Detection of bovine Herpesvirus type 5 in formalin-fixed, paraffin-embedded bovine brain by PCR: a useful adjunct to conventional tissue-based diagnostic test of bovine encephalitis

The aim of this study was to evaluate the application of PCR technique for the detection of BoHV-5 in routinely formalin-fixed, paraffin-embedded brain tissues in 20 naturally infected calves affected by fatal meningoencephalitis. Brains were divided into two halves, one kept fresh for virus isolation and PCR assay, targeting the glycoprotein C gene from BoHV-5 genome. The other half brain, corresponding to posterior cortex region, was submitted to formalin fixation and embedded into paraffin blocks for microscopic evaluation and total DNA isolation. Most of the slides showed severe multifocal non-supurative encephalitis with neuronal degeneration, neurophagia, and no acidophilic intranuclear inclusions could be found in neurons and glial. The 20 fresh samples were confirmed, by virus isolation and PCR assay, as having the BoHV-5 virus and, respective glicoprotein C sequence, while 15 of 20 formalin-fixed, paraffin-embedded samples were considered positive for the same analysis. The results revealed the first description of PCR efficiency, applied to formalin-fixed, paraffin-embedded brain collected from naturally infected calves, improving the detection of BoHV-5 from archival samples in South America. (c) 2007 Published by Elsevier B.V.

Chicken embryo related (CER) cell line for quantification of rabies neutralizing antibody by fluorescent focus inhibition test

Levels of rabies virus neutralization antibody in sera from vaccinated dogs and cattle were either measured by mouse neutralization test (MNT) or by rapid fluorescent focus inhibition test (RFFIT), performed on CER monolayers. The two tests were compared for their ability to detect the 0.5 International Units/ml (I.U.) recommended by the World Health Organization (WHO) as the minimum response for proof of rabies immunization. A significant correlation was found between the two tests (n = 211; r = 0.9949 in dogs and 0.9307 in cows, p < 0.001), good sensitivity (87.5%), specificity (94.7%) and agreement (96.6%) as well. RFFIT method standardized on CER cell system for neutralizing antibodies detection turns the diagnosis easier and less expensive, specially when a great number of samples must be tested from endemic areas as commonly found in Brazil. (c) 2005 the International Association for Biologicals. Published by Elsevier Ltd. All rights reserved.

Comparative evaluation of an indirect ELISA test for diagnosis of swine cysticercosis employing antigen from Taenia solium and Taenia crassiceps metacestodes

Taenia solium cysticercosis is still a serious public health problem in several countries where poverty and lack of hygiene favor transmission. Because pigs are the primary intermediate hosts, prevalence of porcine cysticercosis is a reliable indicator of active transmission zones. Serological diagnostic methods are important tools for epidemiological studies since they can be applied to living animals on a large scale. Four antigen preparations (cyst fluid and crude) from T. solium and T. crassiceps metacestodes were compared for swine cysticercosis diagnosis by indirect ELISA (IE). Twenty-eight serum samples from swine naturally and experimentally infected by cysticerci of T. solium and 56 serum samples from swine reared in commercial herds were tested. Best results of overall sensitivity were obtained by the use of cyst fluid and crude antigen of T. crassiceps metacestode (100 and 96.4%, respectively). Using homologous antigen preparations we have observed higher specificity percentage (98.2% for cyst fluid and 96.4% for crude metacestode T. solium antigen). We concluded that sensitivity is of far more importance than specificity for identification of endemic areas in order to prevent transmission to man. We conclude, therefore, that IE performed with cyst fluid antigen of T. crassiceps metacestode is a better tool for that purpose. (C) 2000 Elsevier B.V. B.V. All rights reserved.

Comparison between RT-PCR and the mouse inoculation test for detection of rabies virus in samples kept for long periods under different conditions

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

Water deprivation-induced sodium appetite and differential expression of encephalic c-Fos immunoreactivity in the spontaneously hypertensive rat

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

Damage to the central amygdala produces differential encephalic c-fos expression in the water deprivation-partial rehydration protocol

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

Bond strength of different endodontic sealers to dentin: push-out test

Objective: The aim of this in vitro study was to evaluate the bond strength of different root canal sealers to dentin. Material and Methods: Forty extracted single-rooted human teeth were examined and the coronal and middle thirds of the canals were prepared with a 1.50 mm post drill (FibreKor Post System, Pentron). The teeth were allocated in two experimental groups, irrigated with 2.5% NaOCl+17% EDTA or saline solution (control group) and instrumented using Race rotary files (FKG) to a size #40 at the working length. Then, the groups were divided into four subgroups and filled with Epiphany sealer (Group 1), EndoREZ (Group 2), AH26 (Group 3) and Grossman's Sealer (Group 4). After 2 weeks of storage in 100% humidity at 37 degrees C, all teeth were sectioned transversally into 2-mm-thick discs. Push-out tests were performed at a cross-head speed of 1 mm/min using a universal testing machine. The maximum load at failure was recorded and expressed in MPa. Results: Means (+/- SD) in root canals irrigated with 2.5% NaOCl and 17% EDTA were: G1 (21.6 +/- 6.0), G2 (15.2 +/- 3.7), G3 (14.6 +/- 4.5) and G4 (11.7 +/- 4.1). Two-way ANOVA and Tukey's test showed the highest bond strength for the Epiphany's group (p < 0.01) when compared to the other tested sealers. Saline solution decreased the values of bond-strength (p < 0.05) for all sealers. Conclusion: Epiphany sealer presented higher bond strength values to dentin in both irrigating protocols, and the use of 2.5% NaOCl and 17% EDTA increased the bond strength values for all sealers.

