Spn Strains Research Articles

1312. Clinical and Economic Burden of Otitis Media in Children under 5 in the United States

Abstract Background Otitis media (OM) is one of the most common childhood illnesses and frequent cause of antibiotic use among patients under 5 years old. Pneumococcal conjugate vaccines (PCVs) led to a reduction in the burden of OM caused by Streptococcus pneumoniae (Spn) strains in the United State (US). Overall, OM incidence was reduced after introduction of PCV7 and plateaued in the late PCV13 era, though there was an increase in OM episodes caused by non-typeable Haemophilus influenzae and non-vaccine-type Spn. The objectives of this study were to assess the incidence and treatment costs of OM episodes and their complications in the US. Methods This retrospective observational study used data from IBM MarketScan to study pediatric OM incidence and complications (tympanostomy tube placement [TTP], hearing loss [HL] and hospitalizations) by age groups and their associated costs. OM episodes occurring between January 1, 2016, and December 31, 2017, were considered in this analysis. Results of the total cost are weighted by the proportion of children covered under commercial insurance and Medicaid, respectively. Results The incidence of OM among children < 5 increased gradually with age, reaching a peak of incidence between 1 and 2 years (figure 1) and more precisely between 6 and 15 months. Overall, most OM episodes were without HL and treated in an outpatient setting without TTP (∼ 94% of cases); OM episodes without HL requiring TTP represented only 4% and the vast majority were performed in an outpatient setting ( > 99%). OM without HL leading to hospitalization was very rare (< 0.025%) for both plans. OM leading to HL corresponds to 2% of all OM episodes. The incidence rate and the mean cost per episode were used to extrapolate an overall economic burden of ∼5 billion USD in 2020, driven by outpatient episodes (50%) followed by OM episodes plus TTP (37%), and OM episodes plus HL (13%). Figure 1:Incidence rates by age groups for (A) Commercial/Medicare; (B) Medicaid Conclusion The burden associated with OM in children < 5 reflects a significant and ongoing unmet need. Thus, prevention of OM not currently covered by existing vaccines is warranted to address the significant burden including the potential emergence of antimicrobial resistance. Funding GlaxoSmithKline Biologicals SA (GSK study identifier: VEO-000264]) Disclosures Lilia Ben Debba, PharmaD, GlaxoSmithKline SA: Advisor/Consultant Dominique Derreumaux, M.Sc, GlaxoSmithKline SA: is employed by the GSK group of companies Jessica Pickett, PhD, GlaxoSmithKline SA: is employed by and hold shares in the GSK group of companies Laura Taddei, M.Sc, GlaxoSmithKline SA: is employed by and hold shares in the GSK group of companies Ekkehard Beck, PhD, GlaxoSmithKline SA: is employed by and hold shares in the GSK group of companies Michael Scherbakov, PhD, GlaxoSmithKline SA: is employed by and hold shares in the GSK group of companies.

Oxidative Reactions Catalyzed by Hydrogen Peroxide Produced by Streptococcus pneumoniae and Other Streptococci Cause the Release and Degradation of Heme from Hemoglobin.

Streptococcus pneumoniae (Spn) strains cause pneumonia that kills millions every year worldwide. Spn produces Ply, a hemolysin that lyses erythrocytes releasing hemoglobin, and also produces the pro-oxidant hydrogen peroxide (Spn-H2O2) during growth. The hallmark of the pathophysiology of hemolytic diseases is the oxidation of hemoglobin, but oxidative reactions catalyzed by Spn-H2O2 have been poorly studied. We characterized the oxidation of hemoglobin by Spn-H2O2. We prepared a series of single-mutant (ΔspxB or ΔlctO), double-mutant (ΔspxB ΔlctO), and complemented strains in TIGR4, D39, and EF3030. We then utilized an in vitro model with oxyhemoglobin to demonstrate that oxyhemoglobin was oxidized rapidly, within 30 min of incubation, by Spn-H2O2 to methemoglobin and that the main source of Spn-H2O2 was pyruvate oxidase (SpxB). Moreover, extended incubation caused the release and the degradation of heme. We then assessed oxidation of hemoglobin and heme degradation by other bacterial inhabitants of the respiratory tract. All hydrogen peroxide-producing streptococci tested caused the oxidation of hemoglobin and heme degradation, whereas bacterial species that produce <1 μM H2O2 neither oxidized hemoglobin nor degraded heme. An ex vivo bacteremia model confirmed that oxidation of hemoglobin and heme degradation occurred concurrently with hemoglobin that was released from erythrocytes by Ply. Finally, gene expression studies demonstrated that heme, but not red blood cells or hemoglobin, induced upregulated transcription of the spxB gene. Oxidation of hemoglobin may be important for pathogenesis and for the symbiosis of hydrogen peroxide-producing bacteria with other species by providing nutrients such as iron.

