RESUMO
Current antifungal drugs present poor effectiveness and there is no available vaccine for fungal infections. Thus, novel strategies to treat or prevent invasive mycosis, such as cryptococcosis, are highly desirable. One strategy is the use of immunomodulators of polysaccharide nature isolated from mushrooms. The purpose of the present work was to evaluate the immunostimulatory activity of ß-(1,3)-glucan-containing exopolysaccharides (EPS) from the edible mushrooms Auricularia auricula in phagocytes and mice infected with Cryptococcus neoformans. EPS triggered macrophages and dendritic cell activation upon binding to Dectin-1, a pattern recognition receptor of the C-type lectin receptor family. Engagement of Dectin-1 culminated in pro-inflammatory cytokine production and cell maturation via its canonical Syk-dependent pathway signaling. Furthermore, upon EPS treatment, M2-like phenotype macrophages, known to support intracellular survival and replication of C. neoformans, repolarize to M1 macrophage pattern associated with enhanced production of the microbicidal molecule nitric oxide that results in efficient killing of C. neoformans. Treatment with EPS also upregulated transcript levels of genes encoding products associated with host protection against C. neoformans and Dectin-1 mediated signaling in macrophages. Finally, orally administrated ß-glucan-containing EPS from A. auricular enhanced the survival of mice infected with C. neoformans. In conclusion, the results demonstrate that EPS from A. auricula exert immunostimulatory activity in phagocytes and induce host protection against C. neoformans, suggesting that polysaccharides from this mushroom may be promising as an adjuvant for vaccines or antifungal therapy.
Assuntos
Agaricales/química , Criptococose/prevenção & controle , Polissacarídeos Fúngicos/imunologia , Fagócitos/efeitos dos fármacos , Fagócitos/imunologia , beta-Glucanas/imunologia , Animais , Criptococose/imunologia , Cryptococcus neoformans/imunologia , Citocinas/imunologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/microbiologia , Fatores Imunológicos/farmacologia , Lectinas Tipo C/imunologia , Pneumopatias Fúngicas , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Fagócitos/microbiologia , Transdução de Sinais , beta-Glucanas/farmacologiaRESUMO
Haemonchus contortus is one of the major gastrointestinal nematodes responsible for significant economic and production losses of sheep. Diseases caused by this species lack effective anthelmintic products, and the search for new compounds to replace synthetic anthelmintics has been extensive. The present investigation assesses the in vitro activity of the essential oil of melaleuca (Melaleuca alternifolia), both free (TTO) and nanostructured (nanoTTO), and terpinen-4-ol (terp-4-ol) on eggs and larvae of H. contortus. Tests of egg hatching (EHT) and inhibition of larval migration (LMIT) were used to assess the in vitro efficacy of TTO, nanoTTO and terp-4-ol. Using EHT, at a concentration of 3.5 mg/ml, 100% inhibition occurred using TTO and terp-4-ol, with LC50 values of 0.43 and 0.63 mg/ml, and LC90 values of 1.75 mg/ml and 3.12 mg/ml, respectively. NanoTTO had lower activity, with 82.6% inhibition at the same concentration. Using LMIT, TTO and nanoTTO had a similar activity with 88.0% and 84.8% inhibition, respectively, at a concentration of 56 mg/ml. Terp-4-ol had a greater effect on larvae, with 85.7% inhibition at a concentration of 56 mg/ml and 82.4% at 3.5 mg/ml, demonstrating high activity at the lowest concentration tested. Therefore, the results indicate that all substances tested showed ovicidal and larvicidal activity against H. contortus. TTO, terp-4-ol and, mainly, nanoTTO may be targeted in in vivo studies, besides being a promising line of research into the control and treatment of veterinary important helminths.
