Currently used nucleic acid amplification tests for the detection of viruses and atypicals in acute respiratory infections

For the detection of respiratory viruses conventional culture techniques are still considered as the gold standard. However, results are mostly available too late to have an impact on patient management. The latest developments include appropriate DNA- and RNA-based amplification techniques (both NASBA and PCR) for the detection of an extended number of agents responsible for LRTI. Real time amplification, the latest technical progress, produces, within a considerable shorter time, results with a lower risk of false positives. As results can be obtained within the same day, patient management with appropriate therapy or reduction of unnecessary antibiotic therapy in LRTI will be possible. A number of technical aspects of these amplification assays, and their advantages are discussed.The availability and use of these new diagnostic tools in virology has contributed to a better understanding of the role of respiratory viruses in LRTI. The increasing importance of the viral agents, Mycoplasma pneumoniae and Chlamydophila pneumoniae in ARI is illustrated. A great proportion of ARI are caused by viruses, but their relative importance depends on the spectrum of agents covered by the diagnostic techniques and on the populations studied, the geographical location and the season. The discovery of new viruses is ongoing; examples are the hMPV and the increasing number of coronaviruses. Indications for the use of these rapid techniques in different clinical situations are discussed. Depending on the possibilities, the laboratory could optimize its diagnostic strategy by applying a combination of immunofluorescence for the detection of RSV an IFL, and a combination of real-time amplification tests for other respiratory viruses and the atypical agents. When implementing a strategy, a compromise between sensitivity, clinical utility, turn around time and cost will have to be found.     

Functional aspects of serotonin transmission in the basal ganglia: a review and an in vivo approach using the push-pull cannula technique

Peripheral deafferentation of the rodent olfactory bulb results in loss of dopamine content, tyrosine hydroxylase activity and immunocytochemical staining for tyrosine hydroxylase in juxtaglomerular dopamine neurons. Reinnervation of the bulb by afferent neurons results in the return of all parameters to control levels suggesting that the dopamine neurons did not degenerate but that the expression of tyrosine hydroxylase enzyme was transneuronally regulated in a static population of juxtaglomerular cells. To evaluate this possibility, we determined the activity and immunocytochemical localization of the second enzyme in the dopamine biosynthetic pathway, DOPA decar?ylase. At a time when tyrosine hydroxylase activity was reduced to 25% of control values, DOPA decar?ylase activity in the lesioned bulb was maintained at about 65% of that in the unlesioned bulb. Immunocytochemical staining with antibodies to both enzymes, performed sequentially in the same sections, demonstrated that in the unlesioned bulb tyrosine hydroxylase and DOPA decar?ylase are co-localized in the same population of juxtaglomerular neurons. Similar results were obtained in adjacent sections each stained with one of the two antibodies. In contrast, in the deafferented bulb, about three times as many neurons were stained with DOPA decar?ylase as with tyrosine hydroxylase antibodies. The DOPA decar?ylase activity measurements and immunocytochemistry argue for the continued presence, in the lesioned olfactory bulb, of a population of tyrosine hydroxylase deficient dopamine neurons.The data suggest that olfactory receptor cell innervation transneuronally regulates the expression of tyrosine hydroxylase by mechanisms separate from those controlling the levels of DOPA decar?ylase.     

New spectral detection and elimination test algorithms of ECG and EOG artefacts in neonatal EEG recordings

New methods for the detection of ECG and EOG artefacts in the EEG are introduced, which can also be used for the evaluation of the quality of the elimination procedure. These algorithms are based on the estimation of the power or coherence spectrum by means of FFT. The advantage of this method is that the EEG spectrum is monitored by the test algorithm. The spectrum will be plotted if no influence of the ECG and EOG can be found. On the other hand, if artefacts are detected the correction of the EEG time series will be carried out and the spectrum of the corrected EEG time series will be plotted after repeated monitoring by the test algorithm.     

Computer-Assisted Detection of Pulmonary Nodules: Preliminary Observations Using a Prototype System with Multidetector-Row CT Data Sets

The continued revolution in multidetector-row CT (MDCT) scanning increases the quality of lung imaging but at the cost of a greater burden of data for review and interpretation. This article discusses our preliminary experience with prototype software for lung nodule detection and characterization using MDCT data sets. We discuss the potential role of computer-assisted detection (CAD) as applied to the automatic detection of lung nodules. We also review the process of CAD, outline its potential results, and explore how it may fit into existing radiology practice. Finally, we discuss MDCT data-acquisition parameters and how they may affect the performance of CAD.     

Oxygen Tension Imaging in the Mouse Retina

A newly developed microscope-based imaging system was used to measure the oxygen tension (PO₂) inside the retinal and choroidal vessels of mice and to generate in vivo maps of retinal PO₂. These maps were generated from the phosphorescence lifetimes of an injected palladium–porphyrin compound using a frequency-domain measurement. The system was fully calibrated and used to produce retinal PO₂ maps at different inspiratory oxygen fractions. PO₂ rose accordingly and predictably as inspiratory O₂ was stepped from hypoxic to hyperoxic conditions. Important experimental and acquisition parameters necessary for applying phosphorescence lifetime imaging to the mouse eye were investigated, including camera exposure and intensifier gain settings. Because of a need to limit light exposure to the retina, PO₂ map quality as measured by the coefficient of determination was investigated as a function of signal-to-noise and accumulated excitation energy deposition. With the development of this technology for use in mice, the potential for investigating the oxygen dynamics in genetically engineered mouse models of retinal disease, including diabetic retinopathy, glaucoma, and age-related macular degeneration, is advanced. © 2003 Biomedical Engineering Society. PAC2003: 4266Ew, 8763Lk, 8719Dd     

