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Background: Bacterial etiologies of acute otitis media (AOM) have shifted from the introduction of pneumococcal conjugate vaccines (PCVs), antibiotic selection and competition among species. We characterized Streptococcus pneumoniae ( Spn ), Haemophilus influenzae ( Hflu ) and Moraxella catarrhalis ( Mcat ) in the nasopharynx during well-child healthy visits and at the onset of AOM, and in middle ear fluid (MEF) of children with AOM to assess anticipated effects of higher-valency PCVs (PCV15 and PCV20)., Methods: From September 2021 to September 2023, we conducted a prospective longitudinal cohort study of PCV13 immunized children 6-36 months old. MEF was collected via tympanocentesis. Serotyping and antibiotic susceptibility testing were performed on Spn , Hflu and Mcat isolates., Results: We obtained 825 nasopharyngeal and 216 MEF samples from 301 children. The order of frequency of nasopharyngeal colonization was Mcat , Spn and Hflu ; Hflu was the predominant otopathogen in MEF. Among Spn isolates, non-PCV15, non-PCV20 serotypes predominated in the nasopharynx and in MEF; the most frequent serotype was 35B. Among MEF samples, 30% of Spn isolates were amoxicillin nonsusceptible; 23% of Hflu isolates and 100% of Mcat isolates were β-lactamase-producing., Conclusion: The majority of Spn isolates among young children were non-PCV15, non-PCV20 serotypes, especially serotype 35B; therefore, the impact of higher-valency PCVs in reducing pneumococcal colonization or AOM is expected to be limited. Hflu continues to be the most frequent AOM pathogen. Antibiotic susceptibility data suggest a high dose of amoxicillin/clavulanate or alternative drugs that are effective against contemporary mix of otopathogens could be considered for optimal empiric selection to provide the best efficacy., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Haemophilus influenzae serotype a (Hia) has recently emerged as an important cause of invasive disease in the North American Arctic and Sub-Arctic regions, mainly affecting young Indigenous children. In this study, we addressed the question of whether the prevalence of Hia and all H. influenzae in the nasopharynx differed between paediatric populations from regions with high versus low incidence of invasive Hia disease. Nasopharyngeal specimens from children with acute respiratory tract infections (ARTI) collected for routine diagnostic detection of respiratory viruses were analysed with molecular-genetic methods to identify and serotype H. influenzae . In Nunavut, a region with a high incidence of invasive Hia disease, all H. influenzae and particularly Hia were found in the nasopharynx of 60.6% and 3.0% children. In Southern Ontario (Hamilton region), where Hia invasive disease is rare, the frequencies of all H. influenzae and Hia detection were 38.5% and 0.6%, respectively. In both cohorts, non-typeable H. influenzae was prevalent (57.0% and 37.9%, respectively). Considering that Hia is an important cause of severe invasive disease in Nunavut children, 3% prevalence of Hia among children with ARTI can reflect continuing circulation of the pathogen in the Northern communities that may result in invasive disease outbreaks.

The Southampton pneumococcal carriage study of children under 5 years old continued during the coronavirus disease 2019 (COVID-19) pandemic. Here, we present data from October 2018 to March 2023 describing prevalence of pneumococci and other pathobionts during the winter seasons before, during, and after the introduction of non-pharmaceutical interventions (NPIs) to prevent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission. Nasopharyngeal swabs were collected from children attending outpatient clinics at a secondary care hospital and community healthcare sites. Pre-NPIs, in 2019/2020, the carriage prevalence of pneumococci at the hospital site was 32% ( n = 161 positive/499 participants). During NPIs, this fell to 19% ( n = 12/64), although based on fewer participants compared to previous years due to COVID-19 restrictions on health-care attendance. In 2021/2022, after NPIs had eased, prevalence rebounded to 33% ( n = 15/46) [compared to NPIs period, χ 2 (1, N = 110) =2.78, P = 0.09]. Carriage prevalence at community healthcare sites fell significantly from 27% ( n = 127/470) in 2019/2020 to 19% during the NPI period ( n = 44/228) in 2020/2021 [ χ 2 (1, N = 698) =4.95, P = 0.026]. No rebound was observed in 2021/2022 [19% ( n = 56/288)]. However, in a multivariate logistic regression model, neither site had a significantly lower carriage prevalence during the NPI period compared to the post NPI period. A reduction in serotype diversity was observed in 2020/2021. Carriage of Haemophilus influenzae was particularly affected by NPIs with a significant reduction observed. In conclusion, among children under 5 years of age, transient, modest, and statistically non-significant alterations in carriage of both Streptococcus pneumoniae and H. influenzae were associated with SARS-CoV-2 NPIs.IMPORTANCE Streptococcus pneumoniae (the pneumococcus) continues to be a major contributor to global morbidity and mortality. Using our long-running pediatric study, we examined changes in pneumococcal carriage prevalence in nearly 3,000 children under the age of 5 years between the winters of 2018/2019 and 2022/2023. This period coincided with the severe acute respiratory syndrome coronavirus 2 pandemic and, in particular, the implementation of national strategies to limit disease transmission in the UK. We observed a transient reduction of both Streptococcus pneumoniae and Haemophilus influenzae in these populations during this period of non-pharmaceutical interventions. This aligned with the reduction in invasive pneumococcal disease seen in the UK and is therefore a likely contributor to this phenomenon., Competing Interests: D.W.C. was a post-doctoral researcher on GSK-funded projects in 2014/2915 and currently receives grant support from Pfizer. S.N.F. receives support from the National Institute for Health Research funding via the NIHR Southampton Clinical Research Facility and the NIHR Southampton Biomedical Research Centre. S.N.F. and S.C.C. act as principal investigators for clinical trials and other studies conducted on behalf of University Hospital Southampton NHS Foundation Trust/University of Southampton that are sponsored by vaccine manufacturers. No personal payments are received from them. S.N.F. and S.C.C. have participated in advisory boards for vaccine manufacturers but receive no personal payments for this work. S.N.F. and S.C.C. have received financial assistance from vaccine manufacturers to attend conferences. All grants and honoraria are paid into accounts within the respective NHS Trusts or Universities or to independent charities. J.C., M.L., K.H., J.S., and B.D.G. are employees of Pfizer and may hold stock and/or stock options.

The nasopharynx is an important reservoir of disease-associated and antimicrobial-resistant bacterial species. This proof-of-concept study assessed the utility of a combined culture, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS), and targeted metagenomic sequencing workflow for the study of the pediatric nasopharyngeal bacterial microbiota. Nasopharyngeal swabs and clinical metadata were collected from Cambodian children during a hospital outpatient visit and then biweekly for 12 weeks. Swabs were cultured on chocolate and blood-gentamicin agar, and all colony morphotypes were identified by MALDI-TOF MS. Metagenomic sequencing was done on a scrape of all colonies from a chocolate agar culture and processed using the mSWEEP pipeline. One hundred one children were enrolled, yielding 620 swabs. MALDI-TOF MS identified 106 bacterial species/40 genera: 20 species accounted for 88.5% (2,190/2,474) of isolates. Colonization by Moraxella catarrhalis (92.1% of children on ≥1 swab), Haemophilus influenzae (87.1%), and Streptococcus pneumoniae (83.2%) was particularly common. In S. pneumoniae -colonized children, a median of two serotypes [inter-quartile range (IQR) 1-2, range 1-4] was detected. For the 21 bacterial species included in the mSWEEP database and identifiable by MALDI-TOF, detection by culture + MALDI-TOF MS and culture + mSWEEP was highly concordant with a median species-level agreement of 96.9% (IQR 86.8%-98.8%). mSWEEP revealed highly dynamic lineage-level colonization patterns for S. pneumoniae which were quite different to those for S. aureus . A combined culture, MALDI-TOF MS, targeted metagenomic sequencing approach for the exploration of the young child nasopharyngeal microbiome was technically feasible, and each component yielded complementary data., Importance: The human upper respiratory tract is an important source of disease-causing and antibiotic-resistant bacteria. However, understanding the interactions and stability of these bacterial populations is technically challenging. We used a combination of approaches to determine colonization patterns over a 3-month period in 101 Cambodian children. The combined approach was feasible to implement, and each component gave complementary data to enable a better understanding of the complex patterns of bacterial colonization., Competing Interests: The authors declare no conflict of interest.

In 2013, Uganda introduced the PCV10 pneumococcal vaccine and it is given to children at 6, 10 and 14 weeks after birth. Carriage prevalence studies post PCV10-introduction are necessary for monitoring the impact of vaccination and trends in antibiotic resistance. Here, we studied carriage/antibiotic resistance of Streptococcus pneumoniae (pneumococcus), Haemophilus influenzae, Moraxella catarrhalis , and Staphylococcus aureus isolated from 194 children at the Mulago Assessment Centre clinic in Kampala-Uganda, 5 years post-PCV10 introduction. Almost all the children were vaccinated with PCV10 (98.5%, 191/194). The overall carriage prevalence (any species) was 62% (120/194), and it was associated with a history of antibiotics use (p=0.0159) and having respiratory symptoms (p=0.0003). The pneumococcus, H. influenzae, M. catarrhalis , and S. aureus carriage prevalence was 46% (90/194), 21% (40/194), 7% (14/194), and 6% (12/194), respectively. Species co-carriage occurred in 32 children (17%, 32/194), predominantly multidrug resistant pneumococcus + H. influenzae (23 children). Furthermore, pneumococci were highly resistant to cotrimoxazole (100%), erythromycin (76%), and tetracycline (52%), 42% being multidrug-resistant. Overall, we note an increase in antibiotic resistance post-PCV10 introduction, and microbial shifts i.e., a decrease in pneumococcus, M. catarrhalis and S. aureus carriage and an increase in H. influenzae carriage suggesting vaccine-associated perturbation of the respiratory ecology., (© 2023 Nshimiyimana T et al.)

