Rapid Visual Authentication Based on DNA Strand Displacement.

Novel ways to track and verify items of a high value or security is an ever-present need. Taggants made from deoxyribonucleic acid (DNA) have several advantageous properties, such as high information density and robust synthesis; however, existing methods require laboratory techniques to verify, limiting applications. Here, we leverage DNA nanotechnology to create DNA taggants that can be validated in the field in seconds to minutes with a simple equipment. The system is driven by toehold-mediated strand-displacement reactions where matching oligonucleotide sequences drive the generation of a fluorescent signal through the potential energy of base pairing. By pooling different "input" oligonucleotide sequences in a taggant and spatially separating "reporter" oligonucleotide sequences on a paper ticket, unique, sequence-driven patterns emerge for different taggant formulations. Algorithmically generated oligonucleotide sequences show no crosstalk and ink-embedded taggants maintain activity for at least 99 days at 60 °C (equivalent to nearly 2 years at room temperature). The resulting fluorescent signals can be analyzed by the eye or a smartphone when paired with a UV flashlight and filtered glasses.

Anti-counterfeiting DNA molecular tagging of pharmaceutical excipients: An evaluation of lactose containing tablets.

The licensed pharmaceutical industry and regulators use many approaches to control counterfeiting, but it remains a very difficult task to differentiate between counterfeit and real products. Moreover, there is a lack of techniques available for providing a batch specific molecular bar code for tablets that has the required traceability, specificity and sensitivity to be fit for purpose. The aim of this study was to evaluate DNA molecular tags as a potential anti-counterfeiting technology in tablets. Lactose tablets (400 mg) were used as a model to investigate incorporation DNA molecular tag into a solid dosage form: DNA authentication was carried out on an Applied DNA SigNify® qPCR instrument. Tablet batches were subjected to accelerated stability conditions (40 °C and 75% RH) for up to 6 months. All batches passed the monograph specifications of the British Pharmacopoeia (hardness, friability and mass uniformity) throughout the storage period. In all of recovery plots, the number of cycles required for DNA detection (Cq values) increased as a function of storage time, which indicated a reduction in tag levels, but it should be noted for all storage experiments the tag was clearly detected. It would appear that DNA molecular tags could feasibly be applied within the pharmaceutical development cycle when a new solid dosage form is brought to the market so as to mitigate the risk and dangers of counterfeiting.

Rapid authentication of pharmaceuticals via DNA tagging and field detection.

A small PCR-generated DNA fragment was introduced into a pharmaceutical grade ink as a molecular taggant, and the DNA tagged ink was delivered onto the surface of capsules by standard high-speed offset printing. The amount of DNA in the ink on each capsule is roughly 10-12 fold lower than that allowed as safe by the United States Food and Drug Administration (FDA) and the WHO with regards to acceptable limits of residual DNA. The printed ink on the capsule surface was sampled by swabbing, followed by direct analysis of the DNA-swab complex, without subsequent DNA purification. It was shown that DNA recovered from the ink by swabbing was suitable for PCR-CE analysis-a widely used method in forensic science and was also suitable for qPCR and isothermal DNA amplification, when coupled with portable devices similar to those used for environmental sampling and food safety testing. The data set a precedent: A small DNA fragment could be introduced as an excipient into a pharmaceutical application, and thereafter tracked through the pharmaceutical supply chain via forensic DNA authentication.

DNA storage under high temperature conditions does not affect performance in human leukocyte antigen genotyping via next-generation sequencing (DNA integrity maintained in extreme conditions).

