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1.
FASEB J ; 38(16): e23888, 2024 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-39157983

RESUMO

Maf1, originally described as a repressor of RNA polymerase III (RNAP III) transcription in yeast, participates in multiple functions across eukaryotes. However, the knowledge about Maf1 in protozoan parasites is scarce. To initiate the study of Maf1 in Leishmania major, we generated a cell line that overexpresses this protein. Overexpression of Maf1 led to a significant reduction in the abundance of tRNAs, 5S rRNA, and U4 snRNA, demonstrating that Maf1 regulates RNAP III activity in L. major. To further explore the roles played by Maf1 in this microorganism, global transcriptomic and proteomic changes due to Maf1 overexpression were determined using RNA-sequencing and label-free quantitative mass spectrometry. Compared to wild-type cells, differential expression was observed for 1082 transcripts (615 down-regulated and 467 up-regulated) and 205 proteins (132 down-regulated and 73 up-regulated) in the overexpressing cells. A correlation of 44% was found between transcriptomic and proteomic results. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that the differentially expressed genes and proteins are mainly involved in transcription, cell cycle regulation, lipid metabolism and transport, ribosomal biogenesis, carbohydrate metabolism, autophagy, and cytoskeleton modification. Thus, our results suggest the involvement of Maf1 in the regulation of all these processes in L. major, as reported in other species, indicating that the functions performed by Maf1 were established early in eukaryotic evolution. Notably, our data also suggest the participation of L. major Maf1 in mRNA post-transcriptional control, a role that, to the best of our knowledge, has not been described in other organisms.


Assuntos
Leishmania major , Proteoma , Transcriptoma , Leishmania major/metabolismo , Leishmania major/genética , Proteoma/metabolismo , Humanos , RNA Polimerase III/metabolismo , RNA Polimerase III/genética , Proteínas de Protozoários/metabolismo , Proteínas de Protozoários/genética , Proteínas Repressoras/metabolismo , Proteínas Repressoras/genética , Regulação da Expressão Gênica
2.
Radiology ; 268(3): 729-37, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23674790

RESUMO

PURPOSE: To (a) determine the diagnostic performance of 64-section multidetector computed tomography (CT) trajectography for penetrating diaphragmatic injury (PDI), (b) determine the diagnostic performance of classic signs of diaphragmatic injury at 64-section multidetector CT, and (c) compare the performance of these signs with that of trajectography. MATERIALS AND METHODS: This HIPAA-compliant retrospective study had institutional review board approval, with a waiver of the informed consent requirement. All patients who had experienced penetrating thoracoabdominal trauma, who had undergone preoperative 64-section multidetector CT of the chest and abdomen, and who had surgical confirmation of findings during a 2.5-year period were included in this study (25 male patients, two female patients; mean age, 32.6 years). After a training session, four trauma radiologists unaware of the surgical outcome independently reviewed all CT studies and scored the probability of PDI on a six-point scale. Collar sign, dependent viscera sign, herniation, contiguous injury on both sides of the diaphragm, discontinuous diaphragm sign, and transdiaphragmatic trajectory were evaluated for sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV). Accuracies were determined and receiver operating characteristic curves were analyzed. RESULTS: Sensitivities for detection of PDI by using 64-section multidetector CT with postprocessing software ranged from 73% to 100%, specificities ranged from 50% to 92%, NPVs ranged from 71% to 100%, PPVs ranged from 68% to 92%, and accuracies ranged from 70% to 89%. Discontinuous diaphragm, herniation, collar, and dependent viscera signs were highly specific (92%-100%) but nonsensitive (0%-60%). Contiguous injury was generally more sensitive (80%-93% vs 73%-100%) but less specific (50%-67% vs 83%-92%) than transdiaphragmatic trajectory when patients with multiple entry wounds were included in the analysis. Transdiaphragmatic trajectory was a much more sensitive sign of PDI than previously reported (73%-100% vs 36%), with NPVs ranging from 71% to 100% and PPVs ranging from 85% to 92%. CONCLUSION: Sixty-four-section multidetector CT trajectography facilitates the identification of transdiaphragmatic trajectory, which accurately rules in PDI when identified. Contiguous injury remains a highly sensitive sign, even when patients with multiple injuries are considered, and is useful for excluding PDI.