We compared the distribution of organisms in the nasopharynx and the middle ear fluid in 164 episodes of acute otitis media in children 6-35 months of age. In contrast to Streptococcus pneumoniae and Haemophilus influenzae , Moraxella catarrhalis is isolated from the middle ear in only 11% of episodes with nasopharyngeal colonization with this organism., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Importance: The large overlap between symptoms of acute sinusitis and viral upper respiratory tract infection suggests that certain subgroups of children being diagnosed with acute sinusitis, and subsequently treated with antibiotics, derive little benefit from antibiotic use., Objective: To assess if antibiotic therapy could be appropriately withheld in prespecified subgroups., Design, Setting, and Participants: Randomized clinical trial including 515 children aged 2 to 11 years diagnosed with acute sinusitis based on clinical criteria. The trial was conducted between February 2016 and April 2022 at primary care offices affiliated with 6 US institutions and was designed to evaluate whether symptom burden differed in subgroups defined by nasopharyngeal Streptococcus pneumoniae, Haemophilus influenzae, or Moraxella catarrhalis on bacterial culture and by the presence of colored nasal discharge., Interventions: Oral amoxicillin (90 mg/kg/d) and clavulanate (6.4 mg/kg/d) (n = 254) or placebo (n = 256) for 10 days., Main Outcomes and Measures: The primary outcome was symptom burden based on daily symptom scores on a validated scale (range, 0-40) during the 10 days after diagnosis. Secondary outcomes included treatment failure, adverse events including clinically significant diarrhea, and resource use by families., Results: Most of the 510 included children were aged 2 to 5 years (64%), male (54%), White (52%), and not Hispanic (89%). The mean symptom scores were significantly lower in children in the amoxicillin and clavulanate group (9.04 [95% CI, 8.71 to 9.37]) compared with those in the placebo group (10.60 [95% CI, 10.27 to 10.93]) (between-group difference, -1.69 [95% CI, -2.07 to -1.31]). The length of time to symptom resolution was significantly lower for children in the antibiotic group (7.0 days) than in the placebo group (9.0 days) (P = .003). Children without nasopharyngeal pathogens detected did not benefit from antibiotic treatment as much as those with pathogens detected; the between-group difference in mean symptom scores was -0.88 (95% CI, -1.63 to -0.12) in those without pathogens detected compared with -1.95 (95% CI, -2.40 to -1.51) in those with pathogens detected. Efficacy did not differ significantly according to whether colored nasal discharge was present (the between-group difference was -1.62 [95% CI, -2.09 to -1.16] for colored nasal discharge vs -1.70 [95% CI, -2.38 to -1.03] for clear nasal discharge; P = .52 for the interaction between treatment group and the presence of colored nasal discharge)., Conclusions: In children with acute sinusitis, antibiotic treatment had minimal benefit for those without nasopharyngeal bacterial pathogens on presentation, and its effects did not depend on the color of nasal discharge. Testing for specific bacteria on presentation may represent a strategy to reduce antibiotic use in this condition., Trial Registration: ClinicalTrials.gov Identifier: NCT02554383.

Background: An association has been reported between early life Staphylococcus aureus nasal carriage and higher risk of childhood eczema, but it is unclear whether this relationship is causal and associations with other bacterial species are unclear., Objective: To examine the associations of early life nasal and nasopharyngeal bacterial carriage with eczema phenotypes, and the direction of any associations identified., Methods: Among 996 subjects of a population-based prospective cohort study, nasal swabs for Staphylococcus  aureus, and nasopharyngeal swabs for Streptococcus pneumoniae, Moraxella catarrhalis and Haemophilus influenzae were collected and cultured from age 6 weeks to 6 years. Never, early, mid-, late transient and persistent eczema phenotypes were identified from parental-reported physician-diagnosed eczema from age 6 months until 10 years. Multinomial regression models and cross-lagged models were applied., Results: Staphylococcus aureus nasal carriage at 6 months was associated with an increased risk of early transient and persistent eczema (OR (95% CI): 2.69 (1.34, 5.39) and 4.17 (1.12, 15.51)). The associations between Staphylococcus aureus nasal carriage and eczema were mostly cross-sectional, and not longitudinal. No associations of Staphylococcus pneumoniae, Moraxella catarrhalis and Haemophilus influenza nasopharyngeal bacterial carriage with eczema and eczema phenotypes were observed (OR range (95% CI): 0.71 (0.35, 1.44) to 1.77 (0.84, 3.73))., Conclusions: Early life Staphylococcus aureus nasal carriage, but not Staphylococcus pneumoniae, Moraxella catarrhalis and Haemophilus influenza nasopharyngeal carriage, was associated with early transient and persistent eczema. Staphylococcus aureus nasal carriage and eczema were mostly cross-sectionally associated, and not longitudinally, making a causal relationship in either direction unlikely., (© 2021 The Authors. Clinical & Experimental Allergy published by John Wiley & Sons Ltd.)

Objectives: Nontypeable Haemophilus influenzae (NTHi) is a chief pathogen in both acute otitis media and otitis media with effusion. Phosphorylcholine (ChoP) is expressed on lipooligosaccharides, and ChoP has phase variation, which is related to its adhesion to and invasion of epithelial cells in the upper airway. However, little is known about the role of ChoP expression. We examined the kinetics of the mucosal clearance of NTHi from the nose and middle ear and the mucosal immune response to NTHi infection by comparing ChoP(+) and ChoP(-) strains in a mouse model of middle ear and nasal challenge., Methods: Six-week-old male BALB/c mice were subjected to bacterial challenge in the middle ear and nasopharynx. Mice were inoculated with a suspension of a ChoP(+) strain or ChoP(-) strain of NTHi. On days 1, 3, and 7 after inoculation, the middle ear wash (MEW) and nasal wash (NW) were harvested from each group. The samples were used for bacterial counts and the supernatant was used to measure the level of cytokines and C-reactive protein (CRP)., Results: MEWs in the ChoP(+) strain group had significantly higher bacterial counts than those in the ChoP(-) strain group on day 1. However, bacteria were eradicated in the ChoP(+) strain group on day 7. NWs in the ChoP(+) strain group had higher bacterial counts than those in the ChoP(-) strain group during the experiment, however, there was no significant difference between the two strains. The levels of cytokines were significantly higher in the ChoP(-) strain group than in the ChoP(+) strain group in MEWs, but these cytokine levels were low in NWs. The CRP concentration in the ChoP(-) group was high on day 7 in the MEWs. In NWs, the CRP concentration was low in all groups during the experiment., Conclusion: ChoP expression of NTHi changes the organism susceptible to killing by CRP, and the ChoP(+) strain might be gradually eradicated from the middle ear via the CRP-complement cascade, but not from nasopharynx. Based on our findings, phase variation by altering Phosphorylcholine expression of nontypeable Haemophilus influenzae affects bacteria clearance and mucosal immune response in the middle ear and nasopharynx., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest associated with this manuscript., (Copyright © 2020. Published by Elsevier B.V.)

Background : We evaluated bacterial nasopharyngeal carriage (NPC) prevalence and cumulative acquisition following 7-valent pneumococcal conjugate vaccine (PCV7) or pneumococcal non-typeable Haemophilus influenzae protein D conjugate vaccine (PHiD-CV) administration. Methods : Participants were children from two clinical trials in a South African center who received PCV7 (n = 250) or PHiD-CV (n = 100) at ~6 weeks, ~14 weeks, and ~9-10 months of age, and were enrolled between Dec2009-Apr2010 and Mar2009-May2010 in the PCV7 and PHiD-CV studies, respectively. Sample collection, most microbiological assessments, and data re-analysis methods were identical. Results : NPC prevalence of any pneumococcal serotype was 18.5% and 17.0% at pre-vaccination, and 63.1% and 67.3% in 24-27 month-old children among PCV7 and PHiD-CV recipients, respectively. In 24-27 month-old children, 96.1% and 99.0% of PCV7 and PHiD-CV recipients had acquired ≥1 pneumococcal serotype, 53.7% and 62.9% ≥1 PCV7 serotype, 1.5%, and 3.1% ≥1 of serotypes 1, 5 or 7F, 23.2% and 19.6% serotype 6A, 23.2% and 21.7% serotype 19A, 88.7%, and 91.0% H. influenzae , and 50.3% and 62.9% Staphylococcus aureus , respectively. Conclusions : This analysis of two concurrent clinical trials did not reveal differences in bacterial NPC prevalence or acquisition in PCV7- and PHiD-CV-vaccinated children. Trial registration : South African National Clinical Trial Register (NHREC DOH-27-0511-299); ClinicalTrials.gov (NCT00829010).

Nasopharyngeal colonization by bacteria is a prerequisite for progression to respiratory disease and an important source of horizontal spread within communities. We aimed to perform quantitative analysis of the bacterial cells and reveal the microbiota of the nasal discharge in children at the species level based on highly accurate 16S rRNA gene sequencing. This study enrolled 40 pediatric patients with rhinorrhea. The bacterial cells in the nasal discharge were counted by epifluorescence microscopic analysis. The microbiota was analyzed by using the 16S rRNA gene clone library sequencing method. We demonstrated that a high abundance (median 2.2 × 10 7 cells/mL) of bacteria was contained in the nasal discharge of children. Of the 40 samples, 37 (92.5%) were dominated by OTUs corresponding to Haemophilus aegyptius/influenzae, Moraxella catarrhalis/nonliquefaciens, or Streptococcus pneumoniae. These samples showed higher cell abundance and lower alpha diversity than the remaining three samples in which the other bacteria coexisted. In addition, 12 sequences with low homology to type strains were considered as previously unknown bacterial lineages. In conclusion, the nasal discharge of most young children contains a large amount of respiratory pathogens and several unknown bacteria, which could not only cause endogenous infection but also be a source of transmission to others.

