Identification and Clinical Significance of Helcococcus kunzii in Human Samples.

From 2008 to 2013, 39 Helcococcus kunzii strains were collected from human clinical specimens (79% from foot ulcers), and 85% of the 39 patients were infected. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry and molecular methods accurately identified all isolates. This large study of clinical observations confirms the potential pathogenic role of H. kunzii.

Identification of clinical Streptococcus pneumoniae isolates among other alpha and nonhemolytic streptococci by use of the Vitek MS matrix-assisted laser desorption ionization-time of flight mass spectrometry system.

Discrimination between Streptococcus pneumoniae and its close relatives of the viridans group is a common difficulty in matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry-based identification. In the present study, the performances of the Vitek MS MALDI-TOF mass spectrometry system were assessed using 334 pneumococci, 166 other S. mitis group streptococci, 184 non-S. mitis group streptococci, and 19 related alpha- and nonhemolytic aerobic Gram-positive catalase-negative coccal isolates. Pneumococci had been identified by means of optochin susceptibility and bile solubility or serotyping, and other isolates mainly by use of RapidID32 Strep strips. In case of discordant or low-discrimination results, genotypic methods were used. The sensitivity of the Vitek MS for the identification of S. pneumoniae was 99.1%, since only three bile-insoluble isolates were misidentified as Streptococcus mitis/Streptococcus oralis. Conversely, two optochin-resistant pneumococci were correctly identified (specificity, 100%). Three Streptococcus pseudopneumoniae isolates were also correctly identified. Among nonpneumococcal isolates, 90.8% (n = 335) were correctly identified to the species or subspecies level and 2.4% (n = 9) at the group level. For the remaining 25 isolates, the Vitek MS proposed a bacterial species included in the list of possible species suggested by genotypic methods, except for 4 isolates which were not identified due to the absence of the species in the database. According to our study, the Vitek MS displays performance similar to that of the optochin susceptibility test for routine identification of pneumococcal isolates. Moreover, the Vitek MS is efficient for the identification of other viridans group streptococci and related isolates, provided that the species are included in the database.

National multidrug-resistant bacteria (MDRB) surveillance in France through the RAISIN network: a 9 year experience.

BACKGROUND: In the mid-1990s, the prevalence rate of multidrug-resistant bacteria (MDRB) in French hospitals was high and control of MDRB spread then became a major priority in the national infection control programme (ICP). METHODS: To evaluate the impact of the ICP, a national coordination of MDRB surveillance was set up in 2002. Data were collected 3 months a year in healthcare facilities (HCFs) on a voluntary basis. All clinical specimens of methicillin-resistant Staphylococcus aureus (MRSA) and extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBLE) were prospectively included. Incidences per 1000 patient days (PDs) were calculated and trends in incidence from 2003 to 2010 were assessed. RESULTS: Participation in the surveillance increased from 478 HCFs in 2002 to 933 in 2010. In 2010, MRSA incidence was 0.40/1000 PDs: 1.14 in intensive care units (ICUs), 0.48 in acute care facilities (ACFs) and 0.27 in rehabilitation and long-term care facilities (RLTCFs). ESBLE incidence was 0.39/1000 PDs: 1.63 in ICUs, 0.46 in ACFs and 0.23 in RLTCFs. MRSA incidence significantly decreased from 0.72/1000 PDs in 2003 to 0.41/1000 PDs in 2010 (P<10(-3)); in contrast, ESBLE incidence significantly increased from 0.17/1000 PDs to 0.48/1000 PDs (P<10(-3)). The most prevalent ESBLE were Enterobacter aerogenes (34%) and Escherichia coli (25%) in 2003 and E. coli (60%) and Klebsiella pneumoniae (18%) in 2010. CONCLUSION: These results demonstrate the positive impact of the national ICP on MRSA rates. In contrast, ESBLE incidence, especially ESBL-producing E. coli, is increasing dramatically and represents a serious threat for hospitals and for the community that deserves specific control actions.

Performances of the Vitek MS matrix-assisted laser desorption ionization-time of flight mass spectrometry system for rapid identification of bacteria in routine clinical microbiology.