Assuntos
Diafragma/diagnóstico por imagem , Diafragma/lesões , Tomografia Computadorizada por Raios X/estatística & dados numéricos , Ferimentos Penetrantes/diagnóstico por imagem , Ferimentos Penetrantes/epidemiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Florida/epidemiologia , Humanos , Incidência , Masculino , Reprodutibilidade dos Testes , Medição de Risco , Sensibilidade e Especificidade , Adulto Jovem
3.
Environ Microbiol ; 14(9): 2495-510, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22564293

RESUMO

In this study we examined the microbial community composition and operating metabolisms on the surface and in the permafrost of Deception Island, (Antarctica) with an on site antibody microarray biosensor. Samples (down to a depth of 4.2 m) were analysed with LDChip300 (Life Detector Chip), an immunosensor containing more than 300 antibodies targeted to bacterial and archaeal antigens. The immunograms showed positive antigen-antibody reactions in all surface samples (lichens, pyroclasts) and the top layer of the permafrost. The results indicated the presence of exopolysaccharides, bacteria belonging to the Alpha-, Delta- and Gammaproteobacteria, Bacteroidetes, Gram-positive Actinobacteria and Firmicutes, as well as archaeal species, most probably Methanobacterium spp. Positive reactions with antibodies to proteins and peptides revealed the presence of nitrogen fixation (NifHD, GlnB, HscA), methanogenic (McrB), iron homeostasis and iron scavenging (ferritins and DPS proteins) proteins, as well as ABC transporters, which indicated that these processes were operating at the time of sampling. These results were validated with other molecular ecology techniques such as oligonucleotide microarrays, 16S bacterial rRNA gene sequence analysis, aerobic viable counts and microscopy. Molecular ecology results showed a differentiated pattern along the depth of the drill, being the top active layer the most diverse, with Acidobacteria, Actinobacteria, Proteobacteria, Bacteroidetes and the phototrophs Cyanobacteria and Chloroflexi as dominant groups. Actinobacteria and Firmicutes were dominant in depths from 0.5 to 2 m, and Betaproteobacteria from 3 to 4.2 m. The geochemical analysis revealed the presence of low molecular weight organic acids (acetate, formate) which could be used by microorganisms as energy sources for sulfate, nitrate and metal reduction under anaerobic conditions.


Assuntos
Archaea , Bactérias , Biodiversidade , Ilhas , Microbiologia do Solo , Regiões Antárticas , Antígenos Arqueais/metabolismo , Antígenos de Bactérias/metabolismo , Archaea/classificação , Archaea/genética , Archaea/metabolismo , Bactérias/classificação , Bactérias/genética , Bactérias/metabolismo , Carga Bacteriana , Biomarcadores/análise , Microscopia Eletrônica de Varredura , Filogenia , RNA Ribossômico 16S/genética , Solo/química
4.
Radiology ; 263(3): 645-60, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22623691

RESUMO

As computed tomography (CT) technology has evolved, multidetector CT has become an integral part of the initial assessment of many injured patients, and the spine is easily included in the total body screening performed in patients with severe blunt polytrauma. Despite all the advantages of multidetector CT, clearing the spine in which injury is suspected continues to be a daily challenge in clinical practice. The purpose of this review is to present the evidence and the controversies surrounding the practice of imaging in patients suspected of having spine injury. The discussion is centered on the increasing reliance on multidetector CT in the work-up of these patients but also considers the important contributions of clinical trials to select patient for appropriate imaging on the basis of risk and probability of injury. Available protocols, injury classification systems, and issues awaiting future research are addressed.


