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J Sirot

Publications and source records attributed to J Sirot.

At least 19 recordsLinked to original sources
PubMed1998

Chromosomally encoded ampC-type beta-lactamase in a clinical isolate of Proteus mirabilis.

A clinical strain of Proteus mirabilis (CF09) isolated from urine specimens of a patient displayed resistance to amoxicillin (MIC >4,096 microg/ml), ticarcillin (4,096 microg/ml), cefoxitin (64 microg/ml), cefotaxime (256 microg/ml), and ceftazidime (128 microg/ml) and required an elevated MIC of aztreonam (4 microg/ml). Clavulanic acid did not act synergistically with cephalosporins. Two beta-lactamases with apparent pIs of 5.6 and 9.0 were identified by isoelectric focusing on a gel. Substrate and inhibition profiles were characteristic of an AmpC-type beta-lactamase with a pI of 9.0. Amplification by PCR with primers for ampC genes (Escherichia coli, Enterobacter cloacae, and Citrobacter freundii) of a 756-bp DNA fragment from strain CF09 was obtained only with C. freundii-specific primers. Hybridization results showed that the ampC gene is only chromosomally located while the TEM gene is plasmid located. After cloning of the gene, analysis of the complete nucleotide sequence (1,146 bp) showed that this ampC gene is close to blaCMY-2, from which it differs by three point mutations leading to amino acid substitutions Glu --> Gly at position 22, Trp --> Arg at position 201, and Ser --> Asn at position 343. AmpC beta-lactamases derived from that of C. freundii (LAT-1, LAT-2, BIL-1, and CMY-2) have been found in Klebsiella pneumoniae, E. coli, and Enterobacter aerogenes and have been reported to be plasmid borne. This is the first example of a chromosomally encoded AmpC-type beta-lactamase observed in P. mirabilis. We suggest that it be designated CMY-3.

Bacterial Proteins↗
PubMed1998

Inhibitor-resistant OXY-2-derived beta-lactamase produced by Klebsiella oxytoca.

Klebsiella oxytoca strains are generally moderately resistant to amoxicillin and ticarcillin due to the activities of the chromosomally encoded OXY-1 and OXY-2 class A beta-lactamase families. These enzymes have the ability to hydrolyze not only penicillins but also cephalosporins, including cefuroxime, ceftriaxone, and aztreonam, and are inhibited by clavulanic acid. A Klebsiella oxytoca strain was isolated from a culture of blood from a patient who had been treated with amoxicillin-clavulanate (3 g/day) for 10 days 1 month earlier. This strain harbored an unusual phenotype characterized by resistance to amoxicillin-clavulanate. It produced an OXY-2-type beta-lactamase (pI 6.3), as confirmed by PCR amplification with primers specific for the OXY-2-encoding gene. Gene sequencing revealed a point mutation (A-->G) corresponding to the amino acid substitution Ser-->Gly at position 130. This mutant enzyme was poorly inhibited by inhibitors, and its kinetic constants compared to those of the parent enzyme were characterized by an increased Km value for ticarcillin, with a drastically reduced activity against cephalosporins, as is observed with inhibitor-resistant TEM enzymes. The substitution Ser-->Gly-130 was previously described in the inhibitor-resistant beta-lactamase SHV-10 derived from an SHV-5 variant, but this is the first report of such a mutant in OXY enzymes from K. oxytoca.

Drug Resistance, Microbial↗
PubMed1998

Non-O157:H7 Stx2-producing Escherichia coli strains associated with sporadic cases of hemolytic-uremic syndrome in adults.

From August 1996 to May 1997, six verotoxin-producing Escherichia coli (VTEC) strains were isolated from stool specimens of adults suffering from hemolytic-uremic syndrome (HUS). All the isolates were stx2 positive and belonged to different serotypes: O6:H4, O91:H10, O91:H21, O rough:H16, OX3:H-, and O nontypeable: H-. The enterohemolysin (Ehly)-encoding genes were detected in two isolates, and none of the isolates harbors the intimin (Eae)-encoding gene. These findings suggest that stx2-positive non-O157:H7 VTEC is a major cause of HUS in adults and that several sources of pathogens are responsible for local endemic infections.

Adhesins, Bacterial↗
PubMed1997

Inhibitor-resistant TEM (IRT) beta-lactamases with different substitutions at position 244.

