Search

Summary Background Preterm infants are at high risk for extended-spectrum-beta-lactamase-producing Enterobacteriaceae (ESBL-E) sepsis and neonatal intensive care unit (NICU) outbreaks. Maternal colonization with ESBL-E may be precursory to mother-to-child transmission. However, there is no consensus regarding surveillance of pregnant women for ESBL-E colonization. Aim To identify pairs of mothers and infants harbouring same-strain ESBL-E colonization and to determine whether maternal transmission may play a role in increasing ESBL-E carriage in preterm infants. Methods This was a one-year analysis from an ongoing, prospective ESBL-E surveillance of mothers of premature infants and their offspring. Mother–infant pairs colonized with the same bacteria underwent strain analysis using pulsed-field gel electrophoresis (PFGE). Clinical parameters were collected from the hospital computerized records. Findings Between January 2015 and January 2016, 313/409 (76.5%) mothers and all 478 (100%) infants were screened for ESBL-E colonization; carriage rates were 21.5% and 14.8%, respectively. Four (5.6%) colonized infants developed late-onset sepsis and two (2.8%) died. Twenty-five mother–infant pairs colonized with the same bacterial strain were identified; a subgroup of 10 pairs of isolates underwent PFGE, and 70% displayed an identical PFGE fingerprint. No similarities were found between isolates recovered from unrelated neonates and mothers. ESBL-E colonization was found significantly earlier in infants of mothers colonized at birth (P Conclusions ESBL-E carriage rates in mothers and NICU infants with non-negligible maternal–neonatal ESBL-E transmission in the study region indicate that maternal colonization surveillance and/or further infection control interventions should be considered.

Israel implemented use of 7- and 13-valent pneumococcal vaccine in 2009 and 2010, respectively. We describe results of prospective, population-based, nationwide active surveillance of Streptococcus pneumoniae serotype 12F (Sp12F) invasive pneumococcal disease (IPD) dynamics in the 7 years after vaccine introduction. Of 4,573 IPD episodes during July 2009–June 2016, a total of 434 (9.5%) were caused by Sp12F. Sp12F IPD rates (cases/100,000 population) increased in children 3.9 since 2011–2012, followed by an increase in all ages. During 2011–2016, Sp12F was the most prevalent IPD serotype. Sp12F isolates were mostly penicillin nonsusceptible (MIC >0.06 µg/mL; MIC50 = 0.12) and predominantly of sequence type 3774), a clone exclusively found in Israel (constituting ≈90% of isolates in 2000–2009). The sharp increase, long duration, and predominance of Sp12F IPD after vaccine implementation reflect a single clone expansion and may represent more than a transient outbreak.

Background In addition to reducing vaccine-type nasopharyngeal carriage rates, pneumococcal conjugate vaccines (PCVs) may decrease carriage density in vaccinated individuals still carrying vaccine serotypes. However, reduction of carriage density has not been systematically studied. This study compared the effect of PCV13 versus PCV7 on carriage density of the serotypes in PCV13 that are not included in PCV7. Methods This randomized, double-blind study was conducted in southern Israel and included Jewish and Bedouin subjects. Per protocol, 881 and 873 infants received PCV13 and PCV7, respectively, at ages 2, 4, 6, and 12 months. Nasopharyngeal cultures at ages 7, 12, 13, 18, and 24 months were plated using the 4-quadrant semiquantitative method and graded 0 (negative) to 4 (growth in all plate quadrants). In this post hoc analysis, the least squares means of cumulative colonization densities per serotype and serotype combination of the total population and each ethnic subpopulation in each vaccine group were calculated, and differences between vaccine groups derived from a linear model. Results PCV13-vaccinated children still carrying the 6 additional PCV13 serotypes unique to PCV13 showed no significant differences in carriage density compared with the PCV7-vaccinated control group. No differences in carriage density were shown between Jewish and Bedouin subpopulations despite higher carriage rates among Bedouin subjects. Conclusions Although PCV13 vaccination reduces vaccine-type carriage compared with PCV7 vaccination by reducing nasopharyngeal acquisition of the additional PCV13 serotypes as previously reported, the current study lacks evidence of a decrease in carriage density of these serotypes when acquired in vaccinated children. Despite the lack of effect on carriage density observed, surveillance data suggest a dramatic decrease in disease rates after PCV implementation. Thus, the current analysis suggests that PCV’s impact on carriage density has minimal or no impact on vaccine success. ( www.ClinicalTrials.gov : NCT00508742)

Background: We compared PCV7 serological response and protection against carriage in infants receiving 3 doses (2, 4, 6 months; 3+0 schedule) to those receiving a booster (12 months; 3+1). Methods: A prospective, randomized controlled study, conducted between 2005 and 2008, before PCVs were implemented in Israel. Healthy infants were randomized 1:1:1 to receive 3+1, 3+0 and 0+2 (control group; 12, 18 months doses). Nasopharyngeal/oropharyngeal swabs were obtained at all visits. Serum serotype-specific IgG concentrations and opsonic activities (OPA) were measured at 2, 7, 13 and 19 months. This study was registered with Current Controlled Trials, Ltd. ISRCTN28445844. Results: Overall, 544 infants were enrolled: 3+1 (n = 178), 3+0 (n = 178) and 0+2 (n = 188). Post-priming (7 months), antibody concentrations were similar in both groups, except for serotype 18C (higher in 3+0). Post-booster (13, 19 months), ELISA and OPA levels were significantly higher in 3+1 than in 3+0 group. Nasopharyngeal/oropharyngeal cultures were positive for Streptococcus pneumoniae in 2673 (543%) visits. Acquisition rates (vaccine and non-vaccine serotypes) were similar for 3+1 and 3+0 groups at 7-30 months and for 0+2 group at 19-30 months. Conclusions: PCV7 booster after 3 priming doses increased substantially IgG concentrations but did not further reduced vaccine-serotype nasopharyngeal acquisition, suggesting that protection from pneumococcal carriage does not depend primarily on serum IgG. (C) 2018 Elsevier Ltd. All rights reserved.

