Centro Nacional de Microbiología (CNM)

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https://hdl.handle.net/20.500.12105/19609

El Centro Nacional de Microbiología (CNM) es uno de los centros pertenecientes al Instituto de Salud Carlos III que proporciona apoyo científico-técnico a la Administración General del Estado, a las Comunidades Autónomas y al Sistema Nacional de Salud (SNS), tal y como se recoge en La Ley General de Sanidad (Ley 14/1986, de 25 de abril) y el Estatuto del Instituto de Salud Carlos III (RD 375/2001, de 6 abril y su posterior reforma, RD 1672/2009, de 6 de noviembre). La función específica del CNM es el control de las enfermedades infecciosas para lo que ofrece servicios de diagnóstico y referencia, manteniendo además programas de investigación, tanto básica como orientada, relacionados con la prevención, el diagnóstico y el tratamiento de estas enfermedades. Dispone de un sistema de gestión de calidad conforme a la norma UNE-EN ISO 9001, certificado por AENOR, para la recepción de muestras biológicas, así como varias técnicas y servicios acreditados por ENAC según la norma UNE-EN ISO 15189.

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Detection of caprine paratuberculosis (Johne's disease) in pre- and post-vaccinated herds: morphological diagnosis, lesion grading, and bacterial identification.
(Frontiers Media, 2024-07-31) Stefanova, Elena Plamenova; Sierra, Eva; Fernández, Antonio; Quesada-Canales, Oscar; Paz-Sánchez, Yania; Colom-Rivero, Ana; Espinosa de Los Monteros, Antonio; Herráez, Pedro; Domínguez, Lucas; Bezos, Javier; Pérez-Sancho, Marta; Risalde, María A; Andrada, Marisa; Moreno-Iruela, Inmaculada; Fundación CajaCanarias; Fundación La Caixa; Gobierno de Canarias (España); Ministerio de Economía (España); Unión Europea. Fondo Europeo de Desarrollo Regional (FEDER/ERDF); Ministerio de Universidades (España); Agencia Canaria de Investigación, Innovación y Sociedad de la Información
Samples from the mesenteric lymph nodes (MS LNs) and ileocecal valves (ICV) of 105 goats, comprising 61 non-vaccinated and 44 vaccinated against subspecies (MAP), were collected at slaughter from a farm with a confirmed history of paratuberculosis (PTB). These goats had subclinical infections. PTB-compatible lesions in the MS LNs, ICV lamina propria (LP), and Peyer's patches (PPs) were graded separately. Furthermore, the load of acid-fast bacilli was quantified using Ziehl-Neelsen staining (ZN), MAP antigens by immunohistochemistry (IHC), and MAP DNA by PCR targeting the IS900 sequence. Gross PTB-compatible lesions were found in 39% of the goats, with 31.72% vaccinated (V) and 68.29% non-vaccinated (nV). Histopathological lesions induced MAP were observed in 58% of the animals, with 36.07% vaccinated and 63.93% non-vaccinated. The inclusion of histopathology as a diagnostic tool led to a 28% increase in diagnosed cases in MS LNs and 86.05% in ICV. Grade IV granulomas with central mineralization and necrosis were the most common lesions in MS LNs. In the ICV, mild granulomatous enteritis with multifocal foci of epithelioid macrophages was predominant, occurring more frequently in the PPs than in the LP. Furthermore, statistical differences in the presence of histopathological lesions between vaccinated and non-vaccinated goats were noted in MS LNs, ICV LPs, and ICV PPs. Non-vaccinated animals showed higher positivity rates in ZN, IHC, and PCR tests, underscoring the benefits of anti-MAP vaccination in reducing PTB lesions and bacterial load in target organs. Our findings emphasize the necessity of integrating gross and histopathological assessments with various laboratory techniques for accurate morphological and etiological diagnosis of PTB in both vaccinated and non-vaccinated goats with subclinical disease. However, further studies are required to refine sampling protocols for subclinical PTB in goats to enhance the consistency of diagnostic tools.
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 & Dohme
Objectives: 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.
Conservation of HLA Spike Protein Epitopes Supports T Cell Cross-Protection in SARS-CoV-2 Vaccinated Individuals against the Potentially Zoonotic Coronavirus Khosta-2
(Multidisciplinary Digital Publishing Institute (MDPI), 2024-05-31) Martin-Galiano, Antonio Javier; Lopez, Daniel; Ministerio de Ciencia e Innovación (España)
Heterologous vaccines, which induce immunity against several related pathogens, can be a very useful and rapid way to deal with new pandemics. In this study, the potential impact of licensed COVID-19 vaccines on cytotoxic and helper cell immune responses against Khosta-2, a novel sarbecovirus that productively infects human cells, was analyzed for the 567 and 41 most common HLA class I and II alleles, respectively. Computational predictions indicated that most of these 608 alleles, covering more than 90% of the human population, contain sufficient fully conserved T-cell epitopes between the Khosta-2 and SARS-CoV-2 spike-in proteins. Ninety percent of these fully conserved peptides for class I and 93% for class II HLA molecules were verified as epitopes recognized by CD8+ or CD4+ T lymphocytes, respectively. These results show a very high correlation between bioinformatic prediction and experimental assays, which strongly validates this study. This immunoinformatics analysis allowed a broader assessment of the alleles that recognize these peptides, a global approach at the population level that is not possible with experimental assays. In summary, these findings suggest that both cytotoxic and helper cell immune protection elicited by currently licensed COVID-19 vaccines should be effective against Khosta-2 virus infection. Finally, by being rapidly adaptable to future coronavirus pandemics, this study has potential public health implications.