Induction of the macrolide-resistance efflux pump Mega inhibits intoxication of Staphylococcus aureus strains by Streptococcus pneumoniae

Streptococcus pneumoniae (Spn) kills Staphylococcus aureus (Sau) through a contact-dependent mechanism that is catalyzed by cations, including iron, to convert hydrogen peroxide (H2O2) to highly toxic hydroxyl radicals (•OH). There are two well-characterized ABC transporters that contribute to the pool of iron in Spn, named Pia and Piu. Some Spn strains have acquired genes mef(E)/mel encoding another ABC trasporter (Mega) that produces an inducible efflux pump for resistance to macrolides. In macrolide-resistant Spn clinical isolates the insertion of Mega class 1. IV and 2. IVc deleted the locus piaABCD and these strains were attenuated for intoxicating Sau. The goal of this study was to investigate if the disruption of iron acquisition, or the antimicrobial-resistance activity of Mega, contributed to inhibiting the killing mechanism. Neither depletion of iron with 2,2'-dipyridyl-d8 (DP) nor incubating with a double knockout mutant SpnΔpiaAΔpiuA, inhibited killing of Sau. Clinical Spn strains carrying Mega1. IV or Mega2. IVc showed a significant delay for killing Sau. An ex vivo recombination system was used to transfer Mega1. IV or Mega2. IVc to reference Spn strains, which was confirmed by whole genome sequencing, and recombinants TIGR4Mega2. IVc, D39Mega2. IVc, and D39Mega1. IV were delayed for killing Sau. We then compared Sau killing of selected Mega-carrying Spn strains when incubated with sub-inhibitory erythromycin (Mega-induced) or sub-inhibitory cefuroxime. Remarkably, killing of Sau was completely inhibited under the Mega-induced condition whereas incubation with cefuroxime did not interfere with killing. Both mef(E) and mel were upregulated > 400-fold, and spxB (encoding an enzyme responsible for production of most H2O2) was upregulated 14.2-fold, whereas transcription of the autolysin (lytA) gene was downregulated when incubated with erythromycin. We demonstrated that erythromycin induction of Mega inhibits the •OH-mediated intoxication of Sau and that the inhibition occurred at the post-translational level suggesting that an imbalance of ions in the membrane inhibits these reactions.

Dynamic changes in otopathogens colonizing the nasopharynx and causing acute otitis media in children after 13-valent (PCV13) pneumococcal conjugate vaccination during 2015-2019.

The otopathogens colonizing the nasopharynx (NP) and causing acute otitis media (AOM) have shown dynamic changes following introduction of pneumococcal conjugate vaccines. Five hundred eighty-nine children were prospectively enrolled, 2015-2019. Two thousand fifty-nine visits (1528 healthy, 393 AOM, and 138 AOM follow-up) were studied. Two thousand forty-two NP and 495 middle ear fluid (MEF) samples by tympanocentesis from 319 AOM cases were cultured for bacterial identification and antibiotic susceptibility. Streptococcus pneumoniae (Spn) isolates were serotyped by Quellung, and multi-locus sequence type (ST) determined by genomic analysis. Haemophilus influenzae (Hi) was the most common otopathogen cultured from MEF during AOM (34% in MEF) followed by Spn (24% in MEF), then Moraxella catarrhalis (Mcat) (15% in MEF). NP isolates during healthy visit were Mcat (39%), Spn (32%), Hi (12%). 48.6% of Hi isolates from MEF were beta-lactamase-producing. Spn non-susceptibility to penicillin and other antibiotics was high. The most common Spn serotypes associated with AOM (and colonizing the NP during healthy visits) were 35B, 23B, and 15B/C. ST558 and ST199 were the most common sequence types. During 2015-2019, Hi was the most common otopathogen cultured from MEF during AOM among young children. Pneumococcal AOM was most commonly caused by non-PCV13 serotypes of Spn, predominantly 35B, 23B, and 15B/C. Resistance to common antibiotics among Spn strains showed an increasing trend.

