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Swine harbors a genetically diverse population of swine influenza A viruses (IAV-S), with demonstrated potential to transmit to the human population, causing outbreaks and pandemics. Here, we describe the development of a one-step, triplex real-time reverse transcription-polymerase chain reaction (rRT-PCR) assay that detects and distinguishes the majority of the antigenically distinct influenza A virus hemagglutinin (HA) clades currently circulating in North American swine, including the IAV-S H1 1A.1 (α), 1A.2 (β), 1A.3 (γ), 1B.2.2 (δ1) and 1B.2.1 (δ2) clades, and the IAV-S H3 2010.1 clade. We performed an in-field test at an exhibition swine show using in-field viral concentration and RNA extraction methodologies and a portable real-time PCR instrument, and rapidly identified three distinct IAV-S clades circulating within the N.A. swine population. Portable sequencing is used to further confirm the results of the in-field test of the swine triplex assay. The IAV-S triplex rRT-PCR assay can be easily transported and used in-field to characterize circulating IAV-S clades in North America, allowing for surveillance and early detection of North American IAV-S with human outbreak and pandemic potential.

One of the key events in autophagy is the formation of a double-membrane phagophore, and many regulatory mechanisms underpinning this remain under investigation. WIPI2b is among the first proteins to be recruited to the phagophore and is essential for stimulating autophagy flux by recruiting the ATG12-ATG5-ATG16L1 complex, driving LC3 and GABARAP lipidation. Here, we set out to investigate how WIPI2b function is regulated by phosphorylation. We studied two phosphorylation sites on WIPI2b, S68 and S284. Phosphorylation at these sites plays distinct roles, regulating WIPI2b's association with ATG16L1 and the phagophore, respectively. We confirm WIPI2b is a novel ULK1 substrate, validated by the detection of endogenous phosphorylation at S284. Notably, S284 is situated within an 18-amino acid stretch, which, when in contact with liposomes, forms an amphipathic helix. Phosphorylation at S284 disrupts the formation of the amphipathic helix, hindering the association of WIPI2b with membranes and autophagosome formation. Understanding these intricacies in the regulatory mechanisms governing WIPI2b's association with its interacting partners and membranes, holds the potential to shed light on these complex processes, integral to phagophore biogenesis., (© 2024. The Author(s).)

Epilepsy is a neurological disease characterised by recurrent seizures with complex aetiology. Temporal lobe epilepsy, the most common form in adults, can be acquired following brain insults including trauma, stroke, infection or sustained status epilepticus. The mechanisms that give rise to the formation and maintenance of hyperexcitable networks following acquired insults remain unknown, yet an extensive body of literature points towards persistent gene and epigenomic dysregulation as a potential mediator of this dysfunction. While much is known about the function of specific classes of epigenetic regulators (writers and erasers) in epilepsy, much less is known about the enzymes, which read the epigenome and modulate gene expression accordingly. Here, we explore the potential role for the epigenetic reader bromodomain and extra-terminal domain (BET) proteins in epilepsy. Using the intra-amygdala kainic acid model of temporal lobe epilepsy, we initially identified widespread dysregulation of important epigenetic regulators including EZH2 and REST as well as altered BRD4 expression in chronically epileptic mice. BRD4 activity was also notably affected by epilepsy-provoking insults as seen by elevated binding to and transcriptional regulation of the immediate early gene Fos. Despite influencing early aspects of epileptogenesis, blocking BET protein activity with JQ1 had no overt effects on epilepsy development in mice but did alter glial reactivity and influence gene expression patterns, promoting various neurotransmitter signalling mechanisms and inflammatory pathways in the hippocampus. Together, these results confirm that epigenetic reader activity is affected by epilepsy-provoking brain insults and that BET activity may exert cell-specific actions on inflammation in epilepsy., (© 2024 The Author(s). European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Psychologists in academic medicine face pressure to juggle multiple roles, and research is often limited by a lack of available resources and funding. In other academic settings, student-led psychology research teams that utilize a tiered mentorship approach are able to produce advances in meaningful research while supporting the development of future professionals in the field. This article identifies the barriers of implementing a tiered mentorship model into an academic medicine setting and reviews a case study of how the model can be effectively adapted and evaluated to promote a self-sustaining, student-led psychology research team., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Care partners of people with serious illness experience significant challenges and unmet needs during the patient's treatment period and after their death. Learning from others with shared experiences can be valuable, but opportunities are not consistently available., Objective: This study aims to design and prototype a regional, facilitated, and web-based peer support network to help active and bereaved care partners of persons with serious illness be better prepared to cope with the surprises that arise during serious illness and in bereavement., Methods: An 18-member co-design team included active care partners and those in bereavement, people who had experienced serious illness, regional health care and support partners, and clinicians. It was guided by facilitators and peer network subject-matter experts. We conducted design exercises to identify the functions and specifications of a peer support network. Co-design members independently prioritized network specifications, which were incorporated into an early iteration of the web-based network., Results: The team prioritized two functions: (1) connecting care partners to information and (2) facilitating emotional support. The design process generated 24 potential network specifications to support these functions. The highest priorities included providing a supportive and respectful community; connecting people to trusted resources; reducing barriers to asking for help; and providing frequently asked questions and responses. The network platform had to be simple and intuitive, provide technical support for users, protect member privacy, provide publicly available information and a private discussion forum, and be easily accessible. It was feasible to enroll members in the ConnectShareCare web-based network over a 3-month period., Conclusions: A co-design process supported the identification of critical features of a peer support network for care partners of people with serious illnesses in a rural setting, as well as initial testing and use. Further testing is underway to assess the long-term viability and impact of the network., (©Elizabeth A O’Donnell, Aricca D Van Citters, Inas S Khayal, Matthew M Wilson, David Gustafson, Amber E Barnato, Andrea C Buccellato, Colleen Young, Megan M Holthoff, Eugene Korsunskiy, Stephanie C Tomlin, Amelia M Cullinan, Alexandra C Steinbaugh, Jennifer J Hinson, Kristen R Johnson, Andrew Williams, Ruth M Thomson, Janet M Haines, Anne B Holmes, Ann D Bradley, Eugene C Nelson, Kathryn B Kirkland. Originally published in JMIR Human Factors (https://humanfactors.jmir.org), 08.05.2024.)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has evolved into numerous lineages with unique spike mutations and caused multiple epidemics domestically and globally. Although COVID-19 vaccines are available, new variants with the capacity for immune evasion continue to emerge. To understand and characterize the evolution of circulating SARS-CoV-2 variants in the U.S., the Centers for Disease Control and Prevention (CDC) initiated the National SARS-CoV-2 Strain Surveillance (NS3) program and has received thousands of SARS-CoV-2 clinical specimens from across the nation as part of a genotype to phenotype characterization process. Focus reduction neutralization with various antisera was used to antigenically characterize 143 SARS-CoV-2 Delta, Mu and Omicron subvariants from selected clinical specimens received between May 2021 and February 2023, representing a total of 59 unique spike protein sequences. BA.4/5 subvariants BU.1, BQ.1.1, CR.1.1, CQ.2 and BA.4/5 + D420N + K444T; BA.2.75 subvariants BM.4.1.1, BA.2.75.2, CV.1; and recombinant Omicron variants XBF, XBB.1, XBB.1.5 showed the greatest escape from neutralizing antibodies when analyzed against post third-dose original monovalent vaccinee sera. Post fourth-dose bivalent vaccinee sera provided better protection against those subvariants, but substantial reductions in neutralization titers were still observed, especially among BA.4/5 subvariants with both an N-terminal domain (NTD) deletion and receptor binding domain (RBD) substitutions K444M + N460K and recombinant Omicron variants. This analysis demonstrated a framework for long-term systematic genotype to antigenic characterization of circulating and emerging SARS-CoV-2 variants in the U.S., which is critical to assessing their potential impact on the effectiveness of current vaccines and antigen recommendations for future updates.

