Artículos (Microbiología)

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Long-Term Impact of an Educational Antimicrobial Stewardship Program on Hospital-Acquired Candidemia and Multidrug-Resistant Bloodstream Infections: A Quasi-Experimental Study of Interrupted Time-Series Analysis
(2017-12-15) Molina, José; Peñalva, Germán; Gil Navarro, María Victoria; Praena, Julia; Lepe Jiménez, José Antonio; Olbrich, Peter; Jiménez-Mejías, Manuel Enrique; Garnacho Montero, José; Valencia, R.; Cordero Matia, María Elisa; Cisneros, José Miguel; García Cabrera, Emilio; Universidad de Sevilla. Departamento de Farmacología, Pediatría y Radiología; Universidad de Sevilla. Departamento de Microbiología; Universidad de Sevilla. Departamento de Medicina Preventiva y Salud Pública; Universidad de Sevilla. Departamento de Medicina; Junta de Andalucía
Background The global crisis of bacterial resistance urges the scientific community to implement intervention programs in healthcare facilities to promote an appropriate use of antibiotics. However, the clinical benefits or the impact on resistance of these interventions has not been definitively proved. Methods We designed a quasi-experimental intervention study with an interrupted time-series analysis. A multidisciplinary team conducted a multifaceted educational intervention in our tertiary-care hospital over a 5-year period. The main activity of the program consisted of peer-to-peer educational interviews between counselors and prescribers from all departments to reinforce the principles of the proper use of antibiotics. We assessed antibiotic consumption, incidence density of Candida and multidrug-resistant (MDR) bacteria bloodstream infections (BSIs) and their crude death rate per 1000 occupied bed days (OBDs). Results A quick and intense reduction in antibiotic consumption occurred 6 months after the implementation of the intervention (change in level, −216.8 defined daily doses per 1000 OBDs; 95% confidence interval, −347.5 to −86.1), and was sustained during subsequent years (average reduction, −19,9%). In addition, the increasing trend observed in the preintervention period for the incidence density of candidemia and MDR BSI (+0.018 cases per 1000 OBDs per quarter; 95% confidence interval, −.003 to .039) reverted toward a decreasing trend of −0.130 per quarter (change in slope, −0.029; −.051 to −.008), and so did the mortality rate (change in slope, −0.015; −.021 to −.008). Conclusions This education-based antimicrobial stewardship program was effective in decreasing the incidence and mortality rate of hospital-acquired candidemia and MDR BSI through sustained reduction in antibiotic use.
Outcomes of the PIRASOA programme, an antimicrobial stewardship programme implemented in hospitals of the Public Health System of Andalusia, Spain: an ecologic study of time-trend analysis
(Elsevier Sci Ltd, 2020) Rodríguez-Baño, Jesús; Pérez-Moreno, M. A.; Peñalva, G.; Garnacho Montero, José; Pinto, C.; Salcedo, I.; Lupión Mendoza, Carmen; Pascual Hernández, Álvaro; Pascual Hernández, Álvaro; Pérez Lozano, María José; Valencia, R.; Cisneros, José Miguel; Universidad de Sevilla. Departamento de Medicina; Universidad de Sevilla. Departamento de Medicina Preventiva y Salud Pública; Universidad de Sevilla. Departamento de Microbiología; Universidad de Sevilla. Departamento de Enfermería
Objectives: Inappropriate antimicrobial use favours the spread of resistance, and multidrug-resistant microorganisms (MDR) are currently of major concern. Antimicrobial stewardship programmes (ASPs) are essential for improving antibiotic use in hospitals. However, their impact on entire healthcare sys tems has not been thoroughly assessed. Our objective was to provide the results of an institutionally supported ASP involving 31 public hospitals in Andalusia, Spain. Methods: We designed an ecologic time-series study from 1 January 2014 to 31 December 2017. Quar terly, data on indicators were collected prospectively, and feedback reports were provided. PIRASOA is an ongoing clinically based quality-improvement programme whose key intervention is the educational interview, regular peer-to-peer interventions between advisors and prescribers to reinforce the appro priate use of antibiotics. Seventy-two indicators were monitored to measure prescribing quality (inap propriate treatments), antimicrobial consumption (defined daily doses per 1000 occupied bed-days), incidence density of MDR per 1000 occupied bed-days and crude mortality rate associated with bloodstream infections. We used Joinpoint regression software to analyse the trends. Results: The quality of antimicrobial prescribing improved markedly, and the inappropriate treatment rate was significantly lower, with quarterly percentage change (QPC) ¼ 3.0%, p < 0.001. Total antimi crobial consumption decreased (QPC ¼ 0.9%, p < 0.001), specifically carbapenems, amoxicillin/clav ulanic acid, quinolones and antifungal agents, whereas antipseudomonal cephalosporin use increased. While the incidence of MDR showed a sustained decreasing trend (QPC ¼ 1.8%; p 0.002), the mortality of patients with bloodstream infections remained stable (QPC ¼ 0.2%, p 0.605). Conclusions: To date, the PIRASOA programme has succeeded in optimizing the use of antimicrobial agents and has had a positive ecologic result on bacterial resistance at level of an entire healthcare system.
