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We describe a 7-month outbreak of nosocomial Burkholderia cepacia bacteremia involving eight children in a pediatric hospital and the results of epidemiological investigations. A B. cepacia strain genotypically identical to the blood isolates was recovered from the upper surface of capped rubber stoppers of bottles of a commercial lipid emulsion used for parenteral nutrition.

An anonymous 14.9-kb rrn -containing HindIII fragment is strongly linked to Escherichia coli strains causing neonatal meningitis. We show in this report that this fragment does not encode new virulence factors but lacks arpA , a gene common in avirulent E. coli strains, and we developed a PCR test to detect this fragment.

Neisseria meningitidis colonizes the nasopharynx and, unlike commensal Neisseria species, is capable of entering the bloodstream, crossing the blood-brain barrier, and invading the meninges. The other pathogenic Neisseria species, Neisseria gonorrhoeae , generally causes an infection which is localized to the genitourinary tract. In order to investigate the genetic basis of this difference in disease profiles, we used a strategy of genomic comparison. We used DNA arrays to compare the genome of N. meningitidis with those of N. gonorrhoeae and Neisseria lactamica , a commensal of the nasopharynx. We thus identified sequences conserved among a representative set of virulent strains which are either specific to N. meningitidis or shared with N. gonorrhoeae but absent from N. lactamica . Though these bacteria express dramatically different pathogenicities, these meningococcal sequences were limited and, in contrast to what has been found in other pathogenic bacterial species, they are not organized in large chromosomal islands.

Background The prevalence of ESBL bacteria in community-acquired UTIs is increasing. This is of concern, since antibiotic therapy would be restricted to a few antibiotics, including carbapenems (in turn, the frequent use of penems leads to carbapenem-resistance), aminoglycosides, colimycin and fosfomycin. Aims To describe the prevalence of ESBL among Gram-negative bacteria causing community-acquired UTIs managed in a tertiary care PED serving an active Department of paediatric urology. Methods Retrospective study of all UTI episodes diagnosed between 1st January and 31th December, 2012. UTIs were retrieved by using the PED and Bacteriology databases. Results 457 (0.6%) community-acquired UTIs have been identified among 78,152 visits in the PED in 2012. 358 (78%) were diagnosed as acute pyelonephritis based on clinical signs and elevated CRP and/or PCT, and 99 (21%) as acute cystitis. Whereas no ESBL bacteria was identified among episodes of cystitis, 16 acute pyelonephritis cases were due to ESBL E.coli (i.e., 4.5% of all E.coli and 3.4% of all UTIs). 13/16 (81%) UTIs occurred in children suffering urinary tract abnormalities. Moreover, one child with vesico-ureteral bilateral reflux had 3 distinct episodes of UTIs due to ESBL K.pneumoniae in 2012. Conclusions The incidence of ESBL E. coli causing community-acquired UTIs remains low (~5%) in a tertiary hospital PED. This reassuring finding comforts the French UTI current recommendations of using as a first-line therapy iv ceftriaxone for 4 days followed by oral cefixime for 6 additional days. However, ESBL bacteria causing UTIs are favoured by urinary malformations, previous hospitalisations and prophylactic antibiotics.

Background Non-typable Haemophilus influenzae (NT-Hi) infection is frequently associated with acute otitis media (AOM) treatment failure, recurrence or chronic otitis media. Persistence of otopathogens in a biofilm-structured community was implicated in these situations. Here, we compared biofilm production by H. influenzae strains obtained by culture of middle ear fluid (MEF) from children with AOM treatment failure and by strains isolated from nasopharyngeal (NP) samples from healthy children or those with AOM (first episode or recurrence). We aimed to evaluate an association of clinical signs and in vitro biofilm formation and establish risk factors of carrying a biofilm-producing strain. Methods We used a modification of the microtiter plate assay with crystal violet staining to compare biofilm production by 216 H. influenzae strains: 41 in MEF from children with AOM treatment failure (group MEF), 43 in NP samples from healthy children (NP group 1), 88 in NP samples from children with a first AOM episode (NP group 2, n = 43) or recurrent (NP group 3, n = 45) and 44 in NP samples from children with AOM associated with conjunctivitis (NP group 4). Results At all, 106/216 (49%) H. influenzae strains produced biofilm as did 26/43 (60.5%) in NP samples from healthy children. Biofilm production in MEF samples and NP samples did not significantly differ (40.5% vs 60.5%, 55.8%, 56.8% and 31.1% for NP groups 1, 2, 3 and 4, respectively). On multivariate analysis, only presence of conjunctivitis was significantly associated with low biofilm production (OR = 0.3, CI [0.16-0.60], p = 0.001). The ampicillin resistance of H. influenzae produced by penicillin-binding protein modification was significantly associated with low biofilm production (p = 0.029). Conclusion We found no association of biofilm production and AOM treatment failure or recurrence. Biofilm production was low from H. influenzae strains associated with conjunctivitis-otitis syndrome and from strains with modified penicillin-binding protein.

