Egg quality in laying hens exposed to Mycoplasma gallisepticum F-strain attenuated vaccine.

Mycoplasma gallisepticum causes coughing, ocular and nasal discharge, reduction in feed intake, lower and uneven growth, decline in egg production and quality, and increase in mortality. Among the attenuated vaccination strains, MGF can reduce clinical signs and lesions in layer hens, stimulate immune responses of cellular and humoral basis, act as an instrument of competitive exclusion in relation to field strains, and reduce the use of antimicrobials. This study aimed to investigate the effects of attenuated MG F-strain vaccination on egg quality in 3 groups of 30 hens each, being one control and 2 vaccinated. Vaccination was applied by ocular route at 8 and 12 wk of age. Comparisons were made among unvaccinated hens; vaccinated at 8 wk of age; and vaccinated at 8 and 12 wk of age. There were no statistical differences in eggshell thickness and weight among groups. Eggs from twice vaccinated birds yielded a Haugh unit significantly lower than the other groups without affecting egg classification. There was no significant difference in ELISA results between the vaccinated groups.

Improving sensitivity of cervical cytology by removal of cervical secretions before sampling: a prospective study in Mexico.

UNLABELLED: Sensitivity of cervical cytology is suboptimal, especially in developing countries such as Mexico, despite available guidelines aimed at improving this. When obtaining cervical samples, whether the samples are taken from the transformation zone and whether abnormal cells are missing must be considered. Cervical secretions (CS) are always present in variable proportions, and when cleaning the cervix, better samples may be obtained. In this study, we analyzed samples obtained with or without cleaning the cervix, and compared their contents in order to determine the sensitivity and specificity of these two methods. METHODS: Of 500 patients who underwent cytology and colposcopy, 271 (54.2%) required a second opinion due to a diagnosis of cervical intraepithelial neoplasia (CIN). CS was removed and compared with the clean, second sample (SS) using in both liquid-based cytology. The quality of samples according to the Bethesda System, the presence of CIN, and inflammatory reactions were recorded. The sensitivity and specificity were calculated using biopsy as the gold standard. RESULTS: The SS resulted in a higher proportion of adequate samples being obtained (97.6% vs. 44.8%), and in increased sensitivity (88.2% vs. 58.8%). CIN was detected in the SS 26% more often than in the CS (34 vs. 27 samples), whereas inflammatory reactions were noted more often in the CS (91.4% vs. 74%). CONCLUSION: Cervical sampling including CS results in lower sensitivity and CIN detection rates, and in more inflammatory reactions. By excluding CS from cervical samples, the sensitivity could be improved and the false negative rate could be reduced.

Breeding of transgenic cattle for human coagulation factor IX by a combination of lentiviral system and cloning.

Recombinant coagulation factor IX must be produced in mammalian cells because FIX synthesis involves translational modifications. Human cell culture-based expression of human coagulation factor IX (hFIX) is expensive, and large-scale production capacity is limited. Transgenic animals may greatly increase the yield of therapeutic proteins and reduce costs. In this study, we used a lentiviral system to obtain transgenic cells and somatic cell nuclear transfer (SCNT) to produce transgenic animals. Lentiviral vectors carrying hFIX driven by 3 bovine ß-casein promoters were constructed. Bovine epithelial mammary cells were transduced by lentivirus, selected with blasticidin, plated on extracellular matrix, and induced by lactogenic hormones; promoter activity was evaluated by quantitative PCR. Transcriptional activity of the 5.335-kb promoter was 6-fold higher than the 3.392- and 4.279-kb promoters, which did not significantly differ. Transgenic bovine fibroblasts were transduced with lentivirus carrying the 5.335-kb promoter and used as donor cells for SCNT. Cloned transgenic embryo production yielded development rates of 28.4%, similar to previous reports on cloned non-transgenic embryos. The embryos were transferred to recipient cows (N = 21) and 2 births of cloned transgenic cattle were obtained. These results suggest combination of the lentiviral system and cloning may be a good strategy for production of transgenic cattle.