Pneumococcal flavin reductase (FlaR) is known to be cell-wall associated and possess age dependent antigenicity in children. This study aimed at characterizing FlaR and elucidating its involvement in pneumococcal physiology and virulence. Bioinformatic analysis of FlaR sequence identified three-conserved cysteine residues, suggesting a transition metal-binding capacity. Recombinant FlaR (rFlaR) bound Fe2+ and exhibited FAD-dependent NADP-reductase activity, which increased in the presence of cysteine or excess Fe2+ and inhibited by divalent-chelating agents. flaR mutant was highly susceptible to H2O2 compared to its wild type (WT) and complemented strains, suggesting a role for FlaR in pneumococcal oxidative stress resistance. Additionally, flaR mutant demonstrated significantly decreased mice mortality following intraperitoneal infection. Interestingly, lack of FlaR did not affect the extent of phagocytosis by primary mouse peritoneal macrophages but reduced adhesion to A549 cells compared to the WT and complemented strains. Noteworthy are the findings that immunization with rFlaR elicited protection in mice against intraperitoneal lethal challenge and anti-FlaR antisera neutralized bacterial virulence. Taken together, FlaR’s roles in pneumococcal physiology and virulence, combined with its lack of significant homology to human proteins, point towards rFlaR as a vaccine candidate.

During the years 1999 through 2008 middle ear fluid (MEF) and nasopharyngeal/oropharyngeal (NP/OP) samples were obtained from Costa Rican children with otitis media (OM). Streptococcus pneumoniae was identified and serotyped according to standard procedures. A total of 1,389 MEF isolates were collected, of which 353 (25%) were S. pneumoniae. Serotyping was performed in 286/353 (81%) isolates, of which 40 (14%) were serogroup 6: serotype 6A (10/40; 25%), 6B (27/40; 67%) and 6C (3/40; 8%). Among 755 NP/OP S. pneumoniae isolated, 537 (71%) were serotyped and serogroup 6 was identified in 112 (21%) isolates: 6A (24/112; 21%), 6B (78/112; 70%), and 6C (10/112; 9%). Twenty percent, 86% and 0% of serotypes 6A, 6B and 6C isolated from the MEF, respectively, were penicillin non-susceptible (MIC ≥ 0.06 μg/mL) and among the NP/OP isolates, 25%, 17.9% and 10%, respectively, were penicillin non-susceptible. Our results show that in Costa Rican children with OM, S. pneumoniae serotype 6C is an infrequent cause of OM and is sporadically found in NP/OP samples.

Streptococcus pneumoniae is a facultative anaerobic pathogen. Although it maintains fermentative metabolism, during aerobic growth pneumococci produce high levels of H2O2, which can have adverse effects on cell viability and DNA, and influence pneumococcal interaction with its host. The pneumococcus is unusual in its dealing with toxic reactive oxygen species (ROS) in that it neither has catalase nor the global regulators of peroxide stress resistance. Previously, we identified pneumococcal thiol peroxidase (TpxD) as the key enzyme for enzymatic removal of H2O2, and showed that TpxD synthesis is up-regulated upon exposure to H2O2. This study aimed to reveal the mechanism controlling TpxD expression under H2O2 stress. We hypothesize that H2O2 activates a transcription factor which in turn up-regulates tpxD expression. Microarray analysis revealed a pneumococcal global transcriptional response to H2O2. Mutation of tpxD abolished H2O2-mediated response to high H2O2 levels, signifying the need for an active TpxD under oxidative stress conditions. Bioinformatic tools, applied to search for a transcription factor modulating tpxD expression, pointed toward CodY as a potential candidate. Indeed, a putative 15-bp consensus CodY binding site was found in the proximal region of tpxD-coding sequence. Binding of CodY to this site was confirmed by EMSA, and genetic engineering techniques demonstrated that this site is essential for TpxD up-regulation under H2O2 stress. Furthermore, tpxD expression was reduced in a ΔcodY mutant. These data indicate that CodY is an activator of tpxD expression, triggering its up-regulation under H2O2 stress. In addition we show that H2O2 specifically oxidizes the 2 CodY cysteines. This oxidation may trigger a conformational change in CodY, resulting in enhanced binding to DNA. A schematic model illustrating the contribution of TpxD and CodY to pneumococcal global transcriptional response to H2O2 is proposed.

This clone has persisted in a post–acute care facility for >5 years., Prolonged outbreaks of multidrug-resistant Streptococcus pneumoniae in health care facilities are uncommon. We found persistent transmission of a fluroquinolone-resistant S. pneumoniae clone during 2006–2011 in a post–acute care facility in Israel, despite mandatory vaccination and fluoroquinolone restriction. Capsular switch and multiple antimicrobial nonsusceptibility mutations occurred within this single clone. The persistent transmission of fluoroquinolone-resistant S. pneumoniae during a 5-year period underscores the importance of long-term care facilities as potential reservoirs of multidrug-resistant streptococci.

SummaryObjectiveTo assess the epidemiological and microbiological characteristics of pneumococcal acute otitis media (AOM) in children in Brasov, Central Romania, before the introduction of pneumococcal conjugate vaccine (PCV) into the routine national immunization program.MethodsAll AOM patients aged 2.0μg/ml). S. pneumoniae non-susceptibility rates to trimethoprim–sulfamethoxazole, erythromycin, and clindamycin were 75.0%, 58.3%, and 35.4%, respectively. All isolates were susceptible to chloramphenicol. Multidrug resistance was found in 33 (68.7%) isolates. The most common S. pneumoniae serotypes were 19F (14, 29.2%), 6B (8, 16.7%), 23F (8, 16.7%), and 14 (6, 12.5%). Serotype 19A was found in three (6.2%) patients and 6A in two (4.1%). Non-PCV13 serotypes represented six (12.6%) of all serotypes (four of them non-susceptible to penicillin). Thirty-six (75.0%) isolates were potentially covered by PCV7, 37 (77.0%) by PCV10, and 42 (87.5%) by PCV13.Conclusions(1) S. pneumoniae was the most prevalent pathogen, with frequent antibiotic resistance and multi-resistance patterns; (2) most pneumococcal AOM and multidrug-resistant episodes could be prevented by PCVs.

