Invasive pneumococcal disease in Latin America and the Caribbean: Serotype distribution, disease burden, and impact of vaccination. A systematic review and meta-analysis.

BACKGROUND: Invasive pneumococcal diseases (IPD) are associated with high morbidity, mortality, and health costs worldwide, particularly in Latin America and the Caribbean (LAC). Surveillance about the distribution of serotypes causing IPD and the impact of pneumococcal vaccination is an important epidemiological tool to monitor disease activity trends, inform public health decision-making, and implement relevant prevention and control measures. OBJECTIVES: To estimate the serotype distribution for IPD and the related disease burden in LAC before, during, and after implementing the pneumococcal vaccine immunization program in LAC. METHODS: Systematic literature review following Cochrane methods of studies from LAC. We evaluated the impact of the pneumococcal vaccine on hospitalization and death during or after hospitalizations due to pneumococcal disease and serotype-specific disease over time. We also analyzed the incidence of serotyped IPD in pneumococcal conjugate vaccine PCV10 and PCV13. The protocol was registered in PROSPERO (ID: CRD42023392097). RESULTS: 155 epidemiological studies were screened and provided epidemiological data on IPD. Meta-analysis of invasive diseases in children <5 years old found that 57%-65% of causative serotypes were included in PCV10 and 66%-84% in PCV13. After PCV introduction, vaccine serotypes declined in IPD, and the emergence of non-vaccine serotypes varied by country. CONCLUSIONS: Pneumococcal conjugate vaccines significantly reduced IPD and shifted serotype distribution in Latin America and the Caribbean. PCV10/PCV13 covered 57-84% of serotypes in children under 5, with marked decline in PCV serotypes post-vaccination. Continuous surveillance remains crucial for monitoring evolving serotypes and informing public health action.

Glucose and Oxygen Levels Modulate the Pore-Forming Effects of Cholesterol-Dependent Cytolysin Pneumolysin from Streptococcus pneumoniae.

A major Streptococcus pneumoniae pathogenic factor is the cholesterol-dependent cytolysin pneumolysin, binding membrane cholesterol and producing permanent lytic or transient pores. During brain infections, vascular damage with variable ischemia occurs. The role of ischemia on pneumolysin's pore-forming capacity remains unknown. In acute brain slice cultures and primary cultured glia, we studied acute toxin lysis (via propidium iodide staining and LDH release) and transient pore formation (by analyzing increases in the intracellular calcium). We analyzed normal peripheral tissue glucose conditions (80 mg%), normal brain glucose levels (20 mg%), and brain hypoglycemic conditions (3 mg%), in combinations either with normoxia (8% oxygen) or hypoxia (2% oxygen). At 80 mg% glucose, hypoxia enhanced cytolysis via pneumolysin. At 20 mg% glucose, hypoxia did not affect cell lysis, but impaired calcium restoration after non-lytic pore formation. Only at 3 mg% glucose, during normoxia, did pneumolysin produce stronger lysis. In hypoglycemic (3 mg% glucose) conditions, pneumolysin caused a milder calcium increase, but restoration was missing. Microglia bound more pneumolysin than astrocytes and demonstrated generally stronger calcium elevation. Thus, our work demonstrated that the toxin pore-forming capacity in cells continuously diminishes when oxygen is reduced, overlapping with a continuously reduced ability of cells to maintain homeostasis of the calcium influx once oxygen and glucose are reduced.

Characterization of bacterial and viral pathogens in the respiratory tract of children with HIV-associated chronic lung disease: a case-control study.