Semirapid detection of piperacillin/tazobactam resistance and extended-spectrum resistance to b-lactams/b-lactamase inhibitors in clinical isolates of escherichia coli
(American Society for Microbiology, 2021-10-20) Rodríguez Villodres, Ángel; Gutiérrez Linares, Alicia; Gálvez Benitez, Lydia; Pachón Díaz, Jerónimo; Lepe Jiménez, José Antonio; Smani, Younes; Universidad de Sevilla. Departamento de Microbiología; Universidad de Sevilla. Departamento de Medicina; Instituto de Salud Carlos III; Ministerio de Ciencia e Innovación (MICIN). España
Piperacillin/tazobactam (TZP) is a b-lactam/b-lactamase inhibitor (BL/BLI) recommended for the empirical treatment of severe infections. The excessive and indiscriminate use of TZP has promoted the emergence of TZP-resistant Escherichia coli isolates. Recently, we demonstrated that TZP may contribute to the development of extended-spectrum resistance to BL/BLI (ESRI) in E. coli isolates that are TZP susceptible but have low-level resistance to BL/BLI (resistance to amoxicillin/clavulanic acid [AMC] and/or ampicillin/sulbactam [SAM]). This raises the need for the development of rapid detection systems. Therefore, the objective of this study was to design and validate a method able to detect TZP resistance and ESRI in E. coli. A colorimetric assay based on b-lactam ring hydrolysis by b-lactamases was designed (ESRI test). A total of 114 E. coli isolates from bloodstream and intra-abdominal sources, characterized according to their susceptibility profiles to BL/BLI, were used. Detection of the three most frequent b-lactamases involved in BL/BLI resistance (blaTEM, blaOXA-1, and blaSHV) was performed by PCR. The ESRI test was able to detect all the TZP-intermediate/-resistant isolates, as well as all the TZP-susceptible isolates with a capacity for ESRI development. Their median times to results were 5 and 30 min, respectively. All the isolates without resistance to BL/BLI displayed a negative result in the ESRI test. blaTEM was the most frequent b-lactamase gene detected, follow by blaSHV and blaOXA-1. These results demonstrate the efficacy of the ESRI test, showing great clinical potential which could lead to reductions in health costs, ineffective treatments, and inappropriate use of BL/BLI.
Secuenciación del genoma del Sars-CoV-2 en Andalucía, metodología y estudio de las variantes
(Consejo Andaluz de Colegios de Médicos, 2022) Salazar, Adolfo de; Fuentes López, Ana; Viñuela, Laura; Camacho Martínez, Pedro; Chueca, Natalia; Merino Díaz, Laura; Pérez Florido, Javier; Casimiro Soriguer, Carlos S.; Dopazo Blázquez, Joaquín; Lorusso, Nicola; García, Federico; Lepe Jiménez, José Antonio; Universidad de Sevilla. Departamento de Microbiología
La incorporación de las técnicas de secuenciación genómica mediante secuenciación de nueva generación ha revolucionado la microbiología clínica, innovando y mejorando el diagnóstico clínico de las enfermedades infecciosas. Hoy en día, la secuenciación de genoma completo en enfermedades infecciosas tiene multitud de aplicaciones en virología, bacteriología, resistencia antibiótica, epidemiología y salud pública. Con la aparición del SARS-CoV-2, se ha visto subrayada la importancia del análisis y estudio de las secuencias genéticas. Desde la identificación inicial del SARS-CoV-2, hasta la fecha, se han compartido, a nivel mundial, más de 414.575 secuencias genómicas completas a través de bases de datos de acceso público. La capacidad de monitorizar la evolución viral casi en tiempo real tiene un impacto directo en la respuesta de salud pública a la pandemia de COVID-19. En este trabajo se presenta la importancia de la secuenciación genómica en microbiología, enfermedades infecciosas, epidemiología y salud pública, y se describe cómo se ha implementado la secuenciación de SARS-CoV-2 en Andalucía, y cuales son los principales resultados hasta la fecha.
Impact of the COVID-19 Pandemic on Survival in the Patients With the Intra-Abdominal Infections
(Frontiers Media S.A., 2021-10-20) Gálvez Benítez, Lydia; Rodríguez Villodres, Ángel; Álvarez Marín, Rocío; Jiménez Rodríguez, Rosa M.; Lepe Jiménez, José Antonio; Pachón Díaz, Jerónimo; Smani, Younes; Universidad de Sevilla. Departamento de Microbiología; Universidad de Sevilla. Departamento de Cirugía; Universidad de Sevilla. Departamento de Medicina; Instituto de Salud Carlos III; European Commission (EC). Fondo Europeo de Desarrollo Regional (FEDER); Ministerio de Economía y Competitividad (MINECO). España
Objective: To analyze the availability and access to the hospital for the patients with intra-abdominal infections (IAIs) by Escherichia coli (E. coli) as a result of the coronavirus disease 2019 (COVID-19) pandemic and the impact of these changes in the diagnosis and their effects on the death of these patients. Methods: Two prospective observational cohorts of the patients with IAI by E. coli were conducted in 2016 (the pre-COVID-19, n = 108) and in 2020 (during the COVID-19, n = 96) at the University Hospital of Seville, Spain. The demographic and clinical variables of the patients were collected and analyzed. The patients were followed-up for 120 days, until the hospital discharge or death. The bivariate and multivariate analyses were performed. Results: Both the cohorts were homogeneous according to age, sex, emergency surgery cause, immunosuppression, neutropenia, acquisition type, and previous intervention. The patients attended during the COVID-19 had significantly higher Charlson comorbidity index and the more McCabe score, required more emergency surgery, had more severe infections with the higher rates of septic shock and sepsis, and the presence of additional care support such as a nasogastric tube. They were diagnosed later; the time intervals between the symptoms onset (SO) to the first medical contact or surgical intervention (SI) and between the first medical contact to the admission or SI were significantly higher. The death rates during the COVID-19 and the pre-COVID-19 were 16.7 and 6.5%, respectively (p = 0.02). Finally, the multivariate analysis in both the cohorts together identified the patients diagnosed during the COVID-19, the longer period from SO to SI, septic shock, and the Charlson comorbidity index as the independent factors associated with death. Conclusion: This study showed the impact of the COVID-19 pandemic on the clinical outcome and death due to IAI with an extension of the time between SO and SI.
