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Microorganisms naturally colonize rock-based materials in outdoor environments, thereby contributing to their degradation. Fungi, especially in tropical environments with abundant water and favorable temperatures, play a significant role in biodegradation. However, many aspects of the microorganism-stone interaction, including fungal colonization dynamics and the impact of treatment applications, remain unclear. This study conducted a four-year in-situ evaluation of fungal community dynamics on limestone surfaces in the Temple of the Warriors at the Chichén Itzá archaeological site in Mexico, focusing on cleaning and treatment using nanoparticles (NPs). These NPs included zinc oxide (ZnO) and CaZn2(OH)6·2H2O (CZ)-based NPs synthesized via sol-gel (CZ-SG) and mechanochemical methods (CZ-MC), as well as CZ/Ca(OH)2-based products (CZ:Ca-SG). The microbial colonization cover was assessed using colorimetric measurements, and the surface was sampled for fungal community isolation and identification. The results demonstrated significant impacts of cleaning and nanomaterial applications on cultivable fungal communities (melanized filamentous, hyaline, and microcolonial fungi), altering composition, dynamics, and stone surface coloration. In particular, ZnO NPs caused 50 % decline in fungal species and individuals, whereas CZ:Ca-SG NPs displaced most species, indicating effective inhibition of the cultivable fungal community. Microcolonial fungi (MCF), known for their tolerance to withstand harsh environmental conditions, were the only fungal group found in the CZ:Ca-SG treatment. In contrast, CZ-SG and CZ-MC increased the abundance of melanized species, resulting in darkening and reduced color intensity. This study highlights the importance of microcolonial fungi that are tolerant to cleaning and coating procedures in the preservation of stone cultural heritage. These findings enhance our understanding of fungal colonization dynamics following treatment and provide valuable, European Union, CONACYT, Depto. de Mineralogía y Petrología, Fac. de Ciencias Geológicas, TRUE, pub

Nanoparticles derived from plant viruses play an important role in nanomedicine due to their biocompatibility, self-assembly and easily-modifiable surface. In this study, we developed a novel platform for increasing antibody sensing using viral nanoparticles derived from turnip mosaic virus (TuMV) functionalized with SARS-CoV-2 receptor binding domain (RBD) through three different methods: chemical conjugation, gene fusion and the SpyTag/SpyCatcher technology. Even though gene fusion turned out to be unsuccessful, the other two constructs were proven to significantly increase antibody sensing when tested with saliva of patients with different infection and vaccination status to SARS-CoV-2. Our findings show the high potential of TuMV nanoparticles in the fields of diagnostics and immunodetection, being especially attractive for the development of novel antibody sensing devices., Ministerio de Universidades (España), Comunidad de Madrid, Universidad Complutense de Madrid, Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Con el objetivo de integrar de manera efectiva los entornos virtuales en las prácticas de Microbiología, se ha analizado la eficacia de las prácticas en formato virtual en comparación con el presencial en la asignatura de Microbiología del Grado de Podología. Tras elegir el formato práctico presencial o virtual, los alumnos realizaron voluntariamente cuestionarios de conocimientos previos y posteriores a las prácticas, con el fin de evaluar su rendimiento. Asimismo, se evaluó el compromiso y satisfacción de los estudiantes. La mayoría de los alumnos eligieron realizar las prácticas en formato presencial, mostrando un mayor compromiso y participación en los cuestionarios. Ambos grupos experimentaron una mejora en el rendimiento, sin diferencias significativas entre formatos. Los estudiantes que participaron en las prácticas expresaron un alto nivel de satisfacción en ambos formatos. Las prácticas virtuales suponen una alternativa adecuada a las prácticas presenciales en la enseñanza de Microbiología., With the aim of effectively integrating virtual environments in Microbiology practices, theeffectiveness of virtual versus face-to-face practices in the Microbiology course of the Degree inPodiatry was analysed. After choosing the face-to-face or virtual practical format, studentsvoluntarily completed pre- and post-practicum knowledge questionnaires to evaluate theirperformance. Students' commitment and satisfaction were also evaluated. Most students chose tocomplete the practices in the face-to-face format and showed a higher level of engagement andparticipation in the questionnaires. Both groups showed improved performance with no significantdifferences between the formats. Students who participated in the practices expressed an elevatedlevel of satisfaction in both formats. Virtual practices are a viable alternative to face-to-face practicesin Microbiology teaching, Depto. de Medicina, Fac. de Medicina, TRUE, pub

Depto. de Medicina, Depto. de Salud Pública y Materno - Infantil, Fac. de Medicina, TRUE, pub

Depto. de Enfermería, Depto. de Medicina, Fac. de Enfermería, Fisioterapia y Podología, Fac. de Medicina, TRUE, pub

Introducción. El objetivo del estudio fue conocer el impacto del consumo de los antibióticos de uso tópico utilizados en oftalmología y sentar las bases para su uso prudente. Material y métodos. Se realizó un estudio cuantitativo descriptivo sobre el consumo de medicamentos entre 1 de enero de 2004 y el 31 de diciembre de 2008. Los medicamentos utilizados para la realización de este estudio, debían cumplir los siguientes criterios: vía de administración tópica y antimicrobianos oftalmológicos sólo o en asociación. Los datos que hacen referencia al número de envases fueron cedidos por Intercontinental Marketing Services (IMS) y los datos de consumo hospitalario se obtuvieron de Sistema Nacional de Salud. Se estudiaron aquellas variables que hacen referencia estrictamente al consumo. Resultados. El mercado de antiinfecciosos oftalmológicos hospitalario y extrahospitalario en 2008 supuso 4,9 y 24,4 millones de euros, respectivamente incrementándose un 5.5% respecto al año anterior. Se consumieron más de 6 millones de envases de los que el 65% se utilizó en preparados monoantibióticos frente al 35% en asociación. Los antibióticos oftálmicos en monoterapia más consumidos fueron los aminoglucósidos (tobramicina y gentamicina) seguido de las quinolonas mientras que en asociación el más consumido fue gramicidina + neomicina + polimixina B. Conclusiones. El mercado de antiinfecciosos oftalmológicos, solos y combinados, utilizados tanto en el medio extrahospitalario como hospitalario, representa casi 30 millones de euros y muestra un crecimiento en torno al 6%. Las rígidas medidas sobre el consumo de antibióticos en general no son tenidas en cuenta en el campo de los antibióticos tópicos oftálmicos., Depto. de Medicina, Depto. de Enfermería, Depto. de Salud Pública y Materno - Infantil, Fac. de Medicina, Fac. de Enfermería, Fisioterapia y Podología, TRUE, pub

Introduction: This study explores effects of pH and inoculum size on imipenem versus tigecycline activity against E. coli, B. fragilis and E. faecalis, both in individual and mixed cultures. Methods: MIC/MBCs (mg/L) of tigecycline and imipenem were 0.12/≥ 16 and 4/4 for E. coli, 0.12/0.5 and ≥ 16/≥ 16 for B. fragilis, and 0.12/≥ 16 and 2/≥ 16 for E. faecalis, respectively. Killing curves in supplemented Brucella broth were performed at pH 7 or 5.8, with two final inocula (≈ 105 or ≈ 107 cfu/ml) of each isolate (individual cultures) and with 1:1:1 mixed inocula. Tubes were 48 h incubated at 37 ºC in anaerobiosis. Final concentrations (estimated concentrations in colon) were 1.50 mg/L for tigecycline and 26.40 mg/L for imipenem, with antibiotic-free curves as controls. Experiments were performed in triplicate. Results: Imipenem showed inoculum effect against E.coli and B. fragilis, with reductions in initial inocula in experiments with standard inocula contrasting with increases in experiments with high inocula (both individual and mixed cultures). Against E. faecalis no inoculum effect for imipenem was observed in individual cultures, with marked reductions in initial inocula regardless inoculum size. However in mixed experiments the indirect protection of E. faecalis by the two gramnegatives resulted in bacterial regrowth. This protection was inoculum-dependant since it occurred with high but not with standard inocula. Tigecycline reduced initial inocula of the three isolates regardless culture type (individual/mixed) or experimental conditions (pH/inocula size), with lower reductions for the tolerant E. faecalis. Conclusion: Carbapenemase activity was inoculum-dependant for self-protection and indirect protection of E. faecalis., Depto. de Medicina, Fac. de Medicina, TRUE, pub

