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1.
Plant Dis ; 2024 Feb 29.
Artigo em Inglês | MEDLINE | ID: mdl-38422440

RESUMO

Xanthomonas phaseoli pv. manihotis (Xpm) is a plant pathogenic bacterium known as the causal agent of cassava bacterial blight (CBB). CBB is the most limiting bacterial disease affecting cassava (Manihot esculenta Crantz), characterized by diverse symptoms including angular water-soaked leaf lesions, blight, wilting, stem exudates, stem cankers and dieback. CBB has been reported in most cassava-growing regions around the world, and, under conducive conditions, crop yield losses can reach up to 100% (Zárate-Chaves et al. 2021). While Xpm genetic diversity is remarkably high in South America (Bart et al. 2012) and cassava originates and was domesticated in the Amazon basin (Allem 2002), reports of CBB in the Amazonian region are missing. To fill this gap, in October 2018 we surveyed for CBB symptoms in cassava fields of the Orellana Province, located in the Amazon forest of the Republic of Ecuador. Adult cassava plants exhibiting typical angular, water-soaked leaf lesions were found in polyculture plots, i.e. intercrops of cassava with other species such as plantains and fruit trees (a.k.a. chakras). After surface disinfection with 5% sodium hypochlorite followed by 70% ethanol, white Xpm-like colonies were isolated from diseased leaf tissues of four plants on YPGA medium (yeast extract, 5 g/l; peptone, 5 g/l; glucose, 5 g/l; agar-agar, 15 g/l) supplemented with cephalexin (40 mg/l) and cycloheximide (50 mg/l). Pathogenicity tests were performed on peat-potted, 2-month-old cassava plants of the cultivar 60444. Bacterial suspensions were adjusted to an OD600 of 0.2 (2 × 108 CFU/ml) in sterile 10-mM MgCl2 and syringe infiltrated in fully-expanded leaves. In parallel, 20 µl of each bacterial suspension adjusted to an OD600 of 0.02 (2 × 107 CFU/ml) were inoculated on stems inside a hole previously punched with a sterile needle in the junction of the third-top petiole. Sterile 10-mM MgCl2 was used for mock inoculations in both leaves and stems, and experiments were replicated in three plants. Plants were incubated in a greenhouse at 28 ± 1°C with a 12-h photoperiod. Infiltrated leaves developed watersoaking 3 days post inoculation, while wilted leaves, stem exudates, and dieback were observed 21 days after stem inoculation. Control plants remained symptomless. White Xpm-like colonies were re-isolated from symptomatic leaves (Fig S1). One colony of each of the four Xpm isolates (before and after re-isolation) was assessed using diagnostic PCRs (Bernal-Galeano et al. 2018; Flores et al. 2019), using strain Xam668 as positive control. All four candidates were positive for both diagnostic tools. The sequences of the housekeeping genes atpD, dnaK, efp, glnA, gyrB and rpoD of our isolates were extracted from full genome sequences obtained through Oxford Nanopore Technologies (ONT) (GenBank OR288194 to OR288217) and compared to their homologs in four close Xanthomonas species and a reference Xpm strain (Table S1). The sequences of the tested strains aligned with that of Xpm CIO151 (GCA_004025275.1) (Arrieta-Ortiz et al. 2013) with nucleotide identity above 99.92% (Fig S2). The four strains were named CIX4169, CIX4170, CIX4171 and CIX4172, stored in the IRD Collection of Xanthomonas, where they are available upon request. To our knowledge, this is the first report of CBB in the Amazonian region and in Ecuador, where cassava is a central element for local culture and economy. Further surveys will be necessary to evaluate the distribution and prevalence of CBB in other ecozones of Ecuador where cassava is cultivated.

