Search

BackgroundReports on medium and long-term sequelae of SARS-CoV-2 infections largely lack quantification of incidence and relative risk. We describe the rationale and methods of the Innovative Support for Patients with SARS-CoV-2 Registry (INSPIRE) that combines patient-reported outcomes with data from digital health records to understand predictors and impacts of SARS-CoV-2 infection.MethodsINSPIRE is a prospective, multicenter, longitudinal study of individuals with symptoms of SARS-CoV-2 infection in eight regions across the US. Adults are eligible for enrollment if they are fluent in English or Spanish, reported symptoms suggestive of acute SARS-CoV-2 infection, and if they are within 42 days of having a SARS-CoV-2 viral test (i.e., nucleic acid amplification test or antigen test), regardless of test results. Recruitment occurs in-person, by phone or email, and through online advertisement. A secure online platform is used to facilitate the collation of consent-related materials, digital health records, and responses to self-administered surveys. Participants are followed for up to 18 months, with patient-reported outcomes collected every three months via survey and linked to concurrent digital health data; follow-up includes no in-person involvement. Our planned enrollment is 4,800 participants, including 2,400 SARS-CoV-2 positive and 2,400 SARS-CoV-2 negative participants (as a concurrent comparison group). These data will allow assessment of longitudinal outcomes from SARS-CoV-2 infection and comparison of the relative risk of outcomes in individuals with and without infection. Patient-reported outcomes include self-reported health function and status, as well as clinical outcomes including health system encounters and new diagnoses.ResultsParticipating sites obtained institutional review board approval. Enrollment and follow-up are ongoing.ConclusionsThis study will characterize medium and long-term sequelae of SARS-CoV-2 infection among a diverse population, predictors of sequelae, and their relative risk compared to persons with similar symptomatology but without SARS-CoV-2 infection. These data may inform clinical interventions for individuals with sequelae of SARS-CoV-2 infection.

OBJECTIVE:Adequate resources are required to rapidly diagnose and treat pediatric musculoskeletal infection (MSKI). The workload MSKI consults contribute to pediatric orthopaedic services is unknown as prior epidemiologic studies are variable and negative work-ups are not included in national discharge databases. The hypothesis was tested that MSKI consults constitute a substantial volume of total consultations for pediatric orthopaedic services across the United States. STUDY DESIGN:Eighteen institutions from the Children's ORthopaedic Trauma and Infection Consortium for Evidence-based Study (CORTICES) group retrospectively reviewed a minimum of 1 year of hospital data, reporting the total number of surgeons, total consultations, and MSKI-related consultations. Consultations were classified by the location of consultation (emergency department or inpatient). Culture positivity rate and pathogens were also reported. RESULTS:87,449 total orthopaedic consultations and 7,814 MSKI-related consultations performed by 229 pediatric orthopaedic surgeons were reviewed. There was an average of 13 orthopaedic surgeons per site each performing an average of 154 consultations per year. On average, 9% of consultations were MSKI related and 37% of these consults yielded positive cultures. Finally, a weak inverse monotonic relationship was noted between percent culture positivity and percent of total orthopedic consults for MSKI. CONCLUSION:At large, academic pediatric tertiary care centers, pediatric orthopaedic services consult on an average of ~3,000 'rule-out' MSKI cases annually. These patients account for nearly 1 in 10 orthopaedic consultations, of which 1 in 3 are culture positive. Considering that 2 in 3 consultations were culture negative, estimating resources required for pediatric orthopaedic consult services to work up and treat children based on culture positive administrative discharge data underestimates clinical need. Finally, ascertainment bias must be considered when comparing differences in culture rates from different institution's pediatric orthopaedics services, given the variability in when orthopaedic physicians become involved in a MSKI workup.

The human milk microbiome (HMM) is hypothesized to be seeded by multiple factors, including the infant oral microbiome during breastfeeding. However, it is not known whether breastfeeding patterns (e.g., frequency or total time) impact the composition of the HMM. As part of the Mother-Infant Microbiomes, Behavior, and Ecology Study (MIMBES), we analyzed data from naturalistic observations of 46 mother-infant dyads living in the US Pacific Northwest and analyzed milk produced by the mothers for its bacterial diversity and composition. DNA was extracted from milk and the V1-V3 region of the 16S rRNA gene was amplified and sequenced. We hypothesized that number of breastfeeding bouts (breastfeeding sessions separated by >30 seconds) and total time breastfeeding would be associated with HMM α-diversity (richness, diversity, or evenness) and differential abundance of HMM bacterial genera. Multiple linear regression was used to examine associations between HMM α-diversity and the number of breastfeeding bouts or total time breastfeeding and selected covariates (infant age, maternal work outside the home, frequency of allomother physical contact with the infant, non-household caregiving network). HMM richness was inversely associated with number of breastfeeding bouts and frequency of allomother physical contact, but not total time breastfeeding. Infants' non-household caregiving network was positively associated with HMM evenness. In two ANCOM-BC analyses, abundances of 5 of the 35 most abundant genera were differentially associated with frequency of breastfeeding bouts (Bifidobacterium, Micrococcus, Pedobacter, Acidocella, Achromobacter); 5 genera (Bifidobacterium, Agreia, Pedobacter, Rugamonas, Stenotrophomonas) were associated with total time breastfeeding. These results indicate that breastfeeding patterns and infant caregiving ecology may play a role in influencing HMM composition. Future research is needed to identify whether these relationships are consistent in other populations and if they are associated with variation in the infant's gastrointestinal (including oral) microbiome.

[This corrects the article DOI: 10.1371/journal.pone.0259299.].

ImportanceSARS-CoV-2 infection can result in ongoing, relapsing, or new symptoms or other health effects after the acute phase of infection; termed post-acute sequelae of SARS-CoV-2 infection (PASC), or long COVID. The characteristics, prevalence, trajectory and mechanisms of PASC are ill-defined. The objectives of the Researching COVID to Enhance Recovery (RECOVER) Multi-site Observational Study of PASC in Adults (RECOVER-Adult) are to: (1) characterize PASC prevalence; (2) characterize the symptoms, organ dysfunction, natural history, and distinct phenotypes of PASC; (3) identify demographic, social and clinical risk factors for PASC onset and recovery; and (4) define the biological mechanisms underlying PASC pathogenesis.MethodsRECOVER-Adult is a combined prospective/retrospective cohort currently planned to enroll 14,880 adults aged ≥18 years. Eligible participants either must meet WHO criteria for suspected, probable, or confirmed infection; or must have evidence of no prior infection. Recruitment occurs at 86 sites in 33 U.S. states, Washington, DC and Puerto Rico, via facility- and community-based outreach. Participants complete quarterly questionnaires about symptoms, social determinants, vaccination status, and interim SARS-CoV-2 infections. In addition, participants contribute biospecimens and undergo physical and laboratory examinations at approximately 0, 90 and 180 days from infection or negative test date, and yearly thereafter. Some participants undergo additional testing based on specific criteria or random sampling. Patient representatives provide input on all study processes. The primary study outcome is onset of PASC, measured by signs and symptoms. A paradigm for identifying PASC cases will be defined and updated using supervised and unsupervised learning approaches with cross-validation. Logistic regression and proportional hazards regression will be conducted to investigate associations between risk factors, onset, and resolution of PASC symptoms.DiscussionRECOVER-Adult is the first national, prospective, longitudinal cohort of PASC among US adults. Results of this study are intended to inform public health, spur clinical trials, and expand treatment options.RegistrationNCT05172024.