Objectif Decrire la prevalence des betalactamases a spectre etendu (BLSE) au sein des enterobacteries responsables d’infections urinaires en 2012 aux urgences pediatriques d’un CHU ayant une activite de chirurgie urologique. Methodes Etude retrospective du 1 er janvier au 31 decembre 2012 aux urgences pediatriques de l’hopital Robert Debre. Nous avons identifie l’ensemble des episodes d’infection urinaire par le biais du codage diagnostic du logiciel des urgences et des ECBU realises dans le laboratoire de microbiologie. Resultats En 2012, 457 episodes d’infections urinaires ont ete identifies (0,6% des passages) dont 99 cystites et 358 pyelonephrites aigues. Aucun episode de cystite n’etait du a un germe producteur de BLSE. 16 episodes de PNA etaient lies a un E. coli BLSE (soit 4,5% des E. coli et 3,4% des infections urinaires). Parmi ces 16 episodes, 13 etaient survenus chez des patients souffrant d’uropathie. Par ailleurs un patient souffrant d’uropathie a presente 3 episodes successifs de PNA a Klebsiella pneumoniae BLSE en 2012. Conclusions La proportion de PNA a E. coli BLSE reste inferieure a 5% en 2012 aux urgences d’un CHU et concerne tres majoritairement des enfants souffrant d’uropathie.

Using the automated Riboprinter system, we have initiated the construction of an electronic Riboprint database composed of 72 ECOR reference strains and 15 archetypal virulent strains in order to provide a new simple molecular characterization method. More than 90% of the ECOR strains clustered in their original phylogenetic group. All but one of the archetypal virulent strains had a profile identical to that of one of the ECOR strains and could be easily affiliated with a phylogenetic group. This method appears to be an accurate and practical tool especially for investigating the genetic relationship between clinical extraintestinal pathogenic strains and B2 subgroup ECOR strains or archetypal pathotype strains.

An intraspecies phylogenetic grouping of 168 human commensal Escherichia coli strains isolated from the stools of three geographically distinct human populations (France, Croatia, Mali) was generated by triplex PCR. The distributions of seven known extraintestinal virulence determinants (ibeA, pap, sfa/foc, afa, hly, cnf1, aer) were also determined by PCR. The data from the three populations were compiled, which showed that strains from phylogenetic groups A (40%) and B1 (34%) were the most common, followed by phylogenetic group D strains (15%). Strains of the phylogenetic group B2 were rare (11%). However, a significant specific distribution for strains of groups A, B1 and B2 within each population was observed, which may indicate the influence of (i) geographic/climatic conditions, (ii) dietary factors and/or the use of antibiotics or (iii) host genetic factors on the commensal flora. Virulence determinants were rarely detected, with only 25·6% of the strains harbouring at least one of the virulence genes tested. The strains with virulence factors most frequently belonged to phylogenetic group B2. The commensal strains of phylogenetic groups A, B1 and D had fewer virulence determinants than pathogenic strains of the corresponding groups when these data were compared with those for previous collections of virulent extraintestinal infection strains studied using the same approach. However, the virulence patterns of commensal and pathogenic B2 phylogenetic group strains were the same. The data thus suggest that strains of the A, B1 and D phylogenetic groups predominate in the gut flora and that these strains must acquire virulence factors to become pathogenic. In contrast, commensal phylogenetic group B2 strains are rare but appear to be potentially virulent.

The prevalence of the ibe10 gene, of the pap, afa, and sfa adhesin-encoding operons, and of a 14.9-kb rrn-containing HindIII fragment was studied for 67 Escherichia coli neonatal meningitis strains, 58 E. coli K1 commensal strains, and 47 E. coli blood isolates from neonates without meningitis. ibe10, sfa, and the 14.9-kb HindIII fragment were observed significantly more often in the meningitis strains than in blood or commensal strains.