BACKGROUND: Stable dry-state storage of DNA is desirable to minimize required storage space and to reduce electrical and shipping costs. DNA purified from various commercially available dry-state stabilization matrices has been used successfully in downstream molecular applications (e.g., quantitative polymerase chain reaction [qPCR], microarray, and sequence-based genotyping). However, standard DNA storage conditions still include freezing of DNA eluted in aqueous buffers or nuclease-free water. Broad implementation of dry-state, long-term DNA storage requires enhancement of such dry-state DNA stabilization products to control for temperature fluctuations at specimen collection, transit, and storage. This study tested the integrity of genomic DNA subjected to long-term storage on GenTegra(™) DNA stabilization matrices (GenTegra LLC, Pleasanton, CA) at extreme conditions, as defined by a 4-year storage period at ambient temperature with an initial incubation for 7 months at 37°C, 56°C, or ambient temperature. Subsequently, purified DNA performance and integrity were measured by qPCR and next-generation sequencing (NGS)-based human leokocyte antigen (HLA) genotyping. RESULTS: High molecular weight genomic DNA samples were recovered from the GenTegra product matrix and exhibited integrity comparable to a highly characterized commercial standard under assessment by qPCR. Samples were genotyped for classical HLA loci using next generation sequencing-based methodolgy on the Roche 454 GS Junior instrument. Amplification efficiency, sequence coverage, and sequence quality were all comparable with those produced from a cell line DNA sequenced as a control. No significant differences were observed in the mean, median, or mode quality scores between samples and controls (p≥0.4). CONCLUSIONS: Next generation HLA genotyping was chosen to test the integrity of GenTegra-treated genomic DNA due to the requirment for long sequence reads to genotype the highly polymorphic classical HLA genes. Experimental results demonstrate the efficacy of the GenTegra product as a suitable genomic DNA preservation tool for collection and long-term biobanking of DNA at fluctuating and high temperatures.

Increased rates of events that activate or deactivate the behavioral approach system, but not events related to goal attainment, in bipolar spectrum disorders.

Research indicates that life events involving goal attainment and goal striving trigger hypomania/mania and that negative life events trigger bipolar depression. These findings are consistent with the behavioral approach system (BAS) dysregulation model of bipolar disorders, which suggests that individuals with bipolar disorders are hypersensitive to cues signaling opportunity for reward and cues signaling failure and loss of rewards. However, no studies to date have investigated whether individuals with bipolar spectrum disorders experience increased rates of these BAS-relevant life events, which would place them at double risk for developing bipolar episodes. The present study found that individuals with bipolar II disorder and cyclothymia experience increased rates of BAS-activating and BAS-deactivating, but not goal-attainment, life events. Finally, for bipolar spectrum individuals only, BAS-activating events predicted BAS-deactivating events' rates.

Behavioral approach system (BAS)-relevant cognitive styles and bipolar spectrum disorders: concurrent and prospective associations.

The authors examined concurrent and prospective associations of behavioral approach system (BAS)-relevant and non-BAS-relevant cognitive styles with bipolar spectrum disorders. Controlling for depressive and hypomanic/manic symptoms, 195 individuals with bipolar spectrum disorders scored higher than 194 demographically similar normal controls on BAS sensitivity and BAS-relevant cognitive dimensions of performance concerns, autonomy, and self-criticism, but not on behavioral inhibition system sensitivity and non-BAS-relevant dimensions of approval seeking, sociotropy, and dependency. Moreover, group differences on autonomy fully mediated the association between higher BAS sensitivity and bipolar status. In addition, only BAS-related cognitive dimensions predicted the likelihood of onset of depressive and hypomanic/manic episodes among the bipolar individuals over a 3.2-year follow-up, controlling for initial symptoms and past history of mood episodes. Higher autonomy and self-criticism predicted a greater likelihood of hypomanic/manic episodes, and higher autonomy predicted a lower likelihood of major depressive episodes. In addition, autonomy mediated the associations between BAS sensitivity and prospective hypomanic/manic episodes. These findings suggest that individuals with bipolar spectrum disorders may exhibit a unique profile of BAS-relevant cognitive styles that influence the course of their mood episodes.

Bipolar spectrum-substance use co-occurrence: Behavioral approach system (BAS) sensitivity and impulsiveness as shared personality vulnerabilities.

Bipolar disorders and substance use disorders (SUDs) show high co-occurrence. One explanation for this co-occurrence may be common personality vulnerabilities involved in both. The authors tested whether high behavioral approach system (BAS) sensitivity and impulsiveness are shared personality vulnerabilities in bipolar spectrum disorders and substance use problems and their co-occurrence in a longitudinal study of 132 individuals on the bipolar spectrum and 153 control participants. At Time 1, participants completed the Behavioral Inhibition System/BAS Scales and the Impulsive Nonconformity Scale. Substance use problems were assessed via the Michigan Alcoholism Screening Test and the Drug Abuse Screening Test at 4-month intervals for 1 year. Participants with bipolar disorder had higher rates of lifetime SUDs and substance use problems during the follow-up, relative to control participants. In line with hypotheses, higher BAS sensitivity and impulsiveness predicted bipolar status and increased substance use problems prospectively. BAS total, BAS Fun Seeking, and impulsiveness mediated the association between bipolar spectrum status and prospective substance use problems, with impulsiveness as the most important mediator. High BAS sensitivity and impulsiveness may represent shared personality vulnerabilities for both disorders and may partially account for their co-occurrence.

