Clinical and Microbiological Characteristics of Invasive Group A Streptococcal Infections Before and After Implementation of a Universal Varicella Vaccine Program.

Since the introduction of the varicella vaccine to the routine immunization schedule, we have observed a 70% reduction in the rate of varicella-associated invasive group A streptococcal infections (IGASI). In the mean time, the clinical presentation of IGASI and microbiological characteristics of GAS strains have changed significantly.

Escherichia coli virulence patterns may help to predict vesicoureteral reflux in paediatric urinary tract infections.

AIM: Ultrasound and biological tools are used to predict high-grade vesicoureteral reflux, but other markers are needed to better select patients who need voiding cystography. Our aim was to determine whether studying Escherichia coli virulence factors would help to predict vesicoureteral reflux in patients with their first acute pyelonephritis. METHODS: We included children presenting with E. coli-related acute pyelonephritis or cystitis. Vesicoureteral reflux was assessed by voiding cystography. Virulence factors were identified by multiplex polymerase chain reaction. Statistical analysis was performed using logistic regression and the mean c-statistic test. RESULTS: We included 198 patients: 30 with cystitis and 168 with acute pyelonephritis, including 46 with vesicoureteral reflux. High-grade reflux was associated with acute pyelonephritis caused by the E. coli lacking virulence factors papGII (82% versus 47%, p < 0.001) or papC (85% versus 53%, p < 0.001) or belonging to phylogenetic group A or B1. When we added genetic data (lack of papGII, fyuA and phylogenetic groups) to classical predictors of vesicoureteral reflux (ultrasound examination, gender, age), the ability to predict high-grade reflux increased, with the c-statistic rising from 0.88 to 0.93. CONCLUSION: Bacterial virulence factors and clinical factors helped to predict high-grade reflux and may help to avoid unnecessary voiding cystographies.

Comparative epidemiology of Streptococcus pyogenes emm-types causing invasive and noninvasive infections in French children by use of high-resolution melting-polymerase chain reaction.

BACKGROUND: This study aims to analyze the epidemiology of Group A streptococci (GAS) emm-types causing invasive and noninvasive infections in French children. METHODS: From September 2009 to May 2011, we analyzed GAS isolates from 585 pharyngitis, 125 invasive infections and, for the first time in France, 32 healthy carriers. M protein gene (emm) typing of the isolates was carried out by a new rapid technique, combining 3 multiplex-polymerase chain reactions (PCRs) coupled to high-resolution melting (HRM) curves, able to detect 13 major emm-types (emm 1, 3, 4, 6, 11, 12, 22, 28, 75, 77, 87, 89 and 102). RESULTS: GAS belonging to emm-type 1 were more frequently found among invasive infections than among pharyngitis (24.0% vs. 11.5%, P < 0.001); emm 4 and 89 were more common in pharyngitis than in invasive infections (emm-type 4, 17.4% vs. 6.4%, P = 0.002 and emm-type 89, 9.9% vs. 2.4%, P = 0.006, respectively) and emm 3 and 4 were more common in cases of pharyngitis associated with scarlet fever (21.6% vs. 6.0%, P < 0.001 and 29.3% vs. 14.5%, P < 0.001, respectively). CONCLUSION: HRM method enables the rapid emm-typing of a large number of isolates in epidemiological studies. Comparison of GAS causing invasive and noninvasive infections in the same population of children displays an unbalanced repartition of emm-types.

Escherichia Coli Meningitis Features in 325 Children From 2001 to 2013 in France.

BACKGROUND: We aimed to describe features of Escherichia coli meningitis in a large population of children and the molecular characteristics of the involved strains to determine factors associated with severe disease or death. METHODS: Between 2001 and 2013, a prospective national survey collected data for 325 children hospitalized with E. coli meningitis. The national reference center genetically characterized 141 isolates. RESULTS: Among the 325 cases, 65.2% were term, 22.4% late preterm, and 12.5% very/extremely preterm infants. Escherichia coli meningitis was 7-fold more frequent in preterm than term infants. Median age at diagnosis was 14 days; 71.1% of infants were neonates, with 2 peaks of infection at age 0-3 days (mostly preterm neonates) and 11-15 days (mostly term neonates); 8.9% were >89 days old. In total, 51.1% patients were considered to have severe disease, and 9.2% died. B2.1 phylogenetic subgroup (56%) and O1 serogroup (27.7%) were the most frequently identified. On multivariate analysis, death was associated with preterm birth (odds ratio [OR], 3.3 [95% confidence interval {CI}, 1.3-8.4], P = .015 for late preterm infants; OR, 7.3 [95% CI, 2.7-20.9], P < .001 for very/extremely preterm infants) and cerebrospinal fluid (CSF) to blood glucose ratio <0.10 (OR, 15.3 [95% CI, 1.8-128.3], P = .012). Death was associated with uncommon O serogroup strains (P = .014) and severe disease with O7 serogroup (P = .034) and PapGII adhesin (OR, 2.3 [95% CI, 1.2-4.5], P = .015). CONCLUSIONS: In this large study of 325 cases of E. coli meningitis, risk factors of severe disease or death were preterm birth, severe hypoglycorrhachia, CSF/blood glucose ratio <0.10, and molecular characteristics of strains, which should help optimize therapeutic management.

