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1.
Artigo em Inglês | MEDLINE | ID: mdl-37463639

RESUMO

Aversive memory extinction comprises a novel learning that blocks retrieving a previously formed traumatic memory. In this sense, aversive memory extinction is an excellent tool for decreasing fear responses. However, this tool it's not effective in the long term because of original memory spontaneous recovery. Thus, searching for alternative strategies that strengthen extinction learning is essential. In the current study, we evaluated the effects of a novel context (i.e., novelty) exposure on aversive memory extinction enhancement over days and the dopaminergic system requirement. Given the purpose, experiments were conducted using 3-month-old male Wistar rats. Animals were trained in inhibitory avoidance (IA). Twenty-four hours later, rats were submitted to a weak extinction protocol. Still, 30 min before the first extinction session, animals were submitted to an exploration of a novel context for 5 min. After, memory retention and persistence were evaluated 24 h, 3, 7, 14, and 21 days later. The exposition of a novel context caused a decrease in aversive responses in all days analyzed and an increase in dopamine levels in the hippocampus. The intrahippocampal infusion of dopamine in the CA1 area or the stimulation of the ventral tegmental area (VTA) by a glutamatergic agonist (NMDA) showed similar effects of novelty. In contrast, VTA inhibition by a gabaergic agonist (muscimol) impaired the persistence of extinction learning induced by novelty exposition and caused a decrease in hippocampal dopamine levels. In summary, we show that novel context exposure promotes persistent aversive memory extinction, revealing the significant role of the dopaminergic system.


Assuntos
Dopamina , Área Tegmentar Ventral , Ratos , Masculino , Animais , Dopamina/farmacologia , Ratos Wistar , Hipocampo , Memória , Extinção Psicológica/fisiologia
2.
Environ Sci Technol ; 56(18): 13019-13028, 2022 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-36053064

RESUMO

The Deepwater Horizon (DWH) disaster released 3.19 million barrels of crude oil into the Gulf of Mexico (GOM) in 2010, overlapping the habitat of pelagic fish populations. Using mahi-mahi (Coryphaena hippurus)─a highly migratory marine teleost present in the GOM during the spill─as a model species, laboratory experiments demonstrate injuries to physiology and behavior following oil exposure. However, more than a decade postspill, impacts on wild populations remain unknown. To address this gap, we exposed wild mahi-mahi to crude oil or control conditions onboard a research vessel, collected fin clip samples, and tagged them with electronic tags prior to release into the GOM. We demonstrate profound effects on survival and reproduction in the wild. In addition to significant changes in gene expression profiles and predation mortality, we documented altered acceleration and habitat use in the first 8 days oil-exposed individuals were at liberty as well as a cessation of apparent spawning activity for at least 37 days. These data reveal that even a brief and low-dose exposure to crude oil impairs fitness in wild mahi-mahi. These findings offer new perspectives on the lasting impacts of the DWH blowout and provide insight about the impacts of future deep-sea oil spills.


Assuntos
Perciformes , Poluição por Petróleo , Petróleo , Hidrocarbonetos Policíclicos Aromáticos , Poluentes Químicos da Água , Animais , Golfo do México , Petróleo/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/análise , Poluentes Químicos da Água/metabolismo
3.
Perspect. nutr. hum ; 24(1): 17-34, ene.-jun. 2022. tab
Artigo em Espanhol | LILACS-Express | LILACS | ID: biblio-1406203

RESUMO

Resumen: Antecedentes: el etiquetado nutricional permite a los consumidores tomar mejores decisiones sobre alimentación, lo que contribuye a combatir las enfermedades crónicas no transmisibles. Objetivo: determinar cómo influye la interpretación del etiquetado nutricional, presente en los alimentos industrializados en Colombia, en la decisión de compra de estudiantes de pregrado del área de la salud de la Universidad CES. Materiales y métodos: se realizó un estudio observacional descriptivo en 76 estudiantes de quinto semestre de programas de pregrado del área de la salud. Resultados: la frecuencia en el uso del etiquetado nutricional se asoció con el sexo (p = 0,036), el tipo de programa (p < 0,001) y el conocimiento de la función del etiquetado nutricional (p = 0,000). La buena interpretación del etiquetado nutricional no condicionó la frecuencia en su uso (p = 0,095); además, el nivel de interpretación del etiquetado nutricional no se asoció con el sexo ni con el tipo de programa de pregrado. Finalmente, se encontró que la decisión de compra estaba influenciada por la costumbre (p = 0,018) y la publicidad (p = 0,008) y no por la interpretación del etiquetado nutricional. Conclusiones: aunque el uso del etiquetado nutricional es más frecuente entre las mujeres, los estudiantes de Nutrición y Dietética y entre quienes conocen la función del etiquetado nutricional, la decisión de compra no está influenciada por su interpretación.


