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Ritah F Mutagonda,1,2 George Bwire,3 Raphael Zozimus Sangeda,2,3 Manase Kilonzi,1 Hamu Mlyuka,1 Joyce Ndunguru,2,4 Agnes Jonathan,2,4 Julie Makani,2,4 Irene Kida Minja,2,5 Paschal Ruggajo,2,6 Emmanuel Balandya,2,7 Appolinary AR Kamuhabwa1 1Department of Clinical Pharmacy and Pharmacology, Muhimbili University of Health and Allied Sciences, Dar es salaam, Tanzania; 2Sickle Cell Programme, Muhimbili University of Health and Allied Sciences, Dar es salaam, Tanzania; 3Department of Pharmaceutical Microbiology, Muhimbili University of Health and Allied Sciences, Dar es salaam, Tanzania; 4Department of Hematology and Blood Transfusion, Muhimbili University of Health and Allied Sciences, Dar es salaam, Tanzania; 5Department of Restorative Dentistry, Muhimbili University of Health and Allied Sciences, Dar es salaam, Tanzania; 6Department of Internal Medicine, Muhimbili University of Health and Allied Sciences, Dar es salaam, Tanzania; 7Department of Physiology, Muhimbili University of Health and Allied Sciences, Dar es salaam, TanzaniaCorrespondence: Ritah F Mutagonda, Department of Clinical Pharmacy and Pharmacology, Muhimbili University of Health and Allied Sciences, P.O BOX 65013, Dar es salaam, Tanzania, Tel +255 713 816481, Email mutagonda@muhas.ac.tz; rittdavisrida@yahoo.comBackground: Bacterial infections contribute significantly to morbidity and mortality in sickle cell disease (SCD) patients, particularly children under five years of age. In Tanzania, prophylaxis against pneumococcal infection among children with SCD advocates the use of both oral penicillin V (PV) and pneumococcal vaccines (PNV). Therefore, this study aimed to investigate nasopharyngeal carriage and antibiogram of Streptococcal pneumoniae (S. pneumoniae) and Staphylococcus aureus (S. aureus) in children with SCD in Tanzania.Methods: This cross-sectional study was undertaken at the two Sickle Pan-African Research Consortium (SPARCO) study sites in Dar es salaam, Tanzania. The study was conducted for six months and enrolled children with SCD between the ages of 6 to 59-months. A semi-structured questionnaire was used to collect patient data. Nasopharyngeal swabs were collected from all participants and cultured for Streptococcal pneumoniae and other bacterial isolates. Antimicrobial susceptibility tests of the isolates were done using the disc diffusion method.Results: Out of 204 participants, the overall prevalence of bacterial carriage was 53.4%, with S. aureus (23.5%), coagulase-negative Staphylococci (CoNS) (23%) and S. pneumoniae (7.8%) being commonly isolated. In antibiotic susceptibility testing, S. aureus isolates were most resistant to penicillin (81.8%), whereas 81.3% of S. pneumoniae isolates were resistant to co-trimoxazole. The least antimicrobial resistance was observed for chloramphenicol for both S. aureus and S. pneumoniae isolates (6.3% versus 0%). The proportion of multi-drug resistance (MDR) was 66.7% for S. aureus isolates and 25% for S. pneumoniae isolates.Conclusion: There are substantially high nasopharyngeal carriage pathogenic bacteria in children with SCD in Dar es Salaam, Tanzania. The presence of MDR strains to the commonly used antibiotics suggests the need to reconsider optimizing antimicrobial prophylaxis in children with SCD and advocacy on pneumococcal vaccines.Keywords: pneumococcal carriage, bacterial pathogens, pneumococcal prophylaxis, antimicrobial resistance

A link between influenza infection and cardiovascular morbidity has been known for almost a century. This narrative review examined the cardiovascular complications associated with influenza and the potential mechanisms behind this relationship. The most common reported cardiovascular complications are cardiovascular death, myocardial infarction, and heart failure hospitalization. There are multiple proposed mechanisms driving the increased risk of cardiovascular complications. These mechanics involve influenza-specific effects such as direct cardiac infection and endothelial dysfunction leading to plaque destabilization and rupture, but also hypoxaemia and systemic inflammatory responses including increased metabolic demand, biomechanical stress, and hypercoagulability. The significance of the individual effects is unclear, and thus whether influenza directly or indirectly causes cardiovascular events is unknown. In conclusion, the risk of acute cardiovascular morbidity and mortality is elevated during influenza infection. The proposed underlying pathophysiological mechanisms support this association, but systemic responses to infection may drive this relationship.

Characterizing the genetic diversity of pathogens within the host promises to greatly improve surveillance and reconstruction of transmission chains. For bacteria, it also informs our understanding of inter-strain competition and how this shapes the distribution of resistant and sensitive bacteria. Here we study the genetic diversity of Streptococcus pneumoniae within 468 infants and 145 of their mothers by deep sequencing whole pneumococcal populations from 3,761 longitudinal nasopharyngeal samples. We demonstrate that deep sequencing has unsurpassed sensitivity for detecting multiple colonization, doubling the rate at which highly invasive serotype 1 bacteria were detected in carriage compared with gold-standard methods. The greater resolution identified an elevated rate of transmission from mothers to their children in the first year of the child's life. Comprehensive treatment data demonstrated that infants were at an elevated risk of both the acquisition and persistent colonization of a multidrug-resistant bacterium following antimicrobial treatment. Some alleles were enriched after antimicrobial treatment, suggesting that they aided persistence, but generally purifying selection dominated within-host evolution. Rates of co-colonization imply that in the absence of treatment, susceptible lineages outcompeted resistant lineages within the host. These results demonstrate the many benefits of deep sequencing for the genomic surveillance of bacterial pathogens. Longitudinal population deep sequencing of Streptococcus pneumoniae sampled from infants and their mothers improves our understanding of the dynamics of colonization, transmission, inter-strain competition and the impact of antibiotic treatment.

Streptococcus pneumoniae is an important and frequently carried respiratory pathogen that has the potential to cause serious invasive diseases, such as pneumonia, meningitis, and sepsis. Young children and older adults are among the most vulnerable to developing serious disease. With the arrival of the COVID-19 pandemic and the concomitant restrictive measures, invasive disease cases caused by respiratory bacterial species, including pneumococci, decreased substantially. Notably, the stringency of the containment measures as well as the visible reduction in the movement of people appeared to coincide with the drop in invasive disease cases. One could argue that wearing protective masks and adhering to social distancing guidelines to halt the spread of the SARS-CoV-2 virus, also led to a reduction in the person-to-person transmission of respiratory bacterial species. Although plausible, this conjecture is challenged by novel data obtained from our nasopharyngeal carriage study which is performed yearly in healthy daycare center attending children. A sustained and high pneumococcal carriage rate was observed amid periods of stringent restrictive measures. This finding prompts us to revisit the connection between nasopharyngeal colonization and invasion and invites us to look closer at the nasopharyngeal microbiome as a whole. ispartof: FRONTIERS IN CELLULAR AND INFECTION MICROBIOLOGY vol:11 ispartof: location:Switzerland status: published

The International Symposium on Pneumococci and Pneumococcal Diseases (ISPPD) is the premier global scientific symposium dedicated to the exchange, advancement and dissemination of the latest research on the pneumococcus, one of the world’s deadliest bacterial pathogens. Since the first ISPPD was held in 1998, substantial progress has been made to control pneumococcal disease, for instance, more than half of surviving infants (78.6 million) from 143 countries now have access to the life-saving pneumococcal conjugate vaccine (PCV). The 11th ISPPD (ISPPD-11) was held in Melbourne, Australia in April 2018 and the proceedings of the symposium are captured in this report.Twenty years on from the first ISPPD, there remain many challenges and unanswered questions such as the continued disparity in disease incidence in Indigenous populations, the slow roll-out of PCV in some regions such as Asia, the persisting burden of disease in adults, serotype replacement and diagnosis of pneumococcal pneumonia. ISPPD-11 also put the spotlight on cutting-edge science including metagenomic, transcriptomic, microscopy, medical imaging and mathematical modelling approaches. ISPPD-11 was highly diverse, bringing together 1184 delegates from 86 countries, representing various fields including academia, primary healthcare, pharmaceuticals, biotechnology, policymakers and public health.

Extracellular polysaccharides and glycoproteins of pathogenic bacteria assist in adherence, autoaggregation, biofilm formation, and host immune system evasion. As a result, considerable research in the field of glycobiology is dedicated to study the composition and function of glycans associated with virulence, as well as the enzymes involved in their biosynthesis with the aim to identify novel antibiotic targets. Especially, insights into the enzyme mechanism, substrate binding, and transition-state structures are valuable as a starting point for rational inhibitor design. An intriguing aspect of enzymes that generate or process polysaccharides and glycoproteins is the level of processivity. The existence of enzymatic processivity reflects the need for regulation of the final glycan/glycoprotein length and structure, depending on the role they perform. In this Review, we describe the currently reported examples of various processive enzymes involved in polymerization and transfer of sugar moieties, predominantly in bacterial pathogens, with a focus on the biochemical methods, to showcase the importance of studying processivity for understanding the mechanism.

