Vancomycin treatment is a risk factor for vancomycin-nonsusceptible Staphylococcus capitis sepsis in preterm neonates.

OBJECTIVES: Multidrug-resistant, vancomycin-nonsusceptible Staphylococcus capitis is an emerging cause worldwide of late-onset sepsis (LOS) in preterm neonates. The pathophysiology and risk factors for S. capitis-related LOS are poorly understood, but we hypothesized that S. capitis LOS follows translocation from the gut microbiota rather than catheter invasion. The objective of this study was to investigate the risk factors of S. capitis LOS and gut colonization. METHODS: We conducted a prospective single-centre cohort study of patients hospitalized in a tertiary-care unit (Lyon, France) from June 2011 to January 2012. S. capitis gut colonization was determined weekly from stool cultures. The determinants of gut colonization and LOS were established by multivariate Cox proportional hazards models. RESULTS: Eighty-three (36.2%) of 229 patients had S. capitis-positive stool culture, and 28 (12.2%) developed S. capitis LOS during hospitalization. Independent risk factors for S. capitis LOS included prior administration of vancomycin independent of a previous LOS episode (hazard ratio 6.44, 95% confidence interval 2.15-19.3, p 0.001) and low birth weight (hazard ratio 0.72 per 100 g increase, 95% confidence interval 0.55-0.95, p 0.02). The prior administration of vancomycin was also an independent risk factor for S. capitis colonization (hazard ratio 3.45, 95% confidence interval 2.07-5.76, p <0.001), particularly in the first week of life and in noncolonized neonates. CONCLUSIONS: Neonates treated with vancomycin are at a higher risk of LOS caused by vancomycin-nonsusceptible S. capitis. The use of vancomycin in neonates must urgently be optimized to limit the selection of vancomycin-nonsusceptible strains, for which alternative antibiotics are lacking.

Comparison of Sofia Legionella FIA and BinaxNOW® Legionella urinary antigen card in two national reference centers.

The Sofia Legionella Fluorescence Immunoassay (FIA; Quidel) is a recently introduced rapid immunochromatographic diagnostic test for Legionnaires' disease using immunofluorescence technology designed to enhance its sensitivity. The aim of this study was to evaluate its performance for the detection of urinary antigens for Legionella pneumophila serogroup 1 in two National Reference Centers for Legionella. The sensitivity and specificity of the Sofia Legionella FIA test were determined in concentrated and nonconcentrated urine samples, before and after boiling, in comparison with the BinaxNOW® Legionella Urinary Antigen Card (UAC; Alere). Compared with BinaxNOW® Legionella UAC, the sensitivity of the Sofia Legionella test was slightly higher in nonconcentrated urine samples and was identical in concentrated urine samples. The specificity of the Sofia Legionella FIA test was highly reduced by the concentration of urine samples. In nonconcentrated samples, a lack of specificity was observed in 2.3 % of samples, all of them resolved by heat treatment. The Sofia Legionella FIA is a sensitive test for detecting Legionella urinary antigens with no previous urine concentration. However, all positive samples have to be re-tested after boiling to reach a high specificity. The reading is automatized on the Sofia analyzer, which can be connected to laboratory information systems, facilitating accurate and rapid reporting of results.

[Primary meningococcal B osteomyelitis and arthritis with multifocal pyomyositis in a child: a case report]. / Ostéoarthrite de hanche primitive et myosite multifocale à méningocoque B chez un enfant de 7 ans.

Neisseria meningitidis is associated with severe invasive infections such as meningitis and fulminant septicemia. Septic arthritis due to N. meningitidis is rare and bone infections have been reported exceptionally. We report the case of a 7-year-old boy who presented with septic arthritis of the right hip associated with a septic location on the pelvis and pyomyositis of the adjacent muscle. Culture of the joint fluid was sterile but universal 16S polymerase chain reaction (PCR) of this fluid revealed group B N. meningitidis. Our patient had never presented any symptoms of meningitis or septicemia and blood cultures were all sterile. Despite appropriate antibiotic treatment, the course of the disease was unusually long and his status did not improve until surgical lavage of the hip was performed. Moreover, MRI imaging showed bilateral hypersignals of the adjacent muscles and revealed an abscess formation in the left gluteus maximus muscle. Presumptive diagnosis bacterial myositis was confirmed by an elevation of creatine phosphokinase in the sera up to 21-fold the normal value but the culture of the abscess, performed 10 days after initiation of antibiotics, was sterile. Despite an initially unfavorable course, this patient's status improved after surgical drainage and he fully recovered 1 month later. This observation illustrates an unusual presentation of invasive meningococcal infection. The respective roles of infection and an inflammatory phenomenon during the course of the disease are discussed. Moreover, this case emphasizes the value of PCR for bacteriological diagnosis of bone and joint infections.