Mycobacterium tuberculosis complex in domestic goats in Southern Spain
(Elsevier, 2024-06) Jiménez-Martín, Débora; García-Bocanegra, Ignacio; Risalde, María A; Napp, Sebastián; Dominguez-Rodriguez, Mercedes; Romero, Beatriz; Moreno-Iruela, Inmaculada; Martínez, Remigio; Cano-Terriza, David; Ministerio de Agricultura, Pesca y Alimentación (España); Ministerio de Economía y Competitividad (España); University of Córdoba (España); Ministerio de Ciencia e Innovación (España); Unión Europea. Fondo Europeo de Desarrollo Regional (FEDER/ERDF); Ministerio de Universidades (España); Regional Government of Andalusia (España); Centro de Investigación Biomédica en Red - CIBERINFEC (Enfermedades Infecciosas)
Tuberculosis (TB) is a zoonotic infectious disease caused by bacteria belonging to the Mycobacterium tuberculosis complex (MTC), which can affect a wide variety of domestic and wild animal species. Although the role of goats as a reservoir of MTC bacteria has been evidenced, information about the circulation of MTC strains in this species is still very scarce. The aim of the present study was to determine the seroprevalence, spatial distribution, risk factors and MTC spoligotypes circulating in goats from Andalusia (Southern Spain), the Spanish region with the largest goat census and a hotspot area of TB in both cattle and wild ungulates. A total of 2155 serum samples from 80 goat flocks were analyzed by an in-house ELISA using the P22 protein complex as a coating antigen. Antibodies against MTC were detected in 473 goats (21.9%, 95% CI: 20.2-23.7) and the true seroprevalence was 22.3% (95% CI: 20.6-24.1). Seropositivity was found in 72 (90.0%) of the 80 flocks analyzed. The generalized estimating equation model showed that the management system (higher seroprevalence on intensive and semi-intensive farms), and the presence of hospital pens inside the regular stables, were risk factors potentially associated with MTC exposure in goats in Southern Spain. The spatial analysis identified a significant spatial cluster (p < 0.001) in Eastern Andalusia. A total of 16 different MTC spoligotypes, including five of M. caprae and eleven of M. bovis, were identified in goats between 2015 and 2022 in the study area, with SB0157 as the most frequently isolated. The results obtained indicate widespread and non-homogeneous spatial distribution of MTC in goat herds from Southern Spain. The high individual and herd-level seroprevalence values found suggest that goats could play a significant role in the maintenance and transmission of MTC in the study area. Our results highlight the importance of implementing control measures in this species.
Microscopy detection and molecular characterisation of Giardia duodenalis infection in outpatients seeking medical care in Egypt
(Frontiers Media, 2024-04) Elmahallawy, Ehab Kotb; Gareh, Ahmed; Ghallab, Marwa M I; Köster, Pamela Carolina; Dashti, Alejandro; Aboelsoued, Dina; Toaleb, Nagwa Ibrahim; Alzaylaee, Hind; Gonzálvez, Moisés; Saleh, Amira A; Alhegaili, Alaa S; Eldehn, Ahmed Fathy; Hernández-Castro, Carolina; Bailo, Begoña; Gonzalez-Barrio, David; Carmena, David; Instituto de Salud Carlos III; Ministerio de Economía y Competitividad (España); Ministry of Higher Education (Egipto); University of Murcia (España); Ministerio de Universidades (España); Unión Europea. Comisión Europea. NextGenerationEU; Ministerio de Ciencia, Innovación y Universidades (España); Fundación Carolina; University of Antioquia (Colombia)
Introduction: Giardiosis remains one of the most prevalent enteric parasitic infections globally. Earlier molecular-based studies conducted in Egypt have primarily focused on paediatric clinical populations and most were based on single genotyping markers. As a result, there is limited information on the frequency and genetic diversity of G. duodenalis infections in individuals of all age groups. Methods: Individual stool samples (n = 460) from outpatients seeking medical care were collected during January-December 2021 in Kafr El-Sheikh governorate, northern Egypt. Initial screening for the presence of G. duodenalis was conducted by coprological examination. Microscopy-positive samples were further confirmed by real-time PCR. A multilocus sequence typing approach targeted amplification of the glutamate dehydrogenase (gdh), beta-giardin (bg), and triose phosphate isomerase (tpi) genes was used for genotyping purposes. A standardised epidemiological questionnaire was used to gather basic sociodemographic and clinical features of the recruited patients. Results: Giardia duodenalis cysts were observed in 5.4% (25/460, 95% CI: 3.6-7.9) of the stool samples examined by conventional microscopy. The infection was more frequent in children under the age of 10 years and in individuals presenting with diarrhoea but without reaching statistical significance. Stool samples collected during the winter period were more likely to harbour G. duodenalis. All 25 microscopy-positive samples were confirmed by real-time PCR, but genotyping data was only available for 56.0% (14/25) of the isolates. Sequence analyses revealed the presence of assemblages A (78.6%, 11/14) and B (21.4%, 3/14). All assemblage A isolates were identified as sub-assemblage AII, whereas the three assemblage B sequences belonged to the sub-assemblage BIII. Patients with giardiosis presenting with diarrhoea were more frequently infected by the assemblage A of the parasite. Conclusion: This is one of the largest epidemiological studies evaluating G. duodenalis infection in individuals of all age groups in Egypt. Our molecular data suggest that G. duodenalis infections in the surveyed population are primarily of anthropic origin. However, because assemblages A and B are zoonotic, some of the infections identified can have an animal origin. Additional investigations targeting animal (domestic and free-living) and environmental (water) samples are warranted to better understand the epidemiology of giardiosis in Egypt.