INTRODUCTION: Chronic lung disease is a major cause of morbidity in African children with HIV infection; however, the microbial determinants of HIV-associated chronic lung disease (HCLD) remain poorly understood. We conducted a case-control study to investigate the prevalence and densities of respiratory microbes among pneumococcal conjugate vaccine (PCV)-naive children with (HCLD +) and without HCLD (HCLD-) established on antiretroviral treatment (ART). METHODS: Nasopharyngeal swabs collected from HCLD + (defined as forced-expiratory-volume/second < -1.0 without reversibility postbronchodilation) and age-, site-, and duration-of-ART-matched HCLD- participants aged between 6-19 years enrolled in Zimbabwe and Malawi (BREATHE trial-NCT02426112) were tested for 94 pneumococcal serotypes together with twelve bacteria, including Streptococcus pneumoniae (SP), Staphylococcus aureus (SA), Haemophilus influenzae (HI), Moraxella catarrhalis (MC), and eight viruses, including human rhinovirus (HRV), respiratory syncytial virus A or B, and human metapneumovirus, using nanofluidic qPCR (Standard BioTools formerly known as Fluidigm). Fisher's exact test and logistic regression analysis were used for between-group comparisons and risk factors associated with common respiratory microbes, respectively. RESULTS: A total of 345 participants (287 HCLD + , 58 HCLD-; median age, 15.5 years [IQR = 12.8-18], females, 52%) were included in the final analysis. The prevalence of SP (40%[116/287] vs. 21%[12/58], p = 0.005) and HRV (7%[21/287] vs. 0%[0/58], p = 0.032) were higher in HCLD + participants compared to HCLD- participants. Of the participants positive for SP (116 HCLD + & 12 HCLD-), 66% [85/128] had non-PCV-13 serotypes detected. Overall, PCV-13 serotypes (4, 19A, 19F: 16% [7/43] each) and NVT 13 and 21 (9% [8/85] each) predominated. The densities of HI (2 × 104 genomic equivalents [GE/ml] vs. 3 × 102 GE/ml, p = 0.006) and MC (1 × 104 GE/ml vs. 1 × 103 GE/ml, p = 0.031) were higher in HCLD + compared to HCLD-. Bacterial codetection (≥ any 2 bacteria) was higher in the HCLD + group (36% [114/287] vs. (19% [11/58]), (p = 0.014), with SP and HI codetection (HCLD + : 30% [86/287] vs. HCLD-: 12% [7/58], p = 0.005) predominating. Viruses (predominantly HRV) were detected only in HCLD + participants. Lastly, participants with a history of previous tuberculosis treatment were more likely to carry SP (adjusted odds ratio (aOR): 1.9 [1.1 -3.2], p = 0.021) or HI (aOR: 2.0 [1.2 - 3.3], p = 0.011), while those who used ART for ≥ 2 years were less likely to carry HI (aOR: 0.3 [0.1 - 0.8], p = 0.005) and MC (aOR: 0.4 [0.1 - 0.9], p = 0.039). CONCLUSION: Children with HCLD + were more likely to be colonized by SP and HRV and had higher HI and MC bacterial loads in their nasopharynx. The role of SP, HI, and HRV in the pathogenesis of CLD, including how they influence the risk of acute exacerbations, should be studied further. TRIAL REGISTRATION: The BREATHE trial (ClinicalTrials.gov Identifier: NCT02426112 , registered date: 24 April 2015).

Elongasome core proteins and class A PBP1a display zonal, processive movement at the midcell of Streptococcus pneumoniae.

Ovoid-shaped bacteria, such as Streptococcus pneumoniae (pneumococcus), have two spatially separated peptidoglycan (PG) synthase nanomachines that locate zonally to the midcell of dividing cells. The septal PG synthase bPBP2x:FtsW closes the septum of dividing pneumococcal cells, whereas the elongasome located on the outer edge of the septal annulus synthesizes peripheral PG outward. We showed previously by sm-TIRFm that the septal PG synthase moves circumferentially at midcell, driven by PG synthesis and not by FtsZ treadmilling. The pneumococcal elongasome consists of the PG synthase bPBP2b:RodA, regulators MreC, MreD, and RodZ, but not MreB, and genetically associated proteins Class A aPBP1a and muramidase MpgA. Given its zonal location separate from FtsZ, it was of considerable interest to determine the dynamics of proteins in the pneumococcal elongasome. We found that bPBP2b, RodA, and MreC move circumferentially with the same velocities and durations at midcell, driven by PG synthesis. However, outside of the midcell zone, the majority of these elongasome proteins move diffusively over the entire surface of cells. Depletion of MreC resulted in loss of circumferential movement of bPBP2b, and bPBP2b and RodA require each other for localization and circumferential movement. Notably, a fraction of aPBP1a molecules also moved circumferentially at midcell with velocities similar to those of components of the core elongasome, but for shorter durations. Other aPBP1a molecules were static at midcell or diffusing over cell bodies. Last, MpgA displayed nonprocessive, subdiffusive motion that was largely confined to the midcell region and less frequently detected over the cell body.

Evaluation of a multiplex-qPCR for paediatric pleural empyema-An observational study in hospitalised children.