Species identification of staphylococci by amplification and sequencing of the tuf gene compared to the gap gene and by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

Staphylococcal species, notably, coagulase-negative staphylococci (CoNS), are frequently misidentified using phenotypic methods. The partial nucleotide sequences of the tuf and gap genes were determined in 47 reference strains to assess their suitability, practicability, and discriminatory power as target molecules for staphylococcal identification. The partial tuf gene sequence was selected and further assessed with a collection of 186 strains, including 35 species and subspecies. Then, to evaluate the efficacy of this genotyping method versus the technology of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS), the 186 strains were identified using MALDI-TOF-MS (Axima® Shimadzu) coupled to the SARAMIS® database (AnagnosTec). The French National Reference Center for Staphylococci identification method was used as a reference. One hundred and eighty-four strains (98.9%) were correctly identified by tuf gene sequencing. Only one strain was misidentified and one was unidentified. MALDI-TOF-MS identified correctly 138 isolates (74.2%). Four strains were misidentified, 39 were unidentified, five were identified at the group (hominis/warneri) level, and one strain was identified at the genus level. These results confirm the value of MALDI-TOF-MS identification for common species in clinical laboratory practice and the value of the partial tuf gene sequence for the identification of all staphylococcal species as required in a reference laboratory.

[Septic osteomyelitis of the hip after Fusobacterium necrophorum pharyngitis]. / Ostéoarthrite de la hanche dans les suites d'une angine àFusobacterium necrophorum.

CASE REPORT: We report the case of a patient presenting with Lemierre's syndrome with an initial atypical presentation due to a predominant hip localization. Despite the adapted antibiotic treatment and surgical drainage, the patient relapsed. The bone and joint biopsy samples were sterile but the presence of the Fusobacterium necrophorum was demonstrated by PCR. CONCLUSION: Lemierre's syndrome may have different clinical presentations, due to the localizations of septic emboli. Careful attention should be paid to common ENT infections with an unfavorable evolution despite antibiotics. PCR has become a very interesting tool to document bone and joints bacterial infections when cultures are negative.

RISA-HPLC analysis of lung bacterial colonizers of cystic fibrosis children.

Microbiological analysis of sputum samples, from children affected by cystic fibrosis (CF) and showing signs of acute or chronic infections, is routinely performed by culture-dependent approaches involving selective media and biochemical tests. These identification schemes are time-consuming, and may lead to false negative results. The aim of this work was to evaluate the efficacy of a Ribosomal Intergenic Spacer Analysis (RISA) coupled to high performance liquid chromatography (HPLC) for the detection and monitoring of CF lung microbial colonizers including Staphylococcus aureus, Haemophilus influenzae, Pseudomonas aeruginosa, the Burkholderia cepacia complex, Stenotrophomonas maltophilia, and Achromobacter xylosoxidans. These RISA-HPLC analyses were performed over a 10-months period on infants (below 18 months) and children that were or were not yet known to be colonised by P. aeruginosa. The RISA-HPLC profiles were found specific of the patients' microbial communities. A specific P. aeruginosa RISA-HPLC peak corresponding to 550 bp PCR products was recorded, and used to investigate P. aeruginosa persistence through time and after various therapeutic treatments. The RISA-HPLC profiles showed the CF children to be colonized by few bacterial species, and sometimes revealed peaks corresponding to bacterial species that were not detected by the selective plating approaches. Significant RISA-HPLC infra-specific variations were observed for most bacterial colonizers of CF lungs except P. aeruginosa. These species could yield as much as 5 distinct RISA-HPLC peaks, with some of these profiles being strain-specific. RISA-HPLC shows a great potential for revealing colonization by novel emerging pathogens, and for evaluating the efficacy of therapeutic treatments on the global bacterial community of CF lungs.