Distribution of Aspergillus species and prevalence of azole resistance in clinical and environmental samples from a Spanish hospital during a three-year study period
(Wiley, 2024-04) Lucio, Jose; Alcazar-Fuoli, Laura; Gil, Horacio; Cano-Pascual, Samuel; Hernandez-Egido, Sara; Cuetara, Maria Soledad; Mellado, Emilia; Instituto de Salud Carlos III; Centro de Investigación Biomédica en Red - CIBERINFEC (Enfermedades Infecciosas)
Background: Surveillance studies are crucial for updating trends in Aspergillus species and antifungal susceptibility information. Objectives: Determine the Aspergillus species distribution and azole resistance prevalence during this 3-year prospective surveillance study in a Spanish hospital. Materials and methods: Three hundred thirty-five Aspergillus spp. clinical and environmental isolates were collected during a 3-year study. All isolates were screened for azole resistance using an agar-based screening method and resistance was confirmed by EUCAST antifungal susceptibility testing. The azole resistance mechanism was confirmed by sequencing the cyp51A gene and its promoter. All Aspergillus fumigatus strains were genotyped using TRESPERG analysis. Results: Aspergillus fumigatus was the predominant species recovered with a total of 174 strains (51.94%). The rest of Aspergillus spp. were less frequent: Aspergillus niger (14.93%), Aspergillus terreus (9.55%), Aspergillus flavus (8.36%), Aspergillus nidulans (5.37%) and Aspergillus lentulus (3.28%), among other Aspergillus species (6.57%). TRESPERG analysis showed 99 different genotypes, with 72.73% of the strains being represented as a single genotype. Some genotypes were common among clinical and environmental A. fumigatus azole-susceptible strains, even when isolated months apart. We describe the occurrence of two azole-resistant A. fumigatus strains, one clinical and another environmental, that were genotypically different and did not share genotypes with any of the azole-susceptible strains. Conclusions: Aspergillus fumigatus strains showed a very diverse population although several genotypes were shared among clinical and environmental strains. The isolation of azole-resistant strains from both settings suggest that an efficient analysis of clinical and environmental sources must be done to detect azole resistance in A. fumigatus.
Crimean-Congo haemorrhagic fever virus in questing non-Hyalomma spp. ticks in Northwest Spain, 2021
(Wiley, 2024-08) Cuadrado-Matías, Raúl; Moraga-Fernández, Alberto; Peralbo-Moreno, Alfonso; Negredo, Anabel; Sánchez-Seco, María Paz; Ruiz-Fons, Francisco; Ministerio de Ciencia e Innovación (España); Agencia Estatal de Investigación (España); Unión Europea. Fondo Europeo de Desarrollo Regional (FEDER/ERDF); Regional Government of Castile-La Mancha (España); Unión Europea. Fondo Social Europeo (ESF/FSE); University of Castilla-La Mancha (España)
Crimean-Congo haemorrhagic fever (CCHF) unexpectedly emerged in humans in Northwest Spain in 2021, and two additional cases were reported in the region in 2022. The 2021 case was associated with a tick bite on the outskirts of the city where the patient lived. PCR analysis of 95 questing ticks collected in the outskirts of that city in 2021, none of the genus Hyalomma, revealed a prevalence of confirmed CCHF virus (CCHFV) infection of 10.5%. Our results in this emerging scenario suggest the need to consider that CCHFV may be effectively spreading to Northwest Spain and to urgently understand any possible role of non-Hyalomma spp. ticks in the eco-epidemiological dynamics of CCHFV.
Could SARS-CoV-1 Vaccines in the Pipeline Have Contributed to Fighting the COVID-19 Pandemic? Lessons for the Next Coronavirus Plague
(Multidisciplinary Digital Publishing Institute (MDPI), 2024-01) Lopez, Daniel; García-Peydró, Marina; Ministerio de Ciencia e Innovación (España)
SARS-CoV-2 caused the devastating COVID-19 pandemic, which, to date, has resulted in more than 800 million confirmed cases and 7 million deaths worldwide. The rapid development and distribution (at least in high-income countries) of various vaccines prevented these overwhelming numbers of infections and deaths from being much higher. But would it have been possible to develop a prophylaxis against this pandemic more quickly? Since SARS-CoV-2 belongs to the subgenus sarbecovirus, with its highly homologous SARS-CoV-1, we propose here that while SARS-CoV-2-specific vaccines are being developed, phase II clinical trials of specific SARS-CoV-1 vaccines, which have been in the pipeline since the early 20th century, could have been conducted to test a highly probable cross-protection between SARS-CoV-1 and SARS-CoV-2.