Molecular and Phylogenetic Characterization of the Monkeypox Outbreak in the South of Spain
(John Wiley & Sons, 2024) Casimiro Soriguer, C. S.; Pérez Florido, J.; Lara, M.; Camacho Martínez, P.; Merino Díaz, L.; Pupo Ledo, I.; de Salazar, A.; Fuentes, A.; Vinuela, L.; Chueca, N.; Lepe Jiménez, José Antonio; Garcia, F.; Universidad de Sevilla. Departamento de Microbiología; Ministerio de Ciencia e Innovación (MICIN). España; Instituto de Salud Carlos III
Background and Aim: Until the May 2022 Monkeypox (MPXV) outbreak, which spread rapidly to many non-endemic countries, the virus was considered a viral zoonosis limited to some African countries. The Andalusian circuit of genomic surveillance was rapidly applied to characterize the MPXV outbreak in the South of Spain. Methods: Whole genome sequencing was used to obtain the genomic profiles of samples collected across the south of Spain, representative of all the provinces of Andalusia. Phylogenetic analysis was used to study the relationship of the isolates and the available sequences of the 2022 outbreak. Results: Whole genome sequencing of a total of 160 MPXV viruses from the different provinces that reported cases were obtained. Interestingly, we report the sequences of MPXV viruses obtained from two patients who died. While one of the isolates bore no noteworthy mutations that explain a potential heightened virulence, in another patient the second consecutive genome sequence, performed after the administration of tecovirimat, uncovered a mutation within the A0A7H0DN30 gene, known to be a prime target for tecovirimat in its Vaccinia counterpart. In general, a low number of mutations were observed in the sequences reported, which were very similar to the reference of the 2022 outbreak (OX044336), as expected from a DNA virus. The samples likely correspond to several introductions of the circulating MPXV viruses from the last outbreak. The virus sequenced from one of the two patients that died presented a mutation in a gene that bears potential connections to drug resistance. This mutation was absent in the initial sequencing before treatment.
A single mutation in Securin induces chromosomal instability and enhances cell invasion
(Elsevier, 2013) Mora Santos, María del Mar; Castilla, Carolina; Herrero Ruiz, Joaquín; Giráldez Macías, Servando; Limón Mortés, María Cristina; Sáez, Carmen; Japón Rodríguez, Miguel Ángel; Tortolero García, María Dolores; Romero Portillo, Francisco; Universidad de Sevilla. Departamento de Microbiología; Ministerio de Ciencia e Innovación; Ministerio de Sanidad. España; Junta de Andalucía
Pituitary tumour transforming gene (pttg1) encodes Securin, a protein involved in the inhibition of sister chromatid separation binding to Separase until the onset of anaphase. Separase is a cysteine-protease that degrades cohesin to segregate the sister chromatids to opposite poles of the cell. The amount of Securin is strongly regulated because it should allow Separase activation when it is degraded by the anaphase promoting complex/cyclosome, should arrest the cell cycle after DNA damage, when it is degraded through SKP1-CUL1-βTrCP ubiquitin ligase, and its overexpression induces tumour formation and correlates with metastasis in multiple tumours. Securin is a phosphoprotein that contains 32 potentially phosphorylatable residues. We mutated and analysed most of them, and found a single mutant, hSecT60A, that showed enhanced oncogenic properties. Our fluorescence activated cell sorting analysis, fluorescence in situ hybridisation assays, tumour cell migration and invasion experiments and gene expression by microarrays analysis clearly involved hSecT60A in chromosomal instability and cell invasion. These results show, for the first time, that a single mutation in pttg1 is sufficient to trigger the oncogenic properties of Securin. The finding of this point mutation in patients might be used as an effective strategy for early detection of cancer.
ginmappeR: an unified approach for integrating gene and protein identifiers across biological sequence databases
(Oxford Univ Press, 2024-08-29) Sola Espinosa, Fernando Luis; Ayala Hernández, Daniel; Pulido, Marina R.; Ayala, Rafael; López Cerero, Lorena; Hernández Salmerón, Inmaculada Concepción; Ruiz Cortés, David; Universidad de Sevilla. Departamento de Lenguajes y Sistemas Informáticos; Universidad de Sevilla. Departamento de Microbiología; Ministerio de Ciencia e Innovación, España; Instituto de Carlos III; Ministerio de Universidades
The proliferation of biological sequence data, due to developments in molecular biology techniques, has led to the creation of numerous open access databases on gene and protein sequencing. However, the lack of direct equivalence between identifiers across these databases difficults data integration. To address this challenge, we introduce ginmappeR, an integrated R package facilitating the translation of gene and protein identifiers between databases. By providing a unified interface, ginmappeR streamlines the integration of diverse data sources into biological workflows, so it enhances efficiency and user experience.
