Browsing by Author "Domenech Lucas, Mirian"
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Publication A national longitudinal study evaluating the activity of cefditoren and other antibiotics against non-susceptible Streptococcus pneumoniae strains during the period 2004-20 in Spain(Oxford University Press, 2022-03-31) Sempere, Julio; Gonzalez-Camacho, Fernando; Domenech Lucas, Mirian; Llamosi, Mirella; Del Rio Menendez, Idoia; Lopez Ruiz, Beatriz; Gimeno, Mercedes; Coronel, Pilar; Yuste, Jose Enrique; Ministerio de Economía, Industria y Competitividad (España); Ministerio de Ciencia e Innovación (España); Meiji Pharma SpainBackground: Surveillance studies including antibiotic resistance and evolution of pneumococcal serotypes are critical to evaluate the susceptibility of commonly used antibiotics and the contribution of conjugate vaccines against resistant strains. Objectives: To determine the susceptibility of clinical isolates of Streptococcus pneumoniae with reduced susceptibility to penicillin to a panel of antibiotics during the period 2004-20 and characterize the impact of pneumococcal conjugate vaccines in the evolution of resistant serotypes. Methods: We selected 3017 clinical isolates in order to determine the minimal inhibitory concentration to penicillin, amoxicillin, cefotaxime, erythromycin, levofloxacin and oral cephalosporins, including cefditoren, cefixime and cefpodoxime. Results: The antibiotics with the lowest proportion of resistant strains from 2004 to 2020 were cefditoren (<0.4%), followed by cefotaxime (<5%), penicillin (<6.5%) and levofloxacin (<7%). Among oral cephalosporins, cefixime was the cephalosporin with the highest MIC90 (32 mg/L) and MIC50 (8-16 mg/L) throughout the study, followed by cefpodoxime with highest values of MIC90 (4 mg/L) and MIC50 (2 mg/L) for the majority of the study period. In contrast, cefditoren was the cephalosporin with the lowest MIC90 (1 mg/L) and MIC50 (0.25-0.5 mg/L). Conclusions: Cefditoren was the antibiotic with the highest proportion of susceptible strains. Hence, more than 80% of the clinical strains were susceptible to cefditoren throughout the period 2004-20. The proportion of resistant isolates to cefditoren and cefotaxime was scarce, being less than 0.4% for cefditoren and lower than 5% for cefotaxime, despite the increased rates of serotypes not covered by the 13-valent pneumococcal conjugate vaccine.Publication Antibacterial activity of a DNA topoisomerase I inhibitor versus fluoroquinolones in Streptococcus pneumoniae.(Public Library of Science (PLOS), 2020) Valenzuela, Myriam V; Domenech Lucas, Mirian; Mateos-Martínez, Patricia; Gonzalez-Camacho, Fernando; de la Campa, Adela G; Garcia, Maria Teresa; Ministerio de Economía y Competitividad (España)The DNA topoisomerase complement of Streptococcus pneumoniae is constituted by two type II enzymes (topoisomerase IV and gyrase), and a single type I enzyme (topoisomerase I). These enzymes maintain the DNA topology, which is essential for replication and transcription. While fluoroquinolones target the type II enzymes, seconeolitsine, a new antimicrobial agent, targets topoisomerase I. We compared for the first time the in vitro effect of inhibition of topoisomerase I by seconeolitsine and of the type II topoisomerases by the fluoroquinolones levofloxacin and moxifloxacin. We used three isogenic non-encapsulated strains and five non-vaccine serotypes isolates belonging to two circulating pneumococcal clones, ST638 (2 strains) and ST1569V (3 strains). Each group contained strains with diverse susceptibility to fluoroquinolones. Minimal inhibitory concentrations, killing curves and postantibiotic effects were determined. Seconeolitsine demonstrated the fastest and highest bactericidal activity against planktonic bacteria and biofilms. When fluoroquinolone-susceptible planktonic bacteria were considered, seconeolitsine induced postantibiotic effects (1.00-1.87 h) similar than levofloxacin (1.00-2.22 h), but longer than moxifloxacin (0.39-1.71 h). The same effect was observed in sessile bacteria forming biofilms. Seconeolitsine induced postantibiotic effects (0.84-2.31 h) that were similar to those of levofloxacin (0.99-3.32 h) but longer than those of moxifloxacin (0.89-1.91 h). The greatest effect was observed in the viability and adherence of bacteria in the postantibiotic phase. Seconeolitsine greatly reduced the thickness of the biofilms formed in comparison with fluoroquinolones: 2.91 ± 0.43 μm (seconeolitsine), 7.18 ± 0.58 μm (levofloxacin), 17.08 ± 1.02 μm (moxifloxacin). When fluoroquinolone-resistant bacteria were considered, postantibiotic effects induced by levofloxacin and moxifloxacin, but not by seconeolitsine, were shorter, decreasing up to 5-fold (levofloxacin) or 2-fold (moxifloxacin) in planktonic cells, and up to 1.7 (levofloxacin) or 1.4-fold (moxifloxacin) during biofilm formation. Therefore, topoisomerase I inhibitors could be an alternative for the treatment of pneumococcal diseases, including those caused by fluoroquinolone-resistant isolates.Publication Clearance of mixed biofilms of Streptococcus pneumoniae and methicillin-susceptible/resistant Staphylococcus aureus by antioxidants N-acetyl-L-cysteine and cysteamine(Nature Publishing Group, 2022-04-23) Sempere, Julio; Llamosi, Mirella; Roman, Federico; Lago, Dario; Gonzalez-Camacho, Fernando; Perez-Garcia, Covadonga; Yuste, Jose Enrique; Domenech Lucas, Mirian; Ministerio de Ciencia e Innovación (España); Ministerio de Economía, Industria y Competitividad (España)Biofilm-associated infections are of great concern because they are associated with antibiotic resistance and immune evasion. Co-colonization by Staphylococcus aureus and Streptococcus pneumoniae is possible and a threat in clinical practice. We investigated the interaction