Assuntos
Anti-Helmínticos/farmacologia , Haemonchus/efeitos dos fármacos , Melaleuca/química , Óleos Voláteis/farmacologia , Terpenos/farmacologia , Zigoto/efeitos dos fármacos , Animais , Anti-Helmínticos/isolamento & purificação , Bioensaio , Haemonchus/crescimento & desenvolvimento , Larva/efeitos dos fármacos , Locomoção/efeitos dos fármacos , Óleos Voláteis/isolamento & purificação , Análise de Sobrevida , Terpenos/isolamento & purificaçãoRESUMO
Drought and rice blast disease caused by Magnaporthe oryzae are two of the most serious threats to global rice production. To explore the mechanisms underlying gene expression induced in rice by stresses, studies involving transcriptome analyses have been conducted over the past few years. Thus, it is crucial to have a reliable set of reference genes to normalize the expression levels of rice genes affected by different stresses. To identify potential reference genes for studies of the differential expression of target genes in rice under M. oryzae infection and drought conditions, the present study evaluated five housekeeping genes for the normalization of gene expression. The stability of the expression of these genes was assessed using the analytical software packages geNorm and NormFinder. For all samples analyzed, the stability rank was UBQ5 > GAPDH > eIF-4α> ß-TUB > 18S rRNA. The data showed that the UBQ5, GAPDH, and eIF-4αgenes are appropriate, high-performing reference genes and will be highly useful in future expression studies of fungal infections and drought in rice.
Assuntos
Regulação da Expressão Gênica de Plantas , Genes Essenciais , Genes de Plantas , Oryza/genética , Reação em Cadeia da Polimerase em Tempo Real/normas , Secas , Fator de Iniciação 4A em Eucariotos/genética , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/genética , Magnaporthe/patogenicidade , Magnaporthe/fisiologia , Oryza/microbiologia , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Padrões de Referência , Estresse Fisiológico , Transcriptoma , Ubiquitina/genéticaRESUMO
STUDY QUESTION: Is the pain associated with levonorgestrel-releasing intrauterine system (LNG-IUS) insertion reduced by intracervical anesthesia in women without previous vaginal birth? SUMMARY ANSWER: Intracervical anesthesia was not associated with reduced pain in women without previous vaginal birth. WHAT IS KNOWN ALREADY: The pain associated with the insertion of intrauterine contraceptives (IUCs) is a limiting factor for the use of these contraceptives by some women. No prophylactic pharmacological intervention has proven efficacy in relieving pain during or after the insertion of IUCs. However, previous studies included women with previous vaginal delivery, and injectable intracervical anesthesia was not evaluated in any of these studies. STUDY DESIGN, SIZE, DURATION: This was a randomized, open, parallel-group clinical trial that evaluated 100 women without previous vaginal delivery who wished to use the LNG-IUS for the first time. These women were evaluated immediately after LNG-IUS insertion and then 2 h and 6 h later. PARTICIPANTS/MATERIALS, SETTING, METHODS: The 100 women were randomized into two groups: (i) use of a non-steroidal anti-inflammatory drug (NSAID) (ibuprofen, 400 mg) 1 h prior to LNG-IUS insertion; or (ii) 2% lidocaine intracervical injection 5 min prior to LNG-IUS insertion. The women were evaluated immediately after LNG-IUS insertion and then 2 h and 6 h after insertion. Two pain scales were used (the visual analogue scale and the facial pain scale) in addition to assessing the ease of insertion (as rated by the provider) and the level of discomfort during the procedure (as rated by the patient). Multivariate logistic regression was performed to analyze the predictors associated with moderate/severe pain. MAIN RESULTS AND THE ROLE OF CHANCE: The pain and discomfort associated with LNG-IUS insertion, and the ease of insertion of the LNG-IUS did not differ between the groups. Nulliparity was more associated with moderate/severe pain [adjusted odds ratio (OR): 3.1 (95% confidence interval (CI): 1.3-7.80]. Injectable intracervical anesthesia use reduced the risk of moderate/severe pain by 40% [adjusted OR: 0.6 (95% CI: 0.2-1.4)]. The difference between the mean pain score in the intracervical anesthesia group and the NSAID group was <10%; thus, the effect size of the intervention was not significant. LIMITATIONS, REASONS FOR CAUTION: Intracervical anesthesia was compared with an oral medication in this study. Intracervical injection of a saline solution or even a dry needling as the placebo for a double-blind study could be a more adequate control; however, this approach was not a protocol approved by the institutional review board. Considering that the majority of the insertions were easy (>80% in both groups), the results may not be extrapolated to difficult insertions with moderate/severe pain where local anesthesia may have a role. WIDER IMPLICATIONS OF THE FINDINGS: The findings can be generalized to most insertions in nulliparous women or in those without a previous vaginal delivery. There is currently no evidence to recommend the routine use of prophylactic intracervical anesthesia prior to LNG-IUS insertion; there is no evidence that this treatment reduces insertion-related pain. STUDY FUNDING/COMPETING INTERESTS: RAF and CSV give occasional lectures for Bayer Healthcare. This study received funding from the National Institute of Hormones and Women's Health, National Council for Scientific and Technological Development (CNPq). TRIAL REGISTRATION NUMBER: NCT02155166.
