[Isolation, morphological and biochemical characteristics of gram-positive bacteria metabolizing beta-caryophyllene]. / Isolement, caractères morphologiques et biochimiques de bactéries à gram positif métabolisant le beta-caryophyllène.

Eleven strains of coryneform bacteria were isolated from soil samples by enrichment culture in a mineral medium containing beta-caryophyllene as the sole energy and carbon source. Ten of them could also metabolize longifolene. Numerical taxonomy, based on the use of 147 characteristics, revealed a large diversity. The DNA G + C content was found to be in the range of 62.5-68.3 mol%. Three strains were placed in "The National Collection of Industrial and Marine Bacteria".

Phenotypic heterogeneity of Pseudomonas syringae van Hall.

The study of phenotypic properties of 108 strains of Pseudomonas syringae pv. syringae van Hall isolated from Cherry laurel (50 strains) and various host plants (58 strains) and 53 strains of other pathovars of P. syringae and fluorescent Pseudomonas showed that the majority of the strains (91/108) were clustered in one phenon (phenon 14) containing strains commonly considered as P.s. pv. syringae. The present type strain of P.s. pv. syringae was distantly related to phenon 14. Other pathovars of P. syringae constituted 13 discrete phenons.

Identification and strain differentiation of Mycobacterium species on the basis of DNA 16S-23S spacer region polymorphism.

Amplification of the region separating the genes coding for the two rRNA species 16S and 23S was performed to identify 56 mycobacterial strains, belonging to eleven species: Mycobacterium tuberculosis, M. avium, M. kansasii, M. gordonae, M. abscessus, M. fortuitum, M. xenopi, M. bovis, M. bovis/BCG, M. africanum and M. intracellulare. Reproducible amplification patterns were obtained with most species with the exception of M. kansasii which showed heterogeneity, confirming the existence of a genetically distinct subspecies within this species. In addition, we used the amplified products as target DNA for restriction endonuclease digestion and RAPD (randomly amplified polymorphic DNA) analysis to compare strains of M. abscessus, M. tuberculosis and M. avium. The discriminatory power of these two typing methods was higher than when whole genomic DNA is used as target. Our results demonstrate that the two-step approach to identification and typing on the basis of the hypervariability of 16S-23S spacer region is reliable, rapid and simple, and consequently could be an epidemiological tool in clinical laboratories.

A one-step microbial DNA extraction method using "Chelex 100" suitable for gene amplification.

"Chelex 100" chelating resin has been previously proposed for the rapid extraction of human DNA for polymerase chain reaction. Protocols are given for the rapid extraction of bacterial and viral DNA from cultures or clinical samples. The DNA samples obtained were suitable for use in polymerase chain reaction.

Variations in DNA concentrations significantly affect the reproducibility of RAPD fingerprint patterns.

The influence of the DNA concentration was tested using two different primers and nine DNA samples. Major modifications in the DNA banding pattern were apparent between successive dilutions. Such differences could be explained by concomitant changes in three different molecular conditions: the presence of perfect priming sites, the amplification of rare sites and the existence of mismatch annealing events. At low DNA concentrations (less than 1 pg/microliter), molecular events occurred at random and had a direct consequence on the reproducibility of RAPD profiles. At the appropriate DNA concentration (between 100 ng/microliters and 10 pg/microliters), reproducibility was adequate at a given concentration, but RAPD profiles differed from one dilution to another. These observations demonstrate the usefulness of the bis-benzimide method for quantification of DNA extracts.

Demonstration of Mycobacterium kansasii species heterogeneity by the amplification of the 16S-23S spacer region.

Amplification of the region separating the genes coding for the two rRNA species 16S and 23S was performed with 56 strains of several mycobacterial species, including 21 clinical isolates of Mycobacterium kansasii and the M. kansasii type strain ATCC 12478. On the basis of PCR product profiles, the previously suggested heterogeneity of M. kansasii species was confirmed. Three subgroups were identified; members of the first subgroup showed the same PCR profile as the reference strain. Different profiles were obtained for the two other subgroups. Amplification of the 16S-23S spacer is rapid and simple and, consequently, may be a helpful tool for identification and characterisation of M. kansasii isolates in epidemiological analysis.

Investigation of outbreaks of Enterobacter aerogenes colonisation and infection in intensive care units by random amplification of polymorphic DNA.

