentration); on the other hand, bephenium was more potent against larvae of isolate PR than larvae of isolate NT (twofold) and nicotine had the same potency against larvae of both isolates. In adults, pyrantel, levamisole, and nicotine were less potent against isolate PR than isolate NT (two- to threefold), but the potency of bephenium against the two isolates was equivalent. Our data indicate a complex pattern of nAchRs in this species and suggest that the two isolates differ in their relative sensitivities to agonists targeting different nAchRs.

scholarly journals Global dissemination of tet(X3) and tet(X6) among livestock-associated Acinetobacter is sporadically mediated by highly diverse plasmidomes

2021 ◽  
Author(s):  
Kai Zhou ◽  
Yingying Cheng ◽  
Yang Liu ◽  
Yong Chen ◽  
Fuman Huang ◽  
...  
Keyword(s):  
Growth Rate ◽  
Phylogenetic Analysis ◽  
Environmental Samples ◽  
One Health ◽  
Clinical Effectiveness ◽  
Fold Increase ◽  
The Other ◽  
Swine Farm ◽  
High Level ◽  
Level Resistance

The emergence of plasmid-borne tet(X) genes mediates high-level resistance of tigecycline largely threatening its clinical effectiveness. Currently, the dissemination pattern of plasmid-borne tet(X) genes remains unclear. In this study, 684 fecal and environmental samples were collected at six livestock farms, and 15 tet(X)-positive Acinetobacter isolates were recovered, mainly including 9 tet(X3)- and 5 tet(X6)-positive A. towneri strains. A clonal dissemination of tet(X3)-positive A. towneri was detected in a swine farm, while the tet(X6)-positive A. towneri strains mainly sporadically disseminated in the same farm. A tet(X3)-carrying plasmid (pAT181) was self-transmissible from a tigecycline-susceptible A. towneri strain to A. baumannii ATCC17978, causing a 128-fold and 64-512-fold increase in the MIC values of tigecycline and the other tetracyclines, respectively. Worrisomely, pAT181 was stably maintained and increased the growth rate of ATCC17978. Further identification of tet(X)s in 10,680 Acinetobacter genomes retrieved from GenBank revealed that, tet(X3) (n=249) followed by tet(X5)-like (n=61) and tet(X6) (n=53) are the prevalent alleles mainly carried by four species, and most of them are livestock associated. Phylogenetic analysis showed that most of tet(X3) and tet(X6)-positive isolates disseminate sporadically. The structures of tet(X3) and tet(X6) plasmidomes are highly diverse and no epidemic plasmids have emerged yet. However, cross-species and cross-region transmissions of tet(X3) might have been mediated by several plasmids in a small proportion of strains. Our study evidence that tet(X3) and tet(X6) currently disseminate sporadically in Acinetobacter. Continuous surveillance for tet(X)s in the context of One Health is necessary to prevent them from transmitting to humans.

scholarly journals Phenotypic and Biochemical Comparison of the Carbapenem-Hydrolyzing Activities of Five Plasmid-Borne AmpC β-Lactamases

2010 ◽  
Vol 54 (11) ◽  
pp. 4556-4560 ◽  
Author(s):  
Hedi Mammeri ◽  
Hélène Guillon ◽  
François Eb ◽  
Patrice Nordmann
Keyword(s):  
Biochemical Analysis ◽  
Carbapenem Resistance ◽  
Catalytic Efficiency ◽  
Wild Type ◽  
E Coli ◽  
Genes Encoding ◽  
High Level ◽  
Biochemical Comparison ◽  
Outer Membrane Permeability

ABSTRACT The CMY-2, ACT-1, DHA-1, ACC-1, and FOX-1 enzymes are representative of five plasmid-mediated AmpC (pAmpC) β-lactamase clusters. Resistance to imipenem has been reported in Enterobacteriaceae as a result of pAmpC expression combined with decreased outer membrane permeability. The aim of this study was to determine the role of different pAmpCs in carbapenem resistance and to define the structure/activity relationship supporting carbapenemase activity. The ampC genes encoding the five pAmpCs and the chromosomal AmpC of Escherichia coli EC6, which was used as a reference cephalosporinase, were cloned and introduced into wild-type E. coli TOP10 and OmpC/OmpF porin-deficient E. coli HB4 strains. The MICs of β-lactams for the recombinant strains revealed that CMY-2, ACT-1, and DHA-1 β-lactamases conferred a high level of resistance to ceftazidime and cefotaxime once expressed in E. coli TOP10 and reduced significantly the susceptibility to imipenem once expressed in E. coli HB4. In contrast, FOX-1 and ACC-1 enzymes did not confer resistance to imipenem. Biochemical analysis showed that CMY-2 β-lactamase and, to a lesser extent, ACT-1 exhibited the highest catalytic efficiency toward imipenem and showed low Km values. A modeling study revealed that the large R2 binding site of these two enzymes may support the carbapenemase activity. Therefore, CMY-2-type, ACT-1-type, and DHA-1-type β-lactamases may promote the emergence of carbapenem resistance in porin-deficient clinical isolates.

