Rapid detection of bovine coronavirus by a semi-nested RT-PCR

Bovine coronavirus (BCoV) is a member of the group 2 of the Coronavirus (Nidovirales: Coronaviridae) and the causative agent of enteritis in both calves and adult bovine, as well as respiratory disease in calves. The present study aimed to develop a semi-nested RT-PCR for the detection of BCoV based on representative up-to-date sequences of the nucleocapsid gene, a conserved region of coronavirus genome. Three primers were designed, the first round with a 463bp and the second (semi-nested) with a 306bp predicted fragment. The analytical sensitivity was determined by 10-fold serial dilutions of the BCoV Kakegawa strain (HA titre: 256) in DEPC treated ultra-pure water, in fetal bovine serum (FBS) and in a BCoV-free fecal suspension, when positive results were found up to the 10-2, 10-3 and 10-7 dilutions, respectively, which suggests that the total amount of RNA in the sample influence the precipitation of pellets by the method of extraction used. When fecal samples was used, a large quantity of total RNA serves as carrier of BCoV RNA, demonstrating a high analytical sensitivity and lack of possible substances inhibiting the PCR. The final semi-nested RT-PCR protocol was applied to 25 fecal samples from adult cows, previously tested by a nested RT-PCR RdRp used as a reference test, resulting in 20 and 17 positives for the first and second tests, respectively, and a substantial agreement was found by kappa statistics (0.694). The high sensitivity and specificity of the new proposed method and the fact that primers were designed based on current BCoV sequences give basis to a more accurate diagnosis of BCoV-caused diseases, as well as to further insights on protocols for the detection of other Coronavirus representatives of both Animal and Public Health importance.

Tree-ring formation, radial increment periodicity, and phenology of tree species from a seasonal semi-deciduous forest in southeast Brazil

Many tropical tree species produce growth rings in response to seasonal environmental factors that influence the activity of the vascular cambium. We applied the following methods to analyze the annual nature of treering formation of 24 tree species from a seasonal semi-deciduous forest of southeast Brazil: describing wood anatomy and phenology, counting tree rings after cambium markings, and using permanent dendrometer bands. After 7 years of systematic observations and measurements, we found the following: the trees lost their leaves during the dry season and grew new leaves at the end of the same season; trunk increment dynamics corresponded to seasonal changes in precipitation, with higher increment (active period) during the rainy season (October-April) and lower increment (dormant period) during the dry season (May-September); the number of tree rings formed after injuries to the cambium coincided with the number of years since the extraction of the wood samples. As a result of these observations, it was concluded that most study trees formed one growth ring per year. This suggests that tree species from the seasonal semi-deciduous forests of Brazil have an annual cycle of wood formation. Therefore, these trees have potential for use in future studies of tree age and radial growth rates, as well as to infer ecological and regional climatic conditions. These future studies can provide important information for the management and conservation of these endangered forests.

Diagnosis and treatment of extraction sockets in preparation for implant placement: report of three cases

This article addresses diagnostic parameters that should be assessed in the treatment of extraction sockets with dental implant placement by presenting three case reports that emphasize the relevance of the amount of remaining bone walls. Diagnosis was based on the analysis of clinical and radiographic parameters (e.g.: bone defect morphology, remaining bone volume, presence of infections on the receptor site). Case 1 presents a 5-wall defect in the maxillary right central incisor region with severe root resorption, which was treated with immediate implant placement. Cases 2 and 3 present, respectively, two- and three-wall bone defects that did not have indication for immediate implants. These cases were first submitted to a guided bone regeneration (GBR) procedure with bone graft biomaterial and membrane barriers, and the implants were installed in a second surgical procedure. The analysis of the preoperative periodontal condition of the adjacent teeth and bone defect morphology is extremely important because these factors determine the choice between immediate implant or GBR treatment followed by implant installation in a subsequent intervention.