Assuntos
Anestésicos Locais/uso terapêutico , Anti-Inflamatórios não Esteroides/uso terapêutico , Colo do Útero/efeitos dos fármacos , Dispositivos Intrauterinos Medicados/efeitos adversos , Lidocaína/uso terapêutico , Dor/tratamento farmacológico , Adulto , Anestésicos Locais/administração & dosagem , Feminino , Humanos , Levanogestrel/administração & dosagem , Lidocaína/administração & dosagem , Pessoa de Meia-Idade , Dor/etiologia , Medição da Dor , Resultado do TratamentoRESUMO
α-Amylases are common enzymes responsible for hydrolyzing starch. Insect-pests, whose larvae develop in seeds, rely obligatorily on α-amylase activity to digest starch, as their major food source. Considering the relevance of insect α-amylases and the natural α-amylase inhibitors present in seeds to protect from insect damage, we report here the molecular cloning and nucleotide sequence of the full-length AmyHha cDNA of the coffee berry borer, Hypothenemus hampei, a major insect-pest of coffee crops. The AmyHha sequence has 1879 bp, containing a 1458 bp open reading frame, which encodes a predicted protein with 485 amino acid residues, with a predicted molecular mass of 51.2 kDa. The deduced protein showed 55-79% identity to other insect α-amylases, including Anthonomus grandis, Ips typographus and Sitophilus oryzae α-amylases. In depth analysis revealed that the highly conserved three amino acid residues (Asp184, Glu220, and Asp285), which compose the catalytic site are also presented in AmyHha amylase. The AmyHha gene seems to be a single copy in the haploid genome and AmyHha transcription levels were found higher in L2 larvae and adult insects, both corresponding to major feeding phases. Modeling of the AmyHha predicted protein uncovered striking structural similarities to the Tenebrio molitor α-amylase also displaying the same amino acid residues involved in enzyme catalysis (Asp184, Glu220 and Asp285). Since AmyHha gene was mostly transcribed in the intestinal tract of H. hampei larvae, the cognate α-amylase could be considered a high valuable target to coffee bean insect control by biotechnological strategies.
Assuntos
Besouros/fisiologia , DNA Complementar/genética , Comportamento Alimentar , alfa-Amilases/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Besouros/classificação , Besouros/enzimologia , Modelos Moleculares , Dados de Sequência Molecular , Fases de Leitura Aberta , Filogenia , Reação em Cadeia da Polimerase em Tempo Real , Homologia de Sequência de Aminoácidos , alfa-Amilases/químicaRESUMO
Microrganismos presentes em dejetos de suínos podem contaminar o meio ambiente. Embora a compostagem seja preconizada como um método eficiente para reduzir este potencial poluidor dos dejetos, existem poucas informações de pesquisa sobre tal processo. O presente trabalho teve o objetivo de avaliar a eficiência da compostagem automatizada dos dejetos líquidos de suínos (DLS) na redução da população de coliformes, usados como indicadores de poluição fecal. Os DLS foram adicionados periodicamente, durante 106 dias, em substrato constituído pela mistura, em partes iguais, de maravalha e serragem. Foram efetuadas 14 adições de DLS, e em cada adição as leiras de compostagem eram revolvidas por meio de uma máquina especialmente desenvolvida para este fim. Foram avaliados dois tratamentos com três repetições, sendo um com e outro sem adição de ácido fosfórico aos dejetos, até pH 6,0. A adição de ácido visou reduzir as perdas de N por volatilização de amônia (NH3) durante a compostagem. A avaliação da população de coliformes foi feita pela técnica do número mais provável (NMP), com uso do caldo Fluorocult, incubado a 37ºC por 24h e posterior leitura em luz ultravioleta. A população de coliformes fecais não foi afetada pela adição de ácido fosfórico. O processo de compostagem automatizada foi eficiente na redução de coliformes fecais, cuja população original passou de 4,2x1010 para 1,2 x 105 ao final da compostagem (156 dias) sem adição de ácido e de 3,8x1010 para 2,3x104 na compostagem com adição de ácido. Essa remoção de coliformes fecais, promovida pela compostagem automatizada dos dejetos líquidos de suínos, corresponde a 99,99 por cento...
