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Biomedical subjects

B de Barbeyrac

Publications and source records attributed to B de Barbeyrac.

At least 37 records · Page 2Linked to original sources

[In vitro activity of a new quinolone, NM 394, against Chlamydia trachomatis].

A study of antimicrobial susceptibility of Chlamydia trachomatis was performed with a new quinolone, NM 394 (Laboratoire Dr. Bouchara). Ten C. trachomatis strains of four different genovars (3G, 4E, 2D, IDv) were grown in confluent McCoy cells on coverslips in multidishes (24 wells). For each C. trachomatis strain, two multidishes were prepared, one for MIC, one for MBC. Three inoculum titrated from 100 to 10000 IFU/ml were inoculated into wells. After incubation 48 h at 37 degrees C, the cultures were fixed in methanol and examined for inclusions after staining with fluorescein-conjugated anti-Chlamydia antibody. The MIC was defined as the lowest antibiotic concentration at which no inclusion formation was observed. For the determination of MBC, the culture media was replaced with sucrose phosphate buffer (2SP) and the cultures were incubated without antibiotic for 48 h and stained. The MBC was the lowest antibiotic concentration at which no inclusion formation was observed after one passage without antibiotic. The increase of the inoculum from 10(3) to 10(5) IFU/ml did not affect significantly the activity of NM 394. All strains were inhibited by 16 mg/l of NM 394. The IC 50 and 90 were 2 and 8 mg/l respectively. The MIC and the MBC values were similar.

Anti-Bacterial Agents↗

Arbitrarily-primed PCR confirms the differentiation of strains of Ureaplasma urealyticum into two biovars.

Fourteen serotypes are currently recognized in the Ureaplasma urealyticum species. These serotypes have been divided into two genomic clusters or biovars by a large number of typing methods. The parvo-biovar includes strains of serotypes 1, 3, 6 and 14 and the T960-biovar, strains belonging to the ten other serotypes. In this study, arbitrarily primed polymerase chain reaction (AP-PCR) has been applied to the analysis of reference strains of the 14 U. urealyticum serotypes. By using two different sets of 10-mer oligonucleotide primers, the method allowed the clear differentiation between the two known biovars of the species. However, further differentiation within a same biovar was only achieved for a few standard strains of the T960-biovar analysed by using a pairwise combination of primers. The reproducibility of AP-PCR profiles was shown on strains tested after repeated subcultures and with different thermal cyclers. Additional experiments were performed on forty isolates of U. urealyticum recovered from subjects of various origins. They confirmed that AP-PCR was able to identify the strains at the biovar level. With reference to the other typing methods, AP-PCR is easy to perform and can be applied to large numbers of strains for epidemiological purposes.

Bacterial Typing Techniques↗

Detection of Chlamydia trachomatis by ligase chain reaction compared with polymerase chain reaction and cell culture in urogenital specimens.

OBJECTIVE: The aim of this study was to evaluate the newly developed ligase chain reaction (LCR) assay for the detection of Chlamydia trachomatis in urogenital specimens using cell culture and Amplicor PCR for comparison. SUBJECTS: Two hundred and eighty patients attending hospital or urban STD clinics (high-risk population, 62 men and 84 women) and obstetric/gynaecology clinics (low-risk population, 134 women) in Bordeaux, France. METHODS: Specimens from men were tested with LCR on urethral swabs and urine, with Amplicor or urine, with cell culture on urethral swabs. Specimens from women were tested with LCR, Amplicor and cell culture on endocervical swabs and with LCR on urine. When the three methods generated different results, the LCR and Amplicor tests were repeated on the remaining samples. Samples with discordant LCR and Amplicor results and a negative culture were further analysed by major outer membrane protein gene omp1-PCR. RESULTS: After analysis of discrepant results, the overall prevalence was 7.5% (21/280) calculated on the basis of an expanded "gold standard" defined as culture positive or LCR plus Amplicor positive or omp1-PCR positive for discrepant results between LCR and Amplicor tests. Of the 21, 20 were detected by LCR, 17 by Amplicor and culture. The specificity of LCR and Amplicor was 99.6%. CONCLUSION: The LCR Chlamydia trachomatis test is a highly sensitive nonculture technique and a good alternative test for the detection of chlamydial infections.