A physiological manoeuvre to improve the positivity of the gastro-oesophageal reflux scintigraphic test

One of the best examinations used routinely to detect gastro-oesophageal reflux (GOR) is scintigraphy and attempts are continuously being made to improve the performance of the test. We have tested a new manoeuvre to improve the results. Three hundred and twenty eight patients (190 males and 138 females, aged 1 month to 84 years, average 4.47 years) were studied after a 12 h fast period with 100 MBq of Tc-99(m)-Phytate, 50 ml volume, directly delivered into the stomach in a mixture of orange juice. All patients were strongly suspected of having GOR. Conventional acquisition was done for 20 min, at a rate of one frame every 20 s. in the middle of the examination, small children were put in the upright position and held there for a few seconds or released in a way that allowed movement; adults were asked to stand up and walk a few metres. The test was then continued for the remaining 10 min. The global index of positivity was 64.6% (76 positive cases in the first half of the examination and 134 positive cases only after the manoeuvre). The mean of GOR episodes observed in the positive cases without the manoeuvre was 0.5 per patient, significantly different from the mean of 1.59 noted after the manoeuvre (P<0.01). In 28 cases the manoeuvre increased either the intensity or the frequency of reflux previously detected without it. We believe that this manoeuvre should be performed in all scintigraphic tests aiming to detect GOR. In the present series, the manoeuvre increased the frequency of GOR episodes. ((C) 2001 Lippincott Williams & Wilkins)].

Evaluation of antimutagenic activity and mechanisms of action of beta-glucan from barley, in CHO-k1 and HTC cell lines using the micronucleus test

Due to the need to identify new antimutagenic agents and to determine their mechanism of action, the present study examined the mechanism of action of the P-glucan with regard to antimutagenicity using the micronucleus assay in CHO-kl and HTC cell lines. The mutagenicity experiments were performed with three different concentrations of P-glucan (5, 10, and 20 mu g/mL), in wich only the highest dose showed mutagenic activity. In the antimutagenicity experiments, the same concentrations of P-glucan were combined with a mutagenic agent, methylmethane sulfonate, or 2-aminoanthracene, using four different treatment protocols: pre-treatment, simultaneous treatment (simple and with pre-incubation), and post-treatment. The results indicate that the CHO-kl cell line treated with MMS presented a chemopreventive activity for all the doses of P-glucan in the different treatment protocols, except for the lowest dose in post-treatment. When HTC cell line treated with MMS is analysed, a chemopreventive activity can be verified for the highest dose in both pre- and post-treatment. For the simple simultaneous treatment, the three doses demonstrated efficacy, while for the simultaneous treatment with pre-incubation only the intermediate concentration was effective. In HTC treated with 2AA both the lowest dose in the pre-treatment protocol and the post-treatment protocol did not show efficacy in preventing DNA damage. The evaluation of the different protocols and the damage decrease percentages observed suggest that P-glucan has both desmutagenic and bioantimutagenic activity. It is necessary, however, to note that efficacy and mechanism of action are subject to variation when compared the two cell lines, since in HTC, representing a drug-metabolizing system, this substance can show a diminished chemopreventive capacity. (c) 2006 Elsevier Ltd. All rights reserved.

Micronucleus test and observation of nuclear alterations in erythrocytes of Nile tilapia exposed to waters affected by refinery effluent

Micronuclei and nuclear alterations tests were performed on erythrocytes of Oreochromis niloticus (Perciformes, Cichlidae) in order to evaluate the water quality from Paraiba do Sul river, in an area affected by effluents from an oil shale processing plant, located in the city of Sao Jose dos Campos, Brazil-SP. Water samples were collected on 2004 May and August (dry season) and on 2004 November and 2005 January (rain season), in three distinct sites, comprising 12 samples. It was possible to detect substances of clastogenic and/or aneugenic potential, as well as cytotoxic substances, chiefly at the point corresponding to the drainage of oil shale plant wastes along the river. The highest incidence of micronuclei and nuclear alterations was detected during May and August, whereas the results obtained in November and January were insignificant. This work shows that the effluent treatment provided by the oil shale plant was not fully efficient to minimize the effect of cytotoxic and mutagenic substances in the test organism surveyed. (c) 2006 Elsevier B.V. All rights reserved.