Background: Otitis media, a disease highly prevalent among children worldwide, manifests clinically in both acute and chronic forms. The manner and time at which chronicity develops among Indian children is unknown., Aim: To study the prevalence, manifestations and risk factors for otitis media in a birth cohort aged 8 years., Methods: A birth cohort of 107 babies was followed up at 8 years of age and ENT evaluation with nasopharyngeal swabbing for detecting Streptococcus pneumoniae and Hemophilus influenzae was performed., Results: The overall prevalence of otitis media was 14%, almost half the prevalence in the first 2 years of life. Eight children (7.5%) with congested, bulging eardrums and no systemic symptoms had asymptomatic acute otitis media. Another five (4.7%) children had otitis media with effusion and 2 (1.9%) had chronic suppurative otitis media. Although 10/15 (66.7%) children with otitis media had positive swabs at 8 years age, only 2 were pneumococcal vaccine (PCV-13) serotypes. Risk factor analysis showed that passive smoking was the only significant parameter associated with otitis media (p = 0.029). Nasopharyngeal swabbing showed that 51/105 (48.6%) children had positive swabs for S. pneumoniae and 5/105 (4.8%) for S.pneumoniae with non-type b H. influenzae. The ten most commonly encountered pneumococcal serotypes were 6A,4,8,16F,33B,35A,35B, 18C, 19F and 23B which together comprised 29 of the 56 (51.8%) isolates. PCV-13 serotypes formed 19/56 (33.9%) to 21/56 (37.5%) of all pneumococcal isolates. Of 6 children who had received PCV-13, 4 tested positive for S. pneumoniae at 8 years of age too. However, none were vaccine serotypes. Four of those with otitis media who had positive swabs had received no immunisation at all and 3 of them had vaccine serotypes, viz. 4, 6A and 18C respectively., Conclusion: Indian children continue to have a high prevalence of otitis media at 8 years age. More than 1/3 of nasopharyngeal isolates at this age are vaccine serotypes. Passive smoking is an important risk factor for childhood otitis media and may contribute to the development of chronicity of the disease., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Airway bacterial carriage might play a role in respiratory disease. We hypothesize that nasal carriage with Staphylococcus aureus or nasopharyngeal carriage with Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae predisposes individuals to adverse respiratory health., Objective: To examine the association of early-life airway bacterial carriage with respiratory tract infections and vice versa, and of early-life airway bacterial carriage with wheezing, lung function, and asthma in later childhood., Methods: We collected upper airway swabs for bacterial culturing for S aureus, H influenzae, M catarrhalis, and H influenzae at six timepoints between the ages of 6 weeks and 6 years among 945 children participating in a population-based prospective cohort study. Information on respiratory tract infections and wheezing until age 6 years, and asthma at age 10 years was obtained by questionnaires. Lung function at age 10 years was measured by spirometry. We tested possible bidirectional associations between airway bacterial carriage and respiratory tract infections by cross-lagged models, and associations of repeatedly measured airway bacterial carriage with wheezing, lung function, and asthma by generalized estimating equations models and regression models., Results: Cross-lagged modeling showed that early-life airway bacterial carriage was not consistently associated with upper and lower respiratory tract infections or vice versa. Nasopharyngeal carriage with any bacteria in infancy was associated with an increased risk of wheezing (OR [95% CI]: 1.66 [1.31, 2.10]). Airway bacterial carriage was not consistently associated with school-age lung function or asthma., Conclusion: Nasopharyngeal carriage with any bacteria is associated with wheezing, but not respiratory tract infections, asthma, or lung function., (© 2020 The Authors. Pediatric Allergy and Immunology published by John Wiley & Sons Ltd.)

Background: The nasopharynx can from time to time accommodate otherwise pathogenic bacteria. This phenomenon is called asymptomatic carriage. However, in case of decreased immunity, viral infection or any other enhancing factors, severe disease can develop. Our aim in this study was to survey the nasal carriage rates of four important respiratory pathogens in three different age groups of children attending nurseries, day-care centres and primary schools. This is the first study from Hungary about the asymptomatic carriage of H. influenzae and M. catarrhalis., Methods: Altogether 580 asymptomatic children were screened in three Hungarian cities. Samples were collected from both nostrils with cotton swabs. The identification was based on both colony morphology and species-specific PCRs. Serotyping was performed for S. pneumoniae, H. influenzae and M. catarrhalis. Antibiotic susceptibility was determined with agar dilution, according to the EUCAST guidelines. Clonality was examined by PFGE., Results and Conclusions: Whereas the carriage rates of S. pneumoniae, H. influenzae and M. catarrhalis clearly decreased with age, that of S. aureus showed an opposite tendency. Multiple carriage was least prevalent if S. aureus was one of the participants. The negative association between this bacterium and the others was statistically significant. For pneumococcus, the overall carriage rate was lower compared to earlier years, and PCV13 serotypes were present in only 6.2% of the children. The majority of H. influenzae isolates was non-typeable and no type b was detected; serotype A was dominant among M. catarrhalis. All four bacteria were more sensitive to antibiotics compared to clinical isolates. No MRSAs were detected, but we found three mupirocin resistant strains. The positive effect of Hib- and PCV-vaccination is undoubted. Continued surveillance of these pathogens is required., Competing Interests: The authors have declared that no competing interests exist.

BackgroundThe current carriage study was set up to reinforce surveillance during/after the PCV13-to-PCVC10 switch in Belgium.AimThis observational study monitored carriage of Streptococcus pneumoniae (Sp) serotypes, particularly those no longer covered (3, 6A, 19A), as well as Haemophilus influenzae (Hi), because PCV10 contains the non-typeable Hi protein D.MethodsA total of 2,615 nasopharyngeal swabs from children (6-30 months old) attending day care were collected in three periods over 2016-2018. Children's demographic and clinical characteristics and vaccination status were obtained through a questionnaire. Sp and Hi were identified by culture and PCR. Pneumococcal strains were tested for antimicrobial (non-)susceptibility by disc diffusion and serotyped by Quellung-reaction (Quellung-reaction and PCR for serotypes 3, 6A, 19A).ResultsThe carriage prevalence of Sp (> 75%) remained stable over the successive periods but that of Hi increased (87.4%, 664 Hi-carriers/760 in 2016 vs 93.9%, 895/953 in 2017-2018). The proportion of non-PCV13 vaccine serotypes decreased (94.6%, 438 isolates/463 in 2016 vs 89.7%, 599/668 in 2017-2018) while that of PCV13-non-PCV10 vaccine serotypes (3 + 6A + 19A) increased (0.9%, 4 isolates/463 in 2016 vs 7.8%, 52/668 in 2017-2018), with serotype 19A most frequently identified (87.9%, 58/66 isolates). Non-susceptibility of pneumococci against any of the tested antibiotics was stable over the study period (> 44%).ConclusionsDuring and after the PCV13-to-PCV10 vaccine switch, the proportion of non-PCV13 serotypes decreased, mainly due to a serotype 19A carriage prevalence increase. These results complement invasive pneumococcal disease surveillance data, providing further basis for pneumococcal vaccination programme policy making.

Introduction . Streptococcus pneumoniae and Haemophilus influenzae are both commensals of the human nasopharynx with a high capacity to cause upper and lower respiratory tract infections. Aim. Molecular testing of nasopharyngeal samples from children at the primary care paediatric department presenting with acute respiratory tract infections (ARTIs). Methodology. From June 2016 to May 2017, 156 nasopharyngeal swabs from children diagnosed with ARTIs who had been admitted to or followed up as outpatients at the Department of Paediatrics, SANADOR Hospital (Bucharest, Romania) were tested for the presence of S. pneumoniae , H. influenzae , Mycoplasma pneumoniae , Chlamydophila pneumoniae , Legionella pneumophila , Bordetella pertussis and Bordetella parapertussis DNA. Results. S. pneumoniae had the highest detection rate (53.8 %, n =84/156), followed by H. influenzae (41 %, n =64/156) and S. pneumoniae/H. influenzae co-detection (26.2 %, n =41/156). Conclusion. A definitive laboratory diagnosis of these micro-organisms can be made for invasive disease, but there are difficulties in establishing the aetiology for mucosal infection. Molecular detection tests could complement culture-based tests by strengthening their surveillance.

This study compared flocked (nylon) swabs and (non-flocked) rayon swabs for the detection of Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis in nasopharyngeal samples from 20 enrolled Indigenous children under the age of 6 years living in remote Australian Aboriginal communities, and determined which swab the child or parent perceived to be more comfortable. There was no evidence of a significant difference between flocked and rayon swabs in the recovery of common respiratory bacteria. Rayon swabs detected presence of S. pneumoniae (90% cf. 74%, p = 0.375), H. influenzae (79% cf. 74%, p = 1.00) and M. catarrhalis (79% cf. 74%, p = 1.00) at higher rates than the flocked swabs. Analysis of semi-quantitative growth scores also showed no significant differences in either the ranked distributions or medians. Rayon swabs median semi-quantitative growth scores were higher for S. pneumoniae (4 [IQR 1-5] cf. 3 [IQR 0-6], p = 0.699), and H. influenzae (2 [IQR1-5] cf. 1 [IQR0-5], p = 0.946). Sixty percent of participants preferred samples to be taken with flocked swabs. This study demonstrates that microbiological outcomes are not compromised when using flocked or rayon swabs in respiratory bacterial carriage studies in this population. Therefore, cost, methodological consistency across studies, and participant preference can be considered when choosing swab type., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Background: Biofilm production by Haemophilus influenzae and Streptococcus pneumoniae has been implicated in the pathogenesis of otitis media, mainly in chronic and recurrent cases. We studied the "in vitro" biofilm production by these 2 species isolated alone or together from the nasopharynx of children with acute otitis media., Methods: The studied strains were from 3 pneumococcal conjugate vaccine (PCV) periods: pre-PCV7, post-PCV7/pre-PCV13 and post-PCV13. A modified microtiter plate assay with crystal violet stain was used to study the biofilm production of 182 H. influenzae and 191 S. pneumoniae strains., Results: Overall, 117/181 (64.6%) H. influenzae and 128/191 (66.8%) S. pneumoniae strains produced biofilm. The proportion of biofilm-producing H. influenzae strains was greater with than without the isolation of S. pneumoniae in the same sample (75.5% vs 52.3%, p = 0.001). Conversely, the proportion of biofilm-producing S. pneumoniae strains was not affected by the presence or not of H. influenzae (66.3% vs 67.4%). S. pneumoniae serotypes 6B, 15B/C, 19A, 35F and 35B were the better biofilm producers (80%). Serotypes 11A, 14, 15A, 19F and 19A were more associated with H. influenzae biofilm-producing strains. Overall, 89/94 (94.6%) of cases with combined isolation showed biofilm production by S. pneumoniae or H. influenzae., Conclusion: This study emphasizes the high proportion of biofilm production by H. influenzae and S. pneumoniae strains isolated from the nasopharynx of children with acute otitis media, which reinforces the results of studies suggesting the importance of biofilm in the pathogenesis of acute otitis media.