Association of Microbiological Factors with Mortality in Escherichia Coli Bacteraemia Presenting with Sepsis/septic Shock: A Prospective Cohort Study
(Elsevier, 2024) Maldonado, Natalia; López Hernández, Inmaculada; López-Cortes, Luis Eduardo; Martínez Pérez-Crespo, Pedro María; Retamar Gentil, Pilar; García Montaner, Andrea; De la Rosa Riestra, Sandra; Sousa Domínguez, Adrián; Goikoetxea, Josune; Pulido Navazo, Ángeles; Pascual Hernández, Álvaro; Rodríguez-Baño, Jesús; Universidad de Sevilla. Departamento de Microbiología
Objectives: This study aimed to determine the association of Escherichia coli microbiological factors with 30-day mortality in patients with bloodstream infection (BSI) presenting with a dysregulated response to infection (i.e. sepsis or septic shock). Methods: Whole-genome sequencing was performed on 224 E coli isolates of patients with sepsis/septic shock, from 22 Spanish hospitals. Phylogroup, sequence type, virulence, antibiotic resistance, and pathogenicity islands were assessed. A multivariable model for 30-day mortality including clinical and epidemiological variables was built, to which microbiological variables were hierarchically added. The predictive capacity of the models was estimated by the area under the receiver operating characteristic curve (AUROC) with 95% confidence intervals (CI). Results: Mortality at day 30 was 31% (69 patients). The clinical model for mortality included (adjusted OR; 95% CI) age (1.04; 1.02e1.07), Charlson index 3 (1.78; 0.95e3.32), urinary BSI source (0.30; 0.16 e0.57), and active empirical treatment (0.36; 0.11e1.14) with an AUROC of 0.73 (95% CI, 0.67e0.80). Addition of microbiological factors selected clone ST95 (3.64; 0.94e14.04), eilA gene (2.62; 1.14e6.02), and astA gene (2.39; 0.87e6.59) as associated with mortality, with an AUROC of 0.76 (0.69e0.82). Discussion: Despite having a modest overall contribution, some microbiological factors were associated with increased odds of death and deserve to be studied as potential therapeutic or preventive targets.
Antimicrobial Activity of Anionic Bis(N-Heterocyclic Carbene) Silver Complexes
(Multidisciplinary Digital Publishing Institute, 2024-09-27) Carrasco Carrasco, Carlos Jesús; Montilla Ramos, Francisco Javier; Villalobo Polo, Eduardo; Angulo, Manuel; Álvarez, Eleuterio; Galindo del Pozo, Agustín; Universidad de Sevilla. Departamento de Microbiología; Ministerio de Ciencia e Innovación (MICIN). España; Universidad de Sevilla
The antimicrobial properties of a series of anionic bis(carbene) silver complexes Na3[Ag(NHCR)2] were investigated (2a–2g and 2c′, where NHCR is a 2,2′-(imidazol-2-ylidene)dicarboxylate-type N-heterocyclic carbene). The complexes were synthesized by the interaction of imidazolium dicarboxylate compounds with silver oxide in the presence of aqueous sodium hydroxide. Complexes 2f,g were characterized analytically and spectroscopically, and the ligand precursor 1f and complexes 2c and 2g were structurally identified by X-ray diffraction methods. The anions of 2c and 2g, [Ag(NHCR)2]3−, showed a typical linear disposition of Ccarbene-Ag-Ccarbene atoms and an uncommonly eclipsed conformation of carbene ligands. The antimicrobial properties of complexes 2a–g, which contains chiral (2b–2e and 2c′) and non-chiral derivatives (2a,f,g), were evaluated against Gram-negative bacteria, Escherichia coli and Pseudomonas aeruginosa, and a Gram-positive bacterium, Staphylococcus aureus. From the observed values of the minimal inhibitory concentration and minimal bactericidal concentration, complexes 2a and 2b showed the best antimicrobial activity against all strains. An interesting chirality–antimicrobial relationship was found, and eutomer 2c′ showed better activity than its enantiomer 2c against the three bacteria. Furthermore, these complexes were investigated experimentally and theoretically by 109Ag nuclear magnetic resonance, and the electronic and steric characteristics of the dianionic carbene ligands were also examined.
Genetic Features of BEL-1-Producing and KPC-2-Producing E. coli from Hospital Wastewater: Human Source or Sewages Adaptation
(Springer Nature, 2024) Romero Oraá, Laura; Pulido, Marina R.; Galán, Fatima; Palacios, María Victoria García; Pascual Hernández, Álvaro; López Cerero, Lorena; Universidad de Sevilla. Departamento de Microbiología; Ministerio de Economía y Competitividad (MINECO). España
Hospital sewage is an ecosystem that facilitates the transfer of antibiotic and heavy metal resistance genes and the interaction of human and environmental bacteria. In this environment, we have detected the presence of 7 KPC-2 and BEL-1 co-producing E. coli isolates of two different clones over a 10-month period in the same hospital. All isolates carried blaKPC-2 and the operon mer on the same IncP plasmid of similar size and an IncN plasmid of different size each clone carrying blaBEL-1. Both IncN-blaBEL-1 plasmids shared a 77 kb region containing blaBEL-1 alongside with fosE, blaOXA-10 and aac(6’)-1b genes in a class 3 integron within a Tn3 transposon. The major IncN plasmid contained in addition a region homolog to P1-like bacteriophage RCS47, including the lytic RepL and lysogenic proteins, but other phage regions were incomplete. The characters such as the temporal persistence in sewage, the absence of colonized patients in the hospital or in the region, the presence of a p1 phage-plasmid fusion and the infrequent class 3 integron as genetic platform would indicate that BEL-1-producing isolates could have been generated in situ by adaptation to human sewage. Part of the microbiota in these discharges could be explained by the interactions of sewage ecosystems and not derive directly from the hospital.
