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Item Caracterização da produção científica sobre DNA barcoding no estudo de plantas(Universidade Federal de Goiás, 2021-04-22) Batista, Eliane Cotrim; Gonçalves, Ariany Rosa; http://lattes.cnpq.br/6401185899301616; Telles, Mariana Pires de Campos; http://lattes.cnpq.br/4648436798023532 ; Telles, Mariana Pires de Campos; Nunes, Rhewter; Braga, Ramila dos SantosThe application of the DNA barcoding has been shown to be successful in studies of biodiversity surveys, biomonitoring of invasive species, forensic analysis to assess food security and medicinal products based on medicinal plants, monitoring the illegal trade in threatened species, studies about food ecology, conservation initiatives, etc. The present study had as main objective to raise and systematize the scientific production on the use of DNA barcoding in plants. For this purpose, a search was carried out in the Web of Science database, using as keywords combined the regions annotated in the chloroplast and nuclear genome of Arabidopsis thaliana (model species), which are used as DNA barcoding markers, associated with the expressions: “DNA barcoding” OR “barcoding” OR “DNA barcode” OR “barcode” OR “metabarcoding”. The search was carried out covering the complete period of publication of the database until 2020. The recovered files were filtered, being considered only original articles, excluding review articles, as well as those that did not correspond to the scope of the study. A total of 1,897 articles were retrieved, of which 1,073 were within the criteria’s inclusion. As the first article was published in 2005, the analysis covered the period between 2005 and 2020. The studies were published in a total of 309 sources, with a general average of publication per year equal to 5.21. Scientific production varied over the period, but always with an increasing trend. The average number of authors per document was 3.49, since each article mostly has more than one author, which explains the average number of coauthors per document (5.34). However, out of a total of 3,747 registered authors, the number of articles per author was low, with an average equal to 0.29. China stands out as the main country, according to the origin of the authors, who has contributed with works involving the application of the DNA barcoding technique in the study of plants, being also the most cited country. The most relevant authors, based on the number of publications, were the Chinese: Chen SL, Song J, Yao H, with the article by Chen et al. (2010) the second most cited globally. The journals that showed greater prominence in relation to the number of articles published, considering the subject, were respectively: Plos One, Molecular Ecology Resources and Scientific Reports. It was possible to characterize four groups of documents regarding the use / application of the technique, respectively: Description/delimitation of species (545), Forensic Genetics (275), Development/methodology (163), Ecology (90) and 75 articles were categorized in up to two different groups. The studies grouped in the most prominent category (Description/delimitation of species), range from delimiting species from complex groups to the validation of species with properties of economic interest and species already cataloged in herbariums. The application possibilities have been expanded due to the development of High Throughput Sequencing technologies, opening doors for studies applied to Ecology, Evolution and Forensic Genetics and which use in addition to DNA barcoding, the metabarcoding.Item Identificação proteômica de alterações metabólicas em Paracoccidioides brasiliensis induzidas por derivado de chalcona(Universidade Federal de Goiás, 2021-09-30) Carvalho Júnior, Marcos Antonio Batista de; Silva, Kleber Santiago Freitas e; http://lattes.cnpq.br/3813868830071259; Pereira, Maristela; http://lattes.cnpq.br/1345781867765758; Pereira, Maristela; Curcio, Juliana Santana de; Soares, Célia Maria de AlmeidaParacoccidioidomycosis is one of the most important systemic mycoses in Latin America. The infection occurs through the inhalation of fungal propagules belonging to Paracoccidioides genus that are present in soils, being largely associated with the contamination of rural workers, who are constantly exposed to this potentially contaminated material. Even more than a century after its discovery, the treatment of this infection with the currently available antifungal arsenal still represents a challenge, due to the long treatment time required, as well as the high toxicity of the drugs used. These questions highlight the need for research to develop and characterize new compounds with the potential to inhibit the growth of these microorganisms. In this sense, a class of molecules called chalcones has shown great versatility for demonstrating broad biological properties, including antifungal activity among them. Through a virtual screening methodology, a chalcone derivative named compound 3 was identified by our group as a promising inhibitor of Paracoccidioides spp. Thus, in order to understand the mode of action of compound 3, we applied a proteomic approach to identify the induced and repressed proteins of P. brasiliensis in the presence of compound 3. In addition, validation assays were performed on the results found. The analysis indicated that the compound can cause an imbalance in the fungal energy homeostasis by reducing the activity of the glycolytic pathway, beta-oxidation and citric acid cycle. In vitro validations also demonstrated that reactive oxygen species accumulate within the cells during exposure to compound 3, which can destabilize cellular components such as the plasma membrane. The molecular docking assay between compound 3 and the enzyme dihydropteroate synthase, which had its expression induced after the treatment, suggest