Microfluidic cartridge with integrated array of amorphous silicon photosensors for chemiluminescence detection of viral DNA

Portable and simple analytical devices based on microfluidics with chemiluminescence detection are particularly attractive for point-of-care applications, offering high detectability and specificity in a simple and miniaturized analytical format. Particularly relevant for infectious disease diagnosis is the ability to sensitively and specifically detect target nucleic acid sequences in biological fluids. To reach the goal of real-life applications for such devices, however, several technological challenges related to full device integration are still to be solved, one key aspect regarding on-chip integration of the chemiluminescence signal detection device. Nowadays, the most promising approach is on-chip integration of thin-film photosensors. We recently proposed a portable cartridge with microwells aligned with an array of hydrogenated amorphous silicon (a-Si:H) photosensors, reaching attomole level limits of detection for different chemiluminescence model reactions. Herein, we explore its applicability and performance for multiplex and quantitative detection of viral DNA. In particular, the cartridge was modified to accommodate microfluidic channels and, upon immobilization of three oligonucleotide probes in different positions along each channel, each specific for a genotype of Parvovirus B19, viral nucleic acid sequences were captured and detected. With this system, taking advantage of oligoprobes specificity, chemiluminescence detectability, and photosensor sensitivity, accurate quantification of target analytes down to 70 pmol L-1 was obtained for each B19 DNA genotype, with high specificity and multiplexing ability. Results confirm the good detection capabilities and assay applicability of the proposed system, prompting the development of innovative portable analytical devices with enhanced sensitivity and multiplexed capabilities

Invisible Events with Radiative Photons at LEP

A study of the radiative neutrino counting reaction $e^+ e^- \to \nu {\bar \nu} \gamma$ at LEP1 and LEP2 energies is presented. An approximate expression for the spectrum of the observed photon is derived within the framework of the $p_t$-dependent structure function approach. This is compared with an exact expression and found in agreement within the foreseen experimental accuracy. This model describing single-photon radiation can be applied to the more general case of initial-state single-photon emission accompanying invisible final-state events. Higher-order QED corrections due to undetected initial-state radiation are also included. The implementation in a Monte Carlo event generator is briefly described.Comment: 10 pages, LaTeX, 7 figures available via anonymous ftp at: ftp://cobra1.pv.infn.it/pub/phot/, files fig#n.ps with #=1,...,

Relevant and selective activity of Pancratium illyricum L. against Candida albicans clinical isolates: a combined effect on yeast growth and virulence

BACKGROUND: Alkaloids present in plants of the Amaryllidaceae family are secondary metabolites of high biological interest, possessing a wide range of pharmacological activities. In the search for new plant-derived compounds with antimicrobial activities, two alkaloid extracts obtained from bulbs and leaves of Pancratium illyricum L., a plant of the Amarillydaceae family, were tested for their effect on bacterial and yeast growth. METHODS: The broth microdilution susceptibility test was applied to study the effect of plant extracts on the growth of reference bacterial strains and Candida albicans reference and clinical isolates strains. Extracts obtained from the different parts of the plant were tested and compared with the pure components identified in the extracts. Since matrix metalloproteinase enzymes play a role in the dissemination process of Candida albicans, the effect of the bulb extract and pure alkaloids on in vitro collagenase activity was tested. Cell viability test was carried out on human embryo lung fibroblasts (HEL 299). RESULTS: Whilst both extracts did not show any inhibitory activity against neither Gram positive nor Gram negative bacteria, a strong antifungal activity was detected, in particular for the bulb extract. All clinical isolates were susceptible to the growth inhibitory activity of the bulb extract, with endpoint IC50 values ranging from 1.22 to 78 μg/mL. The pure alkaloids lycorine and vittatine, identified as components of the extract, were also assayed for their capacity of inhibiting the yeast growth, and lycorine turned very active, with endpoint IC50 values ranging from 0.89 to 28.5 μg/mL. A potent inhibition of the in vitro collagenase activity was found in the presence of the bulb extract, and this effect was much higher than that exerted by the pure alkaloids. Viability of cell lines tested was not affected by the extract. CONCLUSIONS: Taken together, results suggest that the extract of Pancratium illyricum may act as antifungal agent both directly on the yeast growth and by altering the tissue invasion process