Purpose The purpose of this research was to develop, implement, and test the efficacy of a theory-driven, evidence-informed peer leadership program for elementary school students (Grade 6 and 7; age 11–12 years) and the Grade 3/4 students with whom they were partnered. The primary outcome was teacher ratings of their Grade 6/7 students’ transformational leadership behaviors. Secondary outcomes included: Grade 6/7 students’ leadership self-efficacy, as well as Grade 3/4 motivation, perceived competence, general self-concept, fundamental movement skills, school-day physical activity, and program adherence, and program evaluation. Methods We conducted a two-arm cluster randomized controlled trial. In 2019, 6 schools comprising 7 teachers, 132 leaders, and 227 grade 3 and 4 students were randomly allocated to the intervention or waitlist control conditions. Intervention teachers took part in a half-day workshop (January 2019), delivered 7 x 40 minute lessons to Grade 6/7 peer leaders (February and March 2019), and these peer leaders subsequently ran a ten-week physical literacy development program for Grade 3/4 students (2x30 minutes sessions per week). Waitlist-control students followed their usual routines. Assessments were conducted at baseline (January 2019) and immediately post-intervention (June 2019). Results The intervention had no significant effect on teacher ratings of their students’ transformational leadership (b = 0.201, p = .272) after controlling for baseline and gender. There was no significant condition effect for Grade 6/7 student rated transformation leadership (b = 0.077, p = .569) or leadership self-efficacy (b = 3.747, p = .186) while controlling for baseline and gender. There were null findings for all outcomes related to Grade 3 and 4 students. Discussion Adaptions to the delivery mechanism were not effective in increasing leadership skills of older students or components of physical literacy in younger Grade 3/4 students. However, teacher self-reported adherence to the intervention delivery was high. Trial registration This trial was registered on December 19th, 2018 with Clinicaltrials.gov (NCT03783767), https://clinicaltrials.gov/ct2/show/NCT03783767.

BackgroundThis work aims to present a fast, affordable, and reproducible three-cell co-culture system that could represent the different cellular mechanisms of atherosclerosis, extending from atherogenesis to pathological intimal thickening.Methods and resultsWe built four culture models: (i) Culture model #1 (representing normal arterial intima), where human coronary artery endothelial cells were added on top of Matrigel-coated collagen type I matrix, (ii) Culture model #2 (representing atherogenesis), which demonstrated the subendothelial accumulation and oxidative modification of low-density lipoproteins (LDL), (iii) Culture model #3 (representing intimal xanthomas), which demonstrated the monocyte adhesion to the endothelial cell monolayer, transmigration into the subendothelial space, and transformation to lipid-laden macrophages, (iv) Culture model #4 (representing pathological intimal thickening), which incorporated multiple layers of human coronary artery smooth muscle cells within the matrix. Coupling this model with different shear stress conditions revealed the effect of low shear stress on the oxidative modification of LDL and the upregulation of pro-inflammatory molecules and matrix-degrading enzymes. Using electron microscopy, immunofluorescence confocal microscopy, protein and mRNA quantification assays, we showed that the behaviors exhibited by the endothelial cells, macrophages and vascular smooth muscle cells in these models were very similar to those exhibited by these cell types in nascent and intermediate atherosclerotic plaques in humans. The preparation time of the cultures was 24 hours.ConclusionWe present three-cell co-culture models of human atherosclerosis. These models have the potential to allow cost- and time-effective investigations of the mechanobiology of atherosclerosis and new anti-atherosclerotic drug therapies.

Mammalian microRNA expression is dysregulated in human cancer. However, the functional relevance of many microRNAs in the context of tumor biology remains unclear. Using CRISPR-Cas9 technology, we performed a global loss-of-function screen to simultaneously test the functions of individual microRNAs and protein-coding genes during the growth of a myeloid leukemia cell line. This approach identified evolutionarily conserved human microRNAs that suppress or promote cell growth, revealing that microRNAs are extensively integrated into the molecular networks that control tumor cell physiology. miR-155 was identified as a top microRNA candidate promoting cellular fitness, which we confirmed with two distinct miR-155-targeting CRISPR-Cas9 lentiviral constructs. Further, we performed anti-correlation functional profiling to predict relevant microRNA-tumor suppressor gene or microRNA-oncogene interactions in these cells. This analysis identified miR-150 targeting of p53, a connection that was experimentally validated. Taken together, our study describes a powerful genetic approach by which the function of individual microRNAs can be assessed on a global level, and its use will rapidly advance our understanding of how microRNAs contribute to human disease.

Southern Africa spans nearly 7 million km2 and contains approximately 80% of the world’s savannah elephants (Loxodonta africana) mostly living in isolated protected areas. Here we ask what are the prospects for improving the connections between these populations? We combine 1.2 million telemetry observations from 254 elephants with spatial data on environmental factors and human land use across eight southern African countries. Telemetry data show what natural features limit elephant movement and what human factors, including fencing, further prevent or restrict dispersal. The resulting intersection of geospatial data and elephant presences provides a map of suitable landscapes that are environmentally appropriate for elephants and where humans allow elephants to occupy. We explore the environmental and anthropogenic constraints in detail using five case studies. Lastly, we review all the major potential connections that may remain to connect a fragmented elephant metapopulation and document connections that are no longer feasible.

Background Reports on medium and long-term sequelae of SARS-CoV-2 infections largely lack quantification of incidence and relative risk. We describe the rationale and methods of the Innovative Support for Patients with SARS-CoV-2 Registry (INSPIRE) that combines patient-reported outcomes with data from digital health records to understand predictors and impacts of SARS-CoV-2 infection. Methods INSPIRE is a prospective, multicenter, longitudinal study of individuals with symptoms of SARS-CoV-2 infection in eight regions across the US. Adults are eligible for enrollment if they are fluent in English or Spanish, reported symptoms suggestive of acute SARS-CoV-2 infection, and if they are within 42 days of having a SARS-CoV-2 viral test (i.e., nucleic acid amplification test or antigen test), regardless of test results. Recruitment occurs in-person, by phone or email, and through online advertisement. A secure online platform is used to facilitate the collation of consent-related materials, digital health records, and responses to self-administered surveys. Participants are followed for up to 18 months, with patient-reported outcomes collected every three months via survey and linked to concurrent digital health data; follow-up includes no in-person involvement. Our planned enrollment is 4,800 participants, including 2,400 SARS-CoV-2 positive and 2,400 SARS-CoV-2 negative participants (as a concurrent comparison group). These data will allow assessment of longitudinal outcomes from SARS-CoV-2 infection and comparison of the relative risk of outcomes in individuals with and without infection. Patient-reported outcomes include self-reported health function and status, as well as clinical outcomes including health system encounters and new diagnoses. Results Participating sites obtained institutional review board approval. Enrollment and follow-up are ongoing. Conclusions This study will characterize medium and long-term sequelae of SARS-CoV-2 infection among a diverse population, predictors of sequelae, and their relative risk compared to persons with similar symptomatology but without SARS-CoV-2 infection. These data may inform clinical interventions for individuals with sequelae of SARS-CoV-2 infection.

By identifying homogeneity in bone and soft tissue covariation patterns in living hominids, it is possible to produce facial approximation methods with interspecies compatibility. These methods may be useful for producing facial approximations of fossil hominids that are more realistic than currently possible. In this study, we conducted an interspecific comparison of the nasomaxillary region in chimpanzees and modern humans with the aim of producing a method for predicting the positions of the nasal tips of Plio-Pleistocene hominids. We addressed this aim by first collecting and performing regression analyses of linear and angular measurements of nasal cavity length and inclination in modern humans (Homo sapiens; n = 72) and chimpanzees (Pan troglodytes; n = 19), and then performing a set of out-of-group tests. The first test was performed on four subjects that belonged to the same genus as the training sample, i.e., Homo (n = 2) and Pan (n = 2), and the second test, which functioned as an interspecies compatibility test, was performed on Pan paniscus (n = 1), Gorilla gorilla (n = 3), Pongo pygmaeus (n = 1), Pongo abelli (n = 1), Symphalangus syndactylus (n = 3), and Papio hamadryas (n = 3). We identified statistically significant correlations in both humans and chimpanzees with slopes that displayed homogeneity of covariation. Prediction formulae combining these data were found to be compatible with humans and chimpanzees as well as all other African great apes, i.e., bonobos and gorillas. The main conclusion that can be drawn from this study is that our set of regression models for approximating the position of the nasal tip are homogenous among humans and African apes, and can thus be reasonably extended to ancestors leading to these clades.