Pleural empyema is a serious complication of pneumonia in children. Negative bacterial cultures commonly impede optimal antibiotic therapy. To improve bacterial identification, we developed a molecular assay and evaluated its performance compared with bacterial culture. Our multiplex-quantitative PCR to detect Streptococcus pneumoniae, Streptococcus pyogenes, Staphylococcus aureus and Haemophilus influenzae was assessed using bacterial genomic DNA and laboratory-prepared samples (n = 267). To evaluate clinical performance, we conducted the Molecular Assessment of Thoracic Empyema (MATE) observational study, enrolling children hospitalised with empyema. Pleural fluids were tested by bacterial culture and multiplex-qPCR, and performance determined using a study gold standard. We determined clinical sensitivity and time-to-organism-identification to assess the potential of the multiplex-qPCR to reduce the duration of empiric untargeted antibiotic therapy. Using spiked samples, the multiplex-qPCR demonstrated 213/215 (99.1%) sensitivity and 52/52 (100%) specificity for all organisms. During May 2019-March 2023, 100 children were enrolled in the MATE study; median age was 3.9 years (IQR 2-5.6). A bacterial pathogen was identified in 90/100 (90%) specimens by multiplex-qPCR, and 24/100 (24%) by bacterial culture (P <0.001). Multiplex-qPCR identified a bacterial cause in 68/76 (90%) culture-negative specimens. S. pneumoniae was the most common pathogen, identified in 67/100 (67%) specimens. We estimate our multiplex-qPCR would have reduced the duration of untargeted antibiotic therapy in 61% of cases by a median 20 days (IQR 17.5-23, range 1-55). Multiplex-qPCR significantly increased pathogen detection compared with culture and may allow for reducing the duration of untargeted antibiotic therapy.

Therapeutic potential activity of quercetin complexes against Streptococcus pneumoniae.

This study investigates quercetin complexes as potential synergistic agents against the important respiratory pathogen Streptococcus pneumoniae. Six quercetin complexes (QCX1-6) were synthesized by reacting quercetin with various metal salts and boronic acids and characterized using FTIR spectroscopy. Their antibacterial activity alone and in synergism with antibiotics was evaluated against S. pneumoniae ATCC 49619 using disc diffusion screening, broth microdilution MIC determination, and checkerboard assays. Complexes QCX-3 and QCX-4 demonstrated synergy when combined with levofloxacin via fractional inhibitory concentration indices ≤ 0.5 as confirmed by time-kill kinetics. Molecular docking elucidated interactions of these combinations with virulence enzymes sortase A and sialidase. A biofilm inhibition assay found the synergistic combinations more potently reduced biofilm formation versus monotherapy. Additionally, gene-gene interaction networks, biological activity predictions and in-silico toxicity profiling provided insights into potential mechanisms of action and safety.

Parallel detection of multiple biomarkers in a point-of-care-competent device for the prediction of exacerbations in chronic inflammatory lung disease.

Sudden aggravations of chronic inflammatory airway diseases are difficult-to-foresee life-threatening episodes for which advanced prognosis-systems are highly desirable. Here we present an experimental chip-based fluidic system designed for the rapid and sensitive measurement of biomarkers prognostic for potentially imminent asthma or COPD exacerbations. As model biomarkers we chose three cytokines (interleukin-6, interleukin-8, tumor necrosis factor alpha), the bacterial infection marker C-reactive protein and the bacterial pathogen Streptococcus pneumoniae-all relevant factors in exacerbation episodes. Assay protocols established in laboratory environments were adapted to 3D-printed fluidic devices with emphasis on short processing times, low reagent consumption and a low limit of detection in order to enable the fluidic system to be used in point-of-care settings. The final device demonstrator was validated with patient sample material for its capability to detect endogenous as well as exogenous biomarkers in parallel.

Expansion of pneumococcal serotype 23F and 14 lineages with genotypic changes in capsule polysaccharide locus and virulence gene profiles post introduction of pneumococcal conjugate vaccine in Blantyre, Malawi.

Since the introduction of the 13-valent pneumococcal conjugate vaccine (PCV13) in Malawi in 2011, there has been persistent carriage of vaccine serotype (VT) Streptococcus pneumoniae, despite high vaccine coverage. To determine if there has been a genetic change within the VT capsule polysaccharide (cps) loci since the vaccine's introduction, we compared 1022 whole-genome-sequenced VT isolates from 1998 to 2019. We identified the clonal expansion of a multidrug-resistant, penicillin non-susceptible serotype 23F GPSC14-ST2059 lineage, a serotype 14 GPSC9-ST782 lineage and a novel serotype 14 sequence type GPSC9-ST18728 lineage. Serotype 23F GPSC14-ST2059 had an I253T mutation within the capsule oligosaccharide repeat unit polymerase Wzy protein, which is predicted in silico to alter the protein pocket cavity. Moreover, serotype 23F GPSC14-ST2059 had SNPs in the DNA binding sites for the cps transcriptional repressors CspR and SpxR. Serotype 14 GPSC9-ST782 harbours a non-truncated version of the large repetitive protein (Lrp), containing a Cna protein B-type domain which is also present in proteins associated with infection and colonisation. These emergent lineages also harboured genes associated with antibiotic resistance, and the promotion of colonisation and infection which were absent in other lineages of the same serotype. Together these data suggest that in addition to serotype replacement, modifications of the capsule locus associated with changes in virulence factor expression and antibiotic resistance may promote vaccine escape. In summary, the study highlights that the persistence of vaccine serotype carriage despite high vaccine coverage in Malawi may be partly caused by expansion of VT lineages post-PCV13 rollout.