During a 4-month period, 41 isolates of Enterobactor aerogenes were cultured from different specimens from a 14-bed intensive care unit (ICU1). These were obtained from 12 patients out of a total of 187 patients admitted to the ICU. Sixteen E. aerogenes isolates were cultured from another ICU (ICU2) 6 months later. Six non-outbreaks associated strains were included as controls and all the isolates were compared by random amplification of polymorphic DNA (RAPD), with three different 10-mer oligonucleotide primers. RAPD fingerprinting with primer AP12h was as discriminatory as the combined results from all three primers and defined 22 different patterns for the 41 isolates from the ICU1. In nine instances, isolates with indistinguishable RAPD patterns were detected in two-to-five patients over a 3-15-day period, suggesting patient-to-patient transmission. During their stay in ICU1, patients harboured one-to-12 distinguishable isolates. Isolates from ICU2 were indistinguishable by RAPD analysis with the three different primers. These findings suggest that the cluster of colonisations and infections in ICU1 was a 'false outbreak', consisting of successive patient-to-patient transmission of different E. aerogenes strains. In contrast, the outbreak on ICU2 probably involved the extensive spread of a single strain.

Epidemiological investigation of Pseudomonas aeruginosa nosocomial bacteraemia isolates by PCR-based DNA fingerprinting analysis.

Between July 1994 and March 1995, 64 isolates of Pseudomonas aeruginosa were implicated in bacteraemia in 25 cancer patients in five wards of two hospitals. These, together with 24 environmental isolates and one isolate from a bacteraemia in a non-cancer patient were examined by three PCR-based DNA fingerprinting methods: random amplified polymorphic DNA (RAPD), enterobacterial-repetitive intergenic consensus (ERIC)-PCR, and 16S-23S spacer region-based RAPD. These methods were reproducible, discriminatory and showed close agreement; all indicated that 47 isolates that had caused bacteraemia in 19 cancer patients were indistinguishable. Seventeen other isolates that had caused bacteraemia in 10 cancer patients were discriminated into eight further groups, and the 24 environmental and non-cancer patient isolates into further distinct groups. No environmental source of the epidemic strain was found, but it was suspected that the outbreak was related to infusion implants.

A cluster of cases of infections due to Aeromonas hydrophila revealed by combined RAPD and ERIC-PCR.

Two polymerase chain reaction (PCR)-based methods were used for epidemiological typing of Aeromonas hydrophila. Random amplification of polymorphic DNA (RAPD) and enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) were applied to an outbreak involving seven patients. The epidemiological situation appeared complex; with the exception of two clinical isolates, all gave unique patterns with both techniques. These methods demonstrated nosocomial transmission in one unit and permitted the study to exclude a common environmental source in the hospital. The coincidental clustering of patients infected with A. hydrophila probably resulted from an increased prevalence of aeromonads in waters during summer, although no single RAPD or ERIC-PCR pattern was found among both clinical and environmental samples. RAPD and ERIC-PCR proved to be effective for the epidemiological study of A. hydrophila strains.

Serratia marcescens nosocomial outbreak due to contamination of hexetidine solution.

During a 10-week period, 16 patients in a neurosurgery intensive care unit were involved in an outbreak of Serratia marcescens. The epidemic strain was found in several flasks of 1:4 diluted hexetidine solution, an antiseptic used for patient mouth washing. Testing of the bactericidal activity of the diluted antiseptic revealed that all the epidemic strains were able to grow in the diluted antiseptic solution. Strains isolated from clinical samples and from the antiseptic solution were compared by random amplification of polymorphic DNA. Epidemiologic typing data implicated the diluted antiseptic solution as the single source of this S. marcescens outbreak.

Use of random amplified polymorphic DNA for epidemiological typing of Stenotrophomonas maltophilia.

We used the technique of random amplification of polymorphic DNA (RAPD) to type 130 isolates of Stenotrophomonas (Xanthomonas) maltophilia, using four arbitrary short primers. Of the 130 isolates, 51 were from the hospital environment, 48 from clinical specimens and 31 were geographically diverse environmental isolates. DNA amplification with the four sets of primers generated 112 RAPD patterns that differed by two or more bands in one of the four primers. Sixteen pairs of isolates were of the same RAPD pattern and some of these pairs represented clinical strains obtained from patients hospitalized at the same time in the same ward. In three patients, two to three strains of S. maltophilia which gave different RAPD fingerprints were isolated on the same day from different specimens. RAPD fingerprinting demonstrated great genomic diversity within the species S. maltophilia and provided an effective method for the study of the epidemiology of both clinical and environmental strains.

[Diagnosis of familial medullary cancers]. / Dépistage des cancers médullaires familiaux.

Medullary thyroid carcinoma (MTC) is hereditary in 20 to 25% of cases. It is inherited as an autosomal dominant trait. MTC can be considered as a sporadic form only after a clinical and biological survey of the two parents, siblings and children of the patient, using pentagastrin stimulation test. The authors have studied 36 patients from 26 families. Hereditary MTC with different clinical features, were discovered in two kindreds. The systematic investigation leads to the discovery of 7 cases in the first family, and of 3 in the second. The treatment of the disease at the first stage of its evolution has been possible when an early diagnosis had been made, such as in the second family.

