PCR Multiplex fluorescente para detecção de bactérias em sêmen bovino

Este estudo teve como objetivo avaliar o limiar de detecção da técnica de PCR multiplex fluorescente aliada a eletroforese capilar na detecção de agentes infecciosos em amostras de sêmen experimentalmente contaminadas com concentrações decrescentes das bactérias Brucella abortus, Leptospira interrogans sorovar pomona, Campylobacter fetus e Haemophilus somnus. Amostras de sêmen bovino foram experimentalmente contaminadas com concentrações decrescentes de bactérias obtidas através de diluições seriadas na base 10 de modo a obter-se amostras contendo desde 1 vez até 10-7 bactérias/mL a partir da concentração inicial de Leptospira pomona, Brucella abortus, Campylobacter fetus e Haemophilus somnus. As diluições foram efetuadas individualmente para cada bactéria, bem como nas diferentes concentrações necessárias para a padronização do teste de multiplex PCR. As extrações de DNA de todas as soluções contendo espermatozóides e bactérias analisadas no presente estudo foram realizadas segundo protocolo descrito por Heinemann et al. (2000). Os produtos de PCR multiplex foram avaliados por eletroforese em gel de poliacrilamida 8% e separação eletroforética por sistema capilar em equipamento automático de análise de fragmentos de DNA MegaBace. Observou-se a amplificação de fragmentos de 193pb, 330pb, 400pb e 415pb a partir do DNA de B. abortus, L. pomona, H. somnus, C. fetus, respectivamente. Na análise por eletroforese capilar de produtos da PCR multiplex do DNA para detecção simultânea dos quatro patógenos observou-se a sinal de positividade até a diluição de 10-3 bactérias/mL vezes da concentração inicial da solução estoque de cada bactéria. A técnica de PCR multiplex aliada à eletroforese capilar foi usada pela primeira vez para o diagnóstico direto de quatro bactérias patogênicas no sêmen, demonstrando ser um método rápido na detecção de bactérias causadoras de doenças reprodutivas.

Polycyclic aromatic hydrocarbons (PAHs) in raw cane sugar (rapadura) in Brazil

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Assessment of maturity degree of composts from domestic solid wastes by fluorescence and fourier transform infrared spectroscopies

Methods of assessment of compost maturity are needed so the application of composted materials to lands will provide optimal benefits. The aim of the present paper is to assess the maturity reached by composts from domestic solid wastes (DSW) prepared under periodic and permanent aeration systems and sampled at different composting time, by means of excitation-emission matrix (EEM) fluorescence spectroscopy and Fourier transform infrared spectroscopy (FT-IR). EEM spectra indicated the presence of two different fluorophores centered, respectively, at Ex/Em wavelength pairs of 330/425 and 280/330 nm. The fluorescence intensities of these peaks were also analyzed, showing trends related to the maturity of composts. The contour density of EEM maps appeared to be strongly reduced with composting days. After 30 and 45 days of composting, FT-IR spectra exhibited a decrease of intensity of peaks assigned to polysaccharides and in the aliphatic region. EEM and FT-IR techniques seem to produce spectra that correlate with the degree of maturity of the compost. Further refinement of these techniques should provide a relatively rapid method of assessing the suitability of the compost to land application.

Determination of carbamate insecticides in apples, pears, and lettuce by LC with UV detector

A simple and rapid method was developed for determination of 8 carbamate insecticides and 10 of their metabolites in apples, pears, and lettuce by liquid chromatography with UV diode array detector. With this method no derivatization is needed. Carbamates not belonging to the N-methylcarbamate class and metabolites without the N-methyl group can also be determined.

