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.

Rapid spot test analysis for the detection of urotropine in pharmaceutical preparations

The widespread falsification and/or adulteration of commercially available pharmaceutical preparations call for reliable methods of drug identification, preferably through selective and rapid sorting color tests that could be undertaken with minimum equipment remote from laboratory facilities. The present work deals with a convenient adaptation and refinement of a spot test devised by Feigl (1966) for urotropine, based on the hydrolytic cleavage of that substance in the presence of sulfuric acid, splitting out formaldehyde which is identified by its color reaction with chromotropic acid. A simple emergency kit was developed for the quick, efficient, inexpensive and easy performance of urotropine tests by semiskilled personnel even in the drugstore laboratory (or office) as well as in a mobile screening operation. It is shown that when the reagents are added according to the recommended sequence a self-heating system is generated, increasing substantially the reactions' rates and the test sensitivity as well. The identification limit found was 25 mug of urotropine, for both solid and liquid samples. The possible interference of 84 substances/materials was investigated. Interference was noted only for methylene blue, acriflavine, Ponceau Red, Bordeaux Red (these dyes are often included in urotropine dosage forms), pyramidone, dipyrone, quinine and tetracycline. A simple procedure for removing most of the interferences is described. Data for 8 commercial dosage forms and results obtained from their analysis are presented.

RAPID COAGGLUTINATION TEST FOR THE DETECTION AND TYPING OF FOOT-AND-MOUTH-DISEASE VIRUS

Protein A containing Staphylococcus aureus was used to develop a coagglutination (COA) test for the detection and typing of foot and mouth disease virus (FMDV) O, A and C serotypes in infected cells and tissues. Different batches and amounts of guinea pig anti-FMDV sera were assessed to optimize the preparation of COA conjugates. The sensitivity and specificity of the COA Test for the detection of FMDV O, A and C serotypes and heterologous viruses was also characterized. Comparison between the COA Test and complement fixation test for the detection and typing of FMDV obtained from extracts of tongue epithelial tissues from infected cattle revealed high agreement in the results and indicated a potential application of the COA Test for the direct diagnosis of viruses.

Detecção de Leptospira pomona em sêmen bovino por eletroforese capilar fluorescente

This study was performed in order to evaluate the detection limit of PCR with fluorescent capillary electrophoresis for Leptospira pomona diagnosis in bovine semen. Negative bovine semen samples were artificially contaminated with Leptospira pomona (10 0 to 10 7 bacteria/ ml) and DNA was extracted by phenol/chloroform protocol. DNA fragments visualization was done by three electrophoresis methods: under UV light in 2 % agarose gel, silver staining 8% polyacrylamide gel and fluorescent capillary electrophoresis. The detection limit of capillary electrophoresis for Leptospira pomona was 10 2bacteria/ml. Under UV light, in 2 % agarose gel, the detection limit was of 10 4 bacteria/ ml while for silver stained 8 % polyacrylamide gel it was 10 2 bacteria/ ml. PCR with fluorescent capillary electrophoresis is an efficient and rapid diagnostic test for DNA detection of Leptospira in bovine semen and this can be an important tool for herd and semen sanitary control in artificial insemination centers.

Comparative evaluation of several methods for Canine Visceral Leishmaniasis diagnosis

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

Validation of a Leishmania infantum ELISA rapid test for serological diagnosis of Leishmania chagasi in dogs

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Comparison between 100-g glucose tolerance test and two other screening tests for gestational diabetes: combined fasting glucose with risk factors and 50-g glucose tolerance test

CONTEXTO E OBJETIVO: A falta de consenso sobre os protocolos de rastreamento e diagnóstico do diabetes gestacional, associada às dificuldades na realização do teste oral simplificado do diabete gestacional (o teste de tolerância a 100 g de glicose, considerado padrão-ouro) justificam a comparação com alternativas. O objetivo deste trabalho é comparar o teste padrão-ouro a dois testes de rastreamento: associação de glicemia de jejum e fatores de risco (GJ + FR) e o teste oral simplificado de tolerância a 50 g de glicose (TTG 50 g), com o teste de tolerância a 100 g de glicose (TTG 100 g). TIPO DE ESTUDO E LOCAL: Estudo de coorte longitudinal, prospectivo, realizado no Serviço de Ginecologia e Obstetrícia do Hospital Universitário da Universidade Federal de Mato Grosso do Sul. MÉTODOS: 341 gestantes foram submetidas aos três testes. Calcularam-se os índices de sensibilidade (S), especificidade (E), valores preditivos (VPP e VPN), razões de probabilidade (RPP e RPN) e resultados falsos (FP e FN), positivos e negativos da associação GJ + FR e do TTG 50 g em relação ao TTG 100 g. Compararam-se as médias das glicemias de uma hora pós-sobrecarga (1hPS) com 50 e 100 g. Na análise estatística, empregou-se o teste t de Student, com limite de significância de 5%. RESULTADOS: A associação GJ + FR encaminhou mais gestantes (53,9%) para a confirmação diagnóstica que o TTG 50 g (14,4%). Os dois testes foram equivalentes nos índices de S (86,4 e 76,9%), VPN (98,7 e 98,9%), RPN (0,3 e 0,27) e FN (15,4 e 23,1%). As médias das glicemias 1hPS foram semelhantes, 106,8 mg/dl para o TTG 50 g e 107,5 mg/dl para o TTG 100 g. CONCLUSÕES: Os resultados da eficiência diagnóstica associados à simplicidade, praticabilidade e custo referendaram a associação GJ + FR como o mais adequado para o rastreamento. A equivalência das glicemias de 1hPS permitiram a proposição de um novo protocolo de rastreamento e diagnóstico do diabete gestacional, com menores custo e desconforto.