Background: Otitis media (OM) imposes a great burden of disease in indigenous populations around the world, despite a variety of treatment and prevention programs. Improved understanding of the pathogenesis of OM in indigenous populations is required to advance treatment and reduce prevalence. We conducted a systematic review of the literature exploring the upper airway and middle ear microbiota in relation to OM in indigenous children., Methods: Papers targeting microbiota in relation to OM in children < 18 years indigenous to Australia, New Zealand, North America, and Greenland were sought. MEDLINE, CINAHL, EMBASE, Cochrane Library, and Informit databases were searched using key words. Two independent reviewers screened titles, abstracts, and then full-text papers against inclusion criteria according to PRISMA guidelines., Results: Twenty-five papers considering indigenous Australian, Alaskan, and Greenlandic children were included. There were high rates of nasopharyngeal colonization with the three main otopathogens (Haemophilus influenzae, Streptococcus pneumoniae, and Moraxella catarrhalis) in indigenous children with OM. Middle ear samples had lower rates of otopathogen detection, although detection rates increased when molecular methods were used. Pseudomonas aeruginosa and Staphylococcus aureus were commonly detected in middle ear discharge of children with chronic suppurative OM. There was a significant heterogeneity between studies, particularly in microbiological methods, which were largely limited to culture-based detection of the main otopathogens., Conclusions: There are high rates of otopathogen colonization in indigenous children with OM. Chronic suppurative OM appears to be associated with a different microbial profile. Beyond the main otopathogens, the data are limited. Further research is required to explore the entire upper respiratory tract/middle ear microbiota in relation to OM, with the inclusion of healthy indigenous peers as controls.

Paired nasopharynx and midturbinate swabs were obtained from 96 children (mean age: 2.45 years) with upper respiratory tract symptoms. Of these, 38.5% were colonized with Streptococcus pneumoniae only, 16.7% with Haemophilus influenzae only and 9.4% with both bacteria. Bacterial cultures from paired nasopharynx and midturbinate swabs demonstrated agreement for S. pneumoniae (κ = 0.75), H. influenzae (κ = 0.59) and for both pathogens (κ = 0.62). This report compares the results of bacterial cultures obtained from the nasopharynx and midturbinate specimens in children with upper respiratory tract infections.

The nasopharynx is the primary site of colonization by respiratory pathogen that constitutes the port of entrance in the respiratory tract. The role of mucosal respiratory microbiota in infection has been recently emphasized; therefore, we aimed to assess if a specific respiratory microbiota profile was associated with symptomatic infection and/or with presence of respiratory viruses. We performed a case-control study to characterize the healthy respiratory microbiota and its alteration during acute viral infections. Next-generation sequencing of the 16S rRNA gene was applied to 225 nasopharyngeal samples from 177 patients with viral respiratory infection and 48 matched healthy controls. We evidenced an important decrease of bacterial alpha-diversity in patients with symptomatic respiratory infection and a loss of the healthy core microbiota, specifically anaerobes and Prevotella spp. Moreover, eight respiratory pathogens were enriched in these patients, including Staphylococcus aureus, Haemophilus influenzae, Streptococcus pneumoniae, Moraxella catarrhalis, Dol osigranulum pigrum and Corynebacterium propinquum/pseudodiphtheriticum, whose role in respiratory infection is unclear. The asymptomatic carrier of influenza harbors a microbiota similar to healthy subjects, suggesting a critical role of microbiota in the clinical expression of viruses. These data suggest that the commensal microbiota plays a significant role in susceptibility to viral infection. The frequent co-detection of virus and bacteria raises the question of a strategy to prevent bacterial disease, focusing on the prevention of nasopharyngeal colonization through effective antibiotic treatment. In addition to antibiotics, further studies should test preventive or therapeutic interventions for maintaining or restoring a healthy nasopharyngeal microbiota.

Background: We studied Immunoglobulin G (IgG) antibody responses against Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis in young children with acute viral type respiratory infection and analyzed the findings in a multivariate model including age, nasopharyngeal carriage of the tested bacteria and pneumococcal vaccination., Methods: We included 227 children aged 6-23 months with acute respiratory infection. Nasopharyngeal aspirates were tested for bacterial carriage through detection of messenger RNA (mRNA) transcript with nCounter analysis. Acute and convalescent serum samples were tested for IgG antibody response against eight pneumococcal proteins, three proteins from H. influenzae and five proteins from M. catarrhalis in a fluorescent multiplex immunoassay., Results: A two-fold or greater increase in antibodies to S. pneumoniae, H. influenzae and M. catarrhalis was detected in 27.8, 9.7 and 14.1%, respectively. Nasopharyngeal carriage of each of the studied bacteria was not associated with antibody response detection against each respective bacterium. Furthermore, neither age nor pneumococcal vaccination were independently associated to detection of antibody response against the studied bacteria. Children who carried H. influenzae had higher frequency of colonization by M. catarrhalis (175 [80.3%] vs. 2 [22.2%]; p < .001) than those without H. influenzae. Also, children with acute otitis media tended to have higher frequency of antibody response to S. pneumoniae., Conclusion: Nasopharyngeal colonization by S. pneumoniae, H. influenzae and M. catarrhalis did not induce significant increases in antibody levels to these bacteria. Carriage of pathogenic bacteria in the nasopharynx is not able to elicit antibody responses to protein antigens similar to those caused by symptomatic infections.

Background: Nasopharyngeal bacterial colonization is a pre-requisite for developing bacterial mucosal and invasive disease. Pneumococcal conjugate vaccine (PCV) immunization of children reduces their risk of colonization by vaccine-serotypes, which could affect the biome of the nasopharynx in relation to colonization by other bacteria. This study evaluated the association of PCV immunization on the prevalence density of nasopharyngeal colonization by common, potentially pathogenic bacteria., Methods: A multiplex qPCR assay was used to evaluate bacterial nasopharyngeal colonization by 7-valent PCV (PCV7) serotypes, non-vaccine serotypes (NVT), Haemophilus influenzae, Staphylococcus aureus, Moraxella catarrhalis, and Neisseria meningitidis in PCV7-vaccinated and PCV-unvaccinated African children at two time points., Results: PCV7 vaccination was associated with a higher prevalence of NVT and H. influenzae at 9 and 16 months, respectively. While the prevalence of S. aureus was higher in PCV7-vaccinated children at 9 months, no difference was found at 16 months. The density of PCV7 serotypes (3.8 vs. 3.4 log 10 ; p = 0.048), NVT (3.6 vs. 3.1 log 10 ; p = 0.018), H. influenzae (4.34 vs. 3.86 log 10 ; p = 0.008), M. catarrhalis (3.52 vs. 2.98 log 10 ; p < 0.001) and S. aureus (4.02 vs. 3.06 log 10 ; p = 0.02) was higher among PCV-vaccinated compared to PCV-unvaccinated children at 9 months, although, this difference diminished at 16 months of age., Conclusion: The reduction in PCV7-serotype colonization impacted on colonization prevalence and density of other bacterial species of the nasopharynx. The clinical relevance of this needs further exploration in relation to mucosal and invasive disease outcomes, as well as for higher valency PCV vaccines., (Copyright © 2018. Published by Elsevier Ltd.)

Background: Virus is detected in about 80% of upper respiratory tract infections (URTIs) in children and is also detectable in the nasopharynx of 30% of asymptomatic children. The effect of asymptomatic viral infection on the dynamics of bacterial density and colonization of the nasopharynx has not been reported. The current study was performed to assess the presence and density of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis in the nasopharynx of 4-7-year-old children during URTI and when well., Methods: Nasal samples were obtained during 4 surveillance periods when children were asymptomatic and whenever they had symptoms of URTI. Respiratory viruses and bacterial pathogens were identified and quantified using polymerase chain reaction., Results: The proportion of children colonized with all 3 bacteria was higher during visits for acute URTI than during asymptomatic surveillance visits. Mean bacterial densities were significantly higher at all visits for all 3 pathogens when a virus was detected. The differences between the means were 1.0, 0.4, and 0.7 log10 colony-forming unit equivalents per milliliter for S. pneumoniae, H. influenzae, and M. catarrhalis, respectively, compared with visits in which virus was not detected. The percentage of children colonized and density were also higher at asymptomatic visits in which virus was detected than at visits in which virus was not detected., Conclusion: The density and frequency of colonization with S. pneumoniae, H. influenzae, and M. catarrhalis in nasal wash samples increase during periods of both symptomatic and asymptomatic viral infection. Increases in bacterial colonization observed during asymptomatic viral infection were nearly the same magnitude as when children were symptomatic.

Background and Objective: Respiratory syncytial virus (RSV) is the most significant cause of acute respiratory infection (ARI) in early life. RSV and other respiratory viruses are known to stimulate substantial outgrowth of potentially pathogenic bacteria in the upper airways of young children. However, the clinical significance of interactions between viruses and bacteria is currently unclear. The present study aimed to clarify the effect of viral and bacterial co-detections on disease severity during paediatric ARI., Methods: Nasopharyngeal aspirates from children under 2 years of age presenting with ARI to the emergency department were screened by quantitative PCR for 17 respiratory viruses and the bacterial pathogens Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis. Associations between pathogen detection and clinical measures of disease severity were investigated., Results: RSV was the most common virus detected, present in 29 of 58 samples from children with ARI (50%). Detection of S. pneumoniae was significantly more frequent during RSV infections compared to other respiratory viruses (adjusted effect size: 1.8, P: 0.03), and co-detection of both pathogens was associated with higher clinical disease severity scores (adjusted effect size: 1.2, P: 0.03)., Conclusion: Co-detection of RSV and S. pneumoniae in the nasopharynx was associated with more severe ARI, suggesting that S. pneumoniae colonization plays a pathogenic role in young children., (© 2017 Asian Pacific Society of Respirology.)

