Localization of Pantoea ananatis inside lesions of maize White Spot Disease using transmission electron microscopy and molecular techniques

The etiological agent of maize white spot (MWS) disease has been a subject of controversy and discussion. Initially the disease was described as Phaeosphaeria leaf spot caused by Phaeosphaeria maydis. Other authors have Suggested the existence of different fungal species causing similar symptoms. Recently, a bacterium, Pantoea ananatis, was described as the causal agent of this disease. The purpose of this Study was to offer additional information on the correct etiology of this disease by providing visual evidence of the presence of the bacterium in the interior of the MWS lesions by using transmission electron microscopy (TEM) and molecular techniques. The TEM allowed Visualization of a large amount of bacteria in the intercellular spaces of lesions collected from both artificially and naturally infected plants. Fungal structures were not visualized in young lesions. Bacterial primers for the 16S rRNA and rpoB genes were used in PCR reactions to amplify DNA extracted from water-soaked (young) and necrotic lesions. The universal fungal oligonucleotide ITS4 was also included to identity the possible presence of fungal structures inside lesions. Positive PCR products from water-soaked lesions, both from naturally and artificially inoculated plants, were produced with bacterial primers, whereas no amplification was observed when ITS4 oligonucleotide was used. On the other hand, DNA amplification with ITS4 primer was observed when DNA was isolated from necrotic (old) lesions. These results reinforced previous report of P. ananatis as the primary pathogen and the hypothesis that fungal species may colonize lesions pre-established by P. ananatis.

Diversity of endophytic yeasts from sweet orange and their localization by scanning electron microscopy

Endophytes are microorganisms that colonize plant tissues internally without causing harm to the host. Despite the increasing number of studies on sweet orange pathogens and endophytes, yeast has not been described as a sweet orange endophyte. In the present study, endophytic yeasts were isolated from sweet orange plants and identified by sequencing of internal transcribed spacer (ITS) rRNA. Plants sampled from four different sites in the state of Sao Paulo, Brazil exhibited different levels of CVC (citrus variegated chlorosis) development. Three citrus endophytic yeasts (CEYs), chosen as representative examples of the isolates observed, were identified as Rhodotorula mucilaginosa, Pichia guilliermondii and Cryptococcus flavescens. These strains were inoculated into axenic Citrus sinensis seedlings. After 45 days, endophytes were reisolated in populations ranging from 10(6) to 10(9) CFU/g of plant tissue, but, in spite of the high concentrations of yeast cells, no disease symptoms were observed. Colonized plant material was examined by scanning electron microscopy (SEM), and yeast cells were found mainly in the stomata and xylem of plants, reinforcing their endophytic nature. P. guilliermondii was isolated primarily from plants colonized by the causal agent of CVC, Xylella fastidiosa. The supernatant from a culture of P. guilliermondii increased the in vitro growth of X. fastidiosa, suggesting that the yeast could assist in the establishment of this pathogen in its host plant and, therefore, contribute to the development of disease symptoms.

Effects of natural antioxidants on the lipid profile of electron beam-irradiated beef burgers

The purpose of this study was to assess the efficacy of rosemary and oregano extracts in avoiding oxidative changes in beef burgers, and to evaluate the fatty acid profile of these products after electron beam exposition. Extracts, individually or in combination, were added to beef burgers and compared to synthetic antioxidants commonly used in food (butylated hydroxytoluene, butylated hydroxyanisole). The ground beef were submitted to electron beam irradiation at doses of 0, 3.5 and 7 kGy, and stored for 90 days. At regular time intervals, lipid oxidation and fatty acid composition were evaluated through measurement of thiobarbituric acid-reactive substances (TBARS) and gas chromatography, respectively. The results indicate that, although the irradiation process triggers an increase in the lipid oxidation ratio expressed by TBARS values, great changes in the fatty acid profiles were not observed; instead, they continued to present characteristics very similar to that of non-irradiated beef. Thus, as irradiation doses of up to 7 kGy for frozen meat can make foods safe from foodborne pathogens, natural antioxidants derived from spices are able to reduce and avoid lipid changes that may cause a deterioration of the sensory quality of these foods, and these natural extracts offer a good choice for replacing synthetic additives.

Electron beam irradiation of Matricaria chamomilla L. for microbial decontamination

Wild chamomile (Matricaria chamomilla L.) is one of the most popular herbal materials with both internal and external use to cure different health disturbances. As a consequence of its origin, chamomile could carry various microbial contaminants which offer different hazards to the final consumer. Reduction of the microbial load to the in force regulation limits represents an important phase in the technological process of vegetal materials, and the electron beam treatment might be an efficient alternative to the classical methods of hygienic quality assurance. The purpose of the study was to analyze the potential application of the electron beam treatment in order to assure the microbial safety of the wild chamomile. Samples of chamomile dry inflorescences were treated in electron beam (e-beam) of 6 MeV mean energy, at room temperature and ambient pressure. Some loss of the chemical compounds with bioactive role could be noticed, but the number of microorganisms decreased as a function on the absorbed dose. Consequently, the microbial quality of studied vegetal material inflorescences was improved by e-beam. irradiation. (C) 2008 Elsevier B.V. All rights reserved.