Antimicrobial-resistant Neisseria gonorrhoeae in Europe in 2020 compared with in 2013 and 2018: a retrospective genomic surveillance study
(Elsevier, 2024-05) Golparian, Daniel; Cole, Michelle J; Sánchez-Busó, Leonor; Day, Michaela; Jacobsson, Susanne; Uthayakumaran, Thinushaa; Abad, Raquel; Bercot, Beatrice; Caugant, Dominique A; Heuer, Dagmar; Jansen, Klaus; Pleininger, Sonja; Stefanelli, Paola; Aanensen, David M; Bluemel, Benjamin; Unemo, Magnus; Euro-GASP study group; Diaz Franco, Asuncion; Vazquez-Moreno, Julio Alberto; Unión Europea. European Centre for Disease Prevention and Control (ECDC); Örebro University Hospital (Suecia); Örebro County Council Research (Suecia); Instituto de Salud Carlos III; Generalitat Valenciana (España); Ministerio de Ciencia e Innovación (España)
Background: Regular quality-assured whole-genome sequencing linked to antimicrobial resistance (AMR) and patient metadata is imperative to elucidate the shifting gonorrhoea epidemiology, both nationally and internationally. We aimed to examine the gonococcal population in the European Economic Area (EEA) in 2020, elucidate emerging and disappearing gonococcal lineages associated with AMR and patient metadata, compare with 2013 and 2018 whole-genome sequencing data, and explain changes in gonococcal AMR and gonorrhoea epidemiology. Methods: In this retrospective genomic surveillance study, we analysed consecutive gonococcal isolates that were collected in EEA countries through the European Gonococcal Antimicrobial Surveillance Programme (Euro-GASP) in 2020, and made comparisons with Euro-GASP data from 2013 and 2018. All isolates had linked AMR data (based on minimum inhibitory concentration determination) and patient metadata. We performed whole-genome sequencing and molecular typing and AMR determinants were derived from quality-checked whole-genome sequencing data. Links between genomic lineages, AMR, and patient metadata were examined. Findings: 1932 gonococcal isolates collected in 2020 in 21 EEA countries were included. The majority (81·2%, 147 of 181 isolates) of azithromycin resistance (present in 9·4%, 181 of 1932) was explained by the continued expansion of the Neisseria gonorrhoeae sequence typing for antimicrobial resistance (NG-STAR) clonal complexes (CCs) 63, 168, and 213 (with mtrD/mtrR promoter mosaic 2) and the novel NG-STAR CC1031 (semi-mosaic mtrD variant 13), associated with men who have sex with men and anorectal or oropharyngeal infections. The declining cefixime resistance (0·5%, nine of 1932) and negligible ceftriaxone resistance (0·1%, one of 1932) was largely because of the progressive disappearance of NG-STAR CC90 (with mosaic penA allele), which was predominant in 2013. No known resistance determinants for novel antimicrobials (zoliflodacin, gepotidacin, and lefamulin) were found. Interpretation: Azithromycin-resistant clones, mainly with mtrD mosaic or semi-mosaic variants, appear to be stabilising at a relatively high level in the EEA. This mostly low-level azithromycin resistance might threaten the recommended ceftriaxone-azithromycin therapy, but the negligible ceftriaxone resistance is encouraging. The decreased genomic population diversity and increased clonality could be explained in part by the COVID-19 pandemic resulting in lower importation of novel strains into Europe.