A series of novel fluoroquinolone-Safirinium dye hybrids was synthesized by means of tandem Mannich-electrophilic amination reactions from profluorophoric isoxazolones and antibiotics bearing a secondary amino group at position 7 of the quinoline ring. The obtained fluorescent spiro fused conjugates incorporating quaternary nitrogen atoms were characterized by H-1 NMR, IR, MS, and elemental analysis. All the synthetic analogues (3a-h and 4a-h) were evaluated for their in vitro antimicrobial, bactericidal, and antibiofilm activities against a panel of Gram positive and Gram-negative pathogenic bacteria. The most active Safirinium Q derivatives of lomefloxacin (4d) and ciprofloxacin (4e) exhibited molar-based antibacterial activities comparable to the unmodified drugs and displayed considerable inhibitory potencies in E. coli DNA gyrase supercoiling assays with IC50 values in the low micromolar range. Zwiterionic hybrids were noticeably less lipophilic than the parent quinolones in micellar electrokinetic chromatography (MECK) experiments. The tests performed in the presence of phenylalanine-arginine-beta-naphthylamide (PA beta N) or carbonyl cyanide m-chlorophenylhydrazone (CCCP) revealed that the conjugates are to some extent subject to bacterial efflux and cellular accumulation, respectively. Moreover, the hybrids did not exhibit notable cytotoxicity towards the HEK 293 control cell line and demonstrated low propensity for resistance development, as exemplified for compounds 3g and 4b. Finally, molecular docking experiments revealed that the synthesized compounds were able to bind in the fluoroquinolone-binding mode at S. aureus DNA gyrase and S. pneumoniae topoisomerase IV active sites. (C) 2019 Elsevier Masson SAS. All rights reserved.

Introduction: The incidence of human immunodeficiency virus-1 (HIV-1) associated meningitis has been declining in the post-combination antiretroviral treatment (ART) era, although survival rates remain low for the common causes like tuberculosis and cryptococcal disease. Diagnosis and treatment of meningitis in HIV-1 is complicated by atypical clinical presentations, limited accuracy of diagnostic tests, access to diagnostic tests, and therapeutic agents in low- and middle-income countries (LMIC) and immune reconstitution inflammatory syndrome (IRIS). Areas covered: We provide an overview of the common etiologies of meningitis in HIV-1-infected adults, suggest a diagnostic approach based on readily available tests, and review specific chemotherapeutic agents, host-directed therapies, supportive care, timing of ART initiation, and considerations in the management of IRIS with a focus on resource-limited settings. They identify key knowledge gaps and suggest areas for future research. Expert opinion: Evidence-based management of HIV-1-associated meningitis is sparse for common etiologies. More readily available and sensitive diagnostic tests as well as standardized investigation strategies are required in LMIC. There is a lack of availability of recommended drugs in areas of high HIV-1 prevalence and a limited pipeline of novel chemotherapeutic agents. Host-directed therapies have been inadequately studied.

Staphylococcus aureusis a pathogen commonly found in nosocomial environments where infections can easily spread - especially given the reduced immune response of patients and large overlap between personnel in charge of their care. Although antibiotics are available to treat nosocomial infections, the increased occurrence of antibiotic resistance has rendered many treatments ineffective. Such is the case for methicillin resistantS. aureus(MRSA), which has continued to be a threat to public health since its emergence. For this reason, alternative treatment technologies utilizing antimicrobials such as bacteriocins, bacteriophages (phages) and phage endolysins are being developed. These antimicrobials provide an advantage over antibiotics in that many have narrow inhibition spectra, enabling treatments to be selected based on the target (pathogenic) bacterium while allowing for survival of commensal bacteria and thus avoiding collateral damage to the microbiome. Bacterial resistance to these treatments occurs less frequently than with antibiotics, particularly in circumstances where combinatory antimicrobial therapies are used. Phage therapy has been well established in Eastern Europe as an effective treatment against bacterial infections. While there are no Randomized Clinical Trials (RCTs) to our knowledge examining phage treatment ofS. aureusinfections that have completed all trial phases, numerous clinical trials are underway, and several commercial phage preparations are currently available to treatS. aureusinfections. Bacteriocins have primarily been used in the food industry for bio-preservation applications. However, the idea of repurposing bacteriocins for human health is an attractive one considering their efficacy against many bacterial pathogens. There are concerns about the ability of bacteriocins to survive the gastrointestinal tract given their proteinaceous nature, however, this obstacle may be overcome by altering the administration route of the therapy through encapsulation, or by bioengineering protease-resistant variants. Obstacles such as enzymatic digestion are less of an issue for topical/local administration, for example, application to the surface of the skin. Bacteriocins have also shown impressive synergistic effects when used in conjunction with other antimicrobials, including antibiotics, which may allow antibiotic-based therapies to be used more sparingly with less resistance development. This review provides an updated account of known bacteriocins, phages and phage endolysins which have demonstrated an impressive ability to killS. aureusstrains. In particular, examples of antimicrobials with the ability to target MRSA strains and their subsequent use in a clinical setting are outlined.

Altres ajuts: This work also received a grant from the Spanish Society of Pneumology and Thoracic Surgery (SEPAR 024/2016). M.L. gratefully acknowledges funding from the European Respiratory Society Long-term Fellowship grant (LTRF-5934). A.M.E. acknowledges funding from the Royal Society, Department of Medicine (Imperial College), and from the Imperial NIHR Biomedical Research Centre, Imperial College London. B.C.Y is funded by an NIHR Clinical Lectureship. CP acknowledges Programa Germans Trias Sapiens Fundació Catalunya la Pedrera and European Respiratory Society short term research fellowship October 2018 (STRTF201810-00467). Tobacco smoking represents the leading preventable cause of death worldwide. Smoking is a recognised risk factor for several pathologies and is detrimental to host immune surveillance and defence. However, the impact of smoking on microbial residents of the nasopharyngeal cavity, in contact with cigarette smoke (CS), is lacking. Staphylococcus aureus is a major human pathogen that colonises the human nasopharynx and causes a wide range of infections. We investigated the impact of CS on specific virulence phenotypes important in S aureus pathogenesis. We observed strain-dependent differences following exposure to CS, namely growth inhibition, augmented biofilm formation, increased invasion of, and persistence within, bronchial alveolar epithelial cells. Additionally, we confirm the critical role of a functional accessory gene regulator (Agr) system in mediating increased biofilm development and host cell invasion and persistence following CS exposure. Furthermore, CS exposure resulted in reduced toxin production. Importantly, exposure of S aureus to CS accelerated the frequency of mutations and resulted in a significant increase in gentamicin-resistant small colony variant (SCV) formation. Mutational analysis revealed that CS induced SCVs emerge via the SOS response DNA mutagenic repair system. Taken together, our results suggest that CS redirects certain S aureus strains to a virulence profile associated with persistence.

Background Lower respiratory infections are a leading cause of morbidity and mortality around the world. The Global Burden of Diseases, Injuries, and Risk Factors (GBD) Study 2016, provides an up-to-date analysis of the burden of lower respiratory infections in 195 countries. This study assesses cases, deaths, and aetiologies spanning the past 26 years and shows how the burden of lower respiratory infection has changed in people of all ages. Methods We used three separate modelling strategies for lower respiratory infections in GBD 2016: a Bayesian hierarchical ensemble modelling platform (Cause of Death Ensemble model), which uses vital registration, verbal autopsy data, and surveillance system data to predict mortality due to lower respiratory infections; a compartmental meta-regression tool (DisMod-MR), which uses scientific literature, population representative surveys, and healthcare data to predict incidence, prevalence, and mortality; and modelling of counterfactual estimates of the population attributable fraction of lower respiratory infection episodes due to Streptococcus pneumoniae, Haemophilus influenzae type b, influenza, and respiratory syncytial virus. We calculated each modelled estimate for each age, sex, year, and location. We modelled the exposure level in a population for a given risk factor using DisMod-MR and a spatiotemporal Gaussian process regression, and assessed the effectiveness of targeted interventions for each risk factor in children younger than 5 years. We also did a decomposition analysis of the change in LRI deaths from 2000-16 using the risk factors associated with LRI in GBD 2016. Findings In 2016, lower respiratory infections caused 652 572 deaths (95% uncertainty interval [UI] 586 475-720 612) in children younger than 5 years (under-5s), 1 080 958 deaths (943 749-1 170 638) in adults older than 70 years, and 2 377 697 deaths (2145 584-2 512 809) in people of all ages, worldwide. Streptococcus pneumoniae was the leading cause of lower respiratory infection morbidity and mortality globally, contributing to more deaths than all other aetiologies combined in 2016 (1189 937 deaths, 95% UI 690 445-1 770 660). Childhood wasting remains the leading risk factor for lower respiratory infection mortality among children younger than 5 years, responsible for 61.4% of lower respiratory infection deaths in 2016 (95% UI 45.7-69.6). Interventions to improve wasting, household air pollution, ambient particulate matter pollution, and expanded antibiotic use could avert one under-5 death due to lower respiratory infection for every 4000 children treated in the countries with the highest lower respiratory infection burden. Interpretation Our findings show substantial progress in the reduction of lower respiratory infection burden, but this progress has not been equal across locations, has been driven by decreases in several primary risk factors, and might require more effort among elderly adults. By highlighting regions and populations with the highest burden, and the risk factors that could have the greatest effect, funders, policy makers, and programme implementers can more effectively reduce lower respiratory infections among the world's most susceptible populations. Copyright (C) The Author(s). Published by Elsevier Ltd.

Bacterial meningitis (BM) is a severe disease caused by various bacterial pathogens. Toll-like receptors (TLRs) protect humans from invading pathogens. In this study, we determined whether single nucleotide polymorphisms (SNPs) of TLR4 and TLR9 are associated with susceptibility to and outcome of BM in Angolan children. Samples were taken from 241 patients and 265 age-matched ethnic controls. The SNPs TLR4 rs4986790 (896A &gt, G) and TLR9 rs187084 (&minus, 1486T &gt, C) were determined by high-resolution melting analysis (HRMA). The frequency of variant genotypes in TLR4 was significantly higher in patients with Haemophilus influenzae meningitis than controls (odds ratio (OR), 2.5, 95% confidence interval (CI), 1.2&ndash, 5.4, p = 0.021), whereas the frequency of variant genotypes in TLR9 was significantly lower in patients with H. influenzae meningitis than controls (OR, 0.4, 95% CI, 0.2&ndash, 0.9, p = 0.036). No such differences were found with other causative pathogens, such as Streptococcus pneumoniae and Neisseria meningitidis. At the time of discharge, patients with meningitis caused by Gram-negative bacteria who were carriers of variant TLR4 genotypes had a higher risk of ataxia (OR, 12.91, 95% CI, 1.52&ndash, 109.80, p = 0.019) and other neurological sequelae (OR, 11.85, 95% CI, 1.07&ndash, 131.49, p = 0.044) than those with the wild-type TLR4 genotype. Our study suggests an association between H. influenzae meningitis and genetic variation between TLR4 and TLR9 in Angolan children.