In vitro safety assessment of papain on human skin: A qualitative Light and Transmission Electron Microscopy (TEM) study

Papain is a thiol proteolytic enzyme widely used in dermatology that found applications in wound treatment. Recently, papain was also used as absorption enhancer which can modify the peptide/ protein material in the bilayer domain. We investigated papain safety using human skin that was exposed to papain in vitro at different times: 4, 24 and 48 hours. The samples were examined using Light and Transmission Electron Microscopy (TEM) to study of the mechanisms involved in enhancer-skin interaction. After 24 hours, changes occurred in corneosomes. However, samples of 48 hours did not show major changes in agreement with the control. These findings indicated that papain could be used safely onto the skin.

Thoracic aorta calcification detected by electron beam tomography predicts all-cause mortality

Background: The presence of coronary artery calcium (CAC) is an independent marker of increased risk of cardiovascular disease (CVD) events and mortality. However, the predictive value of thoracic aorta calcification (TAC), which can be additionally identified without further scanning during assessment of CAC, is unknown. Methods: We followed a cohort of 8401 asymptomatic individuals (mean age: 53 +/- 10 years, 69% men) undergoing cardiac risk factor evaluation and TAC and CAC testing with electron beam computed tomography. Multivariable Cox proportional hazards models were developed to predict all-cause mortality based on the presence of TAC. Results: During a median follow-up period of 5 years, 124 (1.5%) deaths were observed. Overall survival was 96.9% and 98.9% for those with and without detectable TAC, respectively (p < 0.0001). Compared to those with no TAC, the hazard ratio for mortality in the presence of TAC was 3.25 (95% CI: 2.28-4.65, p < 0.0001) in unadjusted analysis. After adjusting for age, gender, hypertension, dyslipidemia, diabetes mellitus, smoking and family history of premature coronary artery disease, and presence of CAC the relationship remained robust (HR 1.61, 95% CI: 1.10-2.27, p = 0.015). Likelihood ratio chi(2) statistics demonstrated that the addition of TAC contributed significantly in predicting mortality to traditional risk factors alone (chi(2) = 13.62, p = 0.002) as well as risk factors + CAC (chi(2) = 5.84, p = 0.02) models. Conclusion: In conclusion, the presence of TAC was associated with all-cause mortality in our study; this relationship was independent of conventional CVD risk factors as well as the presence of CAC. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Mammography findings following electron intraoperative radiotherapy or external radiotherapy for breast cancer treatment

Radiotherapy following breast cancer conserving surgery decreases the risks of local recurrence. Because 85% of breast cancers relapse in or around the surgical bed there has been some debate on the need for irradiating the whole breast. Electron intraoperative radiotherapy (ELIOT) has been used as a viable alternative for conventional external radiotherapy (RT). While the former requires a single dose of 21 Gy in the tumoral bed, the latter requires 5-6 weeks of irradiation with a total dose of 50 Gy and a boost of 10 Gy that irradiates the surgical bed. Herein, we investigated whether any significant differences exist between the mammography findings obtained from patients submitted to one of the two techniques. Two groups of 30 patients each were included in this study. All patients had mammographies taken at 12 and 24 months after finishing treatment. The mammography findings evaluated were: cutaneous thickening (>2 mm), architectural distortion secondary to fibrosis, edema, calcifications (both benign and malignant), and fat necrosis. For all variables studied, there was no statistical difference between the two groups. This indicates that the mammography findings obtained in either 12- or 24-month follow-up periods after breast cancer conserving surgery are similar, regardless of which of the two radiotherapy techniques (ELIOT or RT) is employed as a treatment for breast cancer. (C) 2010 Published by Elsevier Ireland Ltd.

