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E Cambau

Publications and source records attributed to E Cambau.

At least 37 records · Page 2Linked to original sources
PubMed1998

Clinical utility of an amplification test based on ligase chain reaction in pulmonary tuberculosis.

We evaluated the sensitivity and specificity of a new semiautomated direct amplification test (DAT), the LCx-MTB, for the diagnosis of pulmonary tuberculosis (TB) and assessed its positive predictive value by focusing on patients with high clinical and radiologic suspicion of pulmonary TB. Respiratory tract specimens from 32 consecutive patients with high suspicion of active pulmonary TB (case patients) and from 204 control patients were cultured for Mycobacterium tuberculosis and tested by LCx-MTB. Sensitivity and specificity of LCx-MTB when compared with culture was, respectively, 80 and 98%. Pulmonary TB was confirmed in the 32 case patients without knowledge of the LCx results: 18 patients were smear- and culture-positive for M. tuberculosis, and all gave at least one specimen that was LCx-positive. Eight patients were smear-negative culture-positive, and seven gave at least one LCx-positive specimen. LCx-MTB was negative in all the specimens obtained from six patients with smear- and culture-negative TB. A positive LCx-MTB result in a smear negative specimen was 100% predictive that at least one of the case patients' specimens would yield M. tuberculosis. As a consequence, knowledge of the LCx-MTB results at the time of specimen collection could have hastened the start of the antituberculosis therapy in three (21%) smear-negative case patients and could have avoided unnecessary invasive diagnostic procedures in four (29%). We conclude that the sensitivity of LCx-MTB in detecting M. tuberculosis DNA in respiratory tract specimens is similar to other DATs, that LCx-MTB is a reliable test for confirmation of TB in smear-positive patients and that LCx-MTB could be beneficial as a diagnostic step in smear-negative patients with a high suspicion of pulmonary TB.

Antigens, Bacterial↗
PubMed1998

Use of DNA fingerprinting for primary surveillance of nosocomial tuberculosis in a large urban hospital: detection of outbreaks in homeless people and migrant workers.

SETTING: A large urban teaching hospital in the southeast of Paris. OBJECTIVE: Primary surveillance of nosocomial transmission of tuberculosis (TB) by systematic restriction fragment length polymorphism analysis (RFLP) of isolates (n = 161) recovered from smear-positive pulmonary TB patients identified from 1 March 1993 to 28 February 1994, and from all TB patients (with any form of tuberculous infection) identified from 1 March 1994 to 30 April 1995. RESULTS: Systematic RFLP analysis revealed 12 clusters of patients (n = 40) infected by strains of Mycobacterium tuberculosis showing matching RFLP patterns. None of the isolates were multidrug-resistant. Compared with non-clustered patients, clustered patients were more likely to be homeless (55% vs 19%, P < or = 0.001), or Africans living in hostels for migrant workers (20% vs 6%, P = 0.01), and had fewer previous admissions to hospital (12% vs 28%, P = 0.05). Further epidemiological investigations showed that the clustered TB cases actually resulted not from nosocomial transmission, but from transmission in the community, very likely in homeless shelters and hostels for migrant workers. CONCLUSION: No nosocomial transmission of TB was identified among the patients included during the study period. Systematic RFLP analysis using hospital-based sampling can detect the spread of TB in specific environments in the community where transmission is occurring.

Adult↗
PubMed1997

Nonradioactive single-strand conformation polymorphism analysis for detection of fluoroquinolone resistance in mycobacteria.

A simple, rapid, and nonradioactive method for routine detection of fluoroquinolone resistance in mycobacteria is described. A single-strand conformation polymorphism (SSCP) methodology, based on the use of mini-gels and silver staining of DNA, was optimized for the analysis of denatured DNA products obtained by polymerase chain reaction (PCR) from the gyrA gene involved in fluoroquinolone resistance in mycobacteria. The method was successfully applied to fluoroquinolone-susceptible and -resistant laboratory strains of Mycobacterium smegmatis and to clinical strains of Mycobacterium tuberculosis isolated from patients who developed resistance during the course of fluoroquinolone treatment.

Anti-Infective Agents↗
PubMed1997

[Cutaneous tuberculosis. A study of 4 cases].