between S. aureus and S. pneumoniae in mixed biofilms and tested new antibiofilm therapies with antioxidants N-acetyl-L-cysteine (NAC) and cysteamine (Cys). We developed two in vitro S. aureus-S. pneumoniae mixed biofilms in 96-well polystyrene microtiter plates and we treated in vitro biofilms with Cys and NAC analyzing their effect by CV staining and viable plate counting. S. pneumoniae needed a higher proportion of cells in the inoculum and planktonic culture to reach a similar population rate in the mixed biofilm. We demonstrated the effect of Cys in preventing S. aureus biofilms and S. aureus-S. pneumoniae mixed biofilms. Moreover, administration of 5 mg/ml of NAC nearly eradicated the S. pneumoniae population and killed nearly 94% of MSSA cells and 99% of MRSA cells in the mixed biofilms. The methicillin resistance background did not change the antioxidants effect in S. aureus. These results identify NAC and Cys as promising repurposed drug candidates for the prevention and treatment of mixed biofilms by S. pneumoniae and S. aureus.Publication Clinical Relevance and Molecular Pathogenesis of the Emerging Serotypes 22F and 33F of Streptococcus pneumoniae in Spain(Frontiers Media, 2020) Sempere, Julio; Miguel, Sara de; Gonzalez-Camacho, Fernando; Yuste, Jose Enrique; Domenech Lucas, Mirian; Ministerio de Economía, Industria y Competitividad (España); Centro de Investigación Biomedica en Red - CIBER; Instituto de Salud Carlos IIIStreptococcus pneumoniae is the main bacterial cause of respiratory infections in children and the elderly worldwide. Serotype replacement is a frequent phenomenon after the introduction of conjugated vaccines, with emerging serotypes 22F and 33F as frequent non-PCV13 serotypes in children and adults in North America and other countries. Characterization of mechanisms involved in evasion of the host immune response by these serotypes is of great importance in public health because they are included in the future conjugated vaccines PCV15 and PCV20. One of the main strategies of S. pneumoniae to persistently colonize and causes infection is biofilm formation. In this study, we have evaluated the influence of capsule polysaccharide in biofilm formation and immune evasion by using clinical isolates from different sources and isogenic strains with capsules from prevalent serotypes. Since the introduction of PCV13 in Spain in the year 2010, isolates of serotypes 22F and 33F are rising among risk populations. The predominant circulating genotypes are ST43322F and ST71733F , being CC433 in 22F and CC717 in 33F the main clonal complexes in Spain. The use of clinical isolates of different origin, demonstrated that pediatric isolates of serotypes 22F and 33F formed better biofilms than adult isolates and this was statistically significant. This phenotype was greater in clinical isolates from blood origin compared to those from cerebrospinal fluid, pleural fluid and otitis. Opsonophagocytosis assays showed that serotype 22F and 33F were recognized by the PSGL-1 receptor on leukocytes, although serotype 22F, was more resistant than serotype 33F to phagocytosis killing and more lethal in a mouse sepsis model. Overall, the emergence of additional PCV15 serotypes, especially 22F, could be associated to an enhanced ability to divert the host immune response that markedly increased in a biofilm state. Our findings demonstrate that pediatric isolates of 22F and 33F, that form better biofilm than isolates from adults, could have an advantage to colonize the nasopharynx of children and therefore, be important in carriage and subsequent dissemination to the elderly. The increased ability of serotype 22F to avoid the host immune response, might explain the emergence of this serotype in the last years.Publication Combination of Antibodies and Antibiotics as a Promising Strategy Against Multidrug-Resistant Pathogens of the Respiratory Tract(Frontiers Media, 2018) Domenech Lucas, Mirian; Sempere, Julio; Miguel, Sara de; Yuste, Jose Enrique; Ministerio de Economía, Industria y Competitividad (España)The emergence of clinical isolates associated to multidrug resistance is a serious threat worldwide in terms of public health since complicates the success of the antibiotic treatment and the resolution of the infectious process. This is of great concern in pathogens affecting the lower respiratory tract as these infections are one of the major causes of mortality in children and adults. In most cases where the respiratory pathogen is associated to multidrug-resistance, antimicrobial concentrations both in serum and at the site of infection may be insufficient and the resolution of the infection depends on the interaction of the invading pathogen with the host immune response. The outcome of these infections largely depends on the susceptibility of the pathogen to the antibiotic treatment, although the humoral and cellular immune responses also play an important role in this process. Hence, prophylactic measures or even immunotherapy are alternatives against these multi-resistant pathogens. In this sense, specific antibodies and antibiotics may act concomitantly against the respiratory pathogen. Alteration of cell surface structures by antimicrobial drugs even at sub-inhibitory concentrations might result in greater exposure of microbial ligands that are normally hidden or hardly exposed. This alteration of the bacterial envelope may stimulate opsonization by natural and/or specific antibodies or even by host defense components, increasing the recognition of the microbial pathogen by circulating phagocytes. In this review we will explain the most relevant studies, where vaccination or the use of monoclonal antibodies in combination with antimicrobial treatment has demonstrated to be an alternative strategy to overcome the impact of