Background. The 13-valent pneumococcal conjugate vaccine (PCV13) was licensed to replace the 7-valent pneumococcal conjugate vaccine (PCV7) based on serological noninferiority criteria. To date no randomized PCV13 pediatric trial has included clinical endpoints. Methods. This randomized double-blind trial compared the impact of PCV13 versus PCV7 on nasopharyngeal (NP) colonization and immunogenicity. Healthy infants were randomized (1:1) to receive PCV7 or PCV13 at ages 2, 4, 6, and 12 months; NP swabs were collected at 2, 4, 6, 7, 12, 13, 18, and 24 months, and blood was drawn at 7 and 13 months. Rates of NP acquisition and prevalence, and serotype-specific immunoglobulin G (IgG) concentrations were assessed. Results. The per protocol analysis population included 881 PCV13 and 873 PCV7 recipients. PCV13 significantly reduced NP acquisition of the additional PCV13 serotypes 1, 6A, 7F, and 19A; the cross-reacting serotype 6C; and the common PCV7 serotype 19F. For serotype 3, and the other PCV7 serotypes, there were no significant differences between the vaccine groups. There were too few serotype 5 events to draw inference. The impact on prevalence at predefined time points was similar to that observed with NP acquisition. PCV13 elicited significantly higher IgG responses for PCV13 additional serotypes and serotype 19F, and similar or lower responses for 6/7 PCV7 serotypes. Conclusions. PCV13 resulted in lower acquisition and prevalence of NP colonization than PCV7 did for 4 additional PCV13 serotypes, and serotypes 6C and 19F. It was comparable with PCV7 for all other common serotypes. These findings predict vaccine effectiveness through both direct and indirect protection. Clinical Trials Registration. NCT00508742.

The purpose of this study was to investigate the prevalence of β-lactamase and the genomic clonality of a large collection of Kingella kingae isolates from Israeli patients with a variety of invasive infections and asymptomatic pharyngeal carriers. β-lactamase production was studied by the nitrocefin method and the minimum inhibitory concentrations (MICs) of penicillin and amoxicillin–clavulanate were determined by the epsilon (Etest) method. The genotypic clonality of isolates was investigated by pulsed-field electrophoresis (PFGE). β-lactamase was found in 2 of 190 (1.1 %) invasive isolates and in 66 of 429 (15.4 %) randomly chosen carriage organisms (p < 0.001). Overall, 73 distinct PFGE clones were identified (33 among invasive organisms and 56 among carriage isolates). β-lactamase production was found to be limited to four distinct PFGE clones, which were common among carriage strains but rare among invasive strains, and all organisms in the collection belonging to these four clones expressed β-lactamase. The penicillin MIC of β-lactamase-producing isolates ranged between 0.094 and 2 mcg/mL (MIC50: 0.25 mcg/mL; MIC90: 1.5 mcg/mL) and that of amoxicillin–clavulanate between 0.064 and 0.47 mcg/mL (MIC50: 0.125 mcg/mL; MIC90: 0.125 mcg/mL). The penicillin MIC of β-lactamase non-producing isolates ranged between

Objective: The aim of this study was to analyze the NP/OP S. pneumoniae serotype distribution and potential vaccine coverage in Costa Rican children with Otitis Media (OM) before the introduction of PCV-7 in the National Immunization Program (NIP). Methods: Between 2002 and 2006, NP and OP samples were obtained from 641 children from 6 to 79 months of age, at the time of OM diagnosis. S. pneumoniae serotyping and antimicrobial susceptibility were performed. Results: 386 S. pneumoniae isolates were recovered. The most common S. pneumoniae serotypes (ST) were: ST 6B, ST 14, ST 19F. Penicillin non-susceptibility was observed among 57% of the isolates obtained from children < 24 months of age. 15% strains were multidrug resistant. Potential vaccine coverage was: PCV-7: 60%; PCV-10: 62%; and PCV-13: 76% and against penicillin non-susceptible and multidrug resistant isolates was: PCV-7; 59% and 83%, respectively; PCV-10: 60% and 85%, respectively and PCV-13: 74% and 96%, respectively. Conclusions: S. pneumoniae was isolated from the NP and/or OP in the majority (59%) of studied children with OM. At a statistical significant level, only serotype 3 was more frequently isolated among children >24 months of age. Antibiotic non-susceptibility and MDR were significantly higher in children

In Streptococcus pneumonia, phosphoenolpyruvate protein phosphotransferase (PtsA) is an intracellular protein of the monosaccharide phosphotransferase systems. Biochemical and immunostaining methods were applied to show that PtsA also localizes to the bacterial cell-wall. Thus, it was suspected that PtsA has functions other than its main cytoplasmic enzymatic role. Indeed, recombinant PtsA and anti-rPtsA antiserum were shown to inhibit adhesion of S. pneumoniae to cultured human lung adenocarcinoma A549 cells. Screening of a combinatorial peptide library expressed in a filamentous phage with rPtsA identified epitopes that were capable of inhibiting S. pneumoniae adhesion to A549 cells. The insert peptides in the phages were sequenced, and homologous sequences were found in human BMPER, multimerin1, protocadherin19, integrinβ4, epsin1 and collagen type VIIα1 proteins, all of which can be found in A549 cells except the latter. Six peptides, synthesized according to the homologous sequences in the human proteins, specifically bound rPtsA in the micromolar range and significantly inhibited pneumococcal adhesion in vitro to lung- and tracheal-derived cell lines. In addition, the tested peptides inhibited lung colonization after intranasal inoculation of mice with S. pneumoniae. Immunization with rPtsA protected the mice against a sublethal intranasal and a lethal intravenous pneumococcal challenge. In addition, mouse anti rPtsA antiserum reduced bacterial virulence in the intravenous inoculation mouse model. These findings showed that the surface-localized PtsA functions as an adhesin, PtsA binding peptides derived from its putative target molecules can be considered for future development of therapeutics, and rPtsA should be regarded as a candidate for vaccine development.

