Person: Domínguez Bernal, Gustavo Ramón
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First Name
Gustavo Ramón
Last Name
Domínguez Bernal
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Veterinaria
Department
Sanidad Animal
Area
Sanidad Animal
Identifiers
11 results
Search Results
Now showing 1 - 10 of 11
Item Differences in virulence gene expression between atypical enteropathogenic Escherichia coli strains isolated from diarrheic and healthy ruminants(The Canadian Journal of Veterinary Research, 2012) Carrión Herrero, Francisco Javier; Horcajo Iglesias, María Del Pilar; Domínguez Bernal, Gustavo Ramón; Fuente López, Ricardo De La; Ruiz Santa Quiteria Serrano De La Cruz, José Antonio; Orden Gutiérrez, José AntonioDifferences in the pathogenicity of atypical enteropathogenic Escherichia coli (EPEC) strains may be due, at least partially, to different expression patterns of some virulence genes. To investigate this hypothesis, the virulence gene expression patterns of 6 atypical EPEC strains isolated from healthy and diarrheic ruminants were compared using quantitative real-time reverse transcription polymerase chain reaction after growing the bacteria in culture medium alone or after binding it to HeLa epithelial cells. Some virulence genes in strains from diarrheic animals were upregulated relative to their expression in strains from healthy animals. When bacteria were cultured in the presence of HeLa cells, the ehxA and efa1/lifA genes, previously associated with the production of diarrhea, were expressed at higher levels in strains from diarrheic animals than in strains from healthy animals. Thus, the expression levels of some virulence genes may help determine which atypical EPEC strains cause diarrhea in ruminants.Item Mechanisms of resistance and susceptibility to experimental visceral leishmaniosis: BALB/c mouse versus syrian hamster model(Veterinary Research, 2011) Nieto, Ana; Madrid-Elena, Nadia; Carrión Herrero, Francisco Javier; Domínguez Bernal, Gustavo Ramón; Orden Gutiérrez, José Antonio; Fuente López, Ricardo De LaSeveral animal models have been established to study visceral leishmaniosis (VL), a worldwide vector-borne disease affecting humans and domestic animals that constitutes a serious public health problem. BALB/c mice and Syrian hamsters are the most widely used experimental models. In this paper, we summarize the advantages and disadvantages of these two experimental models and discuss the results obtained using these models in different studies of VL. Studies using the BALB/c mouse model have underscored differences between the liver and spleen in the course of VL, indicating that pathological evaluation of the visceral organs is essential for understanding the immune mechanisms induced by Leishmania infantum infection. The main goal of this review is to collate the relevant literature on Leishmania pathogenesis into a sequence of events, providing a schematic view of the main components of adaptive and innate immunity in the liver and spleen after experimental infection with L. infantum or L. donovani. This review also presents several viewpoints and reflections about some controversial aspects of Leishmania research, including the choice of experimental model, route of administration, inoculum size and the relevance of pathology (intimately linked to parasite persistence): a thorough understanding of which is essential for future VL research and the successful development of efficient control strategies for Leishmania spp.Item Characterisation of the ex vivo virulence of Leishmania infantum isolates from Phlebotomus perniciosus from an outbreak of human leishmaniosis in Madrid, Spain(Parasites and Vectors, 2014) Jiménez, Maribel; Molina, Ricardo; Ordóñez-Gutiérrez, Lara; Carrión Herrero, Francisco Javier; Domínguez Bernal, Gustavo Ramón; Martínez Rodrigo, Abel; Mas Zubiri, Alicia; Cutuli Simón, María TeresaBackground Since mid 2009, an outbreak of human leishmaniosis in Madrid, Spain, has involved more than 560 clinical cases. Many of the cases occurred in people who live in areas around a newly constructed green park (BosqueSur). This periurban park provides a suitable habitat for sand flies (the vectors of Leishmania infantum). Indeed, studies of blood meals from sand flies captured in the area showed a strong association between the insect vector, hares or rabbits, and humans in the area. Interestingly, up to 70% of cases have been found in immunocompetent patients (aged between 46-60 