Inflammatory processes have been shown to modify tryptophan (Trp) metabolism. Gut microbiota appears to play a significant role in the induction of peripheral and central inflammation. Ethanol (EtOH) exposure alters gut permeability, but its effects on Trp metabolism and the involvement of gut microbiota have not been studied. We analyzed several parameters of gut-barrier and of peripheral and central Trp metabolism following 2 different EtOH consumption patterns in mice, the binge model, drinking in the dark (DID), and the chronic intermittent (CI) consumption paradigm. Antibiotic treatment was used to evaluate gut microbiota involvement in the CI model. Mice exposed to CI EtOH intake, but not DID, show bacterial translocation and increased plasma LPS immediately after EtOH removal. Gut-barrier permeability to FITC-dextran is increased by CI, and, furthermore, intestinal epithelial tight-junction (TJ) disruption is observed (decreased expression of zonula occludens 1 and occludin) associated with increased matrix metalloproteinase (MMP)-9 activity and iNOS expression. CI EtOH, but not DID, increases kynurenine (Kyn) levels in plasma and limbic forebrain. Intestinal bacterial decontamination prevents the LPS increase but not the permeability to FITC-dextran, TJ disruption, or the increase in MMP-9 activity and iNOS expression. Although plasma Kyn levels are not affected by antibiotic treatment, the elevation of Kyn in brain is prevented, pointing to an involvement of microbiota in CI EtOH-induced changes in brain Trp metabolism. Additionally, CI EtOH produces depressive-like symptoms of anhedonia, which are prevented by the antibiotic treatment thus pointing to an association between anhedonia and the increase in brain Kyn and to the involvement of gut microbiota.-Giménez-Gómez, P., Pérez-Hernández, M., O'Shea, E., Caso, J. R., Martín-Hernández, D., Cervera, L. A., Centelles. M. L. G.-L., Gutiérrez-Lopez, M. D., Colado, M. I. Changes in brain kynurenine levels via g, Ministerio de Economia y Competitividad (España), Ministerio de Sanidad, Servicios Sociales e Igualdad, Instituto de Salud Carlos III Redes Temáticas de Investigación Cooperativa en Salud-Red de Trastornos Adictivos, Universidad Complutense de Madrid, Depto. de Medicina, Depto. de Farmacología y Toxicología, Fac. de Medicina, TRUE, pub

A crossover trial was conducted to compare hand rub and hand scrub-brush methods for reducing bacterial loads when using propan-1-ol-60% according to European regulations. Both methods significantly reduced the bacterial load immediately after antisepsis, but only the hand rub method achieved significant bacterial load reduction 3 hours after the procedure., Depto. de Medicina, Fac. de Medicina, TRUE, pub

Previous studies have reported the potential emission of nail dust to the indoor air of the podiatrist's workplace during nail drilling procedures. The main objective of this study was to analyse the fungal bioaerosol present in the indoor air of podiatry rooms, including during the nail drilling procedure. Seventeen private clinics were sampled in Madrid during spring and summer. The most prevalent filamentous fungi detected were Cladosporium spp. and Penicillium spp. in both seasons. The presence of Trichophyton rubrum and Trichophyton interdigitale was reported in samples during the drilling procedures, establishing a positive association with the clinical suspicion of onychomycosis (p = 0.002) and the utilization of aspiration systems (p=0.04). The present study concludes that airborne fungal concentrations in podiatrist's clinics are consistent with national and international guidance. Occupational exposure to dermatophytes may briefly occur during definite periods across the daily activity despite the use of dust aspiration systems., Depto. de Medicina, Fac. de Medicina, TRUE, pub

Depto. de Medicina, Fac. de Medicina, TRUE, pub

Analysis of the pulsed-field gel electrophoretic profiles of 82 pneumococcal isolates with reduced susceptibility to ciprofloxacin (RSC) and of 90 co-occurring susceptible isolates indicates a considerable genetic diversity among isolates with RCS and points to a close relation between the two groups. This finding suggests that pneumococci with RCS emerge through independent mutational events., Junta Nacional de Investigação Cientı́fica e Tecnológica (Portugal), Depto. de Medicina, Fac. de Medicina, TRUE, pub

The aim of the study was to explore the structure and growth dynamics of Staphylococcus aureus resistant to methicillin (MRSA) colonies using semithin sections visualized by light microscope. One S. aureus susceptible to methicillin (MSSA) and one MRSA clinical strains were studied. Colonies in agar plates were embedded in epoxy resin after each incubation period (24 h and 48 h) at 37 degrees . Semithin sections of 0.5 µm were stained with toluidine blue and visualized by light microscope. Microscopically, no structural differences were observed between SASM and SARM colonies but differences were observed in both strains between 24 and 48 h incubation periods. Colonies showed two layers clearly differentiated at 24 h independently of the resistance to methicillin: (A) one basal layer with high density of population in contact with culture media, and (B) one superficial layer with a lower density of population. Colonies showed four layers at 48 h: (A) one basal layer with high density of population; (B) one clear layer constituted by very degraded bacterial remains in which can be observed cocci dispersed with their dyeing properties; (C) one mixed layer constituted by viable bacteria and little degraded bacterial remains (D) one superficial layer with a lower density of population than basal layer. Colonial architecture is a complex and time-dependent process., Depto. de Medicina, Fac. de Medicina, TRUE, pub

Objetivo. Valorar si existe una diferencia significativa en tasa de infección tras cirugía de extracción dentaria en dos hospitales de Noruega y España, donde se aplican protocolos de profilaxis quirúrgica diferentes. Material y Métodos. Se realizó un estudio observacional analítico, tipo cohortes retrospectivo, analizando pacientes sanos, sin factores de riesgo, operados de tercer molar incluido en los servicios de maxilofacial de dos hospitales diferentes: St. Olav de Trondheim (Noruega) y Clínico San Carlos de Madrid (España). Se recogieron las variables: edad, número de piezas extraídas, tipo de anestesia, y observaciones recogidas en la historia clínica sobre el curso de la operación. Para valorar el desarrollo infección postoperatoria se recogieron los datos de los pacientes que escogieron el hospital como lugar de retirada de los puntos en el Hospital St. Olav, y en el Hospital Clínico San Carlos se llevó a cabo una encuesta telefónica para conocer el curso de la operación meses después. Resultados. El 11,1% de los pacientes operados en el Hospital St. Olav recibió pauta antibiótica durante una semana tras la operación, mientras que en el Hospital Clínico San Carlos fue del 100%. La tasa de infección tras ésta fue del 15% en el Hospital de St. Olav y del 7,5% en el Hospital Clínico siendo estas diferencias no estadísticamente significativas. Conclusiones. La administración sistemática de antibiótico a pacientes sanos sin factores de riesgo sometidos a extracción quirúrgica del tercer molar retenido es una práctica rutinaria en clínica que no parece estar justificada., Depto. de Medicina, Fac. de Medicina, TRUE, pub