A segmentation algorithm for automated tracking of fast swimming unlabelled cells in three dimensions.

Recent advances in microscopy and cytolabelling methods enable the real time imaging of cells as they move and interact in their real physiological environment. Scenarios in which multiple cells move autonomously in all directions are not uncommon in biology. A remarkable example is the swimming of marine spermatozoa in search of the conspecific oocyte. Imaging cells in these scenarios, particularly when they move fast and are poorly labelled or even unlabelled requires very fast three-dimensional time-lapse (3D+t) imaging. This 3D+t imaging poses challenges not only to the acquisition systems but also to the image analysis algorithms. It is in this context that this work describes an original automated multiparticle segmentation method to analyse motile translucent cells in 3D microscopical volumes. The proposed segmentation technique takes advantage of the way the cell appearance changes with the distance to the focal plane position. The cells translucent properties and their interaction with light produce a specific pattern: when the cell is within or close to the focal plane, its two-dimensional (2D) appearance matches a bright spot surrounded by a dark ring, whereas when it is farther from the focal plane the cell contrast is inverted looking like a dark spot surrounded by a bright ring. The proposed method analyses the acquired video sequence frame-by-frame taking advantage of 2D image segmentation algorithms to identify and select candidate cellular sections. The crux of the method is in the sequential filtering of the candidate sections, first by template matching of the in-focus and out-of-focus templates and second by considering adjacent candidates sections in 3D. These sequential filters effectively narrow down the number of segmented candidate sections making the automatic tracking of cells in three dimensions a straightforward operation.

LC-MS/MS quantitation of plasma progesterone in cattle.

Quantitation of progesterone (P(4)) in biological fluids is often performed by radioimmunoassay (RIA), whereas liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) has been used much less often. Due to its autoconfirmatory nature, LC-MS/MS greatly minimizes false positives and interference. Herein we report and compare with RIA an optimized LC-MS/MS method for rapid, efficient, and cost-effective quantitation of P(4) in plasma of cattle with no sample derivatization. The quantitation of plasma P(4) released from three nonbiodegradable, commercial, intravaginal P(4)-releasing devices (IPRD) over 192 h in six ovariectomized cows was compared in a pairwise study as a test case. Both techniques showed similar P(4) kinetics (P > 0.05) whereas results of P(4) quantitation by RIA were consistently higher compared with LC-MS/MS (P < 0.05) due to interference and matrix effects. The LC-MS/MS method was validated according to the recommended analytical standards and displayed P(4) limits of detection (LOD) and quantitation (LOQ) of 0.08 and a 0.25 ng/mL, respectively. The high selective LC-MS/MS method proposed herein for P(4) quantitation eliminates the risks associated with radioactive handling; it also requires no sample derivatization, which is a common requirement for LC-MS/MS quantitation of steroid hormones. Its application to multisteroid assays is also viable, and it is envisaged that it may provide a gold standard technique for hormone quantitation in animal reproductive science studies.

Analysis of plasma and erythrocyte zinc levels in premenopausal women with breast cancer.

INTRODUCTION: Zinc deficiency has been associated with damage and oxidative changes in DNA that may increase an individual's risk of cancer. Furthermore, zinc metabolism may be affected in cancer patients, leading to alterations in its distribution that would favor carcinogenesis. Plasma and erythrocyte zinc levels in women with breast cancer were evaluated in this cross-sectional, controlled study. MATERIAL AND METHODS: Fifty-five premenopausal women of 25 to 49 years of age with and without breast cancer were divided into two groups: Group A, composed of women without breast cancer (controls, n = 26) and Group B, composed of women with breast cancer (cases, n = 29). Plasma and erythrocyte zinc levels were measured by flame atomic absorption spectrophotometry at γ = 213.9 nm. Diet was assessed using the 3-day diet recall method and analyzed using the NutWin software program, version 1.5. Student's t-test was used to compare means and significance was established at p = 0.05. RESULTS: Mean plasma zinc levels were 69.69 ± 9.00 g/dL in the breast cancer patients and 65.93 ± 12.44 g/dL in the controls (p = 0.201). Mean erythrocyte zinc level was 41.86 ± 8.28 µgZn/gHb in the cases and 47.93 ± 7.00 µgZn/gHb in the controls (p < 0.05). In both groups, dietary zinc levels were above the estimated average requirement. CONCLUSIONS: The present results suggest that zinc levels are lower in the erythrocyte compartment of pre-menopausal women with breast cancer.

