Desarrollo de vacunas contra infecciones nosocomiales

El descubrimiento de los antibióticos y el desarrollo para su uso clínico puede ser considerado uno de los logros biomédicos más importantes del siglo pasado. Desafortunadamente, muchas especies de bacterias patógenas han desarrollado y/o adquirido múltiples tipos de mecanismos de resistencia que pueden limitar severamente la eficacia terapéutica de la terapia antimicrobiana. La vacunación es un método eficaz para prevenir infecciones bacterianas, sin embargo, no se ha aplicado con éxito en las infecciones causadas por algunos de los más problemáticos patógenos multirresistentes. Nuestra investigación desarrollada en el Centro Nacional de Microbiología, Madrid, España, se centra en estudiar el potencial de las vacunas para contribuir a reducir la carga de enfermedades infecciosas causadas por bacterias Gram-negativas multirresistentes. Los obstáculos técnicos, logísticos y sociales que han limitado el desarrollo exitoso de vacunas para estas infecciones en el pasado serán presentados, así como los avances recientes en el desarrollo de vacunas que pueden contribuir a superar estos desafíos. Así mismo, se presentará una síntesis de las tecnologías de vacunas que se han empleado en el desarrollo de vacunas para bacterias Gram-negativas multirresistentes clave, y tecnologías emergentes que pueden contribuir a futuros éxitos. Finalmente, se presentará el desarrollo de una vacuna en contra de los tres patógenos Gram-negativos resistentes a múltiples fármacos más preocupantes, Acinetobacter baumannii, Klebsiella pneumoniae y Pseudomonas aeruginosa, centrándose en estudios recientes realizados en nuestro laboratorio, así como una revisión exhaustiva de los esfuerzos de desarrollo de vacunas durante los últimos 40 años

Effect of Subinhibitory Concentrations of Antibiotics and Disinfectants on ISAba-Mediated Inactivation of Lipooligosaccharide Biosynthesis Genes in Acinetobacter baumannii

Inactivation of the lipooligosaccharide (LOS) biosynthesis genes lpxA, lpxC and lpxD by ISAba insertion elements results in high-level resistance to colistin in A. baumannii. In the present study, we quantify the rate of spontaneous insertional inactivation of LOS biosynthesis genes by ISAba elements in the ATCC 19606-type strain and two multidrug clinical isolates. Using insertional inactivation of lpxC by ISAba11 in the ATCC 19606 strain as a model, we determine the effect of several subinhibitory concentrations of the antibiotics, namely tetracycline, ciprofloxacin, meropenem, kanamycin and rifampicin, as well as the disinfectants ethanol and chlorhexidine on ISAba11 insertion frequencies. Notably, subinhibitory concentrations of tetracycline significantly increased ISAba11 insertion, and rifampicin completely inhibited the emergence of colistin resistance due to ISAba11 inactivation of lpxC. Sequencing of ISAba11 insertion sites within the lpxC gene demonstrated that insertions clustered between nucleotides 382 and 618 (58.3% of unique insertions detected), indicating that this may be a hotspot for ISAba11 insertion. The alignment of insertion sites revealed a semi-conserved AT-rich consensus sequence upstream of the ISAba11 insertion site, suggesting that ISAba11 insertion sites may be sequence-dependent. This study explores previously uncharacterized aspects regarding the acquisition of colistin resistance through insertional activation in LOS biosynthesis genes in A. baumannii.This research was supported by grants MPY 380/18 from the Instituto de Salud Carlos III (ISCIII) awarded to M.J.M. A.C.L. is supported by the Atracción de Talento Program of the Community of Madrid.S

High-affinity chemotaxis to histamine of the human pathogen Pseudomonas aeruginosa

FEMS 2019: 8th congress of European Microbiologists, in Glasgow, Scotland, 7-11 July 201

Towards Control and Oversight of SARS-CoV-2 Diagnosis and Monitoring through Multiplexed Quantitative Electroanalytical Immune Response Biosensors