multidrug resistance in respiratory pathogens.Publication Combination of Cefditoren and N-acetyl-l-Cysteine Shows a Synergistic Effect against Multidrug-Resistant Streptococcus pneumoniae Biofilms(American Society for Microbiology (ASM), 2022-12-21) Llamosi, Mirella; Sempere, Julio; Coronel, Pilar; Gimeno, Mercedes; Yuste, Jose Enrique; Domenech Lucas, Mirian; Ministerio de Ciencia e Innovación (España); Meiji Pharma Spain; Pfizer; Merck, Sharp & DohmeBiofilm formation by Streptococcus pneumoniae is associated with colonization of the upper respiratory tract, including the carrier state, and with chronic respiratory infections in patients suffering from chronic obstructive pulmonary disease (COPD). The use of antibiotics alone to treat recalcitrant infections caused by biofilms is insufficient in many cases, requiring novel strategies based on a combination of antibiotics with other agents, including antibodies, enzybiotics, and antioxidants. In this work, we demonstrate that the third-generation oral cephalosporin cefditoren (CDN) and the antioxidant N-acetyl-l-cysteine (NAC) are synergistic against pneumococcal biofilms. Additionally, the combination of CDN and NAC resulted in the inhibition of bacterial growth (planktonic and biofilm cells) and destruction of the biofilm biomass. This marked antimicrobial effect was also observed in terms of viability in both inhibition (prevention) and disaggregation (treatment) assays. Moreover, the use of CDN and NAC reduced bacterial adhesion to human lung epithelial cells, confirming that this strategy of combining these two compounds is effective against resistant pneumococcal strains colonizing the lung epithelium. Finally, administration of CDN and NAC in mice suffering acute pneumococcal pneumonia caused by a multidrug-resistant strain was effective in clearing the bacteria from the respiratory tract in comparison to treatment with either compound alone. Overall, these results demonstrate that the combination of oral cephalosporins and antioxidants, such as CDN and NAC, respectively, is a promising strategy against respiratory biofilms caused by S. pneumoniae. IMPORTANCE Streptococcus pneumoniae is one of the deadliest bacterial pathogens, accounting for up to 2 million deaths annually prior to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Vaccines have decreased the burden of diseases produced by S. pneumoniae, but the rise of antibiotic-resistant strains and nonvaccine serotypes is worrisome. Pneumococcal biofilms are associated with chronic respiratory infections, and treatment is challenging, making the search for new antibiofilm therapies a priority as biofilms become resistant to traditional antibiotics. In this work, we used the combination of an antibiotic (CDN) and an antioxidant (NAC) to treat the pneumococcal biofilms of relevant clinical isolates. We demonstrated a synergy between CDN and NAC that inhibited and treated pneumococcal biofilms, impaired pneumococcal adherence to the lung epithelium, and treated pneumonia in a mouse pneumonia model. We propose the widely used cephalosporin CDN and the repurposed drug NAC as a new antibiofilm therapy against S. pneumoniae biofilms, including those formed by antibiotic-resistant clinical isolates.Publication Effect of pneumococcal conjugate vaccines and SARS-CoV-2 on antimicrobial resistance and the emergence of Streptococcus pneumoniae serotypes with reduced susceptibility in Spain, 2004-20: a national surveillance study(Elsevier, 2022-10) Sempere, Julio; Llamosi, Mirella; Lopez Ruiz, Beatriz; Del Rio Menendez, Idoia; Perez-Garcia, Covadonga; Lago, Dario; Gimeno, Mercedes; Coronel, Pilar; Gonzalez-Camacho, Fernando; Domenech Lucas, Mirian; Yuste, Jose Enrique; Instituto de Salud Carlos III; Ministerio de Ciencia e Innovación (España); Meiji Pharma SpainBackground: Epidemiological studies are necessary to explore the effect of current pneumococcal conjugate vaccines (PCVs) against antibiotic resistance, including the rise of non-vaccine serotypes that are resistant to antibiotics. Hence, epidemiological changes in the antimicrobial pattern of Streptococcus pneumoniae before and during the first year of the COVID-19 pandemic were studied. Methods: In this national surveillance study, we characterised the antimicrobial susceptibility to a panel of antibiotics in 3017 pneumococcal clinical isolates with reduced susceptibility to penicillin during 2004-20 in Spain. This study covered the early and late PCV7 periods; the early, middle, and late PCV13 periods; and the first year of the COVID-19 pandemic, to evaluate the contribution of PCVs and the pandemic to the emergence of non-vaccine serotypes associated with antibiotic resistance. Findings: Serotypes included in PCV7 and PCV13 showed a decline after the introduction of PCVs in Spain. However, an increase in non-PCV13 serotypes (mainly 11A, 24F, and 23B) that were not susceptible to penicillin promptly appeared. A rise in the proportion of pneumococcal strains with reduced susceptibility to β-lactams and erythromycin was observed in 2020, coinciding with the emergence of SARS-CoV-2. Cefditoren was the β-lactam with the lowest minimum inhibitory concentration (MIC)50 or MIC90 values, and had the highest proportion of susceptible strains throughout 2004-20. Interpretation: The increase in non-PCV13 serotypes associated with antibiotic resistance is concerning, especially the increase of penicillin resistance linked to serotypes 11A and 24F. The future use of PCVs with an increasingly broad spectrum (such as PCV20, which includes serotype 11A) could reduce the impact of antibiotic resistance for non-PCV13 serotypes. The use of antibiotics to prevent co-infections in patients with COVID-19 might have affected the increased proportion of pneumococcal-resistant strains. Cefotaxime as