Purpose: Information on concerns that young adults (YAs) with cancer face when receiving care outside of specialized treatment centers is needed to increase equitable care to YAs at greater risk of marginalization by the health care system. The current study compared distress and unmet needs at the time of clinic visit between YAs receiving care from three different cancer clinics: (1) a National Cancer Institute-designated center, (2) a community-based clinic, and (3) a county hospital outpatient clinic., Methods: The Adolescent and Young Adult Psycho-Oncology Screening Tool (AYA-POST) was administered to measure distress and cancer-related concerns of YAs in active treatment. A one-way analysis of variance (ANOVA) compared distress scores by treatment site. A Fisher's exact test compared the number of participants endorsing each item on the Needs Assessment Checklist from each site. A simple linear regression determined the association between distress and number of items endorsed on the Needs Assessment Checklist., Results: Ninety-seven participants completed the AYA-POST, endorsing, on average, 11 concerns. Fisher's exact test showed significant differences between sites in the proportion of participants endorsing eight items: boredom ( P < .001), eating/appetite ( P < .001), nausea/vomiting ( P < .001), financial concern ( P = .002), hopelessness/helplessness ( P = .03), confidentiality ( P = .04), sibling concern ( P = .04), and insurance ( P = .05). The simple linear regression model was significant (F(1, 94) = 39.772, P < .001, R 2 = 0.297), indicating the number of unmet needs accounted for almost 30% of the variance in distress. The one-way ANOVA was not significant (F(2, 93) = 1.34, P = .267)., Conclusion: Social determinants of health can influence the number and type of unmet needs experienced, affecting distress and other outcomes and underscoring the importance of timely, effective, age-appropriate screening and intervention for distress and unmet needs in YAs with cancer.

Competing Interests: Competing interests The authors declare no competing or financial interests.

Importance: The COVID-19 pandemic was accompanied by an unprecedented surveillance effort. The resulting data were and will continue to be critical for surveillance and control of SARS-CoV-2. However, some genomic surveillance methods experienced challenges as the virus evolved, resulting in incomplete and poor quality data. Complete and quality coverage, especially of the S-gene, is important for supporting the selection of vaccine candidates. As such, we developed a robust method to target the S-gene for amplification and sequencing. By focusing on the S-gene and imposing strict coverage and quality metrics, we hope to increase the quality of surveillance data for this continually evolving gene. Our technique is currently being deployed globally to partner laboratories, and public health representatives from 79 countries have received hands-on training and support. Expanding access to quality surveillance methods will undoubtedly lead to earlier detection of novel variants and better inform vaccine strain selection., Competing Interests: The authors declare no conflict of interest.

Background: In 2019, the Louisiana Department of Health reported an early influenza B/Victoria (B/VIC) virus outbreak., Method: As it was an atypically large outbreak, we deployed to Louisiana to investigate it using genomics and a triplex real-time RT-PCR assay to detect three antigenically distinct B/VIC lineage variant viruses., Results: The investigation indicated that B/VIC V1A.3 subclade, containing a three amino acid deletion in the hemagglutinin and known to be antigenically distinct to the B/Colorado/06/2017 vaccine virus, was the most prevalent circulating virus within the specimens evaluated (86/88 in real-time RT-PCR)., Conclusion: This work underscores the value of portable platforms for rapid, onsite pathogen characterization., Competing Interests: None declared., (© 2024 The Authors. Influenza and Other Respiratory Viruses published by John Wiley & Sons Ltd. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.)