2.
New Phytol ; 238(4): 1593-1604, 2023 05.
Artigo em Inglês | MEDLINE | ID: mdl-36764921

RESUMO

Functional analysis of large gene families in plant pathogens can be cumbersome using classical insertional mutagenesis. Additionally, Cas9 toxicity has limited the application of CRISPR-Cas9 for directed mutagenesis in bacteria. Here, we successfully applied a CRISPR interference strategy to investigate the cryptic role of the transcription activator-like effector (tale) multigene family in several plant-pathogenic Xanthomonas bacterial species, owing to their contribution to pathogen virulence. Single guide RNAs (sgRNAs) designed against Xanthomonas phaseoli pv manihotis tale conserved gene sequences efficiently silenced expression of all tales, with concomitant decrease in virulence and TALE-induced host gene expression. The system is readily translatable to other Xanthomonas species infecting rice, citrus, Brassica, and cassava, silencing up to 16 tales in a given strain using a single sgRNA. Complementation with plasmid-borne designer tales lacking the sgRNA-targeted sequence restored molecular and virulence phenotypes in all pathosystems. Our results evidenced that X. campestris pv campestris CN08 tales are relevant for symptom development in cauliflower. They also show that the MeSWEET10a sugar transporter is surprisingly targeted by the nonvascular cassava pathogen X. cassavae, highlighting a new example of TALE functional convergence between phylogenetically distant Xanthomonas. Overall, this novel technology provides a platform for discovery and rapid functional understanding of highly conserved gene families.


Assuntos
Oryza , Xanthomonas , Efetores Semelhantes a Ativadores de Transcrição/genética , Xanthomonas/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Virulência/genética , Transporte Biológico , Doenças das Plantas/microbiologia , Oryza/genética
3.
Nucleic Acids Res ; 49(19): 11067-11082, 2021 11 08.
Artigo em Inglês | MEDLINE | ID: mdl-34606602

RESUMO

KRAS-activating mutations are oncogenic drivers and are correlated with radioresistance of multiple cancers, including colorectal cancer, but the underlying precise molecular mechanisms remain elusive. Herein we model the radiosensitivity of isogenic HCT116 and SW48 colorectal cancer cell lines bearing wild-type or various mutant KRAS isoforms. We demonstrate that KRAS mutations indeed lead to radioresistance accompanied by reduced radiotherapy-induced mitotic catastrophe and an accelerated release from G2/M arrest. Moreover, KRAS mutations result in increased DNA damage response and upregulation of 53BP1 with associated increased non-homologous end-joining (NHEJ) repair. Remarkably, KRAS mutations lead to activation of NRF2 antioxidant signaling to increase 53BP1 gene transcription. Furthermore, genetic silencing or pharmacological inhibition of KRAS, NRF2 or 53BP1 attenuates KRAS mutation-induced radioresistance, especially in G1 phase cells. These findings reveal an important role for a KRAS-induced NRF2-53BP1 axis in the DNA repair and survival of KRAS-mutant tumor cells after radiotherapy, and indicate that targeting NRF2, 53BP1 or NHEJ may represent novel strategies to selectively abrogate KRAS mutation-mediated radioresistance.


Assuntos
Neoplasias do Colo/genética , Reparo do DNA por Junção de Extremidades , Fator 2 Relacionado a NF-E2/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Tolerância a Radiação/genética , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/genética , Apoptose/genética , Apoptose/efeitos da radiação , Linhagem Celular Tumoral , Proliferação de Células/efeitos da radiação , Neoplasias do Colo/mortalidade , Neoplasias do Colo/patologia , Neoplasias do Colo/radioterapia , Quebras de DNA de Cadeia Dupla , DNA de Neoplasias/genética , DNA de Neoplasias/metabolismo , Pontos de Checagem da Fase G1 do Ciclo Celular/genética , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos da radiação , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos da radiação , Raios gama , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Humanos , Mutação , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Fator 2 Relacionado a NF-E2/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Análise de Sobrevida , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/metabolismo
4.
BMC Pulm Med ; 23(1): 326, 2023 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-37667267

RESUMO

BACKGROUND: Ex vivo lung perfusion (EVLP) constitutes a tool with great research potential due to its advantages over in vivo and in vitro models. Despite its important contribution to lung reconditioning, this technique has the disadvantage of incurring high costs and can induce pulmonary endothelial injury through perfusion and ventilation. The pulmonary endothelium is made up of endothelial glycocalyx (EG), a coating of proteoglycans (PG) on the luminal surface. PGs are glycoproteins linked to terminal sialic acids (Sia) that can affect homeostasis with responses leading to edema formation. This study evaluated the effect of two ex vivo perfusion solutions on lung function and endothelial injury. METHODS: We divided ten landrace swine into two groups and subjected them to EVLP for 120 min: Group I (n = 5) was perfused with Steen® solution, and Group II (n = 5) was perfused with low-potassium dextran-albumin solution. Ventilatory mechanics, histology, gravimetry, and sialic acid concentrations were evaluated. RESULTS: Both groups showed changes in pulmonary vascular resistance and ventilatory mechanics (p < 0.05, Student's t-test). In addition, the lung injury severity score was better in Group I than in Group II (p < 0.05, Mann-Whitney U); and both groups exhibited a significant increase in Sia concentrations in the perfusate (p < 0.05 t-Student) and Sia immunohistochemical expression. CONCLUSIONS: Sia, as a product of EG disruption during EVLP, was found in all samples obtained in the system; however, the changes in its concentration showed no apparent correlation with lung function.