Background A prior 7- and 13-valent pneumococcal conjugate vaccine (PCV7 and PCV13) study provided sufficient data ( N = 1754; Jewish, n = 1154; Bedouin, n = 595; other, n = 5) to investigate the association between nasopharyngeal (NP) acquisition of common PCV7 serotypes and cross-reacting 6A (PCV7 + 6A) and IgG concentrations. Methods Using a logistic regression model, serotype specific association between postinfant series IgG concentration (age 7 months) and new NP acquisition between ages 7 and 24 months was assessed and adjusted for ethnicity. From a subset of subjects with new NP acquisition ( n = 9–152 across serotypes studied), new acquisition percentiles and associated IgG concentrations were calculated. Results For the serotypes studied , new NP acquisition rates decreased as IgG concentrations increased. Ethnicity did not influence these associations despite differences in carriage rates. From the subset with new acquisitions, 50% of the events occurred at IgG concentrations >0.61–5.58 μg/mL; and 10% of the acquisitions occurred at IgG concentrations >2.48–17.69 μg/mL. Conclusion Remarkably high IgG concentrations are required to reduce NP acquisition. These IgG concentrations differ between serotypes. Ethnicity did not influence the association between high IgG concentrations and prevention of carriage despite differences in carriage rates. Since carriage determines transmission, these results may have important implications for herd protection. Trial registration: ClinicalTrials.gov number, NCT00508742 ; http://clinicaltrials.gov/ct2/show/NCT00508742

Background. Despite the increasing recognition of Kingella kingae as an important pathogen of early childhood, the relative frequency and invasiveness of different strains of the organism has not been investigated. A study was conducted to determine the association of K. kingae genotypes with specific clinical syndromes and the temporal and geographic distribution of invasive clones. Methods. A collection of 181 invasive K. kingae strains, isolated between 1991 and 2012 from Israeli patients with bacteremia, skeletal system infections, or endocarditis, were typed by pulsed-field gel electrophoresis (PFGE). In addition, the correspondence between PFGE, multilocus sequence types (MLSTs), and rtxA gene sequencing results was also examined for organisms belonging to the predominant PFGE clones isolated from asymptomatic carriers and patients with invasive infections. Results. A total of 32 different K. kingae clones were identified by PFGE, of which 5 (B, H, K, N, and P) caused 72.9% of all invasive infections, and were recovered during the 21-year period from different regions of the country. Clone K was significantly associated with bacteremia, clone N with skeletal system infections, and clone P with bacterial endocarditis. Strains belonging to the same PFGE clone, either carried asymptomatically or causing different invasive infections, shared MLST complexes and exhibited identical or closely related rtxA alleles. Conclusions. Although K. kingae exhibits noteworthy genetic heterogeneity, a limited number of distinct clones cause the majority of invasive infections in Israel, exhibiting genetic stability, long-term persistence, and wide geographic dispersal. K. kingae strains also show significant association with specific clinical syndromes.

The heptavalent pneumococcal conjugate vaccine (PCV-7) was introduced in high risk children and into the private market in Costa Rica in 2004 (5% annual birth cohort). The aim of this study was to compare the Streptococcus pneumoniae serotype (ST) distribution, antibiotic resistance patterns and potential coverage before and after partial introduction of PCV-7.A comparison between the S. pneumoniae isolates obtained and serotyped from the middle ear fluid (MEF) of Costa Rican children with otitis media between years 1999 and 2003 (before PCV-7 usage) and those isolates obtained from 2004 to 2008.A total of 145 and 218 MEF S. pneumoniae were serotyped between years 1999 and 2003 and 2004 and 2008, respectively. Considering a 19F outbreak observed between years 1999 and 2003, the following statistically significant changes in serotype distribution were detected between 1999 and 2003 and 2004 and 2008: ST 3: 4.8-12.8% (P=0.01); ST 11A: 0-4.1% (P=0.01); ST 14: 3.5-21.1% (P0.001) and ST 19F: 52.4-18.3% (P0.05). Comparison of the two study periods demonstrated that during 2004 and 2008 a statistically significant decrease in penicillin non-susceptible serotypes (36.2-20.4% [P=0.003]) and a statistically significant increase in trimethoprim-sulfametoxazole resistant serotypes (54.9-68.5%, respectively [P=0.03]) was observed. Potential pneumococcal vaccines coverage between 1999 and 2003 and between 2004 and 2008 were: for PCV-7: 77.2-60.5%, respectively (P=0.001); for the 10-valent conjugated vaccine (PCV-10): 78.6-61.4%, respectively (P=0.0008) and for the 13-valent conjugated vaccine (PCV-13): 84.8-79.3%, respectively (P=0.2).Changes in the serotype distribution and antimicrobial susceptibility of MEF S. pneumoniae have been observed in Costa Rican children with OM. Because of the limited use of PCV-7 during the study period, these changes probably cannot be attributed to PCV-7 use. Between 2004 and 2008, PCV-13 offered the highest potential vaccine coverage.

FabF elongation condensing enzyme is a critical factor in determining the spectrum of products produced by the FASII pathway. Its active site contains a critical cysteine-thiol residue, which is a plausible target for oxidation by H2O2. Streptococcus pneumoniae produces exceptionally high levels of H2O2, mainly through the conversion of pyruvate to acetyl-P via pyruvate oxidase (SpxB). We present evidence showing that endogenous H2O2 inhibits FabF activity by specifically oxidizing its active site cysteine-thiol residue. Thiol trapping methods revealed that one of the three FabF cysteines in the wild-type strain was oxidized, whereas in an spxB mutant, defective in H2O2 production, none of the cysteines was oxidized, indicating that the difference in FabF redox state originated from endogenous H2O2. In vitro exposure of the spxB mutant to various H2O2 concentrations further confirmed that only one cysteine residue was susceptible to oxidation. By blocking FabF active site cysteine with cerulenin we show that the oxidized cysteine was the catalytic one. Inhibition of FabF activity by either H2O2 or cerulenin resulted in altered membrane fatty acid composition. We conclude that FabF activity is inhibited by H2O2 produced by S. pneumoniae.