Patients with thalassemia and sickle cell disease (SCD) require blood transfusions as part of their supportive care. However, one of the most serious side effects of this treatment is the risk of red cell alloimmunization. The goal of this study was to assess the prevalence and Specificity of red cell alloimmunization in Egyptian thalassemia and sickle cell anaemia patients. This study included 200 multi transfused Egyptian patients, one hundred and forty patients with transfusion dependent thalassaemia and sixty patients with sickle cell anaemia, who were attending the Paediatric Children Hospital-Cairo University at the period from March 2019 to October 2019. Alloantibody identification was made by Diamed- ID microtyping system. In the studied groups both thalassemia and sickle patients, the prevalence of alloimmunization was 22/200 (11%) patients. The two most often alloantibodies were, antibodies against Kell antigen (37%) and against E antigen (30%). The prevalence of alloimmunization was more in females in comparison to males, but it did not reach statistical significance and patients with thalassemia major had higher alloimmunization rates than other studied groups but was not statistically significant. In the D negative patients in the research group, alloimmunization demonstrated a statistically significant difference ( p  = 0.01). Age, gender, age of transfusion onset and splenectomy were not contributing factors to the antibody presence in the group of patients being investigated. Before receiving blood transfusions, extended red blood cell phenotyping should be thought of as a crucial procedure for hemoglobinopathies patients who would likely have several transfusions. It is advised that haemoglobinopathies patients in Egypt be checked through phenotyping of RBC units for Kell and all Rh antigens to be phenotyped before starting transfusion in these patients which is also standard of care for these patients presently., Competing Interests: Conflict of interestThe authors declare no conflict of interest., (© The Author(s) 2023.)

The epigenetic repressor BMI1 plays an integral role in promoting the self-renewal and proliferation of many adult stem cell populations, and also tumor types, primarily through silencing the Cdkn2a locus, which encodes the tumor suppressors p16 Ink4a and p19 Arf . However, in cutaneous melanoma, BMI1 drives epithelial-mesenchymal transition programs, and thus metastasis, while having little impact on proliferation or primary tumor growth. This raised questions about the requirement and role for BMI1 in melanocyte stem cell (McSC) biology. Here, we demonstrate that murine melanocyte-specific Bmi1 deletion causes premature hair greying and gradual loss of melanocyte lineage cells. Depilation enhances this hair greying defect, accelerating depletion of McSCs in early hair cycles, suggesting that BMI1 acts to protect McSCs against stress. RNA-seq of McSCs, harvested before onset of detectable phenotypic defects, revealed that Bmi1 deletion derepresses p16 Ink4a and p19 Arf , as observed in many other stem cell contexts. Additionally, BMI1 loss downregulated the glutathione S-transferase enzymes, Gsta1 and Gsta2, which can suppress oxidative stress. Accordingly, treatment with the antioxidant N-acetyl cysteine (NAC) partially rescued melanocyte expansion. Together, our data establish a critical function for BMI1 in McSC maintenance that reflects a partial role for suppression of oxidative stress, and likely transcriptional repression of Cdkn2a., (© 2023 The Authors. Pigment Cell & Melanoma Research published by John Wiley & Sons Ltd.)

The coproduction learning health system (CLHS) model extends the definition of a learning health system to explicitly bring together patients and care partners, health care teams, administrators, and scientists to share the work of optimizing health outcomes, improving care value, and generating new knowledge. The CLHS model highlights a partnership for coproduction that is supported by data that can be used to support individual patient care, quality improvement, and research. We provide a case study that describes the application of this model to transform care within an oncology program at an academic medical center., Competing Interests: No conflicts of interest have been declared relating to the manuscript., (Copyright © 2023 The Authors. Published by Wolters Kluwer Health, Inc.)

