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Extrachromosomal circular DNA (ecDNA) have been found in most types of human cancers, and ecDNA incorporating viral genomes has recently been described, specifically in human papillomavirus (HPV)-mediated oropharyngeal cancer (OPC). However, the molecular mechanisms of human-viral hybrid ecDNA (hybrid ecDNA) for carcinogenesis remains elusive. We characterized the epigenetic status of hybrid ecDNA using HPVOPC cell lines and patient-derived tumor xenografts, identifying HPV oncogenes E6/E7 in hybrid ecDNA were flanked by novel somatic DNA enhancers and HPV L1 enhancers, with strong cis-interaction. Targeting of these enhancers by clustered regularly interspaced short palindromic repeats interference or hybrid ecDNA by bromodomain and extra-terminal inhibitor reduced E6/E7 expression, and significantly inhibited in vitro and/or in vivo growth only in ecDNA(+) models. HPV DNA in hybrid ecDNA structures are associated with novel somatic and HPV enhancers in hybrid ecDNA that drive HPV ongogene expression and carcinogenesis, and can be targeted with ecDNA disrupting therapeutics., Competing Interests: Conflict of interest: P.S.M. is a co-founder, chairs the scientific advisory board (SAB) of and has equity interest in Boundless Bio Inc. (BBI). P.S.M. is also an advisor with equity for Asteroid Therapeutics and is an advisor to Sage Therapeutics. V.B. is a co-founder, consultant, SAB member and has equity interest in Boundless Bio and Abterra. J.T.L is an employee of BBI. J.L. previously consulted for BBI. The other authors have no conflicts of interest to disclose.

Trichinella murrelli Pozio and La Rosa, 2000, is the primary sylvatic trichinellid encountered in temperate North America. During a survey for Sarcocystis in wild canids, a single worm matching the morphology of encapsulated Trichinella was observed in a muscle tissue squash from a gray fox male originating from Pennsylvania. The worm was photographed and then separated from the host tissue by artificial digestion, and genomic DNA was extracted from the worm. This DNA was subjected to species-specific multiplex PCR and short-read genomic sequencing. The banding pattern of the multiplex PCR indicated that the worm was T. murrelli, and the sequence of the mitochondrial Cytochrome c oxidase 1 gene and the ribosomal 18S ribosomal RNA, Internal Transcribed Spacer 1, 5.8S ribosomal RNA, Internal Transcribed Spacer 2, and 28S ribosomal RNA confirmed the diagnosis. This is the first report of T. murrelli in gray foxes that includes assays for assigning parasite species. This report confirms suspected data from surveys conducted over 30 yr ago and establishes a new host record for T. murrelli., (© American Society of Parasitologists 2024.)

Abstract: Sarcocystis infections were found for the first time in the muscles of 3 of 3 gray wolves (Canis lupus) from Minnesota. Two kinds (thin-walled and thick-walled) of sarcocysts were detected, based on the appearance of the sarcocyst wall. In wolf 1, sarcocysts were thin-walled (<0.5 μm), and without any visible protrusions. Ultrastructurally, the sarcocyst wall was type 1a and identical to Sarcocystis svanai of the domestic dog (Canis familiaris). The second kind of sarcocyst, with a relatively thicker (>1 μm) sarcocyst wall, was detected in wolves 2 and 3. Ultrastructurally, the sarcocyst wall had undulating, pleomorphic villar protrusion of type 9c; these sarcocysts were identical to Sarcocystis caninum from the domestic dog. Molecularly, the 2 Sarcocystis species were characterized using 18S, 28S, COI, ITS-1, and rpoB genetic markers. All these markers showed 100% identity to either of the 2 species previously described from the domestic dog. The thick-walled sarococyst corresponded to Sarcocystis caninum, whereas the thin-walled sarcocyst corresponded to Sarcocystis svanai., (© American Society of Parasitologists 2024.)

Abstract: Hepatozoon sp. stages were detected in histological sections of the muscles of 11 of 25 bobcats (Lynx rufus). Parasites were detected from the myocardium of 11, tongues of 4, and limb muscles of 5 of these animals, which had been hunted in Mississippi in 2017. The intensity of infection was highest in the heart. Only asexual stages (meronts) were detected. Three types of meronts (types 1, 2, and 3) were detected, based on structure and mode of division. Additionally, individual zoites were found in leukocytes in the blood vessels of the myocardium, but the stage was not identified. Based on genomic DNA characterized from paraffin-embedded myocardium sections from 1 bobcat using 18S rRNA gene, the Hepatozoon species from the bobcat was related to but distinct from other Hepatozoon spp. of felids. This is the first description of the development of Hepatozoon in the muscles of bobcats. A new name, Hepatozoon rufi, is proposed for this parasite in bobcats., (© American Society of Parasitologists 2024.)