An inside out journey: biogenesis, ultrastructure and proteomic characterisation of the ectoparasitic flatworm Sparicotyle chrysophrii extracellular vesicles
(BioMed Central (BMC), 2024-04-03) Riera-Ferrer, Enrique; Mazanec, Hynek; Mladineo, Ivona; Konik, Peter; Piazzon, M Carla; Kuchta, Roman; Palenzuela, Oswaldo; Estensoro, Itziar; Sotillo, Javier; Sitjà-Bobadilla, Ariadna; Consejo Superior de Investigaciones Científicas (España); Conferencia de Rectores de las Universidades Españolas; Ministerio de Ciencia e Innovación (España); Agencia Estatal de Investigación (España); Unión Europea. Fondo Europeo de Desarrollo Regional (FEDER/ERDF); Unión Europea. Comisión Europea. NextGenerationEU; Generalitat Valenciana (España); Czech Science Foundation
Background: Helminth extracellular vesicles (EVs) are known to have a three-way communication function among parasitic helminths, their host and the host-associated microbiota. They are considered biological containers that may carry virulence factors, being therefore appealing as therapeutic and prophylactic target candidates. This study aims to describe and characterise EVs secreted by Sparicotyle chrysophrii (Polyopisthocotyla: Microcotylidae), a blood-feeding gill parasite of gilthead seabream (Sparus aurata), causing significant economic losses in Mediterranean aquaculture. Methods: To identify proteins involved in extracellular vesicle biogenesis, genomic datasets from S. chrysophrii were mined in silico using known protein sequences from Clonorchis spp., Echinococcus spp., Fasciola spp., Fasciolopsis spp., Opisthorchis spp., Paragonimus spp. and Schistosoma spp. The location and ultrastructure of EVs were visualised by transmission electron microscopy after fixing adult S. chrysophrii specimens by high-pressure freezing and freeze substitution. EVs were isolated and purified from adult S. chrysophrii (n = 200) using a newly developed ultracentrifugation-size-exclusion chromatography protocol for Polyopisthocotyla, and EVs were characterised via nanoparticle tracking analysis and tandem mass spectrometry. Results: Fifty-nine proteins involved in EV biogenesis were identified in S. chrysophrii, and EVs compatible with ectosomes were observed in the syncytial layer of the haptoral region lining the clamps. The isolated and purified nanoparticles had a mean size of 251.8 nm and yielded 1.71 × 108 particles · mL-1. The protein composition analysis identified proteins related to peptide hydrolases, GTPases, EF-hand domain proteins, aerobic energy metabolism, anticoagulant/lipid-binding, haem detoxification, iron transport, EV biogenesis-related, vesicle-trafficking and other cytoskeletal-related proteins. Several identified proteins, such as leucyl and alanyl aminopeptidases, calpain, ferritin, dynein light chain, 14-3-3, heat shock protein 70, annexin, tubulin, glutathione S-transferase, superoxide dismutase, enolase and fructose-bisphosphate aldolase, have already been proposed as target candidates for therapeutic or prophylactic purposes. Conclusions: We have unambiguously demonstrated for the first time to our knowledge the secretion of EVs by an ectoparasitic flatworm, inferring their biogenesis machinery at a genomic and transcriptomic level, and by identifying their location and protein composition. The identification of multiple therapeutic targets among EVs' protein repertoire provides opportunities for target-based drug discovery and vaccine development for the first time in Polyopisthocotyla (sensu Monogenea), and in a fish-ectoparasite model.
Activity of cefiderocol and innovative β-lactam/β-lactamase inhibitor combinations against isogenic strains of Escherichia coli expressing single and double β-lactamases under high and low permeability conditions
(Elsevier, 2024-05) Blanco-Martín, Tania; Alonso-García, Isaac; González-Pinto, Lucía; Outeda-García, Michelle; Guijarro-Sánchez, Paula; López-Hernández, Inmaculada; Perez-Vazquez, Maria; Aracil, Belen; López-Cerero, Lorena; Fraile-Ribot, Pablo; Oliver, Antonio; Vázquez-Ucha, Juan Carlos; Beceiro, Alejandro; Bou, Germán; Arca-Suárez, Jorge; GEMARA/SEIMC-CIBERINFEC Study Group on the activity and resistance mechanisms to new β-lactams and β-lactamase inhibitors (PROTECT); Instituto de Salud Carlos III; Unión Europea. Fondo Europeo de Desarrollo Regional (FEDER/ERDF); Merck, Sharp & Dohme; Centro de Investigación Biomédica en Red - CIBERINFEC (Enfermedades Infecciosas); RETICS-Investigación en Patología Infecciosa (REIPI-ISCIII) (España); Plan Nacional de I+D+i (España); Xunta de Galicia (España); Ministerio de Ciencia e Innovación (España)
Objectives: To analyse the impact of the most clinically relevant β-lactamases and their interplay with low outer membrane permeability on the activity of cefiderocol, ceftazidime/avibactam, aztreonam/avibactam, cefepime/enmetazobactam, cefepime/taniborbactam, cefepime/zidebactam, imipenem/relebactam, meropenem/vaborbactam, meropenem/xeruborbactam and meropenem/nacubactam against recombinant Escherichia coli strains. Methods: We constructed 82 E. coli laboratory transformants expressing the main β-lactamases circulating in Enterobacterales (70 expressing single β-lactamase and 12 producing double carbapenemase) under high (E. coli TG1) and low (E. coli HB4) permeability conditions. Antimicrobial susceptibility testing was determined by reference broth microdilution. Results: Aztreonam/avibactam, cefepime/zidebactam, cefiderocol, meropenem/xeruborbactam and meropenem/nacubactam were active against all E. coli TG1 transformants. Imipenem/relebactam, meropenem/vaborbactam, cefepime/taniborbactam and cefepime/enmetazobactam were also highly active, but unstable against most of MBL-producing transformants. Combination of β-lactamases with porin deficiency (E. coli HB4) did not significantly affect the activity of aztreonam/avibactam, cefepime/zidebactam, cefiderocol or meropenem/nacubactam, but limited the effectiveness of the rest of carbapenem- and cefepime-based combinations. Double-carbapenemase production resulted in the loss of activity of most of the compounds tested, an effect particularly evident for those E. coli HB4 transformants in which MBLs were present. Conclusions: Our findings highlight the promising activity that cefiderocol and new β-lactam/β-lactamase inhibitors have against recombinant E. coli strains expressing widespread β-lactamases, including when these are combined with low permeability or other enzymes. Aztreonam/avibactam, cefiderocol, cefepime/zidebactam and meropenem/nacubactam will help to mitigate to some extent the urgency of new compounds able to resist MBL action, although NDM enzymes represent a growing challenge against which drug development efforts are still needed.
