Enhanced production and organic solvent stability of a protease fromBrevibacillus laterosporus strain PAP04

A bacterial strain (PAP04) isolated from cattle farm soil was shown to produce an extracellular, solvent-stable protease. Sequence analysis using 16S rRNA showed that this strain was highly homologous (99%) to Brevibacillus laterosporus. Growth conditions that optimize protease production in this strain were determined as maltose (carbon source), skim milk (nitrogen source), pH 7.0, 40°C temperature, and 48 h incubation. Overall, conditions were optimized to yield a 5.91-fold higher production of protease compared to standard conditions. Furthermore, the stability of the enzyme in organic solvents was assessed by incubation for 2 weeks in solutions containing 50% concentration of various organic solvents. The enzyme retained activity in all tested solvents except ethanol; however, the protease activity was stimulated in benzene (74%) followed by acetone (63%) and chloroform (54.8%). In addition, the plate assay and zymography results also confirmed the stability of the PAP04 protease in various organic solvents. The organic solvent stability of this protease at high (50%) concentrations of solvents makes it an alternative catalyst for peptide synthesis in non-aqueous media.

Optimization of the production of mycorrhizal inoculum on substrate with organic fertilizer

The system for production of inoculum of arbuscular mycorrhizal fungi (AMF) using sand and vermiculite irrigated with nutrient solution is promising. However, organic amendments added to the substrate can stimulate sporulation of AMF and replace the nutrient solution. The aim of this study was to maximize the production of AMF (Acaulospora longula, Claroideoglomus etunicatum, Dentiscutata heterogama and Gigaspora albida) using selected organic substrates (vermicompost, coir dust and Tropstrato) together with sand and vermiculite. The production of spores varied among the tested AMF and according to the organic source added to the substrate. The vermicompost promoted higher sporulation of A. longula in relation to the other AMF and substrates. The Tropstrato® inhibited the sporulation of D. heterogama while the reproduction of C. etunicatum was not affected by the organic compounds. The inoculum of A. longula also showed a high number of infective propagules and promoted biomass accumulation in maize plants. The system of inoculum production using sand and vermiculite + 10% vermicompost favors the production of infective inoculum of A. longula with the fungus benefiting growth of corn plants.

Efficiency of bioaugmentation in the removal of organic matter in aquaculture systems / Eficiência da bioadição na remoção de matéria orgânica em sistemas aquaculturais

Several techniques are currently used to treat effluents. Bioaugmentation is a new bioremediation strategy and has been employed to improve effluent quality by treating the water during the production process. This technology consists basically of the addition of microorganisms able to degrade or remove polluting compounds, especially organic matter and nutrients. The objective of this study was to assess the effects of bioaugmentation on some parameters of organic matter and on the performance of juvenile tilapias in an intensive aquaculture production system. The combination of two bacterial consortiums in a complete randomized design was employed in a factorial analysis with two factors. Statistical differences between treatments were analyzed by the analysis of variance (ANOVA) and Tukey test at the 5 percent level. One of the treatments, heterotrophic bacterial supplementation, was able to reduce biochemical oxygen demand (BOD) by 23 percent, dissolved organic carbon (DOC) by 83.7 percent and phytoplanktonic biomass by 43 percent. On the other hand, no damage was done to either the physical-chemical indicators of water quality or to the growth performance of juvenile tilapias assessed in this study.

Existem diversas tecnologias para tratamento de efluentes, o processo de bioadição consiste em uma vertente da biorremediação e tem sido empregado na melhoria da qualidade dos efluentes através do tratamento da água de produção. Esta tecnologia consiste basicamente na adição de microrganismos com a capacidade de degradar ou remover compostos poluentes, especialmente matéria orgânica e nutrientes. Este estudo objetivou avaliar os efeitos da suplementação de composto bioativo sobre alguns parâmetros de matéria orgânica e de desempenho de juvenis de tilápias em um sistema intensivo de produção aquacultural. Foi empregada a combinação de dois consórcios bacterianos em delineamento inteiramente aleatorizado, em um esquema fatorial com dois fatores. As diferenças estatísticas entre os tratamentos foram analisadas por meio da análise de variância (ANOVA) e do teste de Tukey ao nível de 5 por cento. Verificou-se neste estudo, que a bioadição heterotrófica foi capaz de reduzir em 23 por cento a demanda bioquímica de oxigênio (DBO); em 83,7 por cento, o carbono orgânico dissolvido (COD); e em 43 por cento, a biomassa fitoplanctônica. Por outro lado, não se observou nenhum prejuízo com relação aos parâmetros físico-químicos de qualidade de água bem como ao desempenho de crescimento para juvenis de tilápias avaliados neste estudo.