The Florida Horse Conch, Triplofusus giganteus, one of the largest marine gastropods in the world, has been intensely exploited by shell collectors, curio dealers, and commercial harvest for over a century and is now in decline. Effective management of horse conch populations requires better data on commercial and recreational harvest intensities but also on the species' intrinsic capacity to recover. Here, we use stable oxygen and carbon isotope sclerochronology to investigate the horse conch's life history, including its maximum life span, growth rates, age at first spawning, and number of lifetime spawning seasons. The largest two shells studied (460 and 475 mm linear shell length) grew for 13 and 11 years, respectively. Growth curves for these shells, extrapolated out to the length of the record size shell (606 mm linear shell length) predict a maximum age of just 16 years. Carbon isotopes and field photographs of spawning females suggest that females mature relatively late in life. However, the largest horse conchs remaining in the wild are also smaller and younger than those studied here. Thus, the largest females left in the wild could have few lifetime spawning events. High fecundity can buffer horse conchs from overfishing but only if females reach spawning age and reproductive-age females are protected. Our study highlights the usefulness of stable isotope sclerochronology for characterizing the life histories of molluscan species now too uncommon to study through traditional mark and recapture approaches.

[This corrects the article DOI: 10.1371/journal.pone.0244423.].

BackgroundUganda is a low-income country with high fertility, adolescent birth, and maternal mortality rates. How Ugandan Ministry of Health antenatal education guidelines have been implemented into standardized health education and how pregnant women utilize health facilities remains unclear.ObjectiveWe aimed to determine how women obtain education during pregnancy, what guidelines health educators follow, and what barriers exist to receiving antenatal care in Lweza Village, Uganda.MethodsHousehold surveys were conducted with women in Lweza who were or had previously been pregnant. Focus group discussions were conducted with community members and Lweza Primary School teachers. Interviews were conducted with key informants, including midwives, a traditional birth attendant, a community leader, and a Village Health Team member. Data collection was done in English along with a Luganda translator.ResultsOf the 100 household surveys conducted, 86% of women did not meet the WHO recommendation of 8 antenatal appointments during their pregnancies. Reasons cited for inadequate visits included facing long wait times (>7 h) at health facilities, getting education from family or traditional healers, or being told to delay antenatal care until 6 months pregnant. Informant interviews revealed that no standardized antenatal education program exists. Respondents felt least educated on family planning and postpartum depression, despite 37% of them reporting symptoms consistent with postpartum depression. Education was also lacking on the use of traditional herbs, although most women (60%) reported using them during pregnancy.ConclusionsMost women in Lweza do not receive 8 antenatal appointments during their pregnancies or any standardized antenatal education. Educational opportunities on family planning, postpartum depression, and the safety of traditional herbs during pregnancy exist. Future studies should focus on ways to overcome barriers to antenatal care, which could include implementing community-based education programs to improve health outcomes for women in Lweza Village.

Naegleria fowleri is a pathogenic, thermophilic, free-living amoeba which causes primary amebic meningoencephalitis (PAM). Penetrating the olfactory mucosa, the brain-eating amoeba travels along the olfactory nerves, burrowing through the cribriform plate to its destination: the brain's frontal lobes. The amoeba thrives in warm, freshwater environments, with peak infection rates in the summer months and has a mortality rate of approximately 97%. A major contributor to the pathogen's high mortality is the lack of sensitivity of N. fowleri to current drug therapies, even in the face of combination-drug therapy. To enable rational drug discovery and design efforts we have pursued protein production and crystallography-based structure determination efforts for likely drug targets from N. fowleri. The genes were selected if they had homology to drug targets listed in Drug Bank or were nominated by primary investigators engaged in N. fowleri research. In 2017, 178 N. fowleri protein targets were queued to the Seattle Structural Genomics Center of Infectious Disease (SSGCID) pipeline, and to date 89 soluble recombinant proteins and 19 unique target structures have been produced. Many of the new protein structures are potential drug targets and contain structural differences compared to their human homologs, which could allow for the development of pathogen-specific inhibitors. Five of the structures were analyzed in more detail, and four of five show promise that selective inhibitors of the active site could be found. The 19 solved crystal structures build a foundation for future work in combating this devastating disease by encouraging further investigation to stimulate drug discovery for this neglected pathogen.

In modern humans, facial soft tissue thicknesses have been shown to covary with craniometric dimensions. However, to date it has not been confirmed whether these relationships are shared with non-human apes. In this study, we analyze these relationships in chimpanzees (Pan troglodytes) with the aim of producing regression models for approximating facial soft tissue thicknesses in Plio-Pleistocene hominids. Using CT scans of 19 subjects, 637 soft tissue, and 349 craniometric measurements, statistically significant multiple regression models were established for 26 points on the face and head. Examination of regression model validity resulted in minimal differences between observed and predicted soft tissue thickness values. Assessment of interspecies compatibility using a bonobo (Pan paniscus) and modern human subject resulted in minimal differences for the bonobo but large differences for the modern human. These results clearly show that (1) soft tissue thicknesses covary with craniometric dimensions in P. troglodytes, (2) confirms that such covariation is uniformly present in both extant Homo and Pan species, and (3) suggests that chimp-derived regression models have interspecies compatibility with hominids who have similar craniometric dimensions to P. troglodytes. As the craniometric dimensions of early hominids, such as South African australopithecines, are more similar to P. troglodytes than those of H. sapiens, chimpanzee-derived regression models may be used for approximating their craniofacial anatomy. It is hoped that the results of the present study and the reference dataset for facial soft tissue thicknesses of chimpanzees it provides will encourage further research into this topic.

BackgroundRabies is a viral disease of animals and people causing fatal encephalomyelitis if left untreated. Although effective pre- and post-exposure vaccines exist, they are not widely available in many endemic countries within Africa. Since many individuals in these countries remain at risk of infection, post-exposure healthcare-seeking behaviors are crucial in preventing infection and warrant examination.MethodologyA rabies knowledge, attitudes, and practices survey was conducted at 24 geographically diverse sites in Uganda during 2013 to capture information on knowledge concerning the disease, response to potential exposure events, and vaccination practices. Characteristics of the surveyed population and of the canine-bite victim sub-population were described. Post-exposure healthcare-seeking behaviors of canine-bite victims were examined and compared to the related healthcare-seeking attitudes of non-bite victim respondents. Wealth scores were calculated for each household, rabies knowledge was scored for each non-bitten survey respondent, and rabies exposure risk was scored for each bite victim. Logistic regression was used to determine the independent associations between different variables and healthcare-seeking behaviors among canine-bite victims as well as attitudes of non-bitten study respondents.ResultsA total of 798 households were interviewed, capturing 100 canine-bite victims and a bite incidence of 2.3 per 100 person-years. Over half of bite victims actively sought medical treatment (56%), though very few received rabies post-exposure prophylaxis (3%). Bite victims who did not know or report the closest location where PEP could be received were less likely to seek medical care (p = 0.05). Respondents who did not report having been bitten by a dog with higher knowledge scores were more likely to respond that they would both seek medical care (p = 0.00) and receive PEP (p = 0.06) after a potential rabies exposure event.ConclusionsThere was varying discordance between what respondents who did not report having been bitten by a dog said they would do if bitten by a dog when compared to the behaviors exhibited by canine-bite victims captured in the KAP survey. Bite victims seldom elected to wash their wound or receive PEP. Having lower rabies knowledge was a barrier to theoretically seeking care and receiving PEP among not bitten respondents, indicating a need for effective and robust educational programs in the country.

Breast ductal carcinoma in situ (DCIS) is a non-obligate precursor of invasive ductal carcinoma (IDC). It is still unclear which DCIS will become invasive and which will remain indolent. Patients often receive surgery and radiotherapy, but this early intervention has not produced substantial decreases in late-stage disease. Sprouty proteins are important regulators of ERK/MAPK signaling and have been studied in various cancers. We hypothesized that Sprouty4 is an endogenous inhibitor of ERK/MAPK signaling and that its loss/reduced expression is a mechanism by which DCIS lesions progress toward IDC, including triple-negative disease. Using immunohistochemistry, we found reduced Sprouty4 expression in IDC patient samples compared to DCIS, and that ERK/MAPK phosphorylation had an inverse relationship to Sprouty4 expression. These observations were reproduced using a 3D culture model of disease progression. Knockdown of Sprouty4 in MCF10.DCIS cells increased ERK/MAPK phosphorylation as well as their invasive capability, while overexpression of Sprouty4 in MCF10.CA1d IDC cells reduced ERK/MAPK phosphorylation, invasion, and the aggressive phenotype exhibited by these cells. Immunofluorescence experiments revealed reorganization of the actin cytoskeleton and relocation of E-cadherin back to the cell surface, consistent with the restoration of adherens junctions. To determine whether these effects were due to changes in ERK/MAPK signaling, MEK1/2 was pharmacologically inhibited in IDC cells. Nanomolar concentrations of MEK162/binimetinib restored an epithelial-like phenotype and reduced pericellular proteolysis, similar to Sprouty4 overexpression. From these data we conclude that Sprouty4 acts to control ERK/MAPK signaling in DCIS, thus limiting the progression of these premalignant breast lesions.