Bacteriological Techniques↗

[Molecular typing by pulsed field gel electrophoresis of Pseudomonas (Burkholderia) cepacia isolated from a nosocomial infection].

Because of the emergence of Pseudomonas (Burkholderia) cepacia, isolated among patients from an intensive care unit of the Hôpital Pellegrin (Bordeaux), strains were studied in the aim to determine the source and the mode of transmission of the bacteria. The study was performed on 44 isolates of P. cepacia from 21 patients (23 strains) or from the environment (16 isolates from ventilators and five from the distilled water used for these ventilators). Susceptibility to 23 antibiotics of these strains and endonuclease restriction polymorphism on their total DNA were determined. Pulsed field gel electrophoresis was performed after digestion of the genomic DNA, using three restriction enzymes. Results obtained by analysis of genotypic characteristics demonstrated the clonal origin of the P. cepacia epidemy in the different intensive care units, but did not localize the source of the infection.

Aminoglycosides↗

Genetic variability among Chlamydia trachomatis reference and clinical strains analyzed by pulsed-field gel electrophoresis.

Pulsed-field gel electrophoresis (PFGE) was applied to Chlamydia trachomatis reference strains representing each of the 18 serovars and to 29 clinical isolates from genital specimens collected in Bordeaux, France, or Malmö, Sweden. Comparison of the fingerprint patterns of the reference strains revealed a high level of polymorphism of the total DNA when SmaI was used (14 profiles), whereas the other enzymes, Sse8387I and ApaI, showed fewer differences. Some serovars, considered to be closely related on the basis of their antigenic determinants located on the major outer membrane protein (MOMP), such as D and Da or I and Ia, were shown to be different after PFGE of their genomic DNAs. However, serovars B and Ba and serovars L2 and L2a had identical patterns after analysis with the three endonucleases. When applied to clinical isolates, which were typed by restriction fragment length polymorphism analysis of the MOMP gene, PFGE allowed the detection of intragenotype polymorphisms and showed the identity of two strains successively isolated from the same patient. This technique seems to be an efficient tool for epidemiological studies when used in addition to serotyping or genotyping by restriction fragment length polymorphism analysis of the MOMP gene.

Bacterial Outer Membrane Proteins↗

Evaluation of the Amplicor Chlamydia trachomatis test versus culture in genital samples in various prevalence populations.

OBJECTIVE: To evaluate a newly developed polymerase chain reaction (PCR) assay, Amplicor C trachomatis for the detection of C trachomatis in genital samples using cell culture for comparison. SUBJECTS: 501 patients (431 women and 70 men) attending an STD clinic in Hôpital Pellegrin (high-risk population) and gynaecological clinics (low-risk population) in Bordeaux, France. METHODS: The genital samples (cervical and urethral) were tested for the presence of C trachomatis using the Amplicor test and using standard cell culture identified by the immunofluorescence test using a monoclonal antibody to C trachomatis. Discrepancies between the results of culture and Amplicor were further analysed by major outer membrane protein gene (omp1)-PCR of the specimens taken in transport media and by direct fluorescent antibody (DFA) staining of elementary bodies in culture transport tubes. RESULTS: After analysis of discrepancies, the revised sensitivity and specificity of PCR were 95.3% and 100% and the positive and negative predictive values were 100% and 99.5%, respectively. CONCLUSION: The present results indicate that the Amplicor assay is rapid, specific and more sensitive than the culture method. This test provides an excellent non-culture method for the detection of C trachomatis in various prevalence populations.

Bacteriological Techniques↗

[In situ hybridization of cells infected by Chlamydia trachomatis].

Hybridization in situ of chlamydia trachomatis allows identification of the bacteria. Moreover exact chlamydia trachomatis localization at tissue level can be demonstrated. It seems to us very important to know for instance in conjunctiva were chlamydia trachomatis are located. We used DNA DIGOXIGENIN labelled plasmidic probe of 503 b.p. Detection of chlamydia trachomatis on paraffin embedded cells becomes possible. Specificity is 100% as plasmidic sequence is not found in other bacteria. Sensitivity seems to be good quality on experimental model. At present time, it is under clinical evaluation. This method could complete PCR for chlamydia detection in case of controversial diagnosis.

Chlamydia Infections↗

Potential improvements in therapeutic options for mycoplasmal respiratory infections.