The Lectin-Gold Technique: An Overview of Applications to Pathological Problems

Lectins are proteins, mainly of vegetal origin, which recognize glycosidic residues with high specificity; for this property they have been used for many studies of molecular biology. The colloidal gold represents at present the most popular electron dense marker employed in immunocytochemistry, since it offers intrinsic and unique characteristics which are superior to those displayed by the other markers. The cytochemical method which utilizes the gold-labelled lectins takes advantages from both the two systems, in order to optimize the localization of the glycoconjugates. The present paper reviews both the technical aspects of the preparation of the lectin-gold complex and its application to some selected pathological problems. In particular, the papers concerning the eye and ear tissues, the urinary, reproductive, nervous and digestive systems and the blood cells are quoted

Scanning Electron Microscopy in the Study of Campylobacter Pylori Associated Gastritis

The close association between Campylobacter pylori (CP), gastritis and peptic ulcer is now well established. Moreover increasing evidence has been collected of a major etiological role of CP in type B chronic gastritis. For this reason, searching for CP is essential in all patients with upper gastrointestinal symptoms. Scanning electron microscopy (SEM) is a most reliable technique for studying the distribution of microorganisms and their relationship to the gastric mucosal surface. The aim of this paper is to compare SEM to other routine methods of detection for CP, such as Giemsa staining on histological sections and Urease Microtiter Test (MT) on fresh tissue and to investigate the surface morphology of gastric mucosa colonized by CP and to correlate it with the histopathological picture. Thirty-seven biopsies taken from the gastric body and the antrum of 22 patients were used for each type of determination. The different parameters were graded semiquantitatively. Histology showed a normal mucosa in 4 cases, chronic superficial gastritis in 12 and chronic atrophic gastritis in 21 cases. SEM was more sensitive than histology and Urease MT in detecting Campylobacter pylori. This is due to the patchy distribution of this bacterium on gastric mucosa. For this reason SEM should always be performed when routine tests are negative. The presence of CP correlated significantly (p \u3c 0.001, Spearman Rank Correlation Test) with the neutrophilic infiltrate, thus with the activity of the gastritis. The CP associated gastritis has no distinctive surface features other than the presence of the bacterium SEM morphology of surface gastric mucous cells suggests that CP does not damage the lining epithelium directly. Neutrophils and inflammatory mediators could be involved in the production of the mucosal lesions

Characterization of Mucus Glycoconjugates in Normal Human Conjunctiva by Lectins in Light Microscopy, Transmission and Scanning Electron Microscopy

Maintenance of tear film in normal conditions is dependent on 1) mucus layer integrity and 2) the presence and distribution of conjunctival epithelial cell microvilli. In the present work a new methodology has been developed to gain correlative information about microprojection assessment and mucus composition, from the same specimen, by Light Microscopy (LM), Transmission Electron Microscopy (TEM) and Scanning Electron Microscopy (SEM). We have characterized the glycosidic residues secreted by goblet cells in normal human conjunctiva, by means of four lectins (WGA, ConA, PNA and SBA), conjugated with FITC for LM and with colloidal gold for TEM and SEM. The cytochemical reactions were performed on histological sections of paraffin-embedded material and on semithin and ultrathin sections of both Epon embedded material directly processed for TEM and of blocks recovered from SEM and reprocessed for TEM. WGA, ConA, PNA and SBA receptors were found to be constituents of the mucus produced by goblet cells in human conjunctiva. The granules of the so-called second mucus system (SMS) cells were labelled mainly by WGA. A difference in the quality of glycoconjugates between goblet cells and SMS cells has been also demonstrated. Our results provide an improved method to evaluate alterations of tear film that occur in many conjunctival diseases

Importance of Tests for the Complete Lorentz Structure of the t --> W+ b vertex at Hadron Colliders