Modern bed bugs are resistant to multiple insecticide classes, particularly the pyrethroids. The efficacy of pyrethroid-impregnated mattress liners marketed for bed bug management has been variable. This study evaluated the efficacy of a permethrin-impregnated mattress liner, ActiveGuard, against 24 bed bug strains, consisting of both Cimex hemipterus (F.) and Cimex lectularius L. A 'mat assay', employing an allethrin-impregnated mat, was used to establish the pyrethroid resistance profile of all strains. Three experiments were conducted to evaluate the effect of ActiveGuard exposure on bed bug knockdown: 1) exposing the bed bugs continuously on the liner for up to 24 d, 2) holding the bed bugs on the liner for either 4 or 6 h, and 3) placing a noninsecticide treated fabric above the liner with the bed bugs held continuously on top. Our results indicated that all modern strains (collected within the last 15 years during the current resurgence) were pyrethroid-resistant, although the magnitude of resistance was highly variable between strains. In the continuous exposure study, an incomplete knockdown was recorded for most modern bed bug strains, with some having no knockdown even up to 7 d of constant exposure. In the 4 or 6 h exposure study, the level of knockdown was reduced even further, and very few bed bugs were knocked down in the double fabric study. The results of this study indicate that pyrethroid-impregnated mattress liners are not likely to be effective in the management of most modern bed bug infestations involving either C. hemipterus or C. lectularius., (© The Author(s) 2022. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objectives: A carotid web isdefined as an abnormal shelf-like projection of intimal fibrous tissue into the carotid bulb. Its presence may be an under-recognised source of embolic stroke of undetermined source (ESUS). The aim of this study was to investigate its prevalence in previously reported studies., Materials and Methods: A systematic literature review of Pubmed, EMBASE, and Scopus was conducted up until the 4/12/2021 using variations of the search terms - 'carotid web' and 'ischemic stroke'. Inclusion criteria were studies reporting carotid web prevalence in an ESUS cohort aged >18 years with adequate imaging. Secondary measures such as age, gender, ethnicity, and laterality were recorded. A meta-analysis of proportions was used to summarise the prevalence of webs along with a random-effects model to calculate the relative risk of ipsilateral and contralateral webs in ESUS., Results: The initial search yielded 361 articles, with 11 remaining post the inclusion and exclusion criteria. A meta-analysis of allage groups yielded a total carotid web prevalence among patients with stroke of unknown cause of 9.58 % (95 % CI 5.62 - 15.85). Carotid webs were more often detected in females (76.5 % ± 22.3 %), and in those of African heritage (58 % ± 39 %). In comparison with patients without an ischemic stroke, there was a significant association found for an ipsilateral carotid web (risk ratio of 2.74 (95 % CI: 2.14 - 3.51)) but no association found for contralateral webs (risk ratio of 1.50 (95 % CI: 0.94 - 2.40))., Conclusion: The prevalence of ipsilateral carotid webs associated with ESUS is substantial, and may be more common in females and in individuals of African descent., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Crown Copyright © 2022. Published by Elsevier Ltd. All rights reserved.)

The presence of fecal spots has often been used to verify the existence of a bed bug (Cimex lectularius) infestation. However, no research has been conducted to determine how much fecal material that a bed bug population produces over time. In this study, the number of fecal spots that each nymphal life stage was capable of producing after a bloodmeal was quantified. Adult fecal spots were also quantified to determine if there was consistent production between feedings. During this study, it was discovered that bed bugs produced visible fecal spots and clear spots that were only visible under ultraviolet light. Therefore, three types of fecal spots were quantified: dark feces, light feces, and clear spots. Clear spots were produced in greater amounts (38.5-55.5%) than either dark spots (27.3-40.7%) or light spots (17.3-21.9%). For example, 5th instar bed bugs were thought to produce an average of 21 spots (dark and light) after a single bloodmeal. However, using the ultraviolet light, it was found that the 5th instars actually produced an average of 44.7 spots. Using the total fecal spot data collected during this study, researchers could project contamination potential for an infestation starting with a single gravid female over 30, 60, and 90 d. In addition, the amount of area covered by these spots was projected to be over 12 m2 in just 3 mo, which could greatly reduce the environmental and aesthetic quality of a home., (© The Author(s) 2022. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Globally, the suicide rate is two times higher for males than for females. Previous studies in Newfoundland and Labrador did not examine age-specific rates by sex. The objectives of this study were to determine suicide rates by sex and age group and to compare the demographic and clinical characteristics of males and females who died by suicide., Data and Methods: This observational study analyzed a routinely collected dataset based on all medical examiner-determined suicide deaths among people aged 10 years and older in Newfoundland and Labrador, Canada, between 1997 and 2016. Age-standardized and age-specific suicide rates and rate ratios were calculated based on the number of deaths during the period, and descriptive statistics were used to compare demographic and clinical characteristics between males and females., Results: The age-standardized suicide rate was 4.6 times higher among males than females and was higher for males in most age groups. Rates were highest in the young adult age groups for males (20 to 24 years) and females (35 to 39 years). Males who died by suicide were more likely to be from a rural community and to have died by firearm; females were more likely to die by self-poisoning and to have had a mental illness or substance use history., Interpretation: The results are broadly consistent with previous research, though this is the first study to report age-specific suicide rates among females across the life course in Newfoundland and Labrador. The results underscore the need to design public health and clinical interventions that account for sex differences in suicide risks.

To detect new and changing SARS-CoV-2 variants, we investigated candidate Delta-Omicron recombinant genomes from Centers for Disease Control and Prevention national genomic surveillance. Laboratory and bioinformatic investigations identified and validated 9 genetically related SARS-CoV-2 viruses with a hybrid Delta-Omicron spike protein.

The coronavirus disease 2019 (COVID-19) pandemic has demonstrated a clear need for high-throughput, multiplexed and sensitive assays for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other respiratory viruses and their emerging variants. Here, we present a cost-effective virus and variant detection platform, called microfluidic Combinatorial Arrayed Reactions for Multiplexed Evaluation of Nucleic acids (mCARMEN), which combines CRISPR-based diagnostics and microfluidics with a streamlined workflow for clinical use. We developed the mCARMEN respiratory virus panel to test for up to 21 viruses, including SARS-CoV-2, other coronaviruses and both influenza strains, and demonstrated its diagnostic-grade performance on 525 patient specimens in an academic setting and 166 specimens in a clinical setting. We further developed an mCARMEN panel to enable the identification of 6 SARS-CoV-2 variant lineages, including Delta and Omicron, and evaluated it on 2,088 patient specimens with near-perfect concordance to sequencing-based variant classification. Lastly, we implemented a combined Cas13 and Cas12 approach that enables quantitative measurement of SARS-CoV-2 and influenza A viral copies in samples. The mCARMEN platform enables high-throughput surveillance of multiple viruses and variants simultaneously, enabling rapid detection of SARS-CoV-2 variants., (© 2022. The Author(s).)