An in vivo atlas of host–pathogen transcriptomes during Streptococcus pneumoniae colonization and disease

Streptococcus pneumoniae (Spn) colonizes the nasopharynx and can cause pneumonia. From the lungs it spreads to the bloodstream and causes organ damage. We characterized the in vivo Spn and mouse transcriptomes within the nasopharynx, lungs, blood, heart, and kidneys using three Spn strains. We identified Spn genes highly expressed at all anatomical sites and in an organ-specific manner; highly expressed genes were shown to have vital roles with knockout mutants. The in vivo bacterial transcriptome during colonization/disease was distinct from previously reported in vitro transcriptomes. Distinct Spn and host gene-expression profiles were observed during colonization and disease states, revealing specific genes/operons whereby Spn adapts to and influences host sites in vivo. We identified and experimentally verified host-defense pathways induced by Spn during invasive disease, including proinflammatory responses and the interferon response. These results shed light on the pathogenesis of Spn and identify therapeutic targets.

RETRACTED: Co infection of respiratory syncytial viruses (RSV) and Streptococcus pneumonia modulates pathogenesis and dependent of serotype and phase variant

Streptococcus pneumoniae (pneumococcus) is touted to be the generally found pathogen in patients with respiratory issues and there is an epidemiologic linkage present between Respiratory syncytial virus (RSV). This study aim at investigating the interaction between RSV and two serotypes of S. pneumoniae using a distinct animal model and a well-established colonizing pneumococcal strain. Phase variants phenotype of each strain was determined under oblique light. Co infection model was developed using BALB/c mice housed in a BSL-2 facility. Coinfection experiments were performed and number of bacterial colonies was quantified and phase determination was evaluated. RSV was detected in sample through real-time quantitative PCR. Adherence assays were performed to determine adherence of Spn strains and its knock out ΔNanA to nasopharyngeal carcinoma (NPC) epithelial CNE3 cell line. The biofilm viability was determined and phase composition was counted using plate count. Neuraminidase activity was measured in fluorometircassessed using 2'-(4-methylumbelliferyl)-α-D-N-acetylneuraminic acid (MUAN) as substrate as described in earlier literature. The GraphPad Software version 5.01 i.e., GraphPad Prism was used to conduct the statistical analysis. The extent of bacterial colonization was increased significantly (p<0.05), when the mice were co infected. Nasal epithelium remained intact in mock sample with features of a thick mucociliary border. A small percentage of pneumococci exhibit phase variation between opaque phase and transparent phase. The percentage adherent of both phase were not found to be varying significantly within serotype but it was seen that nonpathogenic type 27 was more adherent. Biofilm formation was selectively more for transparent phase from a mixed-phase inoculum. Adherence of both phase variant of S. pneumoniae to nasopharyngeal epithelial cells 2h post infection expressed as the percentage of adherent bacteria relative to the inoculum. In absence of viral infection, the nasal colonization of the opaque and the transparent variant was increased many folds, which was a significant differences. The extent of nasal colonization by the ΔNanA mutant strain were significantly reduced post-bacterial infection for both type of wild-type (P<0.05). The findings explore insights into the interactions occurring between S. pneumoniae and RSV during respiratory infections and pneumococcal acquisition, indicate that pneumococcal serotypes have different ability to cause infection as well as co infections and potentially follow an unappreciated mechanism. Much more research work is needed to further understand the minutiae of this interaction within co-infection process.