INTRODUCTION: The recent increase in the incidence of tuberculosis has led to the return of cutaneous forms of this disease. In addition, diagnosis can now be made rapidly using genoma amplification. CASE REPORT: Four cases of cutaneous tuberculosis are described in nonimmunosuppressed patients: two cases of lupus vulgaris, including one due to Mycobacterium africanum, and two others of gummas, including one associated with tuberculosis verrucosa. The diagnosis was suggested by epidemiological, clinical, histological and immunological findings and confirmed by culture of the bacilli in 3 cases and by genoma amplification in 1. DISCUSSION: These observations illustrate the difficulties encountered in determining the tuberculosis nature of skin lesions. The clinical presentation, differential diagnosis, the pathophysiology of this disease and the new interest in genoma amplification are discussed.

Adult↗
PubMed1995

Role of mutations in DNA gyrase genes in ciprofloxacin resistance of Pseudomonas aeruginosa susceptible or resistant to imipenem.

In Pseudomonas aeruginosa, resistance to imipenem is mainly related to a lack of protein OprD and resistance to fluoroquinolones is mainly related to alterations in DNA gyrase. However, strains cross resistant to fluoroquinolones and imipenem have been selected in vitro and in vivo with fluoroquinolones. We investigated the mechanisms of resistance to fluoroquinolones in 30 clinical strains of P. aeruginosa resistant to ciprofloxacin (mean MIC, >8 micrograms/ml), 20 of which were also resistant to imipenem (mean MIC, >16 micrograms/ml). By immunoblotting, OprD levels were markedly decreased in all of the imipenem-resistant strains. Plasmids carrying the wild-type gyrA gene (pPAW207) or gyrB gene (pPBW801) of Escherichia coli were introduced into each strain by transformation. MICs of imipenem did not change after transformation, whereas those of ciprofloxacin and sparfloxacin dramatically decreased (25- to 70-fold) for all of the strains. For 28 of them (8 susceptible and 20 resistant to imipenem), complementation was obtained with pPAW207 but not with pPBW801. After complementation, the geometric mean MICs of ciprofloxacin and sparfloxacin (MICs of 0.3 microgram/ml and 0.5 microgram/ml, respectively) were as low as those for wild-type strains. Complementation was obtained only with pPBW801 for one strain and with pPAW207 and pPBW801 for one strain highly resistant to fluoroquinolones. These results demonstrate that in clinical practice, gyrA mutations are the major mechanism of resistance to fluoroquinolones even in the strains of P. aeruginosa resistant to imipenem and lacking OprD, concomitant resistance to these drugs being the result of the addition of at least two independent mechanisms.

Anti-Infective Agents↗
PubMed1995

Sequences of conserved region in the A subunit of DNA gyrase from nine species of the genus Mycobacterium: phylogenetic analysis and implication for intrinsic susceptibility to quinolones.

The sequences of a conserved region in the A subunit of DNA gyrase corresponding to the quinolone resistance-determining region were determined for nine mycobacterial species and were compared. Although the nucleotide sequences were highly conserved, they clearly differentiated one species from another. The results of the phylogenetic analysis based on the sequences of the quinolone resistance-determining regions were compared with those provided by the 16S rRNA sequences. Deduced amino acid sequences were identical within the nine species except for amino acid 83, which was frequently involved in acquired resistance to quinolones in many genera, including mycobacteria. The presence at position 83 of an alanine for seven mycobacterial species (M. tuberculosis, M. bovis BCG, M. leprae, M. avium, M. kansasii, M. chelonae, and M. smegmatis) and of a serine for the two remaining mycobacterial species (M. fortuitum and M. aurum) correlated well with the MICs of ofloxacin for both groups of species, suggesting the role of this residue in intrinsic susceptibility to quinolones in mycobacteria.

4-Quinolones↗
PubMed1994

Amplification and nucleotide sequence of the quinolone resistance-determining region in the gyrA gene of mycobacteria.

Chromosomal DNA of different species of mycobacteria, Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacterium avium and Mycobacterium smegmatis, has been submitted to polymerase chain reaction using two oligonucleotide primers highly homologous to DNA sequences flanking the quinolone resistance-determining region in the gyrA gene of Escherichia coli and Staphylococcus aureus. For each of these mycobacterial species, a 150-bp DNA fragment hybridizing with an intragenic probe of the gyrA gene of E. coli K12 was obtained. The nucleotide sequences of the 108-bp fragments amplified from M. tuberculosis and M. avium were determined. The two sequences were 87% homologous. Except for one residue, their deduced amino acid sequences were identical and shared 67% homology with the quinolone resistance-determining region of the gyrase A subunits of E. coli and S. aureus. Sequencing of the 108-bp fragment amplified from an in vitro mutant of M. avium, highly resistant to fluoroquinolones, showed a point mutation leading to the substitution of Ala for Val at a position corresponding to residues involved in quinolone resistance in E. coli and S. aureus, i.e. Ser 83 for E. coli and Ser 84 for S. aureus.

