A novel universal DNA labeling and amplification system for rapid microarray-based detection of 117 antibiotic resistance genes in Gram-positive bacteria.

A rapid and simple DNA labeling system has been developed for disposable microarrays and has been validated for the detection of 117 antibiotic resistance genes abundant in Gram-positive bacteria. The DNA was fragmented and amplified using phi-29 polymerase and random primers with linkers. Labeling and further amplification were then performed by classic PCR amplification using biotinylated primers specific for the linkers. The microarray developed by Perreten et al. (Perreten, V., Vorlet-Fawer, L., Slickers, P., Ehricht, R., Kuhnert, P., Frey, J., 2005. Microarray-based detection of 90 antibiotic resistance genes of gram-positive bacteria. J.Clin.Microbiol. 43, 2291-2302.) was improved by additional oligonucleotides. A total of 244 oligonucleotides (26 to 37 nucleotide length and with similar melting temperatures) were spotted on the microarray, including genes conferring resistance to clinically important antibiotic classes like β-lactams, macrolides, aminoglycosides, glycopeptides and tetracyclines. Each antibiotic resistance gene is represented by at least 2 oligonucleotides designed from consensus sequences of gene families. The specificity of the oligonucleotides and the quality of the amplification and labeling were verified by analysis of a collection of 65 strains belonging to 24 species. Association between genotype and phenotype was verified for 6 antibiotics using 77 Staphylococcus strains belonging to different species and revealed 95% test specificity and a 93% predictive value of a positive test. The DNA labeling and amplification is independent of the species and of the target genes and could be used for different types of microarrays. This system has also the advantage to detect several genes within one bacterium at once, like in Staphylococcus aureus strain BM3318, in which up to 15 genes were detected. This new microarray-based detection system offers a large potential for applications in clinical diagnostic, basic research, food safety and surveillance programs for antimicrobial resistance.

Detection of back-to-back proton pairs in charged-current neutrino interactions with the ArgoNeuT detector in the NuMI low energy beam line

Short range nucleon-nucleon correlations in nuclei (NN SRC) carry important information on nuclear structure and dynamics. NN SRC have been extensively probed through two-nucleon knock- out reactions in both pion and electron scattering experiments. We report here on the detection of two-nucleon knock-out events from neutrino interactions and discuss their topological features as possibly involving NN SRC content in the target argon nuclei. The ArgoNeuT detector in the Main Injector neutrino beam at Fermilab has recorded a sample of 30 fully reconstructed charged current events where the leading muon is accompanied by a pair of protons at the interaction vertex, 19 of which have both protons above the Fermi momentum of the Ar nucleus. Out of these 19 events, four are found with the two protons in a strictly back-to-back high momenta configuration directly observed in the final state and can be associated to nucleon Resonance pionless mechanisms involving a pre-existing short range correlated np pair in the nucleus. Another fraction (four events) of the remaining 15 events have a reconstructed back-to-back configuration of a np pair in the initial state, a signature compatible with one-body Quasi Elastic interaction on a neutron in a SRC pair. The detection of these two subsamples of the collected (mu- + 2p) events suggests that mechanisms directly involving nucleon-nucleon SRC pairs in the nucleus are active and can be efficiently explored in neutrino-argon interactions with the LAr TPC technology.

Unified detection and tracking of instruments during retinal microsurgery

Methods for tracking an object have generally fallen into two groups: tracking by detection and tracking through local optimization. The advantage of detection-based tracking is its ability to deal with target appearance and disappearance, but it does not naturally take advantage of target motion continuity during detection. The advantage of local optimization is efficiency and accuracy, but it requires additional algorithms to initialize tracking when the target is lost. To bridge these two approaches, we propose a framework for unified detection and tracking as a time-series Bayesian estimation problem. The basis of our approach is to treat both detection and tracking as a sequential entropy minimization problem, where the goal is to determine the parameters describing a target in each frame. To do this we integrate the Active Testing (AT) paradigm with Bayesian filtering, and this results in a framework capable of both detecting and tracking robustly in situations where the target object enters and leaves the field of view regularly. We demonstrate our approach on a retinal tool tracking problem and show through extensive experiments that our method provides an efficient and robust tracking solution.

Field-Applicable Recombinase Polymerase Amplification Assay for Rapid Detection of Mycoplasma capricolum subsp. capripneumoniae.

Contagious caprine pleuropneumonia (CCPP) is a highly contagious disease caused by Mycoplasma capricolum subsp. capripneumoniae that affects goats in Africa and Asia. Current available methods for the diagnosis of Mycoplasma infection, including cultivation, serological assays, and PCR, are time-consuming and require fully equipped stationary laboratories, which make them incompatible with testing in the resource-poor settings that are most relevant to this disease. We report a rapid, specific, and sensitive assay employing isothermal DNA amplification using recombinase polymerase amplification (RPA) for the detection of M. capricolum subsp. capripneumoniae. We developed the assay using a specific target sequence in M. capricolum subsp. capripneumoniae, as found in the genome sequence of the field strain ILRI181 and the type strain F38 and that was further evidenced in 10 field strains from different geographical regions. Detection limits corresponding to 5 × 10(3) and 5 × 10(4) cells/ml were obtained using genomic DNA and bacterial culture from M. capricolum subsp. capripneumoniae strain ILRI181, while no amplification was obtained from 71 related Mycoplasma isolates or from the Acholeplasma or the Pasteurella isolates, demonstrating a high degree of specificity. The assay produces a fluorescent signal within 15 to 20 min and worked well using pleural fluid obtained directly from CCPP-positive animals without prior DNA extraction. We demonstrate that the diagnosis of CCPP can be achieved, with a short sample preparation time and a simple read-out device that can be powered by a car battery, in <45 min in a simulated field setting.