Escherichia coli bacteremia in children: age and portal of entry are the main predictors of severity.

BACKGROUND: Escherichia coli bacteremia is a major cause of severe sepsis in children. Little is known about predictors of severity. METHODS: We analyzed 84 children ≤ 18 years of age with E. coli bacteremia from the prospective COLIBAFI study performed during 2005-2007. The severity of bacteremia was defined as occurrence of death or transfer to intensive care unit. Studied characteristics included age, gender, birth weight, history of prematurity, immunodepression, nosocomial infection, portal of entry, phylogenetic group and subgroup belonging, O-type, virulence gene content and antimicrobial susceptibility. We compared bacterial characteristics in urinary- versus digestive-source bacteremia, in children ≤ 3 versus >3 month of age, and in children versus adults. We also searched for risk factors of severity. RESULTS: Median age was 2.4 months, 57% males. Most frequent portals of entry were urinary (66.2%) and digestive (19.5%) tracts. Most isolates (63.1%) belonged to B2 phylogroup. Strains in children ≤ 3 months of age exhibited more virulence genes, especially neuC and fyuA/irp2, and were less resistant to antibiotics than in children >3 months of age. Comparing community-acquired urinary-source bacteremia between children and adults, we found that bacteremia were less severe in children, whose strains exhibited a specific virulence gene repertoire and had a higher resistance score than in adults. Seventeen children (20.2%) had a severe bacteremia and 8 died. Non-urinary portal of entry and age ≤ 3 months of age were the only risk factors associated with severity. CONCLUSIONS: E. coli strains responsible for bacteremia exhibit specific characteristics according to age of children. However, host characteristics and portal of entry are the main determinants of severity of E. coli bacteremia in children, as observed in adults.

Clonal expansion of the macrolide resistant ST386 within pneumococcal serotype 6C in France.

In France, the use of the 7-valent pneumococcal conjugate vaccine (PCV7) lead to an overall significant decrease in PCV7 invasive pneumococcal disease (IPD) incidence. However, the decrease in vaccine serotype prevalence was partially counterbalanced by the serotype replacement phenomenon. In this study, we analyzed the role of the newly described serotype 6C as one of the replacement serotypes. This work was conducted on a large time scale from the early PCV7 era (2002-2003) to the PCV13 era (2010-2011), both on IPD strains recovered from the whole population and nasopharyngeal colonizing strains isolated in infant less than two years, who are known to be the main reservoir for pneumococci. Serotype 6C took advantage over 6A and 6B serotypes, which both decreased over time. A continuous and significant increase in 6C IPD was observed in adults along the study period; in contrast, in children less than two years, only an increase in 6C nasopharyngeal carriage was found, the prevalence of serotype 6C in IPD remaining very low over time. Among 101 6C invasive and colonizing strains studied by MLST, 24 STs were found to be related to three major clonal complexes, CC395, CC176, and CC315. STs related to CC176 tend to disappear after 2009 and were essentially replaced by ST386 (CC315), which dramatically increased over time. This clonal expansion may be explained by the erythromycin and tetracycline resistances associated with this clone. Finally, the decrease observed in nasopharyngeal 6C carriage since 2010, likely related to the PCV13 introduction in the French immunization schedule, is expected to lead to a decrease in 6C IPD in adults thereafter.

A conserved virulence plasmidic region contributes to the virulence of the multiresistant Escherichia coli meningitis strain S286 belonging to phylogenetic group C.

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.

OXA-48 carbapenemase-producing Klebsiella pneumoniae isolated from Libyan patients.