A glass bead protocol for recovery of host cell free Ehrlichia canis and quantification by Sybr-green real-time PCR

E. canis infection of the canine cell line DH82 is a routine in studies with this bacteria. A protocol for isolation of host cell free bacteria was developed based on the use of glass beads. Improvement of infection with E. canis isolated by this method was detected by real-time PCR.

Development of a new method for diagnosis of Group B Coxsackie genome by reverse transcription loop-mediated isothermal amplification.

Background: Coxsackie B viruses (genus, Enterovirus; family, Picornaviridae) can cause aseptic meningitis, encephalitis, pleurodynia, and fatal myocarditis, and are implicated in the pathogenesis of dilated cardiomyopathy. The differentiation of the group B Coxsackieviruses into their subtypes has potential clinical and epidemiological implications. Objective: In this study, we developed a one-step, single-tube genogroup-specific reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of group B Coxsackie genomes targeting 5′ UTR region. Materials and Methods: The amplification can be obtained in less than 1 hour by incubating all the reagents in a single tube with reverse transcriptase and Bst DNA polymerase at 63°C. Detection of gene amplification could be accomplished by agarose gel electrophoresis and the monitoring of gene amplification can also be visualised with the naked eye by using SYBR green I fluorescent dye. Results: A total of 40 samples comprising 31 positive samples and 9 negative samples were used in this study for comparative evaluation. The results were compared with those from Real-Time Polymerase Chain Reaction (RT-PCR). None of the RT-PCR-positive samples were missed by RT-LAMP, thereby indicating a higher sensitivity of the RT-LAMP assay. Conclusion: Thus, due to easy operation without a requirement of sophisticated equipment and skilled personnel, the RT-LAMP assay reported here is extremely rapid, cost-effective, highly sensitive, and specific and has potential usefulness for rapid detection of non-polio enterovirus (NPEV) not only by well-equipped laboratories but also by peripheral diagnostic laboratories with limited financial resources in developing countries.

[ study on biodegradability enhancement of landfill leachate by fenton oxidation]

Leachate is one of the landfill products and also a wastewater bearing the most advers effects on the environment. Biological methods are usually employed for treatment of young leachate [1-2 years] wich is of high concentration of organic compounds with low molecular weight. However these methods are not approprate for mature leachate [5-10years] due to having high rate of compounds with molecular weight and the presence of materials resistant to biodegradation and toxic compounds. Advanced oxidation process such as Fenton reagent used in treatment or biodegrability improvement of strong wastewater. In the present study the degrability improvement of mature leachate through oxidation fenton process in bench scale and in batch reactor has been investigated. The samples have been taken from Ahwaz landfill and factors such as initial COD and BOD, pH, BOD5/COD were studies as degradability, the amount of Hydrogen Peroxide, Fe[II], optimal reaction time and optimal pH. The highest amount of COD removal was observed in PH=3-3.5 and 90 minutes of reaction time. H2O2=29700mg/land Fe2+=16500 mg/l in [Fe2+]/[H2O2]=1/14.8 molarity ratio were obtained as optimal amounts BOD5/COD was equal to 0.38. This study indicates that Fenton oxidation enhances the biodegradability of leachate

Biotransformation of 3-hydroxydibenzo-alpha-pyrone into 3, 8 dihydroxydibenzo-alpha-pyrone and aminoacyl conjugates by Aspergillus niger isolated from native shilajit

