Computational predictive approaches present themselves as an attractive alternative in the search for new compounds that may present toxic or growth regulatory activity in Aedes aegypti. Aspergillus species is a producer of secondary metabolites with diverse biological activities. In this study, it was possible to evaluate the pharmacokinetic and toxicological properties in silico, as well as the prediction of the insecticidal biological activities of the juvenile hormone of natural substances produced by the genus Aspergillus. Initially, 187 molecules were cataloged from various species of Aspergillus spp., and only seven molecules of indole alkaloids were found to exhibit a toxic dose 50% (TD50) after conducting toxicity predictions. The molecular dynamic simulation trajectories were utilized to study parameters such as root mean square deviation, root mean square fluctuation, radius of gyration and intermolecular hydrogen bond to understand the behavior and stability of proposed compounds in the binding pocket of the target protein. The N-β-acetyltryptamine molecule showed a binding affinity value of –8.100 ± 0.200 kcal mol-1, which classifies the compound as a potential insecticidal agent against the Aedes aegypti vector. silico Initially 18 spp spp. 50 TD50 TD (TD50 predictions deviation fluctuation protein Nβacetyltryptamine N β acetyltryptamine 8100 8 100 –8.10 0200 0 200 0.20 mol1, mol1 mol 1, 1 mol-1 vector 5 TD5 (TD5 810 10 –8.1 020 20 0.2 mol- (TD 81 –8. 02 2 0. –8 –

SIGNIFICANCE: The Freshwater Biodiversity Information System (FBIS) is a powerful, open-access platform for hosting, sharing, and analysing freshwater biodiversity data for South Africa. Platform design and functionality were strongly informed by the requirements of key data end-users and decision-makers. The platform has over 790 000 occurrence records, which are freely available to data-users and are now being converted into impact through incorporation in national decision-support tools. With better data comes better decisions, and access to a data repository like FBIS means that the monitoring, management and conservation of our freshwater ecosystems can be improved, for the long-term betterment of both people and biodiversity.

Microbial fuel cells (MFCs) represent a promising technology to generate bio-electricity and synchronously reduce wastewater pollutants. The presence of exoelectrogens in wastewater is critical for bio-electricity and pollutant reduction, but the performance of exoelectrogens at different pH levels remains unknown. This study aims to bridge this gap by offering an integrated approach to understanding the performance of exoelectrogens under varying substrate pH, particularly in bio-electricity generation and pollutant reduction in sugarbeet processing wastewater (SBWW). Three pH levels (ranging from acidic to alkaline) were studied and MFC's electricity output was measured. Later, current density, power density, and coulombic efficiency (CE) were calculated. Both pre- and post-experiment substrate samples were analysed with inductively coupled plasma (ICP). Furthermore, 16S rRNA gene analysis, DNA amplification, sequencing library preparation, and bioinformatics workflows on post-experiment samples of the substrate and anode samples were conducted. A diverse community of microorganisms was identified, especially Alphaproteobacteria, Gammaproteobacteria, and Deltaproteobacteria (Geobacter). Bacteroidetes and Desulfovibrio were the major exoelectrogens responsible for electricity generation. Among the three pH levels tested, the most alkaline pH level (9.5±0.1) outperformed the others, achieving a 54% higher power density, 21% greater current density, and a 40% higher CE compared to the acidic pH level (6.5±0.1). Around 50-99% of pollutants were removed from the SBWW. The study revealed that Gammaproteobacteria thrive and perform better in alkaline environment.

Abstract The progress in genetic material sequencing enhanced diverse research opportunities related to genomics, transcriptomics and other -omics areas. A need in the development of more accessible and efficient techniques resulted in the second- (NGS) and third-generation technologies (TGS). In this paper we aimed to characterize the main sequencing methods of second- and third-generation. NGS technology is based on the sequencing by biosynthesis of short fragments with high quality and precision, while TGS are specialized in long fragment sequencing by nanopores and single-molecule, real-time technologies. Illumina and PacBio have more precise and high-quality data, compared to Oxford Nanopore Technologies (ONT). However, nanopores long reads address errors in genome assembly, commonly associated with small lectures. All analyzed technologies have production-scale sequencers, but only ONT and Illumina developed benchtop products. ONT also presents portable sequencers for small projects. Latinoamerican countries have less coverage and technical support of sequencing technologies, and only Illumina is sited in this region. Each sequencing technology have advantages and limitations regarding their techniques and their coupled use is promoted to make a deeper analysis of the data, therefore the development of new bioinformatics programs remain as one of the principal obstacles to overcome to enhance the application of these technologies.