Macrolides and tetracyclines are the most active antibiotics against mycoplasmas in vitro. In particular, erythromycin has been widely used for the treatment of infections caused by Mycoplasma pneumoniae. However, improvements in therapy are needed to accommodate specific characteristics of the microorganism or unusual manifestations of the illness it causes. Because no rapid microbiological technique is available for the diagnosis of M. pneumoniae infections, empirical treatment must be effective against other relevant pathogens as well. Macrolides and fluoroquinolones have recently undergone substantial development. The newer macrolides have more favorable pharmacokinetic properties and are better tolerated than older agents of this class, while the newer fluoroquinolones exhibit a broadened spectrum of in vitro activity that in some instances includes M. pneumoniae and Streptococcus pneumoniae. Although the existing data on in vivo efficacy are only preliminary, the newer macrolides and fluoroquinolones are promising agents for the treatment of M. pneumoniae infection and, more generally, for that of community-acquired pneumonia.

Anti-Bacterial Agents↗

Detection of Mycoplasma pneumoniae and Mycoplasma genitalium in clinical samples by polymerase chain reaction.

Respiratory tract specimens, 75 bronchoalveolar lavage (BAL) samples and 19 throat swabs from 75 hospitalized adult patients (of whom 55 were seropositive for human immunodeficiency virus) with pulmonary infiltrates, were examined for Mycoplasma pneumoniae and Mycoplasma genitalium by culture and polymerase chain reaction (PCR). Urogenital specimens (103 from 48 male patients with nongonococcal urethritis and 55 female patients with genital infections) were examined for M. genitalium by the same techniques. Cultures of all the samples were negative. The oligonucleotides were chosen from the published nucleotide sequences of the P1 adhesin gene from M. pneumoniae or the MgPa adhesin gene from M. genitalium. The products of PCR were identified by restriction analysis and hybridization. Results of PCR for respiratory specimens were positive for the following mycoplasmas: M. pneumoniae (3 specimens), M. genitalium (1), and both M. pneumoniae and M. genitalium (3). PCR results for eight urogenital samples from male patients were positive for M. genitalium. Differences concerning the restriction sites were observed between respiratory and genital specimens. The major findings of this study are the presence of DNA from M. pneumoniae in respiratory specimens from patients for whom cultures were negative and DNA from M. genitalium in respiratory and urogenital samples.

Adult↗

Evaluation of molecular typing for epidemiological study of Chlamydia trachomatis genital infections.

Molecular typing and serotyping were compared for 150 Chlamydia trachomatis strains isolated from genital sources, belonging to 10 different serovars. Because of the general agreement of the two methods, molecular omp1 genotyping was applied to the epidemiological study of C. trachomatis isolates from genital infections in Bordeaux (France), during a 29-month period. The most prevalent omp1 genotypes were E (51.7%), F (17.3%), D (8.8%), and G (8.4%). Restriction enzyme analysis allowed identification of a serovar D variant (Dv), whereas serovar E strains were homogeneous.

Bacterial Outer Membrane Proteins↗

[Multicenter study of the in vitro sensitivity of genital mycoplasmas to antibiotics].

The in vitro susceptibility of Ureaplasma urealyticum (Uu) and Mycoplasma hominis (Mh) was evaluated in a multicentric study performed in seven hospitals from different geographic areas in France. During a three month period, 324 Uu and 72 Mh clinical isolates were tested using a system ready for use, SIR Mycoplasma (Sanofi Diagnostics Pasteur). For Uu, the percentage of strains intermediate (I) or resistant (R) was as follows: doxycycline (3), minocycline (2.5), lymecycline (6.7), erythromycin (72, most I), josamycin (0.9), clindamycin (88), pristinamycin (0.3), ofloxacin (34, most I). For Mh, the percentage of strains I or R was respectively: doxycycline (2.7), minocycline (5.5), lymecycline (15.2), erythromycin (100), clindamycin (1.4), ofloxacin (2.7), josamycin (0) and pristinamycin (0). Comparable results were observed in the different geographic areas. The frequency of acquired resistances does not justify modifications in the usual treatment of genital mycoplasma infections but leads to monitor their susceptibility to antibiotics.

Clindamycin↗

Isolation of genital mycoplasmas from blood of febrile obstetrical-gynecologic patients and neonates.