Amino Acid Sequence↗
PubMed1994

Selection of a gyrA mutant of Mycobacterium tuberculosis resistant to fluoroquinolones during treatment with ofloxacin.

A strain of Mycobacterium tuberculosis resistant to ofloxacin was selected in a patient with a long history of multidrug-resistant tuberculosis eventually treated by ofloxacin combined with other second-line drugs. A mutation in the gyrA gene was hypothesized to be the mechanism of acquired resistance to ofloxacin in this strain. Chromosomal DNA of strains MTB1, isolated before treatment and susceptible to ofloxacin (MIC, 1 microgram/mL), and MTB2, isolated during treatment and resistant to ofloxacin (MIC, 32 micrograms/mL), was amplified by polymerase chain reaction (PCR) using two oligonucleotide primers highly homologous to DNA sequences flanking the quinolone resistance-determining region in gyrA of mycobacteria. Comparison of the nucleotide sequences of the 150-bp fragments obtained by PCR revealed a point mutation in MTB2 leading to the substitution of histidine for aspartic acid at a position corresponding to residues involved in quinolone resistance in Escherichia coli (Asp87), Staphylococcus aureus (Glu88), and Campylobacter jejuni (Asp90).

Adult↗
PubMed1994

Characterization of mutations in Mycobacterium smegmatis involved in resistance to fluoroquinolones.

Fluoroquinolone-resistant mutants of Mycobacterium smegmatis have been obtained in vitro by using ofloxacin as a selecting agent. Two types of mutants were identified according to their quinolone resistance patterns. Type 1 showed a low level of resistance to ofloxacin (MIC of 8 micrograms/ml), whereas a high level of resistance to this drug (MICs of 32 to 64 micrograms/ml) characterized type 2. By using two oligonucleotide primers homologous to DNA sequences flanking the quinolone resistance-determining region (QRDR) in the gyrA gene of Escherichia coli and Staphylococcus aureus, a 150-bp DNA fragment was obtained by PCR amplification from total DNA of two wild-type and five mutant strains of M. smegmatis. The nucleotide sequences of the amplified fragments were determined. The deduced amino acid sequence from the wild-type strains showed ca. 79% similarity with the QRDR in the gyrase A subunit from other gram-positive and gram-negative bacteria. The DNA sequences obtained from the fluoroquinolone-resistant mutants of M. smegmatis exhibited nucleotide modifications compared with the wild-type QRDR. The QRDR from type 1 mutants had a C-T or an A-G transition leading to a change from Ala-83 to Val or Asp-87 to Gly, respectively. The QRDR from type 2 mutants had a Val-83 mutation or both Val-83 and Gly-87 mutations detected in the type 1 mutants. These results suggest that point mutations in the QRDR of the mycobacterial gyrA gene are responsible for acquired quinolone resistance in M. smegmatis.

Amino Acid Sequence↗
PubMed1994

Risk factors and clinical relevance of nosocomial maxillary sinusitis in the critically ill.

The incidence of infectious maxillary sinusitis (IMS) and its clinical relevance was prospectively studied in 162 consecutive critically ill patients who were mechanically ventilated for a period longer than 7 d. All had a paranasal computed tomographic (CT) scan within 48 h of admission and were divided into three groups according to the radiologic aspect of their maxillary sinuses: Group 1 = normal maxillary sinuses (n = 40), Group 2 = maxillary mucosal thickening (n = 26), Group 3 = radiologic maxillary sinusitis (RMS) defined as the presence of an air fluid level and/or opacification of maxillary sinuses (n = 96). Group 1 patients were randomized between nasal and oral endotracheal intubation with a gastric intubation performed via the same route and had a second paranasal CT scan 7 d later. Endotracheal and gastric tubes were left in their original position in Group 2 patients and a second paranasal CT scan was performed 7 d later. All patients of Group 3 underwent a transnasal puncture for bacteriologic analysis of maxillary sinus content. Forty-five spontaneously breathing patients served as a control group. In all patients with RMS, the occurrence of bronchopneumonia (BPN) was prospectively assessed for 7 d following the initial CT scan. Upon inclusion, only 25% of the patients had normal maxillary sinuses whereas all patients in the control group had normal paranasal CT scans. After 7 d, 46% of Group 2 patients had evidence of RMS. Risk factors for RMS were nasal placement and duration of endotracheal and gastric intubation.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

[Bacteriological diagnosis of tuberculosis].