Background: Evidence on costs and health benefits of pneumococcal conjugate vaccine (PCV) for children in Asian countries is limited but growing. As a region with a considerably high burden of pneumococcal disease, it is prominent to have a comprehensive overview on the cost-effectiveness of implementing and adopting a PCV vaccination program. Methods: We conducted a systematic review from Pubmed and Embase to identify economic evaluation studies of PCV for children in Asian countries up to May 2020. Data extraction included specific characteristics of the study, input parameters, cost elements, cost-effectiveness results, and key drivers of uncertainty. The Preferred Reporting Items for Systematic Reviews and Meta Analyses (PRISMA) statement was followed for this systematic review. The reporting quality of the included studies was evaluated using the Consolidated Health Economic Evaluation Reporting Standards (CHEERS) statement. Results: After the screening process on both the title and abstract and full text of 518 records, a total of 25 studies fulfilled the inclusion criteria, and were included in the review. The majority of included studies demonstrates that PCV for children is cost-effective in most of the Asian region, and even cost-saving in some countries. Most of the included studies implemented cost utility analysis (CUA) using either quality-adjusted life years (QALYs) or disability-adjusted life years (DALYs). Overall, the main drivers affecting the cost effectiveness were vaccine price, burden regarding pneumonia-related parameters, and the inclusion of herd effects. Conclusion: The children pneumococcal vaccination program appears to be a cost-effective intervention in Asia, and even cost-saving in certain conditions. Vaccine price, pneumonia-related disease burden, and the inclusion of the herd effect are observed as important key drivers in estimating cost-effectiveness in this region. Incorporating PCV in vaccination programs in this region was found to be highly favorable.

Children in Angola are affected by a high burden of disease caused by pneumococcal infections. The 13-valent pneumococcal conjugate vaccine (PCV13) was introduced in the childhood immunization programme in 2013 but the serotype distribution of Streptococcus pneumoniae and antimicrobial susceptibility patterns are unknown. We did a cross-sectional nasopharyngeal carriage study in Luanda and Saurimo, Angola (PCV13 3rd dose coverage 67% and 84%, respectively) during November to December 2017 comprising 940 children aged 4-12 years. The main objective was to assess vaccine serotype coverage and antimicrobial susceptibility rates for S. pneumoniae. Our secondary aim was to characterize colonizinig strains of Haemophilus influenzae and Moraxella catarrhalis. Pneumococcal colonization was found in 35% (95% CI 32-39%) of children (n = 332), with 41% of serotypes covered by PCV13. The most common serotypes were 3 (8%), 18C (6%), 23F (6%), 11A (6%), 34 (6%), 19F (5%) and 16 (5%). Carriage of H. influenzae and M. catarrhalis was detected in 13% (95% CI 11-15%) and 15% (95% CI 13-17%) of children, respectively. Non-susceptibility to penicillin was common among pneumococci (40%), particularly among PCV13-included serotypes (50% vs. 33%; p = 0.003), although the median minimal inhibitory concentration was low (0.19 mg/mL, IQR 0.13-0.25 mg/mL). Most pneumococci and H. influenzae were susceptible to amoxicillin (99% and 88%, respectively). Furthermore, resistance to trimethoprim-sulfamethoxazole was>70% among all three species. Multidrug-resistant pneumococci (non-susceptible to > 3 antibiotics; 7% [n = 24]) were further studied with whole genome sequencing to investigate clonality as an underlying cause for this phenotype. No clearly dominating clone(s) were, however, detected. The results indicate that continued use of PCV13 may have positive direct and herd effects on pneumococcal infections in Angola as carriage of vaccine serotypes was common in the non-vaccinated age group. Finally, amoxicillin is assessed to be a feasible empirical treatment of respiratory tract infections in Angola. (C) 2020 The Author(s). Published by Elsevier Ltd.

Macrophages reside in the body cavities where they maintain serosal homeostasis and provide immune surveillance. Peritoneal macrophages are implicated in the etiology of pathologies including peritonitis, endometriosis and metastatic cancer thus understanding the factors that govern their behavior is vital. Using a combination of fate mapping techniques, we have investigated the impact of sex and age on murine peritoneal macrophage differentiation, turnover and function. We demonstrate that the sexually dimorphic replenishment of peritoneal macrophages from the bone marrow, which is high in males and very low in females, is driven by changes in the local microenvironment that arise upon sexual maturation. Population and single cell RNAseq revealed striking dimorphisms in gene expression between male and female peritoneal macrophages that was in part explained by differences in composition of these populations. By estimating the time of residency of different subsets within the cavity and assessing development of dimorphisms with age and in monocytopenic Ccr2–/– mice, we demonstrate that key sex-dependent features of peritoneal macrophages are a function of the differential rate of replenishment from the bone marrow while others are reliant on local microenvironment signals. Importantly, we demonstrate that the dimorphic turnover of peritoneal macrophages contributes to differences in the ability to protect against pneumococcal peritonitis between the sexes. These data highlight the importance of considering both sex and age in susceptibility to inflammatory and infectious diseases.

Streptococcus pneumoniae can be divided into many strains, each a distinct set of isolates sharing similar core and accessory genomes, which co-circulate within the same hosts. Previous analyses suggested the short-term vaccine-associated dynamics of S. pneumoniae strains may be mediated through multi-locus negative frequency-dependent selection (NFDS), which maintains accessory loci at equilibrium frequencies. Long-term simulations demonstrated NFDS stabilised clonally-evolving multi-strain populations through preventing the loss of variation through drift, based on polymorphism frequencies, pairwise genetic distances and phylogenies. However, allowing symmetrical recombination between isolates evolving under multi-locus NFDS generated unstructured populations of diverse genotypes. Replication of the observed data improved when multi-locus NFDS was combined with recombination that was instead asymmetrical, favouring deletion of accessory loci over insertion. This combination separated populations into strains through outbreeding depression, resulting from recombinants with reduced accessory genomes having lower fitness than their parental genotypes. Although simplistic modelling of recombination likely limited these simulations’ ability to maintain some properties of genomic data as accurately as those lacking recombination, the combination of asymmetrical recombination and multi-locus NFDS could restore multi-strain population structures from randomised initial populations. As many bacteria inhibit insertions into their chromosomes, this combination may commonly underlie the co-existence of strains within a niche.

As one of Gavi, the Vaccine Alliance (previously the Global Alliance for Vaccines and Immunization), graduating countries, Indonesia is still eligible to access Gavi price for PCV13, PCV10 A and B. This study aims to estimate the economic impact of switch from the existing product/presentation of PCV (single-dose of PCV13) to the new product/presentation of PCV (multi-dose of PCV13, PCV10 A and B) since PCV is one of the most expensive vaccines in the Expanded Program on Immunization (EPI) schedule. Assuming that Gavi-Advance Market Commitment (AMC) price for all PCVs can be accessed in 2021, the use of multi-dose PCV13, PCV10 A and PCV10 B with Gavi-AMC price in 2021-2024 were considered as respective scenarios. The result showed that the scenario assuming the use of single-dose of PCV13 with contract price in 2019-2020 that would be switched into multi-dose of PCV10 B with Gavi-AMC price in 2021-2024 resulted in the highest potential saving, compared with other scenarios. Our analysis suggests an economic advantage to switch from single-dose into a multi-dose presentation. Vaccination coverage, vaccine price, vaccine wastage and additional Gavi-AMC vaccine costs were considered to be the most influential parameter affecting the savings in all scenarios. Applying the effectiveness of PCV13 and PCV10 A on reducing the risk for invasive pneumococcal disease (IPD), potential averted incidence of IPD in children under one year of age during 2019-2024 would be 246,164 and 105,587 in both scenarios. Despite the result confirmed that PCV13 may provide an additional benefit, a more comprehensive economic evaluation study is required to investigate further the comparison of cost-effectiveness values among all PCVs in Indonesia.

As one of Gavi, the Vaccine Alliance (previously the Global Alliance for Vaccines and Immunization), graduating countries, Indonesia is still eligible to access Gavi price for PCV13, PCV10 A and B. This study aims to estimate the economic impact of switch from the existing product/presentation of PCV (single-dose of PCV13) to the new product/presentation of PCV (multi-dose of PCV13, PCV10 A and B) since PCV is one of the most expensive vaccines in the Expanded Program on Immunization (EPI) schedule. Assuming that Gavi-Advance Market Commitment (AMC) price for all PCVs can be accessed in 2021, the use of multi-dose PCV13, PCV10 A and PCV10 B with Gavi-AMC price in 2021–2024 were considered as respective scenarios. The result showed that the scenario assuming the use of single-dose of PCV13 with contract price in 2019–2020 that would be switched into multi-dose of PCV10 B with Gavi-AMC price in 2021–2024 resulted in the highest potential saving, compared with other scenarios. Our analysis suggests an economic advantage to switch from single-dose into a multi-dose presentation. Vaccination coverage, vaccine price, vaccine wastage and additional Gavi-AMC vaccine costs were considered to be the most influential parameter affecting the savings in all scenarios. Applying the effectiveness of PCV13 and PCV10 A on reducing the risk for invasive pneumococcal disease (IPD), potential averted incidence of IPD in children under one year of age during 2019–2024 would be 246,164 and 105,587 in both scenarios. Despite the result confirmed that PCV13 may provide an additional benefit, a more comprehensive economic evaluation study is required to investigate further the comparison of cost-effectiveness values among all PCVs in Indonesia.

