Biomedical subjects
Publications and source records attributed to J Etienne.
From 29 June to July 1998, four cases of legionnaires disease in British citizens were reported to the Reseau National de Sante Publique (RNSP) by the statutory notification system (declaration obligatoire (DO)) and by theEuropean Surveillance Scheme for
We report a case of Staphylococcus caprae bone and joint infection, that illustrate difficulties to diagnose coagulase-negative staphylococci (CNS) orthopedic surgery infections, specially following implantation of prostheses. Four of 5 strains successivelly isolated from deep and/or peri-operative specimens during late infection after total hip replacement (THR) have been identified, using commercial systems and conventionnal tests, as S. caprae. Identity of biochemical profile, antibiotype and pulsotype of the 4 isolates confirmed the pathogenicity of this animal CNS, rarely described as a human pathogen. Analysis of the 24 S. caprae human cases previously described evidence a relation ship between this bacteria and bone and joint infections, with implantation of prosthetic material as supplementary risk factor. S. caprae, whose major identification criteria are resumed, may have previously been misidentified as some similar CNS; this bacteria is probably part of our normal flora but may be recognized as an opportunistic pathogen, responsible for both nosocomial and community acquired infections.
Analytical evaluation three models of blood glucose meters (Glucometer IV (Bayer), Glucotrend (Roche Diagnostics) and One touch (Life Scan, Johnson & Johnson)) were compared within a central laboratory to an analyzer (Hitachi 747, Boehringer-Mannheim, Grenoble, France) and to a photometric device (HemoCue B-glucose). Each glucose meters met the criteria of within-run precision defined by the Association Française d'Etudes du Diabète et des Maladies Métaboliques (ALFEDIAM). Furthermore, HemoCue B-glucose met criteria defined by the French Society of Biological Chemistry (SFBC). In the study realized within laboratory, 28 to 66% of glucose tests realized with meters differed by more than 10% from plasma values and 43% of glucose assays realized with different meters differed by more than 10% from HemoCue B-glucose value. Within clinical departments, 46 glucose meters from 5 different models were evaluated. Answers to a questionnaire stored in HemoCue B-glucose underlined the weak development of glucose meters quality assurance. Within clinical departments, 65% of glucose values realized with meters differed by more than 10% from reference values. Development in clinical departments of glucose meters quality assurance programs can be monitored by HemoCue B-glucose, and must allowed improvement of glucose meter quality.
BACKGROUND: Infection remains a severe complication after pacemaker implantation. The purpose of our prospective study was to evaluate the role of the local bacteriologic flora in its occurrence. METHODS AND RESULTS: Specimens were collected at the site of implantation for culture from the skin and the pocket before and after insertion in a consecutive series of patients who underwent elective permanent pacemaker implantation. Microorganisms isolated both at the time of insertion and of any potentially infective complication were compared by using conventional speciation and ribotyping. There were 103 patients (67 men and 36 women) whose age ranged from 16 to 93 years (mean+/-SD, 67+/-15). At the time of pacemaker implantation, a total of 267 isolates were identified. The majority (85%) were staphylococci. During a mean follow-up of 16.5 months (range, 1 to 24), infection occurred in four patients (3.9%). In two of them, an isolate of Staphylococcus schleiferi was recognized by molecular method as identical to the one previously found in the pacemaker pocket. In one patient, Staphylococcus aureus, an organism that was absent at the time of pacemaker insertion, was isolated. In another patient, a Staphylococcus epidermidis was identified both at the time of pacemaker insertion and when erosion occurred; however, their antibiotic resistance profiles were different. CONCLUSIONS: This study strongly supports the hypothesis that pacemaker-related infections are mainly due to local contamination during implantation. S schleiferi appears to play an underestimated role in infectious colonization of implanted biomaterials and should be regarded as an important opportunistic pathogen.
We cloned and sequenced the -976bp promoter of the rat lipoprotein lipase LPL gene. The sequence was compared with the mouse and human sequences. The homology between the rat and mouse LPL nucleotide sequences was not quite as strong in the promoter sequence as in the coding sequence. Among the 976nt promoter there were 118 divergences, i.e. 11.8%, compared to only 5.6% for the LPL coding region. However, within the 200nt immediately 5' to the transcriptional start site (proximal promoter), the divergence was only 4%. New potential cis-elements (such as CACCC, GATA, GC and GA boxes, IRS, Krox, MEF 2, E-box, CCArGG and 1/2 VDRE) were identified in the rat, mouse or human LPL gene.