A panel of recombinant Leishmania donovani cell surface and secreted proteins identifies LdBPK_323600.1 as a serological marker of symptomatic infection
(American Society for Microbiology (ASM), 2024-05-08) Roberts, Adam J; Ong, Han Boon; Clare, Simon; Brandt, Cordelia; Harcourt, Katherine; Takele, Yegnasew; Ghosh, Prakash; Toepp, Angela; Waugh, Max; Matano, Daniel; Färnert, Anna; Adams, Emily; Moreno, Javier; Mbuchi, Margaret; Petersen, Christine; Mondal, Dinesh; Kropf, Pascale; Wright, Gavin J; Wellcome Trust; Unión Europea. Comisión Europea. H2020; Foundation for Innovative New Diagnostics (Suiza); European and Developing Countries Clinical Trials Partnership (EDCTP); Unión Europea. Comisión Europea. Horizonte Europa. ERA-LEARN
Visceral leishmaniasis is a deadly infectious disease and is one of the world's major neglected health problems. Because the symptoms of infection are similar to other endemic diseases, accurate diagnosis is crucial for appropriate treatment. Definitive diagnosis using splenic or bone marrow aspirates is highly invasive, and so, serological assays are preferred, including the direct agglutination test (DAT) or rK39 strip test. These tests, however, are either difficult to perform in the field (DAT) or lack specificity in some endemic regions (rK39), making the development of new tests a research priority. The availability of Leishmania spp. genomes presents an opportunity to identify new diagnostic targets. Here, we use genome data and a mammalian protein expression system to create a panel of 93 proteins consisting of the extracellular ectodomains of the Leishmania donovani cell surface and secreted proteins. We use these panel and sera from murine experimental infection models and natural human and canine infections to identify new candidates for serological diagnosis. We observed a concordance between the most immunoreactive antigens in different host species and transmission settings. The antigen encoded by the LdBPK_323600.1 gene can diagnose Leishmania infections with high sensitivity and specificity in patient cohorts from different endemic regions including Bangladesh and Ethiopia. In longitudinal sampling of treated patients, we observed reductions in immunoreactivity to LdBPK_323600.1 suggesting it could be used to diagnose treatment success. In summary, we have identified new antigens that could contribute to improved serological diagnostic tests to help control the impact of this deadly tropical infectious disease. IMPORTANCE Visceral leishmaniasis is fatal if left untreated with patients often displaying mild and non-specific symptoms during the early stages of infection making accurate diagnosis important. Current methods for diagnosis require highly trained medical staff to perform highly invasive biopsies of the liver or bone marrow which pose risks to the patient. Less invasive molecular tests are available but can suffer from regional variations in their ability to accurately diagnose an infection. To identify new diagnostic markers of visceral leishmaniasis, we produced and tested a panel of 93 proteins identified from the genome of the parasite responsible for this disease. We found that the pattern of host antibody reactivity to these proteins was broadly consistent across naturally acquired infections in both human patients and dogs, as well as experimental rodent infections. We identified a new protein called LdBPK_323600.1 that could accurately diagnose visceral leishmaniasis infections in humans.
Serum GFAP levels correlate with astrocyte reactivity, post-mortem brain atrophy and neurofibrillary tangles
(Oxford University Press, 2024-05-03) Sánchez-Juan, Pascual; Valeriano-Lorenzo, Elizabeth; Ruiz-González, Alicia; Pastor, Ana Belén; Rodrigo Lara, Hector; López-González, Francisco; Zea-Sevilla, María Ascensión; Valentí, Meritxell; Frades, Belen; Ruiz, Paloma; Saiz, Laura; Burgueño-García, Iván; Calero, Miguel; Del Ser, Teodoro; Rábano, Alberto; Fundación Reina Sofía; Unión Europea. Comisión Europea. NextGenerationEU; Instituto de Salud Carlos III; Agencia Estatal de Investigación (España); Ministerio de Ciencia e Innovación (España)
Glial fibrillary acidic protein (GFAP), a proxy of astrocyte reactivity, has been proposed as biomarker of Alzheimer's disease. However, there is limited information about the correlation between blood biomarkers and post-mortem neuropathology. In a single-centre prospective clinicopathological cohort of 139 dementia patients, for which the time-frame between GFAP level determination and neuropathological assessment was exceptionally short (on average 139 days), we analysed this biomarker, measured at three time points, in relation to proxies of disease progression such as cognitive decline and brain weight. Most importantly, we investigated the use of blood GFAP to detect the neuropathological hallmarks of Alzheimer's disease, while accounting for potential influences of the most frequent brain co-pathologies. The main findings demonstrated an association between serum GFAP level and post-mortem tau pathology (β = 12.85; P < 0.001) that was independent of amyloid deposits (β = 13.23; P = 0.02). A mediation analysis provided additional support for the role of astrocytic activation as a link between amyloid and tau pathology in Alzheimer's disease. Furthermore, a negative correlation was observed between pre-mortem serum GFAP and brain weight at post-mortem (r = -0.35; P < 0.001). This finding, together with evidence of a negative correlation with cognitive assessments (r = -0.27; P = 0.005), supports the role of GFAP as a biomarker for disease monitoring, even in the late phases of Alzheimer's disease. Moreover, the diagnostic performance of GFAP in advanced dementia patients was explored, and its discriminative power (area under the receiver operator characteristic curve at baseline = 0.91) in differentiating neuropathologically-confirmed Alzheimer's disease dementias from non-Alzheimer's disease dementias was determined, despite the challenging scenario of advanced age and frequent co-pathologies in these patients. Independently of Alzheimer's disease, serum GFAP levels were shown to be associated with two other pathologies targeting the temporal lobes-hippocampal sclerosis (β = 3.64; P = 0.03) and argyrophilic grain disease (β = -6.11; P = 0.02). Finally, serum GFAP levels were revealed to be correlated with astrocyte reactivity, using the brain GFAP-immunostained area as a proxy (ρ = 0.21; P = 0.02). Our results contribute to increasing evidence suggesting a role for blood GFAP as an Alzheimer's disease biomarker, and the findings offer mechanistic insights into the relationship between blood GFAP and Alzheimer's disease neuropathology, highlighting its ties with tau burden. Moreover, the data highlighting an independent association between serum GFAP levels and other neuropathological lesions provide information for clinicians to consider when interpreting test results. The longitudinal design and correlation with post-mortem data reinforce the robustness of our findings. However, studies correlating blood biomarkers and neuropathological assessments are still scant, and further research is needed to replicate and validate these results in diverse populations.