Whole-genome characterisation of Escherichia coli isolates from patients with bacteraemia presenting with sepsis or septic shock in Spain: a multicentre cross-sectional study
(Elsevier, 2024-04) Maldonado, Natalia; López Hernández, Inmaculada; García Montaner, Andrea; López-Cortes, Luis Eduardo; Martínez Pérez-Crespo, Pedro María; Retamar Gentil, Pilar; Rodríguez-Baño, Jesús; Pascual Hernández, Álvaro; Universidad de Sevilla. Departamento de Medicina; Universidad de Sevilla. Departamento de Microbiología
Background Escherichia coli is the most frequent cause of bloodstream infections (BSIs). About one-third of patients with BSIs due to E coli develop sepsis or shock. The objective of this study is to characterise the microbiological features of E coli blood isolates causing sepsis or septic shock to provide exploratory information for future diagnostic, preventive, or therapeutic interventions. Methods Ecolibloodisolatesfromamulticentrecross-sectionalstudyofpatientsolderthan14yearspresentingwithsepsis or septic shock (according to the Third International Consensus Definitions for Sepsis and Septic Shock criteria) from hospitals in Spain between Oct 4, 2016, and Oct 15, 2017, were studied by whole-genome sequencing. Phylogroups, sequence types (STs), serotype, FimH types, antimicrobial resistance (AMR) genes, pathogenicity islands, and virulence factors were identified. Susceptibility testing was performed by broth microdilution. The main outcome of this study was the characterisation of the E coli blood isolates in terms of population structure by phylogroups, groups (group 1: phylogroups B2, F, and G; group 2: A, B1, and C; group 3: D), and STs and distribution by geographical location and bloodstream infection source. Other outcomes were virulence score and prevalence of virulence-associated genes, pathogenicity islands, AMR, and AMR-associated genes. Frequencies were compared using χ2 or Fisher’s exact tests, and continuous variables using the Mann-Whitney test, with Bonferroni correction for multiple comparisons. Findings We analysed 224 isolates: 140 isolates (63%) were included in phylogenetic group 1, 52 (23%) in group 2, and 32 (14%) in group 3. 85 STs were identified, with four comprising 44% (n=98) of the isolates: ST131 (38 [17%]), ST73 (25 [11%]), ST69 (23 [10%]), and ST95 (12 [5%]). No significant differences in phylogroup or ST distribution were found according to geographical areas or source of bloodstream infection, except for ST95, which was more frequent in urinary tract infections than in other sources (11 [9%] of 116 vs 1 [1%] of 108, p=0⋅0045). Median virulence score was higher in group 1 (median 25⋅0 [IQR 20⋅5–29⋅0) than in group 2 (median 14⋅5[9⋅0–20⋅0]; p<0⋅0001) and group 3 (median 21 [16⋅5–23⋅0]; p<0⋅0001); prevalence of several pathogenicity islands was higher in group 1. No significant differences were found between phylogenetic groups in proportions of resistance to antibiotics. ST73 had higher median virulence score (32 [IQR 29–35]) than the other predominant clones (median range 21–28). Some virulence genes and pathogenicity islands were significantly associated with each ST. ST131 isolates had higher prevalence of AMR and a higher proportion of AMR genes, notably blaCTX-M-15 and blaOXA-1. Interpretation In this exploratory study, the population structure of E coli causing sepsis or shock was similar to previous studies that included all bacteraemic isolates. Virulence genes, pathogenicity islands, and AMR genes were not randomly distributed among phylogroups or STs. These results provide a comprehensive characterisation of invasive E coli isolates causing severe response syndrome. Future studies are required to determine the contribution of these microbiological factors to severe clinical presentation and worse outcomes in patients with E coli bloodstream infection.
Implications of Two-component Systems EnvZ/OmpR and BaeS/BaeR in in Vitro Temocillin Resistance in Escherichia Coli
(Oxford University Press, 2024) Pérez Palacios, Patricia; Rodríguez Ochoa, José Luis; Velázquez Escudero, Ana; Rodríguez-Baño, Jesús; Rodríguez Martínez, José Manuel; Pascual Hernández, Álvaro; Docobo Pérez, Fernando; Universidad de Sevilla. Departamento de Microbiología; Universidad de Sevilla. Departamento de Medicina; Junta de Andalucía; Sociedad Europea de Microbiología Clínica y Enfermedades Infecciosas (ECCMID)
Background: BaeS/BaeR is a two-component system of Escherichia coli that controls the expression of porins and efflux pumps. Its role in beta-lactam resistance is limited. Objectives: To study the role of baeS/baeR two-component system in temocillin resistance in E. coli. Methods: E. coli strain BW25113 and single-gene deletion mutants related to two-component systems were collected from the KEIO collection. Double-gen deletion mutants were generated. Temocillin-resistant mutant frequencies were determined at 32 mg/L. E. coli BW25113 mutants were selected by selective pressure from serial passages. Biological costs were analysed by growth curves. Genomes of the generated mutants were sequenced. The expression level of the mdtA, mdtB, mdtC, acrD and tolC in the ΔbaeS mutant was determined by RT–PCR (with/without temocillin exposure). Results: The frequency of temocillin mutants ranged from 2.12 × 10−8 to 4.51 × 10−8 in single-porin mutants. No mutants were recovered from E. coli BW25113 (>10−9). Selection of temocillin-resistant variants by serial passage yielded mutants up to 128 mg/L. Mutations were found in the baeS gene. Temocillin MICs ranged from 4 to 32 mg/L (highest MICs for ΔbaeS and ΔompR). The efflux pumps mdtA, mdtB, mdtC and acrD pumps were overexpressed 3–10-fold in the presence of temocillin in ΔbaeS compared to control. Conclusions: Mutations in the sensor histidine kinase, baeS, may be involved in temocillin resistance through the expression of the efflux pumps mdtABC and acrD. In addition, the low mutation rate may be a good predictor of temocillin activity.