Bone quality associated with daily intake of coffee: a biochemical, radiographic and histometric study

Caffeine induces loss of calcium and influences the normal development of bone. This study investigated the effects of coffee on bone metabolism in rats by biochemical measurement of calcium, bone densitometry and histometry. Male rats, born of female treated daily with coffee and with coffee intake since born, were anesthetized, subjected to extraction of the upper right incisor, and sacrificed 7, 21 and 42 days after surgery. Blood and urine samples were taken, and their maxilla radiographed and processed to obtain 5-µm-thick semi-serial sections stained with hematoxylin and eosin. The volume and bone quality were estimated using an image-analysis software. The results showed significantly greater amount of calcium in the plasma (9.40 ± 1.73 versus 9.80 ± 2.05 mg%) and urine (1.00 ± 0.50 versus 1.25 ± 0.70 mg/24 h) and significantly less amount in bone (90.0 ± 1.94 versus 86.0 ± 2.12 mg/mg bone), reduced bone mineral density (1.05 ± 0.11 versus 0.65 ± 0.15 mmAL), and lower amount of bone (76.19 ± 1.6 versus 53.41 ± 2.1 %) (ANOVA; p≤0.01) in animals treated with coffee sacrificed after 42 days. It may be concluded that coffee/caffeine intake caused serious adverse effects on calcium metabolism in rats, including increased levels of calcium in the urine and plasma, decreased bone mineral density and lower volume of bone, thus delaying the bone repair process.

Cola beverage consumption delays alveolar bone healing: a histometric study in rats

Epidemiological studies have suggested that cola beverage consumption may affect bone metabolism and increase bone fracture risk. Experimental evidence linking cola beverage consumption to deleterious effects on bone is lacking. Herein, we investigated whether cola beverage consumption from weaning to early puberty delays the rate of reparative bone formation inside the socket of an extracted tooth in rats. Twenty male Wistar rats received cola beverage (cola group) or tap water (control group) ad libitum from the age of 23 days until tooth extraction at 42 days and euthanasia 2 and 3 weeks later. The neoformed bone volume inside the alveolar socket was estimated in semi-serial longitudinal sections using a quantitative differential point-counting method. Histological examination suggested a decrease in the osteogenic process within the tooth sockets of rats from both cola groups, which had thinner and sparser new bone trabeculae. Histometric data confirmed that alveolar bone healing was significantly delayed in cola-fed rats at three weeks after tooth extraction (ANOVA, p = 0.0006, followed by Tukey's test, p < 0.01). Although the results of studies in rats cannot be extrapolated directly to human clinical dentistry, the present study provides evidence that cola beverage consumption negatively affect maxillary bone formation.

Evaluation of DNA extraction protocols for Brucella abortus pcr detection in aborted fetuses or calves born from cows experimentally infected with strain 2308

The objective of the present study was to improve the detection of B. abortus by PCR in organs of aborted fetuses from infected cows, an important mechanism to find infected herds on the eradication phase of the program. So, different DNA extraction protocols were compared, focusing the PCR detection of B. abortus in clinical samples collected from aborted fetuses or calves born from cows challenged with the 2308 B. abortus strain. Therefore, two gold standard groups were built based on classical bacteriology, formed from: 32 lungs (17 positives), 26 spleens (11 positives), 23 livers (8 positives) and 22 bronchial lymph nodes (7 positives). All samples were submitted to three DNA extraction protocols, followed by the same amplification process with the primers B4 and B5. From the accumulated results for organ, the proportion of positives for the lungs was higher than the livers (p=0.04) or bronchial lymph nodes (p=0.004) and equal to the spleens (p=0.18). From the accumulated results for DNA extraction protocol, the proportion of positives for the Boom protocol was bigger than the PK (p<0.0001) and GT (p=0.0004). There was no difference between the PK and GT protocols (p=0.5). Some positive samples from the classical bacteriology were negative to the PCR and viceversa. Therefore, the best strategy for B. abortus detection in the organs of aborted fetuses or calves born from infected cows is the use, in parallel, of isolation by classical bacteriology and the PCR, with the DNA extraction performed by the Boom protocol.

Análise de um ciclone semi-estacionário na costa sul do Brasil associado a bloqueio atmosférico

Na primeira semana de maio de 2008, durante quatro dias, um ciclone em superfície permaneceu semi-estacionário na costa da região sul do Brasil. Este sistema foi responsável por chuvas e ventos fortes no Rio Grande do Sul e Santa Catarina, os quais causaram muitos danos (queda de árvores, enchentes e desabamentos). O objetivo deste trabalho é avaliar o processo de formação e entender os mecanismos responsáveis pelo lento deslocamento do ciclone, já que a maioria dos ciclones nesta região possui deslocamento mais rápido. A equação de desenvolvimento de Sutcliffe mostrou que a advecção de vorticidade absoluta ciclônica na média troposfera e a advecção de ar quente na camada entre 1000-500 hPa foram mecanismos importantes para a ciclogênese. Neste período, o intenso aquecimento diabático também contribuiu para a ciclogênese, à medida que se contrapôs ao intenso resfriamento adiabático devido aos movimentos verticais ascendentes. A advecção de vorticidade absoluta ciclônica que favoreceu a ciclogênese esteve associada a um Vórtice Ciclônico em Altos Níveis (VCAN), que se formou numa região de anomalia de vorticidade potencial. O VCAN se manteve semi-estacionário e compôs o setor norte de um bloqueio do tipo dipolo. Tal bloqueio intensificou um anticiclone em superfície, situado a sul/leste do ciclone, o que contribuiu para o ciclone se manter semi-estacionário. O movimento atípico e lento do ciclone para sul, e em alguns períodos para sudoeste, esteve associado com advecções de vorticidade absoluta ciclônica na média troposfera e de ar quente no seu setor sul. Somente quando o bloqueio em níveis médios e a anomalia de vorticidade potencial em níveis médios/altos se enfraqueceram, o ciclone em superfície se afastou da costa sul do Brasil.