Role of phosphorylcholine in Streptococcus pneumoniae and nontypeable Haemophilus influenzae adherence to epithelial cells

ObjectivePhosphorylcholine (PC) is a structural component of Streptococcus pneumoniae (Spn) and nontypeable Haemophilus influenzae (NTHi), and is known to be associated with adherence through the platelet activating factor receptor (PAF-R). Furthermore, high PC expression is considered to be involved in Spn and NTHi virulence. In this study, we examined the influence of PC expression on the adherence of Spn and NTHi to epithelial cells in order to clarify the potential effectiveness of a vaccine targeting PC. MethodsTwenty-seven strains of Spn and twenty-two strains of NTHi were used, cultured overnight, and PC expression was evaluated by fluorescence activated cell sorting; the strains were divided into two groups: PC low expression (PC-low) and PC high expression (PC-high) groups. Bacterial adherence was then examined using Detroit 562 cells and BALB/c mice. Bacterial invasion was then examined in Detroit 562 cells. ResultsThe adherence of Spn and NTHi and invasion of NTHi in the PC-high group was significantly reduced by pretreatment with a monoclonal anti-PC antibody (TEPC-15), PAF-R antagonist (ABT-491), and PC-keyhole limpet hemocyanin (PC-KLH). However, such findings were not observed in the PC-low group. ConclusionThe present study suggests that PC is involved in the mucosal adhesion of Spn and NTHi, and the mucosal invasion of NTHi with PC-high strains, but not PC-low strains. These results suggest that a PC-targeting mucosal vaccine only affects PC-high Spn and NTHi strains and does not disturb commensal bacterial flora in the upper respiratory tract, which comprises nonpathogenic PC-low bacteria.

Expansion of the multidrug-resistant clonal complex 320 among invasive Streptococcus pneumoniae serotype 19A after the introduction of a ten-valent pneumococcal conjugate vaccine in Brazil.

BackgroundIn 2010, a ten–valent pneumococcal conjugate vaccine (PCV10) was introduced in the routine infant national immunization program in Brazil. Invasive pneumococcal disease (IPD) caused by serotype 19A (Spn19A) increased after the introduction of PCVs in several countries. We compared the frequency, antimicrobial resistance and molecular patterns of invasive Spn19A strains before and after PCV10 introduction in Brazil using data from the national laboratory-based surveillance.MethodsWe analyzed invasive Spn19A strains isolated from 2005–2009 (pre-PCV10 period), 2011–2015 and 2016–2017 (post-PCV10 periods). Antimicrobial susceptibility was performed for all Spn19A strains, and multilocus sequence typing (MLST) was performed for strains isolated in the age groups <5 years and ≥50 years.ResultsAmong the study period, a total of 9,852 invasive Spn strains were analyzed, and 673 (6.8%) belonged to serotype 19A. Overall, the proportion of Spn19A among the total number of IPD strains increased from 2.8% in 2005–2009 to 7.0% and 16.4% in 2011–2015 and 2016–2017, respectively. The relative increase in Spn19A was observed especially in children <5 years old (2005–2009: 3.2%; 2011–2015: 15.5%; 2016–2017: 31.2%). The percentage of penicillin resistance (MIC 2.0–4.0 μg/mL), erythromycin resistance and multidrug resistance (MDR) increased after PCV10 introduction due to the expansion of the MDR clonal complex CC320 (2005–2009: 8.6%; 2011–2015: 56.1%; 2016–2017: 66.5%).ConclusionWe observed an expansion of MDR-CC320 among invasive Spn19A strains after PCV10 introduction in Brazil, probably related to a combination of factors, such as vaccination and antimicrobial pressure. Continued surveillance of Spn19A strains is necessary to monitor the sustainability of this clonal complex in the Brazilian population.

Distribution of Streptococcus pneumoniae serotypes in the northeast macro-region of S\xe3o Paulo state/Brazil after the introduction of conjugate vaccine