Allium cepa test in environmental monitoring: A review on its application

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

Effect of soccer training on the running speed and the blood lactate concentration at the lactate minimum test

Tegtbur et al. [23] devised a new method able to estimate the intensity at maximal lactate steady state termed lactate minimum test. According to Billat et al. [7], no studies have yet been published on the affect of training on highest blood lactate concentration that can be maintained over time without continual blood lactate accumulation. Therefore, the aim of the present study was to verify the effect of soccer training on the running speed and the blood lactate concentration (BLC) at the lactate minimum test (Lac(min)). Thirteen Brazilian male professional soccer players, all members of the same team playing at National level, volunteered for this study. Measurements were carried out before (pre) and after (post) eight weeks of soccer training. The Lac(min) test was adapted to the procedures reported by Tegtbur et al. [23]. The running speed at the Lac(min) test was taken when the gradient of the line was zero. Differences in running speed and blood lactate concentration at the Lac(min) test before (pre) and after (post) the training program were evaluated by Student's paired t-test. The training program increased the running speed at the Lac(min) test (14.94 +/- 0.21 vs. 15.44 +/- 0.42* km(.)h(-1)) and the blood lactate concentration (5.11 +/- 2.31 vs. 6.93 +/- 1.33* mmol(.)L(-1)). The enhance in the blood lactate concentration may be explained by an increase in the lactate/H+ transport capacity of human skeletal muscle verified by other authors.

Determination of anaerobic threshold in rats using the lactate minimum test

The break point of the curve of blood lactate vs exercise load has been called anaerobic threshold (AT) and is considered to be an important indicator of endurance exercise capacity in human subjects. There are few studies of AT determination in animals. We describe a protocol for AT determination by the lactate minimum test in rats during swimming exercise. The test is based on the premise that during an incremental exercise test, and after a bout of maximal exercise, blood lactate decreases to a minimum and then increases again. This minimum value indicates the intensity of the AT. Adult male (90 days) Wistar rats adapted to swimming for 2 weeks were used. The initial state of lactic acidosis was obtained by making the animals jump into the water and swim while carrying a load equivalent to 50% of body weight for 6 min (30-s exercise interrupted by a 30-s rest). After a 9-min rest, blood was collected and the incremental swimming test was started. The test consisted of swimming while supporting loads of 4.5, 5.0, 5.5, 6.0 and 7.0% of body weight. Each exercise load lasted 5 min and was followed by a 30-s rest during which blood samples were taken. The blood lactate minimum was determined from a zero-gradient tangent to a spline function fitting the blood lactate vs workload curve. AT was estimated to be 4.95 ± 0.10% of body weight while interpolated blood lactate was 7.17 ± 0.16 mmol/l. These results suggest the application of AT determination in animal studies concerning metabolism during exercise.

Spectral analysis of the electromyograph of the erector spinae muscle before and after a dynamic manual load-lifting test

The aim of the present study was to assess the spectral behavior of the erector spinae muscle during isometric contractions performed before and after a dynamic manual load-lifting test carried out by the trunk in order to determine the capacity of muscle to perform this task. Nine healthy female students participated in the experiment. Their average age, height, and body mass (± SD) were 20 ± 1 years, 1.6 ± 0.03 m, and 53 ± 4 kg, respectively. The development of muscle fatigue was assessed by spectral analysis (median frequency) and root mean square with time. The test consisted of repeated bending movements from the trunk, starting from a 45º angle of flexion, with the application of approximately 15, 25 and 50% of maximum individual load, to the stand up position. The protocol used proved to be more reliable with loads exceeding 50% of the maximum for the identification of muscle fatigue by electromyography as a function of time. Most of the volunteers showed an increase in root mean square versus time on both the right (N = 7) and the left (N = 6) side, indicating a tendency to become fatigued. With respect to the changes in median frequency of the electromyographic signal, the loads used in this study had no significant effect on either the right or the left side of the erector spinae muscle at this frequency, suggesting that a higher amount and percentage of loads would produce more substantial results in the study of isotonic contractions.

Predicting MAOD Using Only a Supramaximal Exhaustive Test

The objective of this study was to propose an alternative method (MAOD(ALT)) to estimate the maximal accumulated oxygen deficit (MAOD) using only one supramaximal exhaustive test. Nine participants performed the following tests: (a) a maximal incremental exercise test, (b) six submaximal constant workload tests, and (c) a supramaximal constant workload test. Traditional MAOD was determined by calculating the difference between predicted O(2) demand and accumulated O(2) uptake during the supramaximal test. MAOD(ALT) was established by summing the fast component of excess post-exercise oxygen consumption and the O(2) equivalent for energy provided by blood lactate accumulation, both of which were measured during the supramaximal test. There was no significant difference between MAOD (2.82 +/- 0.45 L) and MAOD(ALT) (2.77 +/- 0.37 L) (p = 0.60). The correlation between MAOD and MAOD(ALT) was also high (r = 0.78; p = 0.014). These data indicate that the MAOD(ALT) can be used to estimate the MAOD.