Background Most patients presenting with acute exacerbations of chronic obstructive pulmonary disease (AECOPD) in primary care are prescribed antibiotics, but these may not be beneficial, and they can cause side effects and increase the risk of subsequent resistant infections. Point-of-care tests (POCTs) could safely reduce inappropriate antibiotic prescribing and antimicrobial resistance. Objective To determine whether or not the use of a C-reactive protein (CRP) POCT to guide prescribing decisions for AECOPD reduces antibiotic consumption without having a negative impact on chronic obstructive pulmonary disease (COPD) health status and is cost-effective. Design A multicentre, parallel-arm, randomised controlled open trial with an embedded process, and a health economic evaluation. Setting General practices in Wales and England. A UK NHS perspective was used for the economic analysis. Participants Adults (aged ≥ 40 years) with a primary care diagnosis of COPD, presenting with an AECOPD (with at least one of increased dyspnoea, increased sputum volume and increased sputum purulence) of between 24 hours’ and 21 days’ duration. Intervention CRP POCTs to guide antibiotic prescribing decisions for AECOPD, compared with usual care (no CRP POCT), using remote online randomisation. Main outcome measures Patient-reported antibiotic consumption for AECOPD within 4 weeks post randomisation and COPD health status as measured with the Clinical COPD Questionnaire (CCQ) at 2 weeks. For the economic evaluation, patient-reported resource use and the EuroQol-5 Dimensions were included. Results In total, 653 participants were randomised from 86 general practices. Three withdrew consent and one was randomised in error, leaving 324 participants in the usual-care arm and 325 participants in the CRP POCT arm. Antibiotics were consumed for AECOPD by 212 out of 274 participants (77.4%) and 150 out of 263 participants (57.0%) in the usual-care and CRP POCT arm, respectively [adjusted odds ratio 0.31, 95% confidence interval (CI) 0.20 to 0.47]. The CCQ analysis comprised 282 and 281 participants in the usual-care and CRP POCT arms, respectively, and the adjusted mean CCQ score difference at 2 weeks was 0.19 points (two-sided 90% CI –0.33 to –0.05 points). The upper limit of the CI did not contain the prespecified non-inferiority margin of 0.3. The total cost from a NHS perspective at 4 weeks was £17.59 per patient higher in the CRP POCT arm (95% CI –£34.80 to £69.98; p = 0.408). The mean incremental cost-effectiveness ratios were £222 per 1% reduction in antibiotic consumption compared with usual care at 4 weeks and £15,251 per quality-adjusted life-year gained at 6 months with no significant changes in sensitivity analyses. Patients and clinicians were generally supportive of including CRP POCT in the assessment of AECOPD. Conclusions A CRP POCT diagnostic strategy achieved meaningful reductions in patient-reported antibiotic consumption without impairing COPD health status or increasing costs. There were no associated harms and both patients and clinicians valued the diagnostic strategy. Future work Implementation studies that also build on our qualitative findings could help determine the effect of this intervention over the longer term. Trial registration Current Controlled Trials ISRCTN24346473. Funding This project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 24, No. 15. See the NIHR Journals Library website for further project information.

Neutrophils are key effector cells of innate immunity, rapidly recruited to defend the host against invading pathogens. Neutrophils may kill pathogens intracellularly, following phagocytosis, or extracellularly, by degranulation and the release of neutrophil extracellular traps; all of these microbicidal strategies require the deployment of cytotoxic proteins and proteases, packaged during neutrophil development within cytoplasmic granules. Neutrophils operate in infected and inflamed tissues, which can be profoundly hypoxic. Neutrophilic infiltration of hypoxic tissues characterises a myriad of acute and chronic infectious and inflammatory diseases, and as well as potentially protecting the host from pathogens, neutrophil granule products have been implicated in causing collateral tissue damage in these scenarios. This review discusses the evidence for the enhanced secretion of destructive neutrophil granule contents observed in hypoxic environments and the potential mechanisms for this heightened granule exocytosis, highlighting implications for the host. Understanding the dichotomy of the beneficial and detrimental consequences of neutrophil degranulation in hypoxic environments is crucial to inform potential neutrophil-directed therapeutics in order to limit persistent, excessive, or inappropriate inflammation.

Biofilms are extremely difficult to eradicate due to their decreased antibiotic susceptibility. Inducing biofilm dispersion could be a potential strategy to help combat biofilm-related infections. Mechanisms of biofilm dispersion can basically be divided into two groups, i.e. active and passive dispersion. Active dispersion depends on a decrease in the intracellular c-di-GMP levels, leading to the production of enzymes that degrade the biofilm matrix and promote dispersion. In contrast, passive dispersion relies on triggers that directly release cells from the biofilm. In the present review, several active and passive dispersion strategies are discussed. In addition, the disadvantages and possible consequences of using dispersion as a treatment approach for biofilm-related infections are also reviewed.

Background The current carriage study was set up to reinforce surveillance during/after the PCV13-to-PCVC10 switch in Belgium. Aim This 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. Methods A 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). Results The 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%). Conclusions During 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.

Aim: The purpose of this study was to determine the prevalence of nasopharyngeal carriage of potentially pathogenic bacteria (especially Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis) in healthy 1-7 years old children and risk factors affecting it. Methods: A total of four hundred forty-five healthy children (1-7 years old) from the European side of Istanbul (10 different units) were included in this cross-sectional study. Isolated microorganisms were identified by standard laboratory methods and the results were evaluated. Risk factors affecting the prevalence of nasopharyngeal carriage were also evaluated. Results: In the study, 139 (31.2%) of the samples were positive. S. pneumoniae two (0.4%), H. influenzae 50 (11.2%); of them type b 10 (2.2%), H. parainfluenzae 10 (2.2%), M. catarrhalis 14 (3.1%), S. aureus 35 (7.9%), Group A β hemolytic streptococci (Streptococcus pyogenes) 13 (2.9%), Group B streptococci (S. agalactiae) 18 (4.0%), Non A Non B streptococci 16 (3.6%) were detected. The results obtained from different sites (places) were found to be variable in terms of pathogen density. Conclusion: Nasopharyngeal carriage rates found in our study were generally lower than in some studies on this subject. However, when the samples taken from different places were evaluated one by one, it was seen that the colonization rate reached 45.9% in crowded and poor building properties (small, poorly ventilated, unhealthy buildings). The highest rate of H. influenzae carriage in the 2-3 age group was 33.3%. In this study, crowded and unhealthy school environments and low maternal education level have been determined as risk factors for increased nasopharyngeal carriage rate. Results of such studies vary depending on the region, sample frequency, individual and social factors. Further studies are needed in order to achieve healthier results. Amaç: Bu çalışmada sağlıklı 1-7 yaş çocuklarda (Streptococcuspneumoniae, Haemophilus influenzae, Moraxella catarr halis başta olmak üzere) potansiyel patojen bakterilerin nazofaringeal taşıyıcılık prevalansının ve bunu etkileyen risk faktörlerinin belirlenmesi amaçlanmıştır. Yöntemler: Bu kesitsel çalışmaya İstanbul Avrupa yakasından(10 farklı birimden) 1-7 yaş grubu toplam dört yüz kırk beş sağlıklı çocuk dahil edildi. İzole edilen mikroorganizmalar standart laboratuvar yöntemleri ile tanımlanarak elde edilen sonuçlar değerlendirildi. Ayrıca, nazofaringeal taşıyıcılık prevalansını etkileyen risk faktörleri değerlendirildi.Bulgular: Çalışılan örneklerin 139’u (%31,2) pozitifti. S.pneumoniae iki (%0,4), H. influenzae 50 (%11,2); bunlardantip b 10 (%2,2), Haemophilus parainfluenzae 10 (%2,2), M.catarrhalis 14 (%3,1), S. aureus 35 (%7,9), A grubu β hemolitikstreptokok (Streptococcus pyogenes) 13 (%2,9), B Grubustreptokok (S. agalactiae) 18 (%4,0), Non A Non B streptokok16 (%3,6) oranlarında tespit edildi. Farklı yerlerden elde edilen sonuçlarda, patojen yoğunluğunun ve çeşitliliğinin değişken olduğu görüldü. Sonuç: Çalışmamızda tespit edilen nazofaringeal taşıyıcılıkoranları bu konuda yapılmış bazı çalışmalara göre düşük bulundu. Ancak örnek alınan farklı kurumlar tek tek değerlendirildiğinde mevcudu fazla ve bina özellikleri kötü olan yerlerde (kalabalık, küçük, havalandırılması iyi yapılamayan, sağlıksız binalar) oranın%45,9’a kadar ulaştığı görüldü. H. influenzae taşıyıcılığında ise2-3 yaş grubunda en yüksek oranı %33,3 olarak tespit edildi. Bu tür çalışmaların sonuçları bölgeye, örnekleme sıklığına, bireysel ve sosyal faktörlere göre değişiklik göstermektedir. Daha sağlıklı sonuçlara ulaşmak için daha fazla çalışmaya ihtiyaç vardır. Firat University Scientific Research Projects Management Unit

Introduction Pneumococcal conjugate vaccines (PCVs) have been available in Canada since 2001, with 13-valent PCV (PCV13) added to the infant routine immunization program throughout all Canadian provinces by 2011. The use of PCVs has dramatically reduced the burden of pneumococcal disease in Canada. As a result, decision-makers may consider switching from a more costly, higher-valent vaccine to a lower-cost, lower-valent vaccine in an attempt to allocate funds for other vaccine programs. We assessed the health and economic impact of switching the infant vaccination program from PCV13 to 10-valent PCV (PCV10) in the context of the Canadian health care system. Methods We performed a review of Canadian databases supplemented with published and unpublished data to obtain the historical incidence of pneumococcal disease and direct and indirect medical costs. Observed invasive pneumococcal disease (IPD) trends from surveillance data were used as a basis to forecast the future number of cases of IPD, pneumococcal pneumonia, and acute otitis media given a PCV13- or PCV10-based program. Costs and outcomes over 10 years were then estimated and presented in 2017 Canadian dollars discounted at 3% per year. Results Switching from PCV13 to PCV10 would result in an additional 762,531 cases of pneumococcal disease over 10 years. Although PCV13 has a higher acquisition cost, switching to PCV10 would increase overall costs by over $500 million. Forecasted overall disease incidence was estimated substantially higher with PCV10 than with PCV13 primarily because of the potential reemergence of serotypes 3 and 19A. PCV13 was also cost saving compared with PCV10, even within a 5-year time horizon. Probabilistic sensitivity analysis showed that a PCV13-based program remained cost saving in all simulations. Conclusion Although switching to a PCV10-based infant vaccination program in Canada might result in lower acquisition costs, it would also result in higher public health cost and burden because of serotype reemergence. Funding Pfizer Inc. Electronic supplementary material The online version of this article (10.1007/s40121-018-0206-1) contains supplementary material, which is available to authorized users.