Impact of Breast Density on Computer-Aided Detection in Full-Field Digital Mammography

The goal of this study was to evaluate the performance of a computer-aided detection (CAD) system in full-field digital mammography (Senographe 2000D, General Electric, Buc, France) in finding out carcinomas depending on the parenchymal density. A total of 226 mediolateral oblique (MLO) and 186 craniocaudal (CC) mammographic views of histologically proven cancers were retrospectively evaluated with a digital CAD system (ImageChecker V2.3 R2 Technology, Los Altos, CA, USA). Malignant tumors were detected correctly by CAD in MLO view in 84.85% in breasts with parenchymal tissue density of the American College of Radiology (ACR) type 1, in 70.33% of the ACR type 2, in 68.12% of the ACR type 3, and in 69.70% of the ACR type 4. For the CC view, similar results were found according to the ACR types. Using the chi-square and McNemar tests, there was no statistical significance. However, a trend of better detection could be seen with decreasing ACR type. In conclusion, there seems to be a tendency for breast tissue density to affect the detection rate of breast cancer when using the CAD system.     

Design and application of sensor for recording sounds over human eye and nose

The recording of sounds over the oribt of the eye has been found to be useful in the detection of intracranial aneurysms. A hydrophone for auscultation over the eye has been developed and is tested under controlled conditions. The tests consist of measurement over the eyes in three healthy volunteers at rest, during voluntary breathing, during eyeball movements and during sustained orbicular muscular contractions. Furthermore, measurements are performed at the side of the nose. Major features of the hydrophonic transducer are high sensitivity to physiological sounds and a high degree of insensitivity to environmental sounds propagated through the air. It can be concluded that the hydrophone may be useful for the early detection of intracranial aneurysms and also for apnoea detection.     

Pyrosequencing for detection of mutations in the connexin 26 (GJB2) and mitochondrial 12S RNA (MTRNR1) genes associated with hereditary hearing loss

Hereditary hearing loss (HHL) is one of the most common congenital disorders and is highly heterogeneous. Mutations in the connexin 26 (CX26) gene (GJB2) account for about 20% of all cases of childhood deafness, and approach 50% in documented recessive cases of non-syndromic hearing loss. In addition, a single mitochondrial DNA mutation, mt1555A>G, in the 12S rRNA gene (MTRNR1), is associated with familial cases of progressive deafness. Effective screening of populations for HHL necessitates rapid assessment of several of these potential mutation sites. Pyrosequencing links a DNA synthesis protocol for determining sequence to an enzyme cascade that generates light whenever pyrophosphate is released during primer strand elongation. We assessed the ability of Pyrosequencing to detect common mutations causing HHL. Detection of the most common CX26 mutations in individuals of Caucasian (35delG), Ashkenazi (167delT), and Asian (235delC, V37I) descent was confirmed by Pyrosequencing. A total of 41 different mutations in the CX26 gene and the mitochondrial mt1555A>G mutation were confirmed. Genotyping of up to six different adjacent mutations was achieved, including simultaneous detection of 35delG and 167delT. Accurate and reproducible results were achieved taking advantage of assay flexibility and experimental conditions easily optimized for a high degree of standardization and cost-effectiveness. The standardized sample preparation steps, including target amplification by PCR and preparation of single-stranded template combined with automated sequence reaction and automated genotype scoring, positions this approach as a potentially high throughput platform for SNP/mutation genotyping in a clinical laboratory setting. .     

Detection of DNA from infectious laryngotracheitis virus by colourimetric analyses of polymerase chain reactions

A combination of the polymerase chain reaction and a novel ELISA-type DNA colourimetric assay (developed from studies with a retrovirus from man) was used in a preliminary study to detect DNA from avian infectious laryngotracheitis virus. The method is sensitive, specific and easy to perform. Since it can be readily adapted for the detection of DNA from other sources it could be useful for the identification of a variety of pathogens from other species of veterinary importance.     

“Smart” Materials for Biosensing Devices: Cell-Mimicking Supramolecular Assemblies and Colorimetric Detection of Pathogenic Agents

Mimicking cell membrane and the biomolecular recognition associated with membranes represents a great technical challenge, yet it has opened doors to innovative diagnostic and therapeutic methods. Our work has focused on design and synthesis of a class of smart materials exploiting biological principals for use in biosensors: these materials are functional polymeric assemblies that mimic the cell membrane and conveniently report the presence of pathogens with a color change. Biologically active cell membrane components are incorporated into conjugated polymers with desirable optical properties and the binding of the target molecules onto the material triggers conformational and electronic shifts that are reflected in a chromatic change (a so-called biochromic shift) that is conveniently observed and recorded. Langmuir–Blodgett thin films and vesicle bilayers provide ideal configurations for precise delivery of the biological binding entity to the sensing interface, and for control of molecular orientation for effective biomolecular interaction. Polydiacetylenic membrane-mimicking materials containing cell surface receptor gangliosides and sialic acid residues, respectively were formulated into these architectures and used for colorimetric detection of bacterial toxins and influenza virus. One advantage of these biochromic conjugated polymer (BCP) sensors is that their molecular recognition and signal transduction functionalities are resident in a single functional unit, making them amenable to convenient microfabrication and use.