Assuntos
Lesão Pulmonar , Ácido N-Acetilneuramínico , Animais , Suínos , Respiração , Perfusão , Pulmão , Modelos Teóricos
5.
Mol Pharmacol ; 99(6): 435-447, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33795352

RESUMO

Rearranged during transfection (RET) rearrangements occur in 1% to 2% of lung adenocarcinomas as well as other malignancies and are now established targets for tyrosine kinase inhibitors. We developed three novel RET fusion-positive (RET+) patient-derived cancer cell lines, CUTO22 [kinesin 5B (KIF5B)-RET fusion], CUTO32 (KIF5B-RET fusion), and CUTO42 (echinoderm microtubule-associated protein-like 4-RET fusion), to study RET signaling and response to therapy. We confirmed each of our cell lines expresses the RET fusion protein and assessed their sensitivity to RET inhibitors. We found that the CUTO22 and CUTO42 cell lines were sensitive to multiple RET inhibitors, whereas the CUTO32 cell line was >10-fold more resistant to three RET inhibitors. We discovered that our RET+ cell lines had differential regulation of the mitogen-activated protein kinase and phosphoinositide 3-kinase/protein kinase B (AKT) pathways. After inhibition of RET, the CUTO42 cells had robust inhibition of phosphorylated AKT (pAKT), whereas CUTO22 and CUTO32 cells had sustained AKT activation. Next, we performed a drug screen, which revealed that the CUTO32 cells were sensitive (<1 nM IC50) to inhibition of two cell cycle-regulating proteins, polo-like kinase 1 and Aurora kinase A. Finally, we show that two of these cell lines, CUTO32 and CUTO42, successfully establish xenografted tumors in nude mice. We demonstrated that the RET inhibitor BLU-667 was effective at inhibiting tumor growth in CUTO42 tumors but had a much less profound effect in CUTO32 tumors, consistent with our in vitro experiments. These data highlight the utility of new RET+ models to elucidate differences in response to tyrosine kinase inhibitors and downstream signaling regulation. Our RET+ cell lines effectively recapitulate the interpatient heterogeneity observed in response to RET inhibitors and reveal opportunities for alternative or combination therapies. SIGNIFICANCE STATEMENT: We have derived and characterized three novel rearranged during transfection (RET) fusion non-small cell lung cancer cell lines and demonstrated that they have differential responses to RET inhibition as well as regulation of downstream signaling, an area that has previously been limited by a lack of diverse cell line modes with endogenous RET fusions. These data offer important insight into regulation of response to RET tyrosine kinase inhibitors and other potential therapeutic targets.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-ret/antagonistas & inibidores , Transdução de Sinais , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Proteínas Proto-Oncogênicas c-ret/genética , Proteínas Recombinantes de Fusão/efeitos dos fármacos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
6.
BMC Genomics ; 22(1): 795, 2021 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-34740326

RESUMO

BACKGROUND: Pathogens of the genus Phytophthora are the etiological agents of many devastating diseases in several high-value crops and forestry species such as potato, tomato, cocoa, and oak, among many others. Phytophthora betacei is a recently described species that causes late blight almost exclusively in tree tomatoes, and it is closely related to Phytophthora infestans that causes the disease in potato crops and other Solanaceae. This study reports the assembly and annotation of the genomes of P. betacei P8084, the first of its species, and P. infestans RC1-10, a Colombian strain from the EC-1 lineage, using long-read SMRT sequencing technology. RESULTS: Our results show that P. betacei has the largest sequenced genome size of the Phytophthora genus so far with 270 Mb. A moderate transposable element invasion and a whole genome duplication likely explain its genome size expansion when compared to P. infestans, whereas P. infestans RC1-10 has expanded its genome under the activity of transposable elements. The high diversity and abundance (in terms of copy number) of classified and unclassified transposable elements in P. infestans RC1-10 relative to P. betacei bears testimony of the power of long-read technologies to discover novel repetitive elements in the genomes of organisms. Our data also provides support for the phylogenetic placement of P. betacei as a standalone species and as a sister group of P. infestans. Finally, we found no evidence to support the idea that the genome of P. betacei P8084 follows the same gene-dense/gense-sparse architecture proposed for P. infestans and other filamentous plant pathogens. CONCLUSIONS: This study provides the first genome-wide picture of P. betacei and expands the genomic resources available for P. infestans. This is a contribution towards the understanding of the genome biology and evolutionary history of Phytophthora species belonging to the subclade 1c.