The application of chromogenic media in clinical microbiology.

Since 1990, a wide range of chromogenic culture media has been made commercially available providing useful tools for diagnostic clinical microbiology. By the inclusion of chromogenic enzyme substrates targeting microbial enzymes, such media are able to target pathogens with high specificity. Examples of target pathogens include Staphylococcus aureus, Streptococcus agalactiae, Salmonella spp. and Candida spp. The inclusion of multiple chromogenic substrates into culture media facilitates the differentiation of polymicrobial cultures, thus allowing for the development of improved media for diagnosis of urinary tract infections and media for the enhanced discrimination of yeasts. The purpose of this review is to provide some insight into how such media work and appraise their utility in routine clinical diagnostics, in comparison with conventional media.

Streptococcus pneumoniae thoracic empyema in children: rapid diagnosis by using the Binax NOW immunochromatographic membrane test in pleural fluids.

AIM: To evaluate an immunochromatographic membrane test for Streptococcus pneumoniae antigen (Binax NOW, Inverness medical France) applied to pleural fluid samples. METHODS: Binax NOW was applied to the pleural fluids of 69 children with thoracic empyema, in comparison with conventional culture and molecular techniques. RESULTS: Binax NOW was positive on all 15 pleural fluid samples that yielded S. pneumoniae in culture, on two samples that yielded S. oralis and S. salivarius in culture and on 34 culture-negative samples. Fifteen of these 34 culture-negative samples were retrospectively tested by PCR methods, and 14 were shown to contain S. pneumoniae DNA. Thus, S. pneumoniae was identified by culture in 22% of samples and by Binax NOW in 69% of samples. CONCLUSION: Binax NOW may thus be useful for rapid diagnosis of S. pneumoniae thoracic empyema.

Low incidence of sepsis due to viridans streptococci in a ten-year retrospective study of pediatric acute myeloid leukemia.

BACKGROUND: Infections remain an important cause of morbidity and mortality in children with acute myeloid leukemia (AML), and particularly viridans group streptococci (VGS) sepsis. The present study, conducted between 1993 and 2003 in children with AML, sought to assess the frequency and characteristics of infectious complications (ICs), the incidence of VGS sepsis, the interest of preventive decontamination, and a possible cytarabine dose-effect on the occurrence of ICs. METHODS: Medical charts of 78 children treated according to the EORTC 58921 clinical trial were analyzed retrospectively. Patients were isolated in laminar air flow rooms, received non-absorbable gut decontamination, gum decontamination with vancomycin mouthwash, and trimethoprim-sulfamethoxasole. ICs were categorized as microbiologically documented infections (MDI), clinically documented infections (CDI), or fever of unknown origin (FUO). RESULTS: Overall, 268 ICs occurred: 57.5% FUO, 8.5% CDI, and 34% MDI. Bloodstream infections occurred in 58 febrile episodes: Gram-positive bacteria represented 83% of the pathogens including 66.1% Staphylococcus species and 8.5% Streptococcus species (6.8% VGS), Gram-negative bacteria represented 13.5% of the pathogens and yeasts 3.5%. Five patients died of infection (6.4%). None died from bacterial infection and no case of VGS sepsis required intensive care. Invasive fungal infection was proven in four patients. Number of ICs was significantly different according to gum and gut decontamination status, and according to the cytarabine dose during the first intensification. No resistant strains were detected in spite of the use of local antibiotics. CONCLUSION: The low rate of VGS and enterobacteriaceae sepsis was probably due to the effective decontamination. Our supportive care strategy could potentially help enhance overall survival in children with AML.

Rat bite fever caused by Streptobacillus moniliformis in a child: human infection and rat carriage diagnosed by PCR.

A child owning pet rats developed an eruptive fever with blisters, polyarthritis, and spectacular desquamation of the hands. Streptobacillus moniliformis was identified after culture of the child's blister fluid and was detected in rat samples by molecular methods. Such detection in the pet of a human victim of rat bite fever has not been reported previously.

Routine use of a commercial test, GLABRATA RTT, for rapid identification of Candida glabrata in six laboratories.