that compound 3 may act as an inhibitor of this protein, impairing the folate biosynthesis that participates as a cofactor in synthesis of nucleotides and some amino acids. Therefore, these data support the efficiency of compound 3 in producing imbalances in key pathways for the P. brasiliensis' metabolic maintenance, contributing to its antifungal role.Item Desenvolvimento e caracterização de nanopartículas mistas de quitosana e lecitina contendo melatonina para o tratamento de feridas em ratos diabéticos(Universidade Federal de Goiás, 2020-08-19) Corrêa, Viviane Lopes Rocha; Leite, Liliana Borges de Menezes; http://lattes.cnpq.br/2012543423092393; Amaral, André Corrêa; http://lattes.cnpq.br/8801299423520104; Amaral, André Corrêa; Bocca, Anamélia Lorenzetti; Tedesco, Antônio Cláudio; Diniz, Danielle Guimarães Almeida; Miguel, Marina PachecoWound healing in diabetic patients remains a worldwide problem that can cause amputations and even lead to death. This work aimed to produce melatonin-loaded lecithin-chitosan nanoparticles (MEL-NP) and to evaluate a topical formulation containing these particles for healing in an in vivo animal model for diabetes. For the production of nanoparticles, an ethanolic solution containing soybean lecithin and melatonin was dropwise added to an aqueous solution of chitosan under sonication. The nanoparticles were physical-chemical characterized and evaluated in vivo for toxicity using the Galleria mellonella model and its potential for wound healing in diabetic rats. The MEL-NP were around 160 nm in size and had a zeta potential around 25 mV. The melatonin entrapment efficiency was 27%. Our results demonstrated that treatment with MEL-NP improved wound healing demonstrated by a wound closure earlier than the other treatments evaluated. An appreciated therapeutic effect was achieved by MEL-NP in the induction of fibroblasts and angiogenic proliferation. In addition, it was accompanied by an expressive collagen deposition. Taking the observed data, the MEL-NP produced could be used in the development of promising strategies for wound healing in diabetic people.Item Genoma cloroplastidial de Serjania erecta Raldk: variação no número de genes e análise de seleção de genes de plastomas da família Sapindaceae(Universidade Federal de Goiás, 2022-03-04) Corvalan, Leonardo Carlos Jeronimo; Diniz Filho, José Alexandre Felizola; http://lattes.cnpq.br/0706396442417351; Nunes, Rhewter; http://lattes.cnpq.br/6169806655018346; Nunes, Rhewter; Sobreiro, Mariane Brom; Dias, Renata de OliveiraSerjania erecta from the Sapindaceae family is a plant with medicinal properties. Studies indicate potential for use in the treatment of Alzheimer's disease, gastric diseases and anti-inflammatory use. Although, little is known about its genetic and evolutionary aspects. The goal of this study was to assemble the chloroplast genome of S. erecta, and use it in a comparative analysis with other plastomas of the Sapindaceae family. For this, we sequenced a single specimen of S. erecta from Araxá (MG - Brasil) using Illumina Miseq. The chloroplast genome was assembled using NOVOPlasty v3.2 and annotated using the CHLOROBOX platform. For comparative analysis was used eleven chloroplast genomes from different Sapindaceae family species (Acer buergerianum, Aesculus wangii, Dimocarpus longan, Dipteronia dyeriana, Dodonaea v iscosa, Eurycorymbus cavaleriei, Koelreuteria paniculata, Litchi chinensis, Pometia tomentosa, Sapindus mukorossi, Xanthoceras sorbifolium). The chloroplast genome of S. erecta has a size of 159,297 bp with 132 genes, including 87 are protein-coding genes, 37 are tRNAs and 8 rRNAs. Among twelve chloroplast genomes avalieded, S. erecta has the lowest amount of complex repeats and microsatellites. The structure and order of genes in chloroplast genomes of the order Sapindales was extremely conserved. The variation in numbers of genes was from the 132 genes to 128 genes in the Sapindaceae family. We suggest that three factors cause variation in the number of genes in the family: (1) Inverted repeat region (IR) expansion events cause the duplication of the rpl22, rps3 and rps19 genes; (2) the pseudogenization of the rps2 gene; (3) variation in the number of genes encoding tRNAS. The phylogenetic tree had well supported nodes within Sapindaceae and Serjania formed a clade with Sapindus, Litchi, Dimocarpus, and Pometia genera. Only two genomic regions (ycf1 and ndhF) showed high nucleotide diversity and no one gene is under positive selection (ka/ks > 1). The results obtained in this study provide the assembly and annotation of the chloroplast genome of S. erecta, the first annotation of a species of the genus. It also provides an idea of how chloroplast genomes evolved in the Sapindaceae family.Item Investigação genômica em pacientes com deficiência intelectual ou atraso global do desenvolvimento assistidos na rede pública de saúde do estado de Goiás(Universidade Federal de Goiás, 2022-02-21) D’Ávila, Ana Julia da Cunha Leite; Minasi, Lysa Bernardes; http://lattes.cnpq.br/9057708164796074; Cruz, Aparecido Divino da; http://lattes.cnpq.br/7868817504129985; Cruz, Aparecido Divino da; Silva, Claudio Carlos da; Gigonzac, Thais Cidália Vieira; Costa, Emilia Oliveira Alves; Silva, Daniela de Melo eIntellectual Disability (ID) is a complex and heterogeneous clinical condition that affects about 1 to 3% of children and adolescents worldwide, and that can be caused by a variety of environmental and/or genetic factors. There are a variety of human genetic variants already identified for ID, from small variants to larger structural variants that affect thousands to millions of nucleotides. GTG banding karyotyping and microarray chromosome analysis (CMA) are often used to identify the genetic causes of ID, especially in cases Where the clinical evaluation indicates the syndromic form of ID. The use