Characterizing double-back stutter in low to multi-copy number regimes in forensically relevant STR loci

Modern DNA analysis is possible due to the discovery of repeating microsatellite regions in DNA and successful implementation of the polymerase chain reaction (PCR) in laboratories. PCR amplification chemistries that contain short tandem repeat (STR) loci are sensitive. As a result, the discrimination power within human identification sciences has increased in recent years. Despite these advances, cellular admixtures are commonly collected, and the resultant “DNA mixture profile” is difficult to interpret as it is often encumbered by low-signals and allele drop-out. Regularly detected PCR artifacts can further complicate interpretation. One commonly encountered artifact is stutter, the result of strand slippage during PCR. Stutter can be of two types: forward and reverse. Reverse stutter (or back stutter) is the most prevalent and is one repeat unit shorter (n - 1) than the template strand. In contrast, forward stutter is one repeat unit longer (n + 1). If a reverse stutter amplicon is produced there is the distinct possibility that a stutter product of stutter may occur. This artifact is usually referred to as double-back stutter (DBS) or n - 2 stutter. Recently there has been renewed interest in examining signal approaching baseline levels. As the sensitivity of the process improves, so does the probability of detecting DBS. Therefore, studies that examine the peak height distributions, rarity, stutter signal-to-noise distances and the general impact of DBS on the signal are warranted. Models simulating PCR, and the entire forensic DNA process, have been created by this laboratory. The work presented herein builds upon a preexisting model; specifically, the dynamic model was extended such that DNA profiles consisting of 21 autosomal STRs, consistent with the GlobalFilerTM multiplex, are simulated. Furthermore, this expansion incorporated a three-type Galton-Watson branching process allowing DBS to be added to the simulated electropherogram (EPG). The in silico model was used to simulate the amplification of a 1:43 and 1:73 mixture at a total DNA concentration of 0.3 and 0.5 ng, respectively. We chose these extreme mixture ratios because the signal from these minor contributors would be most susceptible to DBS effects from the major contributor. A total of 1200 alleles from each contributor were simulated at each target, and effects of DBS on the signal from the minor contributor were characterized. At 0.3 and 0.5 ng both the noise and stutter signal histograms are right-skewed and a Kolmogorov-Smirnov (KS) test indicates that the noise and DBS were significantly different (p-value < 4x10-6). The average peak height of DBS for all loci in both scenarios were less than 50 RFU (Relative Fluorescence Units), and the DBS ratios ranged from 0.29 to 2.15% of the main allele, with the median ratios less than 0.5%. A per locus analytical threshold (AT) was calculated for both the 0.3 and 0.5 ng targets using two k-values: 3 and 4. The k-value is chosen based on the Type I risk assessment, wherein increasing the k-value increases AT. The percentage of DBS peaks greater than AT when k = 3 for the mixtures amplified at 0.3 and 0.5 ng ranged from 0 to 7.08% and 0 to 10.50%, respectively. Interestingly, when k = 4 the percentage of DBS peaks greater than AT for 0.3 and 0.5 ng reduced to 0 to 1.08% and 0 to 0.17%, respectively. This suggests that modeling DBS in continuous systems may not be necessary if the laboratory continues to rely on a system that requires an AT of sufficient strength. However, with the advent of Bayesian or machine learning-based approaches to analyzing EPGs, thus removing AT in its entirety, a complete understanding of the prevalence of DBS is necessary. This work shows that DBS from an extreme major using our laboratory protocols is not likely to be in the same signal regime as the signal from alleles; however, it does show that signal from DBS is significantly different from noise. Therefore, the software expert pair should be carefully considered during the validation stage and laboratories should consider DBS during interpretation, especially if enhanced post-PCR parameters are implemented into the forensic laboratory process

