Highly accurate classification of biological spores by culture medium for forensic attribution using multiple chemical signature types and machine learning.

Future proliferation of biological expertise and new technology may increasingly lower the difficulty to produce biological organisms for misuse. Rapid attribution of a biological attack is needed to quickly identify the person or lab responsible and prevent additional attacks by enabling the apprehension of suspects. Here, triplicate batches of Bacillus anthracis Sterne strain (BaSt) spores were grown in a total of seven amateur and professional media. Multiple orthogonal analytical signatures (peptides, metabolites, lipids by fatty acid methyl ester (FAME) analysis, bulk organic profile, and trace elements) were collected from the BaSt spores. The proteomics and metabolomics analyses identified promising attribution signature compounds that are unique to each of the seven production methods. In addition, while each of the signature types showed varying degrees of value individually for attributing BaSt spores to the culture medium used to prepare them, fusing results from all five signatures types to increase sourcing robustness and using a random forest sourcing algorithm yielded 100% hold-one-batch-out cross-validation classification accuracy and an average relative source probability for the correct source 5.5× higher than the most probable incorrect source. These preliminary results provide a proof-of-concept for the development of forensic examinations that can attribute biological agents to production methods for use in future investigations.

Composite Machine Learning Algorithm for Material Sourcing.

This study developed a composite machine learning algorithm for attribution of materials of forensic interest (like ammonium nitrate) to original sources. k-nearest neighbor and random forest models were used for source elimination and classification, respectively, in a two-step, composite algorithm based on particle color, size/shape, and trace element concentration features. Novel approaches for simulation to supplement within-source reference features based on empirically measured multi-lot analyses, an improved hold-one-lot-out method for cross-validation, an assessment of the likelihood of the presence of a reference sample, fusion of the source probabilities from the respective classification models, and the calculation of metrics for assessing ensemble sourcing performance are described. Excellent sourcing predictions were obtained; the sourcing algorithm identified the correct source as the top choice 89% of the time, and the correct source was identified to be an average of 2.7 times more likely than the most likely incorrect source.

Cortical Interneuron Subtypes Vary in Their Axonal Action Potential Properties.

The role of interneurons in cortical microcircuits is strongly influenced by their passive and active electrical properties. Although different types of interneurons exhibit unique electrophysiological properties recorded at the soma, it is not yet clear whether these differences are also manifested in other neuronal compartments. To address this question, we have used voltage-sensitive dye to image the propagation of action potentials into the fine collaterals of axons and dendrites in two of the largest cortical interneuron subtypes in the mouse: fast-spiking interneurons, which are typically basket or chandelier neurons; and somatostatin containing interneurons, which are typically regular spiking Martinotti cells. We found that fast-spiking and somatostatin-expressing interneurons differed in their electrophysiological characteristics along their entire dendrosomatoaxonal extent. The action potentials generated in the somata and axons, including axon collaterals, of somatostatin-expressing interneurons are significantly broader than those generated in the same compartments of fast-spiking inhibitory interneurons. In addition, action potentials back-propagated into the dendrites of somatostatin-expressing interneurons much more readily than fast-spiking interneurons. Pharmacological investigations suggested that axonal action potential repolarization in both cell types depends critically upon Kv1 channels, whereas the axonal and somatic action potentials of somatostatin-expressing interneurons also depend on BK Ca(2+)-activated K(+) channels. These results indicate that the two broad classes of interneurons studied here have expressly different subcellular physiological properties, allowing them to perform unique computational roles in cortical circuit operations. SIGNIFICANCE STATEMENT: Neurons in the cerebral cortex are of two major types: excitatory and inhibitory. The proper balance of excitation and inhibition in the brain is critical for its operation. Neurons contain three main compartments: dendritic, somatic, and axonal. How the neurons receive information, process it, and pass on new information depends upon how these three compartments operate. While it has long been assumed that axons are simply for conducting information from the cell body to the synapses, here we demonstrate that the axons of different types of interneurons, the inhibitory cells, possess differing electrophysiological properties. This result implies that differing types of interneurons perform different tasks in the cortex, not only through their anatomical connections, but also through how their axons operate.

Motor cortex feedback influences sensory processing by modulating network state.

Long-range corticocortical communication may have important roles in context-dependent sensory processing, yet we know very little about how these pathways influence their target regions. We studied the influence of primary motor cortex activity on primary somatosensory cortex in the mouse whisker system. We show that primary motor and somatosensory cortices undergo coherent, context-dependent changes in network state. Moreover, we show that motor cortex activity can drive changes in somatosensory cortex network state. A series of experiments demonstrate the involvement of the direct corticocortical feedback pathway, providing temporally precise and spatially targeted modulation of network dynamics. Cortically mediated changes in network state significantly impact sensory coding, with activated states increasing the reliability of responses to complex stimuli. By influencing network state, corticocortical communication from motor cortex may ensure that during active exploration the relevant sensory region is primed for enhanced sensory discrimination.

