Seroepidemiological study of Q fever in domestic ruminants in semi-extensive grazing systems

<p>Abstract</p> <p>Background</p> <p>Q fever, a worldwide zoonotic disease caused by <it>Coxiella burnetii</it>, is endemic in northern Spain where it has been reported as responsible for large series of human pneumonia cases and domestic ruminants' reproductive disorders. To investigate pathogen exposure among domestic ruminants in semi-extensive grazing systems in northern Spain, a serosurvey was carried out in 1,379 sheep (42 flocks), 626 beef cattle (46 herds) and 115 goats (11 herds). Serum antibodies were analysed by ELISA and positive samples were retested by Complement Fixation test (CFT) to detect recent infections.</p> <p>Results</p> <p>ELISA anti-<it>C. burnetii </it>antibody prevalence was slightly higher in sheep (11.8 ± 2.0%) than in goats (8.7 ± 5.9%) and beef cattle (6.7 ± 2.0%). Herd prevalence was 74% for ovine, 45% for goat and 43% for bovine. Twenty-one percent of sheep flocks, 27% of goat and 14% of cattle herds had a <it>C. burnetii </it>seroprevalence ≥ 20%. Only 15 out of 214 ELISA-positive animals reacted positive by CFT. Age-associated seroprevalence differed between ruminant species with a general increasing pattern with age. No evidence of correlation between abortion history and seroprevalence rates was observed despite the known abortifacient nature of <it>C. burnetii </it>in domestic ruminants.</p> <p>Conclusions</p> <p>Results reported herein showed that sheep had the highest contact rate with <it>C. burnetii </it>in the region but also that cattle and goats should not be neglected as part of the domestic cycle of <it>C. burnetii</it>. This work reports basic epidemiologic patterns of <it>C. burnetii </it>in semi-extensive grazed domestic ruminants which, together with the relevant role of <it>C. burnetii </it>as a zoonotic and abortifacient agent, makes these results to concern both Public and Animal Health Authorities.</p

Multiplex SERS Detection of Metabolic Alterations in Tumor Extracellular Media

The composition and intercellular interactions of tumor cells in the tissues dictate the biochemical and metabolic properties of the tumor microenvironment. The metabolic rewiring has a profound impact on the properties of the microenvironment, to an extent that monitoring such perturbations could harbor diagnostic and therapeutic relevance. A growing interest in these phenomena has inspired the development of novel technologies with sufficient sensitivity and resolution to monitor metabolic alterations in the tumor microenvironment. In this context, surface-enhanced Raman scattering (SERS) can be used for the label-free detection and imaging of diverse molecules of interest among extracellular components. Herein, the application of nanostructured plasmonic substrates comprising Au nanoparticles, self-assembled as ordered superlattices, to the precise SERS detection of selected tumor metabolites, is presented. The potential of this technology is first demonstrated through the analysis of kynurenine, a secreted immunomodulatory derivative of the tumor metabolism and the related molecules tryptophan and purine derivatives. SERS facilitates the unambiguous identification of trace metabolites and allows the multiplex detection of their characteristic fingerprints under different conditions. Finally, the effective plasmonic SERS substrate is combined with a hydrogel-based three-dimensional cancer model, which recreates the tumor microenvironment, for the real-time imaging of metabolite alterations and cytotoxic effects on tumor cells.J.P. acknowledges an FPU fellowship from the Spanish Ministry of Science, Innovation and Universities. L.M.L.-M. acknowledges funding from the European Research Council (ERC AdG 787510, 4DbioSERS) and the Maria de Maeztu Units of Excellence Program from the Spanish State Research Agency (Grant No. MDM-2017-0720). C.G.-A. acknowledges a Juan de la Cierva Fellowship from the Spanish Ministry of Science, Innovation and Universities (FJCI-2016-28887). The authors thank Dr. J. Calvo and Dr. D. Otaegui at CIC biomaGUNE for support with LC/ESI-HRMS measurements. The work of A.C. was supported by the Basque Department of Industry, Tourism and Trade (Elkartek), and the department of education (IKERTALDE IT1106-16, also participated by A. Gomez-Munoz), the BBVA foundation, the MINECO (SAF2016-79381-R (FEDER/EU); Severo Ochoa Excellence Program SEV-2016-0644-18-1; Excellence Networks SAF2016-81975-REDT), European Training Networks Project (H2020-MSCA-ITN-308 2016 721532), the AECC (IDEAS175CARR, GCTRA18006CARR), La Caixa Foundation (HR17-00094), and the European Research Council (starting Grant 336343, PoC 754627). CIBERONC was co-funded with FEDER funds and funded by ISCIII. A.M. acknowledges funding from the European Research Council (Consolidator Grant 819242) and the Spanish Ministry of Science, Innovation and Universities for the excellence program SEV-2015-049