Resumen El avance en la secuenciación de material genético ha favorecido el estudio de nuevas áreas de investigación relacionadas a la genómica, transcriptómica y demás ciencias -ómicas. Esto creó la necesidad de desarrollar nuevas tecnologías más accesibles y eficientes, como las tecnologías de segunda y tercera generación. Por ello, esta revisión caracterizó las principales metodologías de secuenciación de segunda y tercera generación. Las tecnologías de segunda generación se ven representadas por la secuenciación por síntesis fragmentos cortos con alta calidad y precisión. En la tercera generación se especializan en lecturas largas secuenciadas por nanoporos y secuenciación de una sola molécula en tiempo real. Las empresas Illumina y PacBio poseen una mayor calidad y precisión en las lecturas, en comparación a Oxford Nanopore Technologies (ONT). Sin embargo, las lecturas de ONT solucionan problemas del ensamblaje genómico propios de lecturas cortas. Todas las tecnologías analizadas poseen secuenciadores para proyectos de producción a escala; Illumina y ONT también poseen secuenciadores sobremesa, y solamente la última presenta dispositivos portátiles. Con respecto a la accesibilidad, los países centroamericanos poseen poca cobertura por parte de estas empresas, siendo Illumina la única con una distribuidora radicada en la región. Cada tecnología de secuenciación posee ventajas y desventajas asociadas, lo cual impulsa a su uso en conjunto, lo que permite análisis más convenientes y completos. Por lo cual el desarrollo de nuevos programas bioinformáticos para la mejora del análisis corresponde a consideraciones que deben ser abordadas con la intención de fortalecer el uso de las técnicas de secuenciación.

Resumen El cáncer gástrico es un proceso fatal cuyos factores de riesgo incluyen la infección por Helicobacter pylori, la anemia perniciosa, la ingesta de compuestos nitrosos, el abuso de alcohol, el tabaquismo y el sexo masculino. El seguimiento endoscópico ha definido la progresión histológica de lesiones premalignas a lesiones malignas. Sin embargo, los tumores gástricos exhiben diferentes y muy variadas variaciones histológicas, comportamientos clínicos y respuestas a tratamiento. Esta gran heterogeneidad intrapaciente e interpaciente ha obligado a la búsqueda de principios reguladores que gobiernen la evolución del cáncer gástrico. Los programas avanzados de bioinformática, la tecnología de microarreglos de ADN y la genómica funcional han ayudado a integrar la estructura, función y dinámica de moléculas biológicas expresadas o secretadas por las células epiteliales cancerosas que han modificado la clasificación del cáncer gástrico.

Abstract Gastric cancer is a fatal process whose risk factors include infection by Helicobacter pylori, pernicious anemia, nitroso compounds, alcohol abuse, cigarette smoking, and male gender. Endoscopic surveillance has defined the histological progression of premalignant to malignant lesions. Nevertheless, gastric tumors exhibit distinct histologic variations, clinical behaviors, and treatment responses. This “intra and interpatient heterogeneity” has obliged us to search for key principles governing gastric cancer evolution. Advanced bioinformatics programs, DNA microarray technology, and functional genomics have helped to integrate the structure, function, and dynamics of biological molecules expressed or secreted by cancer epithelial cells that have modified the classification of gastric cancer.

Resumen: El gel de Aloe vera es considerado una fuente natural de múltiples beneficios, originados por la acción combinada de vitaminas, aminoácidos, compuestos fenólicos, enzimas, minerales, ácidos orgánicos, lípidos y carbohidratos, que se relacionan con la mejora de enfermedades neuro-degenerativas como Alzheimer. Los ensayos in vitro e in silico permiten confirmar e identificar posibles beneficios de esta planta y sus compuestos en enfermedades. El objetivo del presente trabajo fue evaluar la actividad antioxidante del gel de A. vera y mediante análisis in silico, establecer el potencial terapéutico de sus compuestos bioactivos en la enfermedad de Alzheimer. Se obtuvieron hojas de A. vera, de las que se extrajo el gel, retirando el exocarpio, se liofilizó y almacenó hasta su uso. Se caracterizó la capacidad antioxidante, se cuantificaron los compuestos fenólicos y flavonoides y se analizó la relación que existe entre los parámetros mediante correlación de Pearson. Mediante análisis in silico se evaluó el potencial de interacción de 8 compuestos del gel con la proteína gamma secretasa. El gel de A. vera obtuvo alta capacidad antioxidante por ABTS, DPPH, radical OH y poder reductor, usando bajas concentraciones para inhibir el 50 % de los radicales, y correlaciones positivas con fenoles totales y flavonoides. En el estudio in silico el compuesto que presentó mejor unión con gamma secretasa fue aloe-emodina, con menor energía libre de unión y menor concentración de constante de inhibición, sugiriendo su potencial uso como coadyuvante en el tratamiento de la enfermedad de Alzheimer.