Long-term evolution of Streptococcus mitis and Streptococcus pneumoniae leads to higher genetic diversity within rather than between human populations.

Evaluation of the apportionment of genetic diversity of human bacterial commensals within and between human populations is an important step in the characterization of their evolutionary potential. Recent studies showed a correlation between the genomic diversity of human commensal strains and that of their host, but the strength of this correlation and of the geographic structure among human populations is a matter of debate. Here, we studied the genomic diversity and evolution of the phylogenetically related oro-nasopharyngeal healthy-carriage Streptococcus mitis and Streptococcus pneumoniae, whose lifestyles range from stricter commensalism to high pathogenic potential. A total of 119 S. mitis genomes showed higher within- and among-host variation than 810 S. pneumoniae genomes in European, East Asian and African populations. Summary statistics of the site-frequency spectrum for synonymous and non-synonymous variation and ABC modelling showed this difference to be due to higher ancestral bacterial population effective size (Ne) in S. mitis, whose genomic variation has been maintained close to mutation-drift equilibrium across (at least many) generations, whereas S. pneumoniae has been expanding from a smaller ancestral bacterial population. Strikingly, both species show limited differentiation among human populations. As genetic differentiation is inversely proportional to the product of effective population size and migration rate (Nem), we argue that large Ne have led to similar differentiation patterns, even if m is very low for S. mitis. We conclude that more diversity within than among human populations and limited population differentiation must be common features of the human microbiome due to large Ne.

Environmental and genetic regulation of Streptococcus pneumoniae galactose catabolic pathways.

Efficient utilization of nutrients is crucial for microbial survival and virulence. The same nutrient may be utilized by multiple catabolic pathways, indicating that the physical and chemical environments for induction as well as their functional roles may differ. Here, we study the tagatose and Leloir pathways for galactose catabolism of the human pathogen Streptococcus pneumoniae. We show that galactose utilization potentiates pneumococcal virulence, the induction of galactose catabolic pathways is influenced differentially by the concentration of galactose and temperature, and sialic acid downregulates galactose catabolism. Furthermore, the genetic regulation and in vivo induction of each pathway differ, and both galactose catabolic pathways can be turned off with a galactose analogue in a substrate-specific manner, indicating that galactose catabolic pathways can be potential drug targets.

Clinical characteristics and risk factors for poor outcomes of invasive pneumococcal disease in pediatric patients in China.

BACKGROUND: Invasive pneumococcal disease (IPD) is a significant health concern in children worldwide. In this study, we aimed to analyze the clinical features, antibiotic resistance, and risk variables for poor outcomes in patients with IPD in Hangzhou. METHODS: A retrospective single-centre study was performed using the pediatric intensive care (PIC) database from 2010 to 2018. The clinical characteristics, laboratory data, antimicrobial resistance, and risk factors for in-hospital mortality and sepsis in patients with IPD in intensive care units (ICUs) were analyzed systematically. RESULTS: A total of 178 IPD patients were included in the study. The majority of the IPD children were 2-10 years old. Antimicrobial resistance tests of S. pneumoniae isolates revealed high resistance to erythromycin, tetracycline and compound sulfamethoxazole (SMZ-Co). All the isolates were sensitive to vancomycin, linezolid, moxifloxacin, telithromycin, ofloxacin, and levofloxacin. IPD patients may experience poor outcomes, including death and sepsis. The in-hospital mortality was 3.93%, and 34.27% of patients suffered from sepsis. Temperature (OR 3.80, 95% CI 1.62-8.87; P = 0.0021), Partial Pressure of Oxygen in Arterial Blood (PaO2) (OR 0.99, 95% CI 0.98-1.00; P = 0.0266), and albumin (OR 0.89, 95% CI 0.80-0.99; P = 0.0329) were found to be independent risk factors for sepsis in children with IPD. CONCLUSION: Pediatric IPD deserves attention in China. Appropriate surveillance and antibiotic selection are crucial in managing resistant strains. Early identification of high-risk individuals with risk factors contributes to the development of appropriate treatment strategies.

[Mortality in bacterial pneumonia due to pneumococcus]. / Mortalidad en neumonía bacteriémica por neumococo.