Rapid and cost-effective matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS)-based systems will replace conventional phenotypic methods for routine identification of bacteria. We report here the first evaluation of the new MALDI-TOF MS-based Vitek MS system in a large clinical microbiology laboratory. This system uses an original spectrum classifier algorithm and a specific database designed for the identification of clinically relevant species. We have tested 767 routine clinical isolates representative of 50 genera and 124 species. Vitek MS-based identifications were performed by means of a single deposit on a MALDI disposable target without any prior extraction step and compared with reference identifications obtained mainly with the VITEK2 phenotypic system; if the identifications were discordant, molecular techniques provided reference identifications. The Vitek MS system provided 96.2% correct identifications to the species level (86.7%), to the genus level (8.2%), or within a range of species belonging to different genera (1.3%). Conversely, 1.3% of isolates were misidentified and 2.5% were unidentified, partly because the species was not included in the database; a second deposit provided a successful identification for 0.8% of isolates unidentified with the first deposit. The Vitek MS system is a simple, convenient, and accurate method for routine bacterial identification with a single deposit, considering the high bacterial diversity studied and as evidenced by the low prevalence of species without correct identification. In addition to a second deposit in uncommon cases, expanding the spectral database is expected to further enhance performances.

Pseudomonas aeruginosa may accumulate drug resistance mechanisms without losing its ability to cause bloodstream infections.

In this study, we systematically investigated the resistance mechanisms to beta-lactams, aminoglycosides, and fluoroquinolones of 120 bacteremic strains of Pseudomonas aeruginosa. Pulsed-field gel electrophoresis genotyping showed that 97 of these strains were represented by a single isolate, 10 by 2 and 1 by 3 clonally related isolates, respectively. Seventy-five percent (90 out of 120) of the bacteremic P. aeruginosa strains displayed a significant resistance to one or more of the tested antimicrobials (up to 11 for 1 strain). These strains were found to harbor a great diversity of resistance mechanisms (up to 7 in 1 strain), leading to various levels of drug resistance. Interestingly, 11 and 36% of the isolates appeared to overproduce the MexAB-OprM and MexXY-OprM efflux systems, respectively. Altogether, our results show that P. aeruginosa may accumulate intrinsic (overproduction of cephalosporinase AmpC, increased drug efflux, fluoroquinolone target mutations, and deficient production of porin OprD) and exogenous (production of secondary beta-lactamases and aminoglycoside-modifying enzymes) resistance mechanisms without losing its ability to generate severe bloodstream infections. Consequently, clinicians should be aware that multidrug-resistant P. aeruginosa may remain fully pathogenic.

Retrospective analysis of multidrug-resistant Acinetobacter baumannii strains isolated during a 4-year period in a university hospital.

OBJECTIVE: To describe the epidemiology of Acinetobacter baumannii infection during 2000-2003 and to determine whether the multidrug-resistant strains were already present in our Toulouse hospital before the 2003 French national outbreak. DESIGN: Descriptive molecular and clinical epidemiologic study of A. baumannii isolates using a combination of antibiotyping and pulsed-field gel electrophoresis (PFGE). SETTING: A 1,000-bed university hospital in Toulouse, France. METHODS: All clinical samples that had tested positive for A. baumannii between January 1, 2000, and December 31, 2003, were collected. Multidrug-resistant isolates of A. baumannii were then submitted to PFGE, and clinical characteristics of the source patients were noted. RESULTS: A total of 1,277 A. baumannii samples were identified, 791 of which had not been previously identified; 148 were positive for multidrug-resistant strains. These strains were more likely to have been isolated in the intensive care unit (ICU) than were susceptible strains (P<.001; relative hazard, 3.77). The positive clinical samples were of various types (eg, nonprotected respiratory samples [43%] and blood [5%]), but no difference in type of source was seen between resistant and susceptible strains. A simultaneous analysis of pulsotypes and antibiotypes proved that the outbreak in the ICU in 2003 could be linked to an initially endemic clone that had been isolated in 2001. Furthermore, a second clone responsible for an extended-spectrum beta -lactamase phenotype was sporadically present in our institution. Although the strains isolated in the burn unit were also genetically related one to another, the specific responsible clone only appeared in 2003. CONCLUSION: Several multidrug-resistant clones can coexist endemically for several years and can be detected during an outbreak by close survey of epidemic sources.