years). This study was designed to evaluate the ex vivo virulence of the L. infantum isolates from Phlebotomus perniciosus captured in this area of Madrid. Methods Murine macrophages and dendritic cells were infected ex vivo with L. infantum strain BCN150, isolate BOS1FL1, or isolate POL2FL7. At different times after infection, the infection indices, cytokine production (IL-12p40 and IL-10), NO release and arginase activities were evaluated. Results Using an ex vivo model of infection in murine bone marrow-derived cells, we found that infection with isolates BOS1FL1 and POL2FL7 undermined host immune defence mechanisms in multiple ways. The main factors identified were changes in both the balance of iNOS versus arginase activities and the equilibrium between the production of IL-12 and IL-10. Infection with isolates BOS1FL1 and POL2FL7 also resulted in higher infection rates compared to the BCN150 strain. Infection index values at 24 h were as follows: BCN150-infected cells, 110 for infected MØ and 115 for infected DC; BOS1FL1-infected cells, 300 for infected MØ and 247 for infected DC; and POL2FL7-infected cells, 275 for infected MØ and 292 for infected DC. Conclusions Our data indicate that L. infantum isolates captured from this endemic area exhibited high virulence in terms of infection index, cytokine production and enzymatic activities involved in the pathogenesis of visceral leishmaniosis. Altogether, these data provide a starting point for the study of the virulence behaviour of parasites (BOS1FL1 and POL2FL7) isolated from P. perniciosus during the outbreak of human leishmaniosis in Madrid, Spain, and their involvement in infecting immunocompetent hosts.Item Short communication: Isolation frequency of bacteria causing lymphadenitis and abscesses in small ruminants in central Spain(Small Ruminant Research, 2017) Heras, Moisés de las; Torrijos, Carlos; Carrión Herrero, Francisco Javier; Fuente López, Ricardo De La; Díez De Tejada Martín, María Paloma; Pérez Sancho, Marta; Orden Gutiérrez, José Antonio; Domínguez Bernal, Gustavo RamónInfectious lymphadenitis in small ruminants caused by Corynebacterium pseudotuberculosis and Staphylococcus aureus subsp. anaerobius are widely distributed throughout the world, and result in significant economic losses. Trueperella pyogenes has also been associated with lymphadenitis in sheep and goats. In order to determinate the isolation frequency of the different agents associated with lymphadenitis and abscesses, 171 pus samples (135 from sheep and 36 from goats) from 46 flocks were investigated. Isolated bacteria were identified by MALDI-TOF method. S. aureus subsp. anaerobius was the most frequently detected agent. It was identified in 76 animals (59 sheep and 17 goats) from 24 of the surveyed flocks. Of these infected animal, 25 (32,9%) were over one year old, confirming that abscess disease may occur in a significant percentage of adult animals. C. pseudotuberculosis was identified in 45 of the sampled animals (36 sheep and 9 goats) from 24 flocks. Only 5 of animals suffering caseous lymphadenitis were under one year old. T. pyogenes was isolated from 17 animals (13 sheep and 4 goats) in 11 flocks. Seven of these samples were taken from subcutaneous abscesses located in not lymph nodes regions. A notable finding of this work is the isolation of Actinomyces hyovaginalis from 5 of the samples analyzed all of them taken from subcutaneous abscesses located in superficial lymph nodes regions. Thus, T. pyogenes and A. hyovaginalis should be included in the differential diagnosis of lymphadenitis in small ruminants. In 19 of the 46 surveyed flocks at least two of the four agents were detected, which underlines the need to analyze samples from several animals to reach an accurate diagnosis in flocks affected by lymphadenitis.Item Engineering of a live Salmonella enterica serovar Choleraesuis negative-marker strain that allows serological differentiation between immunised and infected animals(Veterinary Journal, 2016) Herrero-Gil, Aldara; Carrión Herrero, Francisco Javier; Orden Gutiérrez, José Antonio; Fuente López, Ricardo De La; Domínguez Bernal, Gustavo RamónThe usefulness of Salmonella vaccine vehicles is limited by the fact that control programmes relying on Salmonella bacteriology and serology cannot differentiate infected animals from vaccinated ones, an ability referred to as DIVA (differentiating infected from vaccinated animals). As a first step towards Salmonella-based DIVA vaccines, the