Irrigation water contaminated by human fecal material may elevate the risk of produce contamination with the enteric parasite Cyclospora cayetanensis. Oocysts of C. cayetanensis are resistant to commonly used disinfectants and a method of removing C. cayetanensis from irrigation water would mitigate this risk. We evaluated zero valent iron (ZVI) sand filtration as one such method. We sought to determine if sand filters containing ZVI outperformed those without ZVI. We first evaluated the abundant poultry parasites Eimeria maxima , E. tenella and E. acervulina as surrogates for C. cayetanensis . We determined if a miniaturized gravity fed ZVI-sand filter, scaled to evaluate scarce supplies of C. cayetanensis oocysts, provided useful information about the performance of larger filtration systems. Filters were inoculated with oocysts, rinsed, and the resulting filtrate examined microscopically for oocysts. We performed experiments to measure the effect of varying ZVI concentrations, repeated filter use, simulated agricultural water, and oocyst size and condition. We then compared the performance of mini filters to that of larger, gravity-fed pool filters and found that ZVI-sand filtration was far more effective at removing Eimeria spp. from water when compared to sand filtration, at both scales. Sand mini filters retained 13-54 % of E. acervulina oocysts, and pool filters retained 82 %, but when combined with 50 % (mini filter) or 35 % (pool filter) v /v ZVI, mini filters retained 89-99 % of oocysts and pool filters retained >99 %. The effectiveness of the mini filters increased with increasing ZVI concentration, and the addition of ZVI far outweighed the influence of any other measured variable. We then performed experiments including C. cayetanensis , which provided similar results to those utilizing Eimeria ; 59 % of inoculated C. cayetanensis oocysts were retained in sand mini filters, and 97 % in mini filters composed of 35 % v /v ZVI. In sum, ZVI is highly effective in removing oocysts from water and Eimeria is a useful surrogate for C. cayetanensis to assess filtration. ZVI-sand filtration shows promise as a tool to mitigate the risk of C. cayetanensis contamination of irrigation water. Further studies should evaluate the performance of ZVI-sand in pressurized fast filtration systems under a range of field conditions., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Benjamin Rosenthal reports financial support was provided by Center For Produce Safety. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Muscles of 25 bobcats (Lynx rufus) from remote areas of Mississippi in 2017 were tested for parasites. Testing for Sarcocystis infections included microscopic examination of fresh unstained muscle squashes, pepsin digestion of hearts and tongues, and histological sections of paraffin-embedded tissues. Sarcocystis spp. infections were detected in the muscles of 21 (84%) by a combination of methods. Sarcocysts were detected in the unstained tongue squashes of 2 bobcats. Sarcocystis sp. bradyzoites were detected in the pepsin digests of 3 of 19 hearts, and 12 of 19 tongues. In paraffin-embedded histological sections, sarcocysts were detected in 7 of 25 hearts, 17 of 25 tongues, and 5 of 23 limb muscles. Based on the character of the cyst wall, at least 3 morphologic types of sarcocysts were detected: those with small spikes on the cyst wall, corresponding to Sarcocystis felis, those with long villar protrusions, corresponding to Sarcocystis neurona, and those lacking visible cyst wall protrusions, representing an unidentified type of sarcocyst. Myositis associated with sarcocysts was seen in the tongues of 3, and in the limb muscles of 1 bobcat. Multilocus genotyping of the DNA extracted from paraffin-embedded sections from 2 bobcats, employing 18S, 28S, COI, ITS-1, and 5.8S and rpoB genes, diagnosed Sarcocystis caninum, S. felis, Sarcocystis lutrae, and S. neurona. An encapsulated species of Trichinella was identified in the tongue of 1; it represents the first documented occurrences in bobcats from Mississippi. Taken together, these observations suggest intensive exposure of these wild carnivores to Trichinella tissue cysts, implies predation or scavenging on these tissues promotes parasite transmission, and raises caution concerning zoonotic risk when such meat is rendered for human consumption., (© American Society of Parasitologists 2024.)

Phylogenetic evidence indicates that free-living nematodes gave rise to parasitic nematodes where parasitism evolved independently at least 15 times. The high level of genetic and biological diversity among parasites dictates an equally high level of diversity in the transition to parasitism. We previously hypothesized that horizontal gene transfer (HGT) played an important role in the evolution of parasitism among early ancestors of Trichinella, mediated by an interplay of ecological and evolutionary pathways that contributed to persistence and diversification. We propose that host selection may have been associated with the metabolism of ammonia and engender a new paradigm whereby the reprogrammed nurse cell is capable of generating cyanate thereby enabling the importance of the Trichinella cyanase in the longevity of the cell. Parasites and parasitism have revealed considerable resilience against a backdrop of climate change and environmental perturbation. Here we provide a putative link between key periods in the evolution of Trichinella and major geological and climatological events dating back 500 million years. A useful lens for exploring such ideas, the Stockholm Paradigm, integrates Ecological Fitting (a foundation for host colonization and diversification), the Oscillation Hypothesis (recurring shifts between trends in generalization and specialization relative to host range), the Geographic Mosaic Theory of Coevolution (microevolutionary co-adaptive processes), and the Taxon Pulse Hypothesis (alternating events of biotic expansion i.e., exploitation in evolutionary and ecological time). Here we examine how one or more of these interactive theories, in a phylogenetic-historical context and in conjunction with HGT, may help explain the scope and depth of diversity among Trichinella genotypes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