Abstract: Aloe vera gel is considered a natural source of multiple benefits, originated by the combined action of vitamins, amino acids, phenolic compounds, enzymes, minerals, organic acids, lipids and carbohydrates, which are related to the improvement of neuro-degenerative diseases such as Alzheimer’s. In vitro and in silico tests allow us to confirm and identify possible benefits of this plant and its compounds in diseases. The objective of the present study was to evaluate the antioxidant activity of A. vera gel and, through in silico analysis, to establish the therapeutic potential of its bioactive compounds in Alzheimer’s disease. A. vera leaves were obtained, from which the gel was extracted, removing the exocarp, lyophilized and stored until use. The antioxidant capacity was characterized, the phenolic compounds and flavonoids were quantified, and the relationship between the parameters was analyzed using Pearson correlation. The interaction potential of 8 compounds in the gel with the gamma secretase protein was evaluated through in silico analysis. The A. vera gel obtained high antioxidant capacity due to ABTS, DPPH, OH radical and reducing power, using low concentrations to inhibit 50 % of the radicals, and positive correlations with total phenols and flavonoids. In the in silico study, the compound that showed the best binding with gamma secretase was aloe-emodin, with lower binding free energy and lower inhibition constant concentration, suggesting its potential use as an adjuvant in the treatment of Alzheimer’s disease.

Resumen Las displasias esqueléticas son alteraciones del crecimiento óseo y cartilaginoso con amplio espectro clínico y radiológico, cuya prevalencia es 2 a 5 casos por 10 000 recién nacidos. Se presenta el caso de un paciente masculino de 9 años con talla baja asimétrica, neurodesarrollo normal, sin deformidades osteomusculares y familiares con talla baja no estudiada. Se diagnosticó displasia esquelética, sin embargo, ante estudios paraclínicos normales, se indicó estudio molecular que reveló una nueva variante de significancia incierta en el gen TRVP4. Posteriormente, con el uso de bases de datos y de bioinformática, se reclasifica a probablemente patogénica. En valoraciones ulteriores se determinó que no cuenta con manejo dirigido y, ante la ausencia de complicaciones susceptibles de terapia, se indicó seguimiento clínico. Se resalta la importancia del reanálisis de estudios moleculares y del fortalecimiento bioinformático que facilite el diagnóstico oportuno, permitiendo la aplicación de estrategias de detección, seguimiento y tratamiento.

Abstract Skeletal dysplasias are alterations in bone and cartilage growth with a wide clinical and radiological spectrum, whose prevalence is 2 to 5 cases per 10 000 newborns. The case of a 9-year-old male patient with asymmetrical short stature, normal neurodevelopment, without musculoskeletal deformities and relatives with unexplored short stature is presented. Skeletal dysplasia was diagnosed; however, in the presence of normal paraclinical studies, a molecular study was indicated that revealed a new variant of uncertain significance in the TRVP4 gene. Subsequently, with the use of databases and bioinformatics, it was reclassified as likely pathogenic. In subsequent assessments, it was determined that there was no specific management and, given the absence of complications amenable to therapy, clinical follow-up was indicated. The importance of reanalysis of molecular studies and bioinformatic strengthening that facilitates timely diagnosis is highlighted, allowing the application of detection, monitoring and treatment strategies.

Abstract Biological sequence analysis is essential in bioinformatics, but the diversity of available tools presents significant challenges. This article proposes a web application that unifies useful functionalities in biological sequence analysis. Developed in Python and Django, it integrates features from BioPython, BioPandas, 3Dmol.js, and BLAST, among others, making it usable for users without programming knowledge. The tool covers the analysis of DNA and protein sequences, transcription, alignment, sequence search, and more. Its most notable features are efficiency, precision in obtaining quantitative data from the running process, and ease of use, supported by validation and results from the components used. Its modular architecture ensures scalability and adaptability. It is concluded that the tool represents a significant advance in simplifying and improving the processes of biological sequence analysis, promising to enhance efficiency and effectiveness in bioinformatics.