Assuntos
Tomografia Computadorizada Multidetectores/métodos , Traumatismos da Coluna Vertebral/diagnóstico por imagem , Adulto , Ensaios Clínicos como Assunto , Meios de Contraste , Humanos , Traumatismo Múltiplo/diagnóstico por imagem , Medição de Risco , Traumatismos da Coluna Vertebral/classificação , Imagem Corporal Total , Ferimentos não Penetrantes/diagnóstico por imagem
5.
J Vet Diagn Invest ; 34(2): 190-198, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-34852683

RESUMO

During the COVID-19 pandemic, infection of farmed mink has become not only an economic issue but also a widespread public health concern. International agencies have advised the use of strict molecular and serosurveillance methods for monitoring the SARS-CoV2 status on mink farms. We developed 2 ELISAs and a duplex protein microarray immunoassay (MI), all in a double-recognition format (DR), to detect SARS-CoV2 antibodies specific to the receptor-binding domain (RBD) of the spike protein and to the full-length nucleoprotein (N) in mink sera. We collected 264 mink serum samples and 126 oropharyngeal samples from 5 Spanish mink farms. In both of the ELISAs and the MI, RBD performed better than N protein for serologic differentiation of mink from SARS-CoV2-positive and -negative farms. Therefore, RBD was the optimal antigenic target for serosurveillance of mink farms.


Assuntos
COVID-19 , Vison , Animais , Anticorpos Antivirais , COVID-19/veterinária , Fazendas , Imunoensaio/veterinária , Pandemias , RNA Viral , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus
6.
Environ Microbiol ; 13(6): 1421-32, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21401847

RESUMO

The sandwich microarray immunoassay (SMI) is a powerful technique for the analysis and characterization of environmental samples, from the identification of microorganisms to specific bioanalytes. As the number of antibodies increases, however, unspecific binding and cross-reactivity can become a problem. To cope with such difficulties, we present here the concept of antibody graph associated to a sandwich antibody microarray. Antibody graphs give valuable information about the antibody cross-reactivity network and all the players involved in the sandwich format: capturing and tracer antibodies, the antigenic sample and the degree of cross-reactivity between antibodies. Making use of the information contained in the antibody graph, we have developed a deconvolution method that disentangles the antibody cross-reactivity events and gives qualitative information about the composition of the experimental sample under study. We have validated the method by using a 66 antibody-containing microarray to describe known antigenic mixtures as well as natural environmental samples characterized by 16S-RNA gene phylogenetic analysis. The application of our antibody graph and deconvolution method allowed us to discriminate between true specific antigen-antibody reactions and spurious signals on a microarray designed for environmental monitoring.


Assuntos
Anticorpos/análise , Microbiologia Ambiental , Monitoramento Ambiental/métodos , Anticorpos/imunologia , Reações Cruzadas , Imunoensaio/métodos , Filogenia , Análise Serial de Proteínas , RNA Ribossômico 16S
7.
AJR Am J Roentgenol ; 196(3): W309-15, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21343481

RESUMO

OBJECTIVE: CT angiography (CTA) using 64-MDCT enables timely evaluation of injuries associated with blunt neck trauma. The purpose of this article is to familiarize the reader with the most frequent CTA signs of blunt vascular injury. CONCLUSION: CTA is a valuable tool to detect blunt vascular injuries, especially using its multiplanar and 3D reconstruction capabilities.


Assuntos
Angiografia/métodos , Lesões do Pescoço/diagnóstico por imagem , Tomografia Computadorizada por Raios X/métodos , Ferimentos não Penetrantes/diagnóstico por imagem , Humanos , Imageamento Tridimensional
8.
Orig Life Evol Biosph ; 41(4): 331-45, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21161385

