Fast Optimized Cluster Algorithm for Localizations (FOCAL): a spatial cluster analysis for super-resolved microscopy.

MOTIVATION: Single-molecule localization microscopy (SMLM) microscopy provides images of cellular structure at a resolution an order of magnitude below what can be achieved by conventional diffraction limited techniques. The concomitantly larger data sets generated by SMLM require increasingly efficient image analysis software. Density based clustering algorithms, with the most ubiquitous being DBSCAN, are commonly used to quantitatively assess sub-cellular assemblies. DBSCAN, however, is slow, scaling with the number of localizations like O(n log (n)) at best, and it's performance is highly dependent upon a subjectively selected choice of parameters. RESULTS: We have developed a grid-based clustering algorithm FOCAL, which explicitly accounts for several dominant artifacts arising in SMLM image reconstructions. FOCAL is fast and efficient, scaling like O(n), and only has one set parameter. We assess DBSCAN and FOCAL on experimental dSTORM data of clusters of eukaryotic RNAP II and PALM data of the bacterial protein H-NS, then provide a detailed comparison via simulation. FOCAL performs comparable and often superior to DBSCAN while yielding a significantly faster analysis. Additionally, FOCAL provides a novel method for filtering out of focus clusters from complex SMLM images. AVAILABILITY AND IMPLEMENTATION: The data and code are available at: http://www.utm.utoronto.ca/milsteinlab/resources/Software/FOCAL/ CONTACT: josh.milstein@utoronto.ca SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

An exploration of connections between a mother and her unborn in preterm labor.

The purpose of this special feature is to elaborate on a unique physician-patient interaction between a neonatologist and a pediatrician established during the pediatrician's personal experience with the onset of labor between 22 and 23 weeks' gestation. As the two are both versed in conventional aspects of neonatal care including prenatal counseling, the neonatologist thought that he had more to offer the expectant mother holistically. Thus, he decided against approaching her as an information source and contribute to the customary data-dumping format that can be both frightening and dehumanizing. Instead, he wanted to focus on her individual experience of pregnancy and the healing aspects of care. The most important lesson received from the interaction is the value of being present with another. This sense of 'being with' applied to the neonatologist and the mother from their initial phone conversation to their in person times, and to the mother with her unborn permitting interaction in a meaningful way. Fostering each of those connections in alternative ways was valuable in this prenatal experience.

On the role of DNA biomechanics in the regulation of gene expression.

DNA is traditionally seen as a linear sequence of instructions for cellular functions that are expressed through biochemical processes. Cellular DNA, however, is also organized as a complex hierarchical structure with a mosaic of mechanical features, and a growing body of evidence is now emerging to imply that these mechanical features are connected to genetic function. Mechanical tension, for instance, which must be felt by DNA within the heavily constrained and continually fluctuating cellular environment, can affect a number of regulatory processes implicating a role for biomechanics in gene expression complementary to that of biochemical regulation. In this article, we review evidence for such mechanical pathways of genetic regulation.

Bead size effects on protein-mediated DNA looping in tethered-particle motion experiments.

Tethered particle motion (TPM) has become an important tool for single-molecule studies of biomolecules; however, concerns remain that the method may alter the dynamics of the biophysical process under study. We investigate the effect of the attached microsphere on an illustrative biological example: the formation and breakdown of protein-mediated DNA loops in the lac repressor system. By comparing data from a conventional TPM experiment with 800 nm polystyrene beads and dark-field TPM using 50 nm Au nanoparticles, we found that the lifetimes of the looped and unlooped states are only weakly modified, less than two-fold, by the presence of the large bead. This is consistent with our expectation of weak excluded-volume effects and hydrodynamic surface interactions from the cover glass and microsphere.

Protein-mediated DNA loop formation and breakdown in a fluctuating environment.

Living cells provide a fluctuating, out-of-equilibrium environment in which genes must coordinate cellular function. DNA looping, which is a common means of regulating transcription, is very much a stochastic process; the loops arise from the thermal motion of the DNA and other fluctuations of the cellular environment. We present single-molecule measurements of DNA loop formation and breakdown when an artificial fluctuating force, applied to mimic a fluctuating cellular environment, is imposed on the DNA. We show that loop formation is greatly enhanced in the presence of noise of only a fraction of k_{B}T, yet find that hypothetical regulatory schemes that employ mechanical tension in the DNA-as a sensitive switch to control transcription-can be surprisingly robust due to a fortuitous cancellation of noise effects.