Epithelial-mesenchymal transition (EMT) programs operate within carcinoma cells, where they generate phenotypes associated with malignant progression. In their various manifestations, EMT programs enable epithelial cells to enter into a series of intermediate states arrayed along the E-M phenotypic spectrum. At present, we lack a coherent understanding of how carcinoma cells control their entrance into and continued residence in these various states, and which of these states favour the process of metastasis. Here we characterize a layer of EMT-regulating machinery that governs E-M plasticity (EMP). This machinery consists of two chromatin-modifying complexes, PRC2 and KMT2D-COMPASS, which operate as critical regulators to maintain a stable epithelial state. Interestingly, loss of these two complexes unlocks two distinct EMT trajectories. Dysfunction of PRC2, but not KMT2D-COMPASS, yields a quasi-mesenchymal state that is associated with highly metastatic capabilities and poor survival of patients with breast cancer, suggesting that great caution should be applied when PRC2 inhibitors are evaluated clinically in certain patient cohorts. These observations identify epigenetic factors that regulate EMP, determine specific intermediate EMT states and, as a direct consequence, govern the metastatic ability of carcinoma cells., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

This study aimed to determine the levels of contamination in Maumau stream as a result of horticultural activities in its vicinity. The stream was purposefully delineated into three blocks including upstream, midstream and downstream, where water samples were collected and analyzed for physicochemical attributes. Standard analytical procedures for water analysis were followed in laboratory analysis and the collected data was analyzed using Genstat software. Analyzed parameters include total dissolved solids (TDS), salinity, total suspended solids (TSS), sulphates (SO 3 -3 ), phosphates (PO 3 -3 ), nitrates (NO 3 - ), fluoride (Fl - ), turbidity, chloride (Cl - ), magnesium (Mg +2 ), sodium (Na + ), potassium (K + ) and zinc (Zn +2 ). The results were presented in tables and a graph against WHO standards. All measured parameters showed significant differences (p=<0.001) among their means across the sampling sites and control. The pH did not show a clear trend from upstream through midstream to downstream. The concentrations of chloride decreased down the stream with control, midstream and downstream showing no statistical significance. Means of fluoride, magnesium, phosphates, sulphates, total soluble solids and zinc increased down the course of the stream. Increasing concentrations of the physicochemical parameters down the stream was attributed majorly to release and addition of agrochemicals to the stream from the nearby farms. A lucid knowledge of the nexus between land use and water quality was recommended as a prime management implication. In conclusion, the water quality of Maumau stream is being degraded by horticultural activities along the stream. Key policy actions including river pegging should be adopted to protect the water quality., Competing Interests: The authors declare no conflict of interest., (© 2021 Published by Elsevier Ltd.)

The epithelial-mesenchymal transition (EMT) and primary ciliogenesis induce stem cell properties in basal mammary stem cells (MaSCs) to promote mammogenesis, but the underlying mechanisms remain incompletely understood. Here, we show that EMT transcription factors promote ciliogenesis upon entry into intermediate EMT states by activating ciliogenesis inducers, including FGFR1. The resulting primary cilia promote ubiquitination and inactivation of a transcriptional repressor, GLIS2, which localizes to the ciliary base. We show that GLIS2 inactivation promotes MaSC stemness, and GLIS2 is required for normal mammary gland development. Moreover, GLIS2 inactivation is required to induce the proliferative and tumorigenic capacities of the mammary tumor–initiating cells (MaTICs) of claudin-low breast cancers. Claudin-low breast tumors can be segregated from other breast tumor subtypes based on a GLIS2-dependent gene expression signature. Collectively, our findings establish molecular mechanisms by which EMT programs induce ciliogenesis to control MaSC and MaTIC stemness, mammary gland development, and claudin-low breast cancer formation.

Background: The suicide rate in Canada decreased by 24% during the past four decades. However, rates vary between provinces and territories, and not all jurisdictions experienced the same changes. This study examined suicide rates over time in the province of Newfoundland and Labrador., Methods: We used cross-sectional surveillance data from the Canadian Vital Statistics Death Database to examine suicide rates in Newfoundland and Labrador from 1981 to 2018. We calculated annual age-standardized suicide mortality rates and used joinpoint regression to estimate the average annual percent change (AAPC) in suicide rates overall and by sex, age group, and means of suicide., Results: From 1981 to 2018, 1759 deaths by suicide were recorded among people in Newfoundland and Labrador. The age-standardized suicide mortality rate increased more than threefold over the study period, from 4.6 to 15.4 deaths per 100,000. The suicide rate was higher among males than females, and accounted for 83.1% of suicide deaths (n = 1462); the male-to-female ratio of suicide deaths was 4.9 to 1. The average annual percent change in suicide rates was higher among females than males (6.3% versus 2.0%). Age-specific suicide rates increased significantly for all age groups, except seniors (aged 65 or older); the largest increase was among youth aged 10 to 24 years old (AAPC 3.5; 95% CI, 1.6 to 5.5). The predominant means of suicide was hanging/strangulation/suffocation, which accounted for 43.8% of all deaths by suicide., Conclusions: The suicide rate in Newfoundland and Labrador increased steadily between 1981 and 2018, which was in contrast to the national rate decline. The disparity between the provincial and national suicide rates and the variations by sex and age underscore the need for a public health approach to prevention that accounts for geographic and demographic differences in the epidemiology of suicide.

