Permanent Stained Preparations of Synovial Fluid for Detection of Calcium Compounds Using Alizarin Red S

Permanent preparations of air dried synovial fluids were prepared by staining calcium compounds with alizarin red S stain; each slide was coverslipped with Permount. Variables studied were: (a) concentration of the solution of alizarin red S, (b) pH of staining solution, (c) time of incubation in staining solution and aqueous and ethanolic content of staining solution. The staining effect of each solution was tested on calcium pyrophosphate dihydrate, calcium oxalate, apatite and monosodium urate (MSU). Of all the solutions, best results were obtained with 0.25% alizarin red S in 50% ethanol at pH 7.0 for 30 min. With this solution, the calcium-containing compounds were well stained. MSU did not stain and still preserved negative birefringence on polarizaton. Fixation of smears with ethanol served a double purpose: It fixed the slides without dissolving or removing MSU or the calcium compounds, yet it did dissolve five corticosteroids commonly used for intra-articular injection which may interfere with interpretation of compensated polarized light microscopy of synovial fluids.     

Proteomic analysis of phytopathogenic fungus <Emphasis Type="Italic">Botrytis cinerea</Emphasis> as a potential tool for identifying pathogenicity factors,therapeutic targets and for basic research

Botrytis cinerea is a phytopathogenic fungus causing disease in a substantial number of economically important crops. In an attempt to identify putative fungal virulence factors, the two-dimensional gel electrophoresis (2-DE) protein profile from two B. cinerea strains differing in virulence and toxin production were compared. Protein extracts from fungal mycelium obtained by tissue homogenization were analyzed. The mycelial 2-DE protein profile revealed the existence of qualitative and quantitative differences between the analyzed strains. The lack of genomic data from B. cinerea required the use of peptide fragmentation data from MALDI-TOF/TOF and ESI ion trap for protein identification, resulting in the identification of 27 protein spots. A significant number of spots were identified as malate dehydrogenase (MDH) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The different expression patterns revealed by some of the identified proteins could be ascribed to differences in virulence between strains. Our results indicate that proteomic analysis are becoming an important tool to be used as a starting point for identifying new pathogenicity factors, therapeutic targets and for basic research on this plant pathogen in the postgenomic era.     

Hospital costs associated with delirium in older medical patients

Introduction

Delirium is a common and serious complication in older patients, associated with increased, potentially preventable, morbidity and mortality. The aim of this study was to evaluate the associated costs of delirium during hospitalization in a university affiliated hospital in Chile.

Materials and methods

Prospective cohort study of consecutive patients 65 years and older, admitted to a medical ward. A psychogeriatric team assessed patients during the first and every 48 h until discharge using the Confusion Assessment Method (CAM-S), length of hospital stay, pharmacy and total hospitalization costs were analyzed. Statistical analysis was performed using bivariate and multivariate analysis according to delirium diagnosis.

Results

Data from 454 patients was analyzed, 160 of them in a delirium cohort (35.2%) and 294 in a non-delirium cohort (64.8%). The delirium cohort had a longer hospital stay (DATA) and higher mortality (7.0% versus 1.7%). The median of total costs of delirium during hospital stay was 38.7% higher than the non-delirium cohort (P < .001). Total costs were significantly higher in the delirium cohort after adjustment of covariables (P = .01).

Conclusions

This study confirms that delirium is associated with significantly greater costs. Considering that effective delirium prevention is possible, the knowledge of associated costs can help health care providers to justify prevention strategies and finally give better care for older patients.     

A new methodology combining PCR, cloning, and sequencing of clones discriminated by RFLP for the study of microbial populations: application to an UASB reactor sample

This work describes a methodology combining DNA extraction, polymerase chain reaction amplification with primers targeting 16S ribosomal RNA genes, cloning, and sequencing of clones previously analyzed by restriction fragment length polymorphism (RFLP), which can be applied to study the microbial diversity in a given habitat. The methodology allows the minimization of the sequencing effort, which is particularly relevant when analyzing large numbers of clones. The methodology does not require particularly skilled personnel and can easily be adaptable to the molecular characterization of virtually any particular microbial population, provided that both adequate primers and suitable restriction enzymes for RFLP analysis of the clone library have been chosen. An example of application is presented, in which a sample taken from a continuously operating upflow anaerobic sludge blanket reactor was analyzed. RFLP analysis of the initial 162 clones with HaeIII allowed the identification of only 28 distinct profiles. As expected, identical RFLP profiles corresponded to identical nucleotide sequences.     

nAChR‐induced octopamine release mediates the effect of nicotine on a startle response in Drosophila melanogaster