scholarly journals Sequencing of Gyrase and Topoisomerase IV Quinolone-Resistance-Determining Regions of Chlamydia trachomatis and Characterization of Quinolone-Resistant Mutants Obtained In Vitro

1998 ◽  
Vol 42 (10) ◽  
pp. 2474-2481 ◽  
Author(s):  
Sophie Dessus-Babus ◽  
Cécile M. Bébéar ◽  
Alain Charron ◽  
Christiane Bébéar ◽  
Bertille de Barbeyrac
Keyword(s):  
Chlamydia Trachomatis ◽  
Reference Strain ◽  
Fold Increase ◽  
Quinolone Resistance ◽  
Topoisomerase Iv ◽  
Mutant Strains ◽  
Resistant Strains ◽  
Resistant Mutants ◽  
High Level

ABSTRACT The L2 reference strain of Chlamydia trachomatis was exposed to subinhibitory concentrations of ofloxacin (0.5 μg/ml) and sparfloxacin (0.015 μg/ml) to select fluoroquinolone-resistant mutants. In this study, two resistant strains were isolated after four rounds of selection. The C. trachomatis mutants presented with high-level resistance to various fluoroquinolones, particularly to sparfloxacin, for which a 1,000-fold increase in the MICs for the mutant strains compared to the MIC for the susceptible strain was found. The MICs of unrelated antibiotics (doxycycline and erythromycin) for the mutant strains were identical to those for the reference strain. The gyrase (gyrA, gyrB) and topoisomerase IV (parC, parE) genes of the susceptible and resistant strains of C. trachomatis were partially sequenced. A point mutation was found in the gyrAquinolone-resistance-determining region (QRDR) of both resistant strains, leading to a Ser83→Ile substitution (Escherichia coli numbering) in the corresponding protein. ThegyrB, parC, and parE QRDRs of the resistant strains were identical to those of the reference strain. These results suggest that in C. trachomatis, DNA gyrase is the primary target of ofloxacin and sparfloxacin.

High-level resistance to oxidative stress in Lactococcus lactis conferred by Bacillus subtilis catalase KatE

Microbiology ◽  
2005 ◽  
Vol 151 (9) ◽  
pp. 3011-3018 ◽  
Author(s):  
T. Rochat ◽  
A. Miyoshi ◽  
J. J. Gratadoux ◽  
P. Duwat ◽  
S. Sourice ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Bacillus Subtilis ◽  
Lactococcus Lactis ◽  
Fold Increase ◽  
Inducible Promoter ◽  
Optimization Strategy ◽  
High Level ◽  
The Impact ◽  
Level Resistance

Lactococcus lactis, a lactic acid bacterium widely used for food fermentations, is often exposed to damaging stress conditions. In particular, oxidative stress leads to DNA, protein and membrane damages that can be lethal. As L. lactis has no catalase, the impact of production of the Bacillus subtilis haem catalase KatE on its oxidative stress resistance was tested. This cytoplasmic catalase was engineered for extracellular expression in L. lactis with an optimization strategy based on fusion to the nisin-inducible promoter and a lactococcal signal peptide (SPUsp45). The production of KatE by L. lactis conferred an 800-fold increase in survival after 1 h exposure to 4 mM hydrogen peroxide, and a 160-fold greater survival in long-term (3 days) survival of aerated cultures in a cydA mutant, which is unable to respire. The presence of KatE protected DNA from oxidative damage and limited its degradation after long-term aeration in a cydA/recA mutant, defective in DNA repair. L. lactis is thus able to produce active catalase that can provide efficient antioxidant activity.

scholarly journals Resistance to Cefepime and Cefpirome Due to a 4-Amino-Acid Deletion in the Chromosome-Encoded AmpC β-Lactamase of a Serratia marcescens Clinical Isolate

2004 ◽  
Vol 48 (3) ◽  
pp. 716-720 ◽  
Author(s):  
Hedi Mammeri ◽  
Laurent Poirel ◽  
Pascal Bemer ◽  
Henri Drugeon ◽  
Patrice Nordmann
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract Infection ◽  
Amino Acid ◽  
Serratia Marcescens ◽  
Reference Strain ◽  
Sequencing Analysis ◽  
Amino Acid Deletion ◽  
Synergy Test ◽  
High Level ◽  
Level Resistance