Objective: Communication is a key competency for medical education and comprehensive patient care. In rural environments, communication between rural family physicians and urban specialists is an essential pathway for clinical decision making. The aim of this study was to explore rural physicians' perspectives on communication with urban specialists during consultations and referrals., Setting: Newfoundland and Labrador, Canada., Participants: This qualitative study involved semistructured, one-on-one interviews with rural family physicians (n=11) with varied career stages, geographical regions, and community sizes., Results: Four themes specific to communication in rural practice were identified. The themes included: (1) understanding the contexts of rural care; (2) geographical isolation and patient transfer; and (3) respectful discourse; and (4) overcoming communication challenges in referrals and consultations., Conclusions: Communication between rural family physicians and urban specialists is a critical task in providing care for rural patients. Rural physicians see value in conveying unique aspects of rural clinical practice during communication with urban specialists, including context and the complexities of patient transfers., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

During the evolution of SARS-CoV-2 in humans, a D614G substitution in the spike glycoprotein (S) has emerged; virus containing this substitution has become the predominant circulating variant in the COVID-19 pandemic 1 . However, whether the increasing prevalence of this variant reflects a fitness advantage that improves replication and/or transmission in humans or is merely due to founder effects remains unknown. Here we use isogenic SARS-CoV-2 variants to demonstrate that the variant that contains S(D614G) has enhanced binding to the human cell-surface receptor angiotensin-converting enzyme 2 (ACE2), increased replication in primary human bronchial and nasal airway epithelial cultures as well as in a human ACE2 knock-in mouse model, and markedly increased replication and transmissibility in hamster and ferret models of SARS-CoV-2 infection. Our data show that the D614G substitution in S results in subtle increases in binding and replication in vitro, and provides a real competitive advantage in vivo-particularly during the transmission bottleneck. Our data therefore provide an explanation for the global predominance of the variant that contains S(D614G) among the SARS-CoV-2 viruses that are currently circulating.

Epileptic encephalopathies (EE) are severe epilepsy syndromes characterized by multiple seizure types, developmental delay and even regression. This class of disorders are increasingly being identified as resulting from de novo genetic mutations including many identified mutations in the family of chromodomain helicase DNA binding (CHD) proteins. In particular, several de novo pathogenic mutations have been identified in the gene encoding chromodomain helicase DNA binding protein 2 (CHD2), a member of the sucrose nonfermenting (SNF-2) protein family of epigenetic regulators. These mutations in the CHD2 gene are causative of early onset epileptic encephalopathy, abnormal brain function, and intellectual disability. Our understanding of the mechanisms by which modification or loss of CHD2 cause this condition remains poorly understood. Here, we review what is known and still to be elucidated as regards the structure and function of CHD2 and how its dysregulation leads to a highly variable range of phenotypic presentations.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in late 2019, and the outbreak rapidly evolved into the current coronavirus disease pandemic. SARS-CoV-2 is a respiratory virus that causes symptoms similar to those caused by influenza A and B viruses. On July 2, 2020, the US Food and Drug Administration granted emergency use authorization for in vitro diagnostic use of the Influenza SARS-CoV-2 Multiplex Assay. This assay detects influenza A virus at 10 2.0 , influenza B virus at 10 2.2 , and SARS-CoV-2 at 10 0.3 50% tissue culture or egg infectious dose, or as few as 5 RNA copies/reaction. The simultaneous detection and differentiation of these 3 major pathogens increases overall testing capacity, conserves resources, identifies co-infections, and enables efficient surveillance of influenza viruses and SARS-CoV-2.

Influenza pandemics are associated with severe morbidity, mortality, and social and economic disruption. Every summer in the United States, youths attending agricultural fairs are exposed to genetically diverse influenza A viruses (IAVs) circulating in exhibition swine, resulting in over 450 lab-confirmed zoonotic infections since 2010. Exhibition swine represent a small, defined population (∼1.5% of the U.S. herd), presenting a realistic opportunity to mitigate a pandemic threat by reducing IAV transmission in the animals themselves. Through intensive surveillance and genetic sequencing of IAVs in exhibition swine in six U.S. states in 2018 ( n  = 212), we characterized how a heterogeneous circuit of swine shows, comprising fairs with different sizes and geographic coverage, facilitates IAV transmission among exhibition swine and into humans. Specifically, we identified the role of an early-season national show in the propagation and spatial dissemination of a specific virus (H1δ-2) that becomes dominant among exhibition swine and is associated with the majority of zoonotic infections in 2018. These findings suggest that a highly targeted mitigation strategy, such as postponing swine shows for 1 to 2 weeks following the early-season national show, could potentially reduce IAV transmission in exhibition swine and spillover into humans, and this merits further study. IMPORTANCE The varying influenza A virus (IAV) exposure and infection status of individual swine facilitates introduction, transmission, and dissemination of diverse IAVs. Since agricultural fairs bring people into intimate contact with swine, they provide a unique interface for zoonotic transmission of IAV. Understanding the dynamics of IAV transmission through exhibition swine is critical to mitigating the high incidence of variant IAV cases reported in association with agricultural fairs. We used genomic sequences from our exhibition swine surveillance to characterize the hemagglutinin and full genotypic diversity of IAV at early-season shows and the subsequent dissemination through later-season agricultural fairs. We were able to identify a critical time point with important implications for downstream IAV and zoonotic transmission. With improved understanding of evolutionary origins of zoonotic IAV, we can inform public health mitigation strategies to ultimately reduce zoonotic IAV transmission and risk of pandemic IAV emergence., (Copyright © 2020 American Society for Microbiology.)