RESUMO

Clasically, prebiotic chemistry has focused on the production and identification of simple organic molecules, many of them forming part of "intractable polymers" named tholins. In a previous work, we demonstrated that in experiments using an external energy source and inorganic carbon the aqueous aerosols improved the formation of hydrophilic tholins. Herein, we elucidate the role of pH (from 4 to 12) in prebiotic experiments using saline aqueous aerosols, spark discharges and an atmosphere containing CH(4). At all values of pH, the saline aqueous aerosols increased the production of a significant variety of carboxylic acids that could have been present in a primitive Krebs cycle. Moreover, the study for the first time of hydrophilic tholins by 2-D electrophoresis revealed that these are formed by a set of unexpected heavy polymeric species. The initial alkaline conditions significantly increased both the apparent molecular weight of polymeric species up to 80 kDa and their diversity. We propose the term of protobiopolymers to denote those polymeric species fractionated by 2-D electrophoresis since these are formed by biomolecules present in living systems and show diversity in length as well as in functional groups. Thus, aerosols formed in simulated alkaline ocean conditions could provide an optimal medium for the formation of the primeval materials that could be precursors to the emergence of life.


Assuntos
Aerossóis/química , Ácidos Carboxílicos/síntese química , Fracionamento Químico/métodos , Eletroforese em Gel Bidimensional/métodos , Polímeros/síntese química , Atmosfera , Eletricidade , Eletroforese em Gel de Poliacrilamida/métodos , Evolução Química , Cromatografia Gasosa-Espectrometria de Massas/métodos , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Metano/química , Oceanos e Mares , Polímeros/química , Compostos de Amônio Quaternário/química , Salinidade , Espectrofotometria Infravermelho/métodos
9.
Front Microbiol ; 11: 590736, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33391207

RESUMO

Perchlorate anions are produced by chemical industries and are important contaminants in certain natural ecosystems. Perchlorate also occurs in some natural and uncontaminated environments such as the Atacama Desert, the high Arctic or the Antarctic Dry Valleys, and is especially abundant on the surface of Mars. As some bacterial strains are capable of using perchlorate as an electron acceptor under anaerobic conditions, their detection is relevant for environmental monitoring on Earth as well as for the search for life on Mars. We have developed an antibody microarray with 20 polyclonal antibodies to detect perchlorate-reducing bacteria (PRB) strains and two crucial and highly conserved enzymes involved in perchlorate respiration: perchlorate reductase and chlorite dismutase. We determined the cross-reactivity, the working concentration, and the limit of detection of each antibody individually and in a multiplex format by Fluorescent Sandwich Microarray Immunoassay. Although most of them exhibited relatively high sensitivity and specificity, we applied a deconvolution method based on graph theory to discriminate between specific signals and cross-reactions from related microorganisms. We validated the system by analyzing multiple bacterial isolates, crude extracts from contaminated reactors and salt-rich natural samples from the high Arctic. The PRB detecting chip (PRBCHIP) allowed us to detect and classify environmental isolates as well as to detect similar strains by using crude extracts obtained from 0.5 g even from soils with low organic-matter levels (<103 cells/g of soil). Our results demonstrated that PRBCHIP is a valuable tool for sensitive and reliable detection of perchlorate-reducing bacteria for research purposes, environmental monitoring and planetary exploration.

10.
Semin Ultrasound CT MR ; 30(3): 195-204, 2009 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19537052

RESUMO

Penetrating neck trauma may occur from gun shots, stabbings, and accidental injury. Approximately 50% of gunshot and 10%-20% of stab-wound patients are reported as having severe injuries and the mortality from severe vascular injuries is reported as high as 50%. Penetrating traumatic neck injury is no longer best managed with exploratory surgery or conventional angiography in the stable patient. Computed tomography angiography has proven to be a useful, safe, and reliable means of diagnosis. Experience with interpreting and reconstructing computed tomography images, understanding the clinically important findings, and avoiding pitfalls is critical for successful patient care. Therefore, radiologists and clinicians who treat trauma patients need to familiarize themselves with the computed tomography angiography technique and appearances of vascular injury and know when to recommend further evaluation, such as interventional angiography or open surgery.