a parenteral option, and cefditoren as an oral choice, were the antibiotics with the highest activity against non-PCV20 serotypes.Publication Emergence of Amoxicillin-Resistant Variants of Spain9V-ST156 Pneumococci Expressing Serotype 11A Correlates with Their Ability to Evade the Host Immune Response(Public Library of Science (PLOS), 2015) Aguinagalde, Leire; Corsini, Bruno; Domenech, Arnau; Domenech Lucas, Mirian; Cámara, Jordi; Ardanuy, Carmen; García, Ernesto; Liñares, Josefina; Fenoll, Asuncion; Yuste, Jose EnriqueCapsular switching allows pre-existing clones of Streptococcus pneumoniae expressing vaccine serotypes to escape the vaccine-induced immunity by acquisition of capsular genes from pneumococci of a non-vaccine serotype. Here, we have analysed the clonal composition of 492 clinical isolates of serotype 11A causing invasive disease in Spain (2000-2012), and their ability to evade the host immune response. Antibiograms, serotyping and molecular typing were performed. The restriction profiles of pbp2x, pbp1a and pbp2b genes were also analysed. Interaction with the complement components C1q, C3b, C4BP, and factor H was explored whereas opsonophagocytosis assays were performed using a human cell line differentiated to neutrophils. Biofilm formation and the polymorphisms of the major autolysin LytA were evaluated. The main genotypes of the 11A pneumococci were: ST62 (447 isolates, 90.6%), followed by ST6521 (35 isolates, 7.3%) and ST838 (10 isolates, 2.1%). Beta lactam resistant serotype 11A variants of genotypes ST838 and ST6521 closely related to the Spain9V-ST156 clone were first detected in 2005. A different pattern of evasion of complement immunity and phagocytosis was observed between genotypes. The emergence of one vaccine escape variant of Spain9V-ST156 (ST652111A), showing a high potential to avoid the host immune response, was observed. In addition, isolates of ST652111A showed higher ability to produce biofilms than ST83811A or ST6211A, which may have contributed to the emergence of this PEN-resistant ST652111A genotype in the last few years. The emergence of penicillin-resistant 11A invasive variants of the highly successful ST156 clonal complex merits close monitoring.Publication Emerging, Non-PCV13 Serotypes 11A and 35B of Streptococcus pneumoniae Show High Potential for Biofilm Formation In Vitro(Public Library of Science (PLOS), 2015-04-30) Domenech Lucas, Mirian; Damián, Diana; Ardanuy, Carmen; Liñares, Josefina; Fenoll, Asuncion; García, Ernesto; Ministerio de Economía y Competitividad (España); Instituto de Salud Carlos IIIBACKGROUND: Since the use of pneumococcal conjugate vaccines PCV7 and PCV13 in children became widespread, invasive pneumococcal disease (IPD) has dramatically decreased. Nevertheless, there has been a rise in incidence of Streptococcus pneumoniae non-vaccine serotypes (NVT) colonising the human nasopharynx. Nasopharyngeal colonisation, an essential step in the development of S. pneumoniae-induced IPD, is associated with biofilm formation. Although the capsule is the main pneumococcal virulence factor, the formation of pneumococcal biofilms might, in fact, be limited by the presence of capsular polysaccharide (CPS). METHODOLOGY/PRINCIPAL FINDINGS: We used clinical isolates of 16 emerging, non-PCV13 serotypes as well as isogenic transformants of the same serotypes. The biofilm formation capacity of isogenic transformants expressing CPSs from NVT was evaluated in vitro to ascertain whether this trait can be used to predict the emergence of NVT. Fourteen out of 16 NVT analysed were not good biofilm formers, presumably because of the presence of CPS. In contrast, serotypes 11A and 35B formed ≥45% of the biofilm produced by the non-encapsulated M11 strain. CONCLUSIONS/SIGNIFICANCE: This study suggest that emerging, NVT serotypes 11A and 35B deserve a close surveillance.Publication Impact of Pneumococcal Vaccination in the Nasopharyngeal Carriage of Streptococcus pneumoniae in Healthy Children of the Murcia Region in Spain(Multidisciplinary Digital Publishing Institute (MDPI), 2020-12-28) Alfayate Miguélez, Santiago; Yague Guirao, Genoveva; Menasalvas Ruíz, Ana I; Sanchez-Solís, Manuel; Gonzalez-Camacho, Fernando; Ortíz Romero, M Mar; Espejo García, Pilar; Guerrero Gómez, Carmen; Iofrío de Arce, Antonio; Moreno Parrado, Laura; Sánchez Andrada, Rosa M; Cascales Alcolea, Eva; Lorente García, Sebastián; Paredes Reyes, Pedro; Casquet Barceló, Ángela; López Yepes, M Luisa; Vigueras Abellán, Juan José; Sanz Mateo, Gonzalo; Domenech Lucas, Mirian; Murcian Pneumococcal Study Group; Ministerio de Economía y Competitividad (España)An epidemiological study of Streptococcus pneumoniae nasopharyngeal carriage in healthy children was carried out five years after the introduction of the 13-valent pneumococcal conjugate vaccine (PCV13). Study the impact of pediatric vaccination with PCV13, and other associated epidemiological factors on the status of nasopharyngeal carriage, the circulating pneumococcal serotypes, and the antibiotic susceptibility to more frequently used antibiotics. A multi-center study was carried out in Primary Health Care, which included 1821 healthy children aged 1 to 4 years old. All isolates were sent to the Spanish Pneumococcal Reference Laboratory for serotyping and antimicrobial susceptibility testing. At least one dose of PCV13 had been received by 71.9% of children and carriage pneumococcal prevalence was 19.7%. The proportion of PCV13 serotypes was low (14.4%), with an observed predominance of non-vaccine serotypes, 23B, 11A, 10A, 35B/F, and 23A were the five most frequent. A high rate of resistance to penicillin, erythromycin, and trimethoprim sulfamethoxazole was found. A low proportion of PCV13 serotypes were detected, confirming the impact of pediatric vaccination for reducing the serotypes vaccine carriage. High resistance rates to clinically important antibiotics were observed.Publication Nationwide trends of invasive pneumococcal disease in Spain (2009-2019) in children and adults during the pneumococcal conjugate vaccine era.