HPV infection results in changes in host gene methylation which, in turn, are thought to contribute to the neoplastic progression of HPV-associated cancers. The objective of this study was to identify joint and disease-specific genome-wide methylation changes in anal and cervical cancer as well as changes in high-grade pre-neoplastic lesions. Formalin-fixed paraffin-embedded (FFPE) anal tissues (n = 143; 99% HPV+) and fresh frozen cervical tissues (n = 28; 100% HPV+) underwent microdissection, DNA extraction, HPV genotyping, bisulfite modification, DNA restoration (FFPE) and analysis by the Illumina HumanMethylation450 Array. Differentially methylated regions (DMR; t test q0.3) were compared between normal and cancer specimens in partial least squares (PLS) models and then used to classify anal or cervical intraepithelial neoplasia-3 (AIN3/CIN3). In AC, an 84-gene PLS signature (355 significant probes) differentiated normal anal mucosa (NM; n = 9) from AC (n = 121) while a 36-gene PLS signature (173 significant probes) differentiated normal cervical epithelium (n = 10) from CC (n = 9). The CC progression signature was validated using three independent publicly available datasets (n = 424 cases). The AC and CC progression PLS signatures were interchangeable in segregating normal, AIN3/CIN3 and AC and CC and were found to include 17 common overlapping hypermethylated genes. Moreover, these signatures segregated AIN3/CIN3 lesions similarly into cancer-like and normal-like categories. Distinct methylation changes occur across the genome during the progression of AC and CC with overall similar profiles and add to the evidence suggesting that HPV-driven oncogenesis may result in similar non-random methylomic events. Our findings may lead to identification of potential epigenetic drivers of HPV-associated cancers and also, of potential markers to identify higher risk pre-cancerous lesions.

Atherosclerosis is a lipid-driven chronic inflammatory disease that leads to the formation of plaques in the inner lining of arteries. Plaques form over a range of phenotypes, the most severe of which is vulnerable to rupture and causes most of the clinically significant events. In this study, we evaluated the efficacy of nanoparticles (NPs) to differentiate between two plaque phenotypes based on accumulation kinetics in a mouse model of atherosclerosis. This model uses a perivascular cuff to induce two regions of disturbed wall shear stress (WSS) on the inner lining of the instrumented artery, low (upstream) and multidirectional (downstream), which, in turn, cause the development of an unstable and stable plaque phenotype, respectively. To evaluate the influence of each WSS condition, in addition to the final plaque phenotype, in determining NP uptake, mice were injected with NPs at intermediate and fully developed stages of plaque growth. The kinetics of artery wall uptake were assessed in vivo using dynamic contrast-enhanced magnetic resonance imaging. At the intermediate stage, there was no difference in NP uptake between the two WSS conditions, although both were different from the control arteries. At the fully-developed stage, however, NP uptake was reduced in plaques induced by low WSS, but not multidirectional WSS. Histological evaluation of plaques induced by low WSS revealed a significant inverse correlation between the presence of smooth muscle cells and NP accumulation, particularly at the plaque-lumen interface, which did not exist with other constituents (lipid and collagen) and was not present in plaques induced by multidirectional WSS. These findings demonstrate that NP accumulation can be used to differentiate between unstable and stable murine atherosclerosis, but accumulation kinetics are not directly influenced by the WSS condition. This tool could be used as a diagnostic to evaluate the efficacy of experimental therapeutics for atherosclerosis.

The COVID-19 pandemic has reintroduced questions regarding the potential risk of SARS-CoV-2 exposure amongst passengers on an aircraft. Quantifying risk with computational fluid dynamics models or contact tracing methods alone is challenging, as experimental results for inflight biological aerosols is lacking. Using fluorescent aerosol tracers and real time optical sensors, coupled with DNA-tagged tracers for aerosol deposition, we executed ground and inflight testing on Boeing 767 and 777 airframes. Analysis here represents tracer particles released from a simulated infected passenger, in multiple rows and seats, to determine the exposure risk via penetration into breathing zones in that row and numerous rows ahead and behind the index case. We present here conclusions from 118 releases of fluorescent tracer particles, with 40+ Instantaneous Biological Analyzer and Collector sensors placed in passenger breathing zones for real-time measurement of simulated virus particle penetration. Results from both airframes showed a minimum reduction of 99.54% of 1 μm aerosols from the index source to the breathing zone of a typical passenger seated directly next to the source. An average 99.97 to 99.98% reduction was measured for the breathing zones tested in the 767 and 777, respectively. Contamination of surfaces from aerosol sources was minimal, and DNA-tagged 3 μm tracer aerosol collection techniques agreed with fluorescent methodologies.

Accurate maps of species ranges are essential to inform conservation, but time-consuming to produce and update. Given the pace of change of knowledge about species distributions and shifts in ranges under climate change and land use, a need exists for timely mapping approaches that enable batch processing employing widely available data. We develop a systematic approach of batch-processing range maps and derived Area of Habitat maps for terrestrial bird species with published ranges below 125,000 km2 in Central and South America. (Area of Habitat is the habitat available to a species within its range.) We combine existing range maps with the rapidly expanding crowd-sourced eBird data of presences and absences from frequently surveyed locations, plus readily accessible, high resolution satellite data on forest cover and elevation to map the Area of Habitat available to each species. Users can interrogate the maps produced to see details of the observations that contributed to the ranges. Previous estimates of Areas of Habitat were constrained within the published ranges and thus were, by definition, smaller-typically about 30%. This reflects how little habitat within suitable elevation ranges exists within the published ranges. Our results show that on average, Areas of Habitat are 12% larger than published ranges, reflecting the often-considerable extent that eBird records expand the known distributions of species. Interestingly, there are substantial differences between threatened and non-threatened species. Some 40% of Critically Endangered, 43% of Endangered, and 55% of Vulnerable species have Areas of Habitat larger than their published ranges, compared with 31% for Near Threatened and Least Concern species. The important finding for conservation is that threatened species are generally more widespread than previously estimated.

OBJECTIVE:Adequate resources are required to rapidly diagnose and treat pediatric musculoskeletal infection (MSKI). The workload MSKI consults contribute to pediatric orthopaedic services is unknown as prior epidemiologic studies are variable and negative work-ups are not included in national discharge databases. The hypothesis was tested that MSKI consults constitute a substantial volume of total consultations for pediatric orthopaedic services across the United States. STUDY DESIGN:Eighteen institutions from the Children's ORthopaedic Trauma and Infection Consortium for Evidence-based Study (CORTICES) group retrospectively reviewed a minimum of 1 year of hospital data, reporting the total number of surgeons, total consultations, and MSKI-related consultations. Consultations were classified by the location of consultation (emergency department or inpatient). Culture positivity rate and pathogens were also reported. RESULTS:87,449 total orthopaedic consultations and 7,814 MSKI-related consultations performed by 229 pediatric orthopaedic surgeons were reviewed. There was an average of 13 orthopaedic surgeons per site each performing an average of 154 consultations per year. On average, 9% of consultations were MSKI related and 37% of these consults yielded positive cultures. Finally, a weak inverse monotonic relationship was noted between percent culture positivity and percent of total orthopedic consults for MSKI. CONCLUSION:At large, academic pediatric tertiary care centers, pediatric orthopaedic services consult on an average of ~3,000 'rule-out' MSKI cases annually. These patients account for nearly 1 in 10 orthopaedic consultations, of which 1 in 3 are culture positive. Considering that 2 in 3 consultations were culture negative, estimating resources required for pediatric orthopaedic consult services to work up and treat children based on culture positive administrative discharge data underestimates clinical need. Finally, ascertainment bias must be considered when comparing differences in culture rates from different institution's pediatric orthopaedics services, given the variability in when orthopaedic physicians become involved in a MSKI workup.