Abstract: Background: Nutritional labeling enables consumers to make better food choices, helping to fight chronic non-communicable diseases. Objective: To determine the influence of the interpretation of the nutritional labeling, present in industrialized foods in Colombia, on the purchase decision of undergraduate students in the health area of the CES University. Material and Methods: A descriptive observational study was carried out in a population of 76 fifth-semester students enrolled in the undergraduate programs of the health area. Results: The frequency of nutrition labeling use was associated with sex (p=0.036), the type of undergraduate program (p<0.001), and knowledge of the function of the nutrition labeling (p=0.000). A good interpretation of the nutrition labeling does not condition the frequency of its use (p=0,095). Additionally, the level of interpretation of the nutritional labeling was not associated with sex, nor the type of undergraduate program. Finally, it was found that the purchase decision was influenced by custom (p=0.018) and advertising (p=0,008), and not by the interpretation of the nutrition labeling. Conclusions: Although the use of nutrition labeling is more frequent among women, among students of Nutrition and Dietetics, and among those who are aware of the role of nutrition labeling, the purchase decision is not influenced by its interpretation.

4.
J Fish Biol ; 100(1): 325-328, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34655253

RESUMO

Regional migration of the copper shark Carcharhinus brachyurus was recorded for the first time in the Southwest Atlantic (SWA) from Argentina (latitude: -38.1037, longitude: -57.5371) to Brazil (latitude: -20.6833, longitude: -40.2846) as a result of a citizen science tagging project. The recaptured specimen was a female (103 kg weight), with 18 developing embryos within the uterus. The total distance was at least 2566 km, and it is the longest ever recorded for the species. Furthermore, it extends its northern distribution in the SWA.


Assuntos
Tubarões , Animais , Oceano Atlântico , Brasil , Cobre , Feminino , Pesqueiros , Alimentos Marinhos
5.
Front Cell Infect Microbiol ; 11: 772311, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34858879

RESUMO

Until 2015, loss-of-function studies to elucidate protein function in Leishmania relied on gene disruption through homologous recombination. Then, the CRISPR/Cas9 revolution reached these protozoan parasites allowing efficient genome editing with one round of transfection. In addition, the development of LeishGEdit, a PCR-based toolkit for generating knockouts and tagged lines using CRISPR/Cas9, allowed a more straightforward and effective genome editing. In this system, the plasmid pTB007 is delivered to Leishmania for episomal expression or integration in the ß-tubulin locus and for the stable expression of T7 RNA polymerase and Cas9. In South America, and especially in Brazil, Leishmania (Viannia) braziliensis is the most frequent etiological agent of tegumentary leishmaniasis. The L. braziliensis ß-tubulin locus presents significant sequence divergence in comparison with Leishmania major, which precludes the efficient integration of pTB007 and the stable expression of Cas9. To overcome this limitation, the L. major ß-tubulin sequences, present in the pTB007, were replaced by a Leishmania (Viannia) ß-tubulin conserved sequence generating the pTB007_Viannia plasmid. This modification allowed the successful integration of the pTB007_Viannia cassette in the L. braziliensis M2903 genome, and in silico predictions suggest that this can also be achieved in other Viannia species. The activity of Cas9 was evaluated by knocking out the flagellar protein PF16, which caused a phenotype of immobility in these transfectants. Endogenous PF16 was also successfully tagged with mNeonGreen, and an in-locus complementation strategy was employed to return a C-terminally tagged copy of the PF16 gene to the original locus, which resulted in the recovery of swimming capacity. The modified plasmid pTB007_Viannia allowed the integration and stable expression of both T7 RNA polymerase and Cas9 in L. braziliensis and provided an important tool for the study of the biology of this parasite.