Impact of Suppression of the SOS Response on Protein Expression in Clinical Isolates of Escherichia Coli under Antimicrobial Pressure of Ciprofloxacin
(Frontiers Media S.A., 2024) Recacha, Esther; Kuropka, Benno; Díaz Díaz, Sara; García Montaner, Andrea; González Tortuero, Enrique; Docobo Pérez, Fernando; Rodríguez Rojas, Alexandro; Rodríguez Martínez, José Manuel; Universidad de Sevilla. Departamento de Microbiología; Instituto de Salud Carlos III
Introduction/objective: Suppression of the SOS response in combination with drugs damaging DNA has been proposed as a potential target to tackle antimicrobial resistance. The SOS response is the pathway used to repair bacterial DNA damage induced by antimicrobials such as quinolones. The extent of lexA-regulated protein expression and other associated systems under pressure of agents that damage bacterial DNA in clinical isolates remains unclear. The aim of this study was to assess the impact of this strategy consisting on suppression of the SOS response in combination with quinolones on the proteome profile of Escherichia coli clinical strains. Materials and methods: Five clinical isolates of E. coli carrying different chromosomally- and/or plasmid-mediated quinolone resistance mechanisms with different phenotypes were selected, with E. coli ATCC 25922 as control strain. In addition, from each clinical isolate and control, a second strain was created, in which the SOS response was suppressed by deletion of the recA gene. Bacterial inocula from all 12 strains were then exposed to 1xMIC ciprofloxacin treatment (relative to the wild-type phenotype for each isogenic pair) for 1 h. Cell pellets were collected, and proteins were digested into peptides using trypsin. Protein identification and label-free quantification were done by liquid chromatography-mass spectrometry (LC–MS) in order to identify proteins that were differentially expressed upon deletion of recA in each strain. Data analysis and statistical analysis were performed using the MaxQuant and Perseus software. Results: The proteins with the lowest expression levels were: RecA (as control), AphA, CysP, DinG, DinI, GarL, PriS, PsuG, PsuK, RpsQ, UgpB and YebG; those with the highest expression levels were: Hpf, IbpB, TufB and RpmH. Most of these expression alterations were strain-dependent and involved DNA repair processes and nucleotide, protein and carbohydrate metabolism, and transport. In isolates with suppressed SOS response, the number of underexpressed proteins was higher than overexpressed proteins. Conclusion: High genomic and proteomic variability was observed among clinical isolates and was not associated with a specific resistant phenotype. This study provides an interesting approach to identify new potential targets to combat antimicrobial resistance.
Characterizing Replisome Disassembly in Human Cells
(Elsevier, 2024) Jones, Rebecca M.; Herrero Ruiz, Joaquín; caramuzza, Shaun; Nath, Sarmi; Liu, Chaoyu; Henklewska, Marta; Natsume, Toyoaki; Bristow, Robert G.; Romero Portillo, Francisco; Kanemaki, Masato T.; Gambus, Agnieszka; Universidad de Sevilla. Departamento de Microbiología; Wellcome Trust. UK
To ensure timely duplication of the entire eukaryotic genome, thousands of replication machineries (replisomes) act on genomic DNA at any time during S phase. In the final stages of this process, replisomes are unloaded from chromatin. Unloading is driven by polyubiquitylation of MCM7, a subunit of the terminated replicative helicase, and processed by p97/VCP segregase. Most of our knowledge of replication termination comes from model organisms, and little is known about how this process is executed and regulated in human somatic cells. Here we show that replisome disassembly in this system requires CUL2LRR1-driven MCM7 ubiquitylation, p97, and UBXN7 for unloading and provide evidence for “backup” mitotic replisome disassembly, demonstrating conservation of such mechanisms. Finally, we find that small-molecule inhibitors against Cullin ubiquitin ligases (CULi) and p97 (p97i) affect replisome unloading but also lead to induction of replication stress in cells, which limits their usefulness to specifically target replisome disassembly processes.