[Morphological characteristics of cerebral aneurysm of Willis' circle: a direct anatomical study]. / Características morfológicas de aneurimas cerebrales del polígono de Willis: estudio anatómico directo.

OBJECTIVE: To determine the prevalence and morphologic characteristics of unruptured aneurysms of Willis' circle in a sample of mestizo Colombian population. MATERIALS AND METHODS: A mixture of resin and mineral red was injected into cerebral arteries by dissection and canalization of common carotids and vertebral arteries of the encephalons of 125 mestizo male cadavers of 16 to 65 years old. The procedure was carried out during the autopsy course at the Legal Medicine Institute, Bucaramanga - Colombia. Then the encephalons were extracted and fixed. After that, the Willis' circles were identified and the presence of aneurysms at this level was determined with magnifying glass. RESULTS: A total of nine aneurysms were observed in six brains (4.8%). The most frequent location was the supraclinoid segment of the intern carotid artery, with 4 cases (44.4%), three of which were localized in the origin of the anterior choroidal artery. Additionally, three aneurysms were found in the anterior communicating artery (33.3%). From the remaining cases, one (11.1%) was localized in the pre-communicating segment of the anterior cerebral artery, and the other in the bifurcation of the basilar artery. The average size of the aneurysms was 2.49 mm SD 0.37. The affected encephalons presented concomitant variations of the Willis' circle configuration, being predominant the presence of hypoplasic posterior communicanting arteries. CONCLUSION: The aneurysm prevalence in the evaluated sample was similar to the reported in other populations. In this work, the presence of aneurysms on the origin of the anterior chorioid artery, an unusually reported localization, was prominent.

Valor nutritivo do resíduo da indústria processadora de abacaxi (Ananas comosus L.) em dietas para ovinos. 1. Consumo, digestibilidade aparente e balanços energético e nitrogenado / Nutritive value of pineapple by-product (Ananas comosus L.) in diets for sheep. 1. Intake, apparent digestibility, energetic and nitrogenous balance

O presente estudo foi conduzido para avaliar a influência da inclusão do resíduo do processamento de abacaxi (Ananas comosus L.) nas dietas sobre os consumos (por unidade de tamanho metabólico - UTM) e coeficientes de digestibilidade da matéria seca (MS), matéria orgânica (MO), proteína bruta (PB), extrato etéreo (EE), fibra detergente neutro (FDN), fibra detergente ácido (FDA), hemiceluloses (HCEL), celulose (CEL) e energia de dietas experimentais isofibrosas e isoprotéicas. Avaliaram-se também os balanços nitrogenados e de energia dos animais. Vinte ovinos machos, inteiros receberam rações com quatro níveis de inclusão do resíduo (0; 11 por cento; 16 por cento; 27 por cento) em base de matéria seca, em delineamento inteiramente ao acaso, com cinco repetições por tratamento. Maiores consumos de matéria seca foram evidenciados quando as rações continham 11 por cento do resíduo. Maiores consumos de proteína bruta, fibra detergente neutro, fibra detergente ácido e celulose (g/UTM) e energia metabolizável (kcal/UTM) foram apresentados por animais alimentados com dietas contendo 11 por cento de resíduo em relação ao das dietas com 27 por cento de inclusão (P<0,05). Maior coeficiente de digestibilidade da FDA ocorreu para dietas que não continham o resíduo de abacaxi. Os coeficientes de digestibilidade da CEL foram semelhantes. Os balanços energéticos e protéicos foram positivos. Melhores resultados para os parâmetros analisados foram observados para dietas que incluíam até 16 por cento do resíduo de abacaxi na base da matéria seca.(AU)