(Oxford University Press, 2020-09-29) Miguel, Sara de; Domenech Lucas, Mirian; Gonzalez-Camacho, Fernando; Sempere, Julio; Vicioso-Perez, Maria Dolores; Sanz, Juan Carlos; García Comas, Luis; Ardanuy, Carmen; Fenoll, Asuncion; Yuste, Jose Enrique; Ministerio de Economía, Industria y Competitividad (España)Introduction of pneumococcal conjugate vaccines (PCVs) has shown a marked reduction in the disease caused by vaccine serotypes in children providing herd protection to the elderly group. However, the emergence of non-vaccine serotypes is of great concern worldwide. This study includes national laboratory data from invasive pneumococcal disease (IPD) cases affecting pediatric and adult population during 2009-2019. The impact of implementing different vaccine strategies for immunocompetent adults comparing Spanish regions using PCV13 vs regions using PPV23 vaccine was also analyzed for 2017-2019. The overall reductions of IPD cases by PCV13 serotypes in children and adults were 88% and 59% respectively during 2009-2019 with a constant increase of serotype 8 in adults since 2015. IPD cases by additional serotypes covered by PPV23 increased from 20% in 2009 to 52% in 2019. In children, serotype 24F was the most frequent in 2019 whereas in adults, serotypes 3 and 8 accounted for 36% of IPD cases. Introduction of PCV13 or PPV23 in the adult calendar of certain Spanish regions reduced up to 25% and 11% respectively the IPD cases by PCV13 serotypes, showing a decrease of serotype 3 when PCV13 was used. Use of PCV13 in children has shown a clear impact in pneumococcal epidemiology reducing the burden of IPD in children but also in adults by herd protection although the increase of serotype 8 in adults is worrisome. Vaccination with PCV13 in immunocompetent adults seems to control IPD cases by PCV13 serotypes including serotype 3.Publication New live attenuated tuberculosis vaccine MTBVAC induces trained immunity and confers protection against experimental lethal pneumonia(Public Library of Science (PLOS), 2020-04) Tarancón, Raquel; Domínguez-Andrés, Jorge; Uranga, Santiago; Ferreira, Anaísa V; Groh, Laszlo A; Domenech Lucas, Mirian; Gonzalez-Camacho, Fernando; Riksen, Niels P; Aguilo, Nacho; Yuste, Jose Enrique; Martín, Carlos; Netea, Mihai G; Dutch Research Council (Holanda); Ministerio de Ciencia y Universidades (España); Unión Europea. Comisión Europea. European Research Council (ERC)Among infectious diseases, tuberculosis is the leading cause of death worldwide, and represents a serious threat, especially in developing countries. The protective effects of Bacillus Calmette-Guerin (BCG), the current vaccine against tuberculosis, have been related not only to specific induction of T-cell immunity, but also with the long-term epigenetic and metabolic reprogramming of the cells from the innate immune system through a process termed trained immunity. Here we show that MTBVAC, a live attenuated strain of Mycobacterium tuberculosis, safe and immunogenic against tuberculosis antigens in adults and newborns, is also able to generate trained immunity through the induction of glycolysis and glutaminolysis and the accumulation of histone methylation marks at the promoters of proinflammatory genes, facilitating an enhanced response after secondary challenge with non-related bacterial stimuli. Importantly, these findings in human primary myeloid cells are complemented by a strong MTBVAC-induced heterologous protection against a lethal challenge with Streptococcus pneumoniae in an experimental murine model of pneumonia.Publication Pleiotropic effects of cell wall amidase LytA on Streptococcus pneumoniae sensitivity to the host immune response(American Society for Microbiology (ASM), 2015-02) Ramos-Sevillano, Elisa; Urzainqui, Ana; Campuzano, Susana; Moscoso, Miriam; Gonzalez-Camacho, Fernando; Domenech Lucas, Mirian; Rodríguez de Córdoba, Santiago; Sánchez Madrid, Francisco; Brown, Jeremy S; García, Ernesto; Yuste, Jose Enrique; Ministerio de Economía y Competitividad (España); Instituto de Salud Carlos IIIThe complement system is a key component of the host immune response for the recognition and clearance of Streptococcus pneumoniae. In this study, we demonstrate that the amidase LytA, the main pneumococcal autolysin, inhibits complement-mediated immunity independently of effects on pneumolysin by a complex process of impaired complement activation, increased binding of complement regulators, and direct degradation of complement C3. The use of human sera depleted of either C1q or factor B confirmed that LytA prevented activation of both the classical and alternative pathways, whereas pneumolysin inhibited only the classical pathway. LytA prevented binding of C1q and the acute-phase protein C-reactive protein to S. pneumoniae, thereby reducing activation of the classical pathway on the bacterial surface. In addition, LytA increased recruitment of the complement downregulators C4BP and factor H to the pneumococcal cell wall and directly cleaved C3b and iC3b to generate degradation products. As a consequence, C3b deposition and phagocytosis increased in the absence of LytA and were markedly enhanced for the lytA ply double mutant, confirming that a combination of LytA and Ply is essential for the establishment of pneumococcal pneumonia and sepsis in a murine model of infection. These data demonstrate that LytA has pleiotropic effects on complement activation, a finding which, in combination with the effects of pneumolysin on complement to assist with pneumococcal complement evasion, confirms a