Recent research has shown relationships between health outcomes and residence proximity to unconventional oil and natural gas development (UOGD). The challenge of connecting health outcomes to environmental stressors requires ongoing research with new methodological approaches. We investigated UOGD density and well emissions and their association with symptom reporting by residents of southwest Pennsylvania. A retrospective analysis was conducted on 104 unique, de-identified health assessments completed from 2012-2017 by residents living in proximity to UOGD. A novel approach to comparing estimates of exposure was taken. Generalized linear modeling was used to ascertain the relationship between symptom counts and estimated UOGD exposure, while Threshold Indicator Taxa Analysis (TITAN) was used to identify associations between individual symptoms and estimated UOGD exposure. We used three estimates of exposure: cumulative well density (CWD), inverse distance weighting (IDW) of wells, and annual emission concentrations (AEC) from wells within 5 km of respondents' homes. Taking well emissions reported to the Pennsylvania Department of Environmental Protection, an air dispersion and screening model was used to estimate an emissions concentration at residences. When controlling for age, sex, and smoker status, each exposure estimate predicted total number of reported symptoms (CWD, p

ObjectivesTo apply spinal cord injury (SCI) specific waist circumference (WC) cutoff point to identify risks of 1) obesity, 2) metabolic syndrome (MetS), 3) cardiovascular disease (CVD).MethodsThirty-six men with chronic SCI underwent anthropometric measurements, dual-energy x-ray absorptiometry (DXA), and magnetic resonance imaging (MRI) to measure total and regional adiposity. An SCI specific WC cutoff point of 86.5 cm was applied to the existing general population criteria. Pearson chi-square (χ2) analyses tested the difference in the number of participants classified as obese using the SCI specific cutoff point compared to the general population criteria. Sensitivity and specificity analyses relative to percentage body fat mass and visceral adipose tissue was used to assess classification performance of this cutoff point. The interrater reliability for three definitions of MetS was assessed using Cohen's Kappa (κ) values. Linear regression analyses were utilized to propose SCI specific Framingham Coronary Heart Disease Risk Score (FRS) cutoff value.ResultsUsing SCI specific WC cutoff point of 86.5 cm, 36% of participants were classified as obese compared to only 3% when using WC of 102 cm (P < 0.001). Relative to percentage body fat mass, the general population WC cutoff point of 102 cm had a sensitivity of 6.3% and specificity of 100% both which changed to 68.8% and 90%, respectively, with a SCI specific cutoff point of 86.5 cm. Similar results were obtained when using visceral adipose tissue as a reference. The Kappa (κ) values improved substantially after using SCI specific criteria (0.95 ± 0.05) compared to the general population criteria (0.47 ± 0.28) for three definitions of MetS. The SCI specific FRS cutoff value of 6 was predicted after applying a WC cutoff of 86.5 cm.ConclusionsUsing the existing general population criteria underestimated persons with SCI who are at risk of developing obesity, MetS, and CVD. The recommended SCI specific criteria are likely to distinguish those at risks of developing comorbidities and allow healthcare providers to intervene in a timely manner.

The COVID-19 pandemic disrupted the world in 2020 by spreading at unprecedented rates and causing tens of thousands of fatalities within a few months. The number of deaths dramatically increased in regions where the number of patients in need of hospital care exceeded the availability of care. Many COVID-19 patients experience Acute Respiratory Distress Syndrome (ARDS), a condition that can be treated with mechanical ventilation. In response to the need for mechanical ventilators, designed and tested an emergency ventilator (EV) that can control a patient's peak inspiratory pressure (PIP) and breathing rate, while keeping a positive end expiratory pressure (PEEP). This article describes the rapid design, prototyping, and testing of the EV. The development process was enabled by rapid design iterations using additive manufacturing (AM). In the initial design phase, iterations between design, AM, and testing enabled a working prototype within one week. The designs of the 16 different components of the ventilator were locked by additively manufacturing and testing a total of 283 parts having parametrically varied dimensions. In the second stage, AM was used to produce 75 functional prototypes to support engineering evaluation and animal testing. The devices were tested over more than two million cycles. We also developed an electronic monitoring system and with automatic alarm to provide for safe operation, along with training materials and user guides. The final designs are available online under a free license. The designs have been transferred to more than 70 organizations in 15 countries. This project demonstrates the potential for ultra-fast product design, engineering, and testing of medical devices needed for COVID-19 emergency response.

ObjectiveAerobic fitness may be beneficial for neuroanatomical structure. However, few have investigated this in emerging adults while also accounting for potential sex differences. Here we examine aerobic fitness level, sex, and their interaction in relation to cortical thickness, surface area, and volume.MethodSixty-three young adults between the ages of 16-26 were balanced for sex and demonstrated a wide range of aerobic fitness levels. Exclusion criteria included left-handedness, past-year independent Axis-I disorders, major medical/neurologic disorders, prenatal medical issues, prenatal alcohol/illicit drug exposure, or excessive substance use. Participants completed an MRI scan and a graded exercise test to volitional fatigue (VO2 max). Data analyses were run in Freesurfer and data was corrected for multiple comparisons with Monte Carlo simulations at .05.ResultsMales demonstrated higher VO2 values. Higher VO2 values were statistically independently related to thinner lateral occipital, superior parietal, cuneus, precuneus, and inferior parietal regions, smaller lateral occipital volume, and larger inferior parietal surface area. Compared to females, males had larger volume in rostral anterior cingulate, lateral occipital, and superior frontal regions, and greater surface area in fusiform, inferior parietal, rostral and caudal anterior cingulate, and superior parietal regions. VO2*Sex interactions revealed higher-fit females had higher inferior parietal, paracentral, and supramarginal surface area, while lower-fit males showed larger surface area in these same regions.ConclusionsIndividuals with higher aerobic fitness performance had thinner cortices, lower volume, and larger surface area in sensorimotor regions than lower fit individuals, perhaps suggesting earlier neuromaturation in higher fit individuals. Larger surface area was associated with higher-fit females and lower-fit males. Thus both sex and aerobic fitness are important in shaping brain health in emerging adults.

BackgroundPropensity score techniques can reduce confounding and bias in observational studies. Such analyses are able to measure and balance pre-determined covariates between treated and untreated groups, leading to results that can approximate those generated by randomized prospective studies when such trials are not feasible. The most commonly used propensity score -based analytic technique is propensity score matching (PSM). Although PSM popularity has continued to increase in medical literature, improper methodology or methodological reporting may lead to biased interpretation of treatment effects or limited scientific reproducibility and generalizability. In this study, we aim to characterize and assess the quality of PSM methodology reporting in high-impact otolaryngologic literature.MethodsPubMed and Embase based systematic review of the top 20 journals in otolaryngology, as measured by impact factor from the Journal Citations Reports from 2012 to 2018, for articles using PSM analysis throughout their publication history. Eligible articles were reviewed and assessed for quality and reporting of PSM methodology.ResultsOur search yielded 101 studies, of which 92 were eligible for final analysis and review. The proportion of studies utilizing PSM increased significantly over time (p < 0.001). Nearly all studies (96.7%, n = 89) specified the covariates used to calculate propensity scores. Covariate balance was illustrated in 67.4% (n = 62) of studies, most frequently through p-values. A minority (17.4%, n = 16) of studies were found to be fully reproducible according to previously established criteria.ConclusionsWhile PSM analysis is becoming increasingly prevalent in otolaryngologic literature, the quality of PSM methodology reporting can be improved. We provide potential recommendations for authors regarding optimal reporting for analyses using PSM.

Genetically modified mice carrying engrafted human tissues provide useful models to study human cell biology in physiologically relevant contexts. However, there remain several obstacles limiting the compatibility of human cells within their mouse hosts. Among these is inadequate cross-reactvitiy between certain mouse cytokines and human cellular receptors, depriving the graft of important survival and growth signals. To circumvent this problem, we utilized a lentivirus-based delivery system to express physiologically relevant levels of human interleukin-7 (hIL-7) in Rag2-/-gammac-/- mice following a single intravenous injection. hIL-7 promoted homeostatic proliferation of both adoptively transferred and endogenously generated T-cells in Rag2-/-gammac-/- Human Immune System (HIS) mice. Interestingly, we found that hIL-7 increased T lymphocyte numbers in the spleens of HIV infected HIS mice without affecting viral load. Taken together, our study unveils a versatile approach to deliver human cytokines to HIS mice, to both improve engraftment and determine the impact of cytokines on human diseases.