Assuntos
Leishmania braziliensis , Leishmania major , Sistemas CRISPR-Cas , RNA Polimerases Dirigidas por DNA , Edição de Genes , Leishmania braziliensis/genética , Proteínas Virais
6.
Magn Reson Med ; 86(6): 3022-3033, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34254351

RESUMO

PURPOSE: To develop an Off-Resonance Insensitive Orthogonal CSPAMM sequence (ORI-O-CSPAMM) for the acquisition of CSPAMM and MICSR grids in half of the acquisition time. METHODS: Phantom and mid-level left ventricle short-axis tagged images were acquired using CSPAMM, ORI-CSPAMM, O-CSPAMM, and the proposed ORI-O-CPAMM sequences to interrogate and compare its behavior under off-resonance effects produced by vegetable oil and subcutaneous and epicardial fat. The images were compared in terms of signal and the capacity to obtain complex difference and MICSR images. RESULTS: Like ORI-CSPAMM, the proposed ORI-O-CSPAMM sequence removed almost completely the off-resonance artifacts produced during the tagging preparation. Tagging grids without DC components were obtained with ORI-O-CSPAMM using complex difference and MICSR from only two complementary images, which reduced the scan time to a half compared to CSPAMM and ORI-CSPAMM. The removal of off-resonance effects and the capacity to obtain MICSR images are advantages of ORI-O-CSPAMM over the O-CSPAMM sequence. CONCLUSION: We developed a novel and fast tagging sequence designed to remove off-resonance effects during the tagging preparation, and to obtain complex difference and MICSR grids in half of the scan time compared to CSPAMM and ORI-CSPAMM sequences, which could allow its application to clinical protocols.


Assuntos
Ventrículos do Coração , Imageamento por Ressonância Magnética , Artefatos , Imagens de Fantasmas , Cintilografia
7.
Rev. biol. trop ; Rev. biol. trop;69(supl. 2)mar. 2021.
Artigo em Inglês | LILACS, SaludCR | ID: biblio-1507789

RESUMO

Introduction: The bull shark, Carcharhinus leucas, is particularly vulnerable to anthropogenic actions because of its permanence in coastal ecosystems; populations depletion is registered in different places around the world. Aggregations of bull sharks have been reported in the North Pacific of Costa Rica, at Islas Murciélago, within the Guanacaste Conservation Area. Objective: To study the residency of bull sharks at San Pedrillo islet, Islas Murciélago. Methods: During the study period (June 2013 to February 2015) we used passive telemetry to tag 10 bull sharks. Results: All the sharks tagged were females, they were detected on 59 798 occasions by the acoustic receiver deployed in San Pedrillo. Acoustic signals from tagged sharks were received for a total period of 1 to 229 days (mean = 73.9 ± 71.3 days), with the last detections occurring on 9 January 2015. The Residency Index for each tagged shark across the entire monitoring period ranged from 0.41 to 1.00. The bull shark activity showed a significant pattern throughout the day at the receiver that specifically corresponded with the daily light cycle. Conclusions: This study concludes that San Pedrillo is an aggregation site (cleaning station) for bull sharks (C. leucas), possibly related to reproduction and not feeding behaviors.


Introducción: El tiburón toro (Carcharhinus leucas) es una especie tropical y subtropical que habita ríos, estuarios y aguas costeras. Objetivo: Estudiar la residencia del tiburón toro en San Perdillo, Islas Murciélago, Área de Conservación Guanacaste. Métodos: Durante el período de estudio (junio de 2013 a febrero de 2015) se marcaron 10 tiburones toro con telemetría acústica en San Pedrillo. Resultados: Los tiburones marcados fueron todos hembras y fueron detectados en 59 798 ocasiones por el receptor acústico desplegado en San Pedrillo. Las señales acústicas de los tiburones marcados se recibieron durante un período total de 1 a 229 días (media = 73.9 ± 71.3 días), y las últimas detecciones ocurrieron el 9 de enero de 2015. El índice de residencia de cada tiburón marcado durante todo el período de seguimiento osciló entre 0.41 y 1.00. La actividad del tiburón toro mostró un patrón significativo a lo largo del día en el receptor que se corresponde específicamente con el ciclo de luz diario. Conclusiones: Este estudio concluyó que San Pedrillo es un sitio de agregación (estación de limpieza) para el tiburón toro, posiblemente relacionado con comportamientos de reproducción y no de alimentación; y es un sitio crítico para la especie. Se necesitan más estudios para identificar áreas de alimentación y cría en el Área de Conservación de Guanacaste y áreas circundantes.