of next generation sequencing technologies for the diagnosis of ID has made it possible to expand the detection of new mutations and new genes associated with ID. The aim of this study was to perform total exome sequencing with gene panel analysis for intellectual disability, in patients who presented karyotype without structural and/or numerical alterations and did not present pathogenic copy number variations (CNVS) in the CMA. A retrospective cohort of 369 patients of both sexes with clinical indications of DI and/or global developmental delay was evaluated in the study. Peripheral blood samples were collected to perform cytogenetic analysis and gene sequencing. GTG banding was performed for all patients following the standard protocol. CMA was performed for patients who did not present structural and/or numerical alterations in the karyotype. Genomic DNA was isolated from patients and their biological parents in order to determine the origin of the CNVs found. Cases that were not diagnosed after performing the karyotype and CMA were referred for exome sequencing with gene panel analysis for intellectual disability, which encompassed 1,252 genes. With the karyotype, it was possible to identify chromosomal alterations in 34.7% (128/369) of the patients. CMA was performed in 83 patients who had results with no alterations in karyotype (46,XX or 46,XY) with a diagnosis rate of 21.7% (18/83). Exome sequencing for target genes was performed in 19 trios of families that had negative results in previous methodologies. Gene panel analysis of exome data identified mutations in 63.1% (12/19) of cases, of which 75% (9/12) were pathogenic variants, 8.3% (1/12) probably pathogenic and in 16.7% (2/2) VUS. Of the total variants identified, 75% (9/12) were de novo mutations and 25% (3/12) had mutations inherited from healthy parents. The most prevalent variants were missense-type mutations (66.7%) followed by nonsense-type mutations 16.7%), frameshift (8.3%) and loss CNV (8.3%). With the three methodologies applied, it was possible to identify the genetic causes of ID in 42.3% (156/369) of the patients. In conclusion, our results show different methodologies that can be used for the genetic diagnosis of ID and that exome sequencing with gene panel analysis provides an efficient diagnostic strategy when combined with clinical and laboratory screening.Item Mesma informação, novas aplicações: revisitando primers do gene COI em aves e melhorando a identificação por DNA barcoding(Universidade Federal de Goiás, 2021-03-31) Melo, Amanda Alves de; Nunes, Rhewter; http://lattes.cnpq.br/6169806655018346; Telles, Mariana Pires de Campos; http://lattes.cnpq.br/4648436798023532; Telles, Mariana Pires de Campos; Braga, Ramilla dos Santos; Targueta, Cíntia PelegrinetiMolecular identification of species occurs through the comparison of either morphological, biochemical or genetic characteristics. All of these areas of knowledge for delimiting species are connected and should be analyzed as a complement to fill gaps of information of each area. The process of molecular identification of species through comparison of genetic information named DNA barcode consists of analyzing nucleotide sequences of a certain region to compare the differences between species. On the majority of the animals, including birds, the cytochrome c oxidase I (COI) gene is widely used for this purpose and in most cases, it shows the ideal greater variation in its sequence among species rather than within individuals of the same species, providing elucidative information about species differentiation. Choosing primers for a DNA barcode application is a crucial step because it depends on the aim of the study and on the taxonomic group of interest. Primers in silico analysis can evaluate its physicochemical properties, the number of species reached by them and then direct the choice of the best primers for the different research objectives, saving time and money resources. Therefore, this present work aimed to answer questions related to the efficiency of the available primers for the avian COI gene and to the avian mitochondrial genome sequences available in databases: i) What is the taxonomic coverage (number of species) reached by the avian COI gene primers? Are the universal primers actually capable of covering all species?; ii) Which avian orders have the most mitochondrial genome sequences available on the databases and what is its representativeness significance within the total number of species for each order?; iii) Which primers show the ideal physicochemical properties to increase the chances of successful amplification in laboratorial experiments? and; iv) Which primer sets are the most suited to guarantee full species recovery within the avian group?Item Caracterização do genoma cloroplastidial de Caryocar cuneatum (Caryocaraceae)(Universidade Federal de Goiás, 2023-04-28) Mendes, Millena Silva; Nunes, Rhewter; Telles, Mariana Pires de Campos; http://lattes.cnpq.br/4648436798023532; Telles, Mariana Pires de Campos; Pinto, Rafael Barbosa; Brito, Cintia Pelegrineti Targueta de AzevedoThe Caryocaraceae family (1845) belongs to the order Malpighiales, which has 36 families, 716 genera and 16,065 species. Caryocaraceae consists of two genera Anthodiscus G. Mey. with ten species and Caryocar L. with 16 species. The present work aims to carry out the de novo assembly of the chloroplastidial genome of Caryocar cuneatum Wittm. and to carry out comparative analyzes regarding the structure and composition of the genome with other species of Malpighiales. For this, the total DNA of an individual of C. cuneatum was sequenced using the MiSeq platform (Illumina), paired-end (2x300) with the MiSeq V3 600 cycles kit. The plastome was assembled using the NOVOPlasty v3.2 program and annotated using the CHLOROBOX. For comparative analyses, chloroplast genomes