The Ancient Evolutionary History of Polyomaviruses.

Polyomaviruses are a family of DNA tumor viruses that are known to infect mammals and birds. To investigate the deeper evolutionary history of the family, we used a combination of viral metagenomics, bioinformatics, and structural modeling approaches to identify and characterize polyomavirus sequences associated with fish and arthropods. Analyses drawing upon the divergent new sequences indicate that polyomaviruses have been gradually co-evolving with their animal hosts for at least half a billion years. Phylogenetic analyses of individual polyomavirus genes suggest that some modern polyomavirus species arose after ancient recombination events involving distantly related polyomavirus lineages. The improved evolutionary model provides a useful platform for developing a more accurate taxonomic classification system for the viral family Polyomaviridae.

Distinct Viral Lineages from Fish and Amphibians Reveal the Complex Evolutionary History of Hepadnaviruses.

UNLABELLED: Hepadnaviruses (hepatitis B viruses [HBVs]) are the only animal viruses that replicate their DNA by reverse transcription of an RNA intermediate. Until recently, the known host range of hepadnaviruses was limited to mammals and birds. We obtained and analyzed the first amphibian HBV genome, as well as several prototype fish HBVs, which allow the first comprehensive comparative genomic analysis of hepadnaviruses from four classes of vertebrates. Bluegill hepadnavirus (BGHBV) was characterized from in-house viral metagenomic sequencing. The African cichlid hepadnavirus (ACHBV) and the Tibetan frog hepadnavirus (TFHBV) were discovered using in silico analyses of the whole-genome shotgun and transcriptome shotgun assembly databases. Residues in the hydrophobic base of the capsid (core) proteins, designated motifs I, II, and III, are highly conserved, suggesting that structural constraints for proper capsid folding are key to capsid protein evolution. Surface proteins in all vertebrate HBVs contain similar predicted membrane topologies, characterized by three transmembrane domains. Most striking was the fact that BGHBV, ACHBV, and the previously described white sucker hepadnavirus did not form a fish-specific monophyletic group in the phylogenetic analysis of all three hepadnaviral genes. Notably, BGHBV was more closely related to the mammalian hepadnaviruses, indicating that cross-species transmission events have played a major role in viral evolution. Evidence of cross-species transmission was also observed with TFHBV. Hence, these data indicate that the evolutionary history of the hepadnaviruses is more complex than previously realized and combines both virus-host codivergence over millions of years and host species jumping. IMPORTANCE: Hepadnaviruses are responsible for significant disease in humans (hepatitis B virus) and have been reported from a diverse range of vertebrates as both exogenous and endogenous viruses. We report the full-length genome of a novel hepadnavirus from a fish and the first hepadnavirus genome from an amphibian. The novel fish hepadnavirus, sampled from bluegills, was more closely related to mammalian hepadnaviruses than to other fish viruses. This phylogenetic pattern reveals that, although hepadnaviruses have likely been associated with vertebrates for hundreds of millions of years, they have also been characterized by species jumping across wide phylogenetic distances.

Myxobolus axelrodi n. sp. (Myxosporea: Myxobolidae) a parasite infecting the brain and retinas of the cardinal tetra Paracheirodon axelrodi (Teleostei: Characidae).

An investigation of mortalities in a group of cardinal tetras Paracheirodon axelrodi Meyers, 1936, a popular ornamental fish, revealed myxozoan parasites in ventricles of the brains in 3/10 fish and the ocular retina of a fourth. Parasite impacts were unclear, as additional histopathological findings were present, including bacterial dermatitis and meningitis. Ethanol-preserved specimens pooled from multiple fish were used for morphological characterization of myxospores. Elongate, teardrop myxospores were 20.5 ± 0.7-µm (mean ± SD; range = 19.0-21.8 µm) long, 6.6 ± 0.5-µm (5.7-7.9 µm) wide, and 5.1 ± 0.4-µm (4.8-5.9 µm) thick (valvular width). Two, unequally sized, apical, pyriform polar capsules were in the same plane as the sutural ridge. The larger measured 9.9 ± 0.8-µm (8.0-11.2 µm) long and 3.8 ± 0.3-µm (3.2-4.8 µm) wide. The smaller was 4.1 ± 0.3-µm (3.5-4.5 µm) long and 2.0 ± 0.1-µm (1.8-2.3 µm) wide. Identical 1912 bp 18S rRNA sequences were obtained from two pooled spore samples from tetra brains, which did not match any sequences in the NCBI nr/nt database. Phylogenetically, these parasites grouped loosely within a clade containing Myxobolus spp. from other South American characins and Unicauda spp. from siluriform catfish. Myxospores shared some morphological similarities with Myxobolus inaequus from the unrelated glass knifefish (Order: Gymnotiformes), but were genetically divergent (<85 % similarity) from other myxozoan parasites of South American characins and shared few morphological features or tissue predilection sites. Based on host and tissue tropism, spore morphology, and 18S rRNA sequencing, we report this isolate as a previously unknown species, Myxobolus axelrodi n. sp.