Identification of Androgen Receptor Metabolic Correlome Reveals the Repression of Ceramide Kinase by Androgens

Prostate cancer (PCa) is one of the most prevalent cancers in men. Androgen receptor signaling plays a major role in this disease, and androgen deprivation therapy is a common therapeutic strategy in recurrent disease. Sphingolipid metabolism plays a central role in cell death, survival, and therapy resistance in cancer. Ceramide kinase (CERK) catalyzes the phosphorylation of ceramide to ceramide 1-phosphate, which regulates various cellular functions including cell growth and migration. Here we show that activated androgen receptor (AR) is a repressor of CERK expression. We undertook a bioinformatics strategy using PCa transcriptomics datasets to ascertain the metabolic alterations associated with AR activity. CERK was among the most prominent negatively correlated genes in our analysis. Interestingly, we demonstrated through various experimental approaches that activated AR reduces the mRNA expression of CERK: (i) expression of CERK is predominant in cell lines with low or negative AR activity; (ii) AR agonist and antagonist repress and induce CERK mRNA expression, respectively; (iii) orchiectomy in wildtype mice or mice with PCa (harboring prostate-specific Pten deletion) results in elevated Cerk mRNA levels in prostate tissue. Mechanistically, we found that AR represses CERK through interaction with its regulatory elements and that the transcriptional repressor EZH2 contributes to this process. In summary, we identify a repressive mode of AR that influences the expression of CERK in PCa.A.G.-M. is funded by the MINECO (SAF2016-79695-R) and the department of education (IKERTALDE IT1106-16). V.T. is funded by Fundación Vasca de Innovación e Investigación Sanitarias, BIOEF (BIO15/CA/052), the AECC J.P. Bizkaia and the Basque Department of Health (2016111109) and the MINECO RTI2018-097267-B-I00. The work of A. Carracedo is supported by the Basque Department of Industry, Tourism and Trade (Elkartek), the department of education (IKERTALDE IT1106-16) and health (RIS3), the MICINN (PID2019-108787RB-I00 (FEDER/EU); Severo Ochoa Excellence Accreditation SEV-2016-0644; Excellence Networks RED2018-102769-T), the AECC (GCTRA18006CARR), La Caixa Foundation (ID 100010434), under the agreement LCF/PR/HR17/ and the European Research Council (Consolidator Grant 819242). CIBERONC was co-funded with FEDER funds and funded by ISCIII

Molecular method for the characterization of Coxiella burnetii from clinical and environmental samples: variability of genotypes in Spain

BACKGROUND: Coxiella burnetii is a highly clonal microorganism which is difficult to culture, requiring BSL3 conditions for its propagation. This leads to a scarce availability of isolates worldwide. On the other hand, published methods of characterization have delineated up to 8 different genomic groups and 36 genotypes. However, all these methodologies, with the exception of one that exhibited limited discriminatory power (3 genotypes), rely on performing between 10 and 20 PCR amplifications or sequencing long fragments of DNA, which make their direct application to clinical samples impracticable and leads to a scarce accessibility of data on the circulation of C. burnetii genotypes. RESULTS: To assess the variability of this organism in Spain, we have developed a novel method that consists of a multiplex (8 targets) PCR and hybridization with specific probes that reproduce the previous classification of this organism into 8 genomic groups, and up to 16 genotypes. It allows for a direct characterization from clinical and environmental samples in a single run, which will help in the study of the different genotypes circulating in wild and domestic cycles as well as from sporadic human cases and outbreaks. The method has been validated with reference isolates. A high variability of C. burnetii has been found in Spain among 90 samples tested, detecting 10 different genotypes, being those adaA negative associated with acute Q fever cases presenting as fever of intermediate duration with liver involvement and with chronic cases. Genotypes infecting humans are also found in sheep, goats, rats, wild boar and ticks, and the only genotype found in cattle has never been found among our clinical samples. CONCLUSIONS: This newly developed methodology has permitted to demonstrate that C. burnetii is highly variable in Spain. With the data presented here, cattle seem not to participate in the transmission of C. burnetii to humans in the samples studied, while sheep, goats, wild boar, rats and ticks share genotypes with the human population