ompA gene was deleted in live attenuated ΔphoP and ΔrpoS vaccine strains. The ompA gene is present in all Salmonella enterica serovars and it encodes an abundant, highly immunogenic outer membrane protein. The double mutant ΔphoP ΔompA and ΔrpoS ΔompA strains showed similar virulence attenuation, safety and immunogenicity in a mouse model of infection as the parental ΔphoP and ΔrpoS strains. Sera from mice inoculated with the double mutant strains failed to recognise OmpA in Western blots of outer membrane extracts, whereas the protein was recognised by sera from mice inoculated with wild-type Salmonella or a mixture of double mutant and parental strains. These data suggest that OmpA can be a suitable negative marker for DIVA vaccines.Item Alternative strategy for visceral leishmaniosis control: HisAK70-Salmonella Choleraesuis-pulsed dendritic cells(Comparative Immunology, Microbiology and Infectious Diseases, 2017) Carrión Herrero, Francisco Javier; Domínguez Bernal, Gustavo Ramón; Martínez Rodrigo, Abel; Mas Zubiri, Alicia; Blanco Gutiérrez, María Del Mar; Orden Gutiérrez, José Antonio; Fuente López, Ricardo De LaHere, we describe a novel approach that exploits an attenuated mutant of Salmonella enterica serovar Choleraesuis as carrier to deliver a plasmid encoding protein HisAK70. Subsequently, dendritic cells (DCs) were pulsed with this vaccine vector. The aim of this study was to evaluate the effectiveness of the prepared HisAK70-S. Choleraesuis-pulsed DCs (HisAK70-SAL DCs) against visceral leishmaniosis (VL). In our ex vivo model of infection, the prepared formulations could decrease parasite growth by up to 80% by augmenting the production of IL-12p40 and by reducing arginase activity (ARG). Also, BALB/c mice when immunised with this formulation showed significant reduction in parasite burden in both spleen (20% of reduction) and liver (75% of reduction). The balance of the immune ratios IFN-γ/IL-10, TNF-α/IL-10, and IgG2a/IgG1 reflected the acquisition of an improved resistant phenotype in HisAK70-SAL DCs vaccinated mice compared to control mice. Our results suggest that HisAK70-SAL DCs could be a promising alternative approach for vaccine delivery that has the potential to fight Leishmania infantum (L. infantum) infection.Item UCP2 Deficiency Helps to Restrict the Pathogenesis of Experimental Cutaneous and Visceral Leishmaniosis in Mice(PLoS Neglected Tropical Diseases, 2013) Carrión Herrero, Francisco Javier; Abengozar, M. Angeles; Fernández-Reyes, María; Sánchez-Martín, Carlos; Rial, Eduardo; Domínguez Bernal, Gustavo Ramón; González-Barroso, M. Mar; Alain DebrabantBackground: Uncoupling protein 2 (UCP2) is a mitochondrial transporter that has been shown to lower the production of reactive oxygen species (ROS). Intracellular pathogens such as Leishmania upregulate UCP2 and thereby suppress ROS production in infected host tissues, allowing the multiplication of parasites within murine phagocytes. This makes host UCP2 and ROS production potential targets in the development of antileishmanial therapies. Here we explore how UCP2 affects the outcome of cutaneous leishmaniosis (CL) and visceral leishmaniosis (VL) in wild-type (WT) C57BL/6 mice and in C57BL/6 mice lacking the UCP2 gene (UCP2KO). Methodology and Findings: To investigate the effects of host UCP2 deficiency on Leishmania infection, we evaluated parasite loads and cytokine production in target organs. Parasite loads were significantly lower in infected UCP2KO mice than in infected WT mice. We also found that UCP2KO mice produced significantly more interferon-γ (IFN-γ), IL-17 and IL-13 than WT mice (P<0.05), suggesting that UCP2KO mice are resistant to Leishmania infection. Conclusions: In this way, UCP2KO mice were better able than their WT counterparts to overcome L. major and L. infantum infections. These findings suggest that upregulating host ROS levels, perhaps by inhibiting UPC2, may be an effective approach to preventing leishmaniosis.Item Enhancing Control of Leishmania infantum Infection: A Multi-Epitope Nanovaccine for Durable T-Cell Immunity(Animals, 2024) Hurtado Morillas, Clara; Martínez Rodrigo, Abel; Orden Gutiérrez, José Antonio; De Urbina Fuentes, Laura; Mas Zubiri, Alicia; Domínguez Bernal, Gustavo RamónCanine leishmaniosis (CanL) is a growing health problem for which vaccination is a crucial tool for the control of disease. The successful development of an effective vaccine against this disease relies on eliciting a robust and enduring