Resumen El análisis de secuencias biológicas es esencial en bioinformática, pero la diversidad de herramientas disponibles presenta desafíos significativos. Este artículo propone una aplicación web que unifica diferentes funcionalidades útiles en el análisis de secuencias biológicas. Desarrollada en Python y Django, integra funcionalidades de BioPython, BioPandas, 3Dmol.js y BLAST, entre otras, que además posibilita su uso a usuarios sin conocimientos de programación. La herramienta abarca el análisis de secuencias de ADN y proteínas, transcripción, alineación, búsqueda de secuencias y más. Dentro de las características más destacables está la eficiencia, precisión al obtener datos cuantitativos del proceso que se está ejecutando y la facilidad de uso.

Abstract: Sickle cell anemia (SCA) is the most common genetic disease worldwide. There are countries with massive public health programs for early detection of this condition. In the literature, several specific haplotypes or single-base polymorphic variants (SNPs) have been associated with the SCA prognosis. Objective: To demonstrate the significant correlation of SNPs relevant to the diagnosis and prognosis of SCA among different ethnic groups. Methodology: we analyzed population frequencies and correlations of several SNPs related to the prognosis of SCA (i.e., baseline fetal hemoglobin levels), response to hydroxyurea treatment, and response to other drugs used in the SCA treatment, collected from validated genomic databases among different ethnic groups. Results: The calculation of the Hardy-Weinberg equilibrium and the logistic regression was successful in classifying the ethnic groups as African (0 = 0.78, 1 = 0.89), and with a lower efficiency as American (AMR) (0 = 0.88, 1 = 0.00), East Asian (EAS) (0 = 0.80, 1 = 0.00), European (EUR) (0 = 0.79, 1 = 0.00), and South Asian (SAS) (0 = 0.80, 1 = 0.00). Conclusions: The results extend those from previous reports and show that the profile of most of the SNPs studied presented statistically significant distributions among general ethnic groups, pointing to the need to carry out massive early screening of relevant SNPs for SCA in patients diagnosed with this disease. It is concluded that the application of a broad mutation detection program will lead to a more personalized and efficient response in the treatment of SCA.

Resumen: La anemia falciforme (AF) es la enfermedad genética más frecuente del mundo. Hay países con programas de salud pública masivos para la detección temprana esta condición. Varios haplotipos específicos y polimorfismos de una sola base (SNPs) se han asociado en la literatura con la prognosis de AF. Objetivo: Demostrar la correlación significativa de SNPS relevantes para el diagnóstico y prognosis de AF entre diferentes grupos étnicos. Metodología: se analizaron las frecuencias poblacionales y correlaciones entre varios SNPs relevantes para la prognosis de AF (ej: niveles basales de hemoglobina fetal), respuesta a tratamiento con hydroxiurea y respuesta a otras drogas utilizadas en el tratamiento de AF. Los datos fueron obtenidos de bases de datos genómicos validadas entre diferentes grupos étnicos. Resultados: Los cálculos del equilibrio de Hardy-Weinberg y la regresión logística fue exitosa en clasificar los grupos étnicos, Africano (0 = 0,78, 1 = 0,89), y con menor eficiencia; Americano (AMR) (0 = 0,88, 1 = 0,00), Asia del Este (EAS) (0 = 0,80, 1 = 0,00), Europeo (EUR) (0 = 0,79, 1 = 0,00), y Sud-Asiático (SAS) (0 = 0,80, 1 = 0,00). Conclusiones: Estos resultados, extienden los trabajos previos y muestran que el perfil de mutaciones de la mayoría de los SNPs analizados, presenta distribuciones estadísticamente significativas entre grupos étnicos generales, apuntando a la necesidad de realizar programas de testeo masivo de SNPs para AF en pacientes diagnosticados con esta patología. Se concluye que la aplicación de un programa amplio de genotipificación de mutación generara una respuesta más eficiente y personalizada en el tratamiento de AF.