Streptococcus pneumoniae (the pneumococcus) is a common nasopharyngeal commensal that can cause invasive pneumococcal disease (IPD). Each component of current protein–polysaccharide conjugate vaccines (PCVs) generally induces immunity specific to one of the approximately 100 pneumococcal serotypes, and typically eliminates it from carriage and IPD through herd immunity. Overall carriage rates remain stable owing to replacement by non-PCV serotypes. Consequently, the net change in IPD incidence is determined by the relative invasiveness of the pre- and post-PCV-carried pneumococcal populations. In the present study, we identified PCVs expected to minimize the post-vaccine IPD burden by applying Bayesian optimization to an ecological model of serotype replacement that integrated epidemiological and genomic data. We compared optimal formulations for reducing infant-only or population-wide IPD, and identified potential benefits to including non-conserved pneumococcal carrier proteins. Vaccines were also devised to minimize IPD resistant to antibiotic treatment, despite the ecological model assuming that resistance levels in the carried population would be preserved. We found that expanding infant-administered PCV valency is likely to result in diminishing returns, and that complementary pairs of infant- and adult-administered vaccines could be a superior strategy. PCV performance was highly dependent on the circulating pneumococcal population, further highlighting the advantages of a diversity of anti-pneumococcal vaccination strategies. Here the authors have developed a framework to predict which Streptococcus pneumoniae serotypes would be best targeted by vaccination, of children and adults, to prevent the post-vaccine emergence of similarly virulent strains, thereby minimizing the pneumococcal disease burden.

Background The complement system is a vital component of the inflammatory response occurring during bacterial meningitis. Blocking the complement system was shown to improve the outcome of experimental pneumococcal meningitis. Complement factor H (FH) is a complement regulatory protein inhibiting alternative pathway activation but is also exploited by the pneumococcus to prevent complement activation on its surface conferring serum resistance. Methods In a nationwide prospective cohort study of 1009 episodes with community-acquired bacterial meningitis, we analyzed whether genetic variations in CFH influenced FH cerebrospinal fluid levels and/or disease severity. Subsequently, we analyzed the role of FH in our pneumococcal meningitis mouse model using FH knock-out (Cfh−/−) mice and wild-type (wt) mice. Finally, we tested whether adjuvant treatment with human FH (hFH) improved outcome in a randomized investigator blinded trial in a pneumococcal meningitis mouse model. Results We found the major allele (G) of single nucleotide polymorphism in CFH (rs6677604) to be associated with low FH cerebrospinal fluid concentration and increased mortality. In patients and mice with bacterial meningitis, FH concentrations were elevated during disease and Cfh−/− mice with pneumococcal meningitis had increased mortality compared to wild-type mice due to C3 depletion. Adjuvant treatment of wild-type mice with purified human FH led to complement inhibition but also increased bacterial outgrowth which resulted in similar disease outcomes. Conclusion Low FH levels contribute to mortality in pneumococcal meningitis but adjuvant treatment with FH at a clinically relevant time point is not beneficial.

Competition among bacterial members of the same ecological niche is mediated by bacteriocins: antimicrobial peptides produced by bacterial species to kill other bacteria. Bacteriocins are also promising candidates for novel antimicrobials. Streptococcus pneumoniae (the ‘pneumococcus’) is a leading cause of morbidity and mortality worldwide and a frequent coloniser of the human nasopharynx. Here, 14 newly-discovered bacteriocin gene clusters were identified among >6,200 pneumococcal genomes. The molecular epidemiology of the bacteriocin clusters was investigated using a large global and historical pneumococcal dataset dating from 1916. These analyses revealed extraordinary bacteriocin diversity among pneumococci and the majority of bacteriocin clusters were also found in other streptococcal species. Genomic hotspots for the integration of different bacteriocin gene clusters were discovered. Experimentally, bacteriocin genes were transcriptionally active when the pneumococcus was under stress and when two strains were co-cultured in broth. These findings reveal much more diversity among bacterial defence mechanisms than previously appreciated, which fundamentally broaden our understanding of bacteriocins relative to intraspecies and interspecies nasopharyngeal competition and bacterial population structure.

Increased reactive oxygen species (ROS) have been implicated in the pathophysiology of chronic obstructive pulmonary disease (COPD). This study examined the effect of exogenous and endogenous oxidative stress on macrophage phagocytosis in patients with COPD.Monocyte-derived macrophages (MDMs) were generated from non-smoker, smoker and COPD subjects, differentiated in either granulocyte macrophage-colony stimulating factor (G-Mφ) or macrophage-colony stimulating factor (M-Mφ). Alveolar macrophages were isolated from lung tissue or bronchoalveolar lavage fluid. Macrophages were incubated in ±200 µM H2O2 for 24 h, then exposed to fluorescently labelled Haemophilus influenzae or Streptococcus pneumoniae for 4 h, after which phagocytosis, mitochondrial ROS (mROS) and mitochondrial membrane potential (ΔΨm) were measured.Phagocytosis of bacteria was significantly decreased in both G-Mφ and M-Mφ from COPD patients compared with from non-smoker controls. In non-smokers and smokers, bacterial phagocytosis did not alter mROS or ΔΨm; however, in COPD, phagocytosis increased early mROS and decreased ΔΨm in both G-Mφ and M-Mφ. Exogenous oxidative stress reduced phagocytosis in non-smoker and COPD alveolar macrophages and non-smoker MDMs, associated with reduced mROS production.COPD macrophages show defective phagocytosis, which is associated with altered mitochondrial function and an inability to regulate mROS production. Targeting mitochondrial dysfunction may restore the phagocytic defect in COPD.

Background: Patients with recurrent respiratory tract infections and an impaired response to pneumococcal polysaccharide vaccination are diagnosed with a specific antibody deficiency. In adult patients with pneumococcal pneumonia an impaired antibody response to the infecting pneumococcal serotype can sometimes be found. It is unknown whether these patients are unable to produce an adequate anti-polysaccharide antibody response to pneumococcal vaccination after recovery.Case presentation: The authors describe a case of invasive pneumonia caused by Streptococcus pneumoniae serotype 9V in a previously healthy 35-year-old female. This patient did not produce serotype-specific antibodies against the infecting serotype during disease. After pneumococcal polysaccharide vaccination 3 months after recovery, she responded adequately to most other pneumococcal serotypes, but still had no response to the infecting serotype 9V. However, after 9 years (and prior to pneumococcal-conjugate vaccination) normal antibody levels against 9V were found. These antibody levels further increased after pneumococcal-conjugate vaccination.Conclusion: The authors believe that this case is the first description of a temporary deficient response to the infecting pneumococcal serotype in adults, while other reports with similar observations all involved children.

Recent advances in the scale and diversity of population genomic datasets for bacteria now provide the potential for genome-wide patterns of co-evolution to be studied at the resolution of individual bases. Here we describe a new statistical method, genomeDCA, which uses recent advances in computational structural biology to identify the polymorphic loci under the strongest co-evolutionary pressures. We apply genomeDCA to two large population data sets representing the major human pathogens Streptococcus pneumoniae (pneumococcus) and Streptococcus pyogenes (group A Streptococcus). For pneumococcus we identified 5,199 putative epistatic interactions between 1,936 sites. Over three-quarters of the links were between sites within the pbp2x, pbp1a and pbp2b genes, the sequences of which are critical in determining non-susceptibility to beta-lactam antibiotics. A network-based analysis found these genes were also coupled to that encoding dihydrofolate reductase, changes to which underlie trimethoprim resistance. Distinct from these antibiotic resistance genes, a large network component of 384 protein coding sequences encompassed many genes critical in basic cellular functions, while another distinct component included genes associated with virulence. The group A Streptococcus (GAS) data set population represents a clonal population with relatively little genetic variation and a high level of linkage disequilibrium across the genome. Despite this, we were able to pinpoint two RNA pseudouridine synthases, which were each strongly linked to a separate set of loci across the chromosome, representing biologically plausible targets of co-selection. The population genomic analysis method applied here identifies statistically significantly co-evolving locus pairs, potentially arising from fitness selection interdependence reflecting underlying protein-protein interactions, or genes whose product activities contribute to the same phenotype. This discovery approach greatly enhances the future potential of epistasis analysis for systems biology, and can complement genome-wide association studies as a means of formulating hypotheses for targeted experimental work., Author summary Epistatic interactions between polymorphisms in DNA are recognized as important drivers of evolution in numerous organisms. Study of epistasis in bacteria has been hampered by the lack of densely sampled population genomic data, suitable statistical models and inference algorithms sufficiently powered for extremely high-dimensional parameter spaces. We introduce the first model-based method for genome-wide epistasis analysis and use two of the largest available bacterial population genome data sets on Streptococcus pneumoniae (the pneumococcus) and Streptococcus pyogenes (group A Streptococcus) to demonstrate its potential for biological discovery. Our approach reveals interacting networks of resistance, virulence and core machinery genes in the pneumococcus, which highlights putative candidates for novel drug targets. We also discover a number of plausible targets of co-selection in S. pyogenes linked to RNA pseudouridine synthases. Our method significantly enhances the future potential of epistasis analysis for systems biology, and can complement genome-wide association studies as a means of formulating hypotheses for targeted experimental work.