Cerebrospinal fluid (CSF) was taken from 19 children with bacterial meningitis treated with cefotaxime (300 mg/kg of body weight/day) and vancomycin (60 mg/kg/day). Median levels of drugs in CSF were smaller than expected, as follows: 4.4 microg/ml for cefotaxime, 3.2 microg/ml for desacetylcefotaxime, and 1.7 microg/ml for vancomycin. The median CSF bactericidal titer against an intermediately cefotaxime-resistant pneumococcus was 1:4. Our data suggest at least an additive interaction between the drugs used in this study.

Recent isolation of the non-K1 Escherichia coli neonatal meningitis strain S286, belonging to phylogroup C, which is closely related to major group B1, and producing an extended-spectrum beta-lactamase, encouraged us to seek the genetic determinants responsible for its virulence. We show that S286 belongs to the sequence O type ST23O78 and harbors 4 large plasmids. The largest one, pS286colV (~120 kb), not related to resistance, contains genes characteristic of a Conserved Virulence Plasmidic (CVP) region initially identified in B2 extra-intestinal avian pathogenic E. coli (APEC) strains and in the B2 neonatal meningitis E. coli strain S88. The sequence of this CVP region has a strong homology (98%) with that of the recently sequenced plasmid pChi7122-1 of the O78 APEC strain Chi7122. A CVP plasmid-cured variant of S286 was less virulent than the wild type strain in a neonatal rat sepsis model with a significant lower level of bacteremia at 24 h (4.1 ± 1.41 versus 2.60 ± 0.16 log CFU/ml, p = 0.001) and mortality. However, the mortality in the model of adult mice was comparable between wild type and variant indicating that pS286colV is not sufficient by itself to fully explain the virulence of S286. Gene expression analysis of pS286colV in iron depleted environment was very close to that of pS88, suggesting that genes of CVP region may be expressed similarly in two very different genetic backgrounds (group C versus group B2). Screening a collection of 178 human A/B1 extraintestinal pathogenic E. coli (ExPEC) strains revealed that the CVP region is highly prevalent (23%) and MLST analysis indicated that these CVP positive strains belong to several clusters and mostly to phylogroup C. The virulence of S286 is explained in part by the presence of CVP region and this region has spread in different clusters of human A/B1 ExPEC, especially in group C.

We investigated the combination of teicoplanin and ceftriaxone for its bactericidal activity against Staphylococcus aureus, Haemophilus influenzae type b and Streptococcus pneumoniae isolated from bone and joint infections in children. An increase in bactericidal activity was observed against isolates of S. aureus and H. influenzae when the antibiotics were tested at fractional MICs whereas indifference was observed at their MICs. Similar results were obtained at fractional MICs against S. pneumoniae, but the bactericidal activity fell by more than 1 x log 10 cfu/mL when the antibiotics were tested at or above their MICs.

Objective To evaluate the effect of the infection and the obstruction of urinary tract on renal parenchyma, we developed a model of ascending urinary infection in mice with partial unilateral obstructive uropathy. Methods Six week-old CBA/J mice were operated on to perform a right partial ureteral obstruction and challenged transurethrally with Escherichia coli CFT073 (high virulent strain) or E. coli Mez (low virulent strain). Level of infection of urine and kidneys, score of inflammation and fibrosis of kidneys, and kidney weight ratio (KWR) in the short and long term were studied. Results At short-term (72 hours) experimentation, partial ureteral obstruction does not influence the level of kidney infection in terms of bacterial count, and the score of inflammation regardless of whether a virulent or low virulent strain was used. At long-term (42 days) experimentation, E. coli Mez was eliminated from all mice kidneys, but CFT073 persisted in almost all; obstruction did not influence the level of kidney infection with CFT073 but a significant difference of KWR and the inflammation and fibrosis score between obstructed and unobstructed kidneys was found ( P = .0078; P = .036, respectively). Although the E. coli Mez strain did not persist in renal parenchyma, severe damage of the renal parenchyma was observed. Conclusion The proposed model is similar to the obstructive uropathy in children in which ureteral obstruction is present before the onset of infection. The association of obstruction and urinary infection impairs kidney growth and favors the occurrence of renal damage.