The Paris Collection: Reflections on an Interdisciplinary Performance Project

Between September 2013 and January 2014, students and staff of the Technological University Dublin Conservatory of Music and Drama and DIT Dublin School of Creative Arts collaborated on the creation of an opera production which was presented at the National Concert Hall on Jan 23rd. The project was implemented within the modular framework from both colleges. The collaboration was both multi and inter-disciplinary and the students engaged at a professional level, working under staff supervision and in tandem with personnel and resources outside the college environment. The performers were required to audition for their roles and the teams of design students had to prepare and present a \u27pitch\u27 for their concept to be selected. These same design students also took responsibility for the set construction, while the entire cohort had to respond to the logistical and artistic challenges of taking the production into a major venue on the day of performance and being ready to begin in a significantly limited period of time. The delivery of a public performance in a high profile venue meant that the project culminated in full exposure of public scrutiny, and consequently demanded the skills, intelligence, energy and commitment of all involved. This presentation will focus on the collaboration as a relevant case-study, exemplifying a modular-based inter-disciplinary project within the College of Arts and Tourism. It will provide an evaluation of the insights, strengths and benefits gained by all who participated. It will also provide an honest discourse on the requirements, resources and challenges such an undertaking demands, particularly with reference to establishing professional demands within an educational context. With reference to future implementation of other synergies, the presentation will provide recommendations as to how these can be realistically and sufficiently facilitated through creative and flexible programme development

Working with Science Teachers to Transform the Opportunity Landscape for Regional and Rural Youth: A Qualitative Evaluation of the Science in Schools Program

This article reports on a qualitative evaluation of the Science in Schools program; a suite of science based activities delivered by staff of a regional university campus and designed to provide professional development for science teachers working in non-metropolitan schools in a socioeconomically disadvantaged region of Australia. The research identified a range of issues including: the influence of socioeconomic disadvantage and rurality on teachers’ professional learning needs, and the importance of subject specific discourse communities and content knowledge for new and out-of-field teachers. Implications for the design and implementation of school-university partnerships are discussed

Intangible Qualities of Rare Books: Toward a Decision-Making Framework for Preservation Management in Rare Book Collections, Based Upon the Concept of the Book as Object

For rare book collections, a considerable challenge is involved in evaluating collection materials in terms of their inherent value, which includes the textual and intangible information the materials provide for the collection's users. Preservation management in rare book collections is a complex and costly process. As digitization and other technological advances in surrogate technology have provided new forms representation, new dilemmas in weighing the rare book's inherently valuable characteristics against the possibly lesser financial costs of surrogates have arisen. No model has been in wide use to guide preservation management decisions. An initial iteration of such a model is developed, based on a Delphi-like iterative questioning of a group of experts in the field of rare books. The results are used to synthesize a preservation management framework for rare book collections, and a small-scale test of the framework has been completed through two independent analyses of five rare books in a functioning collection. Utilizing a standardized template for making preservation decisions offers a variety of benefits. Preservation decisions may include prioritizing action upon the authentic objects, or developing and maintaining surrogates in lieu of retaining costly original collection materials. The framework constructed in this study provides a method for reducing the subjectivity of preservation decision-making and facilitating the development of a standard of practice for preservation management within rare book collections

A complicated relationship: an introduction to the correspondence between Percival Lowell and Giovanni Virginio Schiaparelli

From 1896 until Schiaparelli’s death in 1910, Percival Lowell and Giovanni Schiaparelli exchanged letters, telegrams, packages, and postcards apprising them of each other’s work and commenting on the other’s latest theories. Lowell, who greatly admired Schiaparelli’s astronomical career, wrote in search of approval for his oft-criticized theories about the perceived presence of canals and vegetation on Mars. Schiaparelli, who courted many theories about Mars but never truly invested himself entirely in one, gently guided Lowell to see other possibilities or recognize faults in his work, criticism that Lowell took with difficulty from others but accepted from Schiaparelli. The correspondence, translated from the original French here and made newly available to the public, shows a complex but respectful, warm, and gentlemanly relationship between two giants of Martian astronomy

Evaluation of a program designed to build science teaching capacity in rural Australia

This article reports on a qualitative evaluation of the Science in Schools program; a suite of science based activities delivered by staff of a regional university campus and designed to provide professional development for science teachers working in non-metropolitan schools in a socioeconomically disadvantaged region of Australia. The research identified a range of issues including: the influence of socioeconomic disadvantage and rurality on teachers' professional learning needs, and the importance of subject specific discourse communities and content knowledge for new and out-of-field teachers. Implications for the design and implementation of school-university partnerships are discussed

Vaccine Uptake in the Era of COVID-19: Associations between Willingness to Receive the Influenza and COVID-19 Vaccines