Whole genome microarray analysis, from neonatal blood cards.

BACKGROUND: Neonatal blood, obtained from a heel stick and stored dry on paper cards, has been the standard for birth defects screening for 50 years. Such dried blood samples are used, primarily, for analysis of small-molecule analytes. More recently, the DNA complement of such dried blood cards has been used for targeted genetic testing, such as for single nucleotide polymorphism in cystic fibrosis. Expansion of such testing to include polygenic traits, and perhaps whole genome scanning, has been discussed as a formal possibility. However, until now the amount of DNA that might be obtained from such dried blood cards has been limiting, due to inefficient DNA recovery technology. RESULTS: A new technology is employed for efficient DNA release from a standard neonatal blood card. Using standard Guthrie cards, stored an average of ten years post-collection, about 1/40th of the air-dried neonatal blood specimen (two 3 mm punches) was processed to obtain DNA that was sufficient in mass and quality for direct use in microarray-based whole genome scanning. Using that same DNA release technology, it is also shown that approximately 1/250th of the original purified DNA (about 1 ng) could be subjected to whole genome amplification, thus yielding an additional microgram of amplified DNA product. That amplified DNA product was then used in microarray analysis and yielded statistical concordance of 99% or greater to the primary, unamplified DNA sample. CONCLUSION: Together, these data suggest that DNA obtained from less than 10% of a standard neonatal blood specimen, stored dry for several years on a Guthrie card, can support a program of genome-wide neonatal genetic testing.

Behavioral Approach System and Behavioral Inhibition System sensitivities and bipolar spectrum disorders: prospective prediction of bipolar mood episodes.

OBJECTIVES: Research has found that bipolar spectrum disorders are associated with Behavioral Approach System (BAS) hypersensitivity and both unipolar and bipolar depression are associated with high Behavioral Inhibition System (BIS) sensitivity, but prospective studies of these relationships are lacking. We tested whether BAS and BIS sensitivities prospectively predicted the time to new onsets of major depressive and hypomanic and manic episodes in bipolar spectrum individuals. METHODS: We followed 136 bipolar II or cyclothymic and 157 demographically matched normal control individuals prospectively for an average of 33 months. Participants completed the BIS/BAS scales and symptom measures at Time 1 and semi-structured diagnostic interviews every four months of follow-up. RESULTS: The bipolar spectrum group exhibited higher Time 1 BAS, but not BIS, scores than the normal controls, controlling for Time 1 symptoms. Among bipolar spectrum participants, high BAS sensitivity prospectively predicted a shorter time to onset of hypomanic and manic episodes, whereas high BIS sensitivity predicted less survival time to major depressive episodes, controlling for initial symptoms. CONCLUSIONS: Consistent with the BAS hypersensitivity model of bipolar disorder, a highly responsive BAS provides vulnerability to onsets of (hypo)manic episodes. In addition, a highly sensitive BIS increases risk for major depressive episodes.

The role of cluster B and C personality disturbance in the course of depression: a prospective study.

The association between personality disturbance and depression has been noted consistently. Prospective tests of personality's impact on the course of depression, however, are lacking. In a sample of 159 undergraduates who experienced at least one prospective depressive episode, dimensional scores for clusters B and C personality disturbance were examined as prospective predictors of four indicators of the course of depression: severity, episode duration, symptomatic chronicity and number of episodes. Cluster C personality disturbance, characterized by anxious and fearful features, predicted depression chronicity. Cluster B, characterized by dramatic, emotional and/or erratic features, predicted severity and duration of depression. The findings are discussed in terms of the possible mechanisms underlying the effects of clusters B and C, as well as implications for future research.

A goal-striving life event and the onset of hypomanic and depressive episodes and symptoms: perspective from the behavioral approach system (BAS) dysregulation theory.