Objectives: the aims of the study were to explore the activity of total and free (according to protein binding) maximal concentrations achieved in serum after multiple doses of voriconazole 400/200 mg and anidulafungin 200/100 mg against Aspergillus fumigatus and Aspergillus flavus and the human albumin or serum effects on antifungal activity. Material and methods: Time-kill curves were performed with two A. fumigatus and two A. flavus strains at voriconazole and anidulafungin Cmax concentrations using different media: a) RPMI broth (Cmax-RPMI); b) RPMI with human serum (Cmax-HS), and c) RPMI with human albumin (Cmax-HAlb). In parallel, free-drug (fCmax) concentrations considering theoretical protein binding were performed in RPMI broth. Aspergillus metabolic activity was measured by the XTT reduction assay. Results: Voriconazol or voriconazole plus anidulafungin reduced >88.4% the metabolic activity of Aspergillus sp. at Cmax-RPMI and fCmax after 48 h of exposition. Anidulafungin alone showed poor metabolic reductions (<80.1% at Cmax- RPMI and <15% at fCmax). Anidulafungin activity, but not voriconazole activity alone or combined decreased in presence of HS or HAlb (more pronounced in A. flavus strains and HAlb). However, anidulafungin Cmax-HS or Cmax-HAlb against A. fumigatus strains were significantly more active (p<0.05) than fCmax in RPMI. These species and culture medium-dependent impact of human protein binding in the activity of anidulafungin was related to macroscopic and microscopic differences among mycelial mat grown in RPMI, HS or HAlb in whose XTT retention was different. Conclusions: Synergism could not be demonstrated due to the high activity showed by voriconazole. Protein binding has not impact on voriconazole activity and this impact is considerably less than predicted by free concentration extrapolated from theoretical binding rate on anidulafungin. The XTT colorimetric assay needs to be standardized for use with Aspergillus spp. since without DM, Depto. de Medicina, Fac. de Medicina, TRUE, pub

Las infecciones de piel y partes blandas (IPPB), y en particular las infecciones del pie de diabético (IPD), presentan complicaciones diagnósticas y terapéuticas, que a menudo desembocan en complicaciones graves. El objetivo de este estudio fue evaluar la eficacia in vitro de cefditoreno y amoxicilina/ ácido clavulánico frente a los microorganismos típicos de las IPD. Se analizaron muestras clínicas de 40 pacientes con IPPB leves mostrando un predominio de especies de Staphylococcus spp. y Streptococcus spp. Cefditoreno mostró frente al 90% de los aislados una potencia superior a la de la amoxicilina/ácido clavulánico. Estos resultados destacan la utilidad de cefditoreno en el tratamiento empírico de la IPD, aunque sería deseable disponer de un mayor tamaño muestral para una mejor validación., Skin and soft tissue infections (SSTIs), and particularly diabetic-related foot infections (DFI), present diagnostic and therapeutic complexities, often leading to severe complications. This study aims to evaluate the in vitro efficacy of cefditoren and amoxicillin/clavulanic acid against typical DFI pathogens. Clinical samples from 40 patients with mild SSTIs were analyzed, revealing a predominance of Staphylococcus spp. and Streptococcus spp. species. Cefditoren exhibited activity against 90% of isolates, with superior potency over amoxicillin/clavulanic acid. These findings underscore the utility of cefditoren in empirical treatment of DFI, although a larger sample size would be desirable for further validation., Meiji Pharma Spain, Depto. de Medicina, Fac. de Medicina, TRUE, pub

Semithin sections of colonies of three ATCC strains (Staphylococcus aureus, Escherichia coli, and Candida albicans) showed that their internal structure had specific patterns that evolved over the time. These patterns generally were defined by the presence of different layers composed of microorganisms with variable population densities and dead cells. The observed structures in this study could be explained as a particular form of biofilm with an air-semisolid interface., Depto. de Medicina, Fac. de Medicina, TRUE, pub

The aim of this study was to determine the survival of human pathogens (Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa) in five natural mineral waters (NMWs) with different properties and mineralization levels. Five NMWs from four Spanish spas with different dry residue at 110 °C were used: A = 76,935 mg/L; B = 1,827 mg/L; C = 808.4 mg/L; D = 283.8 mg/L; and E = 170.4 mg/L. An initial inoculum of 1 × 106 colony forming units (cfu)/mL was used for survival studies. Distilled water, chlorinated tap water and Mueller–Hinton broth were used as controls. Colony counts in all different waters were lower than those achieved with Mueller–Hinton broth over all incubation periods. A direct effect between the bacterial survival and the level of mineralization water was observed. The NMW E with low mineralization level along with the radioactive properties showed the highest antibacterial activity among all NMWs., Depto. de Medicina, Depto. de Radiología, Rehabilitación y Fisioterapia, Fac. de Medicina, TRUE, pub

Objective: To evaluate cefditoren in inducer-substrate combinations to screen for AmpC induction. Methods: 100 clinical isolates (25 P. aeruginosa, 25 E. cloacae, 14 M. morganii, 13 S. marcescens, 12 C. freundii, 7 P. rettgeri, and 4 E. aerogenes) were tested by the Kirby-Bauer disc approximation method using cefditoren and ceftazidime discs as substrates, and cefditoren and imipenem discs as inducers. Results: None of the strains showed induction of AmpC with cefditoren-ceftazidime as inducer-substrate combination. Imipenem-cefditoren as inducer-substrate combination was not useful for evaluating strains of P. aeruginosa since no inhibition zones surrounding the cefditoren disc were found. Among evaluable enterobacteria (those showing substrate inhibition zone), inducible Amp C was detected in 48 out of 63 (76.2%) with cefditoren, and in 33 out of 68 (48.5%) isolates with ceftazidime as substrate. Significantly (p=0.013) higher number of AmpC producers were detected with cefditoren versus ceftazidime (76.2% vs. 48.5%), due to the differences found for E. cloacae (72.8% vs. 21.7%; p=0.0009) and S. marcescens (100% vs. 54.5%; p=0.03). Higher mean reductions of diameters around substrate discs were found for cefditoren (4.17 mm) vs. ceftazidime (3.79 mm), reaching statistical significance (p<0.05) for indol-positive proteae: M. morganii (5.32 mm vs. 3.92 mm) and P. rettgeri (3.47 mm vs. 2.64 mm). Conclusion: Cefditoren showed no induction capability, and when used as substrate (with imipenem as inducer) it offered detection rates of AmpC inducible enterobacteria higher than the imipenem-ceftazidime combination, mainly for Enterobacter spp. and Serratia spp., with higher diameter reductions for indol-positive proteae., Tedec-Meiji Farma S.A., Depto. de Medicina, Fac. de Medicina, TRUE, pub

This study evaluates the dynamic shift in the microbiota at the peri-implant site of titanium (Ti) and zirconia (Zr) implants subjected to experimental peri-implantitis (PI) and, for the first time, of implants made of ceria-stabilized alumina-reinforced zirconia (Ce-TZP/Al), a revolutionary zirconia that is set to play a key role in modern implant dentistry. One- and two-piece (TP) implants, including Ce-TZP/AL TP/G3 glass, were placed bilaterally (six implants/side) in five beagle dogs to mimic a natural vs. ligature-induced PI following a split-mouth design. The experiment spanned 30 weeks from tooth extraction. Both PI models promoted plaque deposition at peri-implant sites. Comparatively, the PI induced by ligatures favored the deposition of anaerobes (p = 0.047 vs. natural). Regardless of the model, the plaque deposition pattern was entirely dependent on the implanted material. Ligated Ti and Zr implant sites accumulated up to 2.14 log CFU/mL unit anaerobic load (p ≤ 0.033 vs. non-ligated implant sites), predominantly comprising obligate anaerobes. Naturally occurring PI induced the deposition of co-occurring networks of obligate anaerobes and less oxygen-dependent bacteria. PI induction favored the enrichment of Ti and Zr sites with bacterial taxa belonging to the orange and red complexes (up to 28% increase naturally and up to 71% in the ligated hemiarch). Anaerobic deposition was significantly lower in ligated Ce-TZP/Al implant sites (p ≤ 0.014 vs. TI and Zr) and independent of the induction model (0.63–1 log units of increase). Facultative bacteria prevailed at Ce-TZP/AL sites. The abundance was lower in the Ce-TZP/AL TP implant. Unlike Ti and Zr sites, taxa from the orange and red complexes were negligible. Biofilms configured at the Ti and Zr sites after ligation-induced PI resemble those found in severe IP. We hypothesize that, although surface properties (surface energy and surface roughness) and physicochemical properties of the substrate play an impo, Unión Europea, Depto. de Medicina, Fac. de Medicina, TRUE, pub