During an outbreak of severe pneumonia among new army recruits, an epidemiological investigation combined with repeated nasopharyngeal/oropharyngeal cultures from sick and healthy contacts subjects was conducted. Fifteen pneumonia cases and 19 influenza-like illness cases occurred among 596 recruits over a 4-week period in December 2005. Pneumonia attack rates reached up to 5.5%. A single pneumococcus serotype 5 clone was isolated from blood or sputum cultures in 4 patients and 30/124 (24.1%) contacts. Immunization with 23-valent polysaccharide vaccine supplemented with a 2-dose azithromycin mass treatment rapidly terminated the outbreak. Carriage rates dropped to

Knowledge of the in vivo physiology and metabolism of Streptococcus pneumoniae is limited, even though pneumococci rely on efficient acquisition and metabolism of the host nutrients for growth and survival. Because the nutrient-limited, hypoxic host tissues favor mixed-acid fermentation, we studied the role of the pneumococcal pyruvate formate lyase (PFL), a key enzyme in mixed-acid fermentation, which is activated posttranslationally by PFL-activating enzyme (PFL-AE). Mutations were introduced to two putative pfl genes, SPD0235 and SPD0420, and two putative pfl A genes, SPD0229 and SPD1774. End-product analysis showed that there was no formate, the main end product of the reaction catalyzed by PFL, produced by mutants defective in SPD0420 and SPD1774, indicating that SPD0420 codes for PFL and SPD1774 for putative PFL-AE. Expression of SPD0420 was elevated in galactose-containing medium in anaerobiosis compared to growth in glucose, and the mutation of SPD0420 resulted in the upregulation of fba and pyk , encoding, respectively, fructose 1,6-bisphosphate aldolase and pyruvate kinase, under the same conditions. In addition, an altered fatty acid composition was detected in SPD0420 and SPD1774 mutants. Mice infected intranasally with the SPD0420 and SPD1774 mutants survived significantly longer than the wild type-infected cohort, and bacteremia developed later in the mutant cohort than in the wild type-infected group. Furthermore, the numbers of CFU of the SPD0420 mutant were lower in the nasopharynx and the lungs after intranasal infection, and fewer numbers of mutant CFU than of wild-type CFU were recovered from blood specimens after intravenous infection. The results demonstrate that there is a direct link between pneumococcal fermentative metabolism and virulence.

Background: Although Kingella kingae is being increasingly recognized as an important pediatric pathogen, our current understanding of the transmission of the organism is limited. The dissemination of K. kingae in the community was studied in 2 ethnic groups living side-by-side in Southern Israel. Methods: Organisms recovered from oropharyngeal cultures, obtained from healthy young Jewish and Bedouin children during a 12-month period, were typed by pulsed-field gel electrophoresis and compared. Results: Isolates from Bedouin children usually differed from those derived from Jews, confirming the relative social isolation of the 2 populations and the importance of close mingling in the spread of K. kingae. Significant clustering of genotypic clones in households and Bedouin neighborhoods was observed, indicating person-to-person transmission through intimate contact. Organisms detected in the study were identical to historical isolates recovered over the last 15 years from respiratory carriers and patients with bacteremia or skeletal infections. Conclusions: The present study demonstrates that children may be asymptomatically colonized in the respiratory tract by virulent K. kingae clones. The organism is transmitted from child-to-child through intimate contact. Some strains exhibit increased fitness and are maintained in the population for prolonged periods.

The prevalence of skin colonisation with Acinetobacter baumannii (ACBA) on admission to the medical intensive care unit (MICU) was studied in an institution endemic for ACBA bloodstream infections (BSIs). The impact of 4% chlorhexidine gluconate (4% CG) whole-body washing on the patients' ACBA skin colonisation was also determined. A prospective cohort trial in a MICU during March 2002 to December 2003 was performed, with a comparison between the prevalence and incidence of ACBA-BSIs obtained after intervention and retrospectively. During the intervention period, ACBA skin-screening swabs were taken from all patients on admission and periodically until discharge. Patients underwent whole-body disinfection with 4% CG immediately after obtaining the initial cultures. Disinfection was carried out on a daily basis until discharge, regardless of colonisation status. Of the 320 patients at ward admission, 55 (17%) yielded ACBA. The prevalence of ACBA colonisation among the remaining MICU patients was 5.5% at 24h and 1% at 48h following the disinfection regimen (P=0.002, OR: 2.4). Following a second screen, 80% of colonised patients were decolonised. Prevalence of ACBA-BSIs decreased from 4.6 to 0.6 per 100 patients (P < or = 0.001; OR: 7.6) and incidence decreased from 7.8 to 1.25 (85% reduction). We conclude that daily whole-body disinfection with 4% CG significantly reduced ACBA skin colonisation. This regimen may be considered in addition to well-known infection control measures, particularly in institutions with endemic rates of multidrug-resistant ACBA-BSIs.

As part of its aerobic metabolism, Streptococcus pneumoniae generates high levels of H2O2 by pyruvate oxidase (SpxB), which can be further reduced to yield the damaging hydroxyl radicals via the Fenton reaction. A universal conserved adaptation response observed among bacteria is the adjustment of the membrane fatty acids to various growth conditions. The aim of the present study was to reveal the effect of endogenous reactive oxygen species (ROS) formation on membrane composition of S. pneumoniae. Blocking carbon aerobic metabolism, by growing the bacteria at anaerobic conditions or by the truncation of the spxB gene, resulted in a significant enhancement in fatty acid unsaturation, mainly cis-vaccenic acid. Moreover, reducing the level of OH· by growing the bacteria at acidic pH, or in the presence of an OH· scavenger (salicylate), resulted in increased fatty acid unsaturation, similar to that obtained under anaerobic conditions. RT-PCR results demonstrated that this change does not originate from a change in mRNA expression level of the fatty acid synthase II genes. We suggest that endogenous ROS play an important regulatory role in membrane adaptation, allowing the survival of this anaerobic organism at aerobic environments of the host.

Objectives To describe the results of the epidemiologic investigation of outbreaks of invasive Kingella kingae infections among attendees at daycare facilities located in 4 closed communities in Israel. Study design The preschool-aged population of communities with clusters of Kingella cases had oropharyngeal cultures performed. K kingae isolates from infected patients and healthy contacts were genotyped by pulsed field gel electrophoresis to determine the spread of outbreak strains. Results The affected closed communities (3 military bases and 1 “kibbutz” commune) were characterized by tight social and family networks and intensive mingling. The outbreaks affected 9 of 51 attendees (attack rate: 17.6%) age 8-19 months (median: 12 months), within a 21-day period. Cases included skeletal system infections (n = 8) and bacteremia (n = 1); K kingae isolates were confirmed by the use of blood culture vials and selective media. Clinical presentation was mild and acute-phase reactants were usually normal or only moderately elevated. Thirty out of 55 (54.5%) asymptomatic children carried the outbreak strains. Analysis of the 3 clusters in which the entire preschool-aged population was cultured revealed that 31 of 71 (43.7%) children younger than 24 months of age were colonized with K kingae organisms compared with 8 of 105 (7.6%) older children (P Conclusions Clusters of invasive K kingae infections characterized by sudden onset, high attack rate, and wide dissemination of the outbreak strain can occur in daycare facilities and closed communities. Because the mild clinical presentation of invasive K kingae infections and the fastidious nature of the organism, a high index of suspicion and use of sensitive detection methods are recommended.