Background: The effect of pneumococcal vaccination is widely variable when measured by nasopharyngeal carriage of vaccine and non-vaccine targets. The aim of this study was to compare the carriage rates and metabolic activity of Streptococcus pneumoniae, Staphylococcus aureus, Haemophilus influenzae and Moraxella catarrhalis among children who were or were not vaccinated with PCV10., Methods: We included children with acute respiratory infection aged 6-23months from a cross-sectional study (CHIADO-IVAS). Nasopharyngeal aspirates were collected and respiratory pathogens were quantified by nCounter digital transcriptomics (Nanostring) and metagenomic sequencing of 16S ribosomal RNA (Illumina). The metabolic rate was calculated by the ratio between RNA transcripts and 16S DNA reads., Results: Out of the 80 patients in this study, 53 were vaccinated with PCV10 and 27 were unvaccinated. There was no difference in nasopharyngeal carriage rates of S. pneumoniae, S. aureus, H. influenzae or M. catarrhalis by either transcriptomic analysis or 16S metagenomics. However, unvaccinated children presented a higher metabolic rate for S. pneumoniae compared to PCV10-vaccinated children (Median [25-75th percentiles]: 126 [22.75-218.41] vs. 0[0-47.83], p=0.004). Furthermore, unvaccinated children presented a positive correlation between mRNA counts and 16S DNA reads for S. pneumoniae (r=0.707; p<0.001) and H. influenzae (r=0.525; p=0.005), in contrast to vaccinated children. No such effect was observed for S. aureus and M. catarrhalis., Conclusions: Vaccination by PCV10 exerts a pathogen-specific effect on pneumococcal metabolic rate. Pathogen RNA/DNA ratio might represent a more sensitive readout for vaccine follow-up, as compared to nasopharyngeal carriage., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Background: Interferences between pathogenic bacteria and specific commensals are known. We determined the interactions between nasopharyngeal microbial pathogens and commensals during viral upper respiratory tract infection (URI) and acute otitis media (AOM) in infants., Methods: We analyzed 971 specimens collected monthly and during URI and AOM episodes from 139 infants. The 16S rRNA V4 gene regions were sequenced on the Illumina MiSeq platform., Results: Among the high abundant genus-level nasopharyngeal microbiota were Moraxella, Haemophilus, and Streptococcus (3 otopathogen genera), Corynebacterium, Dolosigranulum, Staphylococcus, Acinetobacter, Pseudomonas, and Bifidobacterium. Bacterial diversity was lower in culture-positive samples for Streptococcus pneumoniae, and Haemophilus influenzae, compared to cultured-negative samples. URI frequencies were positively associated with increasing trend in otopathogen colonization. AOM frequencies were associated with decreasing trend in Micrococcus colonization. During URI and AOM, there were increases in abundance of otopathogen genera and decreases in Pseudomonas, Myroides, Yersinia, and Sphingomonas. Otopathogen abundance was increased during symptomatic viral infection, but not during asymptomatic infection. The risk for AOM complicating URI was reduced by increased abundance of Staphylococcus and Sphingobium., Conclusion: Otopathogen genera played the key roles in URI and AOM occurrences. Staphylococcus counteracts otopathogens thus Staphylococcal colonization may be beneficial, rather than harmful. While Sphingobium may play a role in preventing AOM complicating URI, the commonly used probiotic Bifidobacterium did not play a significant role during URI or AOM. The role of less common commensals in counteracting the deleterious effects of otopathogens requires further studies.

Objectives: We sought to understand how polymicrobial colonization varies during health, viral upper respiratory infection (URI) and acute upper respiratory bacterial infection to understand differences in infection-prone vs. non-prone patients., Methods: Nasopharyngeal (NP) samples were collected from 74 acute otitis media (AOM) infection-prone and 754 non-prone children during 2094 healthy visits, 673 viral URI visits and 631 AOM visits. Three otopathogens Streptococcus pneumoniae (Spn), Nontypeable Haemophilus influenzae (NTHi), and Moraxella catarrhalis (Mcat) were identified by culture., Results: NP colonization rates of multiple otopathogens during health were significantly lower than during viral URI, and during URI they were lower than at onset of upper respiratory bacterial infection in both AOM infection-prone and non-prone children. AOM infection-prone children had higher polymicrobial colonization rates than non-prone children during health, viral URI and AOM. Polymicrobial colonization rates of AOM infection-prone children during health were equivalent to that of non-prone children during viral URI, and during viral URI were equivalent to that of non-prone during AOM infection. Spn colonization was positively associated with NTHi and Mcat colonization during health, but negatively during AOM infection., Conclusion: The infection-prone patients more frequently have multiple potential bacterial pathogens in the NP than the non-prone patients. Polymicrobial interaction in the NP differs during health and at onset of infection., (Copyright © 2017 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

The efficacy of chocolate agar, versus bacitracin, vancomycin, clindamycin, chocolate agar (BVCCA) for the isolation of non-typeable Haemophilus influenzae (NTHi) from nasopharyngeal swabs was determined. BVCCA cultured NTHi from 97.3% of NTHi-positive swabs, compared to 87.1% for chocolate agar. To maximise culture sensitivity, the use of both media is recommended., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: The accuracy of nasopharyngeal aspirate (NPA) specimens in detecting lower respiratory pathogens remains controversial. The objective of this study was to evaluate the diagnostic accuracy of aspirates (NPAs) specimen in lower respiratory tract infections (LRTIs) in children., Methods: The prospective study was designed to collect the data of paired NPAs and bronchoalveolar lavage fluids from children with acute LRTIs from January 2013 to December 2015. All specimens were subjected to pathogen detection: bacterial detection by culture, Mycoplasma pneumoniae (Mp) detection by polymerase chain reaction assay and virus (influenza A and B viruses, parainfluenza virus [PIV] Types 1 and 3, respiratory syncytial virus, and adenovirus) detection by immunofluorescence assay. The diagnostic accuracy analysis of NPAs was stratified by age ≤3 years (n = 194) and >3 years (n = 294)., Results: We collected paired specimens from 488 children. The positive rate of pathogen was 61.6%. For Streptococcus pneumoniae, NPA culture had the specificity of 89.9% and negative predictive value of 100% in age ≤3 years, the specificity of 97.2% and negative predictive value of 98.9% in age >3 years. For Mp, the positive predictive values of NPA was 77.4% in children ≤3 years, and 89.1% in children >3 years. For PIV III, NPA specimen had the specificity of 99.8% and negative predictive value of 96.5% in children ≤3 years. For adenovirus, NPA had the specificity of 97.8% and negative predictive value of 98.4% in age ≤3 years, the specificity of 98.9% and negative predictive value of 99.3% in age >3 years., Conclusions: NPAs are less invasive diagnostic respiratory specimens, a negative NPA result is helpful in "rule out" lower airway infection; however, a positive result does not reliably "rule in" the presence of pathogens.

Background: This prospective study was performed to evaluate serotype distribution, multilocus sequence typing, and antibiotic susceptibility of Streptococcus pneumoniae identified in Korean children with acute otitis media (AOM) after the introduction of a 7-valent pneumococcal conjugate vaccine (PCV7)., Methods: Nasopharyngeal aspirates were collected from children diagnosed with AOM in seven hospitals in Korea. The bacteria identified in these samples and the serotypes, sequence types (STs), and antibiotic susceptibilities of S. pneumoniae isolates were evaluated., Results: A total of 390 children were enrolled, and bacteria were identified in 376 (96.4%) children. S. pneumoniae, Haemophilus influenzae and Moraxella catarrhalis were identified in 155 (39.7%), 127 (32.6%) and 86 (22.1%) children, respectively. Serotype 19A (22.4%) was the most common S. pneumoniae serotype, with serogroups 11 (14.7%) and 15 (13.5%) following. ST320 (23.5%) was the most common ST; ST166 (17.0%) and ST83 (8.5%) followed. The overall susceptibility rates of S. pneumoniae to oral penicillin V and amoxicillin/clavulanate were 2.6% and 53.2%, respectively. The susceptibility rate to cefditoren was 91.0%; however, the rates for other cephalosporins were less than 10.0%. Compared with other serogroups, S. pneumoniae serogroups 19, 11, and 15 showed significantly lower susceptibility rates to all the antibiotics tested., Conclusion: S. pneumoniae serotype 19A, serogroups 11 and 15 were the major nasopharyngeal-colonizing bacteria in Korean children with AOM after the introduction of PCV7. These relatively prevalent serotype/serogroups showed lower antibiotic susceptibility rates., (Copyright © 2016. Published by Elsevier Ltd.)