The surface adhesion of bacterial cells and the in vivo biocompatibility of a new ceramic–metal composite made of zirconium dioxide and tantalum were evaluated. Within the framework of an in vitro study using the crystal violet staining and colony counting methods, a relatively similar adhesion of Streptococcus oralis to the 3Y-TZP/Ta biocermet (roughness Ra = 0.12 ± 0.04 µm) and Ti-Al6-V4 titanium alloy (Ra = 0.04 ± 0.01 µm) was found. In addition, in an in vivo preliminary study focused on the histological analysis of a series of rods implanted in the jaws of beagle dogs for a six-month period, the absence of any fibrous tissue or inflammatory reaction at the interface between the implanted 3Y-TZP/Ta biocermets and the new bone was found. Thus, it can be concluded that the developed ceramic–metal biocomposite may be a promising new material for use in dentistry., Ministry of Science and Higher Education (Rusia), Depto. de Medicina, Fac. de Medicina, TRUE, pub

Plant viral nanoparticles (VNPs) are attractive to nanomedicine researchers because of their safety, ease of production, resistance, and straightforward functionalization. In this paper, we developed and successfully purified a VNP derived from turnip mosaic virus (TuMV), a well-known plant pathogen, that exhibits a high affinity for immunoglobulins G (IgG) thanks to its functionalization with the Z domain of staphylococcal Protein A via gene fusion. We selected cetuximab as a model IgG to demonstrate the versatility of this novel TuMV VNP by developing a fluorescent nanoplatform to mark tumoral cells from the Cal33 line of a tongue squamous cell carcinoma. Using confocal microscopy, we observed that fluorescent VNP–cetuximab bound selectively to Cal33 and was internalized, revealing the potential of this nanotool in cancer research., Ministerio de Universidades (España), Ministerio de Ciencia e Innovación (España), Comunidad de Madrid, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (España), ERANet, Universidad Complutense de Madrid, Depto. de Genética, Fisiología y Microbiología, Depto. de Biología Celular, Fac. de Ciencias Biológicas, Fac. de Medicina, TRUE, pub

Ministerio de Economía y Competitividad (MINECO), Comunidad de Madrid, Depto. de Bioquímica y Biología Molecular, Fac. de Ciencias Biológicas, TRUE, unpub

The bioindicator potential of the microeukaryotic community in a full-scale A2O wastewater treatment process was assessed after deactivating the internal recirculation in one of the two parallel treatment lines of the plant. The deactivation of the internal recirculation in one of the lines (NIR-line), led to a strong decrease in nitrogen removal efficiency (53 % ammonia-uptake in the NIR-line versus 97 % in the unaltered line or IR-line), as well as to a lower organic matter removal (95 % BOD5 removal versus 97 % in the IR-line). The microeukaryote community structure changed greatly confirming trends observed in experimental studies and showed a decrease in abundance and richness in the NIR-line. However, this line showed a significant higher abundance of certain ciliate species, such as Epistylis camprubii and Vorticellides infusionum, which are common species in conventional activated sludge. Conversely, the IR-line showed useful indicators of internal recirculation, associated with much higher ammonium and BOD5 removal. These results show that identification to ciliate species level provides a more reliable bioindication in nutrient removal systems, supporting the importance of research on biological parameters, mainly protists, as simple and useful tools for plant operators to manage nutrient removal processes. These observations extend the current boundaries of bioindication., Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Producción Científica, In recent years, nutmeg (Myristica fragans Houtt.) has attracted considerable attention in the field of phytochemistry due to its diverse array of bioactive compounds. However, the potential application of nutmeg as a biorational for crop protection has been insufficiently explored. This study investigated the constituents of a nutmeg hydroethanolic extract via gas chromatography-mass spectrometry and vibrational spectroscopy. The research explored the extract’s activity against phytopathogenic fungi and oomycetes, elucidating its mechanism of action. The phytochemical profile revealed fatty acids (including tetradecanoic acid, 9-octadecenoic acid, n-hexadecanoic acid, dodecanoic acid, and octadecanoic acid), methoxyeugenol, and elemicin as the main constituents. Previously unreported phytochemicals included veratone, gelsevirine, and montanine. Significant radial growth inhibition of mycelia was observed against Botrytis cinerea, Colletotrichum acutatum, Diplodia corticola, Phytophthora cinnamomi, and especially against Fusarium culmorum. Mode of action investigation, involving Saccharomyces cerevisiae labeled positively with propidium iodide, and a mutant strain affected in ERG6, encoding sterol C-24 methyltransferase, suggested that the extract induces a necrotic type of death and targets ergosterol biosynthesis. The evidence presented underscores the potential of nutmeg as a source of new antimicrobial agents, showing particular promise against F. culmorum., Fundación Portuguesa para la Ciencia y la Tecnología (FCT) - (projects UIDB/04050/2020 and UIDB/04033/2020), AgrifoodXXI - (NORTE-01-0145-FEDER- 000041), Junta de Castilla y León y Fondo Europeo de Desarrollo Regional (FEDER) - (project VA148P23)

Plant viral nanoparticles (VNPs) have become an attractive platform for the development of novel nanotools in the last years because of their safety, inexpensive production, and straightforward functionalization. Turnip mosaic virus (TuMV) is one example of a plant-based VNP used as a nanobiotechnological platform either as virions or as virus-like particles (VLPs). Their functionalization mainly consists of coating their surface with the molecules of interest via chemical conjugation or genetic fusion. However, because of their limitations, these two methods sometimes result in non-viable constructs. In this paper, we applied the SpyTag/SpyCatcher technology as an alternative for the functionalization of TuMV VLPs with peptides and proteins. We chose as molecules of interest the green fluorescent protein (GFP) because of its good traceability, as well as the vasoactive intestinal peptide (VIP), given the previous unsuccessful attempts to functionalize TuMV VNPs by other methods. The successful conjugation of VLPs to GFP and VIP using SpyTag/SpyCatcher was confirmed through Western blot and electron microscopy. Moreover, the isopeptide bond between SpyTag and SpyCatcher occurred in vivo in co-agroinfiltrated Nicotiana benthamiana plants. These results demonstrated that SpyTag/SpyCatcher improves TuMV functionalization compared with previous approaches, thus implying the expansion of the application of the technology to elongated flexuous VNPs., Unión Europea, Ministerio de Universidades (España), Comunidad de Madrid, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA), Universidad Complutense de Madrid (UCM), Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Two subspecies of Francisella tularensis are recognized: F. tularensis subsp. tularensis (type A) and F. tularensis subsp. holartica (type B). Type A has been subdivided further into A1a, A1b, and A2, which differ geographically and clinically. The aim of this work was to determine whether or not differences among subspecies and clades translate into distinct ecological niches. We used 223 isolates from humans and wildlife representing all six genotypes (type A, B, A1, A2, A1a, or A1b). Ecological-niche models were built independently for each genotype, using the genetic algorithm for ruleset prediction. The resulting models were compared using a non-parametric multivariate analysis-of-variance method. A1 and A2 are ecologically distinct, supporting the previously observed geographic division, whereas ecological niches for types A and B overlapped notably but A1a and A1b displayed no appreciable differences in their ecological niches., U.S. Department of Defense, Centers for Disease Control and Prevention (U.S.A), Microsoft Research, Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