Microorganisms present in pig manure can contaminate the environment. Although composting is recommended as an efficient method to reduce the pollution potential of waste, there is little research information on this process. This study aimed to evaluate the efficiency of automated composting of pig slurry (PS) in reducing the population of coliforms, used as fecal pollution indicators. The PS was added periodically during 106 days in substrate, with a mixture, in equal parts, of wood shavings and sawdust. There were 14 additions of PS and at each addition the compost windrows were revolved through a machine especially developed for this purpose. Two treatments with three replications were evaluated, one with and one without the addition of phosphoric acid to the slurry up to pH 6.0. The acid addition aimed to reduce N losses through the volatilization of ammonia (NH3) during composting. Coliforms were evaluated by the technique of most probable number (MPN) using the Fluorocult broth, incubated at 37 ° C for 24 h and subsequent reading in ultra violet light. The population of fecal coliforms was not affected by the addition of phosphoric acid. The automated composting process was effective in reducing faecal coliforms, whose original population decreased from 4.2 x 1010 to 1.2 x 105 at the end of composting (156 days) without addition of acid and from 3.8 x1010 to 2,3 x104 in compost with added acid. This removal of faecal coliforms, promoted by automated composting of pig slurry, corresponds to 99.99 percent...
Assuntos
Animais , Agroindústria , Coliformes/métodos , Compostagem/métodos , Escherichia coli , Tratamento de Efluentes Industriais , Suínos , Ácidos Fosfóricos/administração & dosagemRESUMO
Microrganismos presentes em dejetos de suínos podem contaminar o meio ambiente. Embora a compostagem seja preconizada como um método eficiente para reduzir este potencial poluidor dos dejetos, existem poucas informações de pesquisa sobre tal processo. O presente trabalho teve o objetivo de avaliar a eficiência da compostagem automatizada dos dejetos líquidos de suínos (DLS) na redução da população de coliformes, usados como indicadores de poluição fecal. Os DLS foram adicionados periodicamente, durante 106 dias, em substrato constituído pela mistura, em partes iguais, de maravalha e serragem. Foram efetuadas 14 adições de DLS, e em cada adição as leiras de compostagem eram revolvidas por meio de uma máquina especialmente desenvolvida para este fim. Foram avaliados dois tratamentos com três repetições, sendo um com e outro sem adição de ácido fosfórico aos dejetos, até pH 6,0. A adição de ácido visou reduzir as perdas de N por volatilização de amônia (NH3) durante a compostagem. A avaliação da população de coliformes foi feita pela técnica do número mais provável (NMP), com uso do caldo Fluorocult, incubado a 37ºC por 24h e posterior leitura em luz ultravioleta. A população de coliformes fecais não foi afetada pela adição de ácido fosfórico. O processo de compostagem automatizada foi eficiente na redução de coliformes fecais, cuja população original passou de 4,2x1010 para 1,2 x 105 ao final da compostagem (156 dias) sem adição de ácido e de 3,8x1010 para 2,3x104 na compostagem com adição de ácido. Essa remoção de coliformes fecais, promovida pela compostagem automatizada dos dejetos líquidos de suínos, corresponde a 99,99 por cento.(AU)
Microorganisms present in pig manure can contaminate the environment. Although composting is recommended as an efficient method to reduce the pollution potential of waste, there is little research information on this process. This study aimed to evaluate the efficiency of automated composting of pig slurry (PS) in reducing the population of coliforms, used as fecal pollution indicators. The PS was added periodically during 106 days in substrate, with a mixture, in equal parts, of wood shavings and sawdust. There were 14 additions of PS and at each addition the compost windrows were revolved through a machine especially developed for this purpose. Two treatments with three replications were evaluated, one with and one without the addition of phosphoric acid to the slurry up to pH 6.0. The acid addition aimed to reduce N losses through the volatilization of ammonia (NH3) during composting. Coliforms were evaluated by the technique of most probable number (MPN) using the Fluorocult broth, incubated at 37 ° C for 24 h and subsequent reading in ultra violet light. The population of fecal coliforms was not affected by the addition of phosphoric acid. The automated composting process was effective in reducing faecal coliforms, whose original population decreased from 4.2 x 1010 to 1.2 x 105 at the end of composting (156 days) without addition of acid and from 3.8 x1010 to 2,3 x104 in compost with added acid. This removal of faecal coliforms, promoted by automated composting of pig slurry, corresponds to 99.99 percent.(AU)
Assuntos
Animais , Suínos , Tratamento de Efluentes Industriais , Agroindústria , Compostagem/métodos , Coliformes/métodos , Escherichia coli , Ácidos Fosfóricos/administração & dosagemRESUMO
We aimed in this study to assess whether serum-decreased bovine cumulus-oocyte complex (COC) steroidogenesis during in vitro maturation (IVM) is caused by deficient androgen milieu. For this approach, bovine COCs were cultured in serum-supplemented IVM medium in the presence or absence of 1 µM androstenedione. After 24 h of culture, medium was collected and analyzed for its content of estradiol-17ß (E2) and progesterone (P4). Medium E2 content markedly increased after incubation of COCs with androstenedione (17.52 ± 1.86 ng/ml to the androgen group; 0.32 ± 0.05 ng/ml to the non-androgen group). No significant difference in the P4 content was detected despite the presence of androstenedione (21.83 ± 1.61 ng/ml to the androgen group; 21.73 ± 1.67 ng/ml to the non-androgen group). Our data provide compelling evidence that bovine COCs steroidogenesis remains functional during culture in serum-supplemented medium and suggest that serum-induced decreased COCs estradiol secretion is caused by deficiency of an aromatizable androgen source.