Assuntos
Lesões do Pescoço/diagnóstico por imagem , Tomografia Computadorizada por Raios X/métodos , Ferimentos Penetrantes/diagnóstico por imagem , Falso Aneurisma/diagnóstico por imagem , Angiografia/métodos , Humanos , Pescoço/anatomia & histologia , Pescoço/irrigação sanguínea , Radiografia Intervencionista
11.
Anal Chem ; 80(21): 7970-9, 2008 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-18837515

RESUMO

Environmental biomonitoring approaches require the measurement of either unequivocal biomarkers or specific biological profiles. Antibody microarrays constitute new tools for fast and reliable analysis of up to hundreds of biomarkers simultaneously. Herein we report 150 new polyclonal antibodies against microbial strains and environmental extracts, as well as the construction and validation of an antibody microarray (EMCHIP200, for "Environmental Monitoring Chip") containing 200 different antibodies. Each antibody was tested against its antigen for its specificity and cross-reactivity by a sandwich microarray immunoassay. The limit of detection was 0.2 ng mL (-1) for some proteins and 10 (4)-10 (5) cells mL (-1) for bacterial cells and spores. Partial biochemical characterization allowed identification of polymeric compounds (proteins and polysaccharides) as some of the targets recognized by the antibodies. We have successfully used the EMCHIP200 for the detection of biological polymers in samples from extreme environments around the world (e.g., a deep South African mine, Antarctica's dry valleys, Yellowstone National Park, Iceland, and Rio Tinto surface and subsurface). Clustering analysis permitted us to associate similar immunoprofiles or patterns to samples from apparently very different environments, indicating that they indeed share similar universal biomarkers. Our EMCHIP200 constitutes a new generation of immunosensors for biomarker detection and profiling, for either environmental, industrial, biotechnological, or astrobiological applications.


Assuntos
Anticorpos/análise , Anticorpos/imunologia , Monitoramento Ambiental/métodos , Imunoensaio/métodos , Análise em Microsséries/métodos , Biomarcadores , Reprodutibilidade dos Testes , Microbiologia do Solo
12.
Astrobiology ; 18(5): 586-606, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-27893284

RESUMO

Oligotrophic glacial lakes in the Andes Mountains serve as models to study the effects of climate change on natural biological systems. The persistent high UV regime and evolution of the lake biota due to deglaciation make Andean lake ecosystems potential analogues in the search for life on other planetary bodies. Our objective was to identify microbial biomarkers and metabolic patterns that represent time points in the evolutionary history of Andean glacial lakes, as these may be used in long-term studies as microscale indicators of climate change processes. We investigated a variety of microbial markers in shallow sediments from Laguna Negra and Lo Encañado lakes (Región Metropolitana, Chile). An on-site immunoassay-based Life Detector Chip (LDChip) revealed the presence of sulfate-reducing bacteria, methanogenic archaea, and exopolymeric substances from Gammaproteobacteria. Bacterial and archaeal 16S rRNA gene sequences obtained from field samples confirmed the results from the immunoassays and also revealed the presence of Alpha-, Beta-, Gamma-, and Deltaproteobacteria, as well as cyanobacteria and methanogenic archaea. The complementary immunoassay and phylogenetic results indicate a rich microbial diversity with active sulfate reduction and methanogenic activities along the shoreline and in shallow sediments. Sulfate inputs from the surrounding volcanic terrains during deglaciation may explain the observed microbial biomarker and metabolic patterns, which differ with depth and between the two lakes. A switch from aerobic and heterotrophic metabolisms to anaerobic ones such as sulfate reduction and methanogenesis in the shallow shores likely reflects the natural evolution of the lake sediments due to deglaciation. Hydrodynamic deposition of sediments creates compartmentalization (e.g., sediments with different structure and composition surrounded by oligotrophic water) that favors metabolic transitions. Similar phenomena would be expected to occur on other planetary lakes, such as those of Titan, where watery niches fed by depositional events would be surrounded by a "sea" of hydrocarbons. Key Words: Glacier lakes-Sedimentation-Prokaryotic metabolisms and biomarkers-Deglaciation-Life detection-Planetary exploration. Astrobiology 18, 586-606.