A novel inhibitor-resistant TEM (IRT) beta-lactamase was detected in an Escherichia coli isolate resistant to amoxicillin-clavulanate and susceptible to cephalothin. The substrate and inhibitor profiles of this beta-lactamase were similar to those of IRT-1 and IRT-2. The novel IRT's bla gene was sequenced, and the deduced amino acid sequence showed the amino acid replacement Arg for His-244 of the TEM-1 sequence. Substitutions for Arg-244 have been reported in three TEM-1 mutants: IRT-1 (which corresponds to TEM-31) (Cys), IRT-2/TEM-30 (Ser), and TEM-41 (Thr). We designated this novel beta-lactamase, which corresponds to TEM-51, IRT-15.

Amino Acid Sequence↗
PubMed1997

Novel extended-spectrum TEM-type beta-lactamase from an Escherichia coli isolate resistant to ceftazidime and susceptible to cephalothin.

A novel extended-spectrum TEM-type beta-lactamase was detected in an Escherichia coli isolate which was resistant to ceftazidime and susceptible to cephalothin. The corresponding bla gene was sequenced. The deduced amino acid sequence showed the following three amino acid replacements with respect to the TEM-2 sequence: Glu-->Lys-104, Arg-->Ser-164, and Glu-->Lys-240. Since it confers a ceftazidimase-type resistance phenotype, we propose for this novel enzyme the designation CAZ-9, corresponding to TEM-46 in the sequential numbering scheme of TEM beta-lactamases.

Ceftazidime↗
PubMed1997

A complex mutant of TEM-1 beta-lactamase with mutations encountered in both IRT-4 and extended-spectrum TEM-15, produced by an Escherichia coli clinical isolate.

Escherichia coli GR102 was isolated from feces of a leukemic patient. It expressed different levels of resistance to amoxicillin or ticarcillin plus clavulanate and to the various cephalosporins tested. The double-disk synergy test was weakly positive. Production of a beta-lactamase with a pI of 5.6 was transferred to E. coli HB101 by conjugation. The nucleotide sequence was determined by direct sequencing of the amplification products obtained by PCR performed with TEM gene primers. This enzyme differed from TEM-1 (blaT-1B gene) by four amino acid substitutions: Met-->Leu-69, Glu-->Lys-104, Gly-->Ser-238 and Asn-->Asp-276. The amino acid susbstitutions Leu-69 and Asp-276 are known to be responsible for inhibitor resistance of the IRT-4 mutant, as are Lys-104 and Ser-238 substitutions for hydrolytic activity of the extended-spectrum beta-lactamases TEM-15, TEM-4, and TEM-3. These combined mutations led to a mutant enzyme which conferred a level of resistance to coamoxiclav (MIC, 64 microg/ml) much lower than that conferred by IRT-4 (MIC, 2,048 microg/ml) but higher than that conferred by TEM-15 or TEM-1 (MIC, 16 microg/ml). In addition, the MIC of ceftazidime for E. coli transconjugant GR202 (1 microg/ml) was lower than that for E. coli TEM-15 (16 microg/ml) and higher than that for E. coli IRT-4 or TEM-1 (0.06 microg/ml). The MICs observed for this TEM-type enzyme were related to the kinetic constants Km and k(cat) and the 50% inhibitory concentration, which were intermediate between those observed for IRT-4 and TEM-15. In conclusion, this new type of complex mutant derived from TEM-1 (CMT-1) is able to confer resistance at a very low level to inhibitors and at a low level to extended-spectrum cephalosporins. CMT-1 received the designation TEM-50.

Anti-Bacterial Agents↗
PubMed1997

[Statistical study of the SIRSCAN computerised camera].

The antibiograms of 1162 bacterial strains, including references, have been performed within four centres. They have been read manually and by the SIRSCAN camera, which yields to 30936 couples of diameters values. A non-concordance, at a 3 mm level, was observed fot 11.14% of the diameters. The mean of difference is 0.82 mm and the standard deviation 3.34 mm. Round Petri dishes gave results less reliable than those obtained with square dishes. A deviation in function of the centres is obtained for wild-strains as for the references. For the whole population a S/R discordance (sensible/résistant confusion) is obtained for 1.76% of the diameters. This value drops to 0.93% for enterobacteriaceae, P. aeruginosa and S. aureus (968 strains).