of 8 species belonging to the Malpighiales order (Caryocar brasiliense, Caryocar glabrum, Lophopyxis maingayi, Drypetes indica, Aspidopterys concava, Byrsonima crassifolia, Balanops balansae, Couepia ovalifolia) were used. The chloroplast genome of C. cuneatum had a size of 165,767 bp, composed of a single copy major region of 83,968 bp, a single copy minor region of 11,854 bp, separated by two inverted repeat regions of 34,973 bp. It has 131 genes, 90 protein coding, 33 transfer RNAs and 8 ribosomal RNAs. The rpl32 gene is not present in species in C. cuneatum and in the other two species of Caryocar. The size of the plastid genome of the genus Caryocar presents itself a lot. For the other species, there was variation in size, structure and genomic composition. The InfA gene is pseudogenized in all analyzed species and was not found in A. concava. In the analysis of synonymous and non-synonymous mutations, the rpl20 gene is under neutrality and the other genes are under negative selection. For the nucleotide diversity in Caryocar genomes, it was possible to identify two hotspot regions: trnS-GCU - trnG-UCC and rbcL - atpB, which can be used as possible molecular markers for DNA barcoding. The phylogenetic tree obtained good support values for nodes in Caryocaraceae, validating the systematic position of C. cuneatum within the family and evidence of its relationship with its sister groups C. brasilense and C. glabrum. In conclusion, this study provides the first genome of C. cuneatum sequenced, information on genomic characterization and the knowledge provided here can be used for further studies and technological.Item Evidência do aumento de desvios mendelianos mediante análise transgeracional de mutações germinativas em população exposta à radiação ionizante de Césio-137(Universidade Federal de Goiás, 2022-08-12) Oliveira, Lorraynne Guimarães; Costa, Emília Oliveira Alves; http://lattes.cnpq.br/2332660855010816; Cruz, Aparecido Divino da; http://lattes.cnpq.br/7868817504129985; Silva, Daniela de Melo e; Silva, Claudio Carlos da; Dias, Renata de Oliveira; Leito Filho, Hugo Pereira; Cruz, Aparecido Divino daOn September 13th, 1987, the largest radiological accident in urban areas occurred in Goiânia, Goiás (Brazil), resulting in human, animal, plant and environmental exposure by Cesium-137. The mutagenic effects on the germline of people exposed to ionizing radiation are of particular concern, as the risk of inherited disorders can be increased. Single Nucleotide Variants are the most common form of human genetic variation and occur in greater abundance in a non-coding region and in recent years, several technologies have been developed to identify these variants. One of these technologies is Next-Generation Sequencing, which offers greater amounts of data, shorter sequencing times and reduced costs. The objective of this work is to establish the spectrum and frequency / rate of base de novo substitutions of germ origin from the trio's new generation sequencing data, corresponding to an F1 generation child and his biological parents, accidentally exposed to high and low doses of IR of cesium-137, contributing to knowledge about the biological effects of exposure to ionizing radiation. Considering case and control, the results of 38 parents, and an offspring of 14 children born from the exposed group and 5 children from the unexposed population were included. Exposed individuals had ~39% increase in global mean DMs compared to controls. Exposed mothers had ~44% increase in global mean DMs compared to controls. The A:T>C:G mutation was the one that showed the greatest statistically significant increase in occurrence in the offspring of exposed individuals. Transition replacement rates were higher than transversions in the offspring of cases and controls, but the difference is not statistically significant. In conclusion, with the methodology and biomarkers used, it was possible to identify the origin of mutation in the parents, as well as the type of substitution and to inform which variant was mutated, it was also possible to detect the frequency of the germline mutation in DM which made it possible to retrospectively study this population exposed to IR.Item Toxicidade de nanopartículas de óxido de ferro (γ-Fe2O3) funcionalizadas com citrato ao longo do desenvolvimento inicial do zebrafish (Danio rerio)(Universidade Federal de Goiás, 2020-01-24) Pereira, Aryelle Canedo; Gonçalves, Bruno Bastos; http://lattes.cnpq.br/3194796958722741; Rocha, Thiago Lopes; http://lattes.cnpq.br/6325937100056775; Rocha, Thiago Lopes; Rodrigues, Gisele Augusto; Franchi, Leonardo PereiraIron oxide nanoparticles (NOFs) are being increasingly used in medical, environmental and technological applications. However, with its growing production and application, concerns arise about its release into the environment and its impact on human and environmental health. In this sense, the zebrafish (Danio rerio) is proving to be an excellent model system for the analysis and classification of nanomaterial toxicity. Thus, the general objective of the present study was to evaluate the toxicity of NOFs (γ-Fe2O3 NPs) functionalized with citrate throughout the initial development of zebrafish, comparing toxicity with iron ions, after static and semi-static exposure. For this purpose, the zebrafish embryos were exposed in static (without medium renewal) and semi-static (medium renewal every 24 h) exposure modes to γ-Fe2O3 NPs functionalized with citrate and iron chloride in different concentrations environmentally relevant iron (0.3, 0.6, 1.25, 2.5, 5.0 and 10 mg L-1) for 144 h, together with the control group maintained in water reconstituted in 24-well plates. The results showed that γ-Fe2O3 NPs were accumulated mainly in the chorion of the embryos and in the digestive system and liver of zebrafish larvae. Γ-Fe2O3 NPs have low toxicity