Epstein-Barr Virus (EBV) detection and typing by PCR: a contribution to diagnostic screening of EBV-positive Burkitt's lymphoma

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Comparison between histochemical and immunohistochemical methods for diagnosis of sporotrichosis

Aims: To compare the efficacy of histochemical and immunohistochemical methods in detecting forms of Sporothrix schenckii in tissue. Methods: Thirty five cutaneous biopsy specimens from 27 patients with sporotrichosis were stained by histochemical haemotoxylin and eosin, periodic acid Schiff, and Gomori's methenamine silver methods and an immunohistochemical (avidin-biotin complex immunoperoxidase) (ABC) technique associated with a newly produced rabbit polyclonal antibody anti-Sporothrix schenckii. Results: A total of 29 (83%) cases were positive by the ABC method used in association with anti-Sporothrix schenckii rabbit polyclonal antibodies. Histochemical methods, using silver staining, periodic acid Schiff, and conventional haematoxylin and eosin detected 37%, 23%, and 23% of forms of S schenckii, respectively. The ABC technique was significantly more reliable than periodic acid Schiff and silver staining techniques. Conclusions: It is concluded that immunostaining is an easy and rapid method which can efficiently increase the accuracy of the diagnosis of sporotrichosis in human tissue.

Determination and persistence of methiocarb in artichokes

A simple and rapid method for the determination of methiocarb in artichokes by high-performance liquid chromatography with UV detection is described. No derivatization is needed and the limit of determination (0.01 ppm) is analogous to that of fluorometric detection. The results of trials carried out with granular and liquid formulations of this active ingredient are also reported. Immediately after treatment with the liquid formulation methiocarb residues averaged 1.47 ppm, while after treatment with the granular formulation residues were considered fortuitous. The decay rate of methiocarb residues in artichokes shows that the decrease and eventual disappearance of this active ingredient can chiefly be ascribed to the dilution effect due to head growth.

Possible effects of sodium chloride treatment on quality of effluents from Alabama channel Catfish ponds

Channel catfish ponds are treated with salt (sodium chloride) to increase chloride concentration and prevent nitrite toxicity in fish. A survey indicated that most farmers try to maintain chloride concentration of 50 to 100 mg/L in ponds by annual salt applications. Averages and standard deviations for selected water quality variables in salt-treated ponds were as follows: chloride, 87.2 ± 37.5 mg/L; total dissolved solids (TDS), 336 ± 96 mg/L; specific conductance, 512 ± 164 μmhos/cm. Maximum values were 189 mg/L for chloride, 481 mg/L for TDS, and 825 μmhos/cm for specific conductance. Good correlations between specific conductance values and both chloride and TDS concentrations suggest that specific conductance can be a rapid method for estimating concentrations of these two variables in surface water. The maximum limit for chloride concentration in Alabama streams allowed by the Alabama Department of Environmental Management is 230 mg/L. The usual recommended upper limit of TDS for protection of aquatic life in freshwater streams is 1,000 mg/L. Based on the observed relationship between TDS concentration and specific conductance in Alabama catfish ponds, 1,000 mg/L TDS corresponds to 1,733 μmhos/cm specific conductance. It is unlikely that effluents from salt-treated catfish ponds would violate the in-stream chloride standard of 230 mg/L or harm aquatic life in streams. Nevertheless, chloride concentrations in ponds should be measured before salt application as a safe guard against excessive salt application and chloride concentrations above the in-stream chloride standard.

Non-aqueous Titration of Gatifloxacin in Pharmaceutical Formulations using Perchloric Acid

An inexpensive, simple, precise and rapid method for the determination of fluoroquinolone gatifloxacin in tablets is described. The procedure is based on the use of volumetric dosage in a non-aqueous medium in glacial acetic acid with 0.1 M perchloric acid. The method validation yielded good results and included the precision, recovery and accuracy. It was also found that the excipients in the commercial tablet preparation did not interfere with the assay.