Assuntos
Sedimentos Geológicos/microbiologia , Lagos/microbiologia , Planetas , Archaea/genética , Archaea/isolamento & purificação , Archaea/metabolismo , Bactérias/genética , Bactérias/isolamento & purificação , Bactérias/metabolismo , Biodiversidade , Biomarcadores/análise , Filogenia , RNA Ribossômico 16S/isolamento & purificação
13.
JSLS ; 10(2): 239-43, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16882428

RESUMO

Injury to the spleen during routine colonoscopy is an extremely rare injury. Diagnosis and management of the injury has evolved with technological advances and experience gained in the management of splenic injuries sustained in trauma. Of the 37 reported cases of colonoscopic splenic injury, 12 had a history of prior surgery or a disease process suggesting the presence of adhesions. Only 6 had noted difficulty during the procedure, and 31 patients experienced pain, shock, or hemoglobin drop as the indication of splenic injury. Since 1989, 21/24 (87.5%) patients have been diagnosed initially using computed tomography or ultrasonography. Overall, only 27.8% have retained their spleens. None have experienced as long a delay as our patient, nor have any had an attempt at percutaneous control of the injury. This report presents an unusual case of a rare complication of colonoscopy and the unsuccessful use of one nonoperative technique, and reviews the experience reported in the world literature, including current day management options.


Assuntos
Colonoscopia/efeitos adversos , Baço/lesões , Baço/cirurgia , Idoso , Feminino , Humanos
14.
Radiol Clin North Am ; 41(3): 599-616, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12797608

RESUMO

The introduction of CT imaging in the 1970s revolutionized all aspects of medical care, perhaps nowhere more so than in the evaluation of acutely injured patients. Just as single-slice helical scanning was a great advance over conventional CT, the capabilities of MSCT are proving to be dramatically superior to single-slice methods. Improved contrast bolus imaging, thinner slices, and isotropic voxels should enable the trauma radiologist to identify both major organ system disruption and subtle injuries more promptly. Multiplanar and three-dimensional reconstructions, a forte of MSCT, facilitate rapid communication of disease states with surgeons and others involved in the care of injured patients. In many centers, whole-body CT is beginning to supplant plain films of the chest and spine in the evaluation of severe trauma victims; the cost-effectiveness of such methods is still under evaluation.


Assuntos
Traumatismos Torácicos/diagnóstico por imagem , Tomografia Computadorizada por Raios X/métodos , Humanos , Processamento de Imagem Assistida por Computador , Imageamento Tridimensional , Radiografia Torácica
15.
PLoS One ; 9(12): e114180, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25531640

RESUMO

In the South African deep mines, a variety of biofilms growing in mine corridor walls as water seeps from intersections or from fractures represents excellent proxies for deep-subsurface environments. However, they may be greatly affected by the oxygen inputs through the galleries of mining activities. As a consequence, the interaction between the anaerobic water coming out from the walls with the oxygen inputs creates new conditions that support rich microbial communities. The inherent difficulties for sampling these delicate habitats, together with transport and storage conditions may alter the community features and composition. Therefore, the development of in situ monitoring methods would be desirable for quick evaluation of the microbial community. In this work, we report the usefulness of an antibody-microarray (EMChip66) immunoassay for a quick check of the microbial diversity of biofilms located at 1.3 km below surface within the Beatrix deep gold mine (South Africa). In addition, a deconvolution method, previously described and used for environmental monitoring, based on graph theory and applied on antibody cross-reactivity was used to interpret the immunoassay results. The results were corroborated and further expanded by 16S rRNA gene sequencing analysis. Both culture-independent techniques coincided in detecting features related to aerobic sulfur-oxidizers, aerobic chemoorganotrophic Alphaproteobacteria and metanotrophic Gammaproteobacteria. 16S rRNA gene sequencing detected phylotypes related to nitrate-reducers and anaerobic sulfur-oxidizers, whereas the EMChip66 detected immunological features from methanogens and sulfate-reducers. The results reveal a diverse microbial community with syntrophic metabolisms both anaerobic (fermentation, methanogenesis, sulphate and nitrate reduction) and aerobic (methanotrophy, sulphur oxidation). The presence of oxygen-scavenging microbes might indicate that the system is modified by the artificial oxygen inputs from the mine galleries.