Assuntos
Phytophthora infestans , Solanum tuberosum , Elementos de DNA Transponíveis , Evolução Molecular , Duplicação Gênica , Filogenia , Phytophthora infestans/genética , Doenças das Plantas , Solanum tuberosum/genética
7.
BMC Microbiol ; 21(1): 14, 2021 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-33407123

RESUMO

BACKGROUND: The type VI protein secretion system (T6SS) is important in diverse cellular processes in Gram-negative bacteria, including interactions with other bacteria and with eukaryotic hosts. In this study we analyze the evolution of the T6SS in the genus Xanthomonas and evaluate its importance of the T6SS for virulence and in vitro motility in Xanthomonas phaseoli pv. manihotis (Xpm), the causal agent of bacterial blight in cassava (Manihot esculenta). We delineate the organization of the T6SS gene clusters in Xanthomonas and then characterize proteins of this secretion system in Xpm strain CIO151. RESULTS: We describe the presence of three different clusters in the genus Xanthomonas that vary in their organization and degree of synteny between species. Using a gene knockout strategy, we also found that vgrG and hcp are required for maximal aggressiveness of Xpm on cassava plants while clpV is important for both motility and maximal aggressiveness. CONCLUSION: We characterized the T6SS in 15 different strains in Xanthomonas and our phylogenetic analyses suggest that the T6SS might have been acquired by a very ancient event of horizontal gene transfer and maintained through evolution, hinting at their importance for the adaptation of Xanthomonas to their hosts. Finally, we demonstrated that the T6SS of Xpm is functional, and significantly contributes to virulence and motility. This is the first experimental study that demonstrates the role of the T6SS in the Xpm-cassava interaction and the T6SS organization in the genus Xanthomonas.


Assuntos
Biologia Computacional/métodos , Sistemas de Secreção Tipo VI/genética , Xanthomonas/patogenicidade , Técnicas de Inativação de Genes , Transferência Genética Horizontal , Mutação , Filogenia , Análise de Sequência de DNA , Virulência , Xanthomonas/classificação , Xanthomonas/genética , Xanthomonas/fisiologia
8.
Proc Natl Acad Sci U S A ; 114(5): 921-925, 2017 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-28096383

RESUMO

Smallholder agricultural systems, strongly dependent on water resources and investments in shared infrastructure, make a significant contribution to food security in developing countries. These communities are being increasingly integrated into the global economy and are exposed to new global climate-related risks that may affect their willingness to cooperate in community-level collective action problems. We performed field experiments on public goods with private and collective risks in 118 small-scale rice-producing communities in four countries. Our results indicate that increasing the integration of those communities with the broader economic system is associated with lower investments in public goods when facing collective risks. These findings indicate that local public good provision may be negatively affected by collective risks, especially in communities more integrated with the market economy.


Assuntos
Agricultura , Participação da Comunidade , Abastecimento de Alimentos , Adulto , China , Mudança Climática , Colômbia , Humanos , Nepal , Oryza , Risco , Tailândia
10.
Phytopathology ; 109(5): 859-869, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30908143