Resumen: La anemia falciforme (AF) es la enfermedad genética más frecuente del mundo. Hay países con programas de salud pública masivos para la detección temprana esta condición. Varios haplotipos específicos y polimorfismos de una sola base (SNPs) se han asociado en la literatura con la prognosis de AF. Objetivo: Demostrar la correlación significativa de SNPS relevantes para el diagnóstico y prognosis de AF entre diferentes grupos étnicos. Metodología: se analizaron las frecuencias poblacionales y correlaciones entre varios SNPs relevantes para la prognosis de AF (ej: niveles basales de hemoglobina fetal), respuesta a tratamiento con hydroxiurea y respuesta a otras drogas utilizadas en el tratamiento de AF. Los datos fueron obtenidos de bases de datos genómicos validadas entre diferentes grupos étnicos. Resultados: Los cálculos del equilibrio de Hardy-Weinberg y la regresión logística fue exitosa en clasificar los grupos étnicos, Africano (0 = 0,78, 1 = 0,89), y con menor eficiencia; Americano (AMR) (0 = 0,88, 1 = 0,00), Asia del Este (EAS) (0 = 0,80, 1 = 0,00), Europeo (EUR) (0 = 0,79, 1 = 0,00), y Sud-Asiático (SAS) (0 = 0,80, 1 = 0,00). Conclusiones: Estos resultados, extienden los trabajos previos y muestran que el perfil de mutaciones de la mayoría de los SNPs analizados, presenta distribuciones estadísticamente significativas entre grupos étnicos generales, apuntando a la necesidad de realizar programas de testeo masivo de SNPs para AF en pacientes diagnosticados con esta patología. Se concluye que la aplicación de un programa amplio de genotipificación de mutación generara una respuesta más eficiente y personalizada en el tratamiento de AF.

Abstract: Sickle cell anemia (SCA) is the most common genetic disease worldwide. There are countries with massive public health programs for early detection of this condition. In the literature, several specific haplotypes or single-base polymorphic variants (SNPs) have been associated with the SCA prognosis. Objective: To demonstrate the significant correlation of SNPs relevant to the diagnosis and prognosis of SCA among different ethnic groups. Methodology: we analyzed population frequencies and correlations of several SNPs related to the prognosis of SCA (i.e., baseline fetal hemoglobin levels), response to hydroxyurea treatment, and response to other drugs used in the SCA treatment, collected from validated genomic databases among different ethnic groups. Results: The calculation of the Hardy-Weinberg equilibrium and the logistic regression was successful in classifying the ethnic groups as African (0 = 0.78, 1 = 0.89), and with a lower efficiency as American (AMR) (0 = 0.88, 1 = 0.00), East Asian (EAS) (0 = 0.80, 1 = 0.00), European (EUR) (0 = 0.79, 1 = 0.00), and South Asian (SAS) (0 = 0.80, 1 = 0.00). Conclusions: The results extend those from previous reports and show that the profile of most of the SNPs studied presented statistically significant distributions among general ethnic groups, pointing to the need to carry out massive early screening of relevant SNPs for SCA in patients diagnosed with this disease. It is concluded that the application of a broad mutation detection program will lead to a more personalized and efficient response in the treatment of SCA.

Resumo A salmonela é uma causa séria de problemas de saúde em humanos e animais em todo o mundo. Salmonella tem sido isolada de diferentes amostras biológicas e é considerada como o principal papel na indução da inflamação do trato gastrointestinal que por sua vez causa diarreia em diferentes espécies. Compreender melhor o envolvimento da Salmonella na contaminação e infecção de ovos frescos e carne de frango caipira. Este estudo teve como objetivo estabelecer as detecções microbiológicas e moleculares de Salmonella na cloaca da galinha caipira e identificar as funções biológicas previstas usando as vias da Enciclopédia de Gene e Genômica de Kyoto (KEGG) e interação proteína-proteína. Suabes cloacais foram coletados de galinhas criadas ao ar livre em fazenda local na cidade de Duhok. Os isolados foram cultivados e o teste bioquímico realizado com XLD e TSI, respectivamente. A detecção molecular e anotação funcional do gene invA foi realizada usando PCR convencional e abordagens de bioinformática. O presente estudo descobriu que Salmonella foi detectada em 36 das 86 amostras usando métodos microbiológicos. Para confirmar esses achados, o gene invA foi utilizado e 9 dos 36 isolados de Salmonella mostraram um sinal positivo de invA pelo gel de agarose. Além disso, a análise bioinformática revelou que o gene invA estava associado principalmente a processos de secreção bacteriana, assim como seus termos KEGG e interação proteína-proteína estavam envolvidos na invasão bacteriana e nas vias de secreção. Esses achados sugeriram que o gene invA desempenha um papel importante na regulação dos processos de colonização e invasão de Salmonella dentro do hospedeiro intestinal na galinha caipira.