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The agr P2 operon in Staphylococcus aureus codes for the elements of a density-sensing cassette made up of a typical two-component signalling system and its corresponding inducer. It is postulated that the autoinducer, a post-translationally modified octapeptide generated from the AgrD peptide, interacts with a receptor protein, coded by agrC, to transmit a signal via AgrA regulating expression of staphylococcal virulence genes through expression of agr RNA III. We show by analysis of PhoA fusions that AgrC is a transmembrane protein, and confirm using Western blotting that a 46 kDa protein corresponding to AgrC is present in the bacterial membrane. This protein is autophosphorylated on a histidine residue only in response to supernatants from an agr+ strain, and can also respond to the purified native octapeptide. A recombinant fusion protein where most of the N-terminal region of AgrC is replaced by the Escherichia coli maltose-binding protein is also autophosphorylated in response to stimulation by agr+ supernatants or purified octapeptide. We conclude that AgrC is the sensor molecule of a typical two-component signal system in S. aureus, and that the ligand-binding site of AgrC is probably located in the third extracellular loop of the protein.
To investigate whether 16S-23S rDNA (rDNA) spacer region length polymorphisms are suitable for the identification of Staphylococcus strains, the 16S-23S rDNA intergenic spacer region lengths of 221 strains belonging to 31 species were studied by using a PCR-based method. Each species presented a specific 16S-23S pattern made of 1-8 fragments ranging from 104-771 bp, with the exception of the species Staphylococcus warnei, Staphylococcus caprae and Staphylococcus piscifermentans, which presented larger or smaller fragments. Very few species showed more than one pattern, Staphylococcus saprophyticus subsp. saprophyticus and Staphylococcus aureus being the most heterogeneous species (five different patterns for eight strains). Five clinical strains that could not be identified at the species level by phenotypical tests were finally identified using this method. Discrimination between some species that showed close patterns (Staphylococcus aureus/Staphylococcus chromogenes/Staphylococcus equorum, Staphylococcus aureus/staphylococcus intermedius, Staphylococcus delphini/Staphylococcus felis, Staphylococcus gallinarum, Staphylococcus delphini/Staphylococcus felis, Staphylococcus vitulus/Staphylococcus auricularis) was further achieved after Dral digestion of the PCR products. Although it does not allow discrimination of subspecies, the use of 16S-23S spacer region length data determined by PCR-mediated amplification is suitable for the identification of the 31 Staphylococcus species tested in this study. The method is rapid, easy and may be a useful tool for the identification of Staphylococcus species in the clinical microbiology laboratory.
Species identification of Legionella in routine laboratory testing is hampered by the lack of highly discriminatory phenotypic tests. Amplification polymorphism of the intergenic 16S-23S spacer regions (ISR) has been previously developed for identification of species within the Legionellaceae [Hookey, J.V., Birtles, R.J. & Saunders, N.A. (1995). J Clin Microbiol 33, 2377-2381], but it did not provide enough resolution to distinguish all members of the bluish-white autofluorescent species and the red autofluorescent group of the Legionellaceae. By choosing new primers that target regions 4 (positions 1521-1541 of Escherichia coli 16S rRNA gene) and 6 (positions 114-132 of E.coli 23S rRNA gene) within the rDNA operon close to the 16S-23S intergenic spacer, 34 profiles were determined among the 79 type and reference strains representing 42 species that were tested. Analysis of the RFLP generated after Hinfl restriction digestion of the PCR products further improved the method, allowing complete discrimination among the species and subspecies of Legionella tested. Twenty-three well-identified strains from unrelated origins belonging to seven species gave amplification patterns identical to that of their type strain. The technique was also tested on 80 field isolates that could not be unequivocally assigned to groups by phenotypic methods. Seventy-two per cent (58/80) of these isolates had a profile identical to that of a type strain, while 27% (22/80) may correspond to new taxa since their ISR-PCR profiles did not match any of the known profiles.
