Biofilmes de "Streptococcus pneumoniae": genética, composición y terapia

Se ha estimado que los biofilmes están implicados en más de la mitad de las infecciones bacterianas humanas y, hasta en un 80%, en las infecciones crónicas. Además, las infecciones causadas por bacterias patógenas asociadas a la formación de biofilmes se muestran refractarias tanto a los agentes antimicrobianos como a los mecanismos de defensa del sistema inmunitario del hospedador. Todo ello nos ha llevado a plantear la necesidad de desarrollar nuevas vías de investigación en bacterias de importancia clínica relevante como es el caso de S. pneumoniae. Hasta el momento, el modo de vida sésil de S. pneumoniae apenas ha sido investigado y, sin embargo, la otitis media, y, posiblemente, la neumonía y la meningitis son enfermedades neumocócicas asociadas a la formación de biofilmes. Teniendo en cuenta lo expuesto anteriormente, los objetivos experimentales que nos planteamos investigar en la presente Tesis son los siguientes: 1. Búsqueda de nuevos genes implicados en la formación de biofilm. 2. Determinar el papel del polisacárido capsular en la formación de biofilm de S. pneumoniae. 3. Investigar la composición de la matriz extracelular de los biofilmes neumocócicos. 4. Estudiar los mecanismos de evasión del sistema inmune por parte de un biofilm. 5. Desarrollar y evaluar posibles medidas terapeúticas contra los biofilmes

Combination of Cefditoren and N-acetyl-l-Cysteine Shows a Synergistic Effect against Multidrug-Resistant Streptococcus pneumoniae Biofilms

Biofilm 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.This work was supported by Ministerio de Ciencia e Innovación (MICINN) (grant PID2020-119298RB-I00) and by Meiji Pharma Spain (grant MVP 119/20). J.Y. has received grants from MSD-USA (MISP Call) and Pfizer that are not related to this work. J.Y. has participated in advisory boards organized by MSD and Pfizer. M.G. and P.C. are members of the Scientific Department of Meiji Pharma Spain. The other authors declare no competing interests.S

Clearance of mixed bioflms of Streptococcus pneumoniae and methicillin‑susceptible/resistant Staphylococcus aureus by antioxidants N‑acetyl‑l‑cysteine and cysteamine.

Bioflm-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 bioflms and tested new antibioflm therapies with antioxidants N-acetyl-l-cysteine (NAC) and cysteamine (Cys). We developed two in vitro S. aureus–S. pneumoniae mixed bioflms in 96-well polystyrene microtiter plates and we treated in vitro bioflms with Cys and NAC analyzing their efect 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 bioflm. We demonstrated the efect of Cys in preventing S. aureus bioflms and S. aureus–S. pneumoniae mixed bioflms. 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 bioflms. The methicillin resistance background did not change the antioxidants efect in S. aureus. These results identify NAC and Cys as promising repurposed drug candidates for the prevention and treatment of mixed bioflms by S. pneumoniae and S. aureus

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

Background: 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.This work was supported by the Spanish Ministry of Science and Innovation (grant PID2020–119298RB-I00), Meiji Pharma Spain (grant MVP 119/20), and internal funding from Instituto de Salud Carlos III.S

Impact of Pneumococcal Vaccination in the Nasopharyngeal Carriage of Streptococcus pneumoniae in Healthy Children of the Murcia Region in Spain.

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.This work was supported by Ministerio de Economía, Industria y Competitividad (MINECO)[grant SAF2017-83388].S

Micromundo@ucm: research and awareness for the stealth pandemics of antibiotic resistance