On the basis of the behavioral approach system (BAS) dysregulation theory of bipolar disorder, this study examined the relation between occurrence of a BAS activation-relevant life event--goal striving--and onset of hypomanic and depressive episodes and symptoms. In particular, the authors examined the relation between preparing for and completing final exams (a goal-striving event) and onset of bipolar spectrum episodes and symptoms in college students with bipolar II disorder or cyclothymia (i.e., "soft" bipolar spectrum conditions). One hundred fifty-nine individuals with either a bipolar spectrum disorder (n=68) or no major affective psychopathology (controls; n=91) were further classified on the basis of whether they were college students (i.e., completed final exams). Consistent with the BAS dysregulation theory, preparing for and completing final exams was associated with an increase in hypomanic but not depressive episodes and symptoms in individuals with a soft bipolar spectrum diagnosis. Furthermore, self-reported BAS sensitivity moderated the presence of certain hypomanic symptoms during final exams.

The course of depression in individuals at high and low cognitive risk for depression: a prospective study.

BACKGROUND: Negative cognitive styles have been shown to prospectively predict depression onset and recurrence. Research has also begun to suggest that cognitive styles may be associated with the course of depression as well. This study examined whether cognitive risk for depression onset also predicts the course of depression in a prospective design. METHODS: One hundred fifty-nine initially nondepressed participants from the Temple-Wisconsin Cognitive Vulnerability to Depression Project who experienced a depressive episode while in the study were followed prospectively for 2.5 years. Four indicators of the course of depression were assessed from diagnostic interviews and questionnaires administered every 6 weeks: the number, severity and duration of episodes and the chronicity of the depression experienced. RESULTS: Cognitive high-risk participants experienced more episodes of depression, more severe episodes, and more chronic courses than low-risk participants. There were no risk group differences observed for the duration of episodes. LIMITATIONS: This study's sample was chosen to include individuals with high and low cognitive vulnerability to depression, potentially limiting the generalizability of the findings to other individuals. Also, the study included some participants with a past history of depression. CONCLUSIONS: Negative cognitive styles predict a worse course of depression as well as rendering an individual prone to depression onset. This highlights that the cognitive factors impacting depression's course overlap, at least partly, with those that initiate depression. Thus, knowledge of a depressed individual's cognitive styles could aid in prognosis and treatment planning.

Prospective incidence of first onsets and recurrences of depression in individuals at high and low cognitive risk for depression.

Do negative cognitive styles provide similar vulnerability to first onsets versus recurrences of depressive disorders, and are these associations specific to depression? The authors followed for 2.5 years prospectively college freshmen (N = 347) with no initial psychiatric disorders at high-risk (HR) versus low-risk (LR) for depression on the basis of their cognitive styles. HR participants had odds of major, minor, and hopelessness depression that were 3.5-6.8 times greater than the odds for LR individuals. Negative cognitive styles were similarly predictive of first onsets and recurrences of major depression and hopelessness depression but predicted first onsets of minor depression more strongly than recurrences. The risk groups did not differ in incidence of anxiety disorders not comorbid with depression or other disorders, but HR participants were more likely to have an onset of anxiety comorbid with depression.

Negative cognitive styles, dysfunctional attitudes, and the remitted depression paradigm: a search for the elusive cognitive vulnerability to depression factor among remitted depressives.

Results from studies using a behavioral high-risk design and approximations to it generally have corroborated the cognitive vulnerability hypothesis of depression, whereas results from remitted depression studies typically have not. Suspecting that design features of previously conducted remitted designs likely precluded them from detecting maladaptive cognitive patterns, the authors conducted a study featuring the remitted design that has been successful in studies of a biological vulnerability for depression. Participants' current depressive symptoms, negative cognitive styles (hopelessness theory), dysfunctional attitudes (Beck's theory), and lifetime prevalence of clinically significant depression were assessed. Participants who had remitted from an episode of clinically significant depression had more negative cognitive styles, but not greater levels of dysfunctional attitudes, than did never depressed individuals.

Proneness to hypomania/mania symptoms or depression symptoms and asymmetrical frontal cortical responses to an anger-evoking event.

The behavioral approach system (BAS) reflects the propensity to respond to signals of reward, including stimuli associated with safety and goal-oriented attack (e.g., anger). Hypomania/mania has been posited to involve increased BAS activity. In contrast, depression has been posited to involve decreased BAS activity. Building on past research, which suggests that increased left frontal cortical activity is a neurophysiological index of BAS activity, the present research tested the hypotheses that proneness toward hypomania/mania symptoms would be related to increased relative left frontal activity and that proneness toward depression symptoms would be related to decreased relative left frontal activity in response to an anger-evoking event. Results from 67 individuals who had completed a measure of proneness toward these affective symptoms and were exposed to an anger-evoking event supported the hypotheses.