RESUMO

Diverse molecular markers have been used to analyze the genetic diversity of plant pathogens. Compared with traditional fingerprinting methods, multiple loci variable number of tandem repeat analyses (MLVAs) have gained importance recently due to their reproducibility, high discriminatory power, ease of performance, low cost, and throughput potential. These characteristics are desirable for continuous pathogen monitoring, especially for pathogens with relatively low genetic diversity, and for disease epidemiology studies. Genetic diversity studies of Xanthomonas phaseoli pv. manihotis, which is the causal agent of cassava bacterial blight, have shown variability and changes in the bacterial population over time. Thus, an easy and fast method needs to be developed to type populations of this pathogen in different countries of the world, especially on small scales. In this study, we developed an MLVA scheme to analyze X. phaseoli pv. manihotis variability on a local scale. The MLVA-15 scheme comprises 15 variable number of tandem repeat loci grouped into four multiplex polymerase chain reaction pools. We showed that the MLVA-15 scheme had slightly higher discriminatory ability at the locality level when compared with amplified fragment length polymorphisms. The MLVA-15 scheme allowed for an accurate determination of the number of genotypes in the sample and showed reproducibility and portability. Additionally, this scheme could be used to analyze numerous strains in a reasonable timeframe. The MLVA-15 scheme was highly specific to X. phaseoli but up to eight tandem repeat loci could be amplified from other Xanthomonas spp. Finally, we assessed the utility of the scheme for analyses of X. phaseoli pv. manihotis genetic variability in the Colombian Caribbean region. MLVA-15 distinguished 88.9% of the haplotypes in our sample. Strains originating from the same field and isolated at the same time could be discriminated. In this study, the advantages of the MLVA-15 scheme targeting 6- or 7-bp repeats were demonstrated. Moreover, this scheme was a fast method that was appropriate for routine monitoring of X. phaseoli pv. manihotis populations on a local scale and, thus, was useful for addressing epidemiological questions.


Assuntos
Genética Populacional , Repetições de Microssatélites , Xanthomonas/genética , Região do Caribe , Colômbia , Doenças das Plantas/microbiologia , Reprodutibilidade dos Testes
12.
J Zoo Wildl Med ; 48(2): 453-460, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28749294

RESUMO

Osteosarcomas are unusual neoplasms in African hedgehogs ( Atelerix albiventris ) and have been reported in extraskeletal and skeletal locations, including mandible, ribs, and vertebra. Five hedgehogs with osteosarcoma submitted to the Pathology Department at Facultad de Medicina Veterinaria y Zootecnia, National Autonomous University of Mexico are reported. In two cases, the neoplasm arose from the skull; one case arose from the ribs with associated compression of the thoracic and abdominal cavity, and another case involved the vertebrae. In the last case, the neoplasm arose from the scapula. Histologic lesions were similar in all cases and consisted of well-demarcated nodules in which neoplastic cells were arranged in sheets of polyhedral to spindle-shaped cells with interspersed areas of necrosis. Numerous trabeculae of osteoid were present throughout the tumors. No metastases were detected. The predominant histologic pattern was osteoblastic, but a telangiectatic-like pattern was observed in the vertebral osteosarcoma. Electron microscopy was performed in two cases, and malignant osteoblasts had features consistent with descriptions in other species, including deposits of hydroxyapatite in osteoid. According to these cases and previously published data, axial osteosarcomas are more frequent in contrast to appendicular osteosarcomas in African hedgehogs, and metastases are rare.


Assuntos
Neoplasias Ósseas/veterinária , Ouriços , Osteossarcoma/veterinária , Animais , Neoplasias Ósseas/patologia , Feminino , Masculino , Osteossarcoma/patologia
13.
BMC Genomics ; 16: 190, 2015 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-25887443

RESUMO

BACKGROUND: Cassava, Manihot esculenta Crantz, is one of the most important crops world-wide representing the staple security for more than one billion of people. The development of dense genetic and physical maps, as the basis for implementing genetic and molecular approaches to accelerate the rate of genetic gains in breeding program represents a significant challenge. A reference genome sequence for cassava has been made recently available and community efforts are underway for improving its quality. Cassava is threatened by several pathogens, but the mechanisms of defense are far from being understood. Besides, there has been a lack of information about the number of genes related to immunity as well as their distribution and genomic organization in the cassava genome. RESULTS: A high dense genetic map of cassava containing 2,141 SNPs has been constructed. Eighteen linkage groups were resolved with an overall size of 2,571 cM and an average distance of 1.26 cM between markers. More than half of mapped SNPs (57.4%) are located in coding sequences. Physical mapping of scaffolds of cassava whole genome sequence draft using the mapped markers as anchors resulted in the orientation of 687 scaffolds covering 45.6% of the genome. One hundred eighty nine new scaffolds are anchored to the genetic cassava map leading to an extension of the present cassava physical map with 30.7 Mb. Comparative analysis using anchor markers showed strong co-linearity to previously reported cassava genetic and physical maps. In silico based searching for conserved domains allowed the annotation of a repertory of 1,061 cassava genes coding for immunity-related proteins (IRPs). Based on physical map of the corresponding sequencing scaffolds, unambiguous genetic localization was possible for 569 IRPs. CONCLUSIONS: This is the first study reported so far of an integrated high density genetic map using SNPs with integrated genetic and physical localization of newly annotated immunity related genes in cassava. These data build a solid basis for future studies to map and associate markers with single loci or quantitative trait loci for agronomical important traits. The enrichment of the physical map with novel scaffolds is in line with the efforts of the cassava genome sequencing consortium.