Thermoresponsive polymers, such as poly(N-isopropyl acrylamide) (PNIPAM), have been identified and used as cell culture substrates, taking advantage of the polymer's lower critical solution temperature (LCST) to mechanically harvest cells. This technology bypasses the use of biochemical enzymes that cleave important cell-cell and cell-matrix interactions. In this study, the process of electrospinning is used to fabricate and characterize aligned PNIPAM nanofiber scaffolds that are biocompatible and thermoresponsive. Nanofiber scaffolds produced by electrospinning possess a 3D architecture that mimics native extracellular matrix, providing physical and chemical cues to drive cell function and phenotype. We present a factorial design of experiments (DOE) approach to systematically determine the effects of different electrospinning process parameters on PNIPAM nanofiber diameter and alignment. Results show that high molecular weight PNIPAM can be successfully electrospun into both random and uniaxially aligned nanofiber mats with similar fiber diameters by simply altering the speed of the rotating mandrel collector from 10,000 to 33,000 RPM. PNIPAM nanofibers were crosslinked with OpePOSS, which was verified using FTIR. The mechanical properties of the scaffolds were characterized using dynamic mechanical analysis, revealing an order of magnitude difference in storage modulus (MPa) between cured and uncured samples. In summary, cross-linked PNIPAM nanofiber scaffolds were determined to be stable in aqueous culture, biocompatible, and thermoresponsive, enabling their use in diverse cell culture applications.

Given the uncertain population status of low-density, widely-occurring raptors, monitoring changes in abundance and distribution is critical to conserving populations. Nest-based monitoring is a common, useful approach, but the difficulty and expense of monitoring raptor nests and importance of reliable trend data to conservation requires that limited resources are allocated efficiently. Power analyses offer a helpful tool to ensure that monitoring programs have the ability to detect trends and to optimize financial resources devoted to monitoring. We evaluated alternative monitoring designs for raptors to identify appropriate survey effort to detect population trends. We used data collected from a territory-occupancy study of ferruginous hawks throughout Wyoming to guide simulations and evaluate the ability to detect trends in occupancy rates. Results suggest that greater gains in precision of trend estimation may be achieved through the addition of more sites and not more visits; statistical power was ≥80% when monitoring lasted 20 years and population declines were 20%; and probability of detection affected statistical power less than rates of population decline. Monitoring at least 150 sites for 20 years would provide reasonable estimates of trend in occupancy given certain rates of detection and occupancy, but only for population declines of 20%. Removal sampling did not result in substantial changes of any metrics used to evaluate simulations, providing little justification for employing the standard design if territory occupancy is the variable of interest. Initial rates of territory occupancy may be biased high, a problem inherent to many studies that monitor territory occupancy. We explored the effects of lower rates of initial occupancy on the ability to detect trends. Although we present data from a study of ferruginous hawks, our simulations can be applied to other raptor species with similar life history and population dynamics to provide guidance for future trend estimation of territory occupancy.

Personal shark deterrents offer the potential of a non-lethal solution to protect individuals from negative interactions with sharks, but the claims of effectiveness of most deterrents are based on theory rather than robust testing of the devices themselves. Therefore, there is a clear need for thorough testing of commercially available shark deterrents to provide the public with information on their effectiveness. Using a modified stereo-camera system, we quantified behavioural interactions between Carcharodon carcharias (white sharks) and a baited target in the presence of a commercially available electric anklet shark deterrent, the Electronic Shark Defense System (ESDS). The stereo-camera system enabled accurate assessment of the behavioural responses of C. carcharias when approaching an ESDS. We found that the ESDS had limited meaningful effect on the behaviour of C. carcharias, with no significant reduction in the proportion of sharks interacting with the bait in the presence of the active device. At close proximity (< 15.5 cm), the active ESDS did show a significant reduction in the number of sharks biting the bait, but this was countered by an increase in other, less aggressive, interactions. The ESDS discharged at a frequency of 7.8 Hz every 5.1 s for 2.5 s, followed by an inactive interval of 2.6 s. As a result, many sharks may have encountered the device in its inactive state, resulting in a reduced behavioural response. Consequently, decreasing the inactive interval between pulses may improve the overall effectiveness of the device, but this would not improve the effective deterrent range of the device, which is primarily a factor of the voltage gradient rather than the stimulus frequency. In conclusion, given the very short effective range of the ESDS and its unreliable deterrent effect, combined with the fact that shark-bite incidents are very rare, it is unlikely that the current device would significantly reduce the risk of a negative interaction with C. carcharias.

Plasma chemistry is widely used in diagnostic and research settings in sea turtles. However, plasma discolorations such as hemolysis are often not considered in data interpretation. The objectives of this study were to (1) evaluate the effects of moderate hemolysis on plasma electrolytes, minerals, and proteins using dry chemistry analysis (DCA) and protein electrophoresis from nesting leatherback sea turtles (Dermochelys coriacea) from Florida and to (2) establish blood analyte reference intervals. Twenty-six plasma samples with absence of hemolysis were selected and sub-divided into one non-hemolytic aliquot and an aliquot that was experimentally manipulated to mimic moderate hemolysis. Plasma samples were analyzed for hemoglobin using a handheld photometer; sodium, potassium, chloride, magnesium, calcium, phosphorus, and total protein using DCA; and protein electrophoresis. Packed cell volume and hemoglobin were measured in corresponding whole blood samples. Reference intervals were established. All analytes except calcium and pre-albumin were significantly higher and the calcium:phosphorus and albumin:globulin ratios were significantly lower in hemolytic plasma compared to non-hemolytic plasma. Alpha2-globulins and potassium were the analytes most impacted by hemolysis, averaging 3.3- and 2.0-fold higher in hemolyzed samples, respectively, indicating that (1) hemoglobin migrates into the alpha2-globulin region in this species and (2) notable intracellular potassium is released into plasma with hemolysis. Attempted conversion factors for compensation of hemolysis were considered inaccurate for 4 of 16 analytes due to non-significant regression lines. We also report that PCV provides an estimate of hemoglobin (g/L) using the formula: (2.59 × PCV) + 24.59. Given the spurious effects of hemolysis, the degree of this artifact should be reported with biochemistry data, and samples with moderate to severe hemolysis should be excluded from datasets when interpreting electrolyte, mineral, and protein results. This will ensure accurate data interpretation for individual turtles in rehabilitation or research settings and population-level data relevant to conservation-focused projects.

Land use influences the prevalence and distribution of ticks due to the intimate relationship of ticks with their environment. This relationship occurs because land use alters two essential tick requirements: vertebrate hosts for blood meals and a suitable microclimate when off-host. Given the risks to human and animal health associated with pathogens transmitted by ticks, there is an ongoing need to understand the impact of environmental drivers on tick distributions. Here, we assessed how landscape features, neighborhood effects, and edges influenced tick occupancy and abundance across an agricultural landscape in southern Africa. We found that Rhipicephalus appendiculatus and Rhipicephalus simus increased in abundance closer to protected savanna, while Haemaphysalis elliptica increased in abundance closer to human habitation. The composition of the landscape surrounding savanna patches also differentially influenced the occupancy of each tick species; H. elliptica was more likely to be found in savanna patches surrounded by subsistence agriculture while R. appendiculatus and R. simus were more likely to be found in savanna surrounded by sugarcane monocultures. At the local scale we found that R. appendiculatus and R. simus avoided savanna edges. The availability of hosts and variation in vegetation structure between commercial agriculture, subsistence agriculture, and savanna likely drove the distribution of ticks at the landscape scale. Understanding how anthropogenic land use influences where ticks occur is useful for land use planning and for assessing public and animal health risks associated with ticks and tick-borne diseases.