Anti-Bacterial Agents↗
PubMed1996

Survey of prevalence of extended spectrum beta-lactamases among enterobacteriaceae.

During the first six months of 1990 and 1994 respectively, 3179 and 2721 non-repetitive strains of Enterobacteriaceae were isolated at the teaching hospital of Clermont-Ferrand. Extended-spectrum beta-lactamases were found in 147 isolates, mainly in Klebsiella pneumoniae (n = 73), Enterobacter aerogenes (n = 37) and Proteus mirabilis (n = 15). TEM-3/CTX-1, SHV-4/CAZ-5 and TEM-24/CAZ-6 were the prevalent enzymes. However, the most frequently involved enzyme in 1990 was TEM-3 (48.6%) whereas in 1994 it was SHV-4 (56.0%). Two salient features emerged from this study: the high frequency of TEM-3 production in P. mirabilis and the production of SHV-4 enzyme, heretofore observed solely in K. pneumoniae, in two other species, E. aerogenes and Escherichia coli.

Anti-Bacterial Agents↗
PubMed1996

Characterization of an inhibitor-resistant enzyme IRT-2 derived from TEM-2 beta-lactamase produced by Proteus mirabilis strains.

Ten clinical isolates of Proteus mirabilis were found to produce an inhibitor-resistant TEM beta-lactamase (IRT) in association with a TEM-1 enzyme. The IRT enzyme was derived from TEM-2. The blaIRT gene differs from blaTEM-2 gene by one point mutation which leads to the amino-acid substitution Arg-->Ser at position 244, as observed for the original IRT-2 enzyme derived from TEM-1 reported in Escherichia coli. This is the first report of an IRT beta-lactamase derived from TEM-2.

Microbial Sensitivity Tests↗
PubMed1996

Enteroadherent Escherichia coli and diarrhea in children: a prospective case-control study.

The relative contribution of diarrheagenic Escherichia coli was examined during a 1-year prospective study of hospitalized children in Clermont-Ferrand, France, including 220 case patients (with diarrhea) and 211 matched controls. Fecal isolates were characterized by means of their pattern of adherence to HEp-2 cells and by colony hybridization with DNA probes specific for the six categories of diarrheagenic E. coli. No enteroinvasive or enterotoxigenic E. coli isolates were isolated. Twenty-eight (6.5%) eae-positive isolates and 39 (9%) enteroaggregative E. coli isolates characterized with the aggregative adherence probe and/or by their adherence pattern were detected; they were equally distributed among the patients and the controls. Diffusely adhering E. coli was the predominant pathotype: 30.7% were detected by their adherence pattern and 13.7% were detected with the daaC probe. They were isolated with similar frequencies from the patients and the controls, thereby showing no association with diarrhea. However, daaC-positive strains were significantly associated with a past record of urinary tract infections. These results suggest that the diffusely adhering E. coli organisms isolated in the present study are not true intestinal pathogens but may be regarded as resident colonic strains.

Adolescent↗
PubMed1996

[Detection of pneumococci with reduced susceptibility to penicillin G using the ATB++STREP strip].

Antibiotic susceptibility of 104 clinical isolates of Streptococcus pneumoniae was routinely tested using ATB STREP strips (Bio-Mérieux), and interpreted with ATB PLUS Expert V 2.3.1 software. In parallel, CMI of penicillin G was determined by Etest, strains with MIC < or = 0.06 mg/l were classified as penicillin susceptible. ATB system proved to be both sensitive and specific for the detection of strains of reduced susceptibility to penicillin G, accurately detecting 70 out of the 71 susceptible strains, and all of the 33 resistant strains. However the level of resistance cannot be deduced from ATB results.

Anti-Bacterial Agents↗
PubMed1995

First characterization of inhibitor-resistant TEM (IRT) beta-lactamases in Klebsiella pneumoniae strains.

Two clinical strains of Klebsiella pneumoniae, TP 01 and TP 02, presented resistance to amoxicillin-clavulanate and were fully susceptible to cephalothin. These strains produced two beta-lactamases, SHV-1 and a TEM enzyme with a pI of 5.2. The previously described changes Arg-244-->Cys and Arg-244-->Ser in IRT-1 and IRT-2 (A. Belaaouaj, C. Lapoumeroulie, M. M. Caniça, G. Vedel, P. Nevot, R. Krishnamoorthy, and G. Paul, FEMS Microbiol. Lett. 120:75-80, 1994) were found in TEM enzymes from the TP 01 and TP 02 strains, respectively. This is the first report of inhibitor-resistant TEM (IRT) in species other than Escherichia coli from the family Enterobacteriaceae.