Assuntos
Archaea/metabolismo , Bactérias/metabolismo , Biofilmes/crescimento & desenvolvimento , Gráficos por Computador , Mineração , Análise Serial de Proteínas , Anticorpos Antiarchaea/metabolismo , Anticorpos Antibacterianos/metabolismo , Archaea/genética , Archaea/imunologia , Archaea/fisiologia , Bactérias/genética , Bactérias/imunologia , Fenômenos Fisiológicos Bacterianos , Ouro , Imunoensaio , Oxigênio/metabolismo , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , África do Sul
16.
PLoS One ; 7(12): e50799, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23272072

RESUMO

The study of cross-reactivity in allergy is key to both understanding. the allergic response of many patients and providing them with a rational treatment In the present study, protein microarrays and a co-sensitization graph approach were used in conjunction with an allergen microarray immunoassay. This enabled us to include a wide number of proteins and a large number of patients, and to study sensitization profiles among members of the LTP family. Fourteen LTPs from the most frequent plant food-induced allergies in the geographical area studied were printed into a microarray specifically designed for this research. 212 patients with fruit allergy and 117 food-tolerant pollen allergic subjects were recruited from seven regions of Spain with different pollen profiles, and their sera were tested with allergen microarray. This approach has proven itself to be a good tool to study cross-reactivity between members of LTP family, and could become a useful strategy to analyze other families of allergens.


Assuntos
Alérgenos/imunologia , Antígenos de Plantas/imunologia , Proteínas de Transporte/imunologia , Proteínas de Plantas/imunologia , Análise Serial de Proteínas/métodos , Alérgenos/química , Reações Cruzadas/imunologia , Eletroforese em Gel de Poliacrilamida , Epitopos/química , Alimentos , Hipersensibilidade Alimentar/imunologia , Geografia , Humanos , Imunoensaio/métodos , Lipídeos/química , Modelos Estatísticos , Pólen , Proteínas Recombinantes/química , Espanha , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
17.
PLoS One ; 7(9): e44088, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22970164

RESUMO

Cross-reactivity of plant foods is an important phenomenon in allergy, with geographical variations with respect to the number and prevalence of the allergens involved in this process, whose complexity requires detailed studies. We have addressed the role of thaumatin-like proteins (TLPs) in cross-reactivity between fruit and pollen allergies. A representative panel of 16 purified TLPs was printed onto an allergen microarray. The proteins selected belonged to the sources most frequently associated with peach allergy in representative regions of Spain. Sera from two groups of well characterized patients, one with allergy to Rosaceae fruit (FAG) and another against pollens but tolerant to food-plant allergens (PAG), were obtained from seven geographical areas with different environmental pollen profiles. Cross-reactivity between members of this family was demonstrated by inhibition assays. Only 6 out of 16 purified TLPs showed noticeable allergenic activity in the studied populations. Pru p 2.0201, the peach TLP (41%), chestnut TLP (24%) and plane pollen TLP (22%) proved to be allergens of probable relevance to fruit allergy, being mainly associated with pollen sensitization, and strongly linked to specific geographical areas such as Barcelona, Bilbao, the Canary Islands and Madrid. The patients exhibited >50% positive response to Pru p 2.0201 and to chestnut TLP in these specific areas. Therefore, their recognition patterns were associated with the geographical area, suggesting a role for pollen in the sensitization of these allergens. Finally, the co-sensitizations of patients considering pairs of TLP allergens were analyzed by using the co-sensitization graph associated with an allergen microarray immunoassay. Our data indicate that TLPs are significant allergens in plant food allergy and should be considered when diagnosing and treating pollen-food allergy.