Acute otitis media (AOM) microbiology was evaluated in children after 7-valent pneumococcal conjugate vaccine (PCV7) introduction in Costa Rica (private sector, 2004; National Immunization Program, 2009). This was a combined prospective and retrospective study conducted in a routine clinical setting in San José, Costa Rica. In the prospective part of the study, which was conducted post-PCV7 introduction (2010–2012), standard bacteriological procedures were used to evaluate the etiology and serotype distribution of middle ear fluid samples collected by tympanocentesis or otorrhea from children aged 3–59 months diagnosed with AOM. E-tests were used to evaluate antimicrobial susceptibility in culture-positive samples. Retrospective data recorded between 1999 and 2004 were used for comparison of bacterial etiology and serotype distribution before and after PCV7 introduction. Statistical significance was evaluated in bivariate analyses at the P-value 1 year of age) versus unvaccinated children. S. pneumoniae non-susceptibility rates were 1.1%, 34.5%, 31.7%, and 50.6% for penicillin, erythromycin, azithromycin, and trimethoprim/sulfamethoxazole (TMP-SMX), respectively. H. influenzae non-susceptibility rate was 66.9% for TMP-SMX. Between pre- and post-PCV7 introduction, H. influenzae became more (20.5% vs 25.9%; P-value

Background In a recent epidemiological study in southern Israel, nontypable Streptococcus pneumoniae isolates were found to be highly associated with sporadic cases of acute conjunctivitis (AC). The purpose of the present study was to evaluate the relative importance in causing AC of the absence of capsule versus genotype properties. Methods DNA typing by pulsed-field gel electrophoresis (PFGE) was performed on 148 nontypable organisms isolated from 3 sites: nasopharynx of healthy children, middle-ear fluid, and conjunctiva. Results Analysis of the PFGE patterns revealed the presence of 6 clusters; 2 clusters that included 44% of the isolates (65/148) were associated with AC, and the remaining 4 were frequently isolated from the nasopharynx. Multilocus sequence typing, performed on representative isolates of the 2 major clusters, confirmed that the organisms were pneumococci; one is a single-locus variant of sequence type (ST) 448, and the other is related to ST344. Both types appear to be members of pneumococcal lineages that have lost capsular loci. The nontypable isolates showed high rates of resistance to antimicrobial agents. Conclusions Our data suggest that the absence of the capsule--along with other, yet-unidentified genetic characteristics--provide S. pneumoniae with a selective virulence advantage in conjunctivitis.

Recent studies have shown that some Streptococcus pneumoniae serotypes possess a higher potential to cause invasive disease than others. However, it is unknown whether disease potential for specific serotypes is similar for mucosal disease. Our objective was to assess the disease potential of individual S. pneumoniae serotypes causing invasive pneumococcal disease (IPD), acute otitis media (AOM) and acute conjunctivitis (AC) in children.Serotypes of pneumococcal isolates from children with IPD, AOM and AC were compared with those carried by healthy children aged3 years. All children resided in the same area and were studied during the same period. Odds ratios for disease were calculated for each diagnosis following multivariate analysis, including gender, age, ethnic group, previous antibiotic treatment and year variability.A total of 5,500 isolates were collected: 189 from blood or cerebrospinal fluid, 3,200 from middle ear fluid, 348 from conjunctiva and 1,763 from nasopharynx of healthy children. A significant positive association with IPD was demonstrated for serotypes 1, 5 and 12F; with AOM for serotypes 1, 3, 5, 12F, 19A and 19F; and with AC for serotype 3 and nontypeable S. pneumoniae. A significant negative association with IPD was demonstrated for nontypeable S. pneumoniae and with AOM for serotypes 6A, 6B, 15A and nontypeable S. pneumoniae.Our results reflect the importance of the polysaccharide capsule in site-specific disease potential and provide useful information regarding disease potential of nonvaccine serotypes shown to be involved in carriage replacement after vaccination with the 7-valent conjugate vaccine.

Fourier transform infrared (FTIR) spectroscopy has shown remarkable ability in distinguishing between bacterial species and identifying bacterial colony structures, when used in tandem with methods such as cluster analysis, principal component analysis, or linear discriminant analysis. The present work was aimed to evaluate the potential of FTIR–microscopy (FTIR-MSP) to distinguish between different serotypes and capsular quantities of Streptococcus pneumoniae. In general, the results obtained have consistently proven that the spectral information at the region 900–1185 cm−1 was sufficient to distinguish between various pneumococcal serotypes. Moreover, the method was able to differentiate between S. pneumoniae phase variants on the basis of their relative carbohydrate content. The unsupervised cluster analysis of the samples showed differences, not only in the carbohydrate content, but also in the region 1350–1480 cm−1, which is dominated by absorptions due to lipids and phospholipids. This approach proved to be useful for the distinction between S. pneumoniae serotypes and between phase variants, which were shown to acquire different pathogenic capacity. © 2006 Wiley Periodicals, Inc. Biopolymers 83:434–442, 2006 This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

The rate of Streptococcus pneumoniae carriage among adults was compared with that among children (age,or =6 years) in the same population. Nasopharyngeal culture results for 1300 adults and 404 children were analyzed. S. pneumoniae was carried by only 4% of the adults, compared with 53% of children in the same community. Young age, day care center attendance, having young siblings, and no antibiotic use during the month before screening were associated with the high carriage rate among children, whereas the only risk factor associated with carriage among adults was the presence of a respiratory infection on the screening day. S. pneumoniae serotype distribution and antibiotic resistance patterns differed between adults and children. Isolates of the same serotype--even of the same clone--differed in their antibiotic susceptibility patterns between children and adults. In a subanalysis of 151 pairs of children and their parents and of 32 pairs of siblings, intrafamilial transmission of S. pneumoniae could not be demonstrated.