Abstract Salmonella is a serious cause of the health issues in human and animal worldwide. Salmonella has been isolated from different biological samples and it considers as the key role in induction of inflammation of gastrointestinal tract which in turn cause diarrhoea in different species. To further understand the involvement of Salmonella in contaminating and infecting fresh eggs and meat of free-range chicken. This study aimed to establish the microbiological and molecular detections of Salmonella in the cloaca of the free-range chicken and to identify predicted biological functions using Kyoto Encyclopedia of Gene and Genomic (KEGG) pathways and protein-protein interaction. Cloacal swabs were collected from free range chicken raised in the local farm in Duhok city. The isolates were cultured and biochemical test performed using XLD and TSI, respectively. Molecular detection and functional annotation of invA gene was carried out using Conventional PCR and bioinformatics approaches. The present study found that Salmonella was detected in 36 out of 86 samples using microbiological methods. To confirm these findings, invA gene was utilised and 9 out of 36 Salmonella isolates have shown a positive signal of invA by agarose gel. In addition, bioinformatic analysis revealed that invA gene was mainly associated with bacterial secretion processes as well as their KEGG terms and Protein-Protein Interaction were involved in bacterial invasion and secretion pathways. These findings suggested that invA gene plays important role in regulating colonization and invasion processes of Salmonella within the gut host in the free range chicken.

Resumo Ligula intestinalis é um parasita cestódeo que acomete peixes de água doce em diversos países do mundo. O presente estudo visa revelar a diversidade filogenética, genética e de haplótipos das sequências do gene mt-CO1 enviadas ao banco de dados do NCBI de diferentes países, por meio de análise in-silico. As sequências gênicas de 105 mt-CO1 (371 pb) de L. intestinalis obtidas do NCBI foram utilizadas para análises bioinformáticas. As sequências foram submetidas a análise filogenética e de haplótipos. Como resultado da análise de haplótipos de L. intestinalis, 38 haplótipos foram obtidos de 13 países diferentes. Hap24 constituiu 44,76% da rede de haplótipos obtida. Mudanças nos nucleotídeos entre os haplótipos ocorreram em 1-84 pontos diferentes. A China e a Turquia apresentam os maiores valores do índice de fixação (Fst), 0,59761, enquanto o menor (-0,10526) foi encontrado entre a Rússia e a Turquia. Este estudo fornece uma linha de base para futuros estudos em larga escala sobre epidemiologia, aspectos ecológicos, padrão de distribuição, dinâmica de transmissão e dispersão populacional de L. intestinalis em todo o mundo.

Abstract Ligula intestinalis is a cestode parasite that affects freshwater fish in different countries of the world. The current study aims to reveal the phylogenetic, genetic and haplotype diversity of mt-CO1 gene sequences sent to the NCBI database from different countries by using in-silico analysis. The 105 mt-CO1 (371 bp) gene sequences of L. intestinalis obtained from NCBI were used for bioinformatics analyses. Sequences were subjected to phylogenetic and haplotype analysis. As a result of the haplotype analysis of L. intestinalis, 38 haplotypes were obtained from 13 different countries. Hap24 constituted 44.76% of the obtained haplotype network. Changes in nucleotides between haplotypes occurred at 1-84 different points. China and Turkey have highest fixation index (Fst) values of 0.59761, while the lowest (-0.10526) was found between Russia and Turkey. This study provides a baseline for future studies on extensive scale on the epidemiology, ecological aspects, distribution pattern, transmission dynamics and population dispersion of L. intestinalis worldwide.

Resumo A pesquisa atual foi projetada para alcançar o potencial de produção de protease extracelular em diferentes cepas de Sordaria fimicola que foram previamente obtidas do Dr. Lamb (Imperial College, Londres) de North Facing Slope e South Facing Slope de Evolution Canyon. Após a triagem inicial e secundária, duas cepas hiperprodutoras S2 e N6 foram selecionadas para fermentação submersa e condições culturais, incluindo temperatura, pH, período de incubação, tamanho do inóculo, concentração de substrato, e diferentes fontes de carbono e nitrogênio foram otimizadas para produção de enzima. A cepa S2 apresentou produção máxima de protease de 3,291 U/mL após 14 dias de incubação a 30 °C com pH 7, concentração de substrato de 1% e inóculo de 1 mL, enquanto a cepa N6 apresentou produção máxima de protease de 1,929 U/mL em condições otimizadas de fermentação. Outro objetivo da presente pesquisa foi sustentar a biodiversidade da genética e modificações pós-tradicionais (PTMs) da protease DPAP (peptidil-aminopeptidase) em Sordaria fimicola. Cinco sítios polimórficos foram observados na sequência de aminoácidos de cepas de S. fimicola com referência a Neurospora crassa. A previsão de PTMs a partir de ferramentas de bioinformática previu 38 locais de fosforilação em resíduos de serina para protease peptidil-aminopeptidase na cepa S1 de S. fimicola, enquanto 45 locais de fosforilação em serina na cepa N7 e 47 modificações de fosforilação de serina foram previstas em N. crassa. A pesquisa atual deu uma ideia de que a mudança na composição genética afetou os PTMs que, em última análise, afetaram a produção da enzima protease em diferentes cepas do mesmo organismo (S. fimicola). A produção e os dados moleculares da pesquisa revelaram que o estresse ambiental tem fortes efeitos sobre genes específicos por meio de mutações que podem causar diversidade genética. S. fimicola é um fungo não patogênico e tem um ciclo de vida curto. Esse fungo pode ser escolhido para produzir enzima protease em escala comercial.

