A Simple Screen to Identify Promoters Conferring High Levels of Phenotypic Noise

Genetically identical populations of unicellular organisms often show marked variation in some phenotypic traits. To investigate the molecular causes and possible biological functions of this phenotypic noise, it would be useful to have a method to identify genes whose expression varies stochastically on a certain time scale. Here, we developed such a method and used it for identifying genes with high levels of phenotypic noise in Salmonella enterica ssp. I serovar Typhimurium (S. Typhimurium). We created a genomic plasmid library fused to a green fluorescent protein (GFP) reporter and subjected replicate populations harboring this library to fluctuating selection for GFP expression using fluorescent-activated cell sorting (FACS). After seven rounds of fluctuating selection, the populations were strongly enriched for promoters that showed a high amount of noise in gene expression. Our results indicate that the activity of some promoters of S. Typhimurium varies on such a short time scale that these promoters can absorb rapid fluctuations in the direction of selection, as imposed during our experiment. The genomic fragments that conferred the highest levels of phenotypic variation were promoters controlling the synthesis of flagella, which are associated with virulence and host–pathogen interactions. This confirms earlier reports that phenotypic noise may play a role in pathogenesis and indicates that these promoters have among the highest levels of noise in the S. Typhimurium genome. This approach can be applied to many other bacterial and eukaryotic systems as a simple method for identifying genes with noisy expression

Complete Genome Sequences of Cluster A Mycobacteriophages BobSwaget, Fred313, KADY, Lokk, MyraDee, Stagni, and StepMih

Seven mycobacteriophages from distinct geographical locations were isolated, using Mycobacterium smegmatis mc2155 as the host, and then purified and sequenced. All of the genomes are related to cluster A mycobacteriophages, BobSwaget and Lokk in subcluster A2; Fred313, KADY, Stagni, and StepMih in subcluster A3; and MyraDee in subcluster A18, the first phage to be assigned to that subcluster

Tracking the international spread of SARS-CoV-2 lineages B.1.1.7 and B.1.351/501Y-V2

Publisher Copyright: © 2021 O'Toole Á et al.Late in 2020, two genetically-distinct clusters of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with mutations of biological concern were reported, one in the United Kingdom and one in South Africa. Using a combination of data from routine surveillance, genomic sequencing and international travel we track the international dispersal of lineages B.1.1.7 and B.1.351 (variant 501Y-V2). We account for potential biases in genomic surveillance efforts by including passenger volumes from location of where the lineage was first reported, London and South Africa respectively. Using the software tool grinch (global report investigating novel coronavirus haplotypes), we track the international spread of lineages of concern with automated daily reports, Further, we have built a custom tracking website (cov-lineages.org/global_report.html) which hosts this daily report and will continue to include novel SARS-CoV-2 lineages of concern as they are detected.Peer reviewe

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Additional file 10: Table S2. of Combining Shigella Tn-seq data with gold-standard E. coli gene deletion data suggests rare transitions between essential and non-essential gene functionality

List of chromosomal SNPs and indels observed in the Shigella strain used here that differ from the GenBank sequence NC004741. (TXT 4 kb

Additional file 12: Figure S8. of Combining Shigella Tn-seq data with gold-standard E. coli gene deletion data suggests rare transitions between essential and non-essential gene functionality

Inferred fragment lengths of perfectly mapped reads across several genomic regions suggest IS-mediated deletions. For each plot, the inferred fragment lengths are arranged by increasing length (ranked on the y-axis). Thus, very long fragments are present at the top of the y-axis. Most fragments have lengths between 100 bp and 400 bp; a small number have lengths over 1000 bp or more. It is very likely that these are not the true insert sizes, but appear that way because of large scale deletions in our Shigella clone compared to the clone present in the NCBI genome database; see Methods for more details. (A) A region of the chromosome in which a complicated series of rearrangements has occurred, leading to paired end reads perfectly matching to different locations in this region. 45 mapped read pairs span more than 1.5 Kbp, a size that is not concordant with the majority of insert sizes. (B) A genomic region where an approximately 10Kbp deletion occurred, removing a region containing the yeaKLMNOP operon. 92 mapped read pairs span more than 8.5Kbp. This region is flanked by two IS elements. (C) A region where an approximately 4Kbp deletion occurred, removing two genes with no E. coli K12 orthologues. 68 mapped read pairs span more than 4 Kbp, and again this region is flanked by two IS elements. (D) A genomic region where an approximately 2Kbp deletion occurred, removing yhdW. 244 mapped read pairs span more than 2Kbp, and the region is flanked by two IS elements. (E) A deletion in the region of the chromosome containing S4145 (yiaN). 232 mapped read pairs spanned more than 1.8 Kbp, and this region is also flanked by two IS elements. (F) A region of the chromosome containing the rfb operon. Most of the genes within this operon are uninterrupted by transposons. However, we find no evidence that this is due to a deletion of this region in our Shigella clone, as we find no reads mapping across the region; a small number of reads map within the region; and the closest IS elements are 15 Kb upstream of rfbJ and 20 Kb downstream of rfbA. The genes in this operon have no orthologues in E. coli K12. (PDF 219 kb

Feasibility, Perceived Impact, and Acceptability of a Socially Assistive Robot to Support Emotion Regulation With Highly Anxious University Students: Mixed Methods Open Trial

BackgroundMental health difficulties among university students have been rising rapidly over the last decade, and the demand for university mental health services commonly far exceeds available resources. Digital interventions are seen as one potential solution to these challenges. However, as in other mental health contexts, digital programs often face low engagement and uptake, and the field lacks usable, engaging, evidence-supported mental health interventions that may be used flexibly when students need them most. ObjectiveThe aim of this study is to investigate the feasibility and acceptability of a new, in situ intervention tool (Purrble) among university students experiencing anxiety. As an intervention, Purrble was designed to provide in situ support for emotion regulation (ER)—a well-known transdiagnostic construct—directly in the moments when individuals are facing emotionally challenging situations. A secondary aim is to consider the perceived impact of Purrble on youth mental health, as reported by students over a 7-week deployment. MethodsA mixed methods open trial was conducted with 78 under- and postgraduate students at Oxford University. Participants were recruited based on moderate to high levels of anxiety measured by Generalized Anxiety Disorder-7 at baseline (mean 16.09, SD 3.03). All participants had access to Purrble for 7 weeks during the spring term with data on their perceived anxiety, emotion dysregulation, ER self-efficacy, and engagement with the intervention collected at baseline (pre), week 4 (mid), and week 8 (postintervention). Qualitative responses were also collected at the mid- and postintervention points. ResultsThe findings demonstrated a sustained engagement with Purrble over the 7-week period, with the acceptability further supported by the qualitative data indicating that students accepted Purrble and that Purrble was well-integrated into their daily routines. Exploratory quantitative data analysis indicated that Purrble was associated with reductions in student anxiety (dz=0.96, 95% CI 0.62-1.29) and emotion dysregulation (dz=0.69, 95% CI 0.38-0.99), and with an increase in ER self-efficacy (dz=–0.56, 95% CI –0.86 to –0.26). ConclusionsThis is the first trial of a simple physical intervention that aims to provide ongoing ER support to university students. Both quantitative and qualitative data suggest that Purrble is an acceptable and feasible intervention among students, the engagement with which can be sustained at a stable level across a 7-week period while retaining a perceived benefit for those who use it (n=32, 61% of our sample). The consistency of use is particularly promising given that there was no clinician engagement or further support provided beyond Purrble being delivered to the students. These results show promise for an innovative intervention model, which could be complementary to the existing interventions