Convecção linearmente organizada na área de Petrolina, semi-árido do Nordeste do Brasil: aspectos em meso e grande escala

É apresentado um estudo sobre sistemas convectivos linearmente organizados e observados por um radar meteorológico banda-C na região semi-árida do Nordeste do Brasil. São analisados três dias (27 a 29) de março de 1985, com ênfase na investigação do papel desempenhado por fatores locais e de grande escala no desenvolvimento dos sistemas. No cenário de grande escala, a área de cobertura do radar foi influenciada por um cavado de ar superior austral no dia 27 e por um vórtice ciclônico de altos níveis no dia 29. A convergência de umidade próxima à superfície favoreceu a atividade convectiva nos dias 27 e 29, enquanto que divergência de umidade próxima à superfície inibiu a atividade convectiva no dia 28. No cenário de mesoescala, foi observado que o aquecimento diurno é um fator importante para a formação de células convectivas, somando-se a ele o papel determinante da orografia na localização dos ecos. De maneira geral, as imagens de radar mostram os sistemas convectivos linearmente organizados em áreas elevadas e núcleos convectivos intensos envolvidos por uma área de precipitação estratiforme. Os resultados indicam que convergência do fluxo de umidade em grande escala e aquecimento radiativo, são fatores determinantes na evolução e desenvolvimento dos ecos na área de estudo.

ScriptLattes: an open-source knowledge extraction system from the Lattes platform

The Lattes platform is the major scientific information system maintained by the National Council for Scientific and Technological Development (CNPq). This platform allows to manage the curricular information of researchers and institutions working in Brazil based on the so called Lattes Curriculum. However, the public information is individually available for each researcher, not providing the automatic creation of reports of several scientific productions for research groups. It is thus difficult to extract and to summarize useful knowledge for medium to large size groups of researchers. This paper describes the design, implementation and experiences with scriptLattes: an open-source system to create academic reports of groups based on curricula of the Lattes Database. The scriptLattes system is composed by the following modules: (a) data selection, (b) data preprocessing, (c) redundancy treatment, (d) collaboration graph generation among group members, (e) research map generation based on geographical information, and (f) automatic report creation of bibliographical, technical and artistic production, and academic supervisions. The system has been extensively tested for a large variety of research groups of Brazilian institutions, and the generated reports have shown an alternative to easily extract knowledge from data in the context of Lattes platform. The source code, usage instructions and examples are available at http://scriptlattes.sourceforge.net/.

Vegetable-origin foam employed in dye extraction in tanning and leather processing facilities

This study addressed the use of conventional and vegetable origin polyurethane foams to extract C. I. Acid Orange 61 dye. The quantitative determination of the residual dye was carried out with an UV/Vis absorption spectrophotometer. The extraction of the dye was found to depend on various factors such as pH of the solution, foam cell structure, contact time and dye and foam interactions. After 45 days, better results were obtained for conventional foam when compared to vegetable foam. Despite presenting a lower percentage of extraction, vegetable foam is advantageous as it is considered a polymer with biodegradable characteristics.