The aim of the study was to determine the presence of human papillomavirus (HPV) in patients with intractable plantar keratosis (IPK) by comparing the histopathological findings of biopsies. A prospective, observational, and concordance study was carried out. Three different specimens were taken from each IPK. A first punch was sent for histopathological examination, and a second punch and a superficial skin scraping were both sent for HPV polymerase chain reaction (PCR) and type determination. A total of 51 patients were included. From the histopathological examination, it was determined that 35 (68.6%) samples were diagnosed as warts and 16 (31.3%) as keratosis. However, the presence of HPV was confirmed by PCR in 49 (96.1%) and in 42 (82.4%) samples obtained by punch and superficial scraping, respectively. In the 49 PCR‐positive samples, the most common HPV types were HPV1, HPV2, HPV27, HPV57, and HPV65, accounting for 81.6% of the samples. In conclusion, this study demonstrates that HPV infection and IPK lesions are very closely related. Although we cannot confirm that HPV is the cause of the development of IPK, the high prevalence of HPV observed in these lesions calls for a change to the procedures for managing IPK., Depto. de Medicina, Fac. de Medicina, TRUE, pub

Background: This study aimed to analyze the relevance of different publications about microbiota on ocular diseases and their authors through a citation network analysis. In addition, the different research areas and the most cited publications have been identified. Methods: The bibliographic search was carried out through the Web of Science (WOS) database, using the following search term: “microbiota AND (vision OR eye OR visual)” for the period between 1995 and December 2022. The Citation Network Explorer and the CiteSpace software have been used to analyze the different publications. Results: 705 publications were found in the field of microbiota on ocular diseases, together with 1014 citation networks. The year 2022 was the year with more publications. The first authors with the highest number of publications in the microbiota on the ocular surface field were Chisari G, Chisari CG, and Li Y. This field is multidisciplinary, highlighting “microbiology” and “ophthalmology” as the main research areas. Publications were clustered into three main groups allowing the identification of the main research topics in this field. The principal was the composition and diversity of the bacterial community on the ocular surface of patients with several pathologies. Conclusion: It could be useful for researchers to choose suitable collaborators or projects to promote their research on the role of microbiota on ocular diseases, as well as to know the main research topics that are of major interest today., Depto. de Optometría y Visión, Depto. de Bioquímica y Biología Molecular, Fac. de Óptica y Optometría, Fac. de Farmacia, TRUE, pub

Since the use of pneumococcal conjugate vaccines PCV7 and PCV13 in children became widespread, invasive pneumococcal disease (IPD) has dramatically decreased. Nevertheless, there has been a rise in incidence of Streptococcus pneumoniae non-vaccine serotypes (NVT) colonising the human nasopharynx. Nasopharyngeal colonisation, an essential step in the development of S. pneumoniae-induced IPD, is associated with biofilm formation. Although the capsule is the main pneumococcal virulence factor, the formation of pneumococcal biofilms might, in fact, be limited by the presence of capsular polysaccharide (CPS). Methodology/Principal Findings We used clinical isolates of 16 emerging, non-PCV13 serotypes as well as isogenic transformants of the same serotypes. The biofilm formation capacity of isogenic transformants expressing CPSs from NVT was evaluated in vitro to ascertain whether this trait can be used to predict the emergence of NVT. Fourteen out of 16 NVT analysed were not good biofilm formers, presumably because of the presence of CPS. In contrast, serotypes 11A and 35B formed ≥45% of the biofilm produced by the non-encapsulated M11 strain. Conclusions/Significance This study suggest that emerging, NVT serotypes 11A and 35B deserve a close surveillance. Since the use of pneumococcal conjugate vaccines PCV7 and PCV13 in children became widespread, invasive pneumococcal disease (IPD) has dramatically decreased. Nevertheless, there has been a rise in incidence of Streptococcus pneumoniae non-vaccine serotypes (NVT) colonising the human nasopharynx. Nasopharyngeal colonisation, an essential step in the development of S. pneumoniae-induced IPD, is associated with biofilm formation. Although the capsule is the main pneumococcal virulence factor, the formation of pneumococcal biofilms might, in fact, be limited by the presence of capsular polysaccharide (CPS). Methodology/Principal Findings We used clinical isolates of 16 emerging, non-PCV13 serotypes as well as isogenic tra, Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Conditions altering the yeast cell wall lead to the activation of an adaptive transcriptional response mainly governed by the cell wall integrity (CWI) mitogen-activated protein kinase (MAPK) pathway. Two high-throughput screenings were developed using the yTHC collection of yeast conditional mutant strains to systematically identify essential genes related to cell wall integrity, and those required for the transcriptional program elicited by cell wall stress. Depleted expression of 52 essential genes resulted in hypersensitivity to the dye Calcofluor white, with chromatin organization, Golgi vesicle transport, rRNA processing, and protein glycosylation processes, as the most highly representative functional groups. Via a flow cytometry-based quantitative assay using a CWI reporter plasmid, 97 strains exhibiting reduced gene-reporter expression levels upon stress were uncovered, highlighting genes associated with RNA metabolism, transcription/translation, protein degradation, and chromatin organization. This screening also led to the discovery of 41 strains displaying a basal increase in CWI-associated gene expression, including mainly putative cell wall-related genes. Interestingly, several members of the RSC chromatin remodelling complex were uncovered in both screenings. Notably, Rsc9 was necessary to regulate the gene expression of CWI-related genes both under stress and non-stress conditions, suggesting distinct requirements of the RSC complex for remodelling particular genes., Ministerio de Ciencia e Innovación, Fondo Social Europeo, Comunidad de Madrid, European Structural and Investment Funds, Consejería de Educación e Investigación de la Comunidad de Madrid, Depto. de Microbiología y Parasitología, Fac. de Farmacia, TRUE, pub