In a prospective study, 1156 blood specimens collected from hospitalized febrile obstetrical-gynecologic patients and neonates with suspected sepsis, were inoculated into a conventional biphasic culture medium, Castaneda S and cultures incubated aerobically. 15-24 h later the broth cultures were subcultured to specific media for detection of mycoplasmas. Genital mycoplasmas were isolated in 15 samples (taken from 8 women) and in 2 from 1 neonate. Mycoplasmas and members of the family Enterobacteriaceae were the most frequent significant bacteria isolated from adult specimens. Mycoplasma isolations were associated with either postpartum or postabortum febrile infections in women. Four of the neonates, whose mothers were infected, showed respiratory distress at birth; 1 of them had mycoplasmas in the blood. All febrile states in obstetrical or gynecological patients, and in neonates, should routinely lead to blood cultures for detection of mycoplasmas and ureaplasmas.

Adult↗

Evaluation of bronchoalveolar lavage for the diagnosis of bacterial pneumonia in ventilated patients.

Fourty-six patients requiring mechanical ventilation and suspected of bacterial pneumonia were examined by fiberoptic bronchoscopy. Specimens recovered by bronchoalveolar lavage (BAL) and using a protected specimen brush (PSB) were quantitatively cultured and the results compared. An assessment of the percentage of cells with intracellular organisms present on cytocentrifuged preparations made from lavage fluid was made to evaluate the utility of this method in early diagnosis of pneumonia. BAL cultures made a correct diagnosis in 43 out of 46 patients and detected 10 false-negative cultures of the PSB. With a threshold of more than 3% of cells with intracellular bacteria, direct microscopic examination diagnosed bacterial pneumonia in 36 out of 46 patients and allowed appropriate antibiotherapy to be instituted earlier. These results demonstrate the ability of BAL to diagnose bacterial pneumonia in ventilated patients.

Adult↗

Susceptibility of Bacteroides ureolyticus to antimicrobial agents and identification of a tetracycline resistance determinant related to tetM.

The in-vitro activity of ten antimicrobial agents was evaluated for 28 clinical isolates of Bacteroides ureolyticus, an obligate anaerobe associated with non-gonococcal urethritis. The isolates were characterized by plasmid DNA profile and PAGE protein pattern. All isolates were inhibited at concentrations equal to or lower than the recommended breakpoint concentration for ampicillin (16 mg/l), metronidazole (16 mg/l) and erythromycin (4 mg/l). Twenty-seven isolates were inhibited by less than or equal to 2 mg/l of ciprofloxacin, pefloxacin and ofloxacin. Four isolates were tetracycline-resistant requiring 2-64 mg/l of tetracycline, minocycline or doxycycline for inhibition. In two tetracycline-resistant isolates tetM was demonstrated by dot-blot and Southern hybridizations. These two isolates did not contain a plasmid and had a PAGE protein pattern type III. These data confirm the spread of the tetM determinant in various bacteria of the genital tract.

Anti-Bacterial Agents↗

Typing of Chlamydia trachomatis by restriction endonuclease analysis of the amplified major outer membrane protein gene.

A procedure was developed for characterization of Chlamydia trachomatis strains by using restriction endonuclease analysis of amplified genes of the major outer membrane protein (MOMP). Reference strains of the 15 serovars (A through K and L1 through L3) and clinical isolates were tested. The nucleotide sequences of the MOMP genes of each of the 15 serovars were arbitrarily constructed by using the sequences of the four variable domains known for each serovar and the constant domains of serovar L1. Computer analysis of these sequences indicated that two restriction digestions performed in parallel, one with AluI and the other with IIpaII, followed by HinfI and EcoRI, would allow the theoretical differentiation of 13 serovars. Serovars Ba and L1 presented the same theoretical restriction profile. Our typing method consisted of polymerase chain reaction amplification of a fragment of about 1,200 bp of the MOMP gene, followed by restriction endonuclease digestion with the aforementioned enzymes. From the 15 serovars, we obtained 14 different patterns; 13 profiles were serovar specific, while serovars B and Ba presented the same pattern. Application of this typing method to C. trachomatis strains isolated from clinical material gave the same results as the immunotyping method for 14 of 17 strains. Furthermore, restriction endonuclease analysis detected differences within a serovar. This method seems to be promising for epidemiological studies.

Bacterial Outer Membrane Proteins↗