compared to iron ions, indicating that the in vivo toxicity of both forms of iron is mediated by different mechanisms of action. However, NPs caused significant changes after semi-static exposure causing a decrease in heart rate, induced blood accumulation and formation of pericardial edema in zebrafish embryos, indicating its cardiotoxic effect. Regarding the exposure method, it was observed that both forms of iron induced high embryotoxicity under semi-static exposure conditions compared to static exposure, indicating that the toxicity depends on the frequency of exposure. This was the first study on the impact of the exposure condition on the toxicity of γ-Fe2O3 NPs on the development of zebrafish.Item Desenvolvimento e testes in vivo de nanopartículas de quitosana contendo insulina na cicatrização de feridas em ratos diabéticos(Universidade Federal de Goiás, 2019-04-16) Ribeiro, Maycon Carvalho; Menezes, Liliana Borges de; http://lattes.cnpq.br/2012543423092393; Amaral, André Corrêa; http://lattes.cnpq.br/8801299423520104; Menezes, Liliana Borges de; Miguel, Marina Pacheco; Mendes, Elizabeth Pereira; Souza, Taís Andrade Dias de; Silva, Luís Antônio Dantas daChitosan has been studied for its ability to accelerate healing and has been tested in the therapy of difficult-to-heal lesions, such as in diabetic patients. Insulin acts by stimulating the signaling pathway for wound healing. The objective of this work was to produce chitosan nanoparticles containing insulin for the evaluation of cicatrizant activity in diabetic rats. For the formation of the nanoparticles, the ionic gelation method was used. The nanoparticles were analyzed by diameter, potential zeta polydispersity index. The degree of deacetylation of chitosan by potentiometric was determined. For the insulin- associated nanoparticles, the mean diameter was 245.9 ± 25.46 nm and zeta potential of 39.3 ± 4.88 mV and PDI of 0.463 ± 0.01. The mean degree of deacetylation found was 72.95%. The Bradford assay revealed that the nanoparticles incorporated 97.19% ± 2.18 of insulin. To evaluate the healing, 72 Wistar rats were divided in four groups: sepigel (S), sepigel with insulin (SI), empty chitosan nanoparticles (QV) and chitosan nanoparticles containing insulin (IQ). The groups were subdivided into three subgroups (n = 6) according to the histological analysis times of the wound (3rd, 7th and 14th day). The induction of diabetes occurred through the intraperitoneal application of alloxan (120mg / kg). After confirmation of the diabetes state, the animals were anesthetized and the wounds were made with an 8.0 mm punch in the dorsal region. Macroscopic and microscopic analyzes were performed. It was possible to produce chitosan nanoparticles by the ionic gelation method, with desired diameter and zeta potential and polydispersity index. No differences were found in the rate of wound retraction among the four groups. The topical use of empty or insulin-containing chitosan nanoparticles in wound healing in diabetic rats was able to stimulate inflammatory cell proliferation and angiogenesis, followed by wound maturation. Differences in wound healing data from the group treated with insulin-containing nanoparticles and from the group treated with free insulin may be related to the high stability of theItem Análise proteômica do fungo patogênico humano Fonsecaea pedrosoi submetido à temperatura do hospedeiro(Universidade Federal de Goiás, 2021-09-29) Rosa, Gabriela Danelli; Bailão, Alexandre Melo; http://lattes.cnpq.br/5415221996976886; Bailão, Alexandre Melo; Lima, Patricia de Sousa; Rocha, Thiago LopesChromoblastomycosis (CBM) is a chronic subcutaneous mycosis, very common in tropical and subtropical regions and affects many men related to rural activities. Lesions of this disease can appear in five clinical forms and treatments are difficult due to the recalcitrant nature of the disease. The main causative agent of CBM is the fungus Fonsecaea pedrosoi, which is a polymorphic, melanized fungus that presents a certain phenotypic plasticity in face of temperature variations. Thermotolerance is one of the virulence factors of virulence for this pathogen, since it survives the temperature increase, like that of the host. However, no work so far has searched to characterize the adapted cellular processes. Proteins are part of the strategies used in response to a variety of stressors, such as host temperature. In this work we mapped the intracellular proteomic profile of F.pedrosoi cultivated under different temperature conditions (28 and 37 °C) for 24 hours. It was possible to identify by means of Liquid Chromatography coupled to Mass Spectrometry (Nano UPLC-MSᴱ) a total of 486 differentially expressed proteins, 101 of which were up-regulated and 385 down-regulated. It was possible to observe that F. pedrosoi seems to respond to temperature stress by repressing central carbon metabolism pathways, such as glycolysis, tricarboxylic acid cycle (TCA) and glyoxylate cycle, cell wall biosynthesis and melanin and induced amino acid degradation enzymes and antioxidant proteins.Item Variantes polimórficas em genes de detoxificação celular e suas relações com o desenvolvimento da esclerose lateral amiotrófica(Universidade Federal de Goiás, 2020-03-05) Santos, Kamilla de Faria; Santos, Rodrigo da Silva; http://lattes.cnpq.br/4806187026900959; Reis, Angela Adamski da Silva; http://lattes.cnpq.br/3243656364470085; Reis, Angela Adamski da Silva; Bailão, Alexandre Melo; Souza, Guilherme Rocha Lino deAmyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease caused by the degeneration of motor neurons, leading to progressive muscular atrophy. Studies suggest the relationship of the disease with environmental and genetic factors, and the involvement of mechanisms, such as oxidative stress, with neuronal degeneration. However, the body has several ways to promote