Virus-associated papillomatous skin lesions in a giant guitarfish Rhynchobatus djiddensis: a case report.

Although elasmobranch species are increasingly displayed in public aquaria, knowledge of disease in wild and captive elasmobranchs, as well as the agents involved, remains limited, and descriptions are often incomplete. This report describes papillomatous skin lesions in a juvenile giant guitarfish Rhynchobatus djiddensis associated with intranuclear viral particles. Skin biopsies were collected from multiple, friable, raised, villonodular skin lesions affecting pigmented and non-pigmented skin of the caudal fin and ventrum, respectively. Microscopic examination revealed papillary proliferation of the epidermis, with widespread marked karyomegaly of squamous epithelial cells. In approximately 75% of nuclei, chromatin was marginated by one to multiple, large, amphophilic inclusions. Large numbers of unencapsulated, 75 nm, icosahedral viral particles were observed to form large arrays in affected nuclei using transmission electron microscopy. Based on intranuclear location, particle size and morphology, a consensus nested-PCR for adenovirus polymerase was attempted. However, no adenoviral gene sequence was amplified. The nature of the involved virus remains unknown and an ongoing area of investigation. Lesions regressed completely over a 6 mo period, during which time the animal showed no signs of systemic illness, and there has been no recrudescence for 6 mo following resolution. Two cohorts of similar age and in close contact with the case animal were unaffected.

Bicycle boulevards and changes in physical activity and active transportation: findings from a natural experiment.

OBJECTIVE: This study evaluates changes in physical activity and active transportation associated with installation of new bicycle boulevards. METHODS: This natural experiment study uses data from a longitudinal panel of adults with children (n=353) in Portland, OR. Activity and active transportation outcomes were measured with GPS and accelerometers worn for up to 5 days in 2010-11 and 2012-13. The effect of the treatment was estimated using difference in differences estimation and multivariate regression models. RESULTS: In five of the seven models, the interaction term was not significant, indicating that after controlling for the main effects of time and exposure separately, there was no correlation between being in a treatment area and minutes of moderate and vigorous physical activity (MVPA) per day, bicycling >10 min, walking >20 min, minutes of walking (if >20), or making a bike trip. Significant covariates included rain, being female, living closer to downtown, and attitudes towards bicycling, walking, and car safety. CONCLUSION: This study could not confirm an increase in physical activity or active transportation among adults with children living near newly installed bicycle boulevards. Additional pre/post studies are encouraged, as well as research on the length of time after installation that behavior change is likely to occur.

Adapting and Developing an Academic and Community Practice Collaborative Care Model for Metastatic Breast Cancer Care (Project ADAPT): Protocol for an Implementation Science-Based Study.

BACKGROUND: Metastatic breast cancer (MBC) remains incurable despite significant treatment advances. Coordinating care for patients with MBC can be challenging given the various treatment options, available clinical trials, and frequent need for ancillary services. To optimize MBC care, we designed a project for adapting and developing an academic and community practice collaborative care model for MBC care (Project ADAPT), based on the Ending Metastatic Breast Cancer for Everyone (EMBRACE) program developed at Dana Farber Cancer Institute. OBJECTIVE: We aim to describe the implementation science-based study design and innovative components of Project ADAPT. METHODS: Project ADAPT uses the Dynamic Adaptation Process informed by the Exploration, Preparation, Implementation, Sustainment framework. Washington University School of Medicine (WUSM) partnered with 3 community hospitals in the St. Louis region covering rural and urban settings. The exploration and preparation phases provide patient and provider feedback on current referral practices to finalize the approach for the implementation phase. At the implementation phase, we will enroll patients with MBC at these 3 community sites to evaluate potential collaborative care at WUSM and assess the impact of this collaborative care model on referral satisfaction and acceptability for patients with MBC and their providers. Patients may then return to their community site for care or continue to receive part of their care at WUSM. We are incorporating virtual and digital health strategies to improve MBC care coordination in order to minimize patient burden. RESULTS: The exploration phase is ongoing. As of August 2021, we have recruited 21 patient and provider participants to complete surveys of the current collaborative care process at WUSM. Using a 2-tailed paired t test, 44 patients (including 10 patients from the exploration phase) and 32 oncologists are required to detect an effect size of 0.5 with 80% power at a level of significance of .05. Throughout this phase and in preparation for the implementation phase, we have iteratively updated and refined our surveys for the implementation phase based on testing of our data collection instruments. Our partner sites are in various stages of the single institutional review board (IRB) approval process. We have ongoing engagement with all partner sites, which has helped solidify our participant recruitment strategies and design patient-friendly recruitment materials. In addition, we have included a patient advocate on the research team. Members of the research team have launched a single IRB Support Network at WUSM to create a repository of the single IRB procedures in order to streamline the partner site onboarding process and facilitate enhanced collaboration across institutions. CONCLUSIONS: With this robust model, we expect that patients with MBC will receive optimal care regardless of geographical location and the model will improve patient and provider experiences when navigating the health system. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/35736.