When evaluated in six clinical laboratories from six countries with 1,174 fresh isolates, including 715 Candida glabrata and 459 non-C. glabrata strains, GLABRATA RTT (Fumouze Diagnostics, Levallois Perret, France) yielded an overall sensitivity and an overall specificity of 95.8 and 98.9%, respectively. The results were consistent from one laboratory to another. The five false-positive results corresponded to C. parapsilosis (n = 2), C. tropicalis, C. guilliermondii, and C. lusitaniae. GLABRATA RTT allows a rapid, cost-effective, and reliable presumptive identification of C. glabrata.

Rapid identification of Candida glabrata with a new commercial test, GLABRATA RTT.

The GLABRATA RTT test (Fumouze Diagnostics, Levallois Perret, France) is based on the ability of Candida glabrata to hydrolyze trehalose but not maltose. It requires an inoculum of only four to six colonies, and the results are available within 20 min. We tested GLABRATA RTT with 330 stock isolates grown in subcultures on four different primary fungal isolation media and obtained a sensitivity of 94 to 98% (depending on the medium used) and a specificity of 97.3 to 98.6%. The false-positive results corresponded to C. tropicalis, C. famata, and C. lusitaniae. GLABRATA RTT thus offers rapid and reliable identification of C. glabrata.

Routine use of a one minute trehalase and maltase test for the identification of Candida glabrata in four laboratories.

AIMS: To evaluate the rapid identification of Candida glabrata using a one minute trehalase and maltase test in four clinical laboratories. METHOD: The test was evaluated with 944 freshly isolated yeasts comprising 572 C glabrata and 372 non-C glabrata strains. These strains were isolated on one of three differential media-Candida ID, CHROMagar Candida, or Albicans ID2 medium-and all strains were fully identified using standard methods. RESULTS: The trehalase and maltase test allowed the overall identification of 550 of 572 C glabrata strains (sensitivity, 96.2%) and only 11 of 372 isolates of other yeast species yielded a false positive result (specificity, 96.8 %). Sensitivity and specificity were consistent from one laboratory to another. Using Candida ID medium, the rapid trehalase and maltase test showed a sensitivity of 95% and specificity of 96.2%. Using CHROMagar Candida, sensitivity and specificity were 95.6% and 98.1%, respectively. Using Albicans ID2 medium (tested by two laboratories), the sensitivity was 100% and 98.5% and specificity was 98.1% and 98.2%. In 60% of cases, the test could be performed directly from the primary isolation medium, thus reducing the time for identification. CONCLUSION: The rapid trehalase and maltase test was highly reliable for the presumptive identification of C glabrata on primary isolation using three different chromogenic media. Direct recognition of C albicans by means of their characteristic colour on chromogenic media coupled with one minute trehalase maltase testing performed only on suspect colonies of C glabrata allowed for rapid presumptive identification of the two yeast species most commonly encountered in clinical samples.

Comparison of four chromogenic media and Hektoen agar for detection and presumptive identification of Salmonella strains in human stools.

Several chromogenic media have been developed to enhance the specificity of Salmonella detection. We compared the performance of four commercial chromogenic media-namely, ABC medium (Lab M. Ltd., Bury, United Kingdom), COMPASS Salmonella agar (Biokar Diagnostics, Beauvais, France), CHROMagar Salmonella agar (CHROMagar Company, Paris, France), and SM ID agar (bioMerieux, Marcy l'Etoile, France)-with conventional Hektoen medium. Nine hundred sixteen stool samples from inpatients at three hospitals were cultured, in parallel, on the five media, both by direct inoculation and after selective enrichment in selenite broth. Sixty-four Salmonella strains with 12 serotypes were isolated on at least one medium. After 48 h of incubation, sensitivity before and after enrichment was 62.5 and 89.1% with ABC medium, 77.1 and 93.8% with COMPASS agar, 66.7 and 89.1% with CHROMagar, 68.8 and 85.9% with SM ID agar, and 85.4 and 98.4% with Hektoen agar, respectively. Broth enrichment and prolonged incubation (48 versus 24 h) increased the sensitivity of all five media. Only one strain was not isolated on Hektoen agar. The number of false-positive isolates was higher with all five media after enrichment in selenite broth and after incubation for 48 h compared to 24 h. The specificity of the four chromogenic media was better than 91% after incubation for 24 h (77.7% with Hektoen agar) and better than 84% after incubation for 48 h (74.8% with Hektoen agar). This higher specificity reduces the need for confirmatory tests, thereby cutting technical time and reagent requirements. Both COMPASS agar and CHROMagar Salmonella, which after simple additional tests showed close efficiencies (96 and 97%, respectively), can be recommended as single-plate media of choice for the detection and presumptive identification of salmonellae in stools.