Vancomycin, a glycopeptide antimicrobial agent, represents the last line of defence against a wide range of multi-resistant Gram-positive pathogens such as enterococci, staphylococci and streptococci. However, vancomycin-resistant enterococci and staphylococci, along with vancomycin-tolerant clinical isolates, are compromising the therapeutic efficacy of vancomycin. It is conceivable that tolerance may emerge during prolonged vancomycin use. It has not been until recently, however, that the molecular basis of this tolerance began to be understood. Superoxide anions might be involved in the bactericidal activity of vancomycin in enterococci, and recent evidence suggests that the stringent response is partly responsible for vancomycin tolerance in Enterococcus faecalis. The mechanism of vancomycin tolerance in Staphylococcus aureus and Streptococcus pneumoniae is sometimes associated with a reduction of autolysin activity. Vancomycin tolerance in S. aureus and S. pneumoniae also appears to be somehow related with the two-component regulatory systems linked to cell envelope stress, although the precise molecular regulatory pathways remain poorly defined., Ministerio de Ciencia e Innovación (MCIN), Dirección General de Investigación Científica y Técnica (España), Instituto de Salud Carlos III. CIBER de Enfermedades Respiratorias (CIBERES), Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Extracellular deoxyribonucleic acid (eDNA) is an essential component of bacterial biofilm matrices, and is required in their formation and maintenance. Extracellular DNA binds to exopolysaccharides or extracellular proteins, affording biofilms greater structural integrity. Recently, we reported evidence of intercellular eDNA−LytC complexes in pneumococcal biofilms. The LytC lysozyme is a member of the choline-binding family of proteins (CBPs) located on the pneumococcal surface. The present work shows that other CBPs, i.e. LytA, LytB, Pce, PspC and CbpF, which have a pI between 5 and 6, can bind DNA in vitro. This process requires the presence of divalent cations other than Mg2+. This DNA binding capacity of CBPs appears to be independent of their enzymatic activity and, at least in the case of LytA, does not require the choline-binding domain characteristic of CBPs. Positively charged, surface-exposed, 25 amino acid-long peptides derived from the catalytic domain of LytB, were also found capable of DNA binding through electrostatic interactions. Confocal laser scanning microcopy revealed the existence of cell-associated LytB−eDNA complexes in Streptococcus pneumoniae biofilms. These and other findings suggest that these surface-located proteins of S. pneumoniae could play roles of varying importance in the colonization and/or invasion of human host where different environmental conditions exist., Ministerio deEconomía y Competitividad, Instituto de Salud Carlos III. CIBER de Enfermedades Respiratorias (CIBERES), Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Rain-fed mountain granite rock basins are temporary habitats conditioned by a fluctuating environment and the unpredictability of precipitation or flooding rates. These small highland freshwater habitats remain largely unexplored at the microbial level. The aim of this work is to report the presence in these habitats of genetic sequences of microbial eukaryotes that are pathogens and potential pathogens of humans, wildlife, cattle, crops as well as of other microorganisms. We sequenced the hypervariable region v4 of the 18S rDNA gene from environmental DNA of sediments taken from 21 rock basins in a National Park in Spain. More than a fifth (21%) of the eukaryotic Operational Taxonomic Units (OTUs) found are ascribed to pathogenic (within 11 Phyla) and potential pathogenic (within 1 phylum, the Chytridiomycota) microorganisms. Some OTUs retrieved are of agro-economic and public health importance (e.g., Pythium spp., Lagenidium spp., Candida spp. and Vermamoeba vermiformis). In 86% of the basins, the most abundant OTUs were affiliated to Chytridiomycota, a broad fungal group including saprozoic and parasitic taxa. Two OTUs affiliated to chytrids were significantly correlated with high concentrations of heavy metals. The high proportion of chytrid-like microbial sequences found emphasises the role of these freshwater habitats for adding knowledge regarding the ecological trade-offs of the still rather unknown Chytridiomycota. Our results show that rain-fed rock basins may be model habitats for the study and surveillance of microbial community dynamics and genetics of (mainly opportunistic) microbial pathogens., Ministerio de Economía y Competitividad (MINECO- Spain), Comunidad de Madrid (Spain), Universidad Complutense de Madrid/ Banco de Santander, Health Research Institute of the Hospital Clínico San Carlos, IdISSC Madrid, Spain, Swiss National Foundation for Research, Swiss NSF, Depto. de Biodiversidad, Ecología y Evolución, Fac. de Ciencias Biológicas, TRUE, pub

Abstract Integrons are genetic platforms that acquire new genes encoded in integron cassettes (ICs), building arrays of adaptive functions. ICs generally encode promoterless genes, whose expression relies on the platform-associated Pc promoter, with the cassette array functioning as an operon-like structure regulated by the distance to the Pc. This is relevant in large sedentary chromosomal integrons (SCIs) carrying hundreds of ICs, like those in Vibrio species. We selected 29 gene-less cassettes in four Vibrio SCIs, and explored whether their function could be related to the transcription regulation of adjacent ICs. We show that most gene-less cassettes have promoter activity on the sense strand, enhancing the expression of downstream cassettes. Additionally, we identified the transcription start sites of gene-less ICs through 5′-RACE. Accordingly, we found that most of the superintegron in Vibrio cholerae is not silent. These promoter cassettes can trigger the expression of a silent dfrB9 cassette downstream, increasing trimethoprim resistance >512-fold in V. cholerae and Escherichia coli. Furthermore, one cassette with an antisense promoter can reduce trimethoprim resistance when cloned downstream. Our findings highlight the regulatory role of gene-less cassettes in the expression of adjacent cassettes, emphasizing their significance in SCIs and their clinical importance if captured by mobile integrons., Depto. de Sanidad Animal, Fac. de Veterinaria, TRUE, pub

Vultures have evolved adaptive mechanisms to prevent infections associated with their scavenging lifestyle. However, food-borne exposure to antimicrobial pharmaceuticals can promote opportunistic infections with adverse outcomes. Here, we used multivariate and network analyses to increase understanding of the behavior of the yeast communities causing oral mycosis outbreaks recently reported in wild nestling cinereous (Aegypius monachus), griffon (Gyps fulvus) and Egyptian (Neophron percnopterus) vultures (CV, GV and EV, respectively) exposed to antibiotics from livestock farming. Common and unique yeast signatures (of Candida, Debaromyces, Diutina, Meyerozyma, Naganishia, Pichia, Rhodotorula, Trichosporon and Yarrowia species) associated with oral mycoses were identified in the three vulture species. Hierarchical clustering analysis (HCA) and principal component analysis (PCA) highlighted that oral lesions from CV and GV shared similar yeast signatures (of major causative pathogens of opportunistic mycoses, such as Candida albicans, Candida parapsilosis and Candida tropicalis), while EV had a distinct yeast signature (of uncommon pathogenic species, such as Candida dubliniensis, Candida zeylanoides, Pichia fermentans and Rhodotorula spp.). Synergistic interactions between yeast species from distinct fungal phyla were found in lesions from CV and GV, but not in EV. These formed co-occurrence subnetworks with partially or fully connected topology. This study reveals that the composition, assembly and co-occurrence patterns of the yeast communities causing oral mycoses differ between vulture species with distinct feeding habits and scavenging lifestyles. Yeast species widely pathogenic to humans and animals, and yeast co-occurrence relationships, are distinctive hallmarks of oral mycoses in CV and GV. These vulture species are more exposed to antibiotics from intensively medicated livestock carcasses provided in supplementary feeding stations and show higher incidence, Ministerio de Economía y Competitividad (MINECO), Fac. de Óptica y Optometría, TRUE, pub

A strain of the protozoan ciliate Tetrahymena thermophila adapted to increasing Pb(II) concentrations over two years has shown that one of the resistance mechanisms to this extreme metal stress is the lead biomineralization to chloropyromorphite, one of the most stable minerals in the earth's crust. Several techniques such as microanalysis coupled to transmission and scanning electron microscopy (X-Ray Energy Disperse Spectroscopy), fluorescence microscopy and X-ray power diffraction analysis have revealed the presence of chloropyromorphite as crystalline aggregates of nano-globular structure, together with the presence of other secondary lead minerals. This is the first time that the existence of this type of biomineralization in a ciliate protozoan is described. The Pb(II) bioremediation capacity of this strain has shown that it can remove >90 % of the toxic soluble lead from the medium. A quantitative proteomic analysis of this strain has revealed the main molecular-physiological elements involved in adaptation to Pb(II) stress: increased activity of proteolytic systems against lead proteotoxicity, occurrence of metallothioneins to immobilize Pb(II) ions, antioxidant enzymes to mitigate oxidative stress, and an intense vesicular trafficking presumably involved in the formation of vacuoles where pyromorphite accumulates and is subsequently excreted, together with an enhanced energy metabolism. As a conclusion, all these results have been compiled into an integrated model that could explain the eukaryotic cellular response to extreme lead stress., Ministerio de Ciencia y Competitividad, Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Fundación Española para la Ciencia y la Tecnología, Universidad Complutense de Madrid, Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