BackgroundInfections caused by Streptococcus pneumoniae (Spn) still challenge health systems around the world, even with advances in vaccination programs. The present study evaluated the frequency of various Spn serotypes isolated in Regional Health Care Network 13 (RRAS 13), which includes the regional health departments (RHDs) of Araraquara, Barretos, Franca and Ribeirão Preto, especially after the introduction of 10-valent pneumococcal conjugate vaccine (PCV10) in 2010.MethodsThe analyzed Spn strains were isolated from patients with invasive pneumococcal diseases (IPDs) and then sent to Adolfo Lutz Institute (ALI) for further confirmative identification tests during the period from 1998 to 2013. The samples were from the cities in RRAS13, which is located in the Northeast region of São Paulo State, and totals 90 municipalities.ResultsWe analyzed strains isolated from 796 patients. They were predominantly: men (58.9%); 20 to 60 years old (32.2%); evaluated from 2003 to 2010 (60.2%); and diagnosed with meningitis (45.7%) and pneumonia (45.0%), the most common invasive pneumococcal diseases. In 2010, serotypes 3, 19F, 1, 23F, 6A and 6B were among the most frequent, while serotypes 3, 12F, 14, 6A, 18C, 8 and 6B were more common after the introduction of PCV10. Serotypes 14, 19F and 3 were more frequent in meningitis, while serotypes 14, 3 and 1 prevailed in pneumonia. After 2010, there was a decrease in serotypes 14, 1, 23F and 5 and an increase in serotypes 3, 12F, 11A and 8, which were not present in the vaccine.ConclusionsThe present study noted the increase in serotypes 3, 12F, 11A and 8 after vaccination. None of those serotypes are included in the available conjugate vaccines, which highlights the importance of continued monitoring of IPDs in order to measure the disease burden in the population in the long term and provide new epidemiological information to determine the impact of PCV10 in Brazil.

Decrease in Pneumococcal Otitis Media Cultures With Concomitant Increased Antibiotic Susceptibility in the Pneumococcal Conjugate Vaccines Era.

To study the change in the incidence and antibiotic susceptibility patterns of Streptococcus pneumoniae (Spn) in cultures obtained from children with otitis media (OM) during the pneumococcal conjugate vaccines (PCVs) era. Retrospective. Secondary medical care center. Children less than 8 years who presented with OM and had positive pneumococcal cultures during January 1, 2007 to December 12, 2014 were identified. Data recorded included demographics, preadmission antibiotics, culture source, and antibiotic susceptibility tests. We compared the pre-PCV years (2007-2008) with the transition years (2009-2011) and the post-PCV13 years (2012-2014). PCV7 (2009), PCV13 (2010), therapeutic. Annual rate of Spn cultures, antibiotic susceptibility patterns. We identified 134 children (76 boys, 57%) who contributed 162 pneumococcal cultures. There was a downward trend in the annual incidence rate of Spn cultures between the pre-PCV years, transition years, and post-PCV13 years: 11.12, 8.48, and 4.11/1000 hospitalized children/year, respectively (p = 0.08, p = 0.04). Had there been no interventions, and based on the 2007 to 2009 average, the observed over the expected Spn cultures ratio rates for 2010 to 2014 were 0.59, 0.45, 0.40, 0.40, and 0.25, respectively. In parallel, the susceptibility of Spn strains to four commonly tested antibiotics significantly increased from the pre-PCV years to the transition years and the post-PCV13 years. In each period, Spn strains were penicillin sensitive in 37, 51, and 100%; for erythromycin, 46, 71, and 82%; for trimethoprim/sulfamethoxazole, 32, 71, and 97%; and for ceftriaxone, 95, 96, and 100%, respectively. The introduction of PCVs significantly decreased the incidence rate of pneumococcal OM, and increased Spn susceptibility to common antibiotics.

Streptococcus pneumoniae Eradicates Preformed Staphylococcus aureus Biofilms through a Mechanism Requiring Physical Contact.

Staphylococcus aureus (Sau) strains are a main cause of disease, including nosocomial infections which have been linked to the production of biofilms and the propagation of antibiotic resistance strains such as methicillin-resistant Staphylococcus aureus (MRSA). A previous study found that Streptococcus pneumoniae (Spn) strains kill planktonic cultures of Sau strains. In this work, we have further evaluated in detail the eradication of Sau biofilms and investigated ultrastructural interactions of the biofilmicidal effect. Spn strain D39, which produces the competence stimulating peptide 1 (CSP1), reduced Sau biofilms within 8 h of inoculation, while TIGR4, producing CSP2, eradicated Sau biofilms and planktonic cells within 4 h. Differences were not attributed to pherotypes as other Spn strains producing different pheromones eradicated Sau within 4 h. Experiments using Transwell devices, which physically separated both species growing in the same well, demonstrated that direct contact between Spn and Sau was required to efficiently eradicate Sau biofilms and biofilm-released planktonic cells. Physical contact-mediated killing of Sau was not related to production of hydrogen peroxide as an isogenic TIGR4ΔspxB mutant eradicated Sau bacteria within 4 h. Confocal micrographs confirmed eradication of Sau biofilms by TIGR4 and allowed us to visualize ultrastructural point of contacts between Sau and Spn. A time-course study further demonstrated spatial colocalization of Spn chains and Sau tetrads as early as 30 min post-inoculation (Pearson's coefficient >0.72). Finally, precolonized biofilms produced by Sau strain Newman, or MRSA strain USA300, were eradicated by mid-log phase cultures of washed TIGR4 bacteria within 2 h post-inoculation. In conclusion, Spn strains rapidly eradicate pre-colonized Sau aureus biofilms, including those formed by MRSA strains, by a mechanism(s) requiring bacterium-bacterium contact, but independent from the production of hydrogen peroxide.