major role of both proteins for the full virulence of the microorganism during septicemia.Publication Pneumococcal Choline-Binding Proteins Involved in Virulence as Vaccine Candidates(Multidisciplinary Digital Publishing Institute (MDPI), 2021-02-20) Sempere, Julio; Llamosi, Mirella; Del Rio Menendez, Idoia; Lopez Ruiz, Beatriz; Domenech Lucas, Mirian; Gonzalez-Camacho, Fernando; Ministerio de Economía, Industria y Competitividad (España)Streptococcus pneumoniae is a pathogen responsible for millions of deaths worldwide. Currently, the available vaccines for the prevention of S. pneumoniae infections are the 23-valent pneumococcal polysaccharide-based vaccine (PPV-23) and the pneumococcal conjugate vaccines (PCV10 and PCV13). These vaccines only cover some pneumococcal serotypes (up to 100 different serotypes have been identified) and are unable to protect against non-vaccine serotypes and non-encapsulated pneumococci. The emergence of antibiotic-resistant non-vaccine serotypes after these vaccines is an increasing threat. Therefore, there is an urgent need to develop new pneumococcal vaccines which could cover a wide range of serotypes. One of the vaccines most characterized as a prophylactic alternative to current PPV-23 or PCVs is a vaccine based on pneumococcal protein antigens. The choline-binding proteins (CBP) are found in all pneumococcal strains, giving them the characteristic to be potential vaccine candidates as they may protect against different serotypes. In this review, we have focused the attention on different CBPs as vaccine candidates because they are involved in the pathogenesis process, confirming their immunogenicity and protection against pneumococcal infection. The review summarizes the major contribution of these proteins to virulence and reinforces the fact that antibodies elicited against many of them may block or interfere with their role in the infection process.Publication PSGL-1 on Leukocytes is a Critical Component of the Host Immune Response against Invasive Pneumococcal Disease(Public Library of Science (PLOS), 2016-03-14) Ramos-Sevillano, Elisa; Urzainqui, Ana; Andres, Belen de; González-Tajuelo, Rafael; Domenech Lucas, Mirian; Gonzalez-Camacho, Fernando; Sánchez Madrid, Francisco; Brown, Jeremy S; García, Ernesto; Yuste, Jose Enrique; Ministerio de Economía y Competitividad (España)Bacterial uptake by phagocytic cells is a vital event in the clearance of invading pathogens such as Streptococcus pneumoniae. A major role of the P-selectin glycoprotein ligand-1 (PSGL-1) on leukocytes against invasive pneumococcal disease is described in this study. Phagocytosis experiments using different serotypes demonstrated that PSGL-1 is involved in the recognition, uptake and killing of S. pneumoniae. Co-localization of several clinical isolates of S. pneumoniae with PSGL-1 was demonstrated, observing a rapid and active phagocytosis in the presence of PSGL-1. Furthermore, the pneumococcal capsular polysaccharide and the main autolysin of the bacterium--the amidase LytA--were identified as bacterial ligands for PSGL-1. Experimental models of pneumococcal disease including invasive pneumonia and systemic infection showed that bacterial levels were markedly increased in the blood of PSGL-1-/- mice. During pneumonia, PSGL-1 controls the severity of pneumococcal dissemination from the lung to the bloodstream. In systemic infection, a major role of PSGL-1 in host defense is to clear the bacteria in the systemic circulation controlling bacterial replication. These results confirmed the importance of this receptor in the recognition and clearance of S. pneumoniae during invasive pneumococcal disease. Histological and cellular analysis demonstrated that PSGL-1-/- mice have increased levels of T cells migrating to the lung than the corresponding wild-type mice. In contrast, during systemic infection, PSGL-1-/- mice had increased numbers of neutrophils and macrophages in blood, but were less effective controlling the infection process due to the lack of this functional receptor. Overall, this study demonstrates that PSGL-1 is a novel receptor for S. pneumoniae that contributes to protection against invasive pneumococcal disease.Publication Pulmonary BCG induces lung-resident macrophage activation and confers long-term protection against tuberculosis(American Association for the Advancement of Science (AAAS), 2021-09-24) Mata, Elena; Tarancon, Raquel; Guerrero, Claudia; Moreo, Eduardo; Moreau, Flavie; Uranga, Santiago; Gomez, Ana Belen; Marinova, Dessislava; Domenech Lucas, Mirian; Gonzalez-Camacho, Fernando; Monzon, Marta; Badiola, Juan; Dominguez-Andres, Jorge; Yuste, Jose Enrique; Anel, Alberto; Peixoto, Antonio; Martin, Carlos; Aguilo, Nacho; Ministerio de Ciencia, Innovación y Universidades (España); Gobierno de Aragón (España); Unión Europea. Fondo Europeo de Desarrollo Regional (FEDER/ERDF)Bacillus Calmette-Guerin (BCG) is an attenuated bacterial vaccine used to protect against Mycobacterium tuberculosis (Mtb) in regions where infections are highly prevalent. BCG is currently delivered by the intradermal route, but alternative routes of administration are of great interest, including intrapulmonary delivery to more closely mimic respiratory Mtb infection. In this study, mice subjected to pulmonary delivery of green fluorescent protein–tagged strains of virulent (Mtb) and attenuated (BCG) mycobacteria were studied to better characterize infected lung cell subsets. Profound differences in dissemination patterns were detected between Mtb and BCG, with a strong tendency of Mtb to disseminate from alveolar macrophages (AMs) to other myeloid subsets, mainly neutrophils and recruited macrophages. BCG mostly remained in AMs, which promoted their activation. These preactivated macrophages were highly efficient in containing