Background: Overcompression of the distal tibiofibular syndesmosis during open reduction and internal fixation of ankle fracture may affect multidirectional flexibility of the ankle bones., Methods: Ten cadaveric lower limbs (78.3±13.0 years, 4 female, 6 male) underwent biomechanical testing in sagittal, coronal, and axial rotation with degrees of motion quantified. The intact force (100%) was the force needed to compress the syndesmosis just beyond the intact position, and overcompression was defined as 150% of the intact force. After intact testing, the anterior inferior tibiofibular ligament (AITFL), interosseus membrane (IOM), and posterior inferior tibiofibular ligament (PITFL) were sectioned and testing was repeated. The IOM and AITFL were reconstructed in sequence and tested at 100% and 150% compression., Results: Overcompression of the syndesmosis did not significantly reduce ROM of the ankle bones for any loading modality ( P  > .05). IOM+AITFL reconstruction restored distal tibiofibular axial rotation to the intact condition. Axial rotation motion was significantly lower with AITFL fixation compared with IOM fixation alone ( P  < .05). The proximal tibiofibular syndesmosis demonstrated significantly higher motion in axial rotation with all distal reconstruction conditions., Conclusion: As assessed by direct visualization, overcompression of the distal tibiofibular syndesmosis did not reduce ROM of the ankle bones. Distal tibiofibular axial rotation was significantly lower with IOM+AITFL fixation compared with IOM augmentation alone. Distal tibiofibular axial rotation did not differ significantly from the intact condition after combined IOM+AITFL fixation. Dynamic fixation of the distal tibiofibular syndesmosis resulted in increased axial rotation at the proximal tibiofibular syndesmosis., Clinical Relevance: These biomechanical data suggest that inadvertent overcompression of the distal tibiofibular syndesmosis when fixing ankle fractures does not restrict subsequent ankle bone ROM. The AITFL is an important stabilizer of the distal tibiofibular syndesmosis in external rotation., Level of Evidence: controlled laboratory study., Competing Interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. Disclosure forms for all authors are available online., (© The Author(s) 2024.)

Ozone is a potent disinfecting agent used to treat potable water and wastewater, effectively clearing protozoa such as Giardia and Cryptosporidium spp. It is unclear whether ozone treatment of water or fresh produce can reduce the spread of the emerging parasite Cyclospora cayetanensis, which causes cyclosporiasis in humans. Obtaining viable C. cayetanensis oocysts to evaluate inactivation methods is challenging because we lack the means to propagate them in vitro, because of delays in case reporting, and because health departments typically add inactivating fixatives to clinical specimens. Research in various surrogate organisms has sought to bolster understanding of the biology of C. cayetanensis. Among these surrogates is the poultry parasite Eimeria acervulina, a closely related and easily cultured parasite of economic significance. We used this surrogate to evaluate the consequences of ozone treatment, using the sporulation state as an indicator of infectious potential. Treating with ozonated water acidified with citric acid reduced sporulation ability in a dose-dependent manner; treatment with up to 4.93 mg/L initial concentration of ozone resulted in a 93% inactivation of sporulation by 7 days posttreatment. This developmental arrest was accompanied by transcriptional changes in genes involved in regulating the response to reactive oxygen species (ROS) in a time course that is consistent with the production of oxygen free radicals. This study shows that ozone is highly effective in preventing sporulation of E. acervulina, a model coccidian used as a surrogate for Cyclospora. Furthermore, ozone exposure induced molecular responses to general oxidative stress, documented with several well-characterized antioxidant enzymes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Purpose: Biosimilar tumor necrosis factor inhibitors (b-TNFi) reduce healthcare costs and maintain equal efficacy when compared to their originator counterparts (o-TNFi). Current practice is to start patients on a slower standard infusion rate during the initial transition from an o-TNFi to a b-TNFi. There is a knowledge gap around switching from rapid originator infusion to rapid biosimilar infusion in the pediatric inflammatory bowel disease (IBD) population., Summary: We present a case series of 8 pediatric patients with IBD who were switched from a rapid-infusion o-TNFi to a rapid-infusion b-TNFi from 2016 through 2022. Our primary interest was safety, which we evaluated based on the occurrence of infusion reactions or need for new premedications within the first 6 months of starting a b-TNFi. We also examined effectiveness through the incidence of IBD-related hospitalizations, TNFi failure, and need for co-medication or dose escalation over the same period. In our cohort, 4 patients had Crohn's disease and 4 had ulcerative colitis. All patients were switched to a biosimilar for nonmedical reasons. During the follow-up period, no patients had infusion reactions necessitating new premedications, serious adverse events, or medication nonresponse., Conclusion: Patients who directly transitioned from a rapid-infusion o-TNFi to a rapid-infusion b-TNFi did not experience serious adverse events. Given the fiscal and patient experience advantages of rapid-rate infusions, larger studies are needed to consider a change in practice., (© American Society of Health-System Pharmacists 2023. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Infections by Sarcocystis in cattle are ubiquitous worldwide. There is considerable debate concerning the identity of Sarcocystis spp. in cattle. Proper diagnosis of Sarcocystis spp. is important to assess their economic and public health importance. Currently there are seven named species: Sarcocystis hirsuta, Sarcocystis cruzi, Sarcocystis hominis, Sarcocystis bovifelis, arcocystis heydorni, Sarcocystis bovini and Sarcocystis rommeli. Additionally, there are unnamed Sarcocystis spp. Two species, S. hominis and S. heydorni, are zoonotic. One out of seven species (S. hirsuta, contracted from cats) forms macroscopic cysts which can be visible during carcass inspection. Current molecular characterization is based on DNA extracted from sarcocysts from naturally infected cattle because DNA was not characterized from tissues of experimentally infected cattle or feces of experimentally infected definitive hosts. Sarcocystis cruzi (transmitted via canids) is recognized as the most pathogenic species and it causes abortion, low milk yield, poor body growth, and outbreaks of clinical sarcocystosis and death. Additionally, Sarcocystis infections have been linked to an inflammatory condition of striated muscles termed bovine eosinophilic myositis (BEM). Cattle affected by BEM appear clinically normal. Diagnosis of BEM at slaughter occurs when inspecting the carcass surface, or once the carcass has been divided into prime cuts or quarters. Sex and breed have no apparent influence on prevalence of BEM. The condition evidently occurs with equal frequency in steers, cows, and heifers. Virtually all striated muscles can be affected including skeletal muscles, the muscles of the eye, larynx, and the heart. In the USA, regulations require condemnation of BEM-affected parts, or (in severe cases) the entire carcass. These aesthetic considerations result in economic losses. Cattle experimentally infected with Sarcocystis did not have BEM at slaughter. Here, we review the status of Sarcocystis spp. and BEM in cattle including prevalence, lesions, epidemiology, and association of BEM with different species of Sarcocystis., (Published by Elsevier Ltd.)