The development of versatile and sensitive biotools to quantify specific SARS-CoV-2 immunoglobulins in SARS-CoV-2 infected and non-infected individuals, built-on the surface of magnetic microbeads functionalized with nucleocapsid (N) and inhouse expressed recombinant spike (S) proteins is reported. Amperometric interrogation of captured N- and S-specific circulating total or individual immunoglobulin (Ig) isotypes (IgG, IgM, and IgA), subsequently labelled with HRP-conjugated secondary antibodies, was performed at disposable single or multiplexed (8) screen-printed electrodes using the HQ/HRP/H2O2 system. The obtained results using N and in-house expressed S ectodomains of five SARS-CoV-2 variants of concern (including the latest Delta and Omicron) allow identification of vulnerable populations from those with natural or acquired immunity, monitoring of infection, evaluation of vaccine efficiency and even identification of the variant responsible for the infection

Familial hypercholesterolaemia in children and adolescents from 48 countries: a cross-sectional study

Background: Approximately 450 000 children are born with familial hypercholesterolaemia worldwide every year, yet only 2·1% of adults with familial hypercholesterolaemia were diagnosed before age 18 years via current diagnostic approaches, which are derived from observations in adults. We aimed to characterise children and adolescents with heterozygous familial hypercholesterolaemia (HeFH) and understand current approaches to the identification and management of familial hypercholesterolaemia to inform future public health strategies. Methods: For this cross-sectional study, we assessed children and adolescents younger than 18 years with a clinical or genetic diagnosis of HeFH at the time of entry into the Familial Hypercholesterolaemia Studies Collaboration (FHSC) registry between Oct 1, 2015, and Jan 31, 2021. Data in the registry were collected from 55 regional or national registries in 48 countries. Diagnoses relying on self-reported history of familial hypercholesterolaemia and suspected secondary hypercholesterolaemia were excluded from the registry; people with untreated LDL cholesterol (LDL-C) of at least 13·0 mmol/L were excluded from this study. Data were assessed overall and by WHO region, World Bank country income status, age, diagnostic criteria, and index-case status. The main outcome of this study was to assess current identification and management of children and adolescents with familial hypercholesterolaemia. Findings: Of 63 093 individuals in the FHSC registry, 11 848 (18·8%) were children or adolescents younger than 18 years with HeFH and were included in this study; 5756 (50·2%) of 11 476 included individuals were female and 5720 (49·8%) were male. Sex data were missing for 372 (3·1%) of 11 848 individuals. Median age at registry entry was 9·6 years (IQR 5·8-13·2). 10 099 (89·9%) of 11 235 included individuals had a final genetically confirmed diagnosis of familial hypercholesterolaemia and 1136 (10·1%) had a clinical diagnosis. Genetically confirmed diagnosis data or clinical diagnosis data were missing for 613 (5·2%) of 11 848 individuals. Genetic diagnosis was more common in children and adolescents from high-income countries (9427 [92·4%] of 10 202) than in children and adolescents from non-high-income countries (199 [48·0%] of 415). 3414 (31·6%) of 10 804 children or adolescents were index cases. Familial-hypercholesterolaemia-related physical signs, cardiovascular risk factors, and cardiovascular disease were uncommon, but were more common in non-high-income countries. 7557 (72·4%) of 10 428 included children or adolescents were not taking lipid-lowering medication (LLM) and had a median LDL-C of 5·00 mmol/L (IQR 4·05-6·08). Compared with genetic diagnosis, the use of unadapted clinical criteria intended for use in adults and reliant on more extreme phenotypes could result in 50-75% of children and adolescents with familial hypercholesterolaemia not being identified. Interpretation: Clinical characteristics observed in adults with familial hypercholesterolaemia are uncommon in children and adolescents with familial hypercholesterolaemia, hence detection in this age group relies on measurement of LDL-C and genetic confirmation. Where genetic testing is unavailable, increased availability and use of LDL-C measurements in the first few years of life could help reduce the current gap between prevalence and detection, enabling increased use of combination LLM to reach recommended LDL-C targets early in life

Molecular basic of chemosensory: Biofilm and cell-to-cell signaling in different species of pseudomonas