Genotyping of <it>Coxiella burnetii</it> from domestic ruminants in northern Spain

Abstract Background Information on the genotypic diversity of Coxiella burnetii isolates from infected domestic ruminants in Spain is limited. The aim of this study was to identify the C. burnetii genotypes infecting livestock in Northern Spain and compare them to other European genotypes. A commercial real-time PCR targeting the IS1111a insertion element was used to detect the presence of C. burnetii DNA in domestic ruminants from Spain. Genotypes were determined by a 6-loci Multiple Locus Variable number tandem repeat analysis (MLVA) panel and Multispacer Sequence Typing (MST). Results A total of 45 samples from 4 goat herds (placentas, N = 4), 12 dairy cattle herds (vaginal mucus, individual milk, bulk tank milk, aerosols, N = 20) and 5 sheep flocks (placenta, vaginal swabs, faeces, air samples, dust, N = 21) were included in the study. Samples from goats and sheep were obtained from herds which had suffered abortions suspected to be caused by C. burnetii, whereas cattle samples were obtained from animals with reproductive problems compatible with C. burnetii infection, or consisted of bulk tank milk (BTM) samples from a Q fever surveillance programme. C. burnetii genotypes identified in ruminants from Spain were compared to those detected in other countries. Three MLVA genotypes were found in 4 goat farms, 7 MLVA genotypes were identified in 12 cattle herds and 4 MLVA genotypes were identified in 5 sheep flocks. Clustering of the MLVA genotypes using the minimum spanning tree method showed a high degree of genetic similarity between most MLVA genotypes. Overall 11 different MLVA genotypes were obtained corresponding to 4 different MST genotypes: MST genotype 13, identified in goat, sheep and cattle from Spain; MST genotype 18, only identified in goats; and, MST genotypes 8 and 20, identified in small ruminants and cattle, respectively. All these genotypes had been previously identified in animal and human clinical samples from several European countries, but some of the MLVA genotypes are described here for the first time. Conclusions Genotyping revealed a substantial genetic diversity among domestic ruminants from Northern Spain.</p