Next generation protein based Streptococcus pneumoniae vaccines

All currently available Streptococcus pneumoniae (Spn) vaccines have limitations due to their capsular serotype composition. Both the 23-valent Spn polysaccharide vaccine (PPV) and 7, 10, or 13-valent Spn conjugate vaccines (PCV-7, 10, -13) are serotype-based vaccines and therefore they elicit only serotype-specific immunity. Emergence of replacement Spn strains expressing other serotypes has consistently occurred following introduction of capsular serotype based Spn vaccines. Furthermore, capsular polysaccharide vaccines are less effective in protection against non-bacteremic pneumonia and acute otitis media (AOM) than against invasive pneumococcal disease (IPD). These shortcomings of capsular polysaccharide-based Spn vaccines have created high interest in development of non-serotype specific protein-based vaccines that could be effective in preventing both IPD and non-IPD infections. This review discusses the progress to date on development of Spn protein vaccine candidates that are highly conserved by all Spn strains, are highly conserved, exhibit maximal antigenicity and minimal reactogenicity to replace or complement the current capsule-based vaccines. Key to development of a protein based Spn vaccine is an understanding of Spn pathogenesis. Based on pathogenesis, a protein-based Spn vaccine should include one or more ingredients that reduce NP colonization below a pathogenic inoculum. Elimination of all Spn colonization may not be achievable or even advisable. The level of expression of a target protein antigen during pathogenesis is another key to the success of protein based vaccines.. As with virtually all currently licensed vaccines, production of a serum antibody response in response to protein based vaccines is anticipated to provide protection from Spn infections. A significant advantage that protein vaccine formulations can offer over capsule based vaccination is their potential benefits associated with natural priming and boosting to all strains of Spn. One of the most universal and comprehensive approaches of identifying novel vaccine candidates is the investigation of human sera from different disease stages of natural infections. Antigens that are robustly reactive in preliminary human serum screening constitute a pathogen-specific antigenome. This strategy has identified a number of Spn protein vaccine candidates that are moving forward in human clinical trials.

New Patterns in the Otopathogens Causing Acute Otitis Media Six to Eight Years After Introduction of Pneumococcal Conjugate Vaccine

To describe NP and AOM otopathogens during the time frame 2007 to 2009, 6 to 8 years after the introduction of 7-valent pneumococcal conjugate (PCV7) in the United States and to compare nasopharyngeal (NP) colonization and acute otitis media (AOM) microbiology in children 6 to 36 months of age having first and second AOM episodes with children who are otitis prone. Prospectively, the microbiology of NP colonization and AOM episodes was determined in 120 children with absent or infrequent AOM episodes. NP samples were collected at 7 routine visits between 6 and 30 months of age and at the time of AOM. For first and subsequent AOM episodes, middle ear fluid (MEF) was obtained by tympanocentesis. Eighty otitis prone children were comparatively studied. All 200 children received age-appropriate doses of PCV7. We found PCV7 serotypes were virtually absent: (0.9% isolated from both NP and MEF) in both study groups. However, non-PCV7 serotypes replaced PCV serotypes such that the frequency of isolation of S. pneumoniae (Spn) was nearly equal to that of non-typeable Haemophilus influenzae (NTHi). M. catarrhalis (Mcat) was less common and Staphylococcus aureus infrequent in the NP and MEF from the 2 groups. The proportion of Spn, NTHi and Mcat causing AOM was similar in children with first and second AOM episodes compared to otitis prone children. However, oxacillin-resistant Spn isolated from the NP and MEF was 19% for the absent/infrequent and 58% for the otitis prone groups, P < 0.0001. Beta-lactamase producing NTHi occurred more frequently in the otitis prone group, P = 0.04. Six to 8 years after widespread use of PCV7, Spn strains expressing vaccine-type serotypes have virtually disappeared from the NP and MEF of vaccinated children. NP colonization and AOM has changed to non-PCV7 strains of Spn. NTHi continues to be a major AOM pathogen. The otopathogens in first and second AOM and in otitis prone children are very similar although Spn and NTHi are more often antibiotic resistant in the otitis prone.