BACKGROUND: Kingella kingae, a normal component of the upper respiratory flora, is being increasingly recognized as an important invasive pathogen in young children. Genetic diversity of this species has not been studied. METHODS: We analyzed 103 strains from different countries and clinical origins by a new multilocus sequence-typing (MLST) schema. Putative virulence gene rtxA, encoding an RTX toxin, was also sequenced, and experimental virulence of representative strains was assessed in a juvenile-rat model. RESULTS: Thirty-six sequence-types (ST) and nine ST-complexes (STc) were detected. The main STc 6, 14 and 23 comprised 23, 17 and 20 strains respectively, and were internationally distributed. rtxA sequencing results were mostly congruent with MLST, and showed horizontal transfer events. Of interest, all members of the distantly related ST-6 (n = 22) and ST-5 (n = 4) harboured a 33 bp duplication or triplication in their rtxA sequence, suggesting that this genetic trait arose through selective advantage. The animal model revealed significant differences in virulence among strains of the species. CONCLUSION: MLST analysis reveals international spread of ST-complexes and will help to decipher acquisition and evolution of virulence traits and diversity of pathogenicity among K. kingae strains, for which an experimental animal model is now available.

We read with interest the comment of C. Miranda, L. Porte, and P. Garcia contributing to the problem of real-time PCR biological diagnosis of whooping cough using IS 481 as the target. The authors detected Bordetella holmesii in 7 patients out of 51 between 0 and 9 years of age, with 3 being less

We evaluated the in vitro killing activities of ceftriaxone, imipenem, vancomycin, gentamicin, fosfomycin, and rifampin, alone and in combination, against 26 Streptococcus pneumoniae strains (penicillin G MICs, > 0.125 to 2 micrograms/ml) isolated from the cerebrospinal fluid of children with meningitis. The antibiotics were tested at clinically achievable concentrations in cerebrospinal fluid. After 5 h of incubation, imipenem was the most effective drug. None of the combinations had synergistic activity. Killing by beta-lactam antibiotics or vancomycin was enhanced by the addition of gentamicin, reduced by the addition of rifampin, and unaffected by the addition of fosfomycin.

La coqueluche, malgre une excellente couverture vaccinale chez les nourrissons et les enfants, demeure la premiere cause de mortalite par infection bacterienne chez le nourrisson de moins de 3 mois. La PCR TR, basee sur la detection de l’IS481, represente la methode diagnostique de reference pour la recherche de B. pertussis. La sensibilite et la specificite de cette technique ont ete largement evaluees, cependant sa performance au cours de l’antibiotherapie n’est pas connue. Objectif Evaluer la persistance de l’ADN de B. pertussis par PCR TR, dans les SNP d’enfants hospitalises et traites pour une coqueluche. Resultats Entre janvier 2005 et janvier 2008, 21 enfants infectes par Bordetella pertussis et hospitalises a l’hopital Robert-Debre, ont eu au moins un deuxieme prelevement realise apres le debut de l’antibiotherapie. Apres 5 jours de traitement les PCR etaient positives pour 100 % des patients. Apres 14 jours d’antibiotherapie 6 des 8 patients analyses (75 %) presentaient encore une PCR positive. La quantification de l’ADN bacterien exprimee en « Cycle Threshold » (CT) indiquait que la clearance de l’ADN etait en moyenne de 1,6 CT/j (extremes : 0,5 a 3,2 CT/js). Discussion Nos resultats indiquent que l’ADN de B. pertussis persiste au-dela de la duree de positivite de la culture (generalement 5 jours) apres le debut de l’antibiotherapie et soulignent l’utilite de la PCR TR dans le diagnostic retrospectif de la coqueluche meme apres 2 semaines d’antibiotherapie. Le suivi quantitatif de l’ADN chez les patients traites pourrait etre evalue comme element predictif d’un echec therapeutique.

Ribotyping randomly amplified polymorphic DNA analysis, and pulsed-field gel electrophoresis were used for the epidemiologic evaluation of eight Pseudomonas aeruginosa O:12 isolates obtained from eight children and two P. aeruginosa O:12 environmental isolates from a hematology ward. Randomly amplified polymorphic DNA analysis and pulsed-field gel electrophoresis were able to discriminate isolates that were indistinguishable by biochemical typing, O serotyping or ribotyping.