This study examined the prevalence of antibiotic-resistant clones that belong to serotypes not included in the pneumococcal conjugate vaccines and that cause a significant percentage of acute otitis media (AOM) in children in southern Israel. During 1998-2001, 2467 pneumococcal isolates, obtained from middle-ear fluid of children

Because of the increasing number of resistant middle ear pathogens and the impact of the new conjugate Streptococcus pneumoniae vaccine, an active surveillance of the microbiology and susceptibility pattern of middle ear pathogens is required.To study the microbiology and susceptibility pattern of middle ear pathogens obtained from Costa Rican children with acute otitis media (AOM), recurrent otitis media (ROM) and therapeutic failure otitis media (FOM).Between 1999 and 2001 middle ear fluid (MEF) was collected from 276 Costa Rican patients. S. pneumoniae serotyping and pulsed field gel electrophoresis analysis was done on available strains.Among the total study population, 102 were AOM patients, 98 were ROM patients and 76 were FOM patients. Overall S. pneumoniae (88 strains) was the most common pathogen isolated followed by Haemophilus influenzae (41 strains) and Streptococcus pyogenes (10 strains). H. influenzae was the most common agent in FOM patients (P = 0.015). Beta-lactamase production was observed in 3 of 41 (7%) H. influenzae strains and 3 of 3 (100%) Moraxella catarrhalis strains. Penicillin-nonsusceptible S. pneumoniae strains were more common in FOM (64%) and ROM (63%) patients than in AOM (42%) patients (P = 0.05). S. pneumoniae serotype 19F was the most prevalent serotype, mainly within one distinct clone.Overall S. pneumoniae serotype 19F was the most common isolate from the middle ear fluid of Costa Rican children. Beta-lactamase-negative H. influenzae was the most prevalent in the subpopulation of patients with FOM. S. pyogenes was the third most common isolate and M. catarrhalis was uncommon.

We describe a child with interleukin-12 receptor beta1 (IL-12Rbeta1) deficiency caused by a homozygous IL12RB1 large deletion who presented at the age of 1 year with recurrent, often asymptomatic episodes of bacteremia caused by group D Salmonella species. No mycobacterial disease or other unusual infection was present. The episodes of salmonellosis were caused by an identical serovar during a period of 18 months. This is the first case of inherited IL-12Rbeta1 deficiency diagnosed after isolated, recurrent salmonellosis.

We describe an outbreak of Acinetobacter baumannii in a neonatal intensive care unit (NICU), and our investigation to determine the source and mode of transmission and identify the population at risk. A case (infected infant) was defined as a patient hospitalized in the NICU during the outbreak period, with clinical signs of sepsis and isolation of A. baumannii. In colonized infants, A. baumannii was isolated from body surfaces without signs of infection. Infected infants were separated and treated by a different medical team. Cultures were taken from working surfaces and along the infant's admission passage from the delivery room to the NICU. The outbreak strain was identified by pulsed-field gel electrophoresis (PFGE). Nine cases and eight colonized infants met the definition criteria. Cases were younger than colonized infants with regard to gestational age and age of diagnosis and had lower birthweights (P

We aimed to study the prevalence of Streptococcus pneumoniae in respiratory samples from institutionalized patients with chronic tracheostomy. A total of 264 pairs of nasopharyngeal and endotracheal cultures were collected. There was no difference in the proportion of positive cultures between children (21%) and adults (18%). However, the proportion of positive endotracheal cultures was higher than that of nasopharyngeal cultures in adults (18% versus 3%, respectively; P < 0.001) but not in children (17% in both sites).

Methicillin-resistant Staphylococcus aureus (MRSA) has recently been reported to emerge in the community setting. We describe the investigation and control of a community-acquired outbreak of MRSA skin infections in a closed community of institutionalized adults with developmental disabilities. In a 9-month period in 1997, 20 (71%) of 28 residents had 73 infectious episodes. Of the cultures, 60% and 32% obtained from residents and personnel, respectively, grew S. aureus; 96% and 27% were MRSA. All isolates were genetically related by pulsed-field gel electrophoresis and belonged to a phage type not previously described in the region. No known risk factors for MRSA acquisition were found. However, 58 antibiotic courses had been administered to 16 residents during the preceding 9 months. Infection control measures, antibiotic restriction, and appropriate therapy resulted in successful termination of this outbreak. Selective antibiotic pressure may result in the emergence, persistence, and dissemination of MRSA strains, causing prolonged disease.

The annual incidence of gonorrhea in Israel has sharply increased during the past 2 years. At the end of 1999, high-level ciprofloxacin-resistant strains of Neisseria gonorrhoeae (MIC90,or =32 mcg/ml) also exhibiting decreased susceptibility to penicillin and tetracycline were isolated for the first time in southern Israel, as well as in other regions of the country. The incidence of male gonococcal urethritis in the south increased in a 1.5-year period from 3/100,000 to 12/100,000 ( P0.05) in correlation with increased isolation of ciprofloxacin-resistant organisms. A marked increase in the incidence of gonorrhea was also encountered in Jerusalem, where ciprofloxacin resistance affected 54.5% of the isolates in 2000. Pulsed-field gel electrophoresis typing of gonococci from different areas of Israel indicated that all of the ciprofloxacin-resistant isolates belonged to identical or related strains. Since fluoroquinolone-resistant gonococci may emerge and disseminate extensively over a short period of time, continuous surveillance of antibiotic susceptibility among gonococcal isolates should be performed to guide empiric therapy.