Biogenic amines (BAs) play a central role in the generation of complex behaviors in vertebrates and invertebrates, including the fly Drosophila melanogaster. The comparative advantages of Drosophila as a genetic model to study the contribution of BAs to behaviors stumble upon the difficulty to access the fly brain to ask relevant physiological questions. For instance, it is not known whether the activation of nicotinic acetylcholine receptors (nAChRs) induces the release of BAs in fly brain, a phenomenon associated to several behaviors in vertebrates. Here, we describe a new preparation to study the efflux of BAs in the adult fly brain by in vitro chronoamperometry. Using this preparation we show that nAChR agonists including nicotine induce a fast, transient, dose‐dependent efflux of endogenous BAs, an effect mediated by α‐bungarotoxin‐sensitive nAChRs. By using different genetic tools we demonstrate that the BA whose efflux is induced by nAChR activation is octopamine (Oct). Furthermore, we show that the impairment of a mechanically induced startle response after nicotine exposure is not observed in flies deficient in Oct transmission. Thus, our data show that the efflux of BAs in Drosophila brain is increased by nAChR activation as in vertebrates, and that then AChR‐induced Oct release could have implications in a nicotine‐induced behavioral response.     

Using Manifold Learning for Atlas Selection in Multi-Atlas Segmentation

Multi-atlas segmentation has been widely used to segment various anatomical structures. The success of this technique partly relies on the selection of atlases that are best mapped to a new target image after registration. Recently, manifold learning has been proposed as a method for atlas selection. Each manifold learning technique seeks to optimize a unique objective function. Therefore, different techniques produce different embeddings even when applied to the same data set. Previous studies used a single technique in their method and gave no reason for the choice of the manifold learning technique employed nor the theoretical grounds for the choice of the manifold parameters. In this study, we compare side-by-side the results given by 3 manifold learning techniques (Isomap, Laplacian Eigenmaps and Locally Linear Embedding) on the same data set. We assess the ability of those 3 different techniques to select the best atlases to combine in the framework of multi-atlas segmentation. First, a leave-one-out experiment is used to optimize our method on a set of 110 manually segmented atlases of hippocampi and find the manifold learning technique and associated manifold parameters that give the best segmentation accuracy. Then, the optimal parameters are used to automatically segment 30 subjects from the Alzheimer’s Disease Neuroimaging Initiative (ADNI). For our dataset, the selection of atlases with Locally Linear Embedding gives the best results. Our findings show that selection of atlases with manifold learning leads to segmentation accuracy close to or significantly higher than the state-of-the-art method and that accuracy can be increased by fine tuning the manifold learning process.     

Population trends for humpback whales (Megaptera novaeangliae) foraging in the Francisco Coloane Coastal‐Marine Protected Area,Magellan Strait,Chile

In 2003 a feeding aggregation of southeastern Pacific humpback whales (Megaptera novaeangliae) was reported in the Magellan Strait. While Chile established its first marine national park in the Strait to protect humpback whale habitat, fatal ship strikes remain a concern because of overlap with a busy shipping lane. To better understand population risk, we estimated abundance and survival for this population using Bayesian robust‐design mark‐recapture models fit to photographic data from 2004 to 2016. Overall, the model estimated a total of 204 whales (95% CI: 199–210) during the last 12 yr, and 93 (95% CI: 86–100) in the 2016/2017 austral summer. The population grew at 2.3% (CI: 2.1%–3.1%), an annual increase of two whales. Annual survival (including calves) was estimated at 0.892 (CI: 0.871–0.910). Our results corroborate a persistent feeding population, but one that is increasing relatively slowly. Owing to its vulnerability stemming from its small size, coupled with significant overlap with a busy shipping lane, we argue this subpopulation is at significant risk from ship strikes and may be one of the few populations where anthropogenic mortalities could regulate population dynamics. We therefore encourage continued monitoring via photographic mark‐resighting surveys, and analyses explicitly investigating potential population‐level ship strike effects.     

The Administration of Cadmium for 2, 3 and 4 Months Causes a Loss of Recognition Memory,Promotes Neuronal Hypotrophy and Apoptosis in the Hippocampus of Rats

Cadmium (Cd) is a toxic metal and classified as a carcinogen whose exposure could affect the function of the central nervous system. There are studies that suggest that Cd promotes neurodegeneration in different regions of the brain, particularly in the hippocampus. It is proposed that its mechanism of toxicity maybe by an oxidative stress pathway, which modifies neuronal morphology and causes the death of neurons and consequently affecting cognitive tasks. However, this mechanism is not yet clear. The aim of the present work was to study the effect of Cd administration on recognition memory for 2, 3 and 4 months, neuronal morphology and immunoreactivity for caspase-3 and 9 in rat hippocampi. The results show that the administration of Cd decreased recognition memory. Likewise, it caused the dendritic morphology of the CA1, CA3 and dentate gyrus regions of the hippocampus to decrease with respect to the time of administration of this heavy metal. In addition, we observed a reduction in the density of dendritic spines as well as an increase in the immunoreactivity of caspase-3 and 9 in the same hippocampal regions of the animals treated with Cd. These results suggest that Cd affects the structure and function of the neurons of the hippocampus, which contribute to the deterioration of recognition memory. Our results suggest that the exposure to Cd represents a critical health problem, which if not addressed quickly, could cause much more serious problems in the quality of life of the human population, as well as in the environment in which they develop.