In single molecule fluorescence studies, background emission from labeled substrates often restricts their concentrations to non-physiological nanomolar values. One approach to address this challenge is the use of zero-mode waveguides (ZMWs), nanoscale holes in a thin metal film that physically and optically confine the observation volume allowing much higher concentrations of fluorescent substrates. Standard fabrication of ZMWs utilizes slow and costly E-beam nano-lithography. Herein, ZMWs are made using a self-assembled mask of polystyrene microspheres, enabling fabrication of thousands of ZMWs in parallel without sophisticated equipment. Polystyrene 1 μm dia. microbeads self-assemble on a glass slide into a hexagonal array, forming a mask for the deposition of metallic posts in the inter-bead interstices. The width of those interstices (and subsequent posts) is adjusted within 100-300 nm by partially fusing the beads at the polystyrene glass transition temperature. The beads are dissolved in toluene, aluminum or gold cladding is deposited around the posts, and those are dissolved, leaving behind an array ZMWs. Parameter optimization and the performance of the ZMWs are presented. By using colloidal self-assembly, typical laboratories can make use of sub-wavelength ZMW technology avoiding the availability and expense of sophisticated clean-room environments and equipment.

The development of automatic methods for segmenting anatomy from medical images is an important goal for many medical and healthcare research areas. Datasets that can be used to train and test computer algorithms, however, are often small due to the difficulties in obtaining experts to segment enough examples. Citizen science provides a potential solution to this problem but the feasibility of using the public to identify and segment anatomy in a medical image has not been investigated. Our study therefore aimed to explore the feasibility, in terms of performance and motivation, of using citizens for such purposes. Public involvement was woven into the study design and evaluation. Twenty-nine citizens were recruited and, after brief training, asked to segment the spine from a dataset of 150 magnetic resonance images. Participants segmented as many images as they could within three one-hour sessions. Their accuracy was evaluated by comparing them, as individuals and as a combined consensus, to the segmentations of three experts. Questionnaires and a focus group were used to determine the citizens' motivation for taking part and their experience of the study. Citizen segmentation accuracy, in terms of agreement with the expert consensus segmentation, varied considerably between individual citizens. The citizen consensus, however, was close to the expert consensus, indicating that when pooled, citizens may be able to replace or supplement experts for generating large image datasets. Personal interest and a desire to help were the two most common reasons for taking part in the study.

Taiwan had been considered rabies free since 1961, until a newly established wildlife disease surveillance program identified rabies virus transmission within the Formosan ferret-badger (Melogale moschata subaurantiaca) in 2013. Ferret-badgers occur throughout southern China and Southeast Asia, but their ecological niche is not well described.As an initial feasibility assessment for potential rabies control measures, field camera trapping and pen assessment of 6 oral rabies vaccine (ORV) baits were conducted in Taiwan in 2013. 46 camera nights were recorded; 6 Formosan ferret-badgers and 14 non-target mammals were sighted. No baits were consumed by ferret-badgers and 8 were consumed by non-target mammals. Penned ferret-badgers ingested 5 of the 18 offered baits. When pen and field trials were combined, and analyzed for palatability, ferret-badgers consumed 1 of 9 marshmallow baits (11.1%), 1 of 21 fishmeal baits (4.8%), 0 of 3 liver baits, and 3 of 3 fruit-flavored baits. It took an average of 261 minutes before ferret-badgers made oral contact with the non-fruit flavored baits, and 34 minutes for first contact with the fruit-based bait. Overall, ferret-badgers sought out the fruit baits 8 times faster, spent a greater proportion of time eating fruit baits, and were 7.5 times more likely to have ruptured the vaccine container of the fruit-based bait.Ferret-badgers are now recognized as rabies reservoir species in China and Taiwan, through two independent 'dog to ferret-badger' host-shift events. Species of ferret-badgers can be found throughout Indochina, where they may be an unrecognized rabies reservoir. Findings from this initial study underscore the need for further captive and field investigations of fruit-based attractants or baits developed for small meso-carnivores. Non-target mammals' competition for baits, ants, bait design, and dense tropical landscape represent potential challenges to effective ORV programs that will need to be considered in future studies.

Insulin-like growth factors (Igfs) are key regulators of key biological processes such as embryonic development, growth, and tissue repair and regeneration. The role of Igf in myogenesis is well documented and, in zebrafish, promotes fin and heart regeneration. However, the mechanism of action of Igf in muscle repair and regeneration is not well understood. Using adult zebrafish extraocular muscle (EOM) regeneration as an experimental model, we show that Igf1 receptor blockage using either chemical inhibitors (BMS754807 and NVP-AEW541) or translation-blocking morpholino oligonucleotides (MOs) reduced EOM regeneration. Zebrafish EOMs regeneration depends on myocyte dedifferentiation, which is driven by early epigenetic reprogramming and requires autophagy activation and cell cycle reentry. Inhibition of Igf signaling had no effect on either autophagy activation or cell proliferation, indicating that Igf signaling was not involved in the early reprogramming steps of regeneration. Instead, blocking Igf signaling produced hypercellularity of regenerating EOMs and diminished myosin expression, resulting in lack of mature differentiated muscle fibers even many days after injury, indicating that Igf was involved in late re-differentiation steps. Although it is considered the main mediator of myogenic Igf actions, Akt activation decreased in regenerating EOMs, suggesting that alternative signaling pathways mediate Igf activity in muscle regeneration. In conclusion, Igf signaling is critical for re-differentiation of reprogrammed myoblasts during late steps of zebrafish EOM regeneration, suggesting a regulatory mechanism for determining regenerated muscle size and timing of differentiation, and a potential target for regenerative therapy.

BACKGROUND:Canine transmitted rabies kills an estimated 59,000 people annually, despite proven methods for elimination through mass dog vaccination. Challenges in directing and monitoring numerous remote vaccination teams across large geographic areas remain a significant barrier to the up-scaling of focal vaccination programmes to sub-national and national level. Smartphone technology (mHealth) is increasingly being used to enhance the coordination and efficiency of public health initiatives in developing countries, however examples of successful scaling beyond pilot implementation are rare. This study describes a smartphone app and website platform, "Mission Rabies App", used to co-ordinate rabies control activities at project sites in four continents to vaccinate over one million dogs. METHODS:Mission Rabies App made it possible to not only gather relevant campaign data from the field, but also to direct vaccination teams systematically in near real-time. The display of user-allocated boundaries on Google maps within data collection forms enabled a project manager to define each team's region of work, assess their output and assign subsequent areas to progressively vaccinate across a geographic area. This ability to monitor work and react to a rapidly changing situation has the potential to improve efficiency and coverage achieved, compared to regular project management structures, as well as enhancing capacity for data review and analysis from remote areas. The ability to plot the location of every vaccine administered facilitated engagement with stakeholders through transparent reporting, and has the potential to motivate politicians to support such activities. RESULTS:Since the system launched in September 2014, over 1.5 million data entries have been made to record dog vaccinations, rabies education classes and field surveys in 16 countries. Use of the system has increased year-on-year with adoption for mass dog vaccination campaigns at the India state level in Goa and national level in Haiti. CONCLUSIONS:Innovative approaches to rapidly scale mass dog vaccination programmes in a sustained and systematic fashion are urgently needed to achieve the WHO, OIE and FAO goal to eliminate canine-transmitted human deaths by 2030. The Mission Rabies App is an mHealth innovation which greatly reduces the logistical and managerial barriers to implementing large scale rabies control activities. Free access to the platform aims to support pilot campaigns to better structure and report on proof-of-concept initiatives, clearly presenting outcomes and opportunities for expansion. The functionalities of the Mission Rabies App may also be beneficial to other infectious disease interventions.