Multiplex SERS Detection of Metabolic Alterations in Tumor Extracellular Media

The composition and intercellular interactions of tumor cells in the tissues dictate the biochemical and metabolic properties of the tumor microenvironment. The metabolic rewiring has a profound impact on the properties of the microenvironment, to an extent that monitoring such perturbations could harbor diagnostic and therapeutic relevance. A growing interest in these phenomena has inspired the development of novel technologies with sufficient sensitivity and resolution to monitor metabolic alterations in the tumor microenvironment. In this context, surface-enhanced Raman scattering (SERS) can be used for the label-free detection and imaging of diverse molecules of interest among extracellular components. Herein, the application of nanostructured plasmonic substrates comprising Au nanoparticles, self-assembled as ordered superlattices, to the precise SERS detection of selected tumor metabolites, is presented. The potential of this technology is first demonstrated through the analysis of kynurenine, a secreted immunomodulatory derivative of the tumor metabolism and the related molecules tryptophan and purine derivatives. SERS facilitates the unambiguous identification of trace metabolites and allows the multiplex detection of their characteristic fingerprints under different conditions. Finally, the effective plasmonic SERS substrate is combined with a hydrogel-based three-dimensional cancer model, which recreates the tumor microenvironment, for the real-time imaging of metabolite alterations and cytotoxic effects on tumor cells.J.P. acknowledges an FPU fellowship from the Spanish Ministry of Science, Innovation and Universities. L.M.L.-M. acknowledges funding from the European Research Council (ERC AdG 787510, 4DbioSERS) and the Maria de Maeztu Units of Excellence Program from the Spanish State Research Agency (Grant No. MDM-2017-0720). C.G.-A. acknowledges a Juan de la Cierva Fellowship from the Spanish Ministry of Science, Innovation and Universities (FJCI-2016-28887). The authors thank Dr. J. Calvo and Dr. D. Otaegui at CIC biomaGUNE for support with LC/ESI-HRMS measurements. The work of A.C. was supported by the Basque Department of Industry, Tourism and Trade (Elkartek), and the department of education (IKERTALDE IT1106-16, also participated by A. Gomez-Munoz), the BBVA foundation, the MINECO (SAF2016-79381-R (FEDER/EU); Severo Ochoa Excellence Program SEV-2016-0644-18-1; Excellence Networks SAF2016-81975-REDT), European Training Networks Project (H2020-MSCA-ITN-308 2016 721532), the AECC (IDEAS175CARR, GCTRA18006CARR), La Caixa Foundation (HR17-00094), and the European Research Council (starting Grant 336343, PoC 754627). CIBERONC was co-funded with FEDER funds and funded by ISCIII. A.M. acknowledges funding from the European Research Council (Consolidator Grant 819242) and the Spanish Ministry of Science, Innovation and Universities for the excellence program SEV-2015-049

Multiplex SERS Detection of Metabolic Alterations in Tumor Extracellular Media

The composition and intercellular interactions of tumor cells in the tissues dictate the biochemical and metabolic properties of the tumor microenvironment. The metabolic rewiring has a profound impact on the properties of the microenvironment, to an extent that monitoring such perturbations could harbor diagnostic and therapeutic relevance. A growing interest in these phenomena has inspired the development of novel technologies with sufficient sensitivity and resolution to monitor metabolic alterations in the tumor microenvironment. In this context, surface‐enhanced Raman scattering (SERS) can be used for the label‐free detection and imaging of diverse molecules of interest among extracellular components. Herein, the application of nanostructured plasmonic substrates comprising Au nanoparticles, self‐assembled as ordered superlattices, to the precise SERS detection of selected tumor metabolites, is presented. The potential of this technology is first demonstrated through the analysis of kynurenine, a secreted immunomodulatory derivative of the tumor metabolism and the related molecules tryptophan and purine derivatives. SERS facilitates the unambiguous identification of trace metabolites and allows the multiplex detection of their characteristic fingerprints under different conditions. Finally, the effective plasmonic SERS substrate is combined with a hydrogel‐based three‐dimensional cancer model, which recreates the tumor microenvironment, for the real‐time imaging of metabolite alterations and cytotoxic effects on tumor cells.J.P. acknowledges an FPU fellowship from the Spanish Ministry of Science, Innovation and Universities. L.M.L.‐M. acknowledges funding from the European Research Council (ERC AdG 787510, 4DbioSERS) and the Maria de Maeztu Units of Excellence Program from the Spanish State Research Agency (Grant No. MDM‐2017‐0720). C.G.‐A. acknowledges a Juan de la Cierva Fellowship from the Spanish Ministry of Science, Innovation and Universities (FJCI‐2016‐28887). The authors thank Dr. J. Calvo and Dr. D. Otaegui at CIC biomaGUNE for support with LC/ESI‐HRMS measurements. The work of A.C. was supported by the Basque Department of Industry, Tourism and Trade (Elkartek), and the department of education (IKERTALDE IT1106‐16, also participated by A. Gomez‐Muñoz), the BBVA foundation, the MINECO (SAF2016‐79381‐R (FEDER/EU); Severo Ochoa Excellence Program SEV‐2016‐0644‐18‐1; Excellence Networks SAF2016‐81975‐REDT), European Training Networks Project (H2020‐MSCA‐ITN‐308 2016 721532), the AECC (IDEAS175CARR, GCTRA18006CARR), La Caixa Foundation (HR17‐00094), and the European Research Council (starting Grant 336343, PoC 754627). CIBERONC was co‐funded with FEDER funds and funded by ISCIII. A.M. acknowledges funding from the European Research Council (Consolidator Grant 819242) and the Spanish Ministry of Science, Innovation and Universities for the excellence program SEV‐2015‐0496.J.P. acknowledges an FPU fellowship from the Spanish Ministry of Science, Innovation and Universities. L.M.L.‐M. acknowledges funding from the European Research Council (ERC AdG 787510, 4DbioSERS) and the Maria de Maeztu Units of Excellence Program from the Spanish State Research Agency (Grant No. MDM‐2017‐0720). C.G.‐A. acknowledges a Juan de la Cierva Fellowship from the Spanish Ministry of Science, Innovation and Universities (FJCI‐2016‐28887). The authors thank Dr. J. Calvo and Dr. D. Otaegui at CIC biomaGUNE for support with LC/ESI‐HRMS measurements. The work of A.C. was supported by the Basque Department of Industry, Tourism and Trade (Elkartek), and the department of education (IKERTALDE IT1106‐16, also participated by A. Gomez‐Muñoz), the BBVA foundation, the MINECO (SAF2016‐79381‐R (FEDER/EU); Severo Ochoa Excellence Program SEV‐2016‐0644‐18‐1; Excellence Networks SAF2016‐81975‐REDT), European Training Networks Project (H2020‐MSCA‐ITN‐308 2016 721532), the AECC (IDEAS175CARR, GCTRA18006CARR), La Caixa Foundation (HR17‐00094), and the European Research Council (starting Grant 336343, PoC 754627). CIBERONC was co‐funded with FEDER funds and funded by ISCIII. A.M. acknowledges funding from the European Research Council (Consolidator Grant 819242) and the Spanish Ministry of Science, Innovation and Universities for the excellence program SEV‐2015‐0496.Peer reviewe