Actualmente se estima que las resistencias antibióticas se cobran 1.270.000 vidas anualmente a nivel global. Es necesario contribuir desde multiples ángulos a preservar la efectividad de os antibióticos y descubrir nuevas etsrategias terapéuticas. MicroMundo es un proyecto de Aprendizaje-Servicio y Ciencia Ciudadana que pretende crear cultura científica y concienciación en cuestiones de Salud Global en los jóvenes. Se pretende que sean los más jóvenes, los estudiantes de ESO y Bachillerato, los responsables de la transmisión de ese conocimiento a la comunidad. Pero un segundo objetivo, no menos importante, es el de generar y potenciar vocaciones STEM e interés por el I+D en Biomedicina. Para conseguir estos objetivos, diversos equipos de estudiantes universitarios imparten y coordinan el proyecto en colegios e institutos de su comunidad, coordinados por sus tutores (profesores e investigadores del área de Microbiología de las Facultades de Farmacia, Medicina, Veterinaria y Biología, cubriendo los tres vértices del triángulo One Health: salud humana, animal y medioambiental). En la UCM, durante el curso 2021-22, treinta y dos equipos de han trabajado en unos treinta colegios e institutos, implicando a unos 600 estudiantes preuniversitarios en el trabajo experimental del proyecto.It is currently estimated that antibiotic resistance annually claims 1,270,000 lives globally. It is necessary to contribute from multiple angles to preserve the effectiveness of antibiotics and discover new therapeutic strategies. MicroMundo is a Service-Learning and Citizen Science project that aims to create a scientific culture and awareness of Global Health issues among young people. It is intended that the youngest, Secondary and Baccalaureate students, be responsible for the transmission of this knowledge to the community. But a second objective, no less important, is to generate and promote STEM vocations and interest in R&D in Biomedicine. To achieve these objectives, various teams of university students teach and coordinate the project in schools and institutes in their community, coordinated by their tutors (professors and researchers from the Microbiology area of ​​the Faculties of Pharmacy, Medicine, Veterinary Medicine and Biology, covering the three vertices of the One Health triangle: human, animal and environmental health). At the UCM, during the 2021-22 academic year, thirty-two teams have worked in thirty-two schools, involving some 600 pre-university students in the experimental work of the project.Depto. de Genética, Fisiología y MicrobiologíaDepto. de Microbiología y ParasitologíaDepto. de Sanidad AnimalSección Dptal. de Nutrición y Ciencia de los Alimentos (Veterinaria)Fac. de Ciencias BiológicasFac. de FarmaciaFac. de VeterinariaTRUEunpu

Subcutaneous anti-COVID-19 hyperimmune immunoglobulin for prevention of disease in asymptomatic individuals with SARS-CoV-2 infection: a double-blind, placebo-controlled, randomised clinical trialResearch in context

Summary: Background: Anti-COVID-19 hyperimmune immunoglobulin (hIG) can provide standardized and controlled antibody content. Data from controlled clinical trials using hIG for the prevention or treatment of COVID-19 outpatients have not been reported. We assessed the safety and efficacy of subcutaneous anti-COVID-19 hyperimmune immunoglobulin 20% (C19-IG20%) compared to placebo in preventing development of symptomatic COVID-19 in asymptomatic individuals with SARS-CoV-2 infection. Methods: We did a multicentre, randomized, double-blind, placebo-controlled trial, in asymptomatic unvaccinated adults (≥18 years of age) with confirmed SARS-CoV-2 infection within 5 days between April 28 and December 27, 2021. Participants were randomly assigned (1:1:1) to receive a blinded subcutaneous infusion of 10 mL with 1 g or 2 g of C19-IG20%, or an equivalent volume of saline as placebo. The primary endpoint was the proportion of participants who remained asymptomatic through day 14 after infusion. Secondary endpoints included the proportion of individuals who required oxygen supplementation, any medically attended visit, hospitalisation, or ICU, and viral load reduction and viral clearance in nasopharyngeal swabs. Safety was assessed as the proportion of patients with adverse events. The trial was terminated early due to a lack of potential benefit in the target population in a planned interim analysis conducted in December 2021. ClinicalTrials.gov registry: NCT04847141. Findings: 461 individuals (mean age 39.6 years [SD 12.8]) were randomized and received the intervention within a mean of 3.1 (SD 1.27) days from a positive SARS-CoV-2 test. In the prespecified modified intention-to-treat analysis that included only participants who received a subcutaneous infusion, the primary outcome occurred in 59.9% (91/152) of participants receiving 1 g C19-IG20%, 64.7% (99/153) receiving 2 g, and 63.5% (99/156) receiving placebo (difference in proportions 1 g C19-IG20% vs. placebo, −3.6%; 95% CI -14.6% to 7.3%, p = 0.53; 2 g C19-IG20% vs placebo, 1.1%; −9.6% to 11.9%, p = 0.85). None of the secondary clinical efficacy endpoints or virological endpoints were significantly different between study groups. Adverse event rate was similar between groups, and no severe or life-threatening adverse events related to investigational product infusion were reported. Interpretation: Our findings suggested that administration of subcutaneous human hyperimmune immunoglobulin C19-IG20% to asymptomatic individuals with SARS-CoV-2 infection was safe but did not prevent development of symptomatic COVID-19. Funding: Grifols