Six multidrug-resistant Klebsiella pneumoniae isolates were recovered from injured Libyan combatants. Production of carbapenemase was screened by using commercial combination tablets from Rosco combined with a temocillin disk. Polymerase chain reaction (PCR) and sequencing were used to detect several carbapenemase genes and to characterize their genetic environment. Genetic support was studied by mating-out assays. Plasmid size was identified by the KADO method. PCR and sequencing allowed characterization of plasmid scaffold. Genotyping was performed by pulse-field gel electrophoresis (PFGE) and multilocus sequence typing. PCR was used to check for the presence of nine genes linked to virulence in K. pneumoniae. No carbapenemase was identified by Rosco disks, but all isolates showed high-level temocillin resistance. All of them harbored blaOXA-48 in the transposon Tn1999.2, on a self-conjugative plasmid of about 60 kb, similar to pOXA-48. PFGE revealed three clusters in which isolates were genetically related: The first comprised FM9 and FM10, and the second comprised FM1, FM4, and FM5. FM2 formed a third distinct clone. Sequence types ST101, ST11, and ST147 were identified in keeping with PFGE results. The entB, ycfM, ybtS, and mrkD genes were detected in all isolates, and kfu gene was present in the three ST101 strains. This work confirms the current and successful spread of blaOXA-48 by horizontal dissemination of a single IncL/M plasmid through different genetic backbones with strong epidemic potential. It also highlights the need for rapid and reliable phenotypic detection methods. Attempts to link virulence factors and the production of this carbapenemase deserve further studies.

Invasive serogroup w meningococcal disease in children: a national survey from 2001 to 2008 in France.

From 2001 to 2008, 119 pediatric cases of meningococcal disease caused by serogroup W were reported in France. They cases represented 4% of all meningococcal disease cases in children and were mostly in infants (54%). Meningitis occurred in 78 (66%) patients but differed by isolate. Isolates of the clonal complex sequence type-22 were associated with nonmeningeal presentation. Further investigations of clinical tropism of meningococcal W isolates are warranted.

Necrotizing pneumonia in children: report of 41 cases between 2006 and 2011 in a French tertiary care center.

Forty-one children hospitalized for necrotizing pneumonia were retrospectively analyzed. Necrotizing pneumonia represented 0.8% of community-acquired pneumonia and 6% of hospitalized community-acquired pneumonia. The chest radiograph revealed necrosis on admission in onethird of cases. Twenty-one cases (51%) were documented, including 13 Staphylococcus aureus, all Panton-Valentine leukocidin positive, 7 Streptococcus pneumoniae and 1 Fusobacterium nucleatum.

Rapid-antigen detection tests for group a streptococcal pharyngitis: revisiting false-positive results using polymerase chain reaction testing.

We investigated mechanisms of the false-positive test results on rapid-antigen detection test (RADT) for group A Streptococcal (GAS) pharyngitis. Most RADT false-positives (76%) were associated with polymerase chain reaction-positive GAS results, suggesting that RADT specificity could be considered close to 100%. Finding that 61% of GAS culture-negative but RADT-positive cases were positive on both GAS polymerase chain reaction and Staphylococcus aureus testing, we posit bacterial inhibition as causative.

Characterization of extended-spectrum-beta-lactamase-producing Escherichia coli strains involved in maternal-fetal colonization: prevalence of E. coli ST131.

Maternal-fetal Escherichia coli infections, such as neonatal bacteremia and meningitis, are important causes of morbidity and mortality. From 2006 to 2010, we studied newborns and their mothers who were colonized with E. coli in a French hospital in order to document (i) the epidemiology and genetic characteristics of extended-spectrum-beta-lactamase (ESBL)-producing E. coli strains, (ii) the prevalence of associated virulence genes, (iii) the prevalence of clone sequence type 131 (ST131), and (iv) the genetic relationship among ESBL-producing strains. Among the 2,755 E. coli cultures recovered from vaginal or neonatal samples, 68 were ESBL producers (2.46%). We found a wide diversity of ESBL genes, with the majority being bla(CTX-M-14), bla(CTX-M-1), and bla(CTX-M-15), distributed among the 4 main phylogenetic groups. Genes encoding virulence factors were found in 90.7% of the isolates, with ≥ 2 virulence genes present in 76% of cases. The prevalence of ST131 among ESBL-producing E. coli isolates was 9.4% (6/64). Five of these 6 ST131 isolates possessed bla(CTX-M-15) enzymes (and also were resistant to quinolones), and one possessed bla(CTX-M-2) enzymes. Two possessed virulence genes, suggesting the presence of pathogenicity island IIJ96 (PAI IIJ96)-like domains. Pulsed-field gel electrophoresis (PFGE) revealed a high level of genomic diversity overall, except for 3 closely related isolates belonging to clonal group ST131. Repetitive PCR showed that the six ST131 isolates were closely related to ST131 control strains (>95% similarity). This study shows a high prevalence of ESBL-producing E. coli strains and clonal group ST131 in the French maternal-fetal population. These results suggest a widespread distribution of ESBL enzymes in the community and highlight the early transmission between mothers and neonates. These findings are worrisome, especially for this particularly vulnerable population.