Design, construction and evaluation of a simple pressurized solvent extraction system

This work describes the construction and testing of a simple pressurized solvent extraction (PSE) system. A mixture of acetone:water (80:20), 80 ºC and 103.5 bar, was used to extract two herbicides (Diuron and Bromacil) from a sample of polluted soil, followed by identification and quantification by high-performance liquid chromatography coupled with diode array detector (HPLC-DAD). The system was also used to extract soybean oil (70 ºC and 69 bar) using pentane. The extracted oil was weighed and characterized through the fatty acid methyl ester analysis (myristic (< 0.3%), palmitic (16.3%), stearic (2.8%), oleic (24.5%), linoleic (46.3%), linolenic (9.6%), araquidic (0.3%), gadoleic (< 0.3%), and behenic (0.3%) acids) using high-resolution gas chromatography with flame ionization detection (HRGC-FID). PSE results were compared with those obtained using classical procedures: Soxhlet extraction for the soybean oil and solid-liquid extraction followed by solid-phase extraction (SLE-SPE) for the herbicides. The results showed: 21.25 ± 0.36% (m/m) of oil in the soybeans using the PSE system and 21.55 ± 0.65% (m/m) using the soxhlet extraction system; extraction efficiency (recovery) of herbicides Diuron and Bromacil of 88.7 ± 4.5% and 106.6 ± 8.1%, respectively, using the PSE system, and 96.8 ± 1.0% and 94.2 ± 3.9%, respectively, with the SLP-SPE system; limit of detection (LOD) and limit of quantification (LOQ) for Diuron of 0.012 mg kg-1 and 0.040 mg kg-1, respectively; LOD and LOQ for Bromacil of 0.025 mg kg-1 and 0.083 mg kg-1, respectively. The linearity used ranged from 0.04 to 1.50 mg L-1 for Diuron and from 0.08 to 1.50 mg L-1 for Bromacil. In conclusion, using the PSE system, due to high pressure and temperature, it is possible to make efficient, fast extractions with reduced solvent consumption in an inert atmosphere, which prevents sample and analyte decomposition.

Estudo da mobilização de metais e elementos traços em ambientes aquáticos do semi-árido brasileiro aplicando análises de componentes principais

The spatial and temporal retention of metals has been studied in water and sediments of the Gavião River, Anagé and Tremedal Reservoirs, located in the semi-arid region, Bahia - Brazil, in order to identify trends in the fluxes of metals from the sediments to the water column. The determination of metals was made by ICP OES and ET AAS. The application of statistical methods showed that this aquatic system presents suitable conditions to move Cd2+ and Pb2+ from the water column to the sediment.

Extraction and concentration of biogenic calcium oxalate from plant leaves

The objective of this study was to extract and concentrate calcium oxalate (CaOx) crystals from plant leaves that form the above mentioned crystals. The chemical and physical studies of CaOx from plant to be performed depend on an adequate amount of the crystals. The plant used in this study was croton (Codiaeum variegatum). The leaves were ground in a heavy duty blender and sieved through a 0.20 mm sieve. The suspension obtained was suspended in distilled water. The crystals were concentrated at the bottom of a test tube. The supernatant must be washed until it is free of plant pigments and other organic substances. Biogenic CaOx crystals have well-defined and sharp peaks, indicating very high crystallinity. Moreover, the CaOx crystals were not damaged during the extraction procedure, as can be seen on the scanning electron microscope images. The porposed method can be considered efficient to extract and concentrate biogenic calcium oxalate.

Protocol for DNA extraction of Cryptosporidium spp. oocysts in fecal samples

Molecular characterization of Cryptosporidium spp.oocysts in clinical samples is useful for public health since it allows the study of sources of contamination as well as the transmission in different geographical regions. Although widely used in developed countries, in Brazil it is restricted to academic studies, mostly using commercial kits for the extraction of genomic DNA, or in collaboration with external reference centers, rendering the method expensive and limited. The study proposes the application of the modifications recently introduced in the method improving feasibility with lower cost. This method was efficient for clinical samples preserved at -20 °C for up to six years and the low number of oocysts may be overcomed by repetitions of extraction.

Power Secant Method applied to natural frequency extraction of Timoshenko beam structures

This work deals with an improved plane frame formulation whose exact dynamic stiffness matrix (DSM) presents, uniquely, null determinant for the natural frequencies. In comparison with the classical DSM, the formulation herein presented has some major advantages: local mode shapes are preserved in the formulation so that, for any positive frequency, the DSM will never be ill-conditioned; in the absence of poles, it is possible to employ the secant method in order to have a more computationally efficient eigenvalue extraction procedure. Applying the procedure to the more general case of Timoshenko beams, we introduce a new technique, named ""power deflation"", that makes the secant method suitable for the transcendental nonlinear eigenvalue problems based on the improved DSM. In order to avoid overflow occurrences that can hinder the secant method iterations, limiting frequencies are formulated, with scaling also applied to the eigenvalue problem. Comparisons with results available in the literature demonstrate the strength of the proposed method. Computational efficiency is compared with solutions obtained both by FEM and by the Wittrick-Williams algorithm.