Assuntos
Animais , Tubarões/crescimento & desenvolvimento , Telemetria , Costa Rica
8.
J Biotechnol ; 327: 18-27, 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33387593

RESUMO

Rapid development of effective biotherapeutics has been a concern during the last couple decades. In our work we designed two novel peptide tags, GMOP and mGMOP, derived from the N-terminal region of human granulocyte and macrophage colony stimulating factor (hGM-CSF), which contain four and six potential O-glycosylation sites, respectively. These peptide tags were fused to the N-terminus of human interferon-α2b (hIFN-α2b), a therapeutic antiviral and antiproliferative protein rapidly cleared from circulation. Two new molecules were obtained which, consistently with the presence of O-glycans, showed higher molecular masses, more negatively charged isoforms, and higher sialic acid content compared to wild-type IFN. In vitro bioactivity of purified chimeras revealed a similar antiviral specific biological activity (SBA) compared to unmodified IFN. A reduction of antiproliferative SBA was only observed for mGMOP-IFN. Pharmacokinetic studies in rats showed a notable improvement in terminal half-life (t1/2elim) (3.3 and 2.8 times-longer) and a marked reduction of the apparent clearance (CLapp, 3.7 and 4.1-fold lower for GMOP-IFN and mGMOP-IFN in comparison with native IFN, respectively). Furthermore, the in vitro thermal and plasma stability of both proteins was improved. Finally, a monoclonal antibody (mAb) that recognizes an N-terminal GM-CSF epitope was able to bind both chimeras in western blots and ELISAs. This demonstrates the potential of both peptides to behave as bifunctional tags to create novel long-acting biotherapeutics and to facilitate detection and purification.


Assuntos
Fator Estimulador de Colônias de Granulócitos e Macrófagos , Peptídeos , Animais , Anticorpos Monoclonais , Antivirais , Glicosilação , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Engenharia de Proteínas , Ratos , Proteínas Recombinantes/genética
9.
Proc Natl Acad Sci U S A ; 117(30): 18029-18036, 2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32665437

RESUMO

Memory reconsolidation occurs when a retrieving event destabilizes transiently a consolidated memory, triggering thereby a new process of restabilization that ensures memory persistence. Although this phenomenon has received wide attention, the effect of new information cooccurring with the reconsolidation process has been less explored. Here we demonstrate that a memory-retrieving event sets a neural tag, which enables the reconsolidation of memory after binding proteins provided by the original or a different contiguous experience. We characterized the specific temporal window during which this association is effective and identified the protein kinase A (PKA) and the extracellular signal-regulated kinase 1 and 2 (ERK 1/2) pathways as the mechanisms related to the setting of the reconsolidation tag and the synthesis of proteins. Our results show, therefore, that memory reconsolidation is mediated by a "behavioral tagging" process, which is common to different memory forms. They represent a significant advance in understanding the fate of memories reconsolidated while being adjacent to other events, and provide a tool for designing noninvasive strategies to attenuate (pathological/traumatic) or improve (education-related) memories.


Assuntos
Comportamento , Consolidação da Memória/fisiologia , Memória/fisiologia , Animais , Biomarcadores , Masculino , Memória/efeitos dos fármacos , Consolidação da Memória/efeitos dos fármacos , Biossíntese de Proteínas/efeitos dos fármacos , Inibidores da Síntese de Proteínas/farmacologia , Ratos
10.
Proc Natl Acad Sci U S A ; 117(3): 1587-1595, 2020 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-31907312

RESUMO

Many large-bodied marine fishes that form spawning aggregations, such as the Nassau grouper (Epinephelus striatus), have suffered regional overfishing due to exploitation during spawning. In response, marine resource managers in many locations have established marine protected areas or seasonal closures to recover these overfished stocks. The challenge in assessing management effectiveness lies largely in the development of accurate estimates to track stock size through time. For the past 15 y, the Cayman Islands government has taken a series of management actions aimed at recovering collapsed stocks of Nassau grouper. Importantly, the government also partnered with academic and nonprofit organizations to establish a research and monitoring program (Grouper Moon) aimed at documenting the impacts of conservation action. Here, we develop an integrated population model of 2 Cayman Nassau grouper stocks based on both diver-collected mark-resight observations and video censuses. Using both data types across multiple years, we fit parameters for a state-space model for population growth. We show that over the last 15 y the Nassau grouper population on Little Cayman has more than tripled in response to conservation efforts. Census data from Cayman Brac, while more sparse, show a similar pattern. These findings demonstrate that spatial and seasonal closures aimed at rebuilding aggregation-based fisheries can foster conservation success.