Femtonewton entropic forces can control the formation of protein-mediated DNA loops.

We show that minuscule entropic forces, on the order of 100 fN, can prevent the formation of DNA loops-a ubiquitous means of regulating the expression of genes. We observe a tenfold decrease in the rate of LacI-mediated DNA loop formation when a tension of 200 fN is applied to the substrate DNA, biasing the thermal fluctuations that drive loop formation and breakdown events. Conversely, once looped, the DNA-protein complex is insensitive to applied force. Our measurements are in excellent agreement with a simple polymer model of loop formation in DNA, and show that an antiparallel topology is the preferred LacI-DNA loop conformation for a generic loop-forming construct.

Methylphenidate-induced impulsivity: pharmacological antagonism by beta-adrenoreceptor blockade.

Noradrenaline-dopamine interactions mediate increases in locomotor activity, development of sensitisation and subjective effects of psychostimulant drugs. However, the modulatory effects of noradrenaline on psychostimulant-induced impulsivity are less clear. This article examined the relative roles of noradrenaline and dopamine in the modulation of methylphenidate-induced impulsive responding in rats performing the 5-choice serial reaction time task. Experiment 1 examined the systemic antagonism of methylphenidate-induced impulsivity with either propranolol, a beta-adrenoreceptor blocker, or prazosin, an alpha1-adrenoreceptor antagonist, which antagonises the locomotor activating effects of amphetamine. Propranolol completely abolished methylphenidate-induced impulsivity. This effect was centrally rather than peripherally mediated, as nadolol, a peripheral beta-blocker failed to affect methylphenidate-induced premature responding. Prazosin partially attenuated the methylphenidate-mediated increase in premature responding. A second experiment examined the effects of selective anti-D beta H saporin-induced cortical noradrenaline depletion on methylphenidate-induced impulsivity. Contrary to the effects of beta-adrenoreceptor blockade, cortical noradrenergic depletion did not alter methylphenidate-induced impulsivity. Other experiments examined the comparative effects of selective dopamine and serotonin receptor blockade. D4 dopamine receptor blockade with systemically administered L-745,870 also attenuated methylphenidate-induced impulsivity. The other antagonists had no effect on methylphenidate-induced impulsivity. Taken together, these studies provide evidence for a modulatory role of beta-adrenoreceptors on methylphenidate-induced impulsive responding.

A statewide quality improvement collaborative to reduce neonatal central line-associated blood stream infections.

OBJECTIVE: The objective of this study was to reduce central line-associated blood stream infections (CLABSIs) among 13 collaborating regional neonatal intensive care units by 25%. We tested the hypothesis that change could be attributed to the quality improvement collaborative by testing for 'special cause' variation. STUDY DESIGN: Our prevention project included five features: (1) leadership commitment, (2) potentially best practices, (3) collaborative processes, (4) audit and feedback tools and (5) quality improvement techniques. Baseline (1 January 2006 to 30 August 2006) data were compared with the intervention (1 September 2006 to 30 June 2007) and post-intervention (1 July 2007 to 30 December 2007) periods and analyzed using statistical process control (SPC) methods. RESULT: We detected special cause variation, suggesting that the collaborative was associated with reduced infection rates, from 4.32 to 3.22 per 1000 line days (a 25% decrease) when comparing the baseline with the follow-up period. CONCLUSION: The collaborative's process was associated with fewer infections. SPC suggested that systematic changes occurred. The remaining challenges include sustaining or even further reducing the infection rate.

Indomethacin-associated neutropenia with subsequent Gram-negative sepsis in a preterm infant. Cause or coincidence?

A preterm male infant with a patent ductus arteriosus developed neutropenia during treatment with indomethacin. Afterward, the mother described her own history of indomethacin-associated neutropenia. During the recovery from the neutropenia, the infant became septic with bacteremia caused by Enterobacter cloacae. Although indomethacin-related neutropenia has been described in adults, no case in a neonate has been reported. If neutropenia occurs after indomethacin therapy in a neonate, a familial history of indomethacin-associated neutropenia should be sought and the increased risk of infection should be considered.

Signatures of resonance superfluidity in a quantum Fermi gas.

We predict a direct and observable signature of the superfluid phase in a quantum Fermi gas, in a temperature regime already accessible in current experiments. We apply the theory of resonance superfluidity to a gas confined in a harmonic potential and demonstrate that a significant increase in density will be observed in the vicinity of the trap center.