Sistema de vigilancia de leishmaniosis en gatos en la Comunidad de Madrid
(Sociedad Española de Sanidad Ambiental (SESA), 2024-05) De la Cruz Pérez, M; Chicharro, Carmen; Moreno-Iruela, Inmaculada; García Benzaquén, N; Mañas Urbón, J
Fulminant septic shock due to community-acquired pneumonia caused by Legionella pneumophila SG1 Olda OLDA ST1. Case report
(Elsevier, 2024-06) de Miguel-Balsa, Eva; Jaimez Navarro, Enrique; Cascajero Díaz, Almudena; Gonzalez-Camacho, Fernando; González-Rubio, Juana María; Instituto de Salud Carlos III
Legionellers' desease accounts for 1-8 % of cases of severe community-acquired pneumonia (CAP). Legionella spp. Is the causative organism that can result in respiratory failure, multi-organ dysfunction, sepsis, and death. Therefore, rapid diagnosis and efficient treatment are crucial. We report the clinical and microbiology study of a patient with community-acquired pneumonia caused by Legionella pneumophila, with fatal outcome. After death, the strain causing the infection was identified as Legionella pneumophila serogroup 1, Olda OLDA phenotype and sequence-type 1. This is the first reported case of septic shock and death associated with an isolate of these characteristics.
Characterisation of Legionella Clinical Isolates in Spain from 2012 to 2022
(Multidisciplinary Digital Publishing Institute (MDPI), 2024-06) González-Rubio, Juana María; Cascajero Díaz, Almudena; Baladron-Jimenez, Beatriz Isabel; Gonzalez-Camacho, Fernando; Instituto de Salud Carlos III
Although cases of Legionnaires’ disease are notifiable, data on the phenotypic and genotypic characterisation of clinical isolates are limited. This retrospective study aims to report the results of the characterisation of Legionella clinical isolates in Spain from 2012 to 2022. Monoclonal antibodies from the Dresden panel were used for phenotypic identification of Legionella pneumophila. Genotypic characterisation and sequence type assignment were performed using the Sequence-Based Typing scheme. Of the 1184 samples, 569 were identified as Legionella by culture. Of these, 561 were identified as L. pneumophila, of which 521 were serogroup 1. The most common subgroups were Philadelphia (n = 107) and Knoxville (n = 106). The SBT analysis revealed 130 different STs, with the most common genotypes being ST1 (n = 87), ST23 (n = 57), ST20 (n = 30), and ST42 (n = 29). Knoxville has the highest variability with 32 different STs. ST23 is mainly found in Allentown/France (n = 46) and ST42 in Benidorm (n = 18), whereas ST1 is widely distributed. The results demonstrate that clinical isolates show high genetic diversity, although only a few sequence types (STs) are responsible for most cases. However, outbreaks can also occur with rare genotypes. More data on LD and associated epidemiological studies are needed to establish the risk of an isolate causing outbreak in the future.
Análisis de una instalación tras 20 años de legionelosis ¿Qué hemos aprendido?