INTRODUCTION: Despite improvements in health care, pneumonia-associated mortality remains high. The objective of this study was to analyze the factors associated with mortality in bacteremic pneumonia caused by pneumococcus. METHODS: Retrospective cohort study in adult patients with pneumonia diagnosis and isolation of pneumococcus in blood cultures, between January 2012 and May 2021, was carried out. Clinical and laboratory variables, radiological involvement, evolution and mortality during hospitalization were analyzed. The group of deceased patients was compared with that of survivors. RESULTS: 152 patients were included. Median age: 58 years; men: 58.9%; 33% presented a CURB-65 > than 2 at admission. Overall mortality: 34% (n=52). Deceased patients were more tachypneic on admission (respiratory rate 26 vs. 22; p=0.003), presented sensory alteration more frequently (58% vs. 14%; p< 0.001), PaO2/fraction of inspired oxygen ratio < 250 (58% vs. 22%; p<0.001), bilateral radiological compromise (50% vs. 32%; p=0.03), needed mechanical ventilation (50% vs 12%; p< 0.001), higher blood creatinine values (1.6 vs. 1.15; p=0.01), lower white blood cell count (10 900 vs 17 400; p=0.002), a lower glucose dosage (111 vs. 120; p=0.01), and fewer days of hospital stay (6 vs. 9; p=0.015). In logistic regression model, significant differences were maintained in the following factors associated with mortality: mechanical ventilation (OR=3.54), altered mental status (OR=5.95), chest X-ray with bilateral compromise (OR 3.20) and PAFI less than 250 (OR=3.62). CONCLUSION: In our series, the factors related to mortality, despite the presence of bacteremia, do not differ from those published in the literature and which are part of the different prognostic scores used in routine practice.

Introducción: A pesar de las mejoras en los cuidados de la salud, la mortalidad asociada a neumonía continúa siendo alta. El objetivo de este estudio fue analizar los factores asociados a mortalidad en neumonía bacteriémica por neumococo. Métodos: Estudio de cohorte retrospectiva en pacientes adultos con diagnóstico de neumonía y neumococo aislado en hemocultivos, entre enero 2012 y mayo 2021. Se analizaron: variables clínicas y de laboratorio, compromiso radiológico, evolución y mortalidad durante la internación. Se comparó el grupo de pacientes fallecidos con el de sobrevivientes. Resultados: Se incluyeron 152 pacientes. La mediana de edad fue de 58 años y el 58.9% fueron hombres. El 33% presentó un CURB-65 mayor a 2 al momento de internación. La mortalidad global fue 34% (n=52). Los pacientes fallecidos se encontraban más frecuentemente taquipneicos al ingreso (frecuencia respiratoria 26 vs. 22; p=0.003), presentaban más frecuentemente alteración del sensorio (58% vs. 14%; p< 0.001), PaO2/fracción inspirada de oxígeno (PAFI) < 250 (58% vs. 22%; p<0.001), compromiso radiológico bilateral (50% vs. 32%; p=0.03), necesidad de asistencia respiratoria mecánica (ARM) (50% vs. 12%; p< 0.001), mayor valor de creatinina en sangre (1.6 vs. 1.15; p=0.01), menor recuento de glóbulos blancos (10 900 vs. 17 400; p=0.002), menor valor de glucemia (111 vs. 120; p=0.01) y menos días de estancia hospitalaria (6 vs. 9; p=0.015). En el análisis de regresión logística multivariable se mantuvieron diferencias significativas en los siguientes factores asociados a mortalidad: ventilación mecánica (OR=3.54), confusión (OR=5.95), radiografía con compromiso bilateral (OR= 3.20) y PAFI < 250 (OR=3.62). Conclusión: Los factores relacionados con mortalidad, a pesar de la presencia de bacteriemia, no difieren de los publicados en la literatura y forman parte de los scores pronósticos de práctica habitual.

Molecular and structural basis of oligopeptide recognition by the Ami transporter system in pneumococci.