Refining the intestinal motility test in mice to reduce animal stress

A new protocol using 3-h fast animal for intestinal motility test was developed in our laboratory aiming the 3R's concept to reduce the stress of animals. Our results may aid in formulating recommendations that can be included in revised guidelines with regard to fasting time of mice.

Relationship between gingival clinical parameters and the reactivity of the BANA test in subgingival samples from children.

Gingivitis is the first manifestation of periodontal disease, and is characterized by painless and slow evolution. Early diagnosis and intervention must be done to avoid the possibility of precocious periodontitis during the childhood or teenage years. The enzymatic BANA test (N-benzoyl-DL-arginine-naphthylamide) was used to evaluate subgingival samples from 54 children between 6 and 9 years of age. Plaque index (PI) and gingival index (GI) were assessed according to the criteria recommended by Löe (1967). Subgingival plaque was collected from the region that featured the greatest periodontal alteration, represented by a higher gingival index. Resulting data were grouped individually according to visible and non-visible plaque and bleeding and non-bleeding gingiva. Results showed that there was no statistically significant correlation between the presence of visible plaque and the positivity of the BANA test, nor was there a statistically significant correlation between the presence of bleeding and the positivity of the BANA test in subgingival samples obtained from children. This study concluded that the BANA test is not an ideal diagnostic test to be applied to children.

Pressões inspiratórias como preditores de sucesso na extubação em prematuros de muito peso

Pós-graduação em Pediatria - FMB

Comparison of diferent diagnostic tests in dogs uninfected and naturally infected with visceral leishmaniasis

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

Taenia saginata: Production and characterization of monoclonal antibodies against Taenia saginata metacestode antigens

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

The safe motherhood referral system to reduce cesarean sections and perinatal mortality - a cross-sectional study [1995-2006]

Background: In 2000, the eight Millennium Development Goals (MDGs) set targets for reducing child mortality and improving maternal health by 2015.Objective: To evaluate the results of a new education and referral system for antenatal/intrapartum care as a strategy to reduce the rates of Cesarean sections (C-sections) and maternal/perinatal mortality.Methods: Design: Cross-sectional study. Setting: Department of Gynecology and Obstetrics, Botucatu Medical School, São Paulo State University/UNESP, Brazil. Population: 27,387 delivering women and 27,827 offspring. Data collection: maternal and perinatal data between 1995 and 2006 at the major level III and level II hospitals in Botucatu, Brazil following initiation of a safe motherhood education and referral system. Main outcome measures: Yearly rates of C-sections, maternal (/100,000 LB) and perinatal (/1000 births) mortality rates at both hospitals. Data analysis: Simple linear regression models were adjusted to estimate the referral system's annual effects on the total number of deliveries, C-section and perinatal mortality ratios in the two hospitals. The linear regression were assessed by residual analysis (Shapiro-Wilk test) and the influence of possible conflicting observations was evaluated by a diagnostic test (Leverage), with p < 0.05.Results: Over the time period evaluated, the overall C-section rate was 37.3%, there were 30 maternal deaths (maternal mortality ratio = 109.5/100,000 LB) and 660 perinatal deaths (perinatal mortality rate = 23.7/1000 births). The C-section rate decreased from 46.5% to 23.4% at the level II hospital while remaining unchanged at the level III hospital. The perinatal mortality rate decreased from 9.71 to 1.66/1000 births and from 60.8 to 39.6/1000 births at the level II and level III hospital, respectively. Maternal mortality ratios were 16.3/100,000 LB and 185.1/100,000 LB at the level II and level III hospitals. There was a shift from direct to indirect causes of maternal mortality.Conclusions: This safe motherhood referral system was a good strategy in reducing perinatal mortality and direct causes of maternal mortality and decreasing the overall rate of C-sections.

Evaluation of cytologic and biochemical variables in blood, plasma, and peritoneal fluid from calves before and after umbilical herniorrhaphy

Objective-To establish reference intervals for cytologic and biochemical variables in peritoneal fluid, whole blood, and plasma in calves with congenital umbilical hernias (CUHs) before and after herniorrhaphy and to assess whether those variables in calves with CUHs were altered, compared with findings in clinically normal calves.Animals-20 Holstein calves with or without a CUH.Procedures-10 calves with CUHs underwent herniorrhaphy. Blood and peritoneal fluid samples from all 20 calves were collected for cytologic and biochemical analyses on days 0 (before surgery), 1, 3, 5, 7, and 15. Data from the 2 groups were compared.Results-Reference intervals for the variables of interest were established for each group, Before surgery, calves with CUHs had significantly greater plasma total protein concentration and creatine kinase (CK) and aspartate aminotransferase activities and peritoneal fluid specific gravity values, compared with values for calves without CUHs. At various time points after surgery, peritoneal fluid total protein concentration; fibrinogen concentration; nucleated cell, polymorphonuclear cell, and lymphocyte counts; specific gravity; and lactate dehydrogenase, aspartate aminotransferase, and CK activities in calves with CUHs were significantly different from values in calves without CUHs. Some plasma and blood variables leg, total protein concentration, neutrophil count, and CK activity were significantly different between the 2 groups.Conclusions and Clinical Relevance-Values of certain cytologic and biochemical variables in peritoneal fluid, blood, and plasma were different between calves with and without CUHs. Thus, determination of reference intervals for these variables is important for interpreting diagnostic test results in calves with CUHs. (Am J Vet Res 2009;70:423-432)