Activity of Cefepime, Carbapenems and New β-lactam/β-lactamase Inhibitor Combinations on Enterobacter Cloacae Complex and Klebsiella Aerogenes in Spain (SMART 2016–2022)
(Oxford University Press, 2024) Rodríguez Villodres, Ángel; Lepe Balsalobre, Esperanza; Ortiz de la Rosa, José Manuel; Giner Almaraz, Salvador; González De Herrero, Elisa; Cercenado, Emilia; García Fernández, Sergio; Benito, Rafael; Ponz Mir, Ricardo; Cantón, Rafael; Lepe Jiménez, José Antonio; Universidad de Sevilla. Departamento de Microbiología; Ministerio de Ciencia, Innovación y Universidades (MICINN). España
Objectives: To analyse the susceptibility profile to cefepime, carbapenems and new β-lactam/β-lactamase inhibitor combinations in Enterobacter cloacae complex and Klebsiella aerogenes isolated from intra-abdominal, urinary, respiratory and bloodstream infections in the SMART (Study for Monitoring Antimicrobial Resistance Trends) surveillance study in Spain. Methods: The susceptibilities of 759 isolates (473 E. cloacae complex and 286 K. aerogenes) collected in 11 Spanish hospitals from 2016 to 2022 were analysed following the EUCAST 2023 criteria. Molecular characterization looking for β-lactamase genes was performed through PCR and DNA sequencing analysis. Results: E. cloacae complex showed resistance to third-generation cephalosporins in 25% of the cases, whereas K. aerogenes was resistant in 35%. Regarding cefepime, resistance in E. cloacae was higher (10%) than in K. aerogenes (2%). Carbapenems showed >85% activity in both microorganisms. Ceftazidime/avibactam, imipenem/relebactam and meropenem/vaborbactam had good activity against these microorganisms (>95%). In contrast, the activity of ceftolozane/tazobactam was lower (80%). A high proportion of the isolates resistant to new β-lactam/β-lactamase inhibitor combinations carried a carbapenemase, mainly OXA-48-like and VIM-1. Conclusions: Ceftazidime/avibactam, imipenem/relebactam and meropenem/vaborbactam show high activity against both E. cloacae complex and K. aerogenes isolates recovered in the SMART-Spain study. In contrast, differences have been found in the case of cefepime, showing more activity against K. aerogenes than E. cloacae complex. These results are useful for antimicrobial stewardship programmes and for the implementation of local and national guidelines.
Survival of infection with TEM beta-lactamase-producing Escherichia coli with Pan-beta-lactam resistance
(Elsevier, 2024) Rodriguez-Villodres, Angel; Ortiz de la Rosa, Jose Manuel; Galvez-Benitez, Lydia; Gascon, Maria Luisa; Penalva, German; Dorado Pardo, Francisco J.; Cisneros, José Miguel; Lepe Jiménez, José Antonio; Universidad de Sevilla. Departamento de Microbiología; Universidad de Sevilla. Departamento de Medicina
Background: Antimicrobial resistance is a critical global health issue, significantly contributing to patient mortality. Recent antibiotic developments have aimed to counteract carbapenemase-producing Enterobacterales; however, the impact of their use on the emergence of antibiotic resistance is unknown. This study investigates the first case of a non-carbapenemase-producing, pan-β-lactam-resistant Escherichia coli strain from a patient previously treated with ceftolozane-tazobactam and cefiderocol. Methods: This study describes the clinical progression of a 39-year-old ICU patient who developed multiple infections, culminating in the isolation of a pan-β-lactam-resistant E. coli strain (EC554). The resistance profile was characterised through MIC determination, whole-genome sequencing, the use of the β-lactam inactivation method, RT-qPCR, efflux pump inhibition assays, outer membrane protein analysis, and blaTEM transformation. Findings: The EC554 isolate displayed resistance to all tested β-lactams and β-lactam-β-lactamase inhibitor combinations. Whole-genome sequencing revealed four plasmids in EC554, with the only β-lactamase gene being blaTEM-252 on the pEC554-PBR-X1-X1 plasmid. We found that the extremely resistant phenotype was attributable to a combination of different mechanisms: a high expression of TEM-252, efflux pump activity, porin loss, and PBP3 mutations. Interpretation: The findings illustrate the complex interplay of multiple resistance mechanisms in E. coli, highlighting the potential for high-level resistance even without carbapenemase production. This study underscores the importance of comprehensively characterising resistance mechanisms in order to inform effective treatment strategies and mitigate the spread of resistant strains.
Effects of drought and increased temperature on phytochemical traits of the edible halophyte Crithmum maritimum: : Perspectives for future climatic scenarios
(Elsevier, 2024-10) Martins Noguerol, Raquel; Rico Jiménez, Diego; Matías Resina, Luis; Pérez Ramos, Ignacio Manuel; Moreira, Xoaquín; Francisco, Marta; Álvarez Morales, Rosario; Gandullo Tovar, Jacinto Manuel; Moreno Pérez, Antonio Javier; Cambrollé Silva, Jesús; Universidad de Sevilla. Departamento de Biología Vegetal y Ecología; Universidad de Sevilla. Departamento de Microbiología; Agencia Estatal de Investigación. España; Ministerio de Ciencia e Innovación (MICIN). España; Junta de Andalucía; European Commission (EC). Fondo Europeo de Desarrollo Regional (FEDER); Universidad de Sevilla
Climate change, characterised by drought events and rising temperatures, exerts a significant threat to crop productivity and global food security. Halophytes, known for their resilience in harsh conditions, offer promising options for sustainable cultivation alternatives. Our study focused on Crithmum maritimum, commonly known as sea fennel, an edible halophyte with potential in the food and nutraceutical industries, to explore the impacts of drought and increased temperatures on its nutritional and antioxidant profiles. Different C. maritimum populations displayed high nutritional qualities, suitable for consumption despite appearing slight differences among localities. While both drought and increased temperatures affected plant growth and phytochemical profiles, their impact on nutritional value was minor. Surprisingly, drought induced an unexpected decline in phenolic content, challenging the assumption of increased antioxidants in response to water scarcity. Different rates of decrease in leaf production were observed among C. maritimum populations under drought, yet overall, they maintained similar levels, suggesting potential suitability for cultivation in environments with limited water availability. Diverse population-specific responses under climatic treatments revealed different alterations in amino acid and oxidative stress profiles, suggesting diverse adaptive strategies. These findings provide critical insights into C. maritimum adaptability to climate-driven changes, offering valuable information for future agricultural practices.