Antiestrogens prescribed to reduce breast cancer risk or recurrence can have undesirable musculoskeletal side effects that may lead to early discontinuation of therapy. Previous studies have not focused on nurse-led assessmen.

Molecular epidemiology using traditional sequencing has been notoriously difficult in inbred parasites due to a lack of genetic variation available for discriminating among parasites. Next generation sequencing techniques offer a solution to this problem by increasing the number of loci that can be sequenced. Here, we introduce Trich-tracker, a tool that makes efficient use of diagnostic variation distributed throughout the genome of Trichinella spiralis to more rapidly, and conclusively, resolve connections and distinctions among focal outbreaks of T. spiralis. In particular, we rapidly characterised genetic variation among a sample of parasites from Polish farms and wildlife, sampling genomic variation using double digest restriction site-associated DNA sequencing (ddRADseq). Approximately 400,000 bases of sequence were generated from each sample and shown to be distributed across the genome with single nucleotide polymorphisms occurring at a frequency of approximately one base in 10,000. Both phylogenetic and Bayesian clustering analyses indicated that ddRADseq genotypes formed distinct clusters for specific outbreaks and were quite distinct from wild boar samples. Two of the investigated outbreaks were more similar to each other than to other outbreak samples, suggesting a link between these outbreaks. Hence, the Trich-tracker procedure identified informative genomic variation which afforded unprecedented epidemiological resolution. Trich-tracker is very flexible tool, quickly and inexpensively mining genomes of even highly inbred populations of T. spiralis to support outbreak investigations. The simplicity of the entire procedure, and time and cost effectiveness of Trich-tracker support its practical application in ongoing Trichinella outbreaks. The discriminating power of this tool is tunable and scalable, allowing application in a variety of epidemiological contexts, and is easily adapted to other parasite systems., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Purpose: A metastatic cancer diagnosis is associated with high levels of distress in patients and caregivers, which may be alleviated by mindfulness interventions. Research on scalable, tailored, online mindfulness training programs is needed. We sought to test the feasibility and acceptability of a remotely delivered 8-week mindfulness-based intervention, Being Present 2.0 (BP2.0)., Methods: We performed a single-arm feasibility study of BP2.0 among patients with any metastatic gastrointestinal cancer receiving chemotherapy, with or without an informal caregiver. Participants were instructed to practice mindfulness using pre-recorded guided meditations 5 times per week using a study-specific website and to attend a weekly live, interactive virtual meeting facilitated by a trained instructor. The web-based platform enabled direct measurement of adherence., Results: The study enrolled 46 of 74 (62%) patients contacted, together with 23 caregivers (69 participants total), from May to October 2018. Median patient age was 52 (range 20-70 years), 39% were male, 67% non-Hispanic white, 65% had colorectal cancer, and 78% lived outside of San Francisco. The top reasons cited for participation were to reduce stress/anxiety and learn how to meditate. Mean baseline National Comprehensive Cancer Network Distress Thermometer (NCCN DT) scores were 4.7 (patients) and 5.8 (caregivers). The study discontinuation rate was 20% (eight patients and six caregivers). Among the remaining 55 participants, 43 (78%) listened to at least one audio recording and/or attended at least one virtual meeting, although adherence data was incomplete. The retention rate was 71%, with 39 participants completing at least one follow-up assessment. In post-intervention qualitative interviews, 88% of respondents reported a positive experience. Compared to baseline, participants reported significantly reduced post-intervention NCCN DT scores (mean 3.1; P = .012)., Conclusion: The BP2.0 online mindfulness-based program is feasible and acceptable for patients with metastatic gastrointestinal cancer and caregivers. These results will guide plans for a follow-up efficacy study. ClinicalTrials.gov Identifier: NCT03528863., Competing Interests: Declaration of conflicting interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (© The Author(s) 2021.)