Assuntos
Mapeamento Cromossômico , Genoma de Planta , Manihot/genética , Imunidade Vegetal/genética , Análise por Conglomerados , Ligação Genética , Marcadores Genéticos , Genótipo , Proteínas de Plantas/genética , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Análise de Sequência de DNA
14.
FASEB J ; 28(1): 195-205, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24036883

RESUMO

Amyloid precursor protein (APP), encoded on Hsa21, functions as a cell adhesion molecule (CAM) in axonal growth cones (GCs) of the developing brain. We show here that axonal GCs of human fetal Down syndrome (DS) neurons (and of a DS mouse model) overexpress APP protein relative to euploid controls. We investigated whether DS neurons generate an abnormal, APP-dependent GC phenotype in vitro. On laminin, which binds APP and ß1 integrins (Itgb1), DS neurons formed enlarged and faster-advancing GCs compared to controls. On peptide matrices that bind APP only, but not on those binding exclusively Itgb1 or L1CAM, DS GCs were significantly enlarged (2.0-fold), formed increased close adhesions (1.8-fold), and advanced faster (1.4-fold). In assays involving alternating stripes of monospecific matrices, human control GCs exhibited no preference for any of the substrates, whereas DS GCs preferred the APP-binding matrix (cross-over decreased significantly from 48.2 to 27.2%). Reducing APP expression in DS GCs with siRNA normalized most measures of the phenotype, including substrate choice. These experiments show that human DS neurons exhibit an APP-dependent, abnormal GC phenotype characterized by increased adhesion and altered contact guidance. The results suggest that APP overexpression may perturb axonal pathfinding and circuit formation in developing DS brain.


Assuntos
Precursor de Proteína beta-Amiloide/metabolismo , Síndrome de Down/metabolismo , Animais , Encéfalo/metabolismo , Moléculas de Adesão Celular/metabolismo , Cones de Crescimento/metabolismo , Hipocampo/metabolismo , Humanos , Técnicas In Vitro , Laminina/metabolismo , Camundongos
15.
Proc Natl Acad Sci U S A ; 109(28): E1972-9, 2012 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-22699502

RESUMO

Cassava bacterial blight (CBB), incited by Xanthomonas axonopodis pv. manihotis (Xam), is the most important bacterial disease of cassava, a staple food source for millions of people in developing countries. Here we present a widely applicable strategy for elucidating the virulence components of a pathogen population. We report Illumina-based draft genomes for 65 Xam strains and deduce the phylogenetic relatedness of Xam across the areas where cassava is grown. Using an extensive database of effector proteins from animal and plant pathogens, we identify the effector repertoire for each sequenced strain and use a comparative sequence analysis to deduce the least polymorphic of the conserved effectors. These highly conserved effectors have been maintained over 11 countries, three continents, and 70 y of evolution and as such represent ideal targets for developing resistance strategies.


Assuntos
Manihot/metabolismo , Manihot/microbiologia , Doenças das Plantas/microbiologia , Análise de Sequência de DNA/métodos , Xanthomonas axonopodis/metabolismo , Área Sob a Curva , Progressão da Doença , Genoma Bacteriano , Genômica , Geografia , Imunidade Inata , Modelos Genéticos , Dados de Sequência Molecular , Filogenia , Doenças das Plantas/genética , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único , Fatores de Tempo
16.
BMC Microbiol ; 14: 161, 2014 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-24946775