Infrastructure, programs, and policies to increase bicycling: an international review.

OBJECTIVES: To assess existing research on the effects of various interventions on levels of bicycling. Interventions include infrastructure (e.g., bike lanes and parking), integration with public transport, education and marketing programs, bicycle access programs, and legal issues. METHODS: A comprehensive search of peer-reviewed and non-reviewed research identified 139 studies. Study methodologies varied considerably in type and quality, with few meeting rigorous standards. Secondary data were gathered for 14 case study cities that adopted multiple interventions. RESULTS: Many studies show positive associations between specific interventions and levels of bicycling. The 14 case studies show that almost all cities adopting comprehensive packages of interventions experienced large increases in the number of bicycle trips and share of people bicycling. CONCLUSIONS: Most of the evidence examined in this review supports the crucial role of public policy in encouraging bicycling. Substantial increases in bicycling require an integrated package of many different, complementary interventions, including infrastructure provision and pro-bicycle programs, supportive land use planning, and restrictions on car use.

Associations between street connectivity and active transportation.

BACKGROUND: Past studies of associations between measures of the built environment, particularly street connectivity, and active transportation (AT) or leisure walking/bicycling have largely failed to account for spatial autocorrelation of connectivity variables and have seldom examined both the propensity for AT and its duration in a coherent fashion. Such efforts could improve our understanding of the spatial and behavioral aspects of AT. We analyzed spatially identified data from Los Angeles and San Diego Counties collected as part of the 2001 California Health Interview Survey. RESULTS: Principal components analysis indicated that ~85% of the variance in nine measures of street connectivity are accounted for by two components representing buffers with short blocks and dense nodes (PRIN1) or buffers with longer blocks that still maintain a grid like structure (PRIN2). PRIN1 and PRIN2 were positively associated with active transportation (AT) after adjustment for diverse demographic and health related variables. Propensity and duration of AT were correlated in both Los Angeles (r = 0.14) and San Diego (r = 0.49) at the zip code level. Multivariate analysis could account for the correlation between the two outcomes.After controlling for demography, measures of the built environment and other factors, no spatial autocorrelation remained for propensity to report AT (i.e., report of AT appeared to be independent among neighborhood residents). However, very localized correlation was evident in duration of AT, particularly in San Diego, where the variance of duration, after accounting for spatial autocorrelation, was 5% smaller within small neighborhoods (approximately 0.01 square latitude/longitude degrees = 0.6 mile diameter) compared to within larger zip code areas. Thus a finer spatial scale of analysis seems to be more appropriate for explaining variation in connectivity and AT. CONCLUSIONS: Joint analysis of the propensity and duration of AT behavior and an explicitly geographic approach can strengthen studies of the built environment and physical activity (PA), specifically AT. More rigorous analytical work on cross-sectional data, such as in the present study, continues to support the need for experimental and longitudinal study designs including the analysis of natural experiments to evaluate the utility of environmental interventions aimed at increasing PA.

Bicycling for transportation and health: the role of infrastructure.

This paper aims to provide insight on whether bicycling for everyday travel can help US adults meet the recommended levels of physical activity and what role public infrastructure may play in encouraging this activity. The study collected data on bicycling behavior from 166 regular cyclists in the Portland, Oregon metropolitan area using global positioning system (GPS) devices. Sixty percent of the cyclists rode for more than 150 minutes per week during the study and nearly all of the bicycling was for utilitarian purposes, not exercise. A disproportionate share of the bicycling occurred on streets with bicycle lanes, separate paths, or bicycle boulevards. The data support the need for well-connected neighborhood streets and a network of bicycle-specific infrastructure to encourage more bicycling among adults. This can be accomplished through comprehensive planning, regulation, and funding.