Genetic variation in pathogen populations provides the means to answer questions in disease ecology and transmission, illuminating interactions between genetic traits, environmental exposures, and disease. Such studies elucidate the phylogeny, evolution, transmission and pathogenesis of viruses, bacteria and parasites. Here, we review how such studies have fostered understanding of the biology and epidemiology of zoonotic nematode parasites in the genus Trichinella spp., which impose considerable economic and health burdens by infecting wildlife, livestock, and people. To use such data to define ongoing chains of local transmission and source traceback, researchers first must understand the extent and distribution of genetic variation resident in regional parasite populations. Thus, genetic variability illuminates a population's past as well as its present. Here we review how such data have helped define population dynamics of Trichinella spp. in wild and domesticated hosts, creating opportunities to harness genetic variation in the quest to prevent, track, and contain future outbreaks., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

While global cases of trichinellosis have fallen since pork regulation began, the disease remains a danger to pork and animal game consumers as well as a liability to producers. Managing food safety risk and supporting agricultural trade requires cost-effective and sensitive diagnostic methods. Several means exist to inspect pork for parasitic infections. Here, we review literature concerning the sensitivity, specificity, and cost of these methods. We found that artificial digestion coupled with optical microscopy to be the best method for verification of Trichinella larva free pork due to its cost efficiency, high specificity, and reliability. Serological techniques such as ELISA are useful for epidemiological surveillance of swine. While current PCR techniques are quick and useful for diagnosing species-specific infections, they are not cost efficient for large-scale testing. However, as PCR techniques, including Lateral Flow- Recombinase Polymerase Amplification (LF-RPA), improve and continue to reduce cost, such methods may ultimately succeed artificial digestion., Competing Interests: All authors declare no competing interests in the content of this review. The work was funded from public sources and performed in the public interest without regard for commercial interests of any kind, and with the sole desire to support science-based decision making and to advance public health., (Published by Elsevier Inc. on behalf of International Association of Food and Waterborne Parasitology.)

Background: Species of Trichinella are globally important foodborne parasites infecting a number of domestic and wild vertebrates, including humans. Free-ranging carnivores can act as sentinel species for detection of Trichinella spp. Knowledge of the epidemiology of these parasites may help prevent Trichinella spp. infections in northern Canadian animals and people. Previous research on Trichinella spp. in wildlife from Yukon did not identify risk factors associated with infection, or the diversity and identity of species of Trichinella in regional circulation, based on geographically extensive sampling with large sample sizes., Methods: In a cross-sectional study, we determined the prevalence, infection intensity, risk factors, and species or genotypes of Trichinella in wolverine (Gulo gulo) in two regions of Yukon, Canada, from 2013-2017. A double separatory funnel digestion method followed by mutiplex PCR and PCR-RFLP were used to recover and identify species of Trichinella, respectively., Results: We found larvae of Trichinella in the tongues of 78% (95% CI 73-82) of 338 wolverine sampled. The odds of adult (≥ 2 years) and yearling (1 year) wolverine being Trichinella spp.-positive were four and two times higher, respectively, compared to juveniles (<1 year). The odds of Trichinella spp. presence were three times higher in wolverine from southeast than northwest Yukon. The mean intensity of infection was 22.6 ± 39 (SD, range 0.1-295) larvae per gram. Trichinella T6 was the predominant genotype (76%), followed by T. nativa (8%); mixed infections with Trichinella T6 and T. nativa (12%) were observed. In addition, T. spiralis was detected in one wolverine. Out of 22 isolates initially identified as T. nativa in multiplex PCR, 14 were analyzed by PCR-RFLP to distinguish them from T. chanchalensis, a recently discovered cryptic species, which cannot be distinguished from the T. nativa on multiplex PCR. Ten isolates were identified either as T. chanchalensis alone (n = 7), or mixed infection with T. chanchalensis and T. nativa (n = 2) or T. chanchalensis and Trichinella T6 (n = 1)]., Conclusions: Wolverine hosted high prevalence, high larval intensity, and multiple species of Trichinella, likely due to their scavenging habits, apex position in the food chain, and wide home range. Wolverine (especially adult males) should be considered as a sentinel species for surveys for Trichinella spp. across their distributional range.