ATP-binding cassette (ABC) transport systems are crucial for bacteria to ensure sufficient uptake of nutrients that are not produced de novo or improve the energy balance. The cell surface of the pathobiont Streptococcus pneumoniae (pneumococcus) is decorated with a substantial array of ABC transporters, critically influencing nasopharyngeal colonization and invasive infections. Given the auxotrophic nature of pneumococci for certain amino acids, the Ami ABC transporter system, orchestrating oligopeptide uptake, becomes indispensable in host compartments lacking amino acids. The system comprises five exposed Oligopeptide Binding Proteins (OBPs) and four proteins building the ABC transporter channel. Here, we present a structural analysis of all the OBPs in this system. Multiple crystallographic structures, capturing both open and closed conformations along with complexes involving chemically synthesized peptides, have been solved at high resolution providing insights into the molecular basis of their diverse peptide specificities. Mass spectrometry analysis of oligopeptides demonstrates the unexpected remarkable promiscuity of some of these proteins when expressed in Escherichia coli, displaying affinity for a wide range of peptides. Finally, a model is proposed for the complete Ami transport system in complex with its various OBPs. We further disclosed, through in silico modelling, some essential structural changes facilitating oligopeptide transport into the cellular cytoplasm. Thus, the structural analysis of the Ami system provides valuable insights into the mechanism and specificity of oligopeptide binding by the different OBPs, shedding light on the intricacies of the uptake mechanism and the in vivo implications for this human pathogen.

Human monoclonal antibodies protect against viral-mediated pneumococcal superinfection.

Introduction: Community-acquired pneumonia (CAP) is a global health concern, with 25% of cases attributed to Streptococcus pneumoniae (Spn). Viral infections like influenza A virus (IAV), respiratory syncytial virus (RSV), and human metapneumovirus (hMPV) increase the risk of Spn, leading to severe complications due to compromised host immunity. Methods: We evaluated the efficacy of an anti-PhtD monoclonal antibody (mAb) cocktail therapy (PhtD3 + 7) in improving survival rates in three viral/bacterial coinfection models: IAV/Spn, hMPV/Spn, and RSV/Spn. Results: The PhtD3 + 7 mAb cocktail outperformed antiviral mAbs, resulting in prolonged survival. In the IAV/Spn model, it reduced bacterial titers in blood and lungs by 2-4 logs. In the hMPV/Spn model, PhtD3 + 7 provided greater protection than the hMPV-neutralizing mAb MPV467, significantly reducing bacterial titers. In the RSV/Spn model, PhtD3 + 7 offered slightly better protection than the antiviral mAb D25, uniquely decreasing bacterial titers in blood and lungs. Discussion: Given the threat of antibiotic resistance, our findings highlight the potential of anti-PhtD mAb therapy as an effective option for treating viral and secondary pneumococcal coinfections.

Effect of erythromycin residuals in food on the development of resistance in Streptococcus pneumoniae: an in vivo study in Galleria mellonella.

Background: The use of antimicrobials to treat food animals may result in antimicrobial residues in foodstuffs of animal origin. The European Medicines Association (EMA) and World Health Organization (WHO) define safe antimicrobial concentrations in food based on acceptable daily intakes (ADIs). It is unknown if ADI doses of antimicrobials in food could influence the antimicrobial susceptibility of human-associated bacteria. Objectives: This aim of this study was to evaluate if the consumption of ADI doses of erythromycin could select for erythromycin resistance in a Galleria mellonella model of Streptococcus pneumoniae infection. Methods: A chronic model of S. pneumoniae infection in G. mellonella larvae was used for the experiment. Inoculation of larvae with S. pneumoniae was followed by injections of erythromycin ADI doses (0.0875 and 0.012 µg/ml according to EMA and WHO, respectively). Isolation of S. pneumoniae colonies was then performed on selective agar plates. Minimum inhibitory concentrations (MICs) of resistant colonies were measured, and whole genome sequencing (WGS) was performed followed by variant calling to determine the genetic modifications. Results: Exposure to single doses of both EMA and WHO ADI doses of erythromycin resulted in the emergence of erythromycin resistance in S. pneumoniae. Emergent resistance to erythromycin was associated with a mutation in rplA, which codes for the L1 ribosomal protein and has been linked to macrolide resistance in previous studies. Conclusion: In our in vivo model, even single doses of erythromycin that are classified as acceptable by the WHO and EMA induced significant increases in erythromycin MICs in S. pneumoniae. These results suggest the need to include the induction of antimicrobial resistance (AMR) as a significant criterion for determining ADIs.

Synthetic BSA-conjugated disaccharide related to the Streptococcus pneumoniae serotype 3 capsular polysaccharide increases IL-17A Levels, γδ T cells, and B1 cells in mice.