Microscopic and Molecular Evidence of the First Elasmobranch Adomavirus, the Cause of Skin Disease in a Giant Guitarfish, Rhynchobatus djiddensis.

Only eight families of double-stranded DNA (dsDNA) viruses are known to infect vertebrate animals. During an investigation of papillomatous skin disease in an elasmobranch species, the giant guitarfish (Rhynchobatus djiddensis), a novel virus, distinct from all known viral families in regard to particle size, morphology, genome organization, and helicase phylogeny was discovered. Large inclusion bodies containing 75-nm icosahedral viral particles were present within epithelial cell nuclei in the proliferative skin lesions. Deep metagenomic sequencing revealed a 22-kb circular dsDNA viral genome, tentatively named guitarfish "adomavirus" (GAdoV), with only distant homology to two other fish viruses, Japanese eel endothelial cell-infecting virus (JEECV) and a recently reported marbled eel virus. Phylogenetic analysis of the helicase domain places the guitarfish virus in a novel clade that is equidistant between members of the Papillomaviridae and Polyomaviridae families. Specific PCR, quantitative PCR, and in situ hybridization were used to detect, quantify, and confirm that GAdoV DNA was localized to affected epithelial cell nuclei. Changes in the viral titer, as well as the presence of a hybridization signal, coincided with the progression and then final resolution of gross and microscopic lesions. The results indicate that GAdoV is the causative agent of the proliferative skin lesions.IMPORTANCE Cartilaginous fish, including the sharks and rays, evolved from ancestral fish species at least 400 million years ago. Even though they are the descendants of one of the most ancient vertebrate lineages, reports of viral diseases in these species are rare and poorly documented. Deep sequencing revealed a highly divergent virus, tentatively named guitarfish adomavirus, that is distantly related to known papillomaviruses and polyomaviruses. Out of the eight predicted viral genes, only the helicase could be identified as viral by sequence homology searches (BLAST), exemplifying the difficulties of discovering novel viruses within seas of unidentifiable "dark matter" associated with deep sequencing data. The novel adomavirus represents the first viral genome shown to cause clinical disease in a cartilaginous fish species, the giant guitarfish. Our findings demonstrate that emerging fish viruses are fertile ground to expand our understanding of viral evolution in vertebrates.

Two new species of betatorqueviruses identified in a human melanoma that metastasized to the brain.

The role of viral infections in the etiology of brain cancer remains uncertain. Prior studies mostly focused on transcriptome or viral DNA integrated in tumor cells. To investigate for the presence of viral particles, we performed metagenomics sequencing on viral capsid-protected nucleic acids from 12 primary and 8 metastatic human brain tumors. One brain tumor metastasized from a skin melanoma harbored two new human anellovirus species, Torque teno mini virus Emory1 (TTMV Emory1) and Emory2 (TTMV Emory2), while the remaining 19 samples did not reveal any exogenous viral sequences. Their genomes share 63-67% identity with other TTMVs, and phylogenetic clustering supports their classification within the Betatorquevirus genus. This is the first identification of betatorqueviruses in brain tumors. The viral DNA was in its expected non-integrated circular form, and it is unclear if the viruses contributed to tumor formation. Whether the viruses originated from blood, or the primary skin tumor could not be ascertained. Overall, our results demonstrate the usefulness of viral metagenomics to detect previously unknown exogenous virus in human brain tumors. They further suggest that active viral infections are rare events in brain tumors, but support a follow-up larger scale study to quantify their frequency in different brain tumor subtypes.

Infection with a novel derogenid trematode in a Flap-necked chameleon (Chamaeleo dilepis).

A 2-year-old female Flap-necked chameleon (Chamaeleo dilepis) was presented with a history of anorexia. On physical examination, three 11×4×2 mm, light tan-pink, raised, gelatinous, freely movable, poorly demarcated, raised nodules were identified within the oral cavity. An impression smear contained multiple, light blue to green structures exhibiting an 18×35 µm, elliptical-shaped anterior portion and a single 1×150 µm filament with a tapered end. Initial differential etiologies included trematode ova, flagellate protozoa, fungal sporangia, plant material, and sperm. Fungal culture yielded no growth. The structures described above did not stain with Gomori methenamine silver stain. A calcofluor white stain yielded positive fluorescence. The DNA was extracted from the impression smears and pan-trematode PCR was performed; products were directly sequenced and phylogenetic analysis revealed maximum similarity with a digenean trematode in the family Derogenidae that was distinct from other trematodes for which a reference sequence was available. To our knowledge, this is the first description of cytologic features of trematode ova from an oral lesion in a chameleon.