RESUMO

BACKGROUND: Molecular typing of pathogen populations is an important tool for the development of effective strategies for disease control. Diverse molecular markers have been used to characterize populations of Xanthomonas axonopodis pv. manihotis (Xam), the main bacterial pathogen of cassava. Recently, diversity and population dynamics of Xam in the Colombian Caribbean coast were estimated using AFLPs, where populations were found to be dynamic, diverse and with haplotypes unstable across time. Aiming to examine the current state of pathogen populations located in the Colombian Eastern Plains, we also used AFLP markers and we evaluated the usefulness of Variable Number Tandem Repeats (VNTRs) as new molecular markers for the study of Xam populations. RESULTS: The population analyses showed that AFLP and VNTR provide a detailed and congruent description of Xam populations from the Colombian Eastern Plains. These two typing strategies clearly separated strains from the Colombian Eastern Plains into distinct populations probably because of geographical distance. Although the majority of analyses were congruent between typing markers, fewer VNTRs were needed to detect a higher number of genetic populations of the pathogen as well as a higher genetic flow among sampled locations than those detected by AFLPs. CONCLUSIONS: This study shows the advantages of VNTRs over AFLPs in the surveillance of pathogen populations and suggests the implementation of VNTRs in studies that involve large numbers of Xam isolates in order to obtain a more detailed overview of the pathogen to improve the strategies for disease control.


Assuntos
Variação Genética , Manihot/microbiologia , Tipagem Molecular/métodos , Doenças das Plantas/microbiologia , Xanthomonas axonopodis/classificação , Xanthomonas axonopodis/genética , Análise do Polimorfismo de Comprimento de Fragmentos Amplificados , Análise por Conglomerados , Colômbia , DNA Bacteriano/química , DNA Bacteriano/genética , Genótipo , Repetições Minissatélites , Dados de Sequência Molecular , Filogenia , Análise de Sequência de DNA , Xanthomonas axonopodis/isolamento & purificação
17.
Microb Ecol ; 68(1): 155-67, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24760168

RESUMO

Cassava bacterial blight, caused by Xanthomonas axonopodis pv. manihotis (Xam), is the most important bacterial disease affecting this crop. A continuous surveillance of the pathogen population dynamics is required to develop an efficient disease management program. During the 1990s, Xam populations showed high levels of genetic variation and relevant migratory processes that were important determinants of the distribution of the pathogen diversity in Colombia. Aiming to characterize the current population structure of the pathogen and the evolutionary forces that shape these populations, sampling collections were carried out from September 2008 until November 2010 in the Colombian Caribbean Region. One hundred and sixty bacterial isolates were characterized using amplified fragment length polymorphism (AFLP) markers. Additionally, a subset of effector genes were sequenced in some isolates to determine their usefulness in Xam population studies and to provide additional information to that obtained with AFLPs. Virulence patterns of ten isolates were determined in nine cassava accessions. Our results show a complex architecture of population and confirm migratory process previously reported in the Caribbean Region. Chinú, one of the locations sampled, presented remarkable features in population dynamics such as longer genetic distances, higher diversity indices, and a genetically differentiated population when it was compared with other locations. Virulence tests showed that MCOL2215, one of the most cultivated cassava varieties in the Caribbean coast, was susceptible to the majority of Xam isolates tested. This study shows the current condition of populations of Xam in the Caribbean Region of Colombia, and it contributes to improve the existing bacterial blight control practices.


Assuntos
Genética Populacional , Manihot/microbiologia , Doenças das Plantas/microbiologia , Xanthomonas axonopodis/genética , Análise do Polimorfismo de Comprimento de Fragmentos Amplificados , Colômbia , DNA Bacteriano/genética , Evolução Molecular , Genes Bacterianos , Marcadores Genéticos , Haplótipos , Polimorfismo Genético , Seleção Genética , Virulência/genética , Xanthomonas axonopodis/patogenicidade
18.
Plant Cell Rep ; 33(11): 1901-12, 2014 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25120000

RESUMO

KEY MESSAGE: An RNAseq-based analysis of the cassava plants inoculated with Xam allowed the identification of transcriptional upregulation of genes involved in jasmonate metabolism, phenylpropanoid biosynthesis and putative targets for a TALE. Cassava bacterial blight, a disease caused by the gram-negative bacterium Xanthomonas axonopodis pv. manihotis (Xam), is a major limitation to cassava production worldwide and especially in developing countries. The molecular mechanisms underlying cassava susceptibility to Xam are currently unknown. To identify host genes and pathways leading to plant susceptibility, we analyzed the transcriptomic responses occurring in cassava plants challenged with either the non-pathogenic Xam strain ORST4, or strain ORST4(TALE1 Xam ) which is pathogenic due to the major virulence transcription activator like effector TALE1 Xam . Both strains triggered similar responses, i.e., induction of genes related to photosynthesis and phenylpropanoid biosynthesis, and repression of genes related to jasmonic acid signaling. Finally, to search for TALE1 Xam virulence targets, we scanned the list of cassava genes induced upon inoculation of ORST4(TALE1 Xam ) for candidates harboring a predicted TALE1 Xam effector binding element in their promoter. Among the six genes identified as potential candidate targets of TALE1 Xam a gene coding for a heat shock transcription factor stands out as the best candidate based on their induction in presence of TALE1 Xam and contain a sequence putatively recognized by TALE1 Xam .