T-cell immune response involving the activation of CD4+ Th1 and CD8+ T-cells. This study aimed to evaluate the immunogenicity and prophylactic efficacy of a novel nanovaccine comprising a multi-epitope peptide, known as HisDTC, encapsulated in PLGA nanoparticles against Leishmania infantum infection in the murine model. The encapsulation strategy was designed to enhance antigen loading and sustain release, ensuring prolonged exposure to the immune system. Our results showed that mice immunized with PLGA-encapsulated HisDTC exhibited a significant reduction in the parasite load in the liver and spleen over both short and long-term duration. This reduction was associated with a cellular immune profile marked by elevated levels of pro-inflammatory cytokines, such as IFN-γ, and the generation of memory T cells. In conclusion, the current study establishes that PLGA-encapsulated HisDTC can promote effective and long-lasting T-cell responses against L. infantum in the murine model. These findings underscore the potential utility of multi-epitope vaccines, in conjunction with appropriate delivery systems, as an alternative strategy for CanL control.Item Subtilase Cytotoxin-Coding Genes in Verotoxin-Producing Escherichia coli Strains from Sheep and Goats Differ from Those from Cattle(Applied and Environmental Microbiology, 2011) Carrión Herrero, Francisco Javier; Orden Gutiérrez, José Antonio; Horcajo Iglesias, María Del Pilar; Fuente López, Ricardo De La; Ruiz Santa Quiteria Serrano De La Cruz, José Antonio; Domínguez Bernal, Gustavo RamónSubtilase cytotoxin (SubAB) from verotoxin (VT)-producing Escherichia coli (VTEC) strains was first described in the 98NK2 strain and has been associated with human disease. However, SubAB has recently been found in two VT-negative E. coli strains (ED 591 and ED 32). SubAB is encoded by two closely linked, cotranscribed genes (subA and subB). In this study, we investigated the presence of subAB genes in 52 VTEC strains isolated from cattle and 209 strains from small ruminants, using PCR. Most (91.9%) VTEC strains from sheep and goats and 25% of the strains from healthy cattle possessed subAB genes. The presence of subAB in a high percentage of the VTEC strains from small ruminants might increase the pathogenicity of these strains for human beings. Some differences in the results of PCRs and in the association with some virulence genes suggested the existence of different variants of subAB. We therefore sequenced the subA gene in 12 strains and showed that the subA gene in most of the subAB-positive VTEC strains from cattle was almost identical (about 99%) to that in the 98NK2 strain, while the subA gene in most of the subAB-positive VTEC strains from small ruminants was almost identical to that in the ED 591 strain. We propose the terms subAB1 to describe the SubAB-coding genes resembling that in the 98NK2 strain and subAB2 to describe those resembling that in the ED 591 strain.Item Subtilase cytotoxin-encoding subAB2 variants in verotoxin-producing Escherichia coli strains isolated from goats and sheep(Research in Veterinary Science, 2016) Orden Gutiérrez, José Antonio; Domínguez Bernal, Gustavo Ramón; Fuente López, Ricardo De La; Carrión Herrero, Francisco JavierSubtilase cytotoxin (SubAB) is a cytotoxin which might contribute to the virulence of verotoxin-producing Escherichia coli (VTEC) strains in humans. Three variants of SubAB encoding genes have been described (subAB1, subAB2-1, and subAB2-2) and it has been suggested that the strains positive for two variants of subAB may be more pathogenic for humans. In this study, 188 subAB2-positive VTEC strains isolated from goats and sheep were investigated for the presence of the subAB2-1 and subAB2-2 variants by PCR. Eighty-one of the 132 (61.4%) caprine strains and 36 of the 56 (64.3%) ovine strains possessed the subAB2-1 variant and all ovine and caprine strains, except one, were positive for the subAB2-2 variant. The results of this study show for first time that the subAB2-1 and subAB2-2 variants are found in caprine subAB2-positive VTEC strains and confirm that both subAB2 variants are detected in ovine subAB2-positive VTEC strains. Since no significant difference in the presence of both subAB2 variants was found among strains belonging to serotypes associated with severe illness in humans and strains not belonging to these serotypes, the occurrence of two subAB2 variants seems not to be associated with a higher risk of severe disease in humans.