Complete Sequence of the Smallest Polyomavirus Genome, Giant Guitarfish (Rhynchobatus djiddensis) Polyomavirus 1.

Polyomaviruses are known to infect mammals and birds. Deep sequencing and metagenomic analysis identified the first polyomavirus from a cartilaginous fish, the giant guitarfish (Rhynchobatus djiddensis). Giant guitarfish polyomavirus 1 (GfPyV1) has typical polyomavirus genome organization, but is the smallest polyomavirus genome (3.96 kb) described to date.

Transport and health: a look at three Latin American cities.

Transport is associated with environmental problems, economic losses, health and social inequalities. A number of European and US cities have implemented initiatives to promote multimodal modes of transport. In Latin America changes are occurring in public transport systems and a number of projects aimed at stimulating non-motorized modes of transport (walking and cycling) have already been implemented. Based on articles from peer-reviewed academic journals, this paper examines experiences in Bogotá (Colombia), Curitiba (Brazil), and Santiago (Chile), and identifies how changes to the transport system contribute to encourage active transportation. Bus rapid transit, ciclovias, bike paths/lanes, and car use restriction are initiatives that contribute to promoting active transportation in these cities. Few studies have been carried out on the relationship between transport and physical activity. Car ownership continues to increase. The public health sector needs to be a stronger activist in the transport policy decision-making process to incorporate health issues into the transport agenda in Latin America.

Evaluation of bike boxes at signalized intersections.

This paper presents a before-after study of bike boxes at 10 signalized intersections in Portland, Oregon. The bike boxes, also known as advanced stop lines or advanced stop boxes, were installed to increase visibility of cyclists and reduce conflicts between motor vehicles and cyclists, particularly in potential "right-hook" situations. Before and after video were analyzed for seven intersections with green bike boxes, three intersections with uncolored bike boxes, and two control intersections. User perceptions were measured through surveys of cyclists passing through five of the bike box intersections and of motorists working downtown, where the boxes were concentrated. Both the observations and survey of motorists found a high rate of compliance and understanding of the markings. Overall, 73% of the stopping motor vehicles did not encroach at all into the bike box. Both motor vehicle and bicycle encroachment in the pedestrian crosswalk fell significantly at the bike box locations compared to the control intersections. The bike boxes had mixed effects on the motorists' encroachment in the bicycle lane. The number of observed conflicts at the bike box locations decreased, while the total number of cyclists and motor vehicles turning right increased. Negative-binomial models based upon the data predict fewer conflicts with the boxes, particularly as right-turning motor vehicle volumes increase. Observations of yielding behavior at two bike box and one control intersection found an improvement in motorists yielding to cyclists at the bike box locations. Differences in the traffic volumes and location contexts make firm conclusions about the effects of green coloring of the boxes difficult. Higher shares of surveyed motorists felt that the bike boxes made driving safer rather than more dangerous, even when the sample was narrowed to respondents who were not also cyclists. Over three-quarters of the surveyed cyclists thought that the boxes made the intersection safer.

The role of health and physical activity in the adoption of innovative land use policy: findings from surveys of local governments.

BACKGROUND: Research has established that built environments, including street networks, bicycle and pedestrian infrastructure, and land uses, can positively affect the frequency and duration of daily physical activity. Attention is now being given to policy frameworks such as zoning codes that set the standards and expectations for this built environment. METHODS: We examined the adoption and implementation of mixed-use and related zoning provisions with specific attention to the role that physical activity serves as a motivation for such policies and to what extent public health agencies influence the adoption process. A sample of planning directors from 53 communities with outstanding examples of mixed-use developments and 145 randomly selected midsized communities were surveyed. RESULTS: Physical activity is not a dominant motivator in master plans and/or zoning codes and public health agencies played minor roles in policy adoption. However, physical activity as a motivation appears to be increasing in recent years and is associated with higher levels of policy innovation. CONCLUSIONS: Recommendations include framing the importance of physical activity in terms of other dominant concerns such as livability, dynamic centers, and economic development. Health agencies are encouraged to work in coalitions to focus arguments on behalf of physical activity.