Shilajit is a panacea in Ayurveda, the Indian traditional system of medicine. The major bioactives of shilajit have been identified as dibenzo-alpha-pyrones (DBPs), its oligomers and aminoacyl conjugated derivatives. These bioactive compounds play a crucial role in energy metabolism in all animal cells including those of man. 3-hydroxydibenzo-alpha-pyrone (3-OH-DBP), a key DBP component of shilajit is converted, among other products, to another active DBP derivative, viz. 3,8-hydroxydibenzo-alpha-pyrone, 3,8(OH)2-DBP, in vivo, when its precursor is ingested. 3,8(OH)2-DBP is then involved in energy synthesis in the mitochondria in the reduction and stabilization of coenzyme Q10 in the electron transport chain. As the chemical synthesis of 3,8(OH)2-DBP is a complex, multi-step process and economically not readily viable, we envisioned the development of a process using microorganisms for bioconversion of 3-OH-DBP to 3,8(OH)2-DBP. In this study, the biotransformation of 3-OH-DBP is achieved using Aspergillus niger, which was involved in the humification process on sedimentary rocks leading to shilajit formation. A 60 percent bioconversion of 3-OH-DBP to 3,8(OH)2-DBP and to its aminoacyl derivatives was achieved. The products were characterized and estimated by high performance liquid chromatography (HPLC), high performance flash chromatography (HPFC) and gas chromatography-mass spectrometry (GC-MS) analyses. Among the Aspergillus species isolated and identified from native shilajit, A. niger was found to be the most efficient for this bioconversion.

Short-term Efficacy of Topical Immunosuppressive Agents on the Survival of Cultivated Allo-Conjunctival Equivalents

PURPOSE: To investigate the short-term efficacy of topical immunosuppressive agents on the survival of cultivated allo-conjunctival equivalents. METHODS: Twenty-five eyes of New Zealand white rabbits were included. Temporal conjunctivae were trephined to a diameter of 7.5 mm, and then cultured allo-conjunctival epithelial cells on amniotic membrane were transplanted onto them. Various immunosuppressants including steroid, cyclosporine, and rapamycin were applied topically four times a day for a week. Epithelial defects and graft edema were graded daily. Numbers of inflammatory cells were measured in H&E. PKH26 and cytokeratin 4 and 7 were immunostained. RESULTS: Earlier epithelialization was observed in 1% steroid-treated eyes and defects persisted significantly in 0.5% CsA applied eyes. In histology, PKH26 positive cells considered as donor cells were only found in 1% steroid or 0.01% rapamycin applied eyes. 1% steroid- or 0.01% rapamycin-applied eyes both showed positive staining for keratin-4 and -7. Inflammatory cells were less found in 1% steroid or 0.01% rapamycin treated eyes. CONCLUSIONS: Topical steroid or rapamycin can help to suppress acute inflammation and enhance the acute survival of transplanted conjunctival cells.

Bacterial growth and DOC consumption in a tropical coastal lagoon

Os objetivos desta pesquisa foram: determinar o principal nutriente limitante ao crescimento bacteriano na lagoa Imboassica, estimar a porcentagem de Carbono Orgânico Dissolvido (COD) disponível para o crescimento bacteriano e determinar a Eficiência de Crescimento Bacteriano (ECB) da comunidade bacteriana. O crescimento bacteriano e o consumo de COD foram avaliados em experimentos de culturas de diluição, nos quais nitrogênio e fósforo foram adicionados às amostras de água, juntos ou separadamente, e um controle foi preparado sem adições de nutrientes. Quando adicionados juntos, N e P estimularam um maior crescimento e produção bacterianos, assim como maiores taxas de consumo de COD. ECB e taxas de consumo de COD foram fortemente dependentes do método utilizado para determinar a produção bacteriana. ECB variou de 11% a 72%, porém apenas uma pequena fração do COD total foi consumida pelas bactérias planctônicas (de 0,7% a 3,4%). Sugere-se que as baixas disponibilidades de fósforo e nitrogênio são os principais fatores responsáveis pela baixa utilização de COD pelas bactérias na lagoa Imboassica.

The effect of the size of particles on mineralization of Oxycaryum cubense (Poepp. & Kunth) Lye