Scanning electron microscopy as an auxiliary method in the study of exhumed bones

Scanning electronmicroscopy (SEM) has been used in forensic science in many ways. The reports of cases in which SEM has been used as an auxiliary method in the investigation of exhumed bones are rare. In this article, we report an exhumation that was made to determine if a seized weapon could have been used in a homicide. We used SEM to analyze a fracture in the interior of the skull of the victim. The findings described in this article showed us that it is possible to develop new researches in this field. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Dorsal cutaneous branch of the ulnar nerve: a light and electron microscopy histometric study

This study describes the normal morphology and morphometry of the dorsal cutaneous branch of the ulnar nerve (DCBU) in humans. Fourteen nerves of eight donors were prepared by conventional techniques for paraffin and epoxy resin embedding. Semiautomatic morphometric analysis was performed by means of specific computer software. Histograms of the myelinated and unmyelinated fiber population and the G-ratio distribution of fibers were plotted. Myelinated fiber density per nerve varied from 5,910 to 10,166 fibers/mm(2), with an average of 8,170 +/- 393 fibers/mm(2). The distribution was bimodal with peaks at 4.0 and 9.5 mu m. Unmyelinated fiber density per nerve varied from 50,985 to 127,108, with an average of 78,474 +/- 6, 610 fibers/mm(2), with a unimodal distribution displaying a peak at 0.8 mu m. This study thus adds information about the fascicles and myelinated and unmyelinated fibers of DCBU nerves in normal people, which may be useful in further studies concerning ulnar nerve neuropathies, mainly leprosy neuropathy.

Characteristics of Filiform, Fungiform and Vallate Papillae and Surface of Interface Epithelium-Connective Tissue of the Maned Sloth Tongue Mucosa (Bradypus torquatus, Iliger, 1811): Light and Scanning Electron Microscopy Study

The study of lingual surfaces and the surface of interface epithelium-connective tissue of the tongue of Bradypus torquatus was performed by employing the light and scanning electron microscopy (SEM) techniques. The results revealed that the rostral part of the tongue presents a round apex and covered by filiform and fungiform lingual papillae and a ventral smooth surface. It was observed that the epithelial layer of the dorsal surface possesses the basal, spinosum, granular and cornified epithelial cells. The lamina propria is characterized by a dense connective tissue forming the long, short and round papillae. Numerous typical filiform papillae are located especially in the rostral part intermingled for few fungiform papillae, which were revealed in three-dimensional SEM images. Usually, the fungiform papillae are located in the border of rostral apex of the tongue exhibiting the rounded form. They are covered by keratinized epithelial cells. In the fungiform papillae, several taste pores were observed on the surface. The vallate papillae presented numerous taste buds in the wall of epithelial cells, being that the major number of taste buds is located on the superior half of vallate papilla. The taste pores are surrounded by several laminae of keratinized epithelial cells. The samples treated with NaOH solution and examined by SEM revealed, after removal of the epithelial layer, the dense connective core in original disposition, presenting different sizes and shapes. The specimens stained with Picrosirius and examined by polarized light microscopy revealed the connective tissue, indicating the collagen fibres type I and type III.

Three-dimensional Characteristics of the Interface Epithelium-Connective Tissue Surface of Finger`s Lamina Propria of Cebus apella Monkey: Scanning Electron Microscopy Study

In the present paper were analysed the three-dimensional characteristics of the interface epithelium-connective tissue surface of finger prints of Cebus apella monkey employing the scanning electron microscopic methods. The connective tissue core (CTC) and epithelial papillae were examined verifying the three-dimensional configuration of the tissue projections. The samples were fixed in Bouin solsution for histologic preparations and in modified Karnovsky for examine to observe in scanning electron microscopy. After treatment in the 10% NaOH solution during 3 to 5 days, the surface of finger prints revealed a distribution of CTC of lamina propria in situ showing original three-dimensional SEM images. The linear and circular dispositions CTC, and the furrows were clearly identified. Each pointed papilla presented a large base and longitudinal disposition of thick collagen fiber bundles and in some areas with a complex reticular formations. The longitudinal furrows between the pointed papillae exhibited a dense layer of connective tissue and showed only low CTC or laminar in shape. The presence of numerous foramina of sweat gland were noted in three-dimensional SEM images.

Light and scanning electron microcopy study of the tongue in Rhea americana

Morphological characteristics of the tongue were studied in adult rhea (Rhea americana). The lingual surface and the surface of epithelium-connective tissue interface of rhea tongue were examined macroscopically and by light and scanning electron microscopy. The rhea tongue revealed a triangular aspect, without adjustment of the inferior bill formation, occupying approximately of the length of the oral cavity. Lingual papilla-like structures were not observed over the lingual surface. The tongue mucosa was composed of a thick non-keratinized stratified squamous epithelium in the dorsal and ventral part, supported by a connective tissue core. The submucosa contained numerous glands with cytoplasmic granules, and luminal secretion was positive for histochemical reaction to Alcian Blue in pH 2.5 and PAS, and negative to Alcian Blue in pH 0.5. Despite the rudimentary characteristic of the tongue in rhea, our results suggest an important role of tongue secretions in food lubrication and humidification during the swallowing process, based on the enormous quantity of lingual glands in the submucosa and the histochemical characteristics of their secretions.