cellular detoxification against harmful compounds. The Glutathione S-transferase (GST) family consists of multifunctional enzymes that act on cell detoxification and elimination of various substances, including oxidative stress products. Among the human cytosolic classes of the GST family, two isoenzymes are highlighted: GSTM1 and GSTT1, both encoded by genes of the same name, these genes have a total deletion polymorphism that results in the absence of enzymatic activity. Therefore, the objective of the present study was to evaluate the deletion polymorphisms in GSTM1 and GSTT1 genes and their association with the risk of developing ALS. A case-control study was conducted, including 101 patients diagnosed with ALS and 119 individuals without diagnosis of neurodegenerative diseases. Peripheral blood samples were collected from both groups and subjected to DNA extraction and subsequent genotyping. The polymorphisms were genotyped by the multiplex real - time PCR (qPCR) technique, with the definition of the null and present genotypes by analysis of the melting curves produced after the amplification. Clinical and demographic data were collected from medical records and questionnaires, including topics such as cigarette use, alcohol intake, age of diagnosis, physical activity practice, occupational history, among others. Among the groups analyzed, alcohol consumption was predominant in patients with ALS, with a significant difference between the case and control groups (p=0.01). However, there was no association of GSTM1 (p=0.85), GSTT1 (p=0.90) deletion polymorphisms and their possible genotypic combinations with the risk of developing ALS. The relationship of polymorphisms with the clinical and demographic profile of patients with the disease was also performed. In this analysis, there was a significant difference in the GSTM1-present genotype with the variables: environmental exposure and smoking (p = 0.02 and 0.03, respectively), in the GSTT1-present genotype with a history of neurodegenerative disease in the family (p=0.01), while the double genotype present also showed a significant difference with the family history of neurodegenerative disease (p=0.02). In this context, the results found in this study demonstrate the non-association of GSTM1 and GSTT1 deletion polymorphisms with the development of ALS.Item Marcadores microssatélites para os genomas nuclear e organelar de amburana cearensis (Allemão) A.C. Smith(Universidade Federal de Goiás, 2021-08-30) Santos, Laís Souza; Mansano, Vidal de Freitas; http://lattes.cnpq.br/5618078551561118; Soares, Thannya Nascimento; http://lattes.cnpq.br/5590256762396056; Soares, Thannya Nascimento; Telles, Mariana Pires de Campos; Vianello, Rosana PereiraAmburana cearensis is a tree species, popularly known in Brazil as cerejeira, widely distributed in Brazil and also in Argentina, Bolivia, Paraguay and Peru. The species is distributed in a range of intense anthropic action and classified as endangered according to the International Union for the Conservation of Nature (IUCN). The economic demand for A. cearensis as a wood source and medicinal source has boosted its predatory exploitation, representing a threat to the survival of the species. In this scenario, conservation genetics is essential for generating knowledge about the genetic diversity of the species and defining appropriate strategies for its conservation and management. The general objective of this work was to design primer pairs with high potential for the development of robust and informative microsatellite markers for the nuclear, chloroplast and mitochondrial genomes of A. cearensis. Moreover, for quickness and cost savings, nuclear microsatellite primer pairs were designed for amplification in multiplex PCR. To achieve the proposed objectives, the genome of A. cearensis was partially sequenced, using the Illumina MiSeq platform. The assembly of the genomic library was performed from the DNA of an individual of the species and the genomic sequences obtained were analyzed using FastQC, Trimmomatic and DipSPAdes software. The search for microsatellite regions was performed using the QDD program. Thirty pairs of primers for nuclear microsatellite regions were designed, organized into five sets for amplification in multiplex PCR. The primer pairs designed in this study have the potential to generate new microsatellite markers with high individual discrimination capacity and transferability to other species of the genus Amburana. These markers can be applied quickly and efficiently for different types of genetic diversity studies of A. cearensis and Amburana genus.Item Malato sintase de Paracoccidioides brasiliensis é uma proteína ligada à superfície que se comporta como uma anchorless adesina(Universidade Federal de Goiás, 2009-05-11) Silva Neto, Benedito Rodrigues da; Pereira, Maristela; http://lattes.cnpq.br/1345781867765758; Pereira, Maristela; Soares, Célia Maria de Almeida; Lenzi, Henrique LeonelThe pathogenic fungus Paracoccidioides brasiliensis causative of Paracoccidioidomycosis (PCM), a pulmonary mycose acquired by inhalation of fungal airborne propagules, which may disseminate to several organs and tissues leading to a severe form of the disease. Adhesion and invasion to host cells are essential steps involved in the internalization and dissemination of pathogens. Inside host, P. brasiliensis use the glyoxylate cycle for intracellular survival. Here, we provide evidence that malate synthase of P. brasiliensis (PbMLS) is localized on the cell wall, and is secreted. PbMLS was overexpressed in Escherichia coli, and polyclonal antibody against this protein was obtained. By using Confocal Laser Scanning Microscopy and Western blot analysis, PbMLS was detected in the cytoplasm and the cell wall of the yeast phase of P. brasiliensis of mother and bud yeast cells. PbMLSr and the respective polyclonal