Mtb bacilli upon challenge and disrupting early bacterial dissemination, which suggests a potential mechanism of protection associated with pulmonary BCG vaccination. Respiratory BCG also protected mice against a lethal Streptococcus pneumoniae challenge, suggesting that BCG-induced innate activation could confer heterologous protection against respiratory pathogens different from Mtb. BCG drove long-term activation of AMs, even after vaccine clearance, and these AMs reacted efficiently upon subsequent challenge. These results suggest the generation of a trained innate memory-like response in AMs induced by pulmonary BCG vaccination.Publication Surveillance of invasive pneumococcal disease in Spain exploring the impact of the COVID-19 pandemic (2019-2023)(Elsevier, 2024-06-19) Perez-Garcia, Covadonga; Sempere, Julio; Miguel, Sara de; Hita, Samantha; Úbeda, Aída; Vidal-Alcántara, Erick Joan; Llorente, Joaquín; Limia, Aurora; Gil-de-Miguel, Ángel; Sanz, Juan Carlos; Martinón-Torres, Federico; Ardanuy, Carmen; Domenech Lucas, Mirian; Yuste, Jose Enrique; Ministerio de Ciencia e Innovación (España); Instituto de Salud Carlos III; Merck, Sharp & DohmeObjectives: Dynamic trends of invasive pneumococcal disease (IPD) including the evolution of prevalent serotypes are very useful to evaluate the impact of current and future pneumococcal conjugate vaccines (PCVs) and the rise of non-vaccine serotypes. In this study, we include epidemiological patterns of S. pneumoniae before and after COVID-19 pandemic. Methods: We characterized all national IPD isolates from children and adults received at the Spanish Pneumococcal Reference Laboratory during 2019-2023. Results: In the first pandemic year 2020, we found a general reduction in IPD cases across all age groups, followed by a partial resurgence in children in 2021 but not in adults. By 2022, IPD cases in children had returned to pre-pandemic levels, and partially in adults. In 2023, IPD rates surpassed those of the last pre-pandemic year. Notably, the emergence of serotype 3 is of significant concern, becoming the leading cause of IPD in both pediatric and adult populations over the last two years (2022-2023). Increase of serotype 4 in young adults occurred in the last epidemiological years. Conclusions: The COVID-19 pandemic led to a temporary decline in all IPD cases during 2020 attributable to non-pharmaceutical interventions followed by a subsequent rise. Employing PCVs with broader coverage and/or enhanced immunogenicity may be critical to mitigate the marked increase of IPD.Publication Trends in invasive bacterial diseases during the first 2 years of the COVID-19 pandemic: analyses of prospective surveillance data from 30 countries and territories in the IRIS Consortium(Elsevier, 2023-09) Shaw, David; Abad, Raquel; Amin-Chowdhury, Zahin; Bautista, Adriana; Bennett, Desiree; Broughton, Karen; Cao, Bin; Casanova, Carlo; Choi, Eun Hwa; Chu, Yiu-Wai; Claus, Heike; Coelho, Juliana; Corcoran, Mary; Cottrell, Simon; Cunney, Robert; Cuypers, Lize; Dalby, Tine; Davies, Heather; de Gouveia, Linda; Deghmane, Ala-Eddine; Demczuk, Walter; Desmet, Stefanie; Domenech Lucas, Mirian; Drew, Richard; du Plessis, Mignon; Duarte, Carolina; Erlendsdóttir, Helga; Fry, Norman K; Fuursted, Kurt; Hale, Thomas; Henares, Desiree; Henriques-Normark, Birgitta; Hilty, Markus; Hoffmann, Steen; Humphreys, Hilary; Ip, Margaret; Jacobsson, Susanne; Johnson, Christopher; Johnston, Jillian; Jolley, Keith A; Kawabata, Aníbal; Kozakova, Jana; Kristinsson, Karl G; Krizova, Pavla; Kuch, Alicja; Ladhani, Shamez; Lâm, Thiên-Trí; León, María Eugenia; Lindholm, Laura; Litt, David; Maiden, Martin C J; Martin, Irene; Martiny, Delphine; Mattheus, Wesley; McCarthy, Noel D; Meehan, Mary; Meiring, Susan; Mölling, Paula; Morfeldt, Eva; Morgan, Julie; Mulhall, Robert; Muñoz-Almagro, Carmen; Murdoch, David; Murphy, Joy; Musilek, Martin; Mzabi, Alexandre; Novakova, Ludmila; Oftadeh, Shahin; Perez-Argüello, Amaresh; Perez-Vazquez, Maria; Perrin, Monique; Perry, Malorie; Prevost, Benoit; Roberts, Maria; Rokney, Assaf; Ron, Merav; Sanabria, Olga Marina; Scott, Kevin J; Sheppard, Carmen; Siira, Lotta; Sintchenko, Vitali; Skoczyńska, Anna; Sloan, Monica; Slotved, Hans-Christian; Smith, Andrew J; Steens, Anneke; Taha, Muhamed-Kheir; Toropainen, Maija; Tzanakaki, Georgina; Vainio, Anni; van der Linden, Mark P G; van Sorge, Nina M; Varon, Emmanuelle; Vohrnova, Sandra; von Gottberg, Anne; Yuste, Jose Enrique; Zanella, Rosemeire; Zhou, Fei; Brueggemann, Angela B; Wellcome Trust; NIHR - Oxford Biomedical Research Centre (Reino Unido); Ministerio de Ciencia e Innovación (España); Korea Disease Control and Prevention Agency (Corea del Sur); Torsten Söderbergs Stiftelse; Swedish Research Council; Federal Ministry for Health (Alemania); Pfizer; Merck KGaA; National Public Health Organization (Grecia); Stockholm County Council; Robert Koch InstituteBackground: The Invasive Respiratory Infection Surveillance (IRIS) Consortium was established to assess the impact of the COVID-19 pandemic on invasive diseases caused by Streptococcus pneumoniae, Haemophilus influenzae, Neisseria meningitidis, and Streptococcus agalactiae. We aimed to analyse the incidence and distribution of these diseases during the first 2 years of the COVID-19 pandemic compared to the 2 years preceding the pandemic. Methods: For this prospective analysis, laboratories in 30 countries and territories representing five continents submitted surveillance data from Jan 1, 2018, to Jan 2, 2022, to private projects within databases in PubMLST. The impact of COVID-19 containment measures on the overall number of cases was analysed, and changes in disease distributions by patient age and serotype or group were examined. Interrupted time-series analyses were done to quantify the impact of pandemic