Streptococcus pneumoniae is able to cause deadly diseases by infecting different tissues, each with distinct environmental and nutritional compositions. We hypothesize that the adaptive capabilities of the microbe is an important facet of pneumococcal survival in fluctuating host environments. Quorum-sensing (QS) mechanisms are pivotal for microbial host adaptation. We previously demonstrated that the TprA/PhrA QS system is required for pneumococcal utilization of galactose and mannose, neuraminidase activity, and virulence. We also showed that the system can be modulated by using linear molecularly imprinted polymers. Due to being a drugable target, we further studied the operation of this QS system in S. pneumoniae. We found that TprA controls the expression of nine different operons on galactose and mannose. Our data revealed that TprA expression is modulated by a complex regulatory network, where the master regulators CcpA and GInR are involved in a sugar dependent manner. Mutants in the TprA/PhrA system are highly attenuated in their survival in nasopharynx and lungs after intranasal infection, and growth in blood after intravenous infection.

Background Point-of-care testing of C-reactive protein (CRP) may be a way to reduce unnecessary use of antibiotics without harming patients who have acute exacerbations of chronic obstructive pulmonary disease (COPD). Methods We performed a multicenter, open-label, randomized, controlled trial involving patients with a diagnosis of COPD in their primary care clinical record who consulted a clinician at 1 of 86 general medical practices in England and Wales for an acute exacerbation of COPD. The patients were assigned to receive usual care guided by CRP point-of-care testing (CRP-guided group) or usual care alone (usual-care group). The primary outcomes were patient-reported use of antibiotics for acute exacerbations of COPD within 4 weeks after randomization (to show superiority) and COPD-related health status at 2 weeks after randomization, as measured by the Clinical COPD Questionnaire, a 10-item scale with scores ranging from 0 (very good COPD health status) to 6 (extremely poor COPD health status) (to show noninferiority). Results A total of 653 patients underwent randomization. Fewer patients in the CRP-guided group reported antibiotic use than in the usual-care group (57.0% vs. 77.4%; adjusted odds ratio, 0.31; 95% confidence interval [CI], 0.20 to 0.47). The adjusted mean difference in the total score on the Clinical COPD Questionnaire at 2 weeks was −0.19 points (two-sided 90% CI, −0.33 to −0.05) in favor of the CRP-guided group. The antibiotic prescribing decisions made by clinicians at the initial consultation were ascertained for all but 1 patient, and antibiotic prescriptions issued over the first 4 weeks of follow-up were ascertained for 96.9% of the patients. A lower percentage of patients in the CRP-guided group than in the usual-care group received an antibiotic prescription at the initial consultation (47.7% vs. 69.7%, for a difference of 22.0 percentage points; adjusted odds ratio, 0.31; 95% CI, 0.21 to 0.45) and during the first 4 weeks of follow-up (59.1% vs. 79.7%, for a difference of 20.6 percentage points; adjusted odds ratio, 0.30; 95% CI, 0.20 to 0.46). Two patients in the usual-care group died within 4 weeks after randomization from causes considered by the investigators to be unrelated to trial participation. Conclusions CRP-guided prescribing of antibiotics for exacerbations of COPD in primary care clinics resulted in a lower percentage of patients who reported antibiotic use and who received antibiotic prescriptions from clinicians, with no evidence of harm. (Funded by the National Institute for Health Research Health Technology Assessment Program; PACE Current Controlled Trials number, ISRCTN24346473. opens in new tab.)

The most frequent form of hemolytic-uremic syndrome (HUS) is associated with infections caused by Shiga-like toxin-producing Enterohaemorrhagic Escherichia coli (STEC). In rarer cases HUS can be triggered by Streptococcus pneumoniae. While production of Shiga-like toxins explains STEC-HUS, the mechanisms of pneumococcal HUS are less well known. S. pneumoniae produces neuraminidases with activity against cell surface sialic acids that are critical for factor H-mediated complement regulation on cells and platelets. The aim of this study was to find out whether S. pneumoniae neuraminidase NanA could trigger complement activation and hemolysis in whole blood. We studied clinical S. pneumoniae isolates and two laboratory strains, a wild-type strain expressing NanA, and a NanA deletion mutant for their ability to remove sialic acids from various human cells and platelets. Red blood cell lysis and activation of complement was measured ex vivo by incubating whole blood with bacterial culture supernatants. We show here that NanA expressing S. pneumoniae strains and isolates are able to remove sialic acids from cells, and platelets. Removal of sialic acids by NanA increased complement activity in whole blood, while absence of NanA blocked complement triggering and hemolytic activity indicating that removal of sialic acids by NanA could potentially trigger pHUS.

Millions of children are exposed to tuberculosis (TB) each year, many of which become infected with Mycobacterium tuberculosis. Most children can immunologically contain or eradicate the organism without pathology developing. However, in a minority, the organism overcomes the immunological constraints, proliferates and causes TB disease. Each year a million children develop TB disease, with a quarter dying. While it is known that young children and those with immunodeficiencies are at increased risk of progression from TB infection to TB disease, our understanding of risk factors for this transition is limited. The most immunologically disruptive process that can happen during childhood is infection with another pathogen and yet the impact of co-infections on TB risk is poorly investigated. Many diseases have overlapping geographical distributions to TB and affect similar patient populations. It is therefore likely that infection with viruses, bacteria, fungi and protozoa may impact on the risk of developing TB disease following exposure and infection, although disentangling correlation and causation is challenging. As vaccinations also disrupt immunological pathways, these may also impact on TB risk. In this article we describe the paediatric immune response to M. tuberculosis and then review the existing evidence of the impact of co-infection with other pathogens, as well as vaccination, on the host response to M. tuberculosis. We focus on the impact of other organisms on the risk of TB disease in children, in particularly evaluating if co-infections drive host immune responses in an age-dependent way. The pathology seen in young children is primarily the result of an insufficient and inadequate response, while the pathology seen in adolescents and adults reflects an over-exuberant response, leading to host tissue damage. We will explore if co-infections drive the host response in these directions. We finally propose priorities for future research in this field. An improved understanding of the impact of co-infections on TB could assist in TB control strategies, vaccine development (for TB vaccines or vaccines for other organisms), TB treatment approaches and TB diagnostics.