RNAIII from Staphylococcus lugdunensis (RNAIII-sl) in a Staphylococcus aureus agr mutant partially restored the Agr phenotype. A chimeric construct consisting of the 5' end of RNAIII-sl and the 3' end of RNAIII from S. aureus restored the Agr phenotype to a greater extent, suggesting the presence of independent regulatory domains.
Two methods were compared for the analysis of 48 unrelated and epidemiologically related Legionella pneumophila serogroup 1 isolates. These are the infrequent-restriction-site PCR (IRS-PCR) assay with adapters designed for XbaI and PstI restriction sites and the pulsed-field gel electrophoresis (PFGE) analysis determined after DNA restriction with SfiI. Both methods demonstrated a high level of discrimination with a similar capacity for differentiating 23 of the 24 unrelated isolates. PFGE analysis and IRS-PCR assay were both able to identify epidemiologically related isolates of L. pneumophila from three outbreaks. Hence, IRS-PCR assay appears to be a reproducible (intergel reproducibility, 100%) and discriminative (discriminatory index, > or = 0.996) tool for typing of Legionella. Compared to PFGE, however, IRS-PCR presented an advantage through ease of performance and with attributes of rapidity and sensitivity of target DNA.
Random amplified polymorphic DNA (RAPD) was used for the identification of Legionella species. Primer SK2 (5'-CGGCGGCGGCGG-3') and standardized RAPD conditions gave the technique a reproducibility of 93 to 100%, depending on the species tested. Species-specific patterns corresponding to the 42 Legionella species were consequently defined by this method; the patterns were dependent on the recognition of a core of common bands for each species. This specificity was demonstrated by testing 65 type strains and 265 environmental and clinical isolates. No serogroup-specific profiles were obtained. A number of unidentified Legionella isolates potentially corresponding to new species were clustered in four groups. RAPD analysis appears to be a rapid and reproducible technique for identification of Legionella isolates to the species level without further restriction or hybridization.
The presence of staphylococcal superantigenic toxins in the supernatants of liquid cultures was detected by an easy and rapid method assessing the activation of T lymphocytes by cytofluorimetric measurement of CD69 expression. Staphylococcus aureus cells were grown in Eagle's minimum essential medium supplemented with 5% heat-inactivated fetal calf serum. Supernatant fluids from all S. aureus strains producing superantigen-related toxins, including enterotoxins A to E, toxic shock syndrome toxin, and exfoliative toxins A and B, induced CD69 expression in a significantly higher number of T cells than a cutoff of 2%. This CD69 assay might be used for initial detection of superantigens from S. aureus strains isolated in the context of staphylococcal toxemia or related chronic human diseases such as atopic dermatitis or Kawasaki syndrome.
Twenty well-characterized isolates of methicillin-resistant Staphylococcus aureus were used to study the optimal resolution and interlaboratory reproducibility of pulsed-field gel electrophoresis (PFGE) of DNA macrorestriction fragments. Five identical isolates (one PFGE type), 5 isolates that produced related PFGE subtypes, and 10 isolates with unique PFGE patterns were analyzed blindly in 12 different laboratories by in-house protocols. In several laboratories a standardized PFGE protocol with a commercial kit was applied successfully as well. Eight of the centers correctly identified the genetic homogeneity of the identical isolates by both the in-house and standard protocols. Four of 12 laboratories failed to produce interpretable data by the standardized protocol, due to technical problems (primarily plug preparation). With the five related isolates, five of eight participants identified the same subtype interrelationships with both in-house and standard protocols. However, two participants identified multiple strain types in this group or classified some of the isolates as unrelated isolates rather than as subtypes. The remaining laboratory failed to distinguish differences between some of the related isolates by utilizing both the in-house and standardized protocols. There were large differences in the relative genome lengths of the isolates as calculated on the basis of the gel pictures. By visual inspection, the numbers of restriction fragments and overall banding pattern similarity in the three groups of isolates showed interlaboratory concordance, but centralized computer analysis of data from four laboratories yielded percent similarity values of only 85% for the group of identical isolates. The differences between the data sets obtained with in-house and standardized protocols could be the experimental parameters which differed with respect to the brand of equipment used, imaging software, running time (20 to 48 h), and pulsing conditions. In conclusion, it appears that the standardization of PFGE depends on controlling a variety of experimental intricacies, as is the case with other bacterial typing procedures.