Mpox in people with advanced HIV infection: a global case series
(Elsevier, 2023-03-18) Mitja, O.; Alemany, A.; Marks, M.; Mora, J. I. L.; Rodríguez-Aldama, J. C.; Silva, M. S. T.; Herrera, E. A. C.; Crabtree-Ramírez, B.; Blanco, J. L.; Girometti, N.; Lepe Jiménez, José Antonio; Orkin, C. M.; SHARE-NET writing group; Universidad de Sevilla. Departamento de Microbiología
Background People living with HIV have accounted for 38–50% of those affected in the 2022 multicountry mpox outbreak. Most reported cases were in people who had high CD4 cell counts and similar outcomes to those without HIV. Emerging data suggest worse clinical outcomes and higher mortality in people with more advanced HIV. We describe the clinical characteristics and outcomes of mpox in a cohort of people with HIV and low CD4 cell counts (CD4 <350 cells per mm3). Methods A network of clinicians from 19 countries provided data of confirmed mpox cases between May 11, 2022, and Jan 18, 2023, in people with HIV infection. Contributing centres completed deidentified structured case report sheets to include variables of interest relevant to people living with HIV and to capture more severe outcomes. We restricted this series to include only adults older than 18 years living with HIV and with a CD4 cell count of less than 350 cells per mm3 or, in settings where a CD4 count was not always routinely available, an HIV infection clinically classified as US Centers for Disease Control and Prevention stage C. We describe their clinical presentation, complications, and causes of death. Analyses were descriptive. Findings We included data of 382 cases: 367 cisgender men, four cisgender women, and ten transgender women. The median age of individuals included was 35 (IQR 30–43) years. At mpox diagnosis, 349 (91%) individuals were known to be living with HIV; 228 (65%) of 349 adherent to antiretroviral therapy (ART); 32 (8%) of 382 had a concurrent opportunistic illness. The median CD4 cell count was 211 (IQR 117–291) cells per mm3, with 85 (22%) individuals with CD4 cell counts of less than 100 cells per mm3 and 94 (25%) with 100–200 cells per mm3. Overall, 193 (51%) of 382 had undetectable viral load. Severe complications were more common in people with a CD4 cell count of less than 100 cells per mm3 than in those with more than 300 cells per mm3, including necrotising skin lesions (54% vs 7%), lung involvement (29% vs 0%) occasionally with nodules, and secondary infections and sepsis (44% vs 9%). Overall, 107 (28%) of 382 were hospitalised, of whom 27 (25%) died. All deaths occurred in people with CD4 counts of less than 200 cells per mm3. Among people with CD4 counts of less than 200 cells per mm3, more deaths occurred in those with high HIV viral load. An immune reconstitution inflammatory syndrome to mpox was suspected in 21 (25%) of 85 people initiated or re-initiated on ART, of whom 12 (57%) of 21 died. 62 (16%) of 382 received tecovirimat and seven (2%) received cidofovir or brincidofovir. Three individuals had laboratory confirmation of tecovirimat resistance. Interpretation A severe necrotising form of mpox in the context of advanced immunosuppression appears to behave like an AIDS-defining condition, with a high prevalence of fulminant dermatological and systemic manifestations and death.
Soil microorganisms buffer the reduction in plant growth and physiological performance under combined abiotic stress in the halophyte Salicornia ramosissima
(Elsevier, 2023-11-08) Mateos Naranjo, Enrique; Pérez Romero, Jesús Alberto; Puglielli, Giacomo; López Jurado, Javier; Mesa Marín, Jennifer; Pajuelo Domínguez, Eloísa; Rodríguez Llorente, Ignacio David; Redondo Gómez, Susana; Universidad de Sevilla. Departamento de Biología Vegetal y Ecología; Universidad de Sevilla. Departamento de Microbiología; Ministerio de Ciencia e Innovación (MICIN). España; Agencia Estatal de Investigación. España; European Commission (EC). Fondo Europeo de Desarrollo Regional (FEDER)
The impact of multifactorial abiotic stress combinations on plant functional responses remains controversial, and general patterns of response are yet to emerge. This knowledge gap is particularly relevant for species with innate tolerance to environmental stress. Using the halophyte Salicornia ramosissima as a model species, we performed a multifactorial study with 16 experimental scenarios that included or not beneficial microorganisms in order to quantify their impact on plant growth, photosynthetic performance, osmotic adjustment and ion homeostasis. The experimental scenarios were characterized by the combination of four factors with two levels (salinity: 171 and 510 mM NaCl; water stress: yes and no; temperature min/max range: 14/25 and 18/29ºC and atmospheric CO2 concentration: 400 and 700 ppm). A plant growth-promoting rhizobacteria (PGPR) consortium was used as a proxy for positive biological interaction. The results revealed that the multifactorial stress combinations triggered unique functional responses, depending on the stress factors involved. However, there was an overall more negative impact on plant functional traits under the most extreme scenario (i.e., 510 mM NaCl + water stress + high temperature). Interestingly, the presence of PGPR was able to reverse this negative influence, although this effect was negligible under non-stressful conditions. Furthermore, the positive effect of PGPR was even magnified when coexisting with elevated atmospheric CO2 concentration. This response is associated with mitigation of the negative impacts of suboptimal factor combinations on plant growth, photosynthetic performance/efficiency, and water/nutrient homeostasis. Therefore, we conclude that the positive impact of microorganisms on halophyte tolerance in complex environmental matrices would only be determinant under extreme conditions in which plant intrinsic tolerance mechanisms would not be sufficient. Remarkably, this effect could be accentuated by increasing atmospheric CO2 concentration.

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