The disaccharide (ß-D-glucopyranosyluronic acid)-(1→4)-ß-D-glucopyranoside represents a repeating unit of the capsular polysaccharide of Streptococcus pneumoniae serotype 3. A conjugate of the disaccharide with BSA (di-BSA conjugate) adjuvanted with aluminum hydroxide induced - in contrast to the non-adjuvanted conjugate - IgG1 antibody production and protected mice against S. pneumoniae serotype 3 infection after intraperitoneal prime-boost immunization. Adjuvanted and non-adjuvanted conjugates induced production of Th1 (IFNγ, TNFα); Th2 (IL-5, IL-13); Th17 (IL-17A), Th1/Th17 (IL-22), and Th2/Th17 cytokines (IL-21) after immunization. The concentration of cytokines in mice sera was higher in response to the adjuvanted conjugate, with the highest level of IL-17A production after the prime and boost immunizations. In contrast, the non-adjuvanted conjugate elicited only weak production of IL-17A, which gradually decreased after the second immunization. After boost immunization of mice with the adjuvanted di-BSA conjugate, there was a significant increase in the number of CD45+/CD19+ B cells, TCR+ γδ T cell, CD5+ В1 cells, and activated cells with MHC II+ expression in the spleens of the mice. IL-17A, TCR+ γδ T cells, and CD5+ В1 cells play a crucial role in preventing pneumococcal infection, but can also contribute to autoimmune diseases. Immunization with the adjuvanted and non-adjuvanted di-BSA conjugate did not elicit autoantibodies against double-stranded DNA targeting cell nuclei in mice. Thus, the molecular and cellular markers associated with antibody production and protective activity in response to immunization with the di-BSA conjugate adjuvanted with aluminum hydroxide are IL-17A, TCR+ γδ T cells, and CD5+ В1 cells against the background of increasing MHC II+ expression.

[Pay attention to new evolution trends in the etiology of community-acquired pneumonia in adults].

Over the past two to three decades, the emergence and re-emergence of new infectious diseases, advances in molecular detection techniques of pathogens, antibiotic resistance, changes in population lifestyle and immune status (including vaccination), and other factors have led to new evolutions in the etiology of community-acquired pneumonia (CAP). (1) Although Streptococcus pneumoniae remains a common pathogen of CAP, it is no longer the leading cause in China and the United States. According to the results of 2 multicenter studies in China in the early 21st century, Streptococcus pneumoniae accounted for 10.3% and 12.0% of adult CAP pathogens, respectively, ranking second. A study on key pathogens of adult CAP in nine cities in mainland China from 2014 to 2019 using real-time quantitative PCR and conventional culture on respiratory and blood specimens showed an overall prevalence of Streptococcus pneumoniae of 7.43%, ranking sixth. However, its ranking varied from third to seventh among the nine cities. (2) Challenges and concerns about viruses have increased. National surveillance of acute respiratory tract infections and epidemiology in China from 2009 to 2019 indicated that the positivity rates for viral infections in adult pneumonia was 20.5%. These rates were similar to the results of the CDC's CAP pathogen study in the United States, although the rankings were different (viruses ranked second in China and first in the United States). Over the past 20 years, the emergence of new viral respiratory infections caused by mutant strains or zoonotic strains has significantly increased the challenges and threats posed by viral respiratory infections. (3) The role of Mycoplasma pneumoniae (M pneumoniae) in adult CAP and the need for routine empirical antibiotic coverage are controversial. In addition to the influence of epidemic cycles, the prevalence of M pneumoniae is influenced by factors such as age, season, study design, and detection methods, and geographical distribution is also an important influencing factor. Although M. pneumoniae ranks first among CAP pathogens in mainland China (11.05%), there are significant regional differences. In Beijing, Xi'an, and Changchun M. pneumoniae ranks first, while in Harbin, Nanjing, and Fuzhou it ranks second to sixth. In Wuhan, Shenzhen, and Chengdu M. pneumoniae ranks after the tenth position. Available evidence supports the notion that routine coverage of M. pneumoniae is not necessary for empirical treatment of CAP, except in severe cases. In regions with a high prevalence of M. pneumoniae, the decision to cover atypical pathogens in patients with mild to moderate CAP should be based on local data and individualized. (4) CAP caused by multidrug-resistant bacteria, especially multidrug-resistant Gram-negative bacilli (GNB), has become a concern. According to a systematic review of Chinese literature, Klebsiella pneumoniae accounted for 8.12% of adult CAP patients, ranking fifth, and Pseudomonas aeruginosa accounted for 4.7% (ninth). The China Antimicrobial Resistance Surveillance System (CARSS) reported an average resistance rate of 27.7% for Klebsiella pneumoniae to third-generation cephalosporins and a resistance rate of 10.0% to carbapenems in 2021. The average resistance rate of Pseudomonas aeruginosa to carbapenems was 16.6%. Early empirical treatment should consider predicting the resistance profile using a "locally validated risk factor" scoring system. (5) Co-infections are common but under-reported. The development of non-culture detection techniques over the past 40 years has significantly increased the detection rate of respiratory pathogens, especially viruses, leading to an increasing number of reports of bacterial-viral co-infections in CAP. It has been reported that co-infections account for 39% of severe CAP cases on ventilators in the ICU. Currently, there is inconsistency and confusion regarding the definition and concept of co-infection, the choice of detection techniques, and the differentiation between co-detection and co-infection. Many reports of co-infections in COVID-19 lacked pathogenic evidence, and some even listed "effective antibiotic treatment" as one of the diagnostic criteria for viral-bacterial co-infections, suggesting to some extent an overuse of antibiotics in COVID-19. Due to the diverse etiological spectrum of CAP between regions in the recent years, it is challenging to develop unified guidelines for the management of CAP in large countries. This article provides recommendations for the development of local guidelines for the diagnosis and treatment of CAP.