Objective: Reconstruction of the mobile spine following total en bloc spondylectomy (TES) of one or multiple vertebral bodies in patients with malignant spinal tumors is a challenging procedure with high failure rates. A common reason for reconstructive failure is nonunion, which becomes more problematic when using local radiation therapy. Radiotherapy is an integral part of the management of primary malignant osseous tumors in the spine. Vascularized grafts may help prevent nonunion in the radiotherapy setting. The authors have utilized free vascularized fibular grafts (FVFGs) for reconstruction of the spine following TES. The purpose of this article is to describe the surgical technique for vascularized reconstruction of defects after TES. Additionally, the outcomes of consecutive cases treated with this technique are reported., Methods: Thirty-nine patients were treated at the authors' tertiary care institution for malignant tumors in the mobile spine using FVFG following TES between 2010 and 2018. Postoperative union, reoperations, complications, neurological outcome, and survival were reported. The median follow-up duration was 50 months (range 14-109 months)., Results: The cohort consisted of 26 males (67%), and the median age was 58 years. Chordoma was the most prevalent tumor (67%), and the lumbar spine was most affected (46%). Complete union was seen in 26 patients (76%), the overall complication rate was 54%, and implant failure was the most common complication, with 13 patients (33%) affected. In 18 patients (46%), one or more reoperations were needed, and the fixation was surgically revised 15 times (42% of reoperations) in 10 patients (26%). A reconstruction below the L1 vertebra had a higher proportion of implant failure (67%; 8 of 12 patients) compared with higher resections (21%; 5 of 24 patients) (p = 0.011). Graft length, number of resected vertebrae, and docking the FVFG on the endplate or cancellous bone was not associated with union or implant failure on univariate analysis., Conclusions: The FVFG is an effective reconstruction technique, particularly in the cervicothoracic spine. However, high implant failure rates in the lumbar spine have been seen, which occurred even in cases in which the graft completely healed. Methods to increase the weight-bearing capacity of the graft in the lumbar spine should be considered in these reconstructions. Overall, the rates of failure and revision surgery for FVFG compare with previous reports on reconstruction after TES.

In less than five months, COVID-19 has spread from a small focus in Wuhan, China, to more than 5 million people in almost every country in the world, dominating the concern of most governments and public health systems. The social and political distresses caused by this epidemic will certainly impact our world for a long time to come. Here, we synthesize lessons from a range of scientific perspectives rooted in epidemiology, virology, genetics, ecology and evolutionary biology so as to provide perspective on how this pandemic started, how it is developing, and how best we can stop it., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Cognitive impairments constitute a core feature of schizophrenia, and a genetic overlap between schizophrenia and cognitive functioning in healthy individuals has been identified. However, due to the high polygenicity and complex genetic architecture of both traits, overlapping biological pathways have not yet been identified between schizophrenia and normal cognitive ability. Network medicine offers a framework to study underlying biological pathways through protein-protein interactions among risk genes. Here, established network-based methods were used to characterize the biological relatedness of schizophrenia and cognition by examining the genetic link between schizophrenia risk genes and genes associated with cognitive performance in healthy individuals, through the protein interactome. First, network separation showed a profound interactome overlap between schizophrenia risk genes and genes associated with cognitive performance (S AB  = -0.22, z-score = -6.80, p = 5.38e-12). To characterize this overlap, network propagation was thereafter used to identify schizophrenia risk genes that are close to cognition-associated genes in the interactome network space (n = 140, of which 54 were part of the direct genetic overlap). Schizophrenia risk genes close to cognition were enriched for pathways including long-term potentiation and Alzheimer's disease, and included genes with a role in neurotransmitter systems important for cognitive functioning, such as glutamate and dopamine. These results pinpoint a subset of schizophrenia risk genes that are of particular interest for further examination in schizophrenia patient groups, of which some are druggable genes with potential as candidate targets for cognitive enhancing drugs., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to declare., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

To assess the role of white-tailed deer (Odocoileus virginianus, WTD) in the epidemiology of toxoplasmosis, we conducted a national survey of WTD across the USA for Toxoplasma gondii infection. To do this, we combined serology with parasite isolation to evaluate the prevalence and genetic diversity of T. gondii in this game species. From October 2012 to March 2019, serum and tissues were collected from 914 WTD across the USA. Serum samples were screened for antibodies to T. gondii, and then the tissues of seropositive WTD were bioassayed in mice. Antibodies were detected in 329 (36%) of 914 WTD tested by the modified agglutination test (positive reaction at 1:25 or higher). Viable T. gondii was isolated from the heart of 36 WTD from 11 states. Three of the 36 isolates were pathogenic but not highly virulent to outbred Swiss Webster mice and all 36 isolates could be propagated further in cell culture and were genotyped. For genotyping, DNA extracted from cell culture-derived tachyzoites was characterized by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using the genetic markers SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico. Genotyping revealed seven ToxoDB PCR-RFLP genotypes, including 24 isolates for genotype #5 (haplogroup 12), four isolates for #2 (type III, haplogroup 3), three isolates for genotypes #1 (type II, haplogroup 2), two isolates for genotypes #3 (type II, haplogroup 2) and one isolate each for #39, #221 and #224. Genotype #5 was the most frequently isolated, accounting for 66.6% (24 of 36) of the isolates. Combining the 36 isolates from this study with previously reported 69 isolates from WTD, 15 genotypes have been identified. Among these, 50.4% (53/105) isolates belong to genotype #5. Our results indicate moderate genetic diversity of T. gondii in WTD. The results also indicate that undercooked venison should not be consumed by humans or fed to cats.