The use of metaproteomics for studying the human gut microbiota can shed light on the taxonomic profile and the functional role of the microbial community. Nevertheless, methods for extracting proteins from stool samples continue to evolve, in the pursuit of optimal protocols for moistening and dispersing the stool sample and for disrupting microbial cells, which are two critical steps for ensuring good protein recovery. Here, we evaluated different stool sample processing (SSP) and microbial cell disruption methods (CDMs). The combination of a longer disintegration period of the stool sample in a tube rotator with sonication increased the overall number of identified peptides and proteins. Proteobacteria, Bacteroidetes, Planctomycetes, and Euryarchaeota identification was favored by mechanical cell disruption with glass beads. In contrast, the relative abundance of Firmicutes, Actinobacteria, and Fusobacteria was improved when sonication was performed before bead beating. Tenericutes and Apicomplexa identification was enhanced by moistening the stool samples during processing and by disrupting cells with medium-sized glass beads combined with or without sonication. Human protein identifications were affected by sonication. To test the reproducibility of these gut metaproteomic analyses, we examined samples from six healthy individuals using a protocol that had shown a good taxonomic diversity and identification of proteins from Proteobacteria and humans. We also detected proteins involved in microbial functions relevant to the host and related mostly to specific taxa, such as B12 biosynthesis and short chain fatty acid (SCFA) production carried out mainly by members in the Prevotella genus and the Firmicutes phylum, respectively. The taxonomic and functional profiles obtained with the different protocols described in this work provides the researcher with valuable information when choosing the most adequate protocol for the study of certain pathologies under suspic, Ministerio de Ciencia, Innovación y Universidades (España), Comunidad de Madrid, Instituto de Salud Carlos III, Fondo Europeo de Desarrollo Regional, Depto. de Microbiología y Parasitología, Fac. de Farmacia, TRUE, pub

Macrophages are involved in the primary human response to Candida albicans. After pathogen recognition, signaling pathways are activated, leading to the production of cytokines, chemokines, and antimicrobial peptides. ATP binding proteins are crucial for this regulation. Here, a quantitative proteomic and phosphoproteomic approach was carried out for the study of human macrophage ATP-binding proteins after interaction with C. albicans. From a total of 547 nonredundant quantified proteins, 137 were ATP binding proteins and 59 were detected as differentially abundant. From the differentially abundant ATP-binding proteins, 6 were kinases (MAP2K2, SYK, STK3, MAP3K2, NDKA, and SRPK1), most of them involved in signaling pathways. Furthermore, 85 phosphopeptides were quantified. Macrophage proteomic alterations including an increase of protein synthesis with a consistent decrease in proteolysis were observed. Besides, macrophages showed changes in proteins of endosomal trafficking together with mitochondrial proteins, including some involved in the response to oxidative stress. Regarding cell death mechanisms, an increase of antiapoptotic over pro-apoptotic signals is suggested. Furthermore, a high pro-inflammatory response was detected, together with no upregulation of key mi-RNAs involved in the negative feedback of this response. These findings illustrate a strategy to deepen the knowledge of the complex interactions between the host and the clinically important pathogen C. albicans., Unión Europea, Ministerio de Economía, Comercio y Empresa (España), Comunidad de Madrid, Instituto de Salud Carlos III, Depto. de Microbiología y Parasitología, Fac. de Óptica y Optometría, Fac. de Farmacia, TRUE, pub

Lactococcus garvieae is a main ichthyopathogen in rainbow trout (Oncorhynchus mykiss, Walbaum) farming, although bacteriocinogenic L. garvieae with antimicrobial activity against virulent strains of this species have also been identified. Some of the bacteriocins characterized, such as garvicin A (GarA) and garvicin Q (GarQ), may show potential for the control of the virulent L. garvieae in food, feed and other biotechnological applications. In this study, we report on the design of Lactococcus lactis strains that produce the bacteriocins GarA and/or GarQ, either alone or together with nisin A (NisA) or nisin Z (NisZ). Synthetic genes encoding the signal peptide of the lactococcal protein Usp45 (SPusp45), fused to mature GarA (lgnA) and/or mature GarQ (garQ) and their associated immunity genes (lgnI and garI, respectively), were cloned into the protein expression vectors pMG36c, which contains the P32 constitutive promoter, and pNZ8048c, which contains the inducible PnisA promoter. The transformation of recombinant vectors into lactococcal cells allowed for the production of GarA and/or GarQ by L. lactis subsp. cremoris NZ9000 and their co-production with NisA by Lactococcus lactis subsp. lactis DPC5598 and L. lactis subsp. lactis BB24. The strains L. lactis subsp. cremoris WA2-67 (pJFQI), a producer of GarQ and NisZ, and L. lactis subsp. cremoris WA2-67 (pJFQIAI), a producer of GarA, GarQ and NisZ, demonstrated the highest antimicrobial activity (5.1- to 10.7-fold and 17.3- to 68.2-fold, respectively) against virulent L. garvieae strains., Ministerio de Ciencia, Innovación y Universidades, Universidad Complutense de Madrid (UCM), Santander/UCM, Sección Dptal. de Nutrición y Ciencia de los Alimentos (Veterinaria), Fac. de Veterinaria, TRUE, pub

Mitochondria play a wide diversity of roles in cell physiology and have a key functional implication in cell bioenergetics and biology of free radicals. As the main cellular source of oxygen radicals, mitochondria have been postulated as the mediators of the cellular decline associated with the biological aging. Recent evidences have shown that mitochondrial free radical production is a highly regulated mechanism contributing to the biological determination of longevity which is species-specific. This mitochondrial free radical generation rate induces a diversity of adaptive responses and derived molecular damage to cell components, highlighting mitochondrial DNA damage, with biological consequences that influence the rate of aging of a given animal species. In this review, we explore the idea that mitochondria play a fundamental role in the determination of animal longevity. Once the basic mechanisms are discerned, molecular approaches to counter aging may be designed and developed to prevent or reverse functional decline, and to modify longevity., Ministerio de Ciencia e Innovación (MICIN), Comunidad de Madrid, Generalitat de Catalunya, Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

The effects of decoquinate (DCQ) and three O-quinoline-carbamate-derivatives were investigated using human foreskin fibroblasts (HFF) infected with Neospora caninum tachyzoites. These compounds exhibited half-maximal proliferation inhibition (IC50s) from 1.7 (RMB060) to 60 nM (RMB055). Conversely, when applied at 5 (DCQ, RMB054) or 10µM (RMB055, RMB060), HFF viability was not affected. Treatments of infected cell cultures at 0.5µM altered the ultrastructure of the parasite mitochondrion and cytoplasm within 24 h, most pronounced for RMB060, and DCQ, RMB054 and RMB060 did not impair the viability of splenocytes from naïve mice. Long-term treatments of N. caninum-infected HFF monolayers with 0.5µM of each compound showed that only exposure to RMB060 over a period of six consecutive days had a parasiticidal effect, while the other compounds were not able to kill all tachyzoites in vitro. Thus, DCQ and RMB060 were comparatively assessed in the pregnant neosporosis mouse model. The oral application of these compounds suspended in corn oil at 10 mg/kg/day for 5 d resulted in a decreased fertility rate and litter size in the DCQ group, whereas reproductive parameters were not altered by RMB060 treatment. However, both compounds failed to protect mice from cerebral infection and did not prevent vertical transmission/pup mortality. Thus, despite the promising in vitro efficacy and safety characteristics of DCQ and DCQ-derivatives, proof of concept for activity against neosporosis could not be demonstrated in the murine model., Swiss National Science Foundation, MRC/National Treasury/Economic Competitiveness and Support Package, SA NRF, Depto. de Sanidad Animal, Fac. de Veterinaria, TRUE, pub