Analysis of polymerization time on abrasive wear of dental resins

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Comparison of the lactate minimum speed and the maximal lactate steady state to determine aerobic capacity in purebred Arabian horses

AIM: To compare five different protocols for estimating the lactate minimum speed (LMS) with that for estimating the maximal lactate steady state (MLSS) in Arabian horses, in order to obtain a more rapid method for monitoring aerobic capacity and prescribing training schedules. METHODS: Eight purebred Arabian horses were conditioned to exercise on a treadmill for 12 days then submitted to three to five exercise sessions to determine the MLSS. Blood samples were collected from a jugular catheter at specific intervals for measurement of lactate concentrations. The MLSS was the velocity maintained during the last 20 minutes of constant submaximal exercise, at which the concentration of lactate increased by no more than 1.0 mmol/L. The LMS test protocols (P1 - P5) included a warm-up period followed by a high-intensity gallop. The speed was then reduced to 4 m/s, and the incremental portion of the test was initiated. In P1, P2, and P3, the velocity increment was 0.5 m/s, and the duration of each incremental stage was three, five and seven minutes, respectively. In P4 and P5, the velocity increments were 1.0 and 1.5 m/s, respectively, and the duration of the stages was fixed at five minutes each. A second-degree polynomial function was fitted to the lactate-velocity curve, and the velocity corresponding to the lowest concentration of lactate was the LMS. RESULTS: Only the mean LMS determined by P1 and P2 did not differ from the velocity determined by the MLSS test (p > 0.1). There was a strong correlation (r >0.6) between P1 and the MLSS velocity. A limits of agreement plot revealed that the best agreement occurred between the MLSS test and P1 (mean bias = 0.14 m/s), followed by P2 (bias = -0.22 m/s). The lactate concentrations associated with the various LMS protocols did not differ. CONCLUSIONS: This study shows the variation between protocols of the LMS test for determining the onset of blood lactate accumulation but also reveals that, at least for Arabian horses, the P1 protocol of the LMS has good agreement with the MLSS. © 2013 Copyright New Zealand Veterinary Association.

Análise quantitativa da imagem ultra-sonográfica de testículos de bovinos jovens da raça Nelore

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Avaliação dos parâmetros de soldagem para compósitos de PPS/fibras contínuas com aplicações aeronáuticas

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Avaliação da eficácia de crioprotetores permeantes e não permeantes no descongelamento rápido e lento do sêmen canino

The aim of this study was to evaluate the association between cryoprotectants little studied in Brazil such as dimethylformamide and trehalose amid thinner, using protocols of fast and slow defrosting. Three adult Labrador Retrievier male, healthy dogs, weekly submitted to one semen collection during five-weeks period, were used. The base diluent medium used in this study was tris-citrate added with 3% of dimethylformamide + 3% glycerol (D1), 3% dimethylformamide and trehalose (D2) and 4% glycerol (D3). At defrosting, half of the semen samples from each diluent medium was defrosted by rapid method in water-bath at 75 °C for seven minutes, followed by a new immersion at 37 °C for 1 minute. The other half of the samples was defrosted by slow method, in water-bath at 37 °C for 1 minute. The semen was evaluated for sperm progressive motility and vigor, besides membrane integrity. For this, the semen samples were submitted to either hyposmotic and membrane integrity tests of the plasmatic membrane and acrosome (fluorescence). The results indicated that the use of glycerol as cryoprotector in TRIS diluter provides greater efficacy in cryopreserving spermatozoa of the canine species, when compared to dimethylformamide associated with trehalose or glycerol.

Efeito do ácido tânico purificado (tanino) sobre o crescimento in vitro de isolados de Pythium insidiosum

Pythiosis is caused by the oomycetous Pythium insidiosum and affect domestic and wild animals and man. The presence of water and vegetal material is fundamental for its life cycle in nature. The biflagellate zoospore are the infective form of this pathogen. The lesions are generally of granulomatous aspect, which frequently may be contaminated by secondary bacterial infection in skin and subcutaneous tissue. Dissemination to systemic tissues may also occur and it may be due to the spread of the pathogen from cutaneous lesions, as well as a primary source of infection. Clinical signs depend on the affected tissue. Diagnosis of pythiosis is based on the clinical manifestations, histopathological sections and culture of the pathogen. Serological tests may also be employed and more recently molecular biology has been introduced as a sensitive, specific and a rapid method for conclusive diagnosis. Treatment is often difficult and extensive surgery procedures are required, however, depending on the anatomic region and size of the lesion, such procedure is unfeasible and relapses are frequent. Due to the climate changes, which has contributed to increase the incidence of pythiosis, it is necessary the search for new therapeutic protocols