Probiotics and prebiotics in preventing episodes of acute otitis media in high-risk children: a randomized, double-blind, placebo-controlled study.

BACKGROUND: Several studies have suggested that probiotics (proB) and/or prebiotics (preB) could reduce the burden of infection in infants and toddlers. We aimed to determine whether follow-up formula supplemented with proB and preB could reduce the risk of acute otitis media (AOM). METHODS: In this double-blind, placebo-controlled trial from November 2007 to April 2009, 37 pediatricians in France enrolled children 7 to 13 months of age with high risk of AOM who were randomly assigned to receive follow-up formula supplemented with proB (Streptococcus thermophilus NCC 2496, Streptococcus salivarius DSM 13084, Lactobacillus rhamnosus LPR CGMCC 1.3724) and preB (Raftilose/Raftiline) or follow-up formula alone (placebo). During 12 months, the 2 groups were compared for number of AOM episodes diagnosed (primary outcome) and secondary outcomes by the Poisson model (incidence rate ratio [IRR]) or logistic regression (odds ratio; and 95% confidence interval [95% CI]) after adjustment on covariates of interest. RESULTS: We enrolled 224 children (112 in each group). All children were vaccinated (4 doses) with the 7-valent pneumococcal conjugate vaccine; demographic characteristics were similar in the 2 groups. In total, 486 AOM episodes were reported, 249 and 237 in the treatment and control groups, respectively. The treatment and control groups did not differ in incidence of AOM (IRR 1.0, 95% CI: 0.8-1.2), lower respiratory tract infections (IRR 0.9, 0.7-1.2) or number of antibiotic treatment courses (IRR = 1.0, 95% CI: 0.8-1.2). Treatment was not associated with recurrent AOM (odds ratio 1.0, 95% CI: 0.5-1.7). With regard to gastrointestinal disorders, both formulas were well tolerated. CONCLUSION: The proB and preB included in follow-up formula given to children at 7 to 13 months of age did not reduce the risk of AOM, recurrent AOM, antibiotic use or lower respiratory tract infections at 1 year.

Comparative genomics of recent Shiga toxin-producing Escherichia coli O104:H4: short-term evolution of an emerging pathogen.

UNLABELLED: The large outbreak of diarrhea and hemolytic uremic syndrome (HUS) caused by Shiga toxin-producing Escherichia coli O104:H4 in Europe from May to July 2011 highlighted the potential of a rarely identified E. coli serogroup to cause severe disease. Prior to the outbreak, there were very few reports of disease caused by this pathogen and thus little known of its diversity and evolution. The identification of cases of HUS caused by E. coli O104:H4 in France and Turkey after the outbreak and with no clear epidemiological links raises questions about whether these sporadic cases are derived from the outbreak. Here, we report genome sequences of five independent isolates from these cases and results of a comparative analysis with historical and 2011 outbreak isolates. These analyses revealed that the five isolates are not derived from the outbreak strain; however, they are more closely related to the outbreak strain and each other than to isolates identified prior to the 2011 outbreak. Over the short time scale represented by these closely related organisms, the majority of genome variation is found within their mobile genetic elements: none of the nine O104:H4 isolates compared here contain the same set of plasmids, and their prophages and genomic islands also differ. Moreover, the presence of closely related HUS-associated E. coli O104:H4 isolates supports the contention that fully virulent O104:H4 isolates are widespread and emphasizes the possibility of future food-borne E. coli O104:H4 outbreaks. IMPORTANCE: In the summer of 2011, a large outbreak of bloody diarrhea with a high rate of severe complications took place in Europe, caused by a previously rarely seen Escherichia coli strain of serogroup O104:H4. Identification of subsequent infections caused by E. coli O104:H4 raised questions about whether these new cases represented ongoing transmission of the outbreak strain. In this study, we sequenced the genomes of isolates from five recent cases and compared them with historical isolates. The analyses reveal that, in the very short term, evolution of the bacterial genome takes place in parts of the genome that are exchanged among bacteria, and these regions contain genes involved in adaptation to local environments. We show that these recent isolates are not derived from the outbreak strain but are very closely related and share many of the same disease-causing genes, emphasizing the concern that these bacteria may cause future severe outbreaks.

Investigation of an outbreak of osteoarticular infections caused by Kingella kingae in a childcare center using molecular techniques.