The influence of increasing levels of pineapple by-product (Ananas comosus L.) on intake (unit metabolic size - UMS) and apparent digestibility ( percent) of dry matter (DM), organic matter (OM), crude protein (CP), ether extract (EE), neutral detergent fiber (NDF), acid detergent fiber (ADF), hemicelluloses (HCEL) and cellulose (CEL) and energy of isofibrous and isoproteic experimental diets of sheep were evaluated. Energetic and nitrogenous balance were also measured. Twenty rams were randomly allotted to four treatments in a completely randomized design with five replications. The treatments were formulated to contain zero; 11, 16 and 27 percent of pineapple by-product on DM basis. Higher dry matter intake was observed on 11 percent pineapple by-product diet. CP, NDF, ADF, CEL intake (g/UMS) and metabolizable energy intake (kcal/UMS) of 11 percent pineapple by-product diet were higher than 27 percent pineapple by-product diet (P<0.05). ADF digestibility was higher for 0 percent pineapple by-product diet. Energy and nitrogenous balances were positives. Diets formulated to contain up to 16 percent of pineapple by-product presented the best results.(AU)

Valor nutritivo do resíduo da indústria processadora de abacaxi (Ananas comosus L.) em dietas para ovinos. 1. Consumo, digestibilidade aparente e balanços energético e nitrogenado / Nutritive value of pineapple by-product (Ananas comosus L.) in diets for sheep. 1. Intake, apparent digestibility, energetic and nitrogenous balance

O presente estudo foi conduzido para avaliar a influência da inclusão do resíduo do processamento de abacaxi (Ananas comosus L.) nas dietas sobre os consumos (por unidade de tamanho metabólico - UTM) e coeficientes de digestibilidade da matéria seca (MS), matéria orgânica (MO), proteína bruta (PB), extrato etéreo (EE), fibra detergente neutro (FDN), fibra detergente ácido (FDA), hemiceluloses (HCEL), celulose (CEL) e energia de dietas experimentais isofibrosas e isoprotéicas. Avaliaram-se também os balanços nitrogenados e de energia dos animais. Vinte ovinos machos, inteiros receberam rações com quatro níveis de inclusão do resíduo (0; 11 por cento; 16 por cento; 27 por cento) em base de matéria seca, em delineamento inteiramente ao acaso, com cinco repetições por tratamento. Maiores consumos de matéria seca foram evidenciados quando as rações continham 11 por cento do resíduo. Maiores consumos de proteína bruta, fibra detergente neutro, fibra detergente ácido e celulose (g/UTM) e energia metabolizável (kcal/UTM) foram apresentados por animais alimentados com dietas contendo 11 por cento de resíduo em relação ao das dietas com 27 por cento de inclusão (P<0,05). Maior coeficiente de digestibilidade da FDA ocorreu para dietas que não continham o resíduo de abacaxi. Os coeficientes de digestibilidade da CEL foram semelhantes. Os balanços energéticos e protéicos foram positivos. Melhores resultados para os parâmetros analisados foram observados para dietas que incluíam até 16 por cento do resíduo de abacaxi na base da matéria seca.

The influence of increasing levels of pineapple by-product (Ananas comosus L.) on intake (unit metabolic size - UMS) and apparent digestibility ( percent) of dry matter (DM), organic matter (OM), crude protein (CP), ether extract (EE), neutral detergent fiber (NDF), acid detergent fiber (ADF), hemicelluloses (HCEL) and cellulose (CEL) and energy of isofibrous and isoproteic experimental diets of sheep were evaluated. Energetic and nitrogenous balance were also measured. Twenty rams were randomly allotted to four treatments in a completely randomized design with five replications. The treatments were formulated to contain zero; 11, 16 and 27 percent of pineapple by-product on DM basis. Higher dry matter intake was observed on 11 percent pineapple by-product diet. CP, NDF, ADF, CEL intake (g/UMS) and metabolizable energy intake (kcal/UMS) of 11 percent pineapple by-product diet were higher than 27 percent pineapple by-product diet (P<0.05). ADF digestibility was higher for 0 percent pineapple by-product diet. Energy and nitrogenous balances were positives. Diets formulated to contain up to 16 percent of pineapple by-product presented the best results.