Background/purpose. Despite efforts to increase flu vaccine uptake, the rate of vaccination remains below target. Additionally, many Virginians remain unvaccinated or have not maintained a booster schedule in line with recommendations. The current study investigated willingness to receive the flu and COVID-19 vaccines during the COVID-19 pandemic, associations between the likelihood of receiving the two vaccines, and differences in uptake by racial group and insurance status. Methods. Participants were adult patients seen for appointments at an ambulatory clinic in Southeast, VA. Participants completed a questionnaire assessing vaccination history and future vaccination intentions. Results. Results revealed significant associations between having received the flu vaccine previously and receiving the current season vaccine as well as receiving the flu vaccine and intentions to receive the COVID-19 vaccine. No significant differences in flu vaccination were found between Black and White adults. Insured participants reported greater intentions to receive the flu vaccine compared to uninsured participants. The primary reason reported for vaccine hesitancy were safety concerns. Discussion. Study participants report a much higher flu vaccination rate compared to the national average. Participants who endorse the efficacy and safety of vaccines are likely to have greater intentions for future vaccine uptake. Conclusions. As new flu and COVID-19 variants emerge, the need to maintain protection via annual boosters will remain. Addressing the reasons behind vaccine hesitancy is essential to increasing vaccine coverage. The study adds to the growing literature on attitudes towards vaccination, the need for continued endorsement of vaccinations, and highlights opportunities to promote vaccination uptake for uninsured patients

Working with science teachers to transform the opportunity landscape for regional and rural youth: a qualitative evaluation of the science in schools program

This article reports on a qualitative evaluation of the Science in Schools program; a suite of science based activities delivered by staff of a regional university campus and designed to provide professional development for science teachers working in non-metropolitan schools in a socioeconomically disadvantaged region of Australia. The research identified a range of issues including: the influence of socioeconomic disadvantage and rurality on teachers’ professional learning needs, and the importance of subject specific discourse communities and content knowledge for new and out-of-field teachers. Implications for the design and implementation of school-university partnerships are discussed

The endoribonuclease YbeY is linked to proper cellular morphology and virulence in 2 Brucella abortus

The endoribonuclease YbeY is one of the most well conserved proteins across the kingdoms of life. In the present study, we demonstrate that YbeY in Brucella abortus is linked to a variety of important activities, including proper cellular morphology, mRNA transcript levels, and virulence. Deletion of ybeY in B. abortus led to a small colony phenotype when the bacteria were grown on agar medium, as well as significant aberrations in the morphology of the bacterial cell as evidenced by electron microscopy. Additionally, compared to the parental strain, the ΔybeY strain was significantly attenuated in both macrophage and mouse models of infection. The ΔybeY strain also showed increased sensitivities to several in vitro applied stressors, including bile acid, hydrogen peroxide, SDS, and paraquat. Transcriptomic analysis revealed that a multitude of mRNA transcripts are dysregulated in the ΔybeY strain, and many of the identified mRNAs encode proteins involved in metabolism, nutrient transport, transcriptional regulation, and flagellum synthesis. We subsequently constructed gene deletion strains of the most highly dysregulated systems, and several of the YbeY-linked gene deletion strains exhibited defects in the ability of the bacteria to survive and replicate in primary murine macrophages. Altogether, these data establish a clear role for YbeY in the biology and virulence of Brucella, and moreover, this work further illuminates the highly varied roles of this widely conserved endoribonuclease in bacteria. Importance Brucella spp. are highly efficient bacterial pathogens of animals and humans, causing significant morbidity and economic loss worldwide, and relapse of disease often occurs following antibiotic treatment of human brucellosis. As such, novel therapeutic strategies to combat Brucella infections are needed. Ribonucleases in the brucellae are understudied, and these enzymes represent elements that may be potential targets for future treatment approaches. The present work demonstrates the importance of the endoribonuclease YbeY for cellular morphology, efficient control of mRNA levels, and virulence in B. abortus. Overall, this study advances our understanding of the critical roles of YbeY in the pathogenesis of the intracellular brucellae and expands our understanding of this highly conserved ribonuclease.National Institute of General Medical Sciences (U.S.) (Grant GM31030