Roles of relSpn in stringent response, global regulation and virulence of serotype 2 Streptococcus pneumoniae D39

RelA/SpoT homologue (RSH) proteins have (p)ppGpp synthetase and hydrolase activities that mediate major global responses to nutrient limitation and other stresses. RSH proteins are conserved in most bacteria and play diverse roles in bacterial pathogenesis. We report here that the RSH protein of Streptococcus pneumoniae, Rel(Spn), can be deleted and is the primary source of (p)ppGpp synthesis in virulent strain D39 under some conditions. A D39 Deltarel(Spn) mutant grew well in complex medium, but did not grow in chemically defined medium unless supplemented with the metals copper and manganese. Transcriptome analysis of D39 rel(+)(Spn) and Deltarel(Spn) strains treated with mupirocin revealed rel(Spn)-independent (translation stress), rel(Spn)-dependent (stringent response) and Deltarel(Spn)-dependent changes, suggesting that rel(Spn) and (p)ppGpp amount play wide-ranging homeostatic roles in pneumococcal physiology, besides adjusting macromolecular synthesis and transport in response to nutrient availability. Notably, the rel(Spn)-dependent response included significant upregulation of the ply operon encoding pneumolysin toxin, whereas the Deltarel(Spn)-dependent response affected expression linked to the VicRK and CiaRH two-component systems. Finally, a D39 Deltarel(Spn) mutant was severely attenuated and displayed a significantly altered course of disease progression in a mouse model of infection, which was restored to normal by an ectopic copy of rel(+)(Spn).

VIGILANCIA EPIDEMIOLÓGICA CENTINELA DE Haemophilus influenzae Y Streptococcus pneumoniae EN MENORES DE 5 AÑOS EN EL PERÚ

Objective: To determine the frequency, serotypes and antimicrobial resistance of H. influenzae (Hi) and S. pneumoniae (Spn) in Peruvian children under 5 years old with meningitis and pneumonia. Materials and methods: Between October 2000 and December 2001 sentinel surveillance por Hi and Spn was implemented in 5 hospitals in Peru. Serotype identification and antimicrobial resitance were performed at the Peruvian National Institute of Health in Lima. Antimicrobial resitance using minimal inhibitory concentration (MIC) was assessed using the plate microdilution method, according to NCCLS standards. Results: 1283 cases were included. 59 strains (31 of Hi and 28 of Spn) were found. Bacterias were detected in 2,3% of pneumonia patients and in 42,5% of meningitis patients. 10/1210 (0,8%) had Hi pneumonia and 18/1210 Spn pneumonia. 21/73 (28,8%) had Hi meningitis and 10/73 (13,7%) Spn meningitis. The following Spn serotypes: 1, 5, 6A, 11, 14, 19, 19F and 20 were identified. Only the serotype b of Hi was identified. We found Spn strains highly resistant to penicillin (3/13), cotrimoxazole (3/13), erithromycin (1/13), chloramphenicol (1/13) and ceftriaxone (1/13). Hi strains highly resistant to cotrimoxazole (4/20) and ampicillin (1/20) were also detected. Conclusions: The frequency of Spn and Hi infection in children under 5 years old with pneumonia and meningitis was very low. We found serotypes already identified in Latin America and some antibiotic used in the standardized treatment of pneumonia and meningitis highly-resistant strains of Spn and Hi. Follow-up of this surveillance system in selected hospitals is recommended. Keys word: Haemophilus influenzae; Streptococcus pneumoniae; Surveillance; Epidemiologic surveillance; Child; Peru (source: BIREME).