'Candidatus Phytoplasma brasiliense', a new phytoplasma taxon associated with hibiscus witches' broom disease.

Hibiscus rosa-sinensis is a valuable ornamental species widely planted in Brazil. Many plants are affected by witches' broom disease, which is characterized by excessive axillary branching, abnormally small leaves, and deformed flowers, symptoms that are characteristic of diseases attributed to phytoplasmas. A phytoplasma was detected in diseased Hibiscus by amplification of rRNA operon sequences by PCRs, and was characterized by RFLP and nucleotide sequence analyses of 16S rDNA. The collective RFLP patterns of amplified 16S rDNA differed from the patterns described previously for other phytoplasmas. On the basis of the RFLP patterns, the hibiscus witches' broom phytoplasma was classified in a new 16S rRNA RFLP group, designated group 16SrXV. A phylogenetic analysis of 16S rDNA sequences from this and other phytoplasmas identified the hibiscus witches' broom phytoplasma as a member of a distinct subclade (designated subclade xiv) of the class Mollicutes. A phylogenetic tree constructed on the basis of 16S rRNA gene sequences was consistent with the hypothesis that there was divergent evolution of hibiscus witches' broom phytoplasma and its closest relatives (members of 16S rRNA RFLP group 16SrII) from a common ancestor. On the basis of unique properties of the DNA from hibiscus witches' broom phytoplasma, it is proposed that it represents a new taxon, namely 'Candidatus Phytoplasma brasiliense'.

First Report of Natural Infection by "Candidatus Phytoplasma brasiliense" in Catharanthus roseus.

Catharanthus roseus (L.) G. Don (periwinkle) is well known as an experimental host for diverse phytoplasmas that are artificially transmitted to it through the use of dodder (Cuscuta sp.), laboratory vector insects, or grafting. However, few phytoplasma taxa have been reported in natural infections of C. roseus, and the role of C. roseus in phytoplasma dissemination and natural disease spread is not clear. In this study, naturally diseased plants of C. roseus exhibiting yellowing and witches' broom symptoms indicative of phytoplasma infection were observed throughout the year in the state of Rio de Janeiro, Brazil. Shoots and leaves of four diseased plants were assayed for the presence of phytoplasma DNA sequences by nested polymerase chain reactions (PCR) as previously described (2,3). Phytoplasma rDNA was amplified from diseased periwinkle plants in PCR primed by primer pair P1/P7 and was reamplified in nested PCR primed by primer pair R16F2n/R16R2 (F2n/R2). The results indicated the presence of phytoplasma in all four diseased plants. Phytoplasma identification was accomplished by restriction fragment length polymorphism (RFLP) analysis, using 11 restriction enzymes, of 16S rDNA amplified in PCR primed by F2n/R2. Phytoplasmas were classified according to the system of Lee et al. (1). On the basis of collective RFLP patterns of 16S rDNA, the phytoplasma infections in the four periwinkle plants could not be distinguished from one another. Furthermore, the collective RFLP patterns were indistinguishable from those reported previously for hibiscus witches' broom phytoplasma, "Candidatus Phytoplasma brasiliense" (2). The phytoplasma found in C. roseus, designated strain HibWB-Cr, was classified in group 16SrXV (hibiscus witches' broom phytoplasma group). HibWB-Cr is tentatively considered a new strain of "Ca. P. brasiliense". C. roseus is the first known, naturally diseased alternate plant host of "Ca. P. brasiliense". The present study identified strain HibWB-Cr in Rio de Janeiro State, where hibiscus witches' broom disease is prevalent (2). How this economically important disease of hibiscus spreads is not known. Our findings raise the possibility that a polyphagous insect vector is involved in the natural transmission of "Ca. P. brasiliense" and that C. roseus or other plant species serve as reservoirs for the spread of this phytoplasma taxon. References: (1) I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998. (2) H. G. Montano et al. Int. J. Syst. Evol. Microbiol. 51:1109, 2001. (3) H. G. Montano et al. Plant Dis. 84:429, 1999.