We describe an outbreak of 5 osteoarticular infections among 24 daycare center attendees. Polymerase chain reaction revealed Kingella kingae in the joint fluid of 1 child and in 85% of throat samples from healthy contacts. Multilocus sequence typing performed on the joint fluid and carriage isolates identified an unique sequence type. Rifampin failed to eradicate K. kingae carriage.

Community faecal carriage of extended-spectrum beta-lactamase-producing Enterobacteriaceae in French children.

BACKGROUND: The increasing incidence of community acquired infection due to Extended-Spectrum Beta-Lactamase (ESBL) -Producing Enterobacteriaceae represent a great concern because there are few therapeutic alternatives. The fecal flora of children in the community can represent a reservoir for ESBLs genes which are located on highly transmissible plasmids and the spread of these genes among bacterial pathogens is concerning. Because intestinal carriage is a key factor in the epidemiology of ESBL-producing Enterobacteriaceae, the study of the prevalence of these resistant bacteria and risk factors in young children is of particular interest. METHODS: We assessed the prevalence and risk factors of community-acquired faecal carriage of extended-spectrum-ß-lactamase (ESBL)-producing Enterobacteriaceae in children aged from 6 to 24 months, by means of rectal swabbing in community pediatric practices. Child's lifestyle and risk factors for carriage of resistant bacteria were noted. RESULTS: Among the 411 children enrolled, 4.6% carried ESBL-producing Enterobacteriaceae. CTX-M-1, CTX-M-15 and CTX-M-14 were the predominant ESBLs. The 18 E. coli isolates were genetically heterogeneous. Recent third-generation oral-cephalosporin exposure was associated with a higher risk of ESBL carriage (AOR=3.52, 95% CI[1.06-11.66], p=0.04). CONCLUSIONS: The carriage rate of ESBL-producing Enterobacteriacae in young children in the French community setting is noteworthy, underlining the importance of this population as a reservoir. Exposure to third-generation oral cephalosporins was associated with a significant risk of ESBL carriage in our study. Because of the significant public health implications including the treatment of community-acquired urinary tract infections, the spread of organisms producing ESBLs in the community merits close monitoring with enhanced efforts for surveillance.

Decline in macrolide-resistant Streptococcus pyogenes isolates from French children.

We studied the macrolide resistance and serotypes of 585 group A streptococcus (GAS) isolates collected from French children with pharyngitis. Nineteen isolates (3.2%) were erythromycin-resistant and harbored the following resistance genes: 31.6% mef(A), 15.8% erm(A), and 52.6% erm(B). The 19 isolates included 7 different emm types (4, 1, 11, 2, 28, 12, and 77) and 7 corresponding multilocus sequence types. The current fall in macrolide consumption has led to a very low rate of GAS macrolide resistance.

Spectrum and inoculum size effect of a rapid antigen detection test for group A streptococcus in children with pharyngitis.

BACKGROUND: The stability of the accuracy of a diagnostic test is critical to whether clinicians can rely on its result. We aimed to assess whether the performance of a rapid antigen detection test (RADT) for group A streptococcus (GAS) is affected by the clinical spectrum and/or bacterial inoculum size. METHODS: Throat swabs were collected from 785 children with pharyngitis in an office-based, prospective, multicenter study (2009-2010). We analysed the effect of clinical spectrum (i.e., the McIsaac score and its components) and inoculum size (light or heavy GAS growth) on the accuracy (sensitivity, specificity, likelihood ratios and predictive values) of a RADT, with laboratory throat culture as the reference test. We also evaluated the accuracy of a McIsaac-score-based decision rule. RESULTS: GAS prevalence was 36% (95CI: 33%-40%). The inoculum was heavy for 85% of cases (81%-89%). We found a significant spectrum effect on sensitivity, specificity, likelihood ratios and positive predictive value (p<0.05) but not negative predictive value, which was stable at about 92%. RADT sensitivity was greater for children with heavy than light inoculum (95% vs. 40%, p<0.001). After stratification by inoculum size, the spectrum effect on RADT sensitivity was significant only in patients with light inoculum, on univariate and multivariate analysis. The McIsaac-score-based decision rule had 99% (97%-100%) sensitivity and 52% (48%-57%) specificity. CONCLUSIONS: Variations in RADT sensitivity only occur in patients with light inocula. Because the spectrum effect does not affect the negative predictive value of the test, clinicians who want to rule out GAS can rely on negative RADT results regardless of clinical features if they accept that about 10% of children with negative RADT results will have a positive throat culture. However, such a policy is more acceptable in populations with very low incidence of complications of GAS infection.