Assuntos
Bass/fisiologia , Conservação dos Recursos Naturais , Animais , Conservação dos Recursos Naturais/métodos , Conservação dos Recursos Naturais/estatística & dados numéricos , Monitoramento Ambiental , Pesqueiros , Peixes/fisiologia , Modelos Biológicos , Oceanos e Mares , Densidade Demográfica , Inquéritos e Questionários , Índias Ocidentais
11.
J Forensic Sci ; 65(1): 67-72, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31373685

RESUMO

The collection of gunshot residue on fabric can be an arduous task due to the microscopic size of particles (blind collection) and sheddability of some fabrics. The introduction of luminescent markers and consequent formation of luminescent gunshot residue (LGSR) can facilitate this analysis. In this study, different fabrics were analyzed in order to verify the persistence of the LGSR on them, the possibility of collecting and analyzing particles by video spectral comparator (VSC) and SEM/EDS. Also, different colored fabrics were used as targets in order to investigate influence of fabric color on LGSR visualization. Furthermore, the influence of the fabric type in the distribution of the LGSR deposited around the projectile´s hole entrance was evaluated. The fabric sheddability did not alter collection of the particles or analysis. It was possible to observe and collect LGSR on all tested fabrics, even after the fabric had been shaken, or in colored fabrics.

12.
Neurobiol Learn Mem ; 166: 107101, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31629785

RESUMO

Extinction is the learned inhibition of retrieval of a previously acquired memory and is a major component of exposure therapy, which has attracted much attention because of the use in the treatment of drug addiction, phobias and particularly fear disorders such as post-traumatic stress disorder (PTSD). Exposure to a novel environment before or after extinction training can enhance the extinction of contextual fear conditioning, however the cellular and molecular substrates are still unclear. Here, we investigated the participation of H2-histaminergic, ß-adrenergic and 5-HT1A-serotonergic receptors of the hippocampus on the enhancement of extinction memory caused by novelty. The infusion into the CA1 region of the serotonin 5-HT1A-receptor agonist, 8-OH-DPAT and the ß-adrenergic blocker, Timolol, after the exposure to the novelty hindered the enhancement of extinction by novelty, while Timolol also hindered the extinction consolidation when infused post-extinction. These impairments were abolished by the coinfusion of 8-OH-DPAT plus the 5-HT1A receptor antagonist, NAN-190 and Timolol plus ß-adrenergic agonist, Isoproterenol. However, Dimaprit and Ranitidine blocked the retrieval of CFC, but did not prevented the extinction learning. Here we elucidated some of the molecular mechanisms that are involved on the enhancement of extinction by novelty, demonstrating that the ß-adrenoreceptors and 5-HT1A serotonergic receptors participate on this process alongside with dopaminergic D1 receptors previously described, while histamine H2 receptors, so ubiquitous in learning-related functions in hippocampus are not involved.


Assuntos
Agonistas Adrenérgicos beta/farmacologia , Antagonistas Adrenérgicos beta/farmacologia , Aprendizagem por Associação/efeitos dos fármacos , Extinção Psicológica/efeitos dos fármacos , Medo/efeitos dos fármacos , Hipocampo/efeitos dos fármacos , Agonistas do Receptor de Serotonina/farmacologia , 8-Hidroxi-2-(di-n-propilamino)tetralina/farmacologia , Animais , Condicionamento Clássico/efeitos dos fármacos , Isoproterenol/farmacologia , Masculino , Ratos Wistar , Timolol/farmacologia
13.
Hippocampus ; 29(6): 491-499, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30295349

RESUMO

Stress is known to have a critical impact on memory processes. In the present work, we focus on the effects of an acute stress event closely associated to an unrelated learning task. Here, we show that acute stress (elevated platform [EP] session) experienced 1 hr after a weak spatial object recognition (SOR) training, which only induces a short-term memory (STM), promoted the formation of SOR-long term memory (SOR-LTM) in rats. The effect induced by stress was dependent on the activation of glucocorticoid- and mineralocorticoid-receptors, brain-derived neurotrophic factor (BDNF) and protein synthesis in the dorsal hippocampus. In contrast, EP after a strong SOR impaired SOR-LTM probably by interfering with the use of necessary resources. Moreover, we show that the EP session before training induced anterograde interference, which it was not reversed by a subsequent exposure to an open field. Our findings provide novel insights into the impact of stress on LTM formation in rodents and they are discussed under the behavioral analogue of the synaptic tagging and capture hypothesis.


Assuntos
Hipocampo/fisiologia , Memória de Longo Prazo/fisiologia , Reconhecimento Psicológico/fisiologia , Estresse Fisiológico/fisiologia , Animais , Fator Neurotrófico Derivado do Encéfalo/fisiologia , Masculino , Memória de Curto Prazo/fisiologia , Ratos , Ratos Wistar , Receptores de Glucocorticoides/fisiologia , Receptores de Mineralocorticoides/fisiologia , Aprendizagem Espacial/fisiologia , Memória Espacial/fisiologia
14.
Forensic Sci Int ; 281: 161-170, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29156218