Mechanically ventilated surgical patients have a variety of bacterial flora that are often undetectable by traditional culture methods. The source of infection in many of these patients remains unclear. To address this clinical problem, the microbiome profile and host inflammatory response in bronchoalveolar lavage samples from the surgical intensive care unit were examined relative to clinical pathology diagnoses. The hypothesis was tested that clinical diagnosis of respiratory tract flora were similar to culture positive lavage samples in both microbiome and inflammatory profile. Bronchoalveolar lavage samples were collected in the surgical intensive care unit as standard of care for intubated individuals with a clinical pulmonary infection score of >6 or who were expected to be intubated for >48 hours. Cytokine analysis was conducted with the Bioplex Pro Human Th17 cytokine panel. The microbiome of the samples was sequenced for the 16S rRNA region using the Ion Torrent. Microbiome diversity analysis showed the culture-positive samples had the lowest levels of diversity and culture negative with the highest based upon the Shannon-Wiener index (culture positive: 0.77 ± 0.36, respiratory tract flora: 2.06 ± 0.73, culture negative: 3.97 ± 0.65). Culture-negative samples were not dominated by a single bacterial genera. Lavages classified as respiratory tract flora were more similar to the culture-positive in the microbiome profile. A comparison of cytokine expression between groups showed increased levels of cytokines (IFN-g, IL-17F, IL-1B, IL-31, TNF-a) in culture-positive and respiratory tract flora groups. Culture-positive samples exhibited a more robust immune response and reduced diversity of bacterial genera. Lower cytokine levels in culture-negative samples, despite a greater number of bacterial species, suggest a resident nonpathogenic bacterial community may be indicative of a normal pulmonary environment. Respiratory tract flora samples were most similar to the culture-positive samples and may warrant classification as culture-positive when considering clinical treatment.

BACKGROUND/OBJECTIVES:To examine associations of different anthropometric measurements of central adiposity to visceral adipose tissue (measured via multi-axial magnetic resonance imaging; MRI) and cardiometabolic disease risk factors in men with spinal cord injury (SCI). Additionally, to determine population-specific seated/supine waist and abdominal circumference cutoffs, which may identify men at increased risk of cardiometabolic disease. PARTICIPANTS/METHODS:Twenty-two men with chronic SCI underwent MRI scans, anthropometric measurements along with assessments of various cardiometabolic risk biomarkers. Pearson/part (accounting for age as a covariate) correlation coefficients were calculated to determine the associations between study variables. Abdominal and waist circumference cutoffs were extrapolated using the slope of linear regression equations. RESULTS:Seated/supine abdominal and waist circumferences were (P < 0.01) associated with MRI visceral fat cross-sectional area (VATCSA), VAT volume and CSA:TotalCSA. Low density lipoprotein, non-high-density lipoprotein and total cholesterol were positively associated with seated/supine abdominal and waist circumferences after controlling for age; r = 0.50-0.61, r = 0.46-0.58, r = 0.52-0.58, P < 0.05, respectively. Tumor necrosis factor alpha was associated with seated/supine abdominal and waist circumferences after accounting for age; r = 0.49-0.51 and r = 0.48-0.56, P < 0.05 respectively. The population-specific cutoffs were 86.5cm and 88.3cm for supine waist and abdominal circumferences, respectively, as well as 89cm and 101cm for seated waist and abdominal circumferences, respectively. After dichotomizing VATCSA (< or ≥ 100cm2), peak oxygen uptake, triglycerides, insulin sensitivity and glycated hemoglobin were different (P < 0.05) between groups. After dichotomizing (< or ≥ 86.5cm) supine waist circumference, VATCSA, triglycerides and insulin sensitivity were different (P < 0.05) between groups. CONCLUSIONS:Seated/supine circumferences are associated with both central adiposity and biomarkers of cardiometabolic disease risk in persons with SCI. Population-specific cutoffs are proposed herein to identify central adiposity and potential cardiometabolic disease risk after SCI.

Somatic complaints are often the presenting symptoms of major depressive disorder (MDD) in the outpatient context, because this may go unrecognized. It is well understood that MDD carries an increased risk of suicide. This study aimed to identify the risk factors and association with both MDD and suicidality among Han Chinese outpatients.A multicenter study was carried out in 5189 outpatient adults (≥18 years old) in four general hospitals in Guangzhou, China. The 1392 patients who had the Patient Health Questionnaire-9 (PHQ-9) score ≥ 5, indicating depressive symptoms were offered an interview with a psychiatrist by the Mini International Neuropsychiatric Interview (MINI); 819 patients consented and completed the MINI interview. MINI module B was used to assess suicidality. Stepwise binary logistic models were used to estimate the relationship between a significant risk factor and suicide or MDD. According to with or without MDD, the secondary analysis was performed using the logistic regression model for the risk of suicidility.The current prevalence of MDD and the one month prevalence of suicidality were 3.7% and 2.3% respectively. The odds ratio of suicidality in women was more than twice that in men (OR = 2.62; 95% CI 1.45-4.76). Other risk factors which were significantly associated with suicidality were: living alone, higher education, self-reported depression, getting psychiatric diagnoses (MDD, anxiety disorders, and bipolar disorders). Significant risk factors for MDD were also noticed, such as comorbid anxiety disorders, self-reported anxiety, insomnia, suicidal ideation.It's a cross-sectional study in outpatient clinics using self-report questionnaires.This study provides valuable data about the risk factors and association of MDD and suicide risk in adult outpatients in Han Chinese. Those factors allow better the employment of preventative measures.

The α6 nicotinic acetylcholine receptor (nAChR) subunit is an attractive drug target for treating nicotine addiction because it is present at limited sites in the brain including the reward pathway. Lynx1 modulates several nAChR subtypes; lynx1-nAChR interaction sites could possibly provide drug targets. We found that dopaminergic cells from the substantia nigra pars compacta (SNc) express lynx1 mRNA transcripts and, as assessed by co-immunoprecipitation, α6 receptors form stable complexes with lynx1 protein, although co-transfection with lynx1 did not affect nicotine-induced currents from cell lines transfected with α6 and β2. To test whether lynx1 is important for the function of α6 nAChRs in vivo, we bred transgenic mice carrying a hypersensitive mutation in the α6 nAChR subunit (α6L9'S) with lynx1 knockout mice, providing a selective probe of the effects of lynx1 on α6* nAChRs. Lynx1 removal reduced the α6 component of nicotine-mediated rubidium efflux and dopamine (DA) release from synaptosomal preparations with no effect on numbers of α6β2 binding sites, indicating that lynx1 is functionally important for α6* nAChR activity. No effects of lynx1 removal were detected on nicotine-induced currents in slices from SNc, suggesting that lynx1 affects presynaptic α6* nAChR function more than somatic function. In the absence of agonist, lynx1 removal did not alter DA release in dorsal striatum as measured by fast scan cyclic voltammetry. Lynx1 removal affected some behaviors, including a novel-environment assay and nicotine-stimulated locomotion. Trends in 24-hour home-cage behavior were also suggestive of an effect of lynx1 removal. Conditioned place preference for nicotine was not affected by lynx1 removal. The results show that some functional and behavioral aspects of α6-nAChRs are modulated by lynx1.

Wikipedia is a gateway to knowledge. However, the extent to which this gateway ends at Wikipedia or continues via supporting citations is unknown. Wikipedia's gateway functionality has implications for information design and education, notably in medicine. This study aims to establish benchmarks for the relative distribution and referral (click) rate of citations-as indicated by presence of a Digital Object Identifier (DOI)-from Wikipedia, with a focus on medical citations. DOIs referred from the English Wikipedia in August 2016 were obtained from Crossref.org. Next, based on a DOI's presence on a WikiProject Medicine page, all DOIs in Wikipedia were categorized as medical (WP:MED) or non-medical (non-WP:MED). Using this categorization, referred DOIs were classified as WP:MED, non-WP:MED, or BOTH, meaning the DOI may have been referred from either category. Data were analyzed using descriptive and inferential statistics. Out of 5.2 million Wikipedia pages, 4.42% (n = 229,857) included at least one DOI. 68,870 were identified as WP:MED, with 22.14% (n = 15,250) featuring one or more DOIs. WP:MED pages featured on average 8.88 DOI citations per page, whereas non-WP:MED pages had on average 4.28 DOI citations. For DOIs only on WP:MED pages, a DOI was referred every 2,283 pageviews and for non-WP:MED pages every 2,467 pageviews. DOIs from BOTH pages accounted for 12% (n = 58,475). The referral of DOI citations found in BOTH could not be assigned to WP:MED or non-WP:MED, as the page from which the referral was made was not provided with the data. While these results cannot provide evidence of greater citation referral from WP:MED than non-WP:MED, they do provide benchmarks to assess strategies for changing referral patterns. These changes might include editors adopting new methods for designing and presenting citations or the introduction of teaching strategies that address the value of consulting citations as a tool for extending learning.