response measures and their relaxation on disease rates, and autoregressive integrated moving average models were used to estimate effect sizes and forecast counterfactual trends by hemisphere. Findings: Overall, 116 841 cases were analysed: 76 481 in 2018-19, before the pandemic, and 40 360 in 2020-21, during the pandemic. During the pandemic there was a significant reduction in the risk of disease caused by S pneumoniae (risk ratio 0·47; 95% CI 0·40-0·55), H influenzae (0·51; 0·40-0·66) and N meningitidis (0·26; 0·21-0·31), while no significant changes were observed for S agalactiae (1·02; 0·75-1·40), which is not transmitted via the respiratory route. No major changes in the distribution of cases were observed when stratified by patient age or serotype or group. An estimated 36 289 (95% prediction interval 17 145-55 434) cases of invasive bacterial disease were averted during the first 2 years of the pandemic among IRIS-participating countries and territories. Interpretation: COVID-19 containment measures were associated with a sustained decrease in the incidence of invasive disease caused by S pneumoniae, H influenzae, and N meningitidis during the first 2 years of the pandemic, but cases began to increase in some countries towards the end of 2021 as pandemic restrictions were lifted. These IRIS data provide a better understanding of microbial transmission, will inform vaccine development and implementation, and can contribute to health-care service planning and provision of policies.Publication Two multi-fragment recombination events resulted in the β-lactam-resistant serotype 11A-ST6521 related to Spain9V-ST156 pneumococcal clone spreading in south-western Europe, 2008 to 2016(European Centre for Disease Prevention and Control (ECDC), 2020-04) González-Díaz, Aida; Machado, Miguel P; Càmara, Jordi; Yuste, Jose Enrique; Varon, Emmanuelle; Domenech Lucas, Mirian; Del Grosso, María; Marimón, José María; Cercenado, Emilia; Larrosa, Nieves; Quesada, María Dolores; Fontanals, Dionisia; El-Mniai, Assiya; Cubero, Meritxell; Carriço, João A; Martí, Sara; Ramirez, Mario; Ardanuy, Carmen; Centro de Investigación Biomedica en Red - CIBER; Instituto de Salud Carlos III; Unión Europea. Fondo Europeo de Desarrollo Regional (FEDER/ERDF); Government of Catalonia (España)BackgroundThe successful pneumococcal clone Spain9V-ST156 (PMEN3) is usually associated with vaccine serotypes 9V and 14.AimOur objective was to analyse the increase of a serotype 11A variant of PMEN3 as cause of invasive pneumococcal disease (IPD) in Spain and its spread in south-western Europe.MethodsWe conducted a prospective multicentre study of adult IPD in Spain (2008-16). Furthermore, a subset of 61 penicillin-resistant serotype 11A isolates from France, Italy, Portugal and Spain were subjected to whole genome sequencing (WGS) and compared with 238 genomes from the European Nucleotide Archive (ENA).ResultsAlthough the incidence of serotype 11A in IPD was stable, a clonal shift was detected from CC62 (penicillin-susceptible) to CC156 (penicillin-resistant). By WGS, three major 11A-CC156 lineages were identified, linked to ST156 (n = 5 isolates; France, Italy and Portugal), ST166 (n = 4 isolates; France and Portugal) and ST838/6521 (n = 52 isolates; France, Portugal and Spain). Acquisition of the 11A capsule allowed to escape vaccine effect. AP200 (11A-ST62) was the donor for ST156 and ST838/6521 but not for ST166. In-depth analysis of ST838/6521 lineage showed two multi-fragment recombination events including four and seven fragments from an 11A-ST62 and an NT-ST344 representative, respectively.ConclusionThe increase in penicillin-resistant serotype 11A IPD in Spain was linked to the spread of a vaccine escape PMEN3 recombinant clone. Several recombination events were observed in PMEN3 acquiring an 11A capsule. The most successful 11A-PMEN3 lineage spreading in south-western Europe appeared after two multi-fragment recombination events with representatives of two major pneumococcal clones (11A-ST62 and NT-ST344).Publication Vaccination with LytA, LytC, or Pce of Streptococcus pneumoniae Protects against Sepsis by Inducing IgGs That Activate the Complement System.(Multidisciplinary Digital Publishing Institute (MDPI), 2021-02-23) Corsini, Bruno; Aguinagalde, Leire; Ruiz, Susana; Domenech Lucas, Mirian; Yuste, Jose Enrique; Ministerio de Economía y Competitividad (España); National Council for Scientific and Technological Development (Brasil)The emergence of non-vaccine serotypes of Streptococcus pneumoniae after the use of vaccines based in capsular polysaccharides demonstrates the need of a broader protection vaccine based in protein antigens and widely conserved. In this study, we characterized three important virulence factors of S. pneumoniae namely LytA, LytC, and Pce as vaccine candidates. These proteins are choline-binding proteins that belong to the cell wall hydrolases' family. Immunization of mice with LytA, LytC, or Pce induced high titers of immunoglobulins G (IgGs) of different subclasses, with IgG1, IgG2a, and IgG2b as the predominant immunoglobulins raised. These antibodies activated the classical pathway of the complement system by increasing the recognition of C1q on the surface of pneumococcal strains of different serotypes. Consequently, the key complement component C3 recognized more efficiently these strains in the presence of specific antibodies elicited by these proteins, activating, therefore, the phagocytosis. Finally, a mouse sepsis model of infection was established, confirming that vaccination with these proteins controlled bacterial replication in the bloodstream, increasing the survival rate. Overall, these results demonstrate that LytA, LytC, and Pce can be protein antigens to be contained in a future universal vaccine against S. pneumoniae.