A prospective study was conducted on 25 Legionella pneumophila culture-positive and 98 culture-negative bronchoalveolar lavage fluid samples to compare two DNA preparation methods: a rapid modified Chelex-based protocol and a proteinase K method. PCR was found to be more sensitive with the Chelex-based method (P = 0.03). N difference was found concerning the inhibition rate.
Within a 1-year period, six surgical-site infections (SSI) caused by Staphylococcus schleiferi were observed in the department of cardiac surgery of Ignatius Hospital, Breda, The Netherlands. Since outbreaks caused by this species of coagulase-negative staphylococci have not been described before, an extensive environmental survey and a case control study were performed in combination with molecular typing of the causative microorganism in order to identify potential sources of infection. Variability, as detected by four different genotyping methods (random amplification of polymorphic DNA [RAPD], conventional and PCR-mediated ribotyping, and pulsed-field gel electrophoresis [PFGE] of DNA macro restriction fragments), appeared to be limited both among the clinical isolates and among several control strains obtained from various unrelated sources. Among unrelated strains, RAPD and PCR-mediated ribotyping identified two types only, whereas seven different types were identified in a relatively concordant manner by conventional ribotyping and PFGE. The latter two procedures proved to be the most useful tools for tracking the epidemiology of S. schleiferi. Four of the outbreak-related strains were identical by both methods, and two isolates showed limited differences. In the search for a potential source of S. schleiferi infection, two slightly different PFGE types were encountered on several occasions in the nose of a single surgeon. These strains were, however, clearly different from the outbreak type. In contrast, S. schleiferi cultures remained negative for two persons identified on the basis of case control analysis. It was demonstrated that SSI caused by S. schleiferi had a clinical impact for patients comparable to that of a wound infection caused by Staphylococcus aureus. This report describes the first well-documented outbreak of S. schleiferi infection. A source of the outbreak was not detected.
The purpose of this study was to determine the influence of local contractile activity on lipoprotein lipase (LPL) regulation in skeletal muscle. Short-term voluntary run training increased LPL mRNA concentration and LPL immunoreactive mass about threefold in white skeletal muscles of the rat hindlimb (all P < 0.01). Training also increased total and heparin-releasable LPL enzyme activity in white hindlimb muscles and in postheparin plasma (P < 0.05). Training did not enhance LPL regulation in a white muscle that was not recruited during running (masseter). LPL levels were already high in red skeletal muscles of control rats, and training did not result in a further rise. In resting rats, local electrical stimulation of a motor nerve to a predominantly white muscle caused a significant rise in LPL mRNA, immunoreactive mass, and enzyme activity relative to the contralateral control muscle of the same animals (all P < 0.01). Finally, LPL expression was several times greater in a red muscle (soleus) of rats with normal postural activity than rats with immobilized hindlimbs (P < 0.01). In summary, these studies support the hypothesis that local contractile activity is required for increasing LPL expression during exercise training and for maintaining a high level of LPL expression in postural muscles.
Laparoscopic splenectomy is considered to be the "gold-standard" treatment of benign hematologic diseases, with normal or slightly enlarged spleens. Laparoscopic treatment of malignant diseases and splenomegalies remains more controversial. The procedure requires a great surgeon's laparoscopic expertise, appropriate positioning of the patient and trocar insertion, and gentle and meticulous dissection of the spleen. The technique is feasible in 91% of the patients with a 0.9% operative mortality and a postoperative complications rate of 12%. When compared with open splenectomy in retrospective case-controlled studies, the laparoscopic approach includes a longer operative time and higher operative room costs. However, advantages include reduced postoperative hospital stay and faster return to normal activities. Despite scarce reported data, long-term hematologic cure rate seems to be equivalent in patients with idiopathic thrombocytopenic purpura. The accuracy of laparoscopic exploration to detect all accessory spleens is however questioned, and residual postoperative accessory splenic tissues have been observed. Prospective randomized controlled trials comparing short- and long-term results of open and laparoscopic splenectomies are required to confirm definitely the role of laparoscopy in the management of hematologic disorders.