BackgroundDuring the 2016/17 influenza season, influenza B/VIC lineage variant viruses emerged with two (K 162 N 163 ) or three (K 162 N 163 D 164 ) amino acid (aa) deletions in the haemagglutinin (HA) protein. There are currently five antigenically distinct HA proteins expressed by co-circulating influenza B viruses: B/YAM, B/VIC V1A (no deletion), B/VIC V1A-2DEL (2 aa deletion) and two antigenically distinguishable groups of B/VIC V1A-3DEL (3 aa deletion). The prevalence of these viruses differs across geographical regions, making it critical to have a sensitive, rapid diagnostic assay that detects and distinguishes these influenza B variant viruses during surveillance.AimOur objective was to develop a real-time RT-PCR (rRT-PCR) assay for detection and discrimination of influenza B/VIC lineage variant viruses.MethodsWe designed a diagnostic assay with one pair of conserved primers and three probes specific to each genetic group. We used propagated influenza B/VIC variant viruses and clinical specimens to assess assay performance.ResultsThis rRT-PCR assay detects and distinguishes the influenza B/VIC V1A, B/VIC V1A-2DEL, and B/VIC V1A-3DEL variant viruses, with no cross-reactivity. This assay can be run as a multiplex reaction, allowing for increased testing efficiency and reduced cost.ConclusionCoupling this assay with the Centers for Disease Control and Prevention's Human Influenza Virus Real-Time RT-PCR Diagnostic Panel Influenza B Lineage Genotyping Kit results in rapid detection and characterisation of circulating influenza B viruses. Detailed surveillance information on these distinct influenza B variant viruses will provide insight into their prevalence and geographical distribution and could aid in vaccine recommendations.

Baloxavir, a new antiviral drug targeting cap-dependent endonuclease activity of polymerase acidic (PA) protein of influenza viruses, is now approved in multiple countries. Several substitutions at isoleucine 38 in PA protein (e.g., PA-I38T) have been associated with decreased baloxavir susceptibility in vitro and in vivo. In recent years, next generation sequencing (NGS) analysis and pyrosequencing have been used by CDC and U.S. Public Health Laboratories to monitor drug susceptibility of influenza viruses. Here we described an improved pyrosequencing assay for detecting influenza A viruses carrying substitutions at PA-38. Cyclic and customized orders of nucleotide dispensation were evaluated, and pyrosequencing results were compared to those generated using NGS. Our data showed that the customized nucleotide dispensation has improved the pyrosequencing assay performance in identification of double mixtures (e.g., PA-38I/T); however, identification of PA-38 variants in triple mixtures remains a challenge. While NGS analysis indicated the presence of PA-I38K in one clinical specimen and isolate, our attempts to detect this mutation by pyrosequencing or recover the virus carrying PA-I38K in cell culture were unsuccessful, raising a possibility of a rarely occurring sequencing error. Overall, pyrosequencing provides a convenient means to detect baloxavir resistant influenza viruses when NGS is unavailable or a faster turnaround time is required., (Published by Elsevier B.V.)

Tissue regeneration relies on adult stem cells (SCs) that possess the ability to self-renew and produce differentiating progeny. In an analogous manner, the development of certain cancers depends on a subset of tumor cells, called cancer stem cells (CSCs), with SC-like properties. In addition to being responsible for tumorigenesis, CSCs exhibit elevated resistance to therapy and thus drive tumor relapse post-treatment. The epithelial-mesenchymal transition (EMT) programs promote SC and CSC stemness in many epithelial tissues. Here, we provide an overview of the mechanisms underlying the relationship between stemness and EMT programs, which may represent therapeutic vulnerabilities for the treatment of cancers., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Purpose: Patients with community-acquired pneumonia (CAP) at low risk of death by CURB-65 scoring system are usually unnecessarily treated as inpatients generating additional economic and clinical burden. We aimed to implement an evidence-based clinical pathway to reduce hospital admissions of low-risk CAP and investigate factors related to mortality and readmissions within 30 days., Patients and Methods: From November 2015 to August 2017, a clinical pathway was implemented at 20 hospitals. We included patients aged >18 years, with a diagnosis of CAP by the attendant physician. The main outcome was the monthly proportion of low-risk CURB-65 admission after the implementation of the clinical pathway. Logistic regression models were performed to assess variables associated with mortality and readmission in the admitted population within 30 days., Results: We included 10,909 participants with suspected CAP. The proportion of low-risk CAP admitted decreased from 22.1% to 12.8% in the period. Among participants with low risk, there has been no perceptible increase in deaths (0.80%) or readmissions (6.92%). Regression analysis identified that CURB-65 variables, presence of pleural effusion (OR= 1.74; 95%CI=1.08-2.8; p=0.02) and leucopenia (OR= 2.47; 95%CI=1.11-5.48; p=0.02) were independently associated with 30-day mortality, whereas a prolonged hospital stay (OR= 2.09; 95%CI=1.14-3.83; p=0.01) was associated with 30-day readmission in the low-risk population., Conclusion: The implementations of a clinical pathway diminished the proportion of low-risk CAP admissions with no apparent increase in clinical outcomes within 30 days. Nonetheless, additional factors influence the clinical decision about the site of care management in low-risk CAP., Competing Interests: The authors declare that they have no competing interests to disclose related to the content of this manuscript., (© 2020 Moreira et al.)