Amoxicillin↗
PubMed1995

Molecular characterization of nine different types of mutants among 107 inhibitor-resistant TEM beta-lactamases from clinical isolates of Escherichia coli.

DNA-DNA hybridization and sequencing were performed to determine the molecular basis of resistance to clavulanic acid in 107 inhibitor-resistant TEM (IRT) enzymes produced by Escherichia coli clinical isolates. These beta-lactamases derived from TEM-1 enzyme focused at pI 5.2 (n = 68) or 5.4 (n = 39) and were very poorly inhibited by clavulanic acid compared with TEM-1 enzyme. Results showed that the amino acid sequences of 84 of the 107 enzymes differ from TEM-1 by one or two substitutions previously described: Arg-244-->Ser (IRT-2) in 22 strains, Met-69-->Leu (TEM-33) in 17 strains, Met-69-->Val (TEM-34) in 14 strains, Met-69-->Ile (IRT-3) in 6 strains, Met-69-->Leu associated with Asn-276-->Asp (IRT-4) in 13 strains, and Met-69-->Val associated with Asn-276-->Asp (TEM-36) in 12 strains. A new combination, Met-69-->Ile with Asn-276-->Asp, was found in 20 strains and was called IRT-8. Two IRT enzymes not previously described were characterized. The substitution Met-69-->Val associated with a novel substitution Arg-275-->Leu occurred in one strain. The combination Met-69-->Leu and Asn-276-->Asp was associated with the novel substitution Trp-165-->Arg in two strains. These two novel enzymes were called IRT-9 and IRT-10, respectively. The implication of these novel mutated positions, 165 and 275, in resistance to inactivation by clavulanate was supported by crystallographic data on the TEM-1 enzyme and results of site-directed mutagenesis. Molecular characterization of these mutants showed great diversity among the genes coding for inhibitor-resistant TEM enzymes produced by clinical E. coli isolates.

Base Sequence↗
PubMed1995

[Detection of beta-lactamases resistant to inhibitors (IRT) by the disk diffusion method].

UNLABELLED: 85 E. coli strains producing various beta-lactamase were studied: TEM low level: n = 11, TEM high level: n = 4, oxacillinase (OXA): n = 5, céphalosporinase (CASE) +/- TEM: n = 5, inhibitor resistant TEM (IRT) n = 50, IRT+TEM: n = 3, IRT+ CASE: n = 7. Their susceptibility to amoxicillin (AMX), amoxicillin+clavulanate (AMC), ticarcillin (TIC), ticarcillin+clavulanate (TCC), piperacillin (PIP), piperacillin+tazobactam (TAZ), mecillinam (MEC), cephalotin (CF), cefoxitin (FOX) et ceftazidime (CAZ) were evaluated by a disk diffusion method, in order to determine the resistance pattern which allows a reliable detection of IRT-producing strains. The phenotype AMX R, TIC R, AMCI/R, TCCI/R et CFS was observed in 46 out of 53 IRT +/- TEM strains, 4/5 of OXA strains and 1/11 of TEM low level strains. A better sensitivity could be obtained by a modification of breakpoints for AMC, TCC and CF, however OXA-producing strains remain indistinguishable from IRT-strains.

Anti-Bacterial Agents↗
PubMed1994

Clinical and bacteriological survey after change in aminoglycoside treatment to control an epidemic of Enterobacter cloacae.