Abstract The current research was designed to reach extracellular protease production potential in different strains of Sordaria fimicola which were previously obtained from Dr. Lamb (Imperial College, London) from North Facing Slope and South Facing Slope of Evolution Canyon. After initial and secondary screening, two hyper-producers strains S2 and N6 were selected for submerged fermentation and cultural conditions including temperature, pH, incubation period, inoculum size, substrate concentration, and different carbon and nitrogen sources were optimized for enzyme production. S2 strain showed maximum protease production of 3.291 U/mL after 14 days of incubation at 30 °C with 7 pH, 1% substrate concentration and 1 mL inoculum, While N6 strain showed maximum protease production of 1.929 U/mL under fermentation optimized conditions. Another aim of the present research was to underpin the biodiversity of genetics and post-translational modifications (PTMs) of protease DPAP (peptidyl-aminopeptidase) in Sordaria fimicola. Five polymorphic sites were observed in amino acid sequence of S. fimicola strains with reference to Neurospora crassa. PTMs prediction from bioinformatics tools predicted 38 phosphorylation sites on serine residues for protease peptidyl-aminopeptidase in S1 strain of S. fimicola while 45 phosphorylation sites on serine in N7 strain and 47 serine phosphorylation modifications were predicted in N. crassa. Current research gave an insight that change in genetic makeup effected PTMs which ultimately affected the production of protease enzyme in different strains of same organism (S. fimicola). The production and molecular data of the research revealed that environmental stress has strong effects on the specific genes through mutations which may cause genetic diversity. S. fimicola is non- pathogenic fungus and has a short life cycle. This fungus can be chosen to produce protease enzyme on a commercial scale.

Resumo Os microRNAs (miRNAs) são reconhecidos como biomarcadores do diabetes mellitus tipo 2 (DM2), úteis para a compreensão do metabolismo da doença, e possuem grande potencial como alvos terapêuticos. O aumento da expressão de BDNF e IGF1 está altamente envolvido nos benefícios as vias de insulina e glicose, porém, são regulados negativamente em condições de resistência à insulina, enquanto seu aumento de expressão está correlacionado com a melhora do metabolismo da glicose e da insulina. Estudos sugerem a regulação desses genes por microRNA em vários contextos diferentes, proporcionando uma nova abordagem de investigação para compreender o metabolismo do DM2 e revelar potenciais alvos terapêuticos. No presente estudo, investigamos em diferentes modelos animais (humanos, ratos e camundongos) miRNAs que têm como alvo BDNF e IGF1 em tecido muscular esquelético com condições fisiológicas de DM2. As análises foram realizadas utilizando ferramentas de bioinformática e bancos de dados para predição de miRNA, homologia molecular, validação experimental de interações, expressão na condição fisiológica estudada e interação em rede. Os resultados mostraram três candidatos a miRNAs para IGF1 (miR-29a, miR-29b e miR-29c) e um para BDNF (miR-206). As avaliações experimentais e a busca pela expressão no músculo esquelético de indivíduos com DM2 confirmaram a interação prevista entre miRNA-mRNA para miR-29b e miR-206 através de modelos humanos, ratos e camundongos. Essa interação foi reafirmada em múltiplas análises de rede. Em conclusão, nossos resultados mostram a relação de regulação entre miR-29b e miR-206 com os genes investigados, em diversos tecidos, sugerindo um padrão de inibição. Contudo, esses dados mostram um grande número de possíveis processos fisiológicos de interação para perspectivas biotecnológicas.