Background Preterm infants are at high risk for extended spectrum β-lactamase producing Enterobacteriaceae (ESBL-E) sepsis and neonatal intensive care unit (NICU) outbreaks. There is no consensus regarding surveillance of pregnant women for ESBL-E colonization. However, neonatal screening is accepted in order to prevent NICU’s transmissions by contact isolation. We hypothesized that a significant proportion of ESBL-E colonized infants in our NICU were due to mother-to-child transmission. The aim of this study was to molecularly identify pairs of mothers and offspring ESBL-E colonization. Methods The Soroka University Medical Center serves the entire population of southern Israel with an estimated 17,000 deliveries per year. This was a one year analysis from an ongoing, prospective, active rectal ESBL-E colonization surveillance of mothers of premature infants and their offspring. Pairs of mothers and infants colonized with the same bacteria underwent molecular identification by pulse-field gel electrophoresis (PFGE). Results Between January 2015 and February 2016, 311(76%) out of 407 mothers and all 477(100%) infants were screened for ESBL-E colonization; carriage rates were 21.5% and 14.9%, respectively. Four (5.6%) carrier infants developed late onset sepsis and 2 (2.8%) died. Mothers of 47% (28) ESBL-E carrier infants were colonized at delivery; comparison of the characteristics between colonized infants of positively and negatively screened mothers is presented in Table 1. Twenty-three (34.3%) colonized mothers delivered 25 infants who were found to be colonized with the same bacterial strain according to species’ identification and antibiogram. A subgroup of 10 mother-infant dyads underwent PFGE and 70% shared identical molecular fingerprint patterns. No similarities were found between isolates recovered from unrelated neonates and mothers. Conclusion High ESBL-E carriage rates in mothers and NICU’S infants, in our region, with a non-negligible maternal-neonatal ESBL-E colonization transmission illuminates the importance of maternal ESBL colonization surveillance and further consideration regarding infectious control preventive measurements. Disclosures All authors: No reported disclosures.

BACKGROUND Pediatric respiratory infections caused by antibiotic-nonsusceptible Streptococcus pneumoniae (ANSP) continue to present an important challenge, even after introduction of 7-valent pneumococcal conjugate vaccine (PCV7). This randomized double-blind trial assessed the potential additional impact of PCV13 over PCV7 on reducing ANSP carriage. METHODS Healthy infants were randomly assigned to receive PCV13 (n = 932) or PCV7 (n = 934) at ages 2, 4, 6, or 12 months. Eight nasopharyngeal specimens were collected by swabbing between ages 2 and 24 months. S. pneumoniae isolates were serotyped and tested for antimicrobial susceptibility by the disk-diffusion method and the Etest. Nasopharyngeal acquisition and prevalence of ANSP during ages 7-24 months were compared between the 2 vaccine groups. RESULTS In general, new acquisition of pneumococci nonsusceptible to penicillin, erythromycin, clindamycin, penicillin plus erythromycin, and multiple drugs (≥3 antibiotics) was significantly lower in the PCV13 group compared with the PCV7 group; the main serotypes contributing to this significant decrease were serotype 19F, present in PCV13 and PCV7, and serotypes 6A and 19A, present in PCV13 only. CONCLUSIONS PCV13 has a significant added benefit over PCV7 in reducing carriage of ANSP. Because carriage determines transmission, these results suggest that PCV13 will provide protection against ANSP disease that exceeds protection provided by PCV7. CLINICAL TRIALS REGISTRATION NCT00508742.

Although antibiotic-resistant pneumococci have been frequently detected among day care center (DCC) attendees, the transmission of these organisms to other members of the community has not been adequately studied. Nasopharyngeal cultures were obtained from 152 children and 244 adult members of a closed community (a kibbutz) in Israel. Serotyping, antibiogram, and pulsed-field gel electrophoresis were performed to determine the relatedness of isolated pneumococci. Twenty (30%) of the 66 isolates from children showed decreased susceptibility to penicillin and 9 isolates (14%) were resistant to > or =3 drugs. Of the 16 isolates from adults, 5 (31%) were intermediately resistant to trimethoprim-sulfamethoxazole. Resistant strains carried by DCC attendees were not isolated either from their parents or from other adult members of the community. Despite the high degree of interpersonal contact occurring in a closed community, resistant pneumococcal strains carried by DCC attendees do not appear to be easily transmitted to the adult population, which suggests the existence of an immunological barrier.

Antibiotic-resistant pneumococci are difficult to eradicate from middle ear fluid (MEF) and the nasopharynx (NP). Bacteriologic eradication from the NP and MEF during acute otitis media (AOM) by 3 common antibiotic drugs was prospectively evaluated. In 19 (16%) of 119 MEF culture-positive patients, an organism susceptible to the treatment drug (Haemophilus influenzae, Streptococcus pneumoniae, or both) was isolated from the initial MEF, whereas resistant S. pneumoniae was present in the NP; in 9 (47%) patients, the initial resistant NP organism (identified by serotyping, resistance to the administered drug, and pulsed-field gel electrophoresis) replaced the susceptible MEF organism within only a few days after initiation of treatment. In regions where resistant pneumococci are prevalent, antibiotics may not only fail to eradicate the organisms, but they may often induce MEF superinfection with resistant pneumococci initially carried in the NP. This is an important mechanism by which, in recently treated patients, AOM infections often become refractory to treatment.

SummaryWe describe the clinical course of a previously healthy 20-month-old toddler admitted with high fever and leukocytosis. Blood culture grew Streptococcus pneumoniae serotype 19A, belonging to the ST663 clone, highly resistant to penicillin, ceftriaxone, and erythromycin. The same clone with identical antibiogram was isolated from the nasopharynx of another three of the other five healthy children attending the same daycare center as the patient. This case exemplifies the potential problems posed by highly-resistant S. pneumoniae serotype 19A, an emerging pathogen worldwide.

In 1996, 19 isolates of serotype 6B Streptococcus pneumoniae with a unique resistance pattern were found in carriers attending daycare centres in Patras, Southwestern Greece. These isolates were penicillin susceptible but resistant to chloramphenicol, tetracycline, erythromycin, clindamycin and trimethoprim-sulphamethoxazole. Subsequently, isolates with the same characteristics were found in 23 additional carriers in central and southern Greece in 1997–98 as well as in 19 carriers in central Italy in 1997, and in seven carriers in southern Israel in 1998. Carriers were all children under 6 years of age, attending daycare centres or outpatient hospital visits. The relatedness of the isolates was determined on representative isolates from the three countries by pulsed-field gel electrophoresis of Sma I digests of chromosomal DNA. Most Greek isolates were identical to each other, while isolates from Italy and Israel showed one to three band differences, with all isolates being closely related to each other as well as to the isolates from Greece. We have therefore documented the presence of this unique clone of S. pneumoniae in these three countries and have named this the ‘Mediterranean' clone. While isolates appear to have a common origin, their source and direction of spread are unknown. However, isolates from Italy showed the most diversity, suggesting that this clone had been present in that country for a longer period than it had been in Greece.