The most general Lorentz-invariant decay-density-matrix for $t\to W^{+}b\to (l^{+}\nu)b$, or for $t\to W^{+}b\to (j_{\bar d}j_u)b$, is expressed in terms of eight helicity parameters. The parameters are physically defined in terms of partial-width-intensities for polarized-final-states in $t\to W^{+}b$ decay. The parameters are the partial width, the $b$ quark's chirality parameter $\xi$, the $W^+$ polarimetry parameter $\sigma$, a "pre-SSB" test parameter $\zeta$, and four $W_{L}$-$W_{T}$ interference parameters $\eta$, $\eta^{'}$, $\omega$, $\omega^{'}$ which test for $\tilde T_{FS}$ violation. They can be used to test for non-CKM-type CP violation, anomalous $\Gamma_{L,T}$'s, top weak magnetism, weak electricity, and second-class currents. By stage-two spin-correlation techniques, percent level statistical uncertainites are typical for measurements at the Tevatron, and several mill level uncertainites are typical at the LHC.Comment: Minor clarifications. Expression for r_{+-} corrected. 19 pages LaTex + Tables + 1 Figur

Hyperfine splittings in the bbˉb\bar{b} system

Recent measurements of the $\eta_b(1S)$, the ground state of the $b\bar{b}$ system, show the splitting between it and the \Up(1S) to be 69.5$\pm$3.2 MeV, considerably larger than lattice QCD and potential model predictions, including recent calculations published by us. The models are unable to incorporate such a large hyperfine splitting within the context of a consistent description of the energy spectrum and decays. We demonstrate that in our model, which incorporates a relativistic kinetic energy term, a linear confining term including its scalar-exchange relativistic corrections, and the complete one-loop QCD short distance potential, such a consistent description, including the measured hyperfine splitting, can be obtained by not softening the delta function terms in the hyperfine potential. We calculate the hyperfine splitting to be 67.5 MeV.Comment: 5 pages, 3 tables, text revision

Antibiotic resistance, virulence factors, phenotyping, and genotyping of non\u2013escherichia coli enterobacterales from the gut microbiota of healthy subjects

Non-Escherichia coli Enterobacterales (NECE) can colonize the human gut and may present virulence determinants and phenotypes that represent severe heath concerns. Most information is available for virulent NECE strains, isolated from patients with an ongoing infection, while the commensal NECE population of healthy subjects is understudied. In this study, 32 NECE strains were isolated from the feces of 20 healthy adults. 16S rRNA gene sequencing and mass spectrometry attributed the isolates to Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter cloacae, Enterobacter aerogenes, Enterobacter kobei, Citrobacter freundii, Citrobacter amalonaticus, Cronobacter sp., and Hafnia alvei, Morganella morganii, and Serratia liquefaciens. Multiplex PCR revealed that K. pneumoniae harbored virulence genes for adhesins (mrkD, ycfM, and kpn) and enterobactin (entB) and, in one case, also for yersiniabactin (ybtS, irp1, irp2, and fyuA). Virulence genes were less numerous in the other NECE species. Biofilm formation was spread across all the species, while curli and cellulose were mainly produced by Citrobacter and Enterobacter. Among the most common antibiotics, amoxicillin-clavulanic acid was the sole against which resistance was observed, only Klebsiella strains being susceptible. The NECE inhabiting the intestine of healthy subjects have traits that may pose a health threat, taking into account the possibility of horizontal gene transfer

Correlative Scanning Electron Microscopy in the Study of Human Gastric Mucosa

We studied two aspects of the human gastric mucosa: - the surface morphology of mucous cells, as viewed by scanning electron microscopy (SEM); - the glycosidic components of intracellular mucins, characterized by means of lectins. The latter were conjugated with fluorescein isothiocyanate and with colloidal gold-silver for the visualization of the reaction products in light microscopy (LM) and in SEM (backscattered mode) respectively. The surface morphology of mucous cells appears to be correlated to the secretory state.In gastric ulcers we found a prevalence of non-secreting cells. A decrease in glycosidic receptors for fucose-binding lectin and galactose-(l-3)-N-acetyl-galactosamine-binding lectin was also observed. This suggests the presence of an impaired mucus secretion which may play a role in the pathogenesis of gastric ulcer. Spiral bacteria, supposed to be aetiologically related to peptic ulcer and gastritis, were easily detected by SEM. Intestinal metaplasia defined complete in LM showed surface morphology and glycosidic components different from those of true intestinal mucosa. This implies the necessity of taking into account also these parameters when classifying this lesion. The same applies to polyps. Our data indicate that correlative SEM may contribute further information on the pathogenesis and pathology of gastric diseases