Alafosfalin as a selective agent for isolation of salmonella from clinical samples.

The selectivity of a range of culture media for the detection of Salmonella was assessed using 435 strains of gram-negative bacteria. These media showed limited ability to inhibit non-Salmonella strains found in stool samples. We report the evaluation of alafosfalin as a selective agent for isolation of Salmonella from stool samples. Susceptibility studies with this agent showed that non-typhi Salmonella strains were relatively resistant (mean MIC, 10.2 mg/liter) compared to many coliforms including Escherichia coli (mean MIC, 0.7 mg/liter). A chromogenic medium, ABC medium, was modified to incorporate alafosfalin and was compared with standard ABC medium and Hektoen enteric agar for the isolation of Salmonella from 1,000 stool samples. On direct culture, modified ABC medium showed higher recovery of Salmonella (53.6%) compared with either ABC medium (35.7%) or Hektoen enteric agar (48.2%). We conclude that alafosfalin is a useful selective agent for the isolation of Salmonella from stool samples.

Comparison of three chromogenic agar plates for isolation and identification of urinary tract pathogens.

OBJECTIVE: To comparatively assess the performance of three chromogenic agar plates, CPS ID2, Chromogenic UTI, and USA, for the detection and enumeration of all urinary tract pathogens and the direct identification of Escherichia coli, Proteus mirabilis and Enterococcus spp. METHODS: Two hundred and forty-three urine specimens prospectively collected from hospitalized patients were randomly inoculated in parallel on the three media. RESULTS: Of the 243 urine specimens, 235 yielded positive cultures, of which 151 were pure cultures and 84 were mixed cultures. CPS ID2, Chromogenic UTI and USA agar gave detection rates of 99.1%, 97.1% and 96.6%, respectively. The main difference in non-detection between CPS ID2 agar and the two new media concerned Staphylococcus spp. strains. Based on the total number of strains detected (n = 348), the total identification rates of E. coli, P. mirabilis and Enterococcus spp. on CPS ID2 agar, Chromogenic UTI agar and USA agar were 60.3%, 61.2% and 59.2%, respectively. CONCLUSION: The detection rates and identification rates of the three media were very close and only minor differences were noted. The lower detection rates for Chromogenic UTI and USA were mainly due to their lesser ability to support growth of Staphylococcus spp.

Identification of Candida glabrata by a 30-second trehalase test.

Rapid (30-s) trehalase tests done with material from colonies of 482 yeasts suspended in a drop of trehalose solution on a commercially supplied glucose test strip were positive for 225 (99.1%) of 227 Candida glabrata isolates grown on either of two differential media, Candida ID medium or CandiSelect medium. The test was positive for only 3 (1.2%) and 12 (4.7%) of 255 isolates of other medically important yeast species grown on the same two media, respectively. A rapid maltase test done with a subset of 255 yeast isolates was negative for all but 1 of 64 trehalase-positive C. glabrata isolates, raising the specificity of the rapid testing for C. glabrata to 98.4 to 100%, depending on the isolation medium used. Rapid trehalase and maltase tests done independently in two laboratories with 217 yeast isolates showed sensitivities of 96.0 to 98.0% and specificities of 98.2 to 99.4% for identification of C. glabrata from colonies grown on Candida ID medium. The specificity was much lower because of frequent false-positive trehalose test results when the source of colonies was Sabouraud agar formulated with 4% glucose. We conclude that direct recognition of C. albicans as blue colonies on Candida ID isolation medium coupled with the performance of the 30-s trehalase and maltase tests for C. glabrata among the white colonies on this medium will allow the rapid presumptive identification of the two yeast species most commonly encountered in clinical samples.