Four-Year Evaluation of the Effect of Vaccination against Coxiella burnetii on Reduction of Animal Infection and Environmental Contamination in a Naturally Infected Dairy Sheep Flock ▿

Vaccination is considered one of the best options for controlling Coxiella burnetii infection in livestock. The efficacy of a phase I vaccine was investigated over 4 years in a sheep flock with confirmed C. burnetii infection. Shedding was not detected in ewes and yearlings in the last 2 years, but C. burnetii still persisted in the environment

Molecular method for the characterization of <it>Coxiella burnetii</it> from clinical and environmental samples: variability of genotypes in Spain

Abstract Background Coxiella burnetii is a highly clonal microorganism which is difficult to culture, requiring BSL3 conditions for its propagation. This leads to a scarce availability of isolates worldwide. On the other hand, published methods of characterization have delineated up to 8 different genomic groups and 36 genotypes. However, all these methodologies, with the exception of one that exhibited limited discriminatory power (3 genotypes), rely on performing between 10 and 20 PCR amplifications or sequencing long fragments of DNA, which make their direct application to clinical samples impracticable and leads to a scarce accessibility of data on the circulation of C. burnetii genotypes. Results To assess the variability of this organism in Spain, we have developed a novel method that consists of a multiplex (8 targets) PCR and hybridization with specific probes that reproduce the previous classification of this organism into 8 genomic groups, and up to 16 genotypes. It allows for a direct characterization from clinical and environmental samples in a single run, which will help in the study of the different genotypes circulating in wild and domestic cycles as well as from sporadic human cases and outbreaks. The method has been validated with reference isolates. A high variability of C. burnetii has been found in Spain among 90 samples tested, detecting 10 different genotypes, being those adaA negative associated with acute Q fever cases presenting as fever of intermediate duration with liver involvement and with chronic cases. Genotypes infecting humans are also found in sheep, goats, rats, wild boar and ticks, and the only genotype found in cattle has never been found among our clinical samples. Conclusions This newly developed methodology has permitted to demonstrate that C. burnetii is highly variable in Spain. With the data presented here, cattle seem not to participate in the transmission of C. burnetii to humans in the samples studied, while sheep, goats, wild boar, rats and ticks share genotypes with the human population.</p