Redescription of Amblyomma varium Koch, 1844 (Acari : Ixodidae) based on light and scanning electron microscopy

Amblyomma varium Koch, 1844 is a Neotropical tick, known as the `sloth`s giant tick`, with records from southern Central America to Argentina. It is found almost exclusively on mammals of the families Bradypodidae and Magalonychidae (Xenarthra). Differences exist in discussions with regard to the dentition of the female hypostome being either 3/3 or 4/4. The male was also originally described as having a short spur on coxa IV, but some specimens recently collected from different Brazilian localities have this spur three times longer. These differences beg the question of whether there is more than one species included under this taxon. In order to answer this question and to clarify the taxonomic characters of this species, 258 adult specimens were examined, and a redescription of male and female based on light and scanning electron microscopy is provided. In addition, DNA was extracted from males with either a long or a short spur on coxa IV to help settle this question for future investigations on their taxonomy. The morphological study showed that the dental formula pattern for males and females is 3/3 and 4/4, respectively. When sequenced, the 12 S rDNA genes of both A. varium males with long and short spurs on coxa IV were found to be identical, indicating that the length of the spurs on coxa IV is likely to be an intraspecifically polymorphic character of this species.

Comparative analysis of root surface smear layer removal by different etching modalities or erbium:yttrium-aluminum-garnet laser irradiation. A scanning electron microscopy study

The purpose of this study was to evaluate the effect of erbium:yttrium-aluminum-garnet (Er:YAG) laser (2.94 mu m) irradiation on the removal of root surface smear layer of extracted human teeth and to compare its efficacy with that of citric acid, ethylenediamine tetra-acetic acid (EDTA), or a gel containing a mixture of tetracycline hydrochloride (HCl) and citric acid, using scanning electron microscopy (SEM). Thirty human dentin specimens were randomly divided into six groups: G1 (control group), irrigated with 10 ml of physiologic saline solution; G2, conditioned with 24% citric acid gel; G3, conditioned with 24% EDTA gel; G4, conditioned with a 50% citric acid and tetracycline gel; G5, irradiated with Er:YAG laser (47 mJ/10 Hz/5.8 J/cm(2)/pulse); G6, irradiated with Er:YAG laser (83 mJ/10 Hz/10.3 J/cm(2)/pulse). Electron micrographs were obtained and analyzed according to a rating system. Statistical analysis was conducted with Kruskal-Wallis and Mann-Whitney tests (P < 0.05). G1 was statistically different from all the other groups; no statistically significant differences were observed between the Er:YAG laser groups and those undergoing the other treatment modalities. When the two Er:YAG laser groups were compared, the fluency of G6 was statistically more effective in smear layer removal than the one used in G5 (Mann-Whitney test, P < 0.01). Root surfaces irradiated by Er:YAG laser had more irregular contours than those treated by chemical agents. It can be concluded that all treatment modalities were effective in smear layer removal. The results of our study suggest that the Er:YAG laser can be safely used to condition diseased root surfaces effectively. Furthermore, the effect of Er:YAG laser irradiation on root surfaces should be evaluated in vivo so that its potential to enhance the healing of periodontal tissues can be assessed.

CO(2) Laser (10.6 mu m) Parameters for Caries Prevention in Dental Enamel

Although CO(2) laser irradiation can decrease enamel demineralisation, it has still not been clarified which laser wavelength and which irradiation conditions represent the optimum parameters for application as preventive treatment. The aim of the present explorative study was to find low-fluence CO(2) laser (lambda = 10.6 mu m) parameters resulting in a maximum caries-preventive effect with the least thermal damage. Different laser parameters were systematically evaluated in 3 steps. In the first experiment, 5 fluences of 0.1, 0.3, 0.4, 0.5 and 0.6 J/cm(2), combined with high repetition rates and 10 mu s pulse duration, were chosen for the experiments. In a second experiment, the influence of different pulse durations (5, 10, 20, 30 and 50 mu s) on the demineralisation of dental enamel was assessed. Finally, 3 different irradiation times (2, 5 and 9 s) were tested in a third experiment. In total, 276 bovine enamel blocks were used for the experiments. An 8-day pH-cycling regime was performed after the laser treatment. Demineralisation was assessed by lesion depth measurements with a polarised light microscope, and morphological changes were assessed with a scanning electron microscope. Irradiation with 0.3 J/cm(2), 5 mu s, 226 Hz for 9 s (2,036 overlapping pulses) increased caries resistance by up to 81% compared to the control and was even significantly better than fluoride application (25%, p < 0.0001). Scanning electron microscopy examination did not reveal any obvious damage caused by the laser irradiation. Copyright (C) 2009 S. Karger AG, Basel