Active action potential propagation but not initiation in thalamic interneuron dendrites.

Inhibitory interneurons of the dorsal lateral geniculate nucleus of the thalamus modulate the activity of thalamocortical cells in response to excitatory input through the release of inhibitory neurotransmitter from both axons and dendrites. The exact mechanisms by which release can occur from dendrites are, however, not well understood. Recent experiments using calcium imaging have suggested that Na/K-based action potentials can evoke calcium transients in dendrites via local active conductances, making the backpropagating action potential a candidate for dendritic neurotransmitter release. In this study, we used high temporal and spatial resolution voltage-sensitive dye imaging to assess the characteristics of dendritic voltage deflections in response to Na/K action potentials in interneurons of the mouse dorsal lateral geniculate nucleus. We found that trains or single action potentials elicited by somatic current injection or local synaptic stimulation rapidly and actively backpropagated throughout the entire dendritic arbor and into the fine filiform dendritic appendages known to release GABAergic vesicles. Action potentials always appeared first in the soma or proximal dendrite in response to somatic current injection or local synaptic stimulation, and the rapid backpropagation into the dendritic arbor depended upon voltage-gated sodium and tetraethylammonium chloride-sensitive potassium channels. Our results indicate that thalamic interneuron dendrites integrate synaptic inputs that initiate action potentials, most likely in the axon initial segment, that then backpropagate with high fidelity into the dendrites, resulting in a nearly synchronous release of GABA from both axonal and dendritic compartments.

Gene expression profiling of whole blood: comparison of target preparation methods for accurate and reproducible microarray analysis.

BACKGROUND: Peripheral blood is an accessible and informative source of transcriptomal information for many human disease and pharmacogenomic studies. While there can be significant advantages to analyzing RNA isolated from whole blood, particularly in clinical studies, the preparation of samples for microarray analysis is complicated by the need to minimize artifacts associated with highly abundant globin RNA transcripts. The impact of globin RNA transcripts on expression profiling data can potentially be reduced by using RNA preparation and labeling methods that remove or block globin RNA during the microarray assay. We compared four different methods for preparing microarray hybridization targets from human whole blood collected in PAXGene tubes. Three of the methods utilized the Affymetrix one-cycle cDNA synthesis/in vitro transcription protocol but varied treatment of input RNA as follows: i. no treatment; ii. treatment with GLOBINclear; or iii. treatment with globin PNA oligos. In the fourth method cDNA targets were prepared with the Ovation amplification and labeling system. RESULTS: We find that microarray targets generated with labeling methods that reduce globin mRNA levels or minimize the impact of globin transcripts during hybridization detect more transcripts in the microarray assay compared with the standard Affymetrix method. Comparison of microarray results with quantitative PCR analysis of a panel of genes from the NF-kappa B pathway shows good correlation of transcript measurements produced with all four target preparation methods, although method-specific differences in overall correlation were observed. The impact of freezing blood collected in PAXGene tubes on data reproducibility was also examined. Expression profiles show little or no difference when RNA is extracted from either fresh or frozen blood samples. CONCLUSION: RNA preparation and labeling methods designed to reduce the impact of globin mRNA transcripts can significantly improve the sensitivity of the DNA microarray expression profiling assay for whole blood samples. While blockage of globin transcripts during first strand cDNA synthesis with globin PNAs resulted in the best overall performance in this study, we conclude that selection of a protocol for expression profiling studies in blood should depend on several factors, including implementation requirements of the method and study design. RNA isolated from either freshly collected or frozen blood samples stored in PAXGene tubes can be used without altering gene expression profiles.

Influence of path integration versus environmental orientation on place cell remapping between visually identical environments.

To assess the effects of interactions between angular path integration and visual landmarks on the firing of hippocampal neurons, we recorded from CA1 pyramidal cells as rats foraged in two identical boxes with polarizing internal cues. In the same-orientation condition, following an earlier experiment by Skaggs and McNaughton, the boxes were oriented identically and connected by a corridor. In the opposite-orientation condition, the boxes were abutted by rotating them 90 degrees in opposite directions, so that their orientations differed by 180 degrees . After 16-23 days of pretraining on the same-orientation condition, three rats experienced both conditions in counterbalanced order on each of two consecutive days. On the third day they ran two opposite-orientation trials. Although Skaggs and McNaughton observed stable partial "remapping" of place fields, none of the fields in this experiment remapped in the same-orientation condition. In the opposite-orientation condition, place fields in the first box were isomorphic with those in the same-orientation condition, whereas in the second box the rats eventually exhibited completely different fields. The rats differed as to the trial in which this first occurred. Once the second box exhibited different fields, it continued to do so in all subsequent opposite-orientation trials, yet fields remained the same in subsequent same-orientation trials. The results demonstrate that when animals move actively between environments, and are thus potentially able to maintain their inertial angular orientation, discordance between environmental orientation and the rat's idiothetic direction sense can profoundly affect the hippocampal map-either immediately, or as a result of cumulative experience.