RESUMO

Gunshot residues (GSR) are important physical evidence in firearm-related crimes. Recently developed non-toxic ammunition, however, requires a new methodology for its characterization. To overcome this drawback, the introduction of noncommercial luminescent markers in ammunition was proposed. These markers, synthesized and added to the gunpowder, presented as a versatile tool for GSR analysis, since they require UV radiation alone to visualize the luminescent GSR (LGSR). This has opened up new perspectives for understanding GSR behavior at a crime scene. This work aims to expand previous studies performed with the luminescent markers in forensic contexts, exploring four different important aspects related to GSR behavior. Using LGSR amount/dispersion and a series of blind tests with marked ammunition, we tried to (1) identify the shooter position; (2) estimate the shooting distance; (3) evaluate the influence of the pistol type on the LGSR distribution on the shooter's hands and guns; and (4) study the transference of LGSR by a chain of handshaking. For this purpose, a portable UV lamp (λ=254nm) and/or techniques such as video spectral comparator (VSC) and scanning electron microscopy with energy dispersive X-ray analysis (SEM/EDS) were used to visualize and analyze the residues. As a result, the observation of the LGSR enabled accurate determination of the shooter position and the firing distance without any chemicals. Besides, the LGSR were visualized on different kinds of pistols, regardless of firing mechanism. And finally, this study evidenced transference of residues from shaking hands with the shooter, which should be carefully considered when interpreting the results of a GSR analysis.

15.
Bio Protoc ; 7(10)2017 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-28758140

RESUMO

To achieve the C-terminal tagging of endogenous proteins in T. cruzi we use the Cas9/pTREX-n vector (Lander et al., 2015) to insert a specific tag sequence (3xHA or 3xc-Myc) at the 3' end of a specific gene of interest (GOI). Chimeric sgRNA targeting the 3' end of the GOI is PCR-amplified and cloned into Cas9/pTREX-n vector. Then a DNA donor molecule to induce DNA repair by homologous recombination is amplified. This donor sequence contains the tag sequence and a marker for antibiotic resistance, plus 100 bp homology arms corresponding to regions located right upstream of the stop codon and downstream of the Cas9 target site at the GOI locus. Vectors pMOTag23M (Oberholzer et al., 2006) or pMOHX1Tag4H (Lander et al., 2016b) are used as PCR templates for DNA donor amplification. Epimastigotes co-transfected with the sgRNA/Cas9/pTREX-n construct and the DNA donor cassette are then cultured for 5 weeks with antibiotics for selection of double resistant parasites. Endogenous gene tagging is finally verified by PCR and Western blot analysis.

16.
Adv Exp Med Biol ; 974: 219-227, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28353239

RESUMO

Shotgun proteomics has been used successfully for more than two decades for measurement of proteins from diverse biological systems. This has led to new insights and identification of potential biomarkers and drug targets for numerous medical conditions. The advent of mass-labeling approaches such as isobaric tagging for relative and absolute quantitation (iTRAQ) has allowed multiplexing of samples to improve the accuracy and throughput of experiments and the reduction of costs. Here, we describe a detailed iTRAQ mass spectrometry protocol for simultaneous analysis of four proteomes from postmortem brain tissue extracts. In addition, a post-labeling procedure for peptide fractionation is presented.


Assuntos
Química Encefálica , Transtornos Mentais/metabolismo , Proteínas do Tecido Nervoso/análise , Espectrometria de Massas em Tandem/métodos , Biomarcadores/análise , Cromatografia Líquida/métodos , Humanos , Peso Molecular , Nanotecnologia/métodos , Proteínas do Tecido Nervoso/isolamento & purificação , Fragmentos de Peptídeos/análise , Espectrometria de Massas por Ionização por Electrospray/métodos , Coloração e Rotulagem/métodos
17.
Univ. sci ; 21(1): 33-52, Jan.-Apr. 2016. ilus, tab
Artigo em Inglês | LILACS-Express | LILACS | ID: lil-783693

RESUMO

The protection of the habitats used by juvenile sharks is a management strategy that has recently caught the attention of fishery biologists. In the present study, we evaluated the population of the lemon shark (Negaprion brevirostris) from Los Roques Archipelago in order to identify the nursery area, describe the size composition, and examine the variation in nocturnal activity of the juvenile individuals. The data analysed came from three different sources: commercial shark fishery, tag-recapture sampling, and visual records. A total of 375 lemon sharks with total lengths between 55 and 281 cm were recorded during the study period. Overall data showed that the area occupied by juvenile lemon sharks was clearly partitioned into primary and secondary nurseries. Additionally, nighttime activity seemed to change according to the size of sharks in the primary nursery, suggesting a reduction of time activity overlapping among juveniles of distinct size/age. Results suggest that the strategy of utilization of the primary nurseries by the lemon shark may lead to important ecological benefits by reducing the competition, predation and natural mortality.