Reduced foveal vision enhances peripheral monitoring and peripheral event detection

Introduction: Although it seems plausible that sports performance relies on high-acuity foveal vision, it could be empirically shown that myoptic blur (up to +2 diopters) does not harm performance in sport tasks that require foveal information pick-up like golf putting (Bulson, Ciuffreda, & Hung, 2008). How myoptic blur affects peripheral performance is yet unknown. Attention might be less needed for processing visual cues foveally and lead to better performance because peripheral cues are better processed as a function of reduced foveal vision, which will be tested in the current experiment. Methods: 18 sport science students with self-reported myopia volunteered as participants, all of them regularly wearing contact lenses. Exclusion criteria comprised visual correction other than myopic, correction of astigmatism and use of contact lenses out of Swiss delivery area. For each of the participants, three pairs of additional contact lenses (besides their regular lenses; used in the “plano” condition) were manufactured with an individual overcorrection to a retinal defocus of +1 to +3 diopters (referred to as “+1.00 D”, “+2.00 D”, and “+3.00 D” condition, respectively). Gaze data were acquired while participants had to perform a multiple object tracking (MOT) task that required to track 4 out of 10 moving stimuli. In addition, in 66.7 % of all trials, one of the 4 targets suddenly stopped during the motion phase for a period of 0.5 s. Stimuli moved in front of a picture of a sports hall to allow for foveal processing. Due to the directional hypotheses, the level of significance for one-tailed tests on differences was set at α = .05 and posteriori effect sizes were computed as partial eta squares (ηρ2). Results: Due to problems with the gaze-data collection, 3 participants had to be excluded from further analyses. The expectation of a centroid strategy was confirmed because gaze was closer to the centroid than the target (all p < .01). In comparison to the plano baseline, participants more often recalled all 4 targets under defocus conditions, F(1,14) = 26.13, p < .01, ηρ2 = .65. The three defocus conditions differed significantly, F(2,28) = 2.56, p = .05, ηρ2 = .16, with a higher accuracy as a function of a defocus increase and significant contrasts between conditions +1.00 D and +2.00 D (p = .03) and +1.00 D and +3.00 D (p = .03). For stop trials, significant differences could neither be found between plano baseline and defocus conditions, F(1,14) = .19, p = .67, ηρ2 = .01, nor between the three defocus conditions, F(2,28) = 1.09, p = .18, ηρ2 = .07. Participants reacted faster in “4 correct+button” trials under defocus than under plano-baseline conditions, F(1,14) = 10.77, p < .01, ηρ2 = .44. The defocus conditions differed significantly, F(2,28) = 6.16, p < .01, ηρ2 = .31, with shorter response times as a function of a defocus increase and significant contrasts between +1.00 D and +2.00 D (p = .01) and +1.00 D and +3.00 D (p < .01). Discussion: The results show that gaze behaviour in MOT is not affected to a relevant degree by a visual overcorrection up to +3 diopters. Hence, it can be taken for granted that peripheral event detection was investigated in the present study. This overcorrection, however, does not harm the capability to peripherally track objects. Moreover, if an event has to be detected peripherally, neither response accuracy nor response time is negatively affected. Findings could claim considerable relevance for all sport situations in which peripheral vision is required which now needs applied studies on this topic. References: Bulson, R. C., Ciuffreda, K. J., & Hung, G. K. (2008). The effect of retinal defocus on golf putting. Ophthalmic and Physiological Optics, 28, 334-344.

Detecting single-target changes in multiple object tracking: The case of peripheral vision

In the current study it is investigated whether peripheral vision can be used to monitor multi-ple moving objects and to detect single-target changes. For this purpose, in Experiment 1, a modified MOT setup with a large projection and a constant-position centroid phase had to be checked first. Classical findings regarding the use of a virtual centroid to track multiple ob-jects and the dependency of tracking accuracy on target speed could be successfully replicat-ed. Thereafter, the main experimental variations regarding the manipulation of to-be-detected target changes could be introduced in Experiment 2. In addition to a button press used for the detection task, gaze behavior was assessed using an integrated eye-tracking system. The anal-ysis of saccadic reaction times in relation to the motor response shows that peripheral vision is naturally used to detect motion and form changes in MOT because the saccade to the target occurred after target-change offset. Furthermore, for changes of comparable task difficulties, motion changes are detected better by peripheral vision than form changes. Findings indicate that capabilities of the visual system (e.g., visual acuity) affect change detection rates and that covert-attention processes may be affected by vision-related aspects like spatial uncertainty. Moreover, it is argued that a centroid-MOT strategy might reduce the amount of saccade-related costs and that eye-tracking seems to be generally valuable to test predictions derived from theories on MOT. Finally, implications for testing covert attention in applied settings are proposed.