antibody produced against this protein inhibited the interaction of P. brasiliensis to in vitro cultured epithelial cells A549. These observations indicated that cell wall-associated MLS of P. brasiliensis could be mediating the binding of fungal cells, thus contributing to the adhesion of fungus to host tissues and to the dissemination of infection.Item Análises proteômicas de cepas de Staphylococcus saprophyticus elucidam diferentes estratégias para virulência e infecção(Universidade Federal de Goiás, 2019-05-13) Silva, Karla Christina Sousa; Rocha, Juliana Alves Parente; http://lattes.cnpq.br/7089231795367245; Rocha, Juliana Alves Parente; Marval, Márcia Giambiagi de; Fontes, Wagner; Kipnis, André; Borges, Clayton LuizStaphylococcus saprophyticus is a gram positive, coagulase negative bacteria and is one of the most common etiological agent of urinary tract infections (UTI) among sexually active young females. The infection may be led mainly by sexual intercourse. Little is known about the molecular features of S. saprophyticus infection model. However, plasticity in the proteomic profile is detected in the genus Staphylococcus and can be related with infection ability. In this study, we aimed to analyze the metabolic differences among three different pathogenic strains of S. saprophyticus (ATCC 15305, 7108 and 9325) using proteomics approaches. The proteomic data revealed variations among the strains that may lead to different responses in the context of an infection. There are differences in the carbohydrate metabolism, amino acid metabolism and pathogen defense against host defenses. Thioredoxin was one of the differentially expressed proteins among strains, thiol level assays showed similar levels of reduced thiol for ATCC15305 and 9325 strains, in contrast 7108 strain showed considerably low levels of reduced thiol levels when compared to the other analyzed strains. Urease is one of the main virulence factors of S. saprophyticus. The cytoplasmatic urease concentrations varied considerably. The 9325 strain had the lowest cytoplasmatic urease activity, however when plated on ureabased agar it showed a high urease activity, which means that probably most of the urease synthesized is exported to the extracellular milieu. Another important feature is related to the purine metabolism operon, which is highly regulated on ATCC15305 and exerts an influence upon the biofilm production in this strain. These results show that, although belonging to the same species, different S. saprophyticus strains present diverse behavior in response to different contexts that the microorganism may face in the host.Item Hibridização in situ por fluorescência: análise histórica, inovações e perspectivas(Universidade Federal de Goiás, 2022-09-05) Silva, Thainá Ferreira; Pinto, Rafael Barbosa; http://lattes.cnpq.br/9944641590654242; Soares, Thannya Nascimento; http://lattes.cnpq.br/5590256762396056; Soares, Thannya Nascimento; Nunes, Rhewter; Machado, Raquel MouraFluorescence in situ hybridization (FISH) is a molecular cytogenetic technique that is based on the use of fluorescence-labeled probes to identify different regions in the genome. From the improvement and development of new technologies, variations of the original technique emerged making it possible to apply it to various issues, from identifying chromosomes through descriptive cytogenetics to making evolutionary inferences or conducting applied research in biotechnology and genetic improvement. Considering the great possibility of applications of the FISH technique and its variations, the present work aims to detail the methodological principles and applications, with emphasis on plants, and to investigate its use in evolutionary studies. For this, the first chapter is a manuscript for the dissemination and popularization of science that aims to discuss the various possibilities of using the FISH technique, ranging from its methodological principles to applications in different areas. From this study, it was verified that FISH can be applied to different biological materials and regions of the genome and, despite being a technique developed for more than three decades, it can generate impacts and contributions through its wide use until today. The second chapter is a systematic review, whose main objective is to summarize and critically analyze the data available in the literature on the applications of FISH for the cytogenetic study in plants, in addition to detailing and discussing the applications of this technique for evolutionary studies in plants. This study was conducted using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. The search was carried out in the Scopus database and the keywords were directed to capture the breadth of the use of the FISH technique aimed at the cytogenetic study of plants. In order to investigate the applications of this technique, the articles found were grouped into different categories, called “Plant Biotechnology”, “Evolutionary Studies”, “Descriptive”, “Techniques/Methodological” and “Gene Expression”. A total of 4006 articles were selected that showed a trend of growth in the number of publications between the years 1975 - 2020. China and the USA concentrated the largest number of articles and Brazil was in 8th position. The analysis of the most used keywords in the articles made it possible to trace a small history of the technique, ranging from the beginning of indirect labeling in plant species, passing through the advances of FISH and its variations, to the use of oligo-synthetic probes today. The categories “Plant Biotechnology” and “Evolutionary Studies” represented more than 60% of the total works, with emphasis on the use of GISH and FISH techniques with repetitive DNA probes and indirect labeling. The Poaceae family stood out in the studies of "Plant Biotechnology" and Leguminosae and Asteraceae were mostly found in the category of "Evolutionary Studies", “Comparative analysis” and “Systems and cytotaxonomy.” Thus, the understanding of the technique, the synthesis and measurement of data and the evolutionary path elaborated in this study allowed the theoretical basis about the versatility of the technique and its use today. demonstrate the main contributions of FISH directed to studies with plants, highlighting the regions of research concentration, variations of the technique, main types of studies and botanical families, as well as contributing to answering different evolutionary questions.Item Montagem e análise do transcritoma do mogno africano (Khaya grandifoliola C. DC.)