International audience

Background After the implementation of pneumococcal conjugate vaccines (PCVs), the marked shift in Streptococcus pneumoniae (Pnc) serotype distribution led to a modification in pneumococcal antibiotic susceptibility. In 2011, the pattern of antibiotic prescription in France for acute otitis media in infants was greatly modified, with decreased use of third-generation cephalosporins and amoxicillin–clavulanate replaced by amoxicillin alone. To assess antibiotic strategies, here we measured the antibiotic susceptibility of Pnc and Haemophilus influenzae (Hi) isolated from nasopharyngeal flora in infants with acute otitis media in the 13-valent PCV (PCV13) era in France. Methods From November 2006 to June 2013, 77 pediatricians obtained nasopharyngeal swabs from infants (6 to 24 months old) with acute otitis media. The swabs were sent for analysis to the national reference centre for pneumococci in France. Demographics, medical history, and physical examination findings were recorded. Results We examined data for 7200 children, 3498 in the pre-PCV13 period (2006–2009) and 3702 in the post-PCV13 period (2010–2013). The Pnc carriage rate decreased from 57.9 % to 54.2 % between the 2 periods, and the proportion of pneumococcal strains with reduced susceptibility to penicillin or resistant to penicillin decreased from 47.1 % to 39 % (P

Background Bordetella pertussis is the main agent of whooping cough. Vaccination with acellular pertussis vaccines has been largely implemented in high-income countries. These vaccines contain 1 to 5 antigens: pertussis toxin (PT), filamentous haemagglutinin (FHA), pertactin (PRN) and/or fimbrial proteins (FIM2 and FIM3). Monitoring the emergence of B. pertussis isolates that might partially escape vaccine-induced immunity is an essential component of public health strategies to control whooping cough. Aim We aimed to investigate temporal trends of fimbriae serotypes and vaccine antigen-expression in B. pertussis over a 23-year period in France (1996–2018). Methods Isolates (n = 2,280) were collected through hospital surveillance, capturing one third of hospitalised paediatric pertussis cases. We assayed PT, FHA and PRN production by Western blot (n = 1,428) and fimbriae production by serotyping (n = 1,058). Molecular events underlying antigen deficiency were investigated by genomic sequencing. Results The proportion of PRN-deficient B. pertussis isolates has increased steadily from 0% (0/38) in 2003 to 48.4% (31/64) in 2018 (chi-squared test for trend, p 481 insertion within the prn gene or a 22 kb genomic inversion involving the prn promoter sequence, indicative of convergent evolution. FIM2-expressing isolates have emerged since 2011 at the expense of FIM3. Conclusions B. pertussis is evolving through the rapid increase of PRN-deficient isolates and a recent shift from FIM3 to FIM2 expression. Excluding PRN, the loss of vaccine antigen expression by circulating B. pertussis isolates is epidemiologically insignificant.

International audience; Objectives:Using registry data to compare fungaemia caused by uncommon yeast species (UYS; i.e. other than Candida albicans , Candida glabrata , Candida parapsilosis , Candida tropicalis and Candida krusei ) and C. albicans -related fungaemia can reveal specific predisposing factors of UYS with potential impact on treatment strategies.Methods:We analysed 338 episodes of UYS fungaemia prospectively collected from 27 hospitals (Paris, France; 1 October 2002-31 December 2014) and compared these with 1998 single episodes of C. albicans fungaemia using univariate and multivariate analyses.Results:The proportion of UYS fungaemia was stable over time. Thirty-five different species were identified (27 ascomycetes, 8 basidiomycetes), 11 had caspofungin MIC 50 >0.25 mg/L and 15 fluconazole MIC 50 >4 mg/L. Haematological malignancies [OR=2.39 (95% CI 1.79-3.18)] and prior exposure to antifungal drugs [OR=1.87 (1.30-2.69)] were independent predisposing factors for UYS infections upon multivariate analysis. However, when considering the genus/species complex level, only infections due to Candida kefyr -related species [OR=4.01 (2.42-6.64)] and to Trichosporon spp. [OR=5.38 (1.72-16.81)] remained associated with haematological malignancies, those due to the GEOTRICHUM group with acute leukaemia [OR=61.29 (19.23-195.36)], and infections with Trichosporon spp. or the GEOTRICHUM group with prior exposure to caspofungin [OR=15.67 (3.62-67.80) and OR=13.17 (3.33-52.03), respectively] but not to fluconazole. The global mortality at day 30 for UYS was similar to that for C. albicans (35.4%, and 39.9%, respectively), but very divergent results were observed according to the specific UYS.Conclusions:UYS encompass a high diversity of species, each with its own behaviour and predisposing factors for human infections. This variety makes it important to rapidly identify an isolate to the species level in order to optimize antifungal treatment.