La protección de los hábitats utilizados por tiburones juveniles es una estrategia de manejo que ha capturado recientemente la atención de los biólogos de pesquerías. En el presente estudio se evaluó la población del tiburón limón (Negaprion brevirostris) del Archipiélago Los Roques con el fin de identificar su zona de cría, describir su composición de tamaño y examinar la variación en la actividad nocturna de los juveniles. Los datos analizados provienen de 3 fuentes distintas: pesquería comercial de tiburones, muestreo de captura-marcaje-recaptura y registros visuales. Durante el período de estudio se registró un total de 375 tiburones limón, con longitudes totales entre 55 y 281 cm. En general, los datos mostraron que el área ocupada por los tiburones limón estaba claramente dividida entre zonas de cría primarias y secundarias. Adicionalmente, la actividad nocturna parecía cambiar de acuerdo con el tamaño de los tiburones en la zona de cría primaria, lo cual sugiere una reducción de la coincidencia entre juveniles de distinto tamaño/edad. Los resultados sugieren que la estrategia de utilización de la zona de cría primaria por el tiburón limón puede conducir a importantes beneficios ecológicos por la reducción de competencia, predación y mortalidad natural.


A protecao dos habitats utilizados pelos tubaroes jovens é uma estratégia de manejo que recentemente tem chamado á atenção dos biólogos pesqueiros. No presente estudo avaliamos a populacao do tubarao limao (Negaprion brevirostris) no Arquipélago de Los Roques, com o propósito de identificar a área de berçário, descrever a composição de tamanhos, e examinar a variação da atividade noturna dos individuos jovens. Os dados analisados foram originados a partir de tres fontes diferentes: Foram analisados dados da pescaría comercial de tubaroes, trabalhos de marcação e recaptura, e registos visuais. Um total de 375 tubaroes limáo com comprimento total entre 55 e 281 cm foi registado durante o período de estudo. Os dados mostram que a zona ocupada pelos tubaroes limao jovens se encontra claramente dividida em áreas de berçários primaria e secundária. Adicionalmente, a atividade noturna dentro da área de berçário primaria pareceu mudar de acordo com o tamanho dos tubaroes, sugerindo uma separação desta atividade por grupos de idade. Os resultados sugerem que a estratégia de utilizacao da área de berçário primaria do tubarao limao poderia oferecer importantes benefícios ecológicos, como a reducao da competicao, predacao e mortalidade natural.

18.
J Fish Biol ; 88(2): 800-4, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26694077

RESUMO

Photo-tagging, i.e. using a specific software to match colour patterns on photographs, was tested as a means to identify individual Indo-Pacific Pterois volitans to assist with population and movement studies of this invasive species. The stripe pattern on the flank of adult P. volitans (n = 48) was the most individually distinctive of three body regions tested, leading to correct individual identification on 68 and 82% of tests with a single and two images of the reference individual, respectively. Photo-tagging is inexpensive, logistically simple and can involve citizen scientists, making it a viable alternative to traditional tagging to provide information on P. volitans distribution, movement patterns and recolonization rates after removals.


Assuntos
Sistemas de Identificação Animal , Perciformes , Fotografação , Animais , Espécies Introduzidas , Software
19.
Neotrop. ichthyol ; 14(1)apr. 2016. ilus, tab, graf
Artigo em Inglês | VETINDEX | ID: vti-339471

RESUMO

Photo-identification allows individual recognition of animal species based on natural marks, being an alternative to other more stressful artificial tagging/marking techniques. An increasing number of studies with different animal groups has shown that photo-identification can successfully be used in several situations, but its feasibility to study freshwater fishes is yet to be explored. We demonstrate the potential use of photo-identification for intraspecific recognition of individuals in the stream-dwelling loricariid Rineloricaria aequalicuspis . We tested photo-identification in laboratory and field conditions based on the interindividual variability in abdominal bony plates. Our test yielded high correct matches in both laboratory (100%) and field conditions (> 97%), comparable to other reliable techniques and to studies that successfully used photo-identification in other animals. In field conditions, the number of correct matches did not differ statistically between computer-assisted and naked-eye identification. However, the average time expended to conclude computer-assisted photo evaluations was about half of the time expended to conclude naked-eye evaluations. This result may be exacerbated when using database with large number of images. Our results indicate that photo-identification can be a feasible alternative technique to study freshwater fish species, allowing for a wider use of mark-recapture in ecological and behavioral studies.(AU)