Lactococcus garvieae is a main ichthyopathogen in rainbow trout (Oncorhynchus mykiss, Walbaum) farming, although bacteriocinogenic L. garvieae with antimicrobial activity against virulent strains of this species have also been identified. Some of the bacteriocins characterized, such as garvicin A (GarA) and garvicin Q (GarQ), may show potential for the control of the virulent L. garvieae in food, feed and other biotechnological applications. In this study, we report on the design of Lactococcus lactis strains that produce the bacteriocins GarA and/or GarQ, either alone or together with nisin A (NisA) or nisin Z (NisZ). Synthetic genes encoding the signal peptide of the lactococcal protein Usp45 (SPusp45), fused to mature GarA (lgnA) and/or mature GarQ (garQ) and their associated immunity genes (lgnI and garI, respectively), were cloned into the protein expression vectors pMG36c, which contains the P32 constitutive promoter, and pNZ8048c, which contains the inducible PnisA promoter. The transformation of recombinant vectors into lactococcal cells allowed for the production of GarA and/or GarQ by L. lactis subsp. cremoris NZ9000 and their co-production with NisA by Lactococcus lactis subsp. lactis DPC5598 and L. lactis subsp. lactis BB24. The strains L. lactis subsp. cremoris WA2-67 (pJFQI), a producer of GarQ and NisZ, and L. lactis subsp. cremoris WA2-67 (pJFQIAI), a producer of GarA, GarQ and NisZ, demonstrated the highest antimicrobial activity (5.1- to 10.7-fold and 17.3- to 68.2-fold, respectively) against virulent L. garvieae strains., Ministerio de Ciencia, Innovación y Universidades (MCIU), Universidad Complutense de Madrid and Banco de Santander, Universidad Complutense de Madrid, Sección Dptal. de Nutrición y Ciencia de los Alimentos (Veterinaria), Fac. de Veterinaria, TRUE, pub

Algunos agentes biológicos, pueden ser empleados como armas ofensivas (bacterias, rickettsias, virus, hongos filamentosos, protistas o toxinas), obtenidos a partir de cepas naturales o por modificaciones genéticas o selección en el laboratorio de alguno de ellos. Todos ellos presentan una serie de características que permiten que puedan ser considerados como potenciales armas y que resultan determinantes para su elección. Factores intrínsecos (propios del agente) como su nivel de patogenicidad, grado de virulencia, tiempo de incubación o forma de transmisión y extrínsecos (factores medioambientales) viento, humedad, temperatura, luminosidad, vías de entrada. Otra serie de factores a tener en cuenta para la elección de un agente biológico están relacionados con su facilidad y bajo coste para ser producidos (temperatura de crecimiento, requerimientos nutricionales, escalado de producción, estabilización del agente, etc.). También hay que evaluar las ventajas de su uso, los inconvenientes de su utilización, determinar cuál va a ser el lugar idóneo para su empleo y qué propiedades tiene desde el punto de vista ofensivo. Es muy importante conocer los tipos de patologías que pueden producir. Entre los agentes que potencialmente podrían ser empleados en un ataque biológico, los más probables evaluados por un análisis exhaustivo de los factores anteriores se encontrarían Bacillius anthracis (agente productor del anthrax), Yersinia pestis (productor de la peste), el virus de la Variola (productor de la viruela) y el virus productor de la viruela del mono y la intoxicación por ricina. Este estudio puede resultar de utilidad de cara a una evaluación razonada para determinar el potencial de distintos agentebiológicos, así como estimar sus riesgos de empleo., Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

El avance tecnológico ocurrido en los últimos años ha permitido un avance significativo en el campo de las ciencias de la vida. En un mundo globalizado que cuenta con tecnología interconectada y con aplicaciones múltiples, el desarrollo de una determinada aplicación para un sector en concreto rápidamente se expande a otros sectores y ámbitos, incrementando de manera exponencial la versatilidad de estos avances. Los diferentes avances en informática ocurridos desde mediados del siglo XX han permitido el diseño de nuevos procesos, aplicaciones y protocolos de análisis y manejo de datos biológicos, en especial en estas primeras décadas del siglo XXI. Los avances en el conocimiento sobre la estructura, función e ingeniería de los ácidos nucleicos, en especial ADN, han sido el motor que han permitido, y permiten, la rápida evolución de las tecnologías de ingeniería genética que permiten la edición de microorganismos haciendo que estos adquieran nuevas funciones, o incluso, crearlos desde cero aprovechando información genética depositada en bases de datos y generando microorganismos de manera sintética. La democratización de la ciencia, los protocolos y el conocimiento que la componen, ha hecho que numerosos actores, ya sean científicos de carrera o aficionados, puedan contribuir de manera sustancial al progreso. El riesgo de estas nuevas tecnologías no reside en ellas mismas, si no en el uso que determinados actores maliciosos puedan hacer de ellas. Cualquier avance tecnológico diseñado para hacer el bien, en cualquier momento, se puede emplear para causar un daño, intencionado o no., Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

El uso malintencionado de los agentes biológicos por parte del ser humano se conoce desde la antigüedad. Los primeros textos documentados sobre estos hechos se remontan al siglo VI a.C., empleando de forma empírica, plantas, toxinas extraídas de las mismas o incluso empleando cadáveres contaminados con microorganismos altamente patógenos. Tanto en la Edad Media como en la llegada a América por los europeos se utilizaron estos agentes, conociendo claramente los efectos que podrían tener sobre la población. El descubrimiento de los microorganismos en el siglo XVII por A. van Leewenhoek y posteriormente, los grandes descubrimientos que hacen la microbiología y la bioquímica en el siglo XIX abre nuevas posibilidades al empleo de los microorganismos y las toxinas con fines bélicos. El comienzo de la Primera Guerra Mundial supuso un punto de inflexión en la investigación por parte de los países de la Triple Alianza en el desarrollo y uso de microorganismos con fines bélicos. El periodo entre guerras y la Segunda Guerra Mundial hacen que investigadores tanto japoneses como norteamericanos, ingleses y rusos desarrollen a nivel nacional sus programas de desarrollo de armas biológicas. El Protocolo de Ginebra de 1925 puso las bases sobre la no proliferación del armamento biológico por los países beligerantes, que más tarde se vería refrendado por la Convención sobre Armas Biológicas (CAB) de 1972, que entró en vigor en 1975, refrendado actualmente por más de 160 países del mundo. El gran reto actual radica en la lucha contra el bioterrorismo por parte de las instituciones internacionales., Depto. de Genética, Fisiología y Microbiología, Fac. de Ciencias Biológicas, TRUE, pub

Producción Científica, Two strains of Saccharomyces cerevisiae (Sc01 and Sc02) and one strain of Wickerhamomyces anomalus (Wa) were isolated from organic Verdejo spontaneous fermentations and used for the development of experimental winemaking. Sc01 and Sc02 represented 52.7% of the population of the Saccharomyces strains isolated throughout the fermentation process. W. anomalus appeared as the predominant species among the non-Saccharomyces yeasts. Wa turned out to be the strain of this species with the shortest lag phase and positive enzymatic activities, and it was selected for white wine production. Fermentations with unique inoculation of S. cerevisiae strains were compared with sequential inoculation with W. anomalus. The results showed that the sequential inoculations did not affect the fermentation kinetics or physicochemical characteristics of the wines compared with the unique inoculations. However, this study identified a significant impact on the aromatic profiles of the produced wines due to the sequential inoculations. This modification resulted in a similar new aromatic profile in both sequential inoculations, demonstrating common characteristics related to the contribution of W. anomalus. In general, the sequential fermentations were mainly characterized by lower levels of acetate esters and an increase in ethyl acetate levels, whereas lower levels of ethyl octanoate and ethyl dec-9-enoate were detected. Propan-1-ol and butan-1-ol showed an increase in the sequential fermentations, while 4-methylpentan-1-ol and 2-phenylethanol were found in lower concentrations. These results highlight the great influence that the presence of specific strains of native non-Saccharomyces yeasts exerts on the characteristics of elaborate wines., Universidad de Valladolid (UVa) y Bodegas Belondrade S. L. - (grant 061/100671)