ABSTRACT A multiresistant Serratia marcescens strain, HD, isolated from a patient with a urinary tract infection, was resistant to amino-, carboxy-, and ureidopenicillins, ceftazidime, and cefepime and was susceptible to cefotaxime and ceftriaxone, according to the guidelines of the NCCLS. No synergy was found between expanded-spectrum cephalosporins and clavulanic acid, according to the double-disk synergy test. The bla AmpC gene of the strain was amplified by PCR and cloned into Escherichia coli DH10B, giving rise to high-level resistance to ceftazidime, cefepime, and cefpirome. Sequencing analysis revealed that the bla AmpC gene from S. marcescens HD had a 12-nucleotide deletion compared to the bla AmpC gene from reference strain S. marcescens S3, leading to a 4-amino-acid deletion located in the H-10 helix of the β-lactamase. Kinetic analysis showed that this enzyme significantly hydrolyzed ceftazidime, cefepime, and cefpirome. This work underlined that resistance to the latest expanded-spectrum cephalosporins may be mediated by structurally modified AmpC-type β-lactamases.

scholarly journals Rapid Development of High-Level Resistance to Dolutegravir With Emergence of T97A Mutation in 2 Treatment-Experienced Individuals With Baseline Partial Sensitivity to Dolutegravir

2018 ◽  
Vol 5 (10) ◽  
Author(s):  
Jomy M George ◽  
Safia S Kuriakose ◽  
Nicola Dee ◽  
Pam Stoll ◽  
Tahaniyat Lalani ◽  
...  
Keyword(s):  
Salvage Therapy ◽  
Rapid Development ◽  
Fold Increase ◽  
Strand Transfer ◽  
Hiv Integrase ◽  
Integrase Strand Transfer Inhibitors ◽  
Single Addition ◽  
High Level ◽  
Level Resistance

Abstract HIV integrase mutation T97A emerges after suboptimal therapy with integrase strand transfer inhibitors (INSTIs), but the contribution of T97A to dolutegravir resistance remains uncertain. Here we report >10-fold increase in dolutegravir resistance after the single addition of T97A in 2 individuals with prior INSTI resistance receiving dolutegravir salvage therapy.

scholarly journals A Novel Class A Extended-Spectrum β-Lactamase (BES-1) in Serratia marcescens Isolated in Brazil

2000 ◽  
Vol 44 (11) ◽  
pp. 3061-3068 ◽  
Author(s):  
R. Bonnet ◽  
J. L. M. Sampaio ◽  
C. Chanal ◽  
D. Sirot ◽  
C. De Champs ◽  
...  
Keyword(s):  
Serratia Marcescens ◽  
Functional Group ◽  
Inhibitory Concentration ◽  
Structural Features ◽  
Cloning And Sequencing ◽  
Gene Encoding ◽  
Class A ◽  
Extended Spectrum ◽  
Related Enzyme ◽  
High Level

ABSTRACT Serratia marcescens Rio-5, one of 18 extended-spectrum β-lactamase (ESBL)-producing strains isolated in several hospitals in Rio de Janeiro (Brazil) in 1996 and 1997, exhibited a high level of resistance to aztreonam (MIC, 512 μg/ml) and a distinctly higher level of resistance to cefotaxime (MIC, 64 μg/ml) than to ceftazidime (MIC, 8 μg/ml). The strain produced a plasmid-encoded ESBL with a pI of 7.5 whose bla gene was not related to those of other plasmid-mediated Ambler class A ESBLs. Cloning and sequencing revealed a bla gene encoding a novel class A β-lactamase in functional group 2be, designated BES-1 (Brazil extended-spectrum β-lactamase). This enzyme had 51% identity with chromosomal class A penicillinase of Yersinia enterocolitica Y56, which was the most closely related enzyme and 47 to 48% identity with CTX-M-type β-lactamases, which were the most closely related ESBLs. In common with CTX-M enzymes, BES-1 exhibited high cefotaxime-hydrolyzing activity (k cat, 425 s−1). However, BES-1 differed from CTX-M enzymes by its significant ceftazidime-hydrolyzing activity (k cat, 25 s−1), high affinity for aztreonam (Ki , 1 μM), and lower susceptibility to tazobactam (50% inhibitory concentration [IC50], 0.820 μM) than to clavulanate (IC50, 0.045 μM). Likewise, certain characteristic structural features of CTX-M enzymes, such as Phe-160, Ser-237, and Arg-276, were observed for BES-1, which, in addition, harbored different residues (Ala-104, Ser-171, Arg-220, Gly-240) and six additional residues at the end of the sequence. BES-1, therefore, may be an interesting model for further investigations of the structure-function relationships of class A ESBLs.

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