Background: Blueberries are dietary sources of polyphenols, specifically anthocyanins. Anthocyanins have been identified as having a strong association with type 2 diabetes risk reduction; however, to date few human clinical trials have evaluated the potential beneficial health effects of blueberries in populations with type 2 diabetes., Objectives: We investigated the effects of blueberry consumption for 8 wk on cardiometabolic parameters in men with type 2 diabetes., Methods: In a double-blind, parallel-arm, randomized controlled trial, 52 men who are US veterans [mean baseline characteristics: age, 67 y (range: 51-75 y); weight, 102 kg (range: 80-130 kg); BMI (in kg/m 2 ), 34 (range: 26-45)] were randomly assigned to 1 of 2 intervention groups. The interventions were either 22 g freeze-dried blueberries or 22 g placebo. The study participants were asked to consume 11 g freeze-dried blueberries or placebo with each of their morning and evening meals along with their typical diet., Results: Mean ± SE hemoglobin A1c (7.1% ± 0.1% compared with 7.5% ± 0.2%; P  = 0.03), fructosamine (275.5 ± 4.1 compared with 292.4 ± 7.9 µmol/L; P  = 0.04), triglycerides (179.6 ± 10.1 compared with 199.6 ± 19.9 mg/dL; P  = 0.03), aspartate transaminase (23.2 ± 1.4 compared with 30.5 ± 2.7 units/L; P  = 0.02), and alanine transaminase (35.6 ± 1.5 compared with 48.3 ± 2.9 units/L; P  = 0.0003) were significantly lower for those consuming blueberries for 8 wk than for those consuming the placebo. Fasting plasma glucose concentrations; serum insulin, total cholesterol, LDL-cholesterol, HDL-cholesterol, and C-reactive protein concentrations; blood pressure; and body weight were not significantly different after 8 wk consumption of blueberries compared with the placebo., Conclusions: Consumption of 22 g freeze-dried blueberries for 8 wk may beneficially affect cardiometabolic health parameters in men with type 2 diabetes.This trial was registered at clinicaltrials.gov as NCT02972996., (Copyright © The Author(s) 2020.)

While working overnight at a swine exhibition, we identified an influenza A virus (IAV) outbreak in swine, Nanopore sequenced 13 IAV genomes from samples we collected, and predicted in real time that these viruses posed a novel risk to humans due to genetic mismatches between the viruses and current prepandemic candidate vaccine viruses (CVVs). We developed and used a portable IAV sequencing and analysis platform called Mia (Mobile Influenza Analysis) to complete and characterize full-length consensus genomes approximately 18 h after unpacking the mobile lab. Exhibition swine are a known source for zoonotic transmission of IAV to humans and pose a potential pandemic risk. Genomic analyses of IAV in swine are critical to understanding this risk, the types of viruses circulating in swine, and whether current vaccines developed for use in humans would be predicted to provide immune protection. Nanopore sequencing technology has enabled genome sequencing in the field at the source of viral outbreaks or at the bedside or pen-side of infected humans and animals. The acquired data, however, have not yet demonstrated real-time, actionable public health responses. The Mia system rapidly identified three genetically distinct swine IAV lineages from three subtypes, A(H1N1), A(H3N2), and A(H1N2). Analysis of the hemagglutinin (HA) sequences of the A(H1N2) viruses identified >30 amino acid differences between the HA1 of these viruses and the most closely related CVV. As an exercise in pandemic preparedness, all sequences were emailed to CDC collaborators who initiated the development of a synthetically derived CVV. IMPORTANCE Swine are influenza virus reservoirs that have caused outbreaks and pandemics. Genomic characterization of these viruses enables pandemic risk assessment and vaccine comparisons, though this typically occurs after a novel swine virus jumps into humans. The greatest risk occurs where large groups of swine and humans comingle. At a large swine exhibition, we used Nanopore sequencing and on-site analytics to interpret 13 swine influenza virus genomes and identified an influenza virus cluster that was genetically highly varied to currently available vaccines. As part of the National Strategy for Pandemic Preparedness exercises, the sequences were emailed to colleagues at the CDC who initiated the development of a synthetically derived vaccine designed to match the viruses at the exhibition. Subsequently, this virus caused 14 infections in humans and was the dominant U.S. variant virus in 2018.

Background: Therapeutic cannabis is being more widely used by patients to manage multiple symptoms, but the patterns of use in the palliative care population are not well defined. Objective: The primary aim of this pilot study was to describe the use of cannabis among patients attending a palliative care clinic (PCC). Design: The study was a retrospective chart review of patients seen at four different interval points during 2017 and 2018 in an ambulatory palliative care setting. Setting/Subjects: The study was conducted at a 396-bed rural academic medical center in the PCC, where the majority of patients have oncological diseases. Results: Clinicians saw 299 unique patients during the four one-month time periods reviewed. Eighty-three patients (27%) reported use of any form of cannabis. The most common reasons for cannabis use were pain ( n  = 49, 59%), anorexia ( n  = 16, 19%), insomnia ( n  = 14, 17%), nausea ( n  = 13, 16%), anxiety ( n  = 8, 10%), and depression ( n  = 5, 6%). Twenty-six patients (31%) used cannabis for more than one symptom. Among the 83 patients using cannabis, 60 (72%) were also prescribed opioids with 32% on immediate-release only and 25% on both immediate- and extended-release opioids. These 60 patients on opioids and cannabis represent 33% of all patients prescribed opioids in this clinic. Tetrahydrocannabinol was present in 25% of the 73 urine drug screens. Conclusions: Our data show a significant minority of patients in a PCC use cannabis. Further research should focus on more detailed information about formulation use, methods of ingestion, perceived efficacy, side effects, cost, and standardization of clinical practices. Given the prevalence of cannabis use, further research into its efficacy, side effects, and safety is needed, including whether patients with prior/active substance use receive more or less benefit or harm from cannabis use.