Feral swine are known reservoirs of various pathogens, including Toxoplasma gondii. Here, we report the first national survey of viable T. gondii in feral swine in the USA. We paired serological surveys with parasite isolation and bioassay to evaluate the prevalence and genetic diversity of these parasites. From 2012-2017, sera and tissues from 1517 feral swine across the USA were collected for the isolation of viable T. gondii. Serum samples were initially screened for antibodies to T. gondii, and then the tissues of seropositive feral swine were bioassayed in mice. Antibodies were detected in 27.7% of feral swine tested by the modified agglutination test (1:25 or higher). Antibody positive rates increased significantly with age, with 10.1% of juveniles, 16.0% of sub-adults and 38.4% of adults testing seropositive. Myocardium (50 g) from 232 seropositive feral swine was digested in pepsin and bioassayed in mice. Viable T. gondii was isolated from 78 feral swine from 21 states. Twelve of the 78 isolates were pathogenic to outbred Swiss Webster mice and 76 of the 78 isolates could be propagated further in cell culture and were genotyped. For genotyping, deoxyribonucleic acid extracted from cell culture-derived tachyzoites was characterized by polymerase chain reaction restriction fragment length polymorphism using the genetic markers SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico. Genotyping revealed 15 ToxoDB genotypes, including 43 isolates for genotype #5 (haplogroup 12), 11 isolates for #24, four isolates for #2 (haplogroup 3), two isolates for each of genotypes #3 (haplogroup 2), #4 (haplogroup 12), #216, #221, #289 and #297 and one isolate for each of genotypes #1 (haplogroup 2), #39, #66, #260, #261 and #299. Genotype #5 was the most frequently isolated, accounted for 57% (43/76) of the isolates, followed by #24, accounted for 14% (11/76). Genotypes #260, #289, #297 and #299 are new types. Genotype #289 was highly virulent to mice and originated from feral swine collected in Louisiana on the same day at the same location. Genotype #216 was previously demonstrated to be highly virulent to mice. Our results indicate moderate genetic diversity of T. gondii in feral swine in the USA, with the genotype #5 (haplogroup 12) dominant in the continental USA, whereas genotype #24 (10/14) was dominant in Hawaii, suggesting different population structures of the parasites among the two distinct geographical locations.

Besnoitia darlingi, B. neotomofelis and B. oryctofelisi are closely related coccidian parasites with cats as definitive hosts. While B. darlingi uses opossums as intermediate hosts, B. neotomofelis and B. oryctofelisi have been described in Southern Plains woodrats ( Neotoma micropus ) from the USA and in domestic rabbits from Argentina, respectively. A comparison of the Internal Transcribed Spacer-1 (ITS-1) region of the ribosomal DNA (rDNA) of these Besnoitia spp. showed only a few differences. The present study aimed at developing a real-time PCR to detect B. darlingi, B. neotomofelis and B. oryctofelisi in tissues of intermediate and in faeces of definitive hosts in order to support studies of these organisms' epidemiology and pathogenesis. The established PCR was based on primer regions distinct from the ITS-1 sequences of ungulate Besnoitia spp. and made use of a Besnoitia universal probe. To monitor inhibition, a heterologous internal control was established based on the enhanced green fluorescent protein gene. The real-time PCR reacted with B. darlingi , B. neotomofelis and B. oryctofelisi, while the novel PCR did not recognize ungulate Besnoitia spp. ( B. besnoiti, B. bennetti, B. tarandi ). DNA of Apicomplexa ascribed to other Besnoitia-related genera, including other gut parasites of cats ( Cryptosporidium parvum , Giardia duodenalis , Tritrichomonas foetus ), was not recognized. The real-time PCR had an analytic sensitivity of less than 1 tachyzoite per reaction. In feline faeces spiked with B. darlingi oocysts, the limit of detection was a DNA amount equivalent to 1 oocyst per PCR reaction. In B. darlingi infected ɣ-interferon knock-out mice, the lung was identified as the predilection organ. In conclusion, this real-time PCR should advance further studies on these parasites and may inspire research on related species, not only in the Americas, but also in other parts of the world., Competing Interests: The study described is original and is not under consideration by any other journal. All authors approved the final manuscript and its submission. The authors declare that they have no conflict of interest., (© 2020 The Author(s).)