BACKGROUND: Prior to introducing pneumococcal conjugate vaccines (PCVs), Streptococcus pneumoniae was most commonly isolated from middle ear fluid of children with acute otitis media (AOM). Reducing nasopharyngeal colonisation of this bacterium by PCVs may lead to a decline in AOM. The effects of PCVs deserve ongoing monitoring since studies from the post-PCV era report a shift in causative otopathogens towards non-vaccine serotypes and other bacteria. This updated Cochrane Review was first published in 2002 and updated in 2004, 2009, and 2014. The review title was changed (to include the population, i.e. children) for this update.OBJECTIVES: To assess the effect of PCVs in preventing AOM in children up to 12 years of age.SEARCH METHODS: We searched CENTRAL, MEDLINE, Embase, CINAHL, LILACS, Web of Science, and trials registers (ClinicalTrials.gov and WHO ICTRP) to 29 March 2019.SELECTION CRITERIA: Randomised controlled trials of PCV versus placebo or control vaccine.DATA COLLECTION AND ANALYSIS: We used the standard methodological procedures expected by Cochrane. The primary outcomes were frequency of all-cause AOM and adverse effects. Secondary outcomes included frequency of pneumococcal AOM and frequency of recurrent AOM (defined as three or more AOM episodes in six months or four or more in one year). We used GRADE to assess the quality of the evidence.MAIN RESULTS: We included 14 publications of 11 trials (60,733 children, range 74 to 37,868 per trial) of 7- to 11-valent PCVs versus control vaccines (meningococcus type C vaccine in three trials, and hepatitis A or B vaccine in eight trials). We included two additional trials for this update. We did not find any relevant trials with the newer 13-valent PCV. Most studies were funded by pharmaceutical companies. Overall, risk of bias was low. In seven trials (59,415 children) PCVs were administered in early infancy, while four trials (1318 children) included children aged one year and over who were either healthy or had a history of respiratory illness. There was considerable clinical heterogeneity across studies, therefore we did not perform meta-analyses.Adverse eventsNine trials reported on adverse effects (77,389 children; high-quality evidence). Mild local reactions and fever were common in both groups, and occurred more frequently in PCV than in control vaccine groups: redness (< 2.5 cm): 5% to 20% versus 0% to 16%; swelling (< 2.5 cm): 5% to 12% versus 0% to 8%; and fever (< 39 °C): 15% to 44% versus 8% to 25%. More severe redness (> 2.5 cm), swelling (> 2.5 cm), and fever (> 39 °C) occurred less frequently (0% to 0.9%, 0.1% to 1.3%, and 0.4% to 2.5%, respectively in children receiving PCV) and did not differ significantly between PCV and control vaccine groups. Pain or tenderness, or both was reported more frequently in PCV than in control vaccine groups: 3% to 38% versus 0% to 8%. Serious adverse events judged causally related to vaccination were rare and did not differ significantly between groups, and no fatal serious adverse event judged causally related to vaccination was reported.PCV administered in early infancyPCV7The effect of a licenced 7-valent PCV with CRM197 as carrier protein (CRM197-PCV7) on all-cause AOM varied from -5% (95% confidence interval (CI) -25% to 12%) relative risk reduction (RRR) in high-risk infants (1 trial; 944 children; moderate-quality evidence) to 6% (95% CI -4% to 16%; 1 trial; 1662 children) and 6% (95% CI 4% to 9%; 1 trial; 37,868 children) RRR in low-risk infants (high-quality evidence). PCV7 with the outer membrane protein complex of Neisseria meningitidis serogroup B as carrier protein (OMPC-PCV7), was not associated with a reduction in all-cause AOM (RRR -1%, 95% CI -12% to 10%; 1 trial; 1666 children; high-quality evidence).CRM197-PCV7 and OMPC-PCV7 were associated with 20% (95% CI 7% to 31%) and 25% (95% CI 11% to 37%) RRR in pneumococcal AOM, respectively (2 trials; 3328 children; high-quality evidence) and CRM197-PCV7 with 9% (95% CI -12% to 27%) to 10% (95% CI 7% to 13%) RRR in recurrent AOM (2 trials; 39,530 children; high-quality evidence).PHiD-CV10/11The effect of a licenced 10-valent PCV conjugated to protein D, a surface lipoprotein of Haemophilus influenzae, (PHiD-CV10) on all-cause AOM varied from 6% (95% CI -6% to 17%; 1 trial; 5095 children) to 15% (95% CI -1% to 28%; 1 trial; 7359 children) RRR in healthy infants (moderate-quality evidence). PHiD-CV11 was associated with 34% (95% CI 21% to 44%) RRR in all-cause AOM (1 trial; 4968 children; high-quality evidence).PHiD-CV10 and PHiD-CV11 were associated with 53% (95% CI 16% to 74%) and 52% (95% CI 37% to 63%) RRR in pneumococcal AOM (2 trials; 12,327 children; high-quality evidence) and PHiD-CV11 with 56% (95% CI -2% to 80%) RRR in recurrent AOM (1 trial; 4968 children; moderate-quality evidence).PCV administered at later agePCV7We found no evidence of a beneficial effect on all-cause AOM of administering CRM197-PCV7 in children aged 1 to 7 years with a history of respiratory illness or frequent AOM (2 trials; 457 children; high-quality evidence) and CRM197-PCV7 combined with a trivalent influenza vaccine in children aged 18 to 72 months with a history of respiratory tract infections (1 trial; 597 children; high-quality evidence).CRM197-PCV9In 1 trial including 264 healthy day-care attendees aged 1 to 3 years, CRM197-PCV9 was associated with 17% (95% CI -2% to 33%) RRR in parent-reported all-cause OM (low-quality evidence).AUTHORS' CONCLUSIONS: Administration of the licenced CRM197-PCV7 and PHiD-CV10 during early infancy is associated with large relative risk reductions in pneumococcal AOM. However, the effects of these vaccines on all-cause AOM is far more uncertain. We found no evidence of a beneficial effect on all-cause AOM of administering PCVs in high-risk infants, after early infancy (i.e. in children one year and above), and in older children with a history of respiratory illness. Compared to control vaccines, PCVs were associated with an increase in mild local reactions (redness, swelling), fever, and pain and/or tenderness. We found no evidence of a difference in more severe local reactions, fever, or serious adverse events judged causally related to vaccination.

Background: Pneumococcal conjugate vaccines have reduced the incidence of invasive pneumococcal disease, caused by vaccine serotypes, but non-vaccine-serotypes remain a concern. We used whole genome sequencing to study pneumococcal serotype, antibiotic resistance and invasiveness, in the context of genetic background. Methods: Our dataset of 13,454 genomes, combined with four published genomic datasets, represented Africa (40%), Asia (25%), Europe (19%), North America (12%), and South America (5%). These 20,027 pneumococcal genomes were clustered into lineages using PopPUNK, and named Global Pneumococcal Sequence Clusters (GPSCs). From our dataset, we additionally derived serotype and sequence type, and predicted antibiotic sensitivity. We then measured invasiveness using odds ratios, relating prevalence in disease to carriage. Findings: The combined collections (n=20,027) were clustered into 621 GPSCs. Thirty-five GPSCs observed in our dataset were represented by >100 isolates, and subsequently classed as dominant-GPSCs. In 22/35 (63%) of dominant-GPSCs both non-vaccine serotypes and vaccine serotypes were observed pre-PCV. Penicillin and multidrug resistance were higher (p

The standard workhorse for genomic analysis of the evolution of bacterial populations is phylogenetic modelling of mutations in the core genome. However, a notable amount of information about evolutionary and transmission processes in diverse populations can be lost unless the accessory genome is also taken into consideration. Here, we introduce PANINI (Pangenome Neighbour Identification for Bacterial Populations), a computationally scalable method for identifying the neighbours for each isolate in a data set using unsupervised machine learning with stochastic neighbour embedding based on the t-SNE (t-distributed stochastic neighbour embedding) algorithm. PANINI is browser-based and integrates with the Microreact platform for rapid online visualization and exploration of both core and accessory genome evolutionary signals, together with relevant epidemiological, geographical, temporal and other metadata. Several case studies with single- and multi-clone pneumococcal populations are presented to demonstrate the ability to identify biologically important signals from gene content data. PANINI is available at http://panini.pathogen.watch and code at http://gitlab.com/cgps/panini.

Background There is a scarcity of data on pneumococcal serotypes carried by children in Ethiopia. We studied pneumococcal nasopharyngeal carriage rate, serotypes, and risk factors among children with community acquired pneumonia (CAP). Methods A prospective observational cohort study was performed in children with CAP, aged 0–15 years, in 2 pediatric emergency departments in Addis Ababa, Ethiopia. Nasopharyngeal swabs were cultured, and serotypes of Streptococcus pneumoniae were determined by sequencing the cpsB gene and by the Quellung reaction. Risk factors were analyzed by using binary logistic regression. Results Nasopharyngeal swabs were collected from 362 children with CAP. Pneumococcal carriage rate was 21.5% (78 of 362). The most common serotypes were 19A (27%), 16F (8.5%), and 6A (4.9%). In addition, 8.5% of the pneumococcal isolates were nontypeable. In bivariate analysis, children with a parent that smokes were more likely to carry pneumococci (crude odds ratio, 3.9; 95% confidence interval [CI], 1.2–12.3; P = .023) than those with parents that do not smoke. In multivariable analysis, living in a house with ≥2 rooms (adjusted odds ratio [AOR], 0.48; 95% CI, 0.28–0.82; P = .007) and vaccination with ≥2 doses of 10-valent pneumococcal conjugate vaccine (PCV10) (AOR, 0.37; 95% CI, 0.15–0.92; P = .033) were protective of pneumococcal carriage. Conclusions Five years after introduction of PCV10 in Ethiopia, the vaccine-related serotype 19A was predominant in the nasopharynx of children with CAP. Continued evaluation of the direct and indirect impact of PCV10 on pneumococcal serotype distribution in Ethiopia is warranted.

The 13-valent pneumococcal conjugate vaccine (PCV-13) was introduced in the United States in 2010. Using a large paediatric carriage sample collected from shortly after the introduction of PCV-7 to several years after the introduction of PCV-13, we investigate alterations in the composition of the pneumococcal population following the introduction of PCV-13, evaluating the extent to which the post-vaccination non-vaccine type (NVT) population mirrors that from prior to vaccine introduction and the effect of PCV-13 on vaccine type lineages. Draft genome assemblies from 736 newly sequenced and 616 previously published pneumococcal carriage isolates from children in Massachusetts between 2001 and 2014 were analysed. Isolates were classified into one of 22 sequence clusters (SCs) on the basis of their core genome sequence. We calculated the SC diversity for each sampling period as the probability that any two randomly drawn isolates from that period belong to different SCs. The sampling period immediately after the introduction of PCV-13 (2011) was found to have higher diversity than preceding (2007) or subsequent (2014) sampling periods {Simpson’s D 2007: 0.915 [95 % confidence interval (CI) 0.901, 0.929]; 2011: 0.935 [0.927, 0.942]; 2014 : 0.912 [0.901, 0.923]}. Amongst NVT isolates, we found the distribution of SCs in 2011 to be significantly different from that in 2007 or 2014 (Fisher’s exact test P=0.018, 0.0078), but did not find a difference comparing 2007 to 2014 (Fisher’s exact test P=0.24), indicating greater similarity between samples separated by a longer time period than between samples from closer time periods. We also found changes in the accessory gene content of the NVT population between 2007 and 2011 to have been reduced by 2014. Amongst the new serotypes targeted by PCV-13, four were present in our sample. The proportion of our sample composed of PCV-13-only vaccine serotypes 19A, 6C and 7F decreased between 2007 and 2014, but no such reduction was seen for serotype 3. We did, however, observe differences in the genetic composition of the pre- and post-PCV-13 serotype 3 population. Our isolates were collected during discrete sampling periods from a small geographical area, which may limit the generalizability of our findings. Pneumococcal diversity increased immediately following the introduction of PCV-13, but subsequently returned to pre-vaccination levels. This is reflected in the distribution of NVT lineages, and, to a lesser extent, their accessory gene frequencies. As such, there may be a period during which the population is particularly disrupted by vaccination before returning to a more stable distribution. The persistence and shifting genetic composition of serotype 3 is a concern and warrants further investigation.

