Tracking bacterial pathogens with genetically-encoded reporters

AbstractDuring the infectious process, bacterial pathogens are subject to changes in environmental conditions such as nutrient availability, immune response challenges, bacterial density and physical contacts with targeted host cells. These conditions occur in the colonized organs, in diverse regions within infected tissues or even at the subcellular level for intracellular pathogens. Integration of environmental cues leads to measurable biological responses in the bacterium required for adaptation. Recent progress in technology enabled the study of bacterial adaptation in situ using genetically encoded reporters that allow single cell analysis or whole body imaging based on fluorescent proteins, alternative fluorescent assays or luciferases. This review presents a historical perspective and technical details on the methods used to develop transcriptional reporters, protein–protein interaction assays and secretion detection assays to study pathogenic bacteria adaptation in situ. Finally, studies published in the last 5years on gram positive and gram negative bacterial adaptation to the host during infection are discussed. However, the methods described here could easily be extended to study complex microbial communities within host tissue and in the environment

The `Parahippocampal Place Area' Responds Selectively to High Spatial Frequencies

Defining the exact mechanisms by which the brain processes visual objects and scenes remains an unresolved challenge. Valuable clues to this process have emerged from the demonstration that clusters of neurons (“modules”) in inferior temporal cortex apparently respond selectively to specific categories of visual stimuli, such as places/scenes. However, the higher-order “category-selective” response could also reflect specific lower-level spatial factors. Here we tested this idea in multiple functional MRI experiments, in humans and macaque monkeys, by systematically manipulating the spatial content of geometrical shapes and natural images. These tests revealed that visual spatial discontinuities (as reflected by an increased response to high spatial frequencies) selectively activate a well-known place-selective region of visual cortex (the “parahippocampal place area”) in humans. In macaques, we demonstrate a homologous cortical area, and show that it also responds selectively to higher spatial frequencies. The parahippocampal place area may use such information for detecting object borders and scene details during spatial perception and navigation.National Institutes of Health (U.S.) (NIH Grant R01 MH6752)National Institutes of Health (U.S.) (grant R01 EY017081)Athinoula A. Martinos Center for Biomedical ImagingNational Center for Research Resources (U.S.)Mind Research Institut

On the effectiveness of noise masks: Naturalistic vs. un-naturalistic image statistics

AbstractIt has been argued that the human visual system is optimized for identification of broadband objects embedded in stimuli possessing orientation averaged power spectra fall-offs that obey the 1/fβ relationship typically observed in natural scene imagery (i.e., β=2.0 on logarithmic axes). Here, we were interested in whether individual spatial channels leading to recognition are functionally optimized for narrowband targets when masked by noise possessing naturalistic image statistics (β=2.0). The current study therefore explores the impact of variable β noise masks on the identification of narrowband target stimuli ranging in spatial complexity, while simultaneously controlling for physical or perceived differences between the masks. The results show that β=2.0 noise masks produce the largest identification thresholds regardless of target complexity, and thus do not seem to yield functionally optimized channel processing. The differential masking effects are discussed in the context of contrast gain control

A neural signature of the unique hues

Since at least the 17th century there has been the idea that there are four simple and perceptually pure “unique” hues: red, yellow, green, and blue, and that all other hues are perceived as mixtures of these four hues. However, sustained scientific investigation has not yet provided solid evidence for a neural representation that separates the unique hues from other colors. We measured event-related potentials elicited from unique hues and the ‘intermediate’ hues in between them. We find a neural signature of the unique hues 230 ms after stimulus onset at a post-perceptual stage of visual processing. Specifically, the posterior P2 component over the parieto-occipital lobe peaked significantly earlier for the unique than for the intermediate hues (Z = -2.9, p = .004). Having identified a neural marker for unique hues, fundamental questions about the contribution of neural hardwiring, language and environment to the unique hues can now be addressed

Reducing Crowding by Weakening Inhibitory Lateral Interactions in the Periphery with Perceptual Learning

We investigated whether lateral masking in the near-periphery, due to inhibitory lateral interactions at an early level of central visual processing, could be weakened by perceptual learning and whether learning transferred to an untrained, higher-level lateral masking known as crowding. The trained task was contrast detection of a Gabor target presented in the near periphery (4°) in the presence of co-oriented and co-aligned high contrast Gabor flankers, which featured different target-to-flankers separations along the vertical axis that varied from 2λ to 8λ. We found both suppressive and facilitatory lateral interactions at target-to-flankers distances (2λ - 4λ and 8λ, respectively) that were larger than those found in the fovea. Training reduces suppression but does not increase facilitation. Most importantly, we found that learning reduces crowding and improves contrast sensitivity, but has no effect on visual acuity (VA). These results suggest a different pattern of connectivity in the periphery with respect to the fovea as well as a different modulation of this connectivity via perceptual learning that not only reduces low-level lateral masking but also reduces crowding. These results have important implications for the rehabilitation of low-vision patients who must use peripheral vision to perform tasks, such as reading and refined figure-ground segmentation, which normal sighted subjects perform in the fovea

Tilt aftereffect following adaptation to translational Glass patterns

Glass patterns (GPs) consist of randomly distributed dot pairs (dipoles) whose orientations are determined by specific geometric transforms. We assessed whether adaptation to stationary oriented translational GPs suppresses the activity of orientation selective detectors producing a tilt aftereffect (TAE). The results showed that adaptation to GPs produces a TAE similar to that reported in previous studies, though reduced in amplitude. This suggests the involvement of orientation selective mechanisms. We also measured the interocular transfer (IOT) of the GP-induced TAE and found an almost complete IOT, indicating the involvement of orientation selective and binocularly driven units. In additional experiments, we assessed the role of attention in TAE from GPs. The results showed that distraction during adaptation similarly modulates the TAE after adapting to both GPs and gratings. Moreover, in the case of GPs, distraction is likely to interfere with the adaptation process rather than with the spatial summation of local dipoles. We conclude that TAE from GPs possibly relies on visual processing levels in which the global orientation of GPs has been encoded by neurons that are mostly binocularly driven, orientation selective and whose adaptation-related neural activity is strongly modulated by attention

What do contrast threshold equivalent noise studies actually measure? Noise vs. nonlinearity in different masking paradigms

The internal noise present in a linear system can be quantified by the equivalent noise method. By measuring the effect that applying external noise to the system’s input has on its output one can estimate the variance of this internal noise. By applying this simple “linear amplifier” model to the human visual system, one can entirely explain an observer’s detection performance by a combination of the internal noise variance and their efficiency relative to an ideal observer. Studies using this method rely on two crucial factors: firstly that the external noise in their stimuli behaves like the visual system’s internal noise in the dimension of interest, and secondly that the assumptions underlying their model are correct (e.g. linearity). Here we explore the effects of these two factors while applying the equivalent noise method to investigate the contrast sensitivity function (CSF). We compare the results at 0.5 and 6 c/deg from the equivalent noise method against those we would expect based on pedestal masking data collected from the same observers. We find that the loss of sensitivity with increasing spatial frequency results from changes in the saturation constant of the gain control nonlinearity, and that this only masquerades as a change in internal noise under the equivalent noise method. Part of the effect we find can be attributed to the optical transfer function of the eye. The remainder can be explained by either changes in effective input gain, divisive suppression, or a combination of the two. Given these effects the efficiency of our observers approaches the ideal level. We show the importance of considering these factors in equivalent noise studies

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field