Assuntos
Reações Cruzadas/imunologia , Hipersensibilidade Alimentar/imunologia , Proteínas de Plantas/imunologia , Análise Serial de Proteínas , Adolescente , Adulto , Criança , Feminino , Alimentos/efeitos adversos , Hipersensibilidade Alimentar/sangue , Frutas/imunologia , Geografia , Humanos , Imunização , Imunoensaio , Imunoglobulina E/imunologia , Masculino , Pessoa de Meia-Idade , Peso Molecular , Proteínas de Plantas/isolamento & purificação , Pólen/imunologia , Espanha , Adulto Jovem
19.
Astrobiology ; 11(8): 759-73, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22007740

RESUMO

Antibody microarrays are becoming frequently used tools for analytical purposes. A key factor for optimal performance is the stability of the immobilized (capturing) antibodies as well as those that have been fluorescently labeled to achieve the immunological test (tracers). This is especially critical for long-distance transport, field testing, or planetary exploration. A number of different environmental stresses may affect the antibody integrity, such as dryness, sudden temperature shift cycles, or, as in the case of space science, exposure to large quantities of the highly penetrating gamma radiation. Here, we report on the effect of certain stabilizing solutions for long-term storage of printed antibody microarrays under different conditions. We tested the effect of gamma radiation on printed and freeze- or vacuum-dried fluorescent antibodies at working concentrations (tracer antibodies), as well as the effect of multiple cycles of sudden and prolonged temperature shifts on the stability of fluorescently labeled tracer antibody cocktails. Our results show that (i) antibody microarrays are stable at room temperature when printed on stabilizing spotting solutions for at least 6 months, (ii) lyophilized and vacuum-dried fluorescently labeled tracer antibodies are stable for more than 9 months of sudden temperature shift cycles (-20°C to 25°C and 50°C), and (iii) both printed and freeze- or vacuum-dried fluorescent tracer antibodies are stable after several-fold excess of the dose of gamma radiation expected during a mission to Mars. Although different antibodies may exhibit different susceptibilities, we conclude that, in general, antibodies are suitable for use in planetary exploration purposes if they are properly treated and stored with the use of stabilizing substances.


Assuntos
Anticorpos/análise , Corantes Fluorescentes , Raios gama , Análise Serial de Proteínas , Voo Espacial , Temperatura , Anticorpos/efeitos da radiação , Anticorpos Imobilizados/análise , Anticorpos Imobilizados/efeitos da radiação , Liofilização , Soluções
20.
Astrobiology ; 11(10): 969-96, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22149750

RESUMO

The Atacama Desert has long been considered a good Mars analogue for testing instrumentation for planetary exploration, but very few data (if any) have been reported about the geomicrobiology of its salt-rich subsurface. We performed a Mars analogue drilling campaign next to the Salar Grande (Atacama, Chile) in July 2009, and several cores and powder samples from up to 5 m deep were analyzed in situ with LDChip300 (a Life Detector Chip containing 300 antibodies). Here, we show the discovery of a hypersaline subsurface microbial habitat associated with halite-, nitrate-, and perchlorate-containing salts at 2 m deep. LDChip300 detected bacteria, archaea, and other biological material (DNA, exopolysaccharides, some peptides) from the analysis of less than 0.5 g of ground core sample. The results were supported by oligonucleotide microarray hybridization in the field and finally confirmed by molecular phylogenetic analysis and direct visualization of microbial cells bound to halite crystals in the laboratory. Geochemical analyses revealed a habitat with abundant hygroscopic salts like halite (up to 260 g kg(-1)) and perchlorate (41.13 µg g(-1) maximum), which allow deliquescence events at low relative humidity. Thin liquid water films would permit microbes to proliferate by using detected organic acids like acetate (19.14 µg g(-1)) or formate (76.06 µg g(-1)) as electron donors, and sulfate (15875 µg g(-1)), nitrate (13490 µg g(-1)), or perchlorate as acceptors. Our results correlate with the discovery of similar hygroscopic salts and possible deliquescence processes on Mars, and open new search strategies for subsurface martian biota. The performance demonstrated by our LDChip300 validates this technology for planetary exploration, particularly for the search for life on Mars.


Assuntos
Archaea/isolamento & purificação , Bactérias/isolamento & purificação , Técnicas Biossensoriais , Clima Desértico , Consórcios Microbianos , Biomarcadores/análise , Chile , Ecossistema , Marte , Salinidade
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