Objective: The study sought to develop and evaluate an electronic health record-based child abuse clinical decision support system in 2 general emergency departments., Materials and Methods: A combination of a child abuse screen, natural language processing, physician orders, and discharge diagnoses were used to identify children <2 years of age with injuries suspicious for physical abuse. Providers received an alert and were referred to a physical abuse order set whenever a child triggered the system. Physician compliance with clinical guidelines was compared before and during the intervention., Results: A total of 242 children triggered the system, 86 during the preintervention and 156 during the intervention. The number of children identified with suspicious injuries increased 4-fold during the intervention (P < .001). Compliance was 70% (7 of 10) in the preintervention period vs 50% (22 of 44) in the intervention, a change that was not statistically different (P = .55). Fifty-two percent of providers said that receiving the alert changed their clinical decision making. There was no relationship between compliance and provider or patient demographics., Conclusions: A multifaceted child abuse clinical decision support system resulted in a marked increase in the number of young children identified as having injuries suspicious for physical abuse in 2 general emergency departments. Compliance with published guidelines did not change; we hypothesize that this is related to the increased number of children identified with suspicious, but less serious injuries. These injuries were likely missed preintervention. Tracking compliance with guidelines over time will be important to assess whether compliance increases as physician comfort with evaluation of suspected physical abuse in young children improves., (© The Author(s) 2019. Published by Oxford University Press on behalf of the American Medical Informatics Association. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Species of Trichinella are a globally distributed assemblage of nematodes, often with distinct host ranges, which include people, domestic, and wild animals. Trichinella spp. are important in northern Canada, where dietary habits of people and methods of meat preparation (drying, smoking, fermenting as well as raw) increase the risk posed by these foodborne zoonotic parasites. Outbreaks in the arctic and subarctic regions of Canada and the United States are generally attributed to T. nativa (T2) or the T6 genotype, when genetic characterization is performed. We report the discovery of Trichinella pseudospiralis (T4), a non-encapsulated species, in a wolverine ( Gulo gulo ) from the Northwest Territories of Canada. This parasite has been previously reported elsewhere from both mammals and carnivorous birds, but our findings represent new host and geographic records for T. pseudospiralis . Multiplex PCR and sequencing of fragments of Cytochrome Oxidase Subunit I (COI) and D3 rDNA confirmed the identification. Phylogenetically, Canadian isolates linked with each other and others derived from Palearctic or Neotropical regions, but not elsewhere in the Nearctic (continental USA). We suggest that migratory birds might have played a role in the dispersal of this pathogen 1000's of km to northwestern Canada. Wolverines are not typically consumed by humans, and thus should not pose a direct food safety risk for trichinellosis. However, the current finding suggests that they may serve as an indicator of a broader distribution for T. pseudospiralis . Along with infection risk already recognized for T. nativa and Trichinella T6, our observations emphasize the need for further studies using molecular diagnostics and alternative methods to clarify if this is a solitary case, or if T. pseudospiralis and other freeze susceptible species of Trichinella (such as T. spiralis ) circulate more broadly in wildlife in Canada, and elsewhere.

As massage therapy experiences a resurgence of use for hospitalized patients, it is appropriate to consider the competencies needed by practitioners to practice safely and effectively in the inpatient setting. Hospitals differ vastly from other massage practice locations such as private offices, spas, and sports clubs. The variety of conditions encountered in an acute care setting require the knowledge and ability to adapt massage protocols appropriately. The Academic Collaborative for Integrative Health (ACIH) created the Hospital Based Massage Therapy (HBMT) Task Force to determine if there is a need for HBMT specific competencies and then, if needed, to develop peer reviewed competencies that hospital staff, massage therapy educators, and massage therapists all may find useful. The members of the task force identified massage therapists who worked in hospitals generally, as well as in hospitals known to have HBMT programs. A spreadsheet was created listing the individuals and a survey was distributed to those on the spreadsheet. These individuals were also asked to identify others who might be interested in participating in the project. The purpose of the survey was to assess various elements of HBMT programs such as educational/experience requirements, employment model, orientation, and supervision. 32 out of 37 hospitals (87%) completed the survey. The Task Force considered the high response rate and the extent to which respondents provided in-depth answers to the open-ended questions as evidence of the need for specific competencies for safe and effective massage therapy for hospitalized patients. In addition to the survey, the task force used a Delphi technique to engage survey participants and other experts in the field to shape the initial draft of the competencies. As these competencies are shared with hospitals, massage therapists, and massage educators, the Task Force members expect that additional development of the competencies will take place as various groups implement them., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The pancreatic islets of Langerhans regulate glucose homeostasis. The loss of insulin-producing β cells within islets results in diabetes, and islet transplantation from cadaveric donors can cure the disease. In vitro production of whole islets, not just β cells, will benefit from a better understanding of endocrine differentiation and islet morphogenesis. We used single-cell mRNA sequencing to obtain a detailed description of pancreatic islet development. Contrary to the prevailing dogma, we find islet morphology and endocrine differentiation to be directly related. As endocrine progenitors differentiate, they migrate in cohesion and form bud-like islet precursors, or "peninsulas" (literally "almost islands"). α cells, the first to develop, constitute the peninsular outer layer, and β cells form later, beneath them. This spatiotemporal collinearity leads to the typical core-mantle architecture of the mature, spherical islet. Finally, we induce peninsula-like structures in differentiating human embryonic stem cells, laying the ground for the generation of entire islets in vitro., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: Routine child abuse screening is an approach to early identification of abuse. Previous studies evaluated paper-based screens; the widespread use of electronic health records suggests that