Background Of over 90 serotypes of Streptococcus pneumoniae, only seven were included in the first pneumococcal conjugate vaccine (PCV). While PCV reduced the disease incidence, in part because of a herd immunity effect, a replacement effect was observed whereby disease was increasingly caused by serotypes not included in the vaccine. Dynamic transmission models can account for these effects to describe post-vaccination scenarios, whereas economic evaluations can enable decision-makers to compare vaccines of increasing valency for implementation. Aim The aim of this review was to examine epidemiological and economic models and their assumptions for their potential contributions to future research and immunisation policy. Sources Pubmed, Scopus, Ovid, ISI Web of Knowledge, Centre of Reviews and Dissemination (CRD) databases were searched. Content Twenty-three dynamic transmission models and 21 economic models were retrieved and reviewed. Published models employed various templates, revealing several key uncertainties regarding the biology and epidemiology of pneumococcal infection. While models suggested that PCVs will reduce the burden of disease, the extent to which they are predicted to do so depended on various assumptions regarding features of pneumococcal infection and epidemiology that governed PCV cost-effectiveness as well. Such features include the duration of protection and competitive interactions between serotypes, which are unclear at present, but which directly relate to herd immunity and serotype replacement. Implications Economic evaluations are not typically based on transmission dynamic models and hence omit indirect herd immunity effects. The two tools could be used in conjunction to inform decision-makers on vaccine implementation, but so far there have been few attempts to build economic evaluations on transmission dynamic models, and none in this field. Future directions for research could include studies to evaluate key parameters for the models involving herd immunity, serotype competition and the natural history of infection.

Constantly surrounded by kin or alien organisms in nature, eukaryotes and prokaryotes developed various communication systems to coordinate adaptive multi-entity behavior. In complex and overcrowded environments, they require to discriminate relevant signals in a myriad of pheromones to execute appropriate responses. In the human gut commensal Streptococcus salivarius, the cytoplasmic Rgg/RNPP regulator ComR couples competence to bacteriocin-mediated predation. Here, we describe a paralogous sensor duo, ScuR and SarF, which circumvents ComR in order to disconnect these two physiological processes. We highlighted the recurring role of Rgg/RNPP in the production of antimicrobials and designed a robust genetic screen to unveil potent/optimized peptide pheromones. Further mutational and biochemical analyses dissected the modifiable selectivity toward their pheromone and operating sequences at the subtle molecular level. Additionally, our results highlight how we might mobilize antimicrobial molecules while silencing competence in endogenous populations of human microflora and temper gut disorders provoked by bacterial pathogens.

Bacterial growth and cell division requires precise spatiotemporal regulation of the synthesis and remodelling of the peptidoglycan layer that surrounds the cytoplasmic membrane. GpsB is a cytosolic protein that affects cell wall synthesis by binding to the cytoplasmic mini-domains of peptidoglycan synthases to ensure their correct subcellular localisation. Here we have discovered critical structural features for the interaction of GpsB with peptidoglycan synthases from three different bacteria and demonstrated their importance for cell wall growth and viability. We have used these structural motifs to predict and confirm novel partners of GpsB in Bacillus subtilis, illuminating the role of this key regulator of peptidoglycan synthesis. GpsB thus functions as an adaptor, to mediate the interaction between membrane proteins, scaffolding proteins, signalling proteins and enzymes to generate larger protein complexes at specific sites in a bacterial cell cycle-dependent manner. Given the importance of GpsB in pathogenic bacteria, this study has not only revealed mechanistic details of how cell wall synthesis is co-ordinated with the bacterial cell cycle but could also represent a starting point for the design of much needed new antibiotics.

Pneumococcal whole cell vaccines (WCVs) could cost-effectively protect against a greater strain diversity than current capsule-based vaccines. Immunoglobulin G (IgG) responses to a WCV were characterised by applying longitudinally-sampled sera, available from 35 adult placebo-controlled phase I trial participants, to a panproteome microarray. Despite individuals maintaining distinctive antibody ‘fingerprints’, responses were consistent across vaccinated cohorts. Seventy-two functionally distinct proteins were associated with WCV-induced increases in IgG binding. These shared characteristics with naturally immunogenic proteins, being enriched for transporters and cell wall metabolism enzymes, likely unusually exposed on the unencapsulated WCV’s surface. Vaccine-induced responses were specific to variants of the diverse PclA, PspC and ZmpB proteins, whereas PspA- and ZmpA-induced antibodies recognised a broader set of alleles. Temporal variation in IgG levels suggested a mixture of anamnestic and novel responses. These reproducible increases in IgG binding to a limited, but functionally diverse, set of conserved proteins indicate WCV could provide species-wide immunity. Clinical trial registration: The trial was registered with ClinicalTrials.gov with Identifier NCT01537185; the results are available from https://clinicaltrials.gov/ct2/show/results/NCT01537185., eLife digest Streptococcus pneumoniae is a bug that causes pneumonia and meningitis, killing around a million people each year. Vaccines now exist to protect young children against these diseases, but they are expensive and do not work against all the strains of the bacteria. This is because these shots train the body’s immune system to recognize and attack the bacterium’s capsule, a layer of sugars that surrounds the microbe and is often different between strains. One possible solution could be a cheap, whole cell vaccine. These injections expose the body to genetically modified S. pneumoniae that do not carry the capsule. Such treatment has now been tested in a small number of people during a clinical trial. Here, Campo et al. use a technique known as panproteome array to scan samples collected during this trial, and identify which elements the body learns to recognize when it is exposed to the genetically manipulated strain of S. pneumoniae. The results show that when volunteers receive this vaccine, their body targets proteins that the capsule normally shields from the immune system. Many of these proteins are very similar across all strains of S. pneumoniae, which means that the whole cell vaccine could potentially better protect against a broad spectrum of bacteria. However, further studies are needed to assess whether this is the case, especially in infants.

Summary Streptococcus pneumoniae can acquire antibiotic resistance by activation of competence and subsequent DNA uptake. Here, we demonstrate that aztreonam (ATM) and clavulanic acid (CLA) promote competence. We show that both compounds induce cell chain formation by targeting the d,d-carboxypeptidase PBP3. In support of the hypothesis that chain formation promotes competence, we demonstrate that an autolysin mutant (ΔlytB) is hypercompetent. Since competence is initiated by the binding of a small extracellular peptide (CSP) to a membrane-anchored receptor (ComD), we wondered whether chain formation alters CSP diffusion kinetics. Indeed, ATM or CLA presence affects competence synchronization by shifting from global to local quorum sensing, as CSP is primarily retained to chained cells, rather than shared in a common pool. Importantly, autocrine-like signaling prolongs the time window in which the population is able to take up DNA. Together, these insights demonstrate the versatility of quorum sensing and highlight the importance of an accurate antibiotic prescription., Graphical Abstract, Highlights • Identification of a mechanism by which antibiotics induce competence in S. pneumoniae • Antibiotics targeting penicillin-binding protein 3 promote chain formation • Cell chains retain, rather than diffuse, the quorum-sensing peptide CSP • Chaining populations feature a longer competence and transformation time window, Streptococcus pneumoniae can take up exogenous DNA by activating competence. Aztreonam and clavulanic acid can induce competence by targeting PBP3, leading to cell chaining. Cell chaining reshapes quorum sensing to autocrine-like signaling and increases the time window in which cells can take up DNA, potentially accelerating the spread of antibiotic resistance.

Acute respiratory infection (ARI) is the most frequent cause for hospitalization in infants and young children. Using multiplexed nCounter technology to digitally quantify 600 human mRNAs in parallel with 14 virus- and 5 bacterium-specific RNAs, we characterized viral and bacterial presence in nasopharyngeal aspirates (NPA) of 58 children with ARI and determined the corresponding in situ immune profiles. NPA contained different groups of organisms and these were classified into bacterial (n = 27), viral (n = 5), codetection [containing both viral and bacterial transcripts (n = 21), or indeterminate intermediate where microbial load is below threshold (n = 5)]. We then identified differentially expressed immune transcripts (DEITs) comparing NPAs from symptomatic children vs. healthy controls, and comparing children presenting NPAs with detectable microbial load vs. indeterminate. We observed a strong innate immune response in NPAs, due to the presence of evolutionarily conserved type I Interferon (IFN)-stimulated genes (ISG), which was correlated with total bacterial and/or viral load. In comparison with indeterminate NPAs, adaptive immunity transcripts discriminated among viral, bacterial, and codetected microbial profiles. In viral NPAs, B cell transcripts were significantly enriched among DEITs, while only type III IFN was correlated with viral load. In bacterial NPAs, myeloid cells and coinhibitory transcripts were enriched and significantly correlated with bacterial load. In conclusion, digital nCounter transcriptomics provide a microbial and immunological in situ "snapshot" of the nasopharyngeal interface in children with ARI. This enabled discrimination among viral, bacterial, codetection, and indeterminate transcripts in the samples using non-invasive sampling. ispartof: FRONTIERS IN MICROBIOLOGY vol:9 issue:NOV ispartof: location:Switzerland status: published

Streptococcus pneumoniae is an opportunistic human pathogen that typically colonizes the nasopharyngeal passage and causes lethal disease in other host niches such as the lung or the meninges. How pneumococcal genes are expressed and regulated at the different stages of its life cycle, as commensal or as pathogen, has not been entirely described. To chart the transcriptional responses of S. pneumoniae, we quantified the transcriptome under 22 different infection-relevant conditions. The transcriptomic compendium exposed a high level of dynamic expression and, strikingly, all annotated pneumococcal genomic features were expressed in at least one of the studied conditions. By computing the correlation of gene expression of every two genes across all studied conditions, we created a co-expression matrix that provides valuable information on both operon structure and regulatory processes. The co-expression data is highly consistent with well-characterized operons and regulons, such as the PyrR, ComE and ComX regulons, and has allowed us to identify a new member of the competence regulon. Finally, we created an interactive data center named PneumoExpress (www.veeninglab.com/pneumoexpress) that enables users to access the expression data as well as the co-expression matrix in an intuitive and efficient manner, providing a valuable resource to the pneumococcal research community.