Haemophilus influenzae harbours a complex array of factors to resist human complement attack. As non-typeable H. influenzae (NTHi) strains do not possess a capsule, their serum resistance mainly depends on other mechanisms including LOS decoration. In this report, we describe the identification of a highly serum resistant, nasopharyngeal isolate (NTHi23) by screening a collection of 77 clinical isolates. For NTHi23, we defined the MLST sequence type 1133, which matches the profile of a previously published invasive NTHi isolate. A detailed genetic analysis revealed that NTHi23 shares several complement evading mechanisms with invasive disease isolates. These mechanisms include the functional expression of a retrograde phospholipid trafficking system and the presumable decoration of the LOS structure with sialic acid. By screening the NTHi23 population for spontaneous decreased serum resistance, we identified a clone, which was about 10 3 -fold more sensitive to complement-mediated killing. Genome-wide analysis of this isolate revealed a phase variation in the N'-terminal region of lpsA, leading to a truncated version of the glycosyltransferase (LpsA). We further showed that a NTHi23 lpsA mutant exhibits a decreased invasion rate into human alveolar basal epithelial cells. Since only a small proportion of the NTHi23 population expressed the serum sensitive phenotype, resulting from lpsA phase-off, we conclude that the nasopharyngeal environment selected for a population expressing the intact and functional glycosyltransferase., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

Background: In 2010, Greenland introduced the 13-valent pneumococcal conjugate vaccine (Prevnar 13®- PCV-13) in the childhood immunisation program. The authors aimed to evaluate the impact of PCV-13 on nasopharyngeal carriage of bacteria frequently associated with respiratory infections in children., Method: In 2013 a cross-sectional population-based study of nasopharyngeal carriage was conducted among Greenlandic children aged 0-6 years and results were compared with an equivalent study from 2011. Nasopharyngeal swab samples were tested for Streptococcus pneumoniae, non-typeable Haemophilus influenzae, Moraxella catarrhalis and Staphylococcus aureus. Pneumococcal serotyping was performed by Quellung reaction and serotype-specific antisera. Statistical analysis included logistic regression models, adjusting for known risk factors., Result: A total of 377 nasopharyngeal samples were collected. Overall carriage rate of S. pneumoniae remained unchanged from 2011 to 2013 (51% and 56%, p=0.13), but significant serotype shifts were observed among both vaccinated and unvaccinated children with marked reductions in carriage of vaccine-type pneumococci, counterbalanced by increasing carriage of non-vaccine types. Carriage rate of S. aureus decreased significantly among vaccinated children whereas that of M. catarrhalis increased., Conclusion: PCV-13 introduction in Greenland is associated with significant changes in nasopharyngeal bacterial carriage. Continued surveillance is warranted to clarify whether these changes are persistent, and affect the pattern of respiratory and invasive diseases in Greenland.

In March 2010, Brazil introduced the 10-valent pneumococcal conjugate vaccine (PCV10) in the routine infant immunization program using a 4-dose schedule and catch-up for children <23months. We investigated PCV10 effect on nasopharyngeal carriage with vaccine-type Streptococcus pneumoniae (Spn) and non-typeable Haemophilus influenzae (NTHi) among children in São Paulo city. Cross-sectional surveys were conducted in 2010 (baseline) and 2013 (post-PCV10). Healthy PCV-naïve children aged 12-23months were recruited from primary health centers during immunization campaigns. Nasopharyngeal swabs were collected and tested for Hi; for Spn, all baseline and a stratified random sample of 400 post-PCV10 swabs were tested. We compared vaccine-type Spn and NTHi carriage prevalence pre-/post-PCV10, and used logistic regression to estimate PCV10 effectiveness (1-adjusted odds ratio×100%). Overall 501 children were included in the baseline and 1167 in the post-PCV10 survey (including 400 tested for Spn). Spn was detected in 40.3% of children at baseline and 48.8% post-PCV10; PCV10 serotypes were found in 19.8% and 1.8% respectively, representing a decline of 90.9% (p<0.0001). Carriage of vaccine-related serotypes increased (10.8-21.0%, p<0.0001), driven primarily by a rise in serotype 6C (1.8-11.2%, p<0.0001); carriage of serotypes 6A and 19A did not significantly change. PCV10 effectiveness (4 doses) against vaccine-type carriage was 97.3% (95% confidence interval 88.7-99.3). NTHi prevalence increased from 26.0% (130/501) to 43.6% (509/1167, p<0.0001); PCV10 vaccination seemed significantly associated with NTHi carriage, even after adjusting for other known risk factors. Carriage with PCV10 serotypes among toddlers declined dramatically following PCV10 introduction in São Paulo, Brazil. No protection of PCV10 against NTHi was observed. Our findings contribute to a growing body of evidence of PCV10 impact on vaccine-type carriage and highlight the importance of PCV10 as a tool to reduce the burden of pneumococcal disease in Brazil and globally., (Copyright © 2016. Published by Elsevier Ltd.)

Background: Healthy condition and chronic diseases may be associated with microbiota composition and its properties. The prevalence of respiratory haemophili with respect to their phenotypes including the ability to biofilm formation in patients with sarcoidosis was assayed., Methods: Nasopharynx and sputum specimens were taken in 31 patients with sarcoidosis (average age 42.6 ± 13), and nasopharynx specimens were taken in 37 healthy people (average age 44.6 ± 11.6). Haemophili were identified by API-NH microtest and by the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) system. Biofilm was visualised by crystal violet staining and confocal scanning laser microscopy (CSLM). The statistical analysis was performed with Statgraphics Plus for Windows., Results: In total, 30/31 patients with sarcoidosis and 31/37 healthy people were colonized by Haemophilus influenzae (6/30 vs. 1/31) and Haemophilus parainfluenzae (28/30 vs. 31/31) in the nasopharynx. The overall number of nasopharyngeal haemophili isolates was 59 in patients with sarcoidosis and 67 in healthy volunteers (H. influenzae 6/59 vs. 1/67, P = 0.05; H. parainfluenzae 47/59 vs. 65/67, P = 0.0032). Moreover, the decreased number of H. parainfluenzae biofilm-producing isolates was shown in nasopharyngeal samples in patients with sarcoidosis as compared to healthy people (19/31 vs. 57/65, P = 0.006), especially with respect to isolates classified as strong and very strong biofilm-producers (8/31 vs. 39/65, P = 0.002)., Conclusions: The obtained data suggest that the qualitative and quantitative changes within the respiratory microbiota concerning the overall prevalence of H. influenzae together with the decreased number of H. parainfluenzae strains and the decreased rate of H. parainfluenzae biofilm-producing isolates as compared to healthy people may be associated with sarcoidosis.

Background: In West Africa, the carriage of Group B Streptococcus (GBS), among infants is poorly characterised. We investigated co-carriage of GBS with other respiratory pathogens in the infants' nasopharynx in The Gambia., Methods: We assessed the carriage, serotypes and antibiotic susceptibility of Beta-haemolytic Streptococci (BHS) groups A-G; along with the carriage of Streptococcus pneumoniae; Haemophilus influenzae; Staphylococcus aureus and Moraxella catarrhalis in 1200 two-month old infants., Results: The BHS prevalence was 20.0 % and GBS dominated (13.8 %), particularly serotypes V and II; serotype V being negatively associated with H. Influenzae carriage (OR 0.41 [95 % CI: 0.18-0.93], p = 0.033). Although co-colonization of GBS and other BHS was not seen, colonization with GBS was positively associated with S. aureus (OR 1.89 [95 % CI: 1.33-2.69], P < 0.001) and negatively associated with S. pneumoniae (OR 0.47 [95 % CI: 0.33-0.67], p < 0.001) and M. catarrhalis (OR 0.61 [95 % CI: 0.40-0.92], p = 0.017). ≥ 89 % of GBS isolates were susceptible to most antibiotics tested, except for tetracycline resistance, which was 89 %., Conclusion: This study provides baseline data on the carriage of GBS in two month old infants from West Africa. The dominant serotypes of GBS in this setting are serotypes V and II. This may be important for future GBS vaccine development for the West African sub-region.

After introduction of the 7-valent pneumococcal conjugate vaccine (PCV7) in the infant national immunization program (NIP) in the Netherlands in 2006, Streptococcus pneumoniae strains of the non-vaccine serotype 19A emerged and became the dominant serotype in carriage in children and their parents. Similar patterns were observed in other European countries and the United States. Increases in carriage rates of Staphylococcus aureus and non-typeable (NT) Haemophilus influenzae were also observed. After switching of PCV7 to 10-valent vaccine (PCV10) in 2011, a new carriage surveillance study was performed in the winter of 2012/2013. Nasopharyngeal carriage of S. pneumoniae, H. influenzae, S. aureus, and Moraxella catarrhalis was determined by conventional culture in 330 PCV10-vaccinated 11-month-old children, 330 PCV7-vaccinated 24-month-old children, and their parents. Carriage prevalence was compared with similar carriage studies conducted in 2005, 2009, and 2010/2011. Although serotype 19A remained the most frequently carried pneumococcal serotype in children, prevalence of 19A significantly declined in PCV7-vaccinated 24-month-old children (14% to 8%, p=0.01), but less in PCV10-vaccinated 11-month-old children (12% to 9%, p=0.31). Carriage of H. influenzae remained stable at an elevated level (65% in 11-month-olds and 69% in 24-month-olds), while the carriage of S. aureus returned to pre-PCV7 levels in 11-month-old children (14% in 2010/2011 to 7% in 2012/2013), but not in 24-month-olds (remained at 7%). Our results might indicate a new balance between replacing non-vaccine pneumococcal serotypes and other potential pathogenic bacteria in nasopharyngeal carriage. Carriage studies are valuable tools in assessing vaccine effects on pathogens circulating in the population, for evaluation of PCV impact, and in predicting changes in respiratory and invasive disease., (Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Bacterial vaccines can reduce carriage rates. Colonization is usually a binary endpoint. Real time quantitative PCR (qPCR) can quantify bacterial DNA in mucosal samples over a wide range. Using culture and single-gene species-specific qPCRs for Streptococcus pneumoniae (lytA), Streptococcus pyogenes (ntpC), Moraxella catarrhalis (ompJ), Haemophilus influenzae (hdp) and Staphylococcus aureus (nuc) and standard curves against log-phase reference strain broth cultures we described frequency and peak density distributions of carriage in nasopharyngeal swabs from 161 healthy 2-4 y old children collected into STGG broth. In general, detection by qPCR and culture was consistent. Discordance mostly occurred at lower detection thresholds of both methods, although PCR assays for S. pyogenes and S. aureus were less sensitive. Density varied across 5-7 orders of magnitude for the 5 species with the abundant species skewed toward high values (modes: S. pneumoniae log3-4, M. catarrhalis & H. influenzae log4-5 CFU/ml broth). Wide ranges of bacterial DNA concentrations in healthy children carrying these bacteria could mean that different individuals at different times vary greatly in infectiousness. Understanding the host, microbial and environmental determinants of colonization density will permit more accurate prediction of vaccine effectiveness.