(Sociedad Española de Sanidad Ambiental (SESA), 2024-05) Gonzalez-Camacho, Fernando; González-Rubio, Juana María; Cancino-Muñoz, Irving; Coscollá, Mireia
Vigilancia microbiológica del Legionella spp. a partir de muestras de origen ambiental en el periodo 2010-2023 en España
(Sociedad Española de Sanidad Ambiental (SESA), 2024-05) González-Rubio, Juana María; Bocero-Guerrero, Adrián; Cascajero Díaz, Almudena; Gonzalez-Camacho, Fernando
Immune response against the SARS-CoV-2 spike protein in cancer patients after COVID-19 vaccination during the Omicron wave: a prospective study
(Elsevier, 2024-07) Muñoz-Gómez, María José; Ryan, Pablo; Quero-Delgado, Marta; Martin-Vicente, Maria; Cuevas, Guillermo; Valencia, Jorge; Jiménez, Eva; Blanca-López, Natalia; Lara-Álvarez, Miguel Ángel; Hernández-Rivas, José Ángel; Redondo, Gerardo; Mas-Lloret, Vicente; Sepulveda-Crespo, Daniel; Vazquez-Alcaraz, Monica; Torres-Macho, Juan; Martinez, Isidoro; Resino, Salvador; Centro de Investigación Biomédica en Red - CIBERINFEC (Enfermedades Infecciosas); Instituto de Salud Carlos III
Background: Cancer patients often have weakened immune systems, resulting in a lower response to vaccines, especially those receiving immunosuppressive oncological treatment (OT). We aimed to assess the impact of OT on the humoral and T-cell response to the B.1 lineage and Omicron variant following COVID-19 vaccination in patients with solid and hematological neoplasms. Methods: We conducted a prospective study on cancer patients, stratified into OT and non-OT groups, who received a two-dose series of the COVID-19 mRNA vaccine and a booster six months later. The outcomes measured were the humoral (anti-SARS-CoV-2 S IgG titers and ACE2-S interaction inhibition capacity) and cellular (SARS-CoV-2 S-specific T-cell spots per million PBMCs) responses against the B.1 lineage and Omicron variant. These responses were evaluated four weeks after the second dose (n = 98) and eight weeks after the booster dose (n = 71). Results: The humoral response after the second vaccine dose against the B.1 lineage and Omicron variant was significantly weaker in the OT group compared to the non-OT group (q-value<0.05). A booster dose of the mRNA-1273 vaccine significantly improved the humoral response in the OT group, making it comparable to the non-OT group. The mRNA-1273 vaccine, designed for the original Wuhan strain, elicited a weaker humoral response against the Omicron variant compared to the B.1 lineage, regardless of oncological treatment or vaccine dose. In contrast, T-cell responses against SARS-CoV-2, including the Omicron variant, were already present after the second vaccine dose and were not significantly affected by oncological treatments. Conclusions: Cancer patients, particularly those receiving immunosuppressive oncological treatments, should require booster doses and adapted COVID-19 vaccines for new SARS-CoV-2 variants like Omicron. Future studies should evaluate the durability of the immune response and the efficacy of individualized regimens.
COVID-19-associated pulmonary aspergillosis (CAPA) in hematological patients: Could antifungal prophylaxis be necessary? A nationwide study
(Elsevier, 2024-06) Tamayo-Velasco, Álvaro; López-Herrero, Rocío; Gómez-García, Lara María; Sánchez-de Prada, Laura; Aguilar-Monserrate, Gerardo; Martín-Fernández, Marta; Bardají-Carrillo, Miguel; Álvaro-Meca, Alejandro; Tamayo, Eduardo; Resino, Salvador; Miramontes-González, José Pablo; Peñarrubia-Ponce, María Jesús; Instituto de Salud Carlos III
Background: COVID-19-associated pulmonary aspergillosis (CAPA) has emerged as a relatively common complication. Multiple studies described this relationship in critical patients, however its incidence and outcome in other risk groups such as immunosuppressed patients remains unknown. In this sense, we aimed to evaluate the rates and outcomes of CAPA in hematological patients and according to the different hematological malignances, comparing to invasive pulmonary aspergillosis (IPA) in non-COVID-19 ones. Methods: Nationwide, population-based and retrospective observational cohort study including all adult patients with hematological malignancies admitted in Spain since March 1, 2020 to December 31, 2021. The main outcome variable was the diagnosis of IPA during hospitalization in hematological patients with or without COVID-19 at admission. The rate of CAPA compared to IPA in non-COVID-19 patients in each hematological malignancy was also performed, as well as survival curve analysis. Findings: COVID-19 was diagnosed in 3.85 % (4367 out of 113,525) of the hematological adult inpatients. COVID-19 group developed more fungal infections (5.1 % vs. 3 %; p < 0.001). Candida spp. showed higher rate in non-COVID-19 (74.2 % vs. 66.8 %; p = 0.015), meanwhile Aspergillus spp. confirmed its predominance in COVID-19 hematological patients (35.4 % vs. 19.1 %; p < 0.001). IPA was diagnosed in 703 patients and 11.2 % (79 cases) were CAPA. The multivariate logistic regression analysis found that the diagnosis of COVID-19 disease at hospital admission increased more than two-fold IPA development [OR: 2.5, 95CI (1.9-3.1), p < 0.001]. B-cell malignancies - specifically B-cell non-Hodgkin lymphoma, multiple myeloma, chronic lymphocytic leukemia and acute lymphoblastic leukemia - showed between four- and six-fold higher CAPA development and 90-day mortality rates ranging between 50 % and 72 %. However, myeloid malignancies did not show higher CAPA rates compared to IPA in non-COVID-19 patients. Conclusion: COVID-19 constitutes an independent risk factor for developing aspergillosis in B-cell hematological malignancies and the use of antifungal prophylaxis during hospitalizations may be warranted.

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