A multiple-antibiotic resistance-independent active chloramphenicol efflux in an Escherichia coli clinical isolate.

The clinical isolate, Escherichia coli 1941, exhibits high resistance to chloramphenicol and tetracycline (minimum inhibitory concentrations of 512 micrograms/ml). Neither resistance is linked to the large conjugative plasmid present in the strain. The intracellular accumulation of radiolabeled chloramphenicol increased about 9-fold after the addition of the energy uncoupler carbonyl cyanide m-chlorophenol-hydrazone to an E. coli 1941 culture, indicating the presence of an active efflux mechanism. Sequence analysis and expression study suggested that the multiple-antibiotic resistance marRAB locus and the AcrAB drug-efflux pump were not involved in this active efflux of chloramphenicol.

Pseudomonas aeruginosa and cystic fibrosis: intrastrain variability in relation to intensity of immune response and clinical outcome.

Over periods exceeding two years, 63 Pseudomonas aeruginosa strains isolated from three children with cystic fibrosis were characterized using a 154 character auxanogram. Numerical analysis was carried out to establish intrastrain variability in relation to intensity of immune response and clinical outcome. For two patients, strains segregated in two distinct phenons. For the last patient, strains did not cluster. The degree of intrastrain variability during the infection was correlated with an increasing number of precipitins and with an unfavorable clinical outcome. This would suggest that strains which have a singular potential for antigenic variability are more likely to promote a host-bacteria conflict.

A rapid DNA extraction method from culture and clinical samples. Suitable for the detection of human cytomegalovirus by the polymerase chain reaction.

We propose an one-step DNA extraction method suitable for the polymerase chain reaction. This procedure utilizes Chelex 100, a chelating in exchange resin. This technique was compared with a traditional technique (proteinase K lysis, phenol-chloroform extraction and ethanol precipitation) for isolation of human cytomegalovirus DNA from clinical samples. The procedure using Chelex 100 appeared to be a simple and fast extraction method for human cytomegalovirus DNA.

[Identification of coagulase-negative Staphylococci. Significance of miniaturized systems]. / Identification des staphylocoques à coagulase négative. Intérêt des galeries miniaturisées.

To evaluate accuracy of API Staph. and Staph. Ident. systems for identification of coagulase-negative Staphylococci, the authors have tested 235 clinical and environmental isolates. The results obtained were compared with those of conventional method of Kloos and Schleifer. API Staph. and Staph. Ident. correctly identified respectively 64 and 53 p. cent of the strains. Diagnosis ability coefficient (Descamps and Véron) are close. It should be necessary to compare identification obtained using analytical profile index with described characteristics of species.

[Identification of Mycobacterium tuberculosis and Mycobacterium avium by non-radioactive probes: evaluation of the Snap Syngene system]. / Identification de Mycobacterium tuberculosis et des mycobactéries aviaires par sondes non radioactives: évaluation du système Snap Syngène.

We evaluated an alkaline phosphatase-labeled oligonucleotide probe for the rapid identification of Mycobacterium tuberculosis and mycobacteria belonging to the M avium and M intracellulare complex (MAIS). Sixty-two strains of mycobacteria and eight strains belonging to related genera were studied. All M tuberculosis strains hybridized with the tuberculosis probe. All M avium and M intracellulare gave a strong signal with their probes. However the 3 M xenopi strains tested hybridized with all probes for MAIS complex.

[Pulmonary actinomycosis. Apropos of a case]. / Actinomycose pulmonaire. A propos d'un cas.

The authors report a case of pseudo-tumoral thoracic actinomycosis with lysis of a rib in a young man who was a heavy smoker and drinker. This observation is an opportunity to review the usual difficulties of diagnosis in patients with this rare disease occurring most often in a patient who is debilitated with important disease in the teeth and gingival margins. The standard treatment is penicillin G, which leads to a cure within one month.

[Separation by high-performance liquid chromatography of phenylthiohydantoin derivatives of amino acids released by Edman degradation (author's transl)]. / Séparation par chromatographie en phase liquide rapide des dérivés phénylthiohydantoine des amino-acides rencontrés lors de la dégradation d'Edman

The separation by high-performance liquid chromatography of seventeen phenylthiohydantoin derivatives of amino acids released after Edman degradation of proteins is described. Sixteen phenylthiohydantoin derivatives of amino acids are differentiated within 40 min by liquid-liquid partition on a column packed with Micropak CN (moderately polar alkyl nitrile phase bonded on 10mum porous silica microparticles) and gradient elution with hexane=methylene chloride-isopropanol mixtures.