Previously, the Fiji Pneumococcal Project (FiPP) evaluated reduced dose immunization schedules that incorporated pneumococcal protein conjugate and/or polysaccharide vaccine (PCV7 and 23vPPV, respectively). Immune hyporesponsiveness was observed in children vaccinated with 23vPPV at 12 months of age compared with children who did not receive 23vPPV. Here we assess the long-term impact of 23vPPV vaccination on nasopharyngeal carriage rates and densities of Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus and Moraxella catarrhalis. Nasopharyngeal swabs (n=194) were obtained from healthy children who participated in FiPP (now aged 5-7 years). S. pneumoniae were isolated and identified by standard culture-based methods, and serotyped using latex agglutination and the Quellung reaction. Carriage rates and densities of S. pneumoniae, H. influenzae, S. aureus and M. catarrhalis were determined using real-time quantitative PCR. There were no differences in the rate or density of S. pneumoniae, H. influenzae or M. catarrhalis carriage by PCV7 dose or 23vPPV vaccination in the vaccinated participants overall. However, differences were observed between the two main ethnic groups: Fijian children of Indian descent (Indo-Fijian) were less likely to carry S. pneumoniae, H. influenzae and M. catarrhalis, and there was evidence of a higher carriage rate of S. aureus compared with indigenous Fijian (iTaukei) children. Polysaccharide vaccination appeared to have effects that varied between ethnic groups, with 23vPPV vaccination associated with a higher carriage rate of S. aureus in iTaukei children, while there was a lower carriage rate of S. pneumoniae associated with 23vPPV vaccination in Indo-Fijian children. Overall, polysaccharide vaccination had no long-term impact on pneumococcal carriage, but may have impacted on S. aureus carriage and have varying effects in ethnic groups, suggesting current WHO vaccine schedule recommendations against the use of 23vPPV in children under two years of age are appropriate., (Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objectives: Animal studies suggest that RSV increases nasopharyngeal (NP) bacterial colonization facilitating bacterial infections. We investigated the influence of antibiotic treatment and colonization with potentially pathogenic bacteria on inflammatory markers and disease severity in RSV-infected in infants., Methods: Healthy young infants hospitalized with RSV bronchiolitis (n = 136) and age-matched healthy controls (n = 23) were enrolled and NP samples cultured for potentially pathogenic bacteria including: Gram-positive bacteria (GPB): Staphylococcus aureus, Streptococcus pneumoniae, β-hemolytic Streptococcus; and Gram-negative bacteria (GNB): Moraxella catarrhalis and Haemophilus influenzae. Clinical parameters and plasma IL-8, IL-6 and TNF-α concentrations were compared according to the bacterial class and antibiotic treatment., Results: Antibiotic treatment decreased by 10-fold NP bacterial recovery. Eighty-one percent of RSV infants who did not receive antibiotics before sample collection were colonized with pathogenic bacteria. Overall, GNB were identified in 21% of patients versus 4% of controls who were mostly colonized with GPB. Additionally, in RSV patients NP white blood cell counts (p = 0.026), and blood neutrophils (p = 0.02) were higher in those colonized with potentially pathogenic bacteria versus respiratory flora. RSV patients colonized with GNB had higher plasma IL-8 (p = 0.01) and IL-6 (p < 0.01) concentrations than controls, and required longer duration of oxygen (p = 0.049)., Conclusions: Infants with RSV bronchiolitis colonized with potentially pathogenic bacteria had increased numbers of mucosal and systemic inflammatory cells. Specifically, colonization with GNB was associated with higher concentrations of proinflammatory cytokines and a trend towards increased disease severity., (Copyright © 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

Background: Streptococcus pneumoniae, Haemophilus influenzae and Staphylococcus aureus are all potentially pathogenic, which frequently colonize the nasopharynx (NP) prior to causing disease. We studied bacterial NP-colonization in 321 HIV-infected and 243 HIV-uninfected children vaccinated with 7-valent pneumococcal conjugate vaccine (PCV7) at 6, 10 and 14 weeks of age., Methods: HIV-uninfected infants included those born to HIV-uninfected (HUU) and HIV-infected women (HEU); HIV-infected children with CD4+ lymphocyte ≥25% were randomized to initiate antiretroviral therapy immediately (ART-Immed) or when clinically indicated (ART-Def). Nasopharyngeal swabs for bacterial culture were taken prior to each PCV7 dose (Visits 1-3) and at 20, 39, 47 and 67 weeks of age (Visits 4-7). Swabs were cultured by standard methods and pneumococcal serotyping done by the Quellung method., Results: Colonization patterns for pneumococcus, H. influenzae and S. aureus did not differ between HUU and HEU children; and were also generally similar between ART-Def and ART-Immed children. Prevalence of PCV7-serotype colonization was similar between HIV-infected and HIV-uninfected children, however, overall pneumococcal and specifically non-vaccine serotype colonization tended to be lower in HIV-infected children. HIV-infected children also had a 44% lower prevalence of S. aureus colonization at Visit-1 (p=0.010); and H. influenzae colonization was also lower among HIV-infected than HIV-uninfected children at Visit-2, Visit-3, Visit-6 and Visit-7., Conclusion: Vaccine-serotype colonization is similar in PCV-immunized HIV-infected and HIV-uninfected children. We, however, identified a lower prevalence of overall-pneumococcal and H. influenzae colonization in HIV-infected children post-PCV vaccination, the clinical-relevance of which warrants further study., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Haemophili are pathogenic or opportunistic bacteria often colonizing the upper respiratory tract mucosa. The prevalence of Haemophilus influenzae (with serotypes distribution), and H. parainfluenzae in the nasopharynx and/or the adenoid core in children with recurrent pharyngotonsillitis undergoing adenoidectomy was assessed. Haemophili isolates were investigated for their ability to biofilm production.Nasopharyngeal swabs and the adenoid core were collected from 164 children who underwent adenoidectomy (2-5 years old). Bacteria were identified by the standard methods. Serotyping of H. influenzae was performed using polyclonal and monoclonal antisera. Biofilm formation was detected spectrophotometrically using 96-well microplates and 0.1% crystal violet.Ninety seven percent (159/164) children who underwent adenoidectomy were colonized by Haemophilus spp. The adenoid core was colonized in 99.4% (158/159) children, whereas the nasopharynx in 47.2% (75/159) children (P < 0.0001). In 32% (51/159) children only encapsulated (typeable) isolates of H. influenzae were identified, in 22.6% (36/159) children only (nonencapsulated) H. influenzae NTHi (nonencapsulated) isolates were present, whereas 7.5% (12/159) children were colonized by both types. 14.5% (23/159) children were colonized by untypeable (rough) H. influenzae. In 22% (35/159) children H. influenzae serotype d was isolated. Totally, 192 isolates of H. influenzae, 96 isolates of H. parainfluenzae and 14 isolates of other Haemophilus spp. were selected. In 20.1% (32/159) children 2 or 3 phenotypically different isolates of the same species (H. influenzae or H. parainfluenzae) or serotypes (H. influenzae) were identified in 1 child. 67.2% (129/192) isolates of H. influenzae, 56.3% (54/96) isolates of H. parainfluenzae and 85.7% (12/14) isolates of other Haemophilus spp. were positive for biofilm production. Statistically significant differences (P = 0.0029) among H. parainfluenzae biofilm producers and nonproducers in the adenoid core and the nasopharynx were detected.H. influenzae and H. parainfluenzae carriage rate was comparatively higher in the adenoid core than that in the nasopharynx in children undergoing adenoidectomy, suggesting that their involvement in chronic adenoiditis. The growth in the biofilm seems to be an important feature of haemophili colonizing the upper respiratory tract responsible for their persistence.

Sulfadoxine-pyrimethamine (SP) plus azithromycin (AZ) (SPAZ) has the potential for intermittent preventive treatment of malaria in pregnancy (IPTp), but its use could increase circulation of antibiotic-resistant bacteria associated with severe pediatric infections. We evaluated the effect of monthly SPAZ-IPTp compared to a single course of SP plus chloroquine (SPCQ) on maternal nasopharyngeal carriage and antibiotic susceptibility of Streptococcus pneumoniae, Haemophilus influenzae, and Staphylococcus aureus at delivery among 854 women participating in a randomized controlled trial in Papua New Guinea. Serotyping was performed, and antibiotic susceptibility was evaluated by disk diffusion and Etest. Potential risk factors for carriage were examined. Nasopharyngeal carriage at delivery of S. pneumoniae (SPAZ, 7.2% [30/418], versus SPCQ, 19.3% [84/436]; P<0.001) and H. influenzae (2.9% [12/418] versus 6.0% [26/436], P=0.028), but not S. aureus, was significantly reduced among women who had received SPAZ-IPTp. The number of macrolide-resistant pneumococcal isolates was small but increased in the SPAZ group (13.3% [4/30], versus SPCQ, 2.2% [2/91]; P=0.033). The proportions of isolates with serotypes covered by the 13-valent pneumococcal conjugate vaccine were similar (SPAZ, 10.3% [3/29], versus SPCQ, 17.6% [16/91]; P=0.352). Although macrolide-resistant isolates were rare, they were more commonly detected in women who had received SPAZ-IPTp, despite the significant reduction of maternal carriage of S. pneumoniae and H. influenzae observed in this group. Future studies on SPAZ-IPTp should evaluate carriage and persistence of macrolide-resistant S. pneumoniae and other pathogenic bacteria in both mothers and infants and assess the clinical significance of their circulation., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Aim: This study aims to describe the microbiology of middle ear fluid (MEF) in a cohort of children vaccinated with Streptococcus pneumoniae conjugate vaccine (PCV7) having ventilation tube insertion. Nasopharyngeal (NP) carriage of otopathogens in these children is compared with children without history of otitis media., Methods: Between May and November 2011, MEF and NP samples from 325 children aged <3 years were collected in three major centres in New Zealand at the time of ventilation tube insertion. An age-matched non-otitis-prone comparison group of 137 children had NP samples taken. A questionnaire was completed by both groups., Results: Immunisation coverage with at least one dose of PCV7 was 97%. Haemophilus influenzae was cultured in 19.4% of MEF and was polymerase chain reaction (PCR) positive in 43.4%. S. pneumoniae and Moraxella catarrhalis were cultured in <10% of MEF samples but were PCR positive for 23.1% and 38.7%, respectively. H. influenzae was the most common organism isolated from NP samples (60%) in the grommet group, while M. catarrhalis (56%) was the most common in the non-otitis prone group. S. pneumoniae was more commonly found in the nasopharynx of children with ear disease (41% vs. 29%). 19F was the most prominent S. pneumoniae serotype in NP samples of both groups, but no serotype dominated in MEF. Ninety-five per cent of H. influenzae isolates were confirmed to be non-typeable H. influenzae., Conclusion: In this cohort of children with established ear disease requiring surgical intervention, non-typeable H. influenzae is the dominant pathogen in both the nasopharynx and MEF., (© 2014 The Authors. Journal of Paediatrics and Child Health published by Wiley Publishing Asia Pty Ltd on behalf of Paediatrics and Child Health Division. (The Royal Australasian College of Physicians).)