Risk of emergence of Pseudomonas aeruginosa resistance to beta-lactam antibiotics in intensive care units.

OBJECTIVE: The emergence of Pseudomonas aeruginosa resistance to antimicrobial drugs is frequent in intensive care units and may be correlated with the use of some specific drugs. The purpose of our study was to identify a relationship between the use of various beta-lactam antibiotics and the emergence of resistance and to characterize the mechanism of resistance involved. DESIGN: We conducted an open prospective study over a 3-yr period by including all patients in whom P. aeruginosa had been isolated from one or more specimens: bronchial aspiration, blood cultures, catheters, and urinary cultures. SETTING: General intensive care unit. PATIENTS: One hundred and thirty-two intensive care unit patients. INTERVENTIONS: The antibiotics studied were amoxiclav, piperacillin-tazobactam, cefotaxime, ceftazidime, cefepim, and imipenem. The mechanisms of resistance studied were production of penicillinase or cephalosporinase, nonenzymatic mechanisms, and loss of porin OprD2. Analysis was performed using Cox proportional-hazard regression with time-dependant variables. MEASUREMENTS AND MAIN RESULTS: Forty-two strains became resistant, 30 to one antibiotic, nine to two, and three to three, leading to the study of 57 resistant strains. Imipenem (hazard ratio 7.8; 95% confidence interval, 3.4-18.1), piperacillin-tazobactam (hazard ratio 3.9; 95% confidence interval, 1.3-11.9), and cefotaxim (hazard ratio 9.3; 95% confidence interval, 2.9-30.2) were strongly linked to the emergence of resistance. The use of imipenem (p<.0001) was associated with the loss of porin OprD2. Thirty-six strains from nine patients, assayed by pulsed-field gel electrophoresis, showed that for any one patient, all the strains were genetically related. CONCLUSIONS: Our results show that there is a high risk of the emergence of drug resistance during treatment with cefotaxime, imipenem, and piperacillin-tazobactam. This has to be taken into account in the therapeutic choice and in the patient's surveillance.

A nosocomial outbreak of Acinetobacter baumannii isolates expressing the carbapenem-hydrolysing oxacillinase OXA-58.

OBJECTIVE: The aim of this study was to analyse the spread of the bla(OXA-58) gene as a source of carbapenem resistance in Acinetobacter baumannii in the burns unit of a university hospital in Toulouse in 2003-2004. METHODS: Six carbapenem-resistant A. baumannii isolates from six patients, and a carbapenem-resistant environmental A. baumannii isolate were collected in the burns unit of the Rangueil hospital (Toulouse). Susceptibility tests were carried out by disc diffusion and agar dilution methods. The detection of the bla(OXA-58) gene was conducted by PCR followed by sequence analysis. Plasmids were extracted and hybridized with a probe specific for bla(OXA-58). DNA fingerprints were obtained by pulsed-field gel electrophoresis of ApaI- or SmaI-digested chromosomal DNA of the tested strains. RESULTS: The multidrug-resistant clinical isolates had a similar 30 kb plasmid that encoded the carbapenem-hydrolysing beta-lactamase OXA-58. These isolates were clonally related. The unrelated environmental carbapenem-resistant A. baumannii isolate had a similar bla(OXA-58)-carrying plasmid, suggesting spread of this gene. CONCLUSIONS: A novel oxacillinase was the source of carbapenem resistance in the A. baumannii isolates. Its gene was plasmid-, but not integron-borne.

Bactericidal properties of group IIa secreted phospholipase A(2) against Pseudomonas aeruginosa clinical isolates.