(Universidade Federal de Goiás, 2019-07-25) Soares, Sabrina Delgado; Novaes, Evandro; http://lattes.cnpq.br/0568272239145336; Novaes, Evandro; Telles, Mariana Pires de Campos; Soares, Thannya Nascimento; Coelho, Alexandre Siqueira Guedes; Vianello, Rosana PereiraKhaya grandifoliola C. DC. is a tropical species with high value wood in international markets. Despite the high interest for reforestation and timber production, virtually nothing is known about the genetic and genomic aspects of the species. With the development of next-generation sequencing platforms (NGS), the characterization of transcriptomes can be performed on a large scale and at affordable cost. In this context, the objective of the work was to discover K. grandifoliola genes by using RNA-Seq technique and to identify those differentially expressed between the leaf and the xylem K. grandifoliola tissues. Total RNA was extracted from leaf and xylem samples of four trees planted in the experimental area of the UFG. mRNA samples were sequenced on the Illumina HiSeq 2000 platform, in paired-end mode 2 × 100 bp. Sequencing resulted in a total of 857 million sequences which, after quality control, decreased to 808 million with an average size of 24 to 85 bp. Trinity program was used to reassemble the transcriptome, obtaining 116,289 transcripts for a total of 36,271 genes. Transcripts ranged from 524 to 12,031 bp with a mean of 1,366 bp and N50 of 1,657 bp. Analyses of gene expression identified 13,626 differentially expressed genes (DEGs) (FDR <0.05) between the two tissues. Of these, 5,438 had higher expression in leaves and 8,188 in xylem. A functional enrichment analysis of Gene Ontology categories among the DEGs was performed by the Blast2GO program. This analysis identified 26 GO terms enriched among the most active DEGs in xylem. Among genes up-regulated in leaves, 48 enriched terms were found. As expected, the categories identified as enriched between DGEs point to biological processes and cellular components specific of each tissue. For example, among the most active genes in leaf tissue, terms associated with thylakoids (GO:0009579), plastids (GO:0009536) and photosynthesis (GO:0015979) were identified. On the other hand, for the xylem, terms related to carbohydrate metabolism (GO:0005975), cell wall (GO:0071554) and cytoskeletal proteins (GO:0008092) were significantly enriched. To the best of our knowledge, this is the first report of large-scale gene sequences for K. grandifoliola. Differential expression analysis allowed us to identify which genes are most active in leaf and xylem. Analysis in iPath3 identified the metabolic routes activated in leaf and xylem tissues. A BlastX search against the orange proteome (Citrus sinensis) revealed the presence of 95 DEGs annotated for the enzymes of the limonoid, a secondary metabolite with medicinal properties.Item Homeostase de ferro em Paracoccidioides spp.: novos alvos de estudo e estabelecimento de HSP30 como proteína ligante de hemoglobina(Universidade Federal de Goiás, 2021-06-28) Souza, Aparecido Ferreira de; Soares, Célia Maria de Almeida; http://lattes.cnpq.br/8539946335852637; Soares, Célia Maria de Almeida; Schrank, Augusto; Parente, Ana Flávia Alves; Bailão, Elisa Flávia Luiz Cardoso; Bailão, Alexandre MeloUnderstanding the mechanisms that govern host-pathogen interaction is crucial for the development of new therapeutic and diagnostic approaches. Fungi of Paracoccidioides genus are the etiological agents of paracoccidioidomycosis, a systemic mycosis and, when in the host, fungi find a hostile environment that is scarce in nutrients and micronutrients, such as Fe, which is indispensable for the survival of the pathogen. Previous studies have shown that fungi of this genus, in response to Fe deprivation, are able to synthesize and capture siderophores (Fe3+ chelators), use host proteins that contain Fe as metal’s source and use a non-canonical reductive pathway for assimilation of Fe. Despite all these findings, there are still gaps that need to be filled regarding the pathogen's response to Fe deprivation. In the present work, targets that still need to be studied in this context were defined, such as the establishment of the protein functions of the fungus that contain the CFEM domain and the determination of the specificity of siderophore receptors. Additionally, it was shown that the exposure of Paracoccidioides to hemoglobin promotes changes at fungal cell wall proteome level, which is illustrated by the increased ability of the fungus to interact with macrophages. This approach allowed the bioprospecting of the hemoglobin-binding protein HSP30, which is present on the cell surface and has its expression regulated positively when the fungus is exposed to hemoglobin. It has been shown that silencing of hsp30 causes decreased growth of the fungus after exposure to hemoglobin. It is also necessary to investigate the function of HSP30 as heme oxygenase. The detailed observation of those open questions can promote the expansion of knowledge about the biology of Paracoccidioides spp. and, consequently, promote the bioprospecting of new targets for differential diagnosis and/or therapy of PCM.