Ensaios foram realizados para avaliar os efeitos de tamanho dos detritos na mineralização de uma macrófita aquática, a Oxycaryum cubense. Foram coletadas amostras de planta e de água de uma lagoa marginal, a lagoa do Infernão (21° 35' S e 47° 51' W), localizada na planície de inundação do rio Mogi Guaçu. As plantas foram levadas ao laboratório, lavadas em água corrente, secas (50 °C) e fracionadas em seis grupos de acordo com o tamanho: 100, 10, 1,13, 0,78, 0,61 e 0,25 mm. Foram preparadas câmaras de decomposição adicionando aproximadamente 1,0 g de fragmentos de planta em 4,1 L de água da lagoa. Na seqüência, as incubações foram aeradas e as concentrações de oxigênio dissolvido, o pH, a condutividade elétrica e a temperatura foram monitorados durante 120 dias. A ocorrência de processos de anaeróbios foi evitada por reoxigenação das soluções. Os resultados experimentais foram ajustados a um modelo cinético de primeira ordem e o consumo de oxigênio dissolvido decorrente da mineralização foi obtido. De maneira geral, o processo físico de fragmentação não tende a envolver maiores quantidades de oxigênio durante a mineralização aeróbia dos detritos, significando que a fragmentação não interfere no balanço de oxigênio dissolvido deste sistema aquático; entretanto favoreceu a acidificação e também a liberação de íons dissolvidos na lagoa do Infernão.

Seasonal variations in compostability and production of vermiprotein by Eisenia fetida.

The potential of E. fetida to degrade wastes into vermicompost and to produce vermiprotein in the form of worm-biomass during different seasons was evaluated. Results revealed that the environmental factors prevailing during different seasons did influence directly the life activities of the worm and indirectly the compostability of the wastes. Feeding activities of E. fetida reduced the time of production of an efficient organic pool with energy reserves as vermicompost. Further, the amount of vermicompost produced by the worm activity depended primarily on the environmental factors and secondarily on the nature of organic wastes.

Importância de compostos organossulfurados em sementes de leguminosas e em outros vegetais / Importance of organosulfur compounds in seeds of legumes and in other vegetables

Os compostos organossulfurados, representados pelos peptídeos ´GAMA`-glutamilsulfurados, são encontrados em grande quantidade em vegetais das famílias Alliaceae, Cruciferae e Leguminosae. A seus produtos de metabolização se atribui propriedades organolépticas típicas e quando são ingeridos por mamíferos, causam efeitos biológicos diversos. Estes efeitos são diferentes nestas três famílias, devido à formação de metabólitos secundários que dependem da presença ou ausência de enzimas específicas nestes vegetais. Na família Alliaceae, representada pelo alho e pela cebola, são produzidos compostos sulfurados, que além de serem precursores de aroma e de flavor característicos após a ruptura do tecido vegetal e a ação enzimática, se tornaram conhecidos pelos efeitos benéficos para a saúde humana. A via metabólica percorrida pelos compostos organossulfurados na família Cruciferae, dá origem, além do flavor, a compostos secundários diversos, incluindo o metanotiol e dissulfetos, altamente tóxicos para ruminantes, causando anemia hemolítica. Na família Leguminosae, os dipeptídeos sulfurados são encontrados principalmente nas sementes, sendo que o feijão Phaseolus vulgaris, L., dentre os gêneros estudados, os possui em maior concentração. A ´BETA’-glutamil-S-metil-L-cisteina e a S-metil-L-cisteina livre, podem em conjunto, representar ate ´50 POR CENTO1’ do conteúdo total de metionina presente no feijão, desaparecendo somente por ocasião da sua germinação...

Nitrógeno, fósforo y cociente C/N en los sedimentos superficiales de la laguna de Chacopata, Sucre, Venezuela / Nitrogen, phosphorus and the C/N ratio in superficial sediments of Chacopata lagoon from Sucre, Venezuela

The basal behaviour and relationship with organic carbon (Corg) content and prevailing granulometric fractions, of organic nitrogen (Norg), total phosphorus (P-total) and C/N ratio in surface sediments of Chacopata lagoon, Sucre State, Venezuela, were studied. Concentrations and spatial gradients were determined in sixteen stations following a common method for marine sediments. Norg concentrations varied between 0.102 and 0.510% (x = 0.237%), total phosphorus between 0.012 and 0.094% (x = 0.058%) and C/N ratio between 9.27 and 44.47 (x = 20.53). The higher contents of Norg and total phosphorus are from sections with an abundance of mangroves, marine phanerogams, macroalgae, benthonic biomass and migratory birds shelters. The C/N ratio shows the typical values for carbonated sediments, indicating that the nitrogenous compounds are rapidly degraded and the organic matter presents itself as humic substances. This parameter was moderately associated with silt and clay, and showed antagonism with sand, whereas Norg and phosphorus showed no correlation with them. The organic contribution is purely autochthonous: no anthropogenous contributions were found.