Pneumococcal hydrogen peroxide regulates host cell kinase activity.

Introduction: Protein kinases are indispensable reversible molecular switches that adapt and control protein functions during cellular processes requiring rapid responses to internal and external events. Bacterial infections can affect kinase-mediated phosphorylation events, with consequences for both innate and adaptive immunity, through regulation of antigen presentation, pathogen recognition, cell invasiveness and phagocytosis. Streptococcus pneumoniae (Spn), a human respiratory tract pathogen and a major cause of community-acquired pneumoniae, affects phosphorylation-based signalling of several kinases, but the pneumococcal mediator(s) involved in this process remain elusive. In this study, we investigated the influence of pneumococcal H2O2 on the protein kinase activity of the human lung epithelial H441 cell line, a generally accepted model of alveolar epithelial cells. Methods: We performed kinome analysis using PamGene microarray chips and protein analysis in Western blotting in H441 lung cells infected with Spn wild type (SpnWT) or with SpnΔlctOΔspxB -a deletion mutant strongly attenuated in H2O2 production- to assess the impact of pneumococcal hydrogen peroxide (H2O2) on global protein kinase activity profiles. Results: Our kinome analysis provides direct evidence that kinase activity profiles in infected H441 cells significantly vary according to the levels of pneumococcal H2O2. A large number of kinases in H441 cells infected with SpnWT are significantly downregulated, whereas this no longer occurs in cells infected with the mutant SpnΔlctOΔspxB strain, which lacks H2O2. In particular, we describe for the first time H2O2-mediated downregulation of Protein kinase B (Akt1) and activation of lymphocyte-specific tyrosine protein kinase (Lck) via H2O2-mediated phosphorylation.

Elixhauser comorbidity method in predicting death of Spanish inpatients with asplenia and pneumococcal pneumonia.

BACKGROUND: Pneumococcal pneumonia (PP) is a serious infection caused by Streptococcus pneumoniae (pneumococcus), with a wide spectrum of clinical manifestations. The aim of this study was to analyze the comorbidity factors that influenced the mortality in patients with asplenia according to PP. METHODS: Discharge reports from the Spanish Minimum Basic Data Set (MBDS) was used to retrospectively analyze patients with asplenia and PP, from 1997 to 2021. Elixhauser Comorbidity Index (ECI) was calculated to predict in-hospital mortality (IHM). RESULTS: 97,922 patients with asplenia were included and 381 cases of PP were identified. The average age for men was 63.87 years and for women 65.99 years. In all years, ECI was larger for splenectomized than for non-splenectomized patients, with men having a higher mean ECI than women. An association was found between risk factors ECI, splenectomy, age group, sex, pneumococcal pneumonia, and increased mortality (OR = 0.98; 95% CI: 0.97-0.99; p < 0.001). The IHM increased steadily with the number of comorbidities and index scores in 1997-2021. CONCLUSIONS: Asplenia remain a relevant cause of hospitalization in Spain. Comorbidities reflected a great impact in patients with asplenia and PP, which would mean higher risk of mortality.

Estimating geographical spread of Streptococcus pneumoniae within Israel using genomic data.

Understanding how pathogens spread across geographical space is fundamental for control measures such as vaccination. Streptococcus pneumoniae (the pneumococcus) is a respiratory bacterium responsible for a large proportion of infectious disease morbidity and mortality globally. Even in the post-vaccination era, the rates of invasive pneumococcal disease (IPD) remain stable in most countries, including Israel. To understand the geographical spread of the pneumococcus in Israel, we analysed 1174 pneumococcal genomes from patients with IPD across multiple regions. We included the evolutionary distance between pairs of isolates inferred using whole-genome data within a relative risk (RR) ratio framework to capture the geographical structure of S. pneumoniae. While we could not find geographical structure at the overall lineage level, the extra granularity provided by whole-genome sequence data showed that it takes approximately 5 years for invasive pneumococcal isolates to become fully mixed across the country.This article contains data hosted by Microreact.