Microscopic examination and culture are still today essential elements of the bacteriological diagnosis of tuberculosis. Microscopic examination of a Ziehl fuchsin or auramine stained specimen allows detection of most strains in less than an hour. Culture on Löwenstein-Jensen medium is more sensitive than the microscopic examination and is required for identification and to measure sensitivity to antibiotics. Mycobacterium colonies, generally the causal agent in tuberculosis, usually grow within 28 days and are easily recognized by their "cauliflower" aspect. The niacin test is used for formal identification. Currently, radiometric respirometry allows detection of M. tuberculosis growth and provides antibiotic sensitivity results more rapidly, usually within 10 days. Use of this technique is however limited because the culture medium contains radioactive carbon. Genetic probes are on the other hand quite easy to use and allow identification of cultured bacteria in only a few hours. After polymerization chain reaction gene amplification, M. tuberculosis strains can be detected directly in the specimen within 2 or 3 hours, but in practice, this method has not become a routine laboratory technique, particularly due to lack of sufficient specificity and sensitivity. No other serologic tests are currently reliable enough for the diagnosis of tuberculosis. For cases with low-count specimens, there still is no reliable "on-the-spot" diagnostic test.

Bacteriological Techniques↗
PubMed1993

Novel gyrA point mutation in a strain of Escherichia coli resistant to fluoroquinolones but not to nalidixic acid.

We have previously described a clinical isolate of Escherichia coli (Q2) that is highly resistant to fluoroquinolones (MIC of ciprofloxacin, 16 micrograms/ml) but susceptible to nalidixic acid (MIC of nalidixic acid, 4 micrograms/ml) (N. Moniot-Ville, J. Guibert, N. Moreau, J.F. Acar, E. Collatz, and L. Gutmann, Antimicrob. Agents Chemother. 35:519-523, 1991). Transformation of strain Q2 with a plasmid carrying the wild-type gyrA gene from E. coli K-12(pAFF801) resulted in a 32-fold decrease in the MIC of ciprofloxacin, suggesting that at least one mutation in gyrA was involved in the resistance of Q2. Intragenic gyrA fragments of 668 and 2,500 bp from strain Q2 were amplified by the polymerase chain reaction. We sequenced the 668-bp fragment and identified a single novel point mutation (transition from G to A at position 242), leading to an amino acid substitution (Gly-81 to Asp) in the gyrase A subunit. We constructed hybrid plasmids by substituting either the 668-bp fragment or the 2,500-bp fragment from Q2 DNA, both of which contained the gyrA point mutation, for the corresponding fragments in wild-type gyrA (2,625 bp) of E. coli K-12. When introduced into E. coli KNK453 (gyrA temperature sensitive), both plasmids conferred an eightfold increase in the MIC of ciprofloxacin, but only a twofold increase in the MIC of nalidixic acid. When introduced into E. coli Q2, neither plasmid conferred any change in the MICs of ciprofloxacin or nalidixic acid, suggesting that only the point mutation found in gyrA was involved in the resistance that we observed.

Anti-Infective Agents↗
PubMed1993

Mechanisms of resistance to quinolones.

Mechanisms of resistance to the quinolones have been described for several bacterial species, but mainly for Escherichia coli and Staphylococcus aureus. Two principal mechanisms have been described: 1) alteration of the DNA gyrase, which is the target site of the quinolones; and 2) diminished accumulation in the cell as a result of either decreased uptake or increased efflux. Alteration of DNA gyrase is usually the result of a mutation in the gyrA, or more rarely, the gyrB gene. All substitutions in subunit A of the gyrase are located in the 67 to 106 amino-acid domain and are clustered around Ser-83 in E. coli and Ser-84 in S. aureus. A decrease in uptake has been described for Gram-negative bacteria such as Enterobacteriaceae and Pseudomonas aeruginosa. It has almost always been correlated with a modified electrophoretic profile of outer membrane proteins of the quinolone-resistant mutants. In E. coli, a decrease in OmpF seemed to be linked to the activation of the micF operon in most of the mutants described. These mutants were cross-resistant to unrelated antibiotics, such as trimethoprim, chloramphenicol, tetracycline, and some beta-lactams. In all these mutants the normal or enhanced efflux of quinolones increased the level of resistance. Enhanced efflux has been described as the second mechanism of resistance in S. aureus. Acquired resistance to the quinolones was thought, until recently, to result from chromosomal mutation. Plasmid-mediated resistance associated with an enhanced efflux has been described in S. aureus, but this needs to be confirmed. When a high level of resistance is observed, 2 or 3 mechanisms may be involved.(ABSTRACT TRUNCATED AT 250 WORDS)

4-Quinolones↗