A foto-identificação permite o reconhecimento individual de espécies de animais baseando-se em marcas naturais, sendo uma alternativa a outras técnicas de marcação artificial mais estressantes comumente usadas. O número crescente de estudos que usam foto-identificação em diferentes grupos animais mostra que esta técnica pode ser utilizada com sucesso, mas a viabilidade em estudos com peixes de água doce ainda não foi avaliada. Nós demonstramos o uso potencial da foto-identificação para o reconhecimento individual de peixes com indivíduos do loricarídeo Rineloricaria aequalicuspis. Nós testamos foto-identificação em condições de laboratório e de campo com base na variabilidade inter-individual das placas ósseas abdominais. O teste resultou em elevada porcentagem de acerto nas comparações, tanto para a condição de laboratório (100%) quanto para a de campo (> 97%), o que é comparável com outras técnicas confiáveis e com outros estudos que empregaram foto-identificação com sucesso. No teste de campo, o número de acertos não diferiu estatisticamente entre auxílio de computador e olho nu. Entretanto, o tempo médio despendido para concluir as avaliações com o auxílio de computador foi cerca da metade do tempo despendido para as avaliações a olho nu. Esse resultado pode ser exacerbado em avaliações com um grande número de imagens. Nossos resultados indicam que a foto-identificação pode ser uma técnica alternativa viável para estudar peixes de água doce e possibilita um uso mais amplo da marcação e recaptura para estudos ecológicos e comportamentais.(AU)


Assuntos
Animais , Sistemas de Identificação Animal/métodos , Sistemas de Identificação Animal , Sistemas de Identificação Animal/veterinária , Peixes-Gato
20.
Neotrop. ichthyol ; 14(1)2016. ilus, tab, graf
Artigo em Inglês | LILACS | ID: lil-794413

RESUMO

Photo-identification allows individual recognition of animal species based on natural marks, being an alternative to other more stressful artificial tagging/marking techniques. An increasing number of studies with different animal groups has shown that photo-identification can successfully be used in several situations, but its feasibility to study freshwater fishes is yet to be explored. We demonstrate the potential use of photo-identification for intraspecific recognition of individuals in the stream-dwelling loricariid Rineloricaria aequalicuspis . We tested photo-identification in laboratory and field conditions based on the interindividual variability in abdominal bony plates. Our test yielded high correct matches in both laboratory (100%) and field conditions (> 97%), comparable to other reliable techniques and to studies that successfully used photo-identification in other animals. In field conditions, the number of correct matches did not differ statistically between computer-assisted and naked-eye identification. However, the average time expended to conclude computer-assisted photo evaluations was about half of the time expended to conclude naked-eye evaluations. This result may be exacerbated when using database with large number of images. Our results indicate that photo-identification can be a feasible alternative technique to study freshwater fish species, allowing for a wider use of mark-recapture in ecological and behavioral studies.


A foto-identificação permite o reconhecimento individual de espécies de animais baseando-se em marcas naturais, sendo uma alternativa a outras técnicas de marcação artificial mais estressantes comumente usadas. O número crescente de estudos que usam foto-identificação em diferentes grupos animais mostra que esta técnica pode ser utilizada com sucesso, mas a viabilidade em estudos com peixes de água doce ainda não foi avaliada. Nós demonstramos o uso potencial da foto-identificação para o reconhecimento individual de peixes com indivíduos do loricarídeo Rineloricaria aequalicuspis. Nós testamos foto-identificação em condições de laboratório e de campo com base na variabilidade inter-individual das placas ósseas abdominais. O teste resultou em elevada porcentagem de acerto nas comparações, tanto para a condição de laboratório (100%) quanto para a de campo (> 97%), o que é comparável com outras técnicas confiáveis e com outros estudos que empregaram foto-identificação com sucesso. No teste de campo, o número de acertos não diferiu estatisticamente entre auxílio de computador e olho nu. Entretanto, o tempo médio despendido para concluir as avaliações com o auxílio de computador foi cerca da metade do tempo despendido para as avaliações a olho nu. Esse resultado pode ser exacerbado em avaliações com um grande número de imagens. Nossos resultados indicam que a foto-identificação pode ser uma técnica alternativa viável para estudar peixes de água doce e possibilita um uso mais amplo da marcação e recaptura para estudos ecológicos e comportamentais.


Assuntos
Animais , Sistemas de Identificação Animal/métodos , Sistemas de Identificação Animal , Sistemas de Identificação Animal/veterinária , Peixes-Gato
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