La investigación descrita en esta tesis se centra en tres aspectos importantes de la señalización en Pseudomonas, como son: Las vías quimiosensoras involucradas en quimiotaxis o en funciones celulares alternativas, la formación de biopelículas y la comunicación bacteriana a través de sistemas de quorum sensing. Las bacterias utilizan diversas moléculas de pequeño peso molecular para monitorear su entorno, así como su estado intracelular. Esta capacidad, a su vez permite a las bacterias adaptarse rápidamente a los cambios. Aunque está bien establecido que los sistemas de regulación basados en uno y dos componentes participan en la regulación de la formación de biopelículas, también existe evidencia que sugiere que las vías quimiosensoras están involucradas en dicha regulación. Sin embargo, existe poca información sobre qué quimiorreceptores de la vía quimiosensora y cuales señales modulan este proceso. Por lo tanto, uno de los objetivos principales de esta tesis era investigar el papel que tienen los quimiorreceptores en la formación de biopelículas.Research described in this dissertation focuses on three important aspects of Pseudomonas signaling: Chemosensory pathways involved in chemotaxis or with alternative cellular functions (ACF), biofilm formation and bacterial communication via quorum sensing (QS).Tesis Univ. Granada. Programa Oficial de Doctorado en: Biología Fundamental y de Sistema

Molecular basic of chemosensory: Biofilm and cell-to-cell signaling in different species of pseudomonas

La investigación descrita en esta tesis se centra en tres aspectos importantes de la señalización en Pseudomonas, como son: Las vías quimiosensoras involucradas en quimiotaxis o en funciones celulares alternativas, la formación de biopelículas y la comunicación bacteriana a través de sistemas de quorum sensing. Las bacterias utilizan diversas moléculas de pequeño peso molecular para monitorear su entorno, así como su estado intracelular. Esta capacidad, a su vez permite a las bacterias adaptarse rápidamente a los cambios. Aunque está bien establecido que los sistemas de regulación basados en uno y dos componentes participan en la regulación de la formación de biopelículas, también existe evidencia que sugiere que las vías quimiosensoras están involucradas en dicha regulación. Sin embargo, existe poca información sobre qué quimiorreceptores de la vía quimiosensora y cuales señales modulan este proceso. Por lo tanto, uno de los objetivos principales de esta tesis era investigar el papel que tienen los quimiorreceptores en la formación de biopelículas.Research described in this dissertation focuses on three important aspects of Pseudomonas signaling: Chemosensory pathways involved in chemotaxis or with alternative cellular functions (ACF), biofilm formation and bacterial communication via quorum sensing (QS).Tesis Univ. Granada. Programa Oficial de Doctorado en: Biología Fundamental y de Sistema

The plant compound rosmarinic acid induces a broad quorum sensing response in Pseudomonas aeruginosa PAO1

The interference of plant compounds with bacterial quorum sensing (QS) is a major mechanism through which plants and bacteria communicate. However, little is known about the modes of action and effects on signal integrity during this type of communication. We have recently shown that the plant compound rosmarinic acid (RA) specifically binds to the Pseudomonas aeruginosa RhlR QS receptor. To determine the effect of RA on expression patterns, we carried out global RNA-seq analysis. The results show that RA induces the expression of 128 genes, amongst which many virulence factor genes. RA triggers a broad QS response because 88% of the induced genes are known to be controlled by QS, and because RA stimulated genes were found to be involved in all four QS signalling systems within P. aeruginosa. This finding was confirmed through the analysis of transcriptional fusions transferred to wt and a rhlI/lasI double mutant. RA did not induce gene expression in the rhlI/lasI/rhlR triple mutant indicating that the effects observed are due to the RA-RhlR interaction. Furthermore, RA induced seven sRNAs that were all encoded in regions close to QS and/or RA induced genes. This work significantly enhances our understanding of plant bacteria interaction.We would like to thank Dr. M. Cámara (Nottingham University) and Dr. Junichi Kato (Hiroshima University) for their openness and generosity in providing fellow scientists with strains and plasmids. We thank Pedro Carmona and Jordi Martorell from the GENYO Research Center (Granada,Spain) for conducting RNA-seq experiments, data analysis and their help in writing this manuscript. This work was supported by FEDER funds and Fondo Social Europeo through grants to T. Krell from the Junta de Andalucía (Grant No. CVI-7335) and the Spanish Ministry for Economy and Competitiveness (Grant Nos. BIO2013-42297 and BIO2016-76779-P).Peer Reviewe