Assuntos
Perfilação da Expressão Gênica , Manihot/genética , Doenças das Plantas/genética , Xanthomonas axonopodis/crescimento & desenvolvimento , Álcoois Benzílicos/metabolismo , Análise por Conglomerados , Genes de Plantas/genética , Interações Hospedeiro-Patógeno , Manihot/microbiologia , Análise de Sequência com Séries de Oligonucleotídeos , Fotossíntese/genética , Doenças das Plantas/microbiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Análise de Componente Principal , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Virulência , Xanthomonas axonopodis/patogenicidade , Xanthomonas axonopodis/fisiologia
19.
Trop Anim Health Prod ; 46(3): 555-61, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24488621

RESUMO

A wide spectrum of welfare issues is encountered on working equids. The objectives of this study were: (i) to determine the prevalence of cutaneous lesions in working equids in tropical regions of the North-Central of Veracruz State, Mexico, (ii) to identify the most prevalent cutaneous pathological conditions in this population of working equids, and (iii) to identify risk factors associated to the presence of work-related skin wounds. A number of 467 working equids presented for treatment at Donkey Sanctuary Mobile Clinic Program - National Autonomous University of Mexico from seven different villages in the North-Central region of Veracruz State were evaluated. Animals with signs of cutaneous pathology underwent a physical examination and samples were obtained of exudates, hair, and tissue for cytology examination, fungal cultures, and dermo-histopathology examination. The overall prevalence of cutaneous pathological conditions in working equids was 20.6 %. The prevalence per species was 22.6 % for horses, 18.2 % for donkeys, and 14.3 % for mules. The most common cause of skin lesions was trauma (abrasions, lacerations, and abscesses) followed in frequency by parasitic dermatitis and sarcoids. No strong associations among species were observed (P > 0.20). Multivariate analysis showed that there was greater association between BCS and age with the presence of skin wounds (P < 0.05). Equids who's BCS was rated as "moderate" as well as adult animals showed significantly more skin wounds compared with the young and the elderly (P < 0.05). These findings can help to develop a better management practices to improve the welfare of equids in tropical regions of Veracruz, Mexico.


Assuntos
Bem-Estar do Animal , Equidae , Dermatopatias/veterinária , Ferimentos e Lesões/veterinária , Criação de Animais Domésticos , Animais , Estudos Transversais , Coleta de Dados , Feminino , Masculino , México/epidemiologia , Prevalência , Fatores de Risco , Dermatopatias/epidemiologia , Dermatopatias/patologia , Inquéritos e Questionários , Ferimentos e Lesões/epidemiologia
20.
J Zoo Wildl Med ; 45(3): 682-5, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25314843

RESUMO

A sebaceous carcinoma was diagnosed, together with a mammary carcinoma, in an adult African hedgehog (Atelerix albiventris). The first neoplasm was located in the subcutaneous tissue of the neck and extended towards the axillary area of the chest. The second was located in the subcutaneous left caudal abdominal region. The purpose of this paper is to report the histopathologic and ultrastructural features of these neoplasms. Although there is little information about diseases affecting this species, it is known that neoplastic disorders are fairly common in African hedgehogs. The mammary carcinoma is considered to be the most common neoplasm in these animals; however, the presentation of sebaceous carcinoma is rare. In hedgehogs, the simultaneous presence of two neoplasms is common, which is why special attention should be paid to the presentation of other tumors during the early detection of a neoplastic process as this will greatly facilitate the optimal treatment and improve the long-term prognosis of affected animals.


Assuntos
Carcinoma/veterinária , Ouriços , Neoplasias Mamárias Animais/patologia , Neoplasias das Glândulas Sebáceas/veterinária , Animais , Carcinoma/patologia , Feminino , Neoplasias das Glândulas Sebáceas/patologia
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