The effect of a change in the first line antibiotic treatment in a neonatal unit was studied. A total of 238 neonates (G1), admitted between 1 January and 31 July 1989, and treated with gentamicin, were compared with 398 (G2) admitted between 1 August 1989 and 31 July 1990 who received amikacin, in the combination of ampicillin plus an aminoglycoside. This change was implemented in an attempt to prevent the spread of an epidemic strain of Enterobacter cloacae resistant to third generation cephalosporins and all aminoglycosides, except amikacin. The change in treatment had no effect on the incidence of nosocomial infections [19.7% (G1) vs. 16.3% (G2) RR = 1.21 (0.86-1.70)], but the proportion of patients with nosocomial infections caused by the E. cloacae decreased (6.3% vs. 2.0% RR 3.14 CI 1.35-7.28). Certain trends in the bacterial ecology emerged: E. aerogenes and Enterococci increased in G2. The proportion of gentamicin-resistant strains such as E. cloacae or Staphylococci decreased and there was no increase in aminoglycoside-resistant strains, except in Escherichia coli, in which resistance to amikacin rose from 0 to 3%. This study illustrates the influence of antimicrobial therapy on the species and the resistance of strains isolated in nosocomial infections. It also highlights the need for epidemiologic surveillance, and poses the question of how best to modify antibiotic policy.

Amikacin↗
PubMed1994

Clinical isolates of Escherichia coli producing multiple TEM mutants resistant to beta-lactamase inhibitors.

Twenty clinical isolates of Escherichia coli resistant to amoxycillin and ticarcillin, both in combination with clavulanic acid, were studied. The ranges of MICs for these strains as determined by the agar dilution method were as follows: amoxycillin, 2048- > 4096 mg/L; ticarcillin, 512- > 4096 mg/L; piperacillin, 32-256 mg/L; mecillinam, 0.5-8 mg/L; and cephalothin 4-16 mg/L. Combining amoxycillin with beta-lactamase inhibitors, each at a fixed concentration of 4 mg/L, had only modest potentiating effects on the activities of this agent, the ranges of MICs falling to 256- > 2048 mg/L in the presence of clavulanic acid or sulbactam and to 64-1024 mg/L and 128-2048 mg/L in the presence of tazobactam and brobactam respectively. The pI values for the beta-lactamases produced by the 20 isolates were 5.2 for 15 strains, 5.4 for four strains and 7.4 for a single strain. Colony hybridization with oligonucleotides was performed in order to detect substitutions of arginine at position 241 (Arg-241) and methionine at position 67 (Met-67). Based on this technique, the four beta-lactamases with pI values of 5.4 were grouped into two oligotypes (+ = hybridization, - = non-hybridization)-Arg-241+, Met-67- (n = 3) and Arg-241+, Met-67+ (n = 1); in one of the three mutants which did not hybridize with the Met-67 probe, leucine had been substituted for methionine at position 67. The beta-lactamases with pI values of 5.2 which were identified in 15 strains were grouped into the following three oligotypes: Arg-241-, Met-67+ (n = 7); Arg-241-, Met-67- (n = 6); and Arg-241+, Met-67- (n = 2). In three of the 13 mutants which failed to hybridize with the Arg-241 probe, serine residues had replaced arginine residues at position 241. Substitutions of Arg-241 or Met-67 led to reduced affinities of the mutant enzymes for the beta-lactams tested. The results of the hybridization studies demonstrate that, amongst E. coli clinical isolates, there is a diversity of mutant TEM enzymes mediating resistance to beta-lactamase inhibitors.

Amoxicillin↗
PubMed1994

Frequency of inhibitor-resistant TEM beta-lactamases in Escherichia coli isolates from urinary tract infections in France.

Over a six-month period in 1993, 2972 non-duplicated isolates of Escherichia coli causing urinary tract infections were collected in a French teaching hospital (n = 785) and in three private laboratories (n = 2187). The resistance rate to amoxycillin-clavulanate combination (MIC > 16 mg/l) was 25.0% in the hospital isolates and 10.0% in the community isolates. Respectively, 27.5% and 45.0% of hospital and community isolates resistant to amoxycillin-clavulanate exhibited an unusual beta-lactam resistance pattern, suggesting inhibitor-resistant TEM (IRT) beta-lactamase production. These isolates were highly resistant to amoxycillin-clavulanate (MIC90 > 1024 mg/L), but were susceptible to cephalosporins (MIC < 32 mg/L). Enzyme extracts of these IRT-producing strains focused at pI 5.2 (n = 100) or 5.4 (n = 53). DNA-DNA hybridization confirmed that the beta-lactamases involved in this resistance mechanism were TEM-1 derived and contained variations in the altered positions described in IRT enzymes. This study shows a total frequency of 4.9% of IRT-producing isolates among E. coli isolated from urine specimens.

Amoxicillin↗