It has been shown that human group IIa secreted phospholipase A(2) (sPLA(2)), found at high levels in inflammatory fluids, displays direct bactericidal properties against Gram-positive bacteria, while activity against Gram-negative bacteria requires the complement system or additional co-factors produced by neutrophils. Pseudomonas aeruginosa, an increasingly prevalent opportunistic human pathogen, is the most common Gram-negative rod found in cystic fibrosis lung infections, where it is associated with an inflammatory environment. Because murine intestinal group II sPLA(2) produced by Paneth cells has been shown to be directly bactericidal against Gram-negative bacteria, IIa sPLA(2) activity against P. aeruginosa clinical isolates was evaluated and provides the first evidence that the enzyme can be fully bactericidal in a concentration- and time-dependent manner against Gram-negative rods. Furthermore, it was demonstrated that these bactericidal properties were unaffected by high protein and salt concentrations, as observed in cystic fibrosis secretions, and that bacterial killing paralleled phospholipid hydrolysis. Finally, no cytotoxicity was observed when IIa sPLA(2) was incubated with human pulmonary cells, highlighting its potential use to synergize bactericidal antibiotics by promoting sublethal alterations of the bacterial cell wall.

Characteristics of French methicillin-resistant Staphylococcus aureus isolates with decreased susceptibility or resistance to glycopeptides.

According to the French Society of Microbiology, Staphylococcus aureus isolates are suspected to have decreased susceptibility to glycopeptide(s) when at least one colony is able to grow from an inoculum of 10 microL of 2 McFarland bacterial suspension plated on Mueller-Hinton agar containing 5 mg/L teicoplanin and incubated for 48 h at 35-37 degrees C. We analysed 89 methicillin-resistant S. aureus isolates (MRSA), collected in 2000-2001 from 24 hospitals located in 18 French cities, which were able to grow on this selective medium. These isolates were distributed into six groups on the basis of their glycopeptide resistance phenotypes: (A) glycopeptide susceptible (GSSA, 21 isolates); (B) heterogeneous teicoplanin intermediately resistant (hetero-TISA, 24 isolates); (C) heterogeneous and intermediately resistant to both glycopeptides, teicoplanin and vancomycin (hetero-GISA, six isolates); (D) heterogeneous vancomycin intermediately resistant/teicoplanin intermediately resistant (hetero-VISA/TISA, 30 isolates); (E) GISA (four isolates); (F) TISA (four isolates). Despite the persistent decrease in gentamicin-resistant MRSA isolates in French hospitals since 1993, their prevalence is very high in groups D, E and F. Moreover, most of the group C, D and E isolates exhibiting decreased susceptibility to both glycopeptides belong to the same major SmaI genotype, which has been detected in Europe since at least 1989.

Colonization of the oral cavity by Candida species: risk factors in long-term geriatric care.

The population of elderly people in hospitals for long-term geriatric care presents many risk factors for nosocomial infection by Candida species. The aim of this work was to reduce the risk of C. albicans nosocomial infections starting from colonization of the oral cavity. The population of concern was the patients in long-stay geriatrics units; a sample of 110 people was selected by drawing lots. The clinical and biological parameters of each patient included in the study were recorded. The oral cavity was colonized by Candida spp in 67% of cases. The distribution of the strains showed that C. albicans was the most frequently identified strain, followed by C. glabrata; of the 73 patients with at least one strain of Candida spp., 47 had a clinically diagnosed candidiasis (64.4%). The wearing of dentures was not statistically linked with the development of oral candidiasis. Detecting which patients have been colonized, identifying the risk factors and applying preventive measures should reduce the probability of elderly people falling into the vicious circle of infection-malnutrition-immune-depression.

[Epidemiology of nosocomial infections after cataract surgery and role of the Infection Control Committee in prevention]. / Epidémiologie des infections nosocomiales après opération de la cataracte et rôle du CLIN dans la prévention.

Despite of a low incidence (3 for 1,000), post-cataract surgery endophthalmitis remains a serious complication with a poor prognosis. The nosocomial definition is almost always present despite the endogenous origin; the latter is associated with the risk prone operative procedure, and the presence of numerous normal flora on skin and conjunctiva. Within this context, the incriminated bacteria are coagulase-negative Staphylococcus. Micro-outbreaks of postoperative Pseudomonas aeruginosa endophthalmitis have an exogenous origin. In order to prevent these nosocomial infections, the role of the "infection control committee" and the operational hygiene team is very important: 1) survey of new infectious cases and of the hospital environment; 2) infection control measures about pre and intra operative preparation of the patient; 3) operating room maintenance.