Abstract microRNAs (miRNAs) are recognized as diabetes mellitus type 2 (T2DM) biomarkers useful for disease metabolism comprehension and have great potential as therapeutics targets. BDNF and IGF1 increased expression are highly involved in the benefits of insulin and glucose paths, however, they are down-regulated in insulin resistance conditions, while their expression increase is correlated to the improvement of glucose and insulin metabolism. Studies suggest the microRNA regulation of these genes in several different contexts, providing a novel investigation approach for comprehending T2DM metabolism and revealing potential therapeutic targets. In the present study, we investigate in different animal models (human, rat, and mouse) miRNAs that target BDNF and IGF1 in skeletal muscle tissue with T2DM physiological conditions. Bioinformatics tools and databases were used to miRNA prediction, molecular homology, experimental validation of interactions, expression in the studied physiological condition, and network interaction. The findings showed three miRNAs candidates for IGF1(miR-29a, miR-29b, and miR-29c) and one for BDNF (miR-206). The experimental evaluations and the search for the expression in skeletal muscle from T2DM subjects confirmed the predicted interaction between miRNA-mRNA for miR-29b and miR-206 through human, rat, and mouse models. This interaction was reaffirmed in multiple network analyses. In conclusion, our results show the regulation relationship between miR-29b and miR-206 with the investigated genes, in several tissues, suggesting an inhibition pattern. Nevertheless, these data show a large number of possible interaction physiological processes, for future biotechnological prospects.

Resumo A produção de plantas transgênicas com características agronômicas e hortícolas desejadas vem ganhando grande importância por atender às demandas da crescente população. A transformação genética também é um passo fundamental para estudar os fundamentos das ciências das plantas. Foram desenvolvidos e utilizados diferentes protocolos de transformação, que são confiáveis e eficientes. Esses protocolos usaram genes de resistência a antibióticos ou herbicidas incorporados ao gene de interesse para identificar plantas transformadas e não transformadas. Esses genes marcadores podem representar uma ameaça ao ser humano e ao meio ambiente. O uso de marcadores visuais permite a observação direta e fácil de plantas transformadas com mais precisão. No estudo atual, foi projetado um cassete de genes com o gene da hidroxilase de produção de pigmentos (PPH) sob promotor específico de fibra (GhSCFP) e terminador NOS a jusante. Após verificar a eficiência estrutural e funcional do gene otimizado por códons usando ferramentas de bioinformática, o cassete foi enviado para síntese química a partir de fonte comercial. O cassete do gene do pigmento (PPH_CEMB), clonado em pCAMBIA-1301, foi transformado em Agrobacterium por eletroporação. O método de imersão floral mediado por Agrobacterium foi usado para transformar a inflorescência de Camelina sativa. Após a formação de sementes, foi observado um total de 600 sementes com mudança de cor, das quais 19 desenvolveram uma coloração marrom-avermelhada, enquanto as 581 restantes permaneceram amarelas. A eficiência de transformação calculada com base na mudança de cor foi de 1%. A análise de PCR das folhas obtidas após a semeadura de sementes avermelhadas confirmou a transformação do gene produtor de pigmentos, enquanto não foi observada amplificação por PCR em folhas de plantas de sementes do tipo selvagem. A partir dos resultados, é evidente que a transformação da inflorescência de C. sativa mediada por Agrobacterium é uma técnica muito eficiente e favorável ao ambiente não só para a detecção de plantas transformadas, mas também para estudar processos celulares básicos.

Abstract Production of transgenic plants with desired agronomic and horticultural traits has gained great importance to fulfill demands of the growing population. Genetic transformation is also a fundamental step to study basics of plant sciences. Different transformation protocols have been developed and used which are reliable and efficient. These protocols used antibiotic or herbicide resistance genes incorporated along with gene of interest to identify transformed plants from non-transformed ones. These marker genes may pose a threat to human and environment. Use of visual markers enables direct and easier observation of transformed plants with more precision. In current study a gene cassette with ‘pigment production hydroxylase (PPH) gene under fiber specific promoter (GhSCFP) and downstream Nos-terminator was designed. After checking the structural and functional efficiency of codon optimized gene using bioinformatics tools, the cassette was sent for chemical synthesis from commercial source. The pigment gene cassette (PPH_CEMB), cloned in pCAMBIA-1301, was transformed into Agrobacterium through electroporation. Agrobacterium-mediated floral dip method was used to transform Camelina sativa inflorescence. After seed setting a total of 600 seed were observed for change in color and out of these, 19 seeds developed a reddish-brown coloration, while the remaining 581 seeds remained yellow. The transformation efficiency calculated on basis of color change was 1.0%. PCR analysis of leaves obtained after sowing reddish seeds confirmed the transformation of pigment production gene, while no PCR amplification was observed in leaves of plants from wild type seeds. From the results it is evident that Agrobacterium-mediated transformation of C. sativa inflorescence is very efficient and environment friendly technique not only for detection of transformed plants but also to study basic cellular processes.