Background: Haemophilus influenzae is one of the main pathogens that cause community-acquired respiratory infections in children. Our previous study showed that H. influenzae is the second most common pathogen causing pneumonia and accounts for 30-50% of bacterial meningitis among Chinese children. H. influenzae carriage in children and its resistance to commonly used antimicrobials varies widely both geographically and over time., Results: Surveys of the nasopharyngeal carriage of H. influenzae in children younger than 5 years of age with acute respiratory tract infection (ARI) were conducted in Beijing Children's Hospital, China in 2000, 2002, 2010, and 2012. The overall annual carriage rates of H. influenzae among children younger than 5 years of age with ARI were 35.5%, 20.6%, 14.4%, and 18.7%, and the percentages of H. influenzae isolates producing β-lactamase were 4%, 13%, 27.1%, and 31%, respectively. The percentages of susceptibility to ampicillin progressively decreased from 96% (2000) to 87% (2002) to 63% (2010) to 61% (2012). All of the ampicillin-resistant isolates were found to be beta-lactamase producers. The susceptibility to tetracycline increased from 54% (2000) to 60% (2002) to 91.5% (2010) to 94.5% (2012). No statistically significant differences were observed in the susceptibility to cefaclor, cefuroxime, sulfamethoxazole, and chloramphenicol. Amoxicillin/clavulanic acid and ceftriaxone were the most effective antimicrobials for the isolates of H. influenzae across the 10-year period., Conclusions: This report on the H. influenzae carriage rates in children and the susceptibility of these bacteria to commonly used antibiotics showed that H. influenzae carriage decreased from 2000 to 2012. Additionally, the percentage of β-lactamase-producing isolates increased while their susceptibility to ampicillin progressively decreased during this time. These results indicate that the appropriate empirical antimicrobial therapy should be changed for pediatric patients in China.

Conclusion: Although children vaccinated with heptavalent pneumococcal conjugate vaccine (PCV) had fewer episodes of acute otitis media (AOM), this trial was unable to prove a simultaneous decrease in nasopharyngeal carriage., Objective: Carriage rates of AOM pathogens in the nasopharynx are high among children, and colonization is the first step towards infection. The possible impact of PCV on carriage is therefore of interest, particularly in children with recurrent AOM. The aims of this study were to examine the effect of heptavalent PCV on carriage of AOM pathogens in children at high risk of developing recurrent disease, and to monitor carriage of resistant pathogens in vaccinated and unvaccinated children., Methods: A total of 109 children with an onset of AOM before 6 months of age, 89 of whom developed recurrent disease, were enrolled in a trial. Fifty-two children were vaccinated and all were closely monitored for 3 years., Results: There was no difference statistically between vaccinated children and controls concerning the carriage of any of the major AOM pathogens. There was evidence of within-child clustering for S. pneumoniae (p = 0.002) and H. influenzae (p < 0.001), indicating that children continued to carry either species over time. Resistance rates were generally low and comparable with national levels.

Background: We sought to determine if nasopharyngeal (NP) cultures taken at times of healthy visits or at onset of acute otitis media (AOM) could predict the otopathogen mix and antibiotic-susceptibility of middle ear isolates as determined by middle ear fluid (MEF) cultures obtained by tympanocentesis., Methods: During a 7-year-prospective study of 619 children from Jun 2006-Aug 2013, NP cultures were obtained from 6-30 month olds at healthy visits and NP and MEF (by tympanocentesis) at onset of AOM episodes., Results: 2601 NP and 530 MEF samples were collected. During healthy visits, S. pneumoniae (Spn) was isolated from 656 (31.7%) NP cultures compared to 253 (12.2%) for Nontypeable Haemophilus influenzae (NTHi) and 723 (34.9%) for Moraxella catarrhalis (Mcat). At onset of AOM 256 (48.3%) of 530 NP samples were culture positive for Spn, 223 (42%) for NTHi and 251 (47.4%) for Mcat, alone or in combinations. At 530 AOM visits, Spn was isolated from 152 (28.7%) of MEF compared to 196 (37.0%) for NTHi and 104 (19.6%) for Mcat. NP cultures collected at onset of AOM but not when children were healthy had predictive value for epidemiologic antibiotic susceptibility pattern assessments., Conclusions: NP cultures at onset of AOM more closely correlate with otopathogen mix than NP cultures at healthy visits using MEF culture as the gold standard, but the correlation was too low to allow NP cultures to be recommended as a substitute for MEF culture. For epidemiology purposes, antibiotic susceptibility of MEF isolates can be predicted by NP culture results when samples are collected at onset of AOM.

Nontypeable Haemophilus influenzae (NTHi) are Gram-negative coccobacilli that colonize the human pharynx, their only known natural reservoir. Adherence to the host epithelium facilitates NTHi colonization and marks one of the first steps in NTHi pathogenesis. Epithelial cell attachment is mediated, in part, by a pair of high molecular weight (HMW) adhesins that are highly immunogenic, antigenically diverse, and display a wide range of amino acid diversity both within and between isolates. In this study, the prevalence of hmwA, which encodes the HMW adhesin, was determined for a collection of 170 NTHi isolates recovered from the middle ears of children with otitis media (OM isolates) or throats or nasopharynges of healthy children (commensal isolates) from Finland, Israel, and the U.S. Overall, hmwA was detected in 61% of NTHi isolates and was significantly more prevalent (P=0.004) among OM isolates than among commensal isolates; the prevalence ratio comparing hmwA prevalence among ear isolates with that of commensal isolates was 1.47 (95% CI (1.12, 1.92)). Ninety-five percent (98/103) of the hmwA-positive NTHi isolates possessed two hmw loci. To advance our understanding of hmwA binding sequence diversity, we determined the DNA sequence of the hmwA binding region of 33 isolates from this collection. The average amino acid identity across all hmwA sequences was 62%. Phylogenetic analyses of the hmwA binding revealed four distinct sequence clusters, and the majority of hmwA sequences (83%) belonged to one of two dominant sequence clusters. hmwA sequences did not cluster by chromosomal location, geographic region, or disease status., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Objectives: Bacterial carriage in the upper respiratory tract is usually asymptomatic but can lead to respiratory tract infection (RTI), meningitis and septicaemia. We aimed to provide a baseline measure of Streptococcus pneumoniae, Moraxella catarrhalis, Pseudomonas aeruginosa, Staphylococcus aureus, Haemophilus influenzae and Neisseria meningitidis carriage within the community. Self-swabbing and healthcare professional (HCP) swabbing were compared., Design: Cross-sectional study., Setting: Individuals registered at 20 general practitioner practices within the Wessex Primary Care Research Network South West, UK., Participants: 10,448 individuals were invited to participate; 5394 within a self-swabbing group and 5054 within a HCP swabbing group. Self-swabbing invitees included 2405 individuals aged 0-4 years and 3349 individuals aged ≥5 years. HCP swabbing invitees included 1908 individuals aged 0-4 years and 3146 individuals aged ≥5 years., Results: 1574 (15.1%) individuals participated, 1260 (23.4%, 95% CI 22.3% to 24.5%) undertaking self-swabbing and 314 (6.2%, 95% CI 5.5% to 6.9%) undertaking HCP-led swabbing. Participation was lower in young children and more deprived practice locations. Swab positivity rates were 34.8% (95% CI 32.2% to 37.4%) for self-taken nose swabs (NS), 19% (95% CI 16.8% to 21.2%) for self-taken whole mouth swabs (WMS), 25.2% (95% CI 20.4% to 30%) for nasopharyngeal swabs (NPS) and 33.4% (95% CI 28.2% to 38.6%) for HCP-taken WMS. Carriage rates of S. aureus were highest in NS (21.3%). S. pneumoniae carriage was highest in NS (11%) and NPS (7.4%). M. catarrhalis carriage was highest in HCP-taken WMS (28.8%). H. influenzae and P. aeruginosa carriage were similar between swab types. N. meningitidis was not detected in any swab. Age and recent RTI affected carriage of S. pneumoniae and H. influenzae. Participant costs were lower for self-swabbing (£41.21) versus HCP swabbing (£69.66)., Conclusions: Higher participation and lower costs of self-swabbing as well as sensitivity of self-swabbing favour this method for use in large population-based respiratory carriage studies., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

During a 5-year prospective study of nasopharyngeal (NP) colonization and acute otitis media (AOM) infections in children during the 7-valent pneumococcal conjugate vaccine (PCV) era (July 2006-June 2011) we studied risk factors for NP colonization and AOM. NP samples were collected at ages 6, 9, 12, 15, 18, 24, and 30 months during well-child visits. Additionally, NP and middle ear fluid (MEF) samples were collected at onset of every AOM episode. From 1825 visits (n = 464 children), 5301 NP and 570 MEF samples were collected and analysed for potential otopathogens. Daycare attendance, NP colonization by Moraxella catarrhalis, and siblings aged <5 years increased the risk of Streptococcus pneumoniae NP colonization. NP colonization with S. pneumoniae, M. catarrhalis, or Haemophilus influenzae and a family history of OM increased the risk of AOM. Risk factors that increase the risk of pneumococcal AOM will be important to reassess as we move into a new 13-valent PCV era, especially co-colonization with other potential otopathogens.