Assuntos
Androstenodiona/farmacologia , Estradiol/metabolismo , Oócitos/metabolismo , Soro , Animais , Bovinos , Células Cultivadas , Meios de Cultura/química , Meios de Cultura/farmacologia , Estradiol/biossíntese , Feminino , Técnicas de Maturação in Vitro de Oócitos/métodos , Oócitos/efeitos dos fármacos , Folículo Ovariano , Progesterona/metabolismoRESUMO
The present study aimed to evaluate the survival, growth, antrum formation, oocyte extrusion, and hormone production during in vitro culture of caprine preantral follicles isolated from pure-breed (Saanen) or crossbreed (F1 generation: ½ Saanen + ½ Anglo-Nubian) goats. Secondary follicles (diameter: 150-250 µm) from Saanen or crossbreed (Saanen × Anglo-Nubian) goats were isolated from the ovarian cortex by microdissection and cultured in vitro for 18 days in α-modified minimum essential medium (α-MEM+) supplemented with vascular endothelial growth factor and increasing concentrations of follicle-stimulating hormone. Every six days follicular morphology, growth, antrum formation, and follicular extrusion were evaluated. In addition, on days 2, 6, 12, and 18 of culture the medium samples were collected and stored at -20°C for further measurement of estradiol and progesterone. The follicular survival, antrum formation, and oocyte extrusion were analyzed by the Chi square test. Follicular diameter and hormone assays were compared using the Kruskal-Wallis test. Survival rates, growth, antral follicle formation, and oocyte extrusion of preantral follicles cultured in vitro were similar between the different genetic groups. The production of estradiol and progesterone indicated the maintenance of cell viability throughout the culture. In conclusion, preantral follicles from pure-breed or crossbreed goats can be used with the same efficiency for in vitro culture of isolated caprine preantral follicles.(AU)
Assuntos
Animais , Oócitos/citologia , Estradiol/análise , Progesterona/análise , Cabras/classificaçãoRESUMO
The present study aimed to evaluate the survival, growth, antrum formation, oocyte extrusion, and hormone production during in vitro culture of caprine preantral follicles isolated from pure-breed (Saanen) or crossbreed (F1 generation: ½ Saanen + ½ Anglo-Nubian) goats. Secondary follicles (diameter: 150-250 µm) from Saanen or crossbreed (Saanen × Anglo-Nubian) goats were isolated from the ovarian cortex by microdissection and cultured in vitro for 18 days in α-modified minimum essential medium (α-MEM+) supplemented with vascular endothelial growth factor and increasing concentrations of follicle-stimulating hormone. Every six days follicular morphology, growth, antrum formation, and follicular extrusion were evaluated. In addition, on days 2, 6, 12, and 18 of culture the medium samples were collected and stored at -20°C for further measurement of estradiol and progesterone. The follicular survival, antrum formation, and oocyte extrusion were analyzed by the Chi square test. Follicular diameter and hormone assays were compared using the Kruskal-Wallis test. Survival rates, growth, antral follicle formation, and oocyte extrusion of preantral follicles cultured in vitro were similar between the different genetic groups. The production of estradiol and progesterone indicated the maintenance of cell viability throughout the culture. In conclusion, preantral follicles from pure-breed or crossbreed goats can be used with the same efficiency for in vitro culture of isolated caprine preantral follicles.