How is mechanical ventilation employed in the intensive care unit? An international utilization review.

A 1-d point-prevalence study was performed with the aim of describing the characteristics of conventional mechanical ventilation in intensive care units ICUs from North America, South America, Spain, and Portugal. The study involved 412 medical-surgical ICUs and 1,638 patients receiving mechanical ventilation at the moment of the study. The main outcome measures were characterization of the indications for initiation of mechanical ventilation, the artificial airways used to deliver mechanical ventilation, the ventilator modes and settings, and the methods of weaning. The median age of the study patients was 61 yr, and the median duration of mechanical ventilation at the time of the study was 7 d. Common indications for the initiation of mechanical ventilation included acute respiratory failure (66%), acute exacerbation of chronic obstructive pulmonary disease (13%), coma (10%), and neuromuscular disorders (10%). Mechanical ventilation was delivered via an endotracheal tube in 75% of patients, a tracheostomy in 24%, and a facial mask in 1%. Ventilator modes consisted of assist/control ventilation in 47% of patients and 46% were ventilated with synchronized intermittent mandatory ventilation, pressure support, or the combination of both. The median tidal volume setting was 9 ml/kg in patients receiving assist/control and the median setting of pressure support was 18 cm H(2)O. Positive end-expiratory pressure was not employed in 31% of patients. Method of weaning varied considerably from country to country, and even within a country several methods were in use. We conclude that the primary indications for mechanical ventilation and the ventilator settings were remarkably similar across countries, but the selection of modes of mechanical ventilation and methods of weaning varied considerably from country to country.

Identification and Phylogenetic Analysis of a New Phytoplasma from Diseased Chayote in Brazil.

Chayote (Sechium edule) (Cucurbitaceae), also known as vegetable pear, mirliton, or mango squash, is a commercially important vegetable crop in Brazil, where it is affected by chayote witches'-broom disease. Affected plants exhibit witches'-broom growths and other symptoms characteristic of plant diseases caused by phytoplasmas. Since previous electron microscopic studies revealed the association of a phytoplasma with chayote witches'-broom, the present work was aimed at detecting and classifying the phytoplasma that may be the causal agent of the disease. Strains of a phytoplasma belonging to group 16SrIII (X-disease phytoplasma group) were discovered in chayote affected by witches'-broom disease and in diseased plants of Momordica charantia that were growing as weeds in fields of chayote in Brazil. On the basis of results from restriction fragment length polymorphism and nucleotide sequence analyses of 16S rDNA, the phytoplasma was classified in a new subgroup, designated subgroup III-J. This classification was supported by a phylogenetic tree constructed by the Neighbor-Joining method.

Biosorption of metal ions byAzotobacter vinelandii.

Azotobacter vinelandii was better than eitherDerxia gummosa orRhizobium trifolii for sorption of UO 2 (2+) . Its maximum binding capacity was 0.25 mmol UO 2 (2+) /g dry biomass with an affinity constant of 333 l/mmol at pH 4.1 according to the Langmuir model. In a semisynthetic medium,A. vinelandii showed the highest sorption capacity in the early stationary phase. The binding of UO 2 (2+) , Cu(2+), Ca(2+) and Zn(2+) was affected by the pH of the solution. With HCl as eluent, virtually all the sorbed UO 2 (2+) was released. The presence of Cu(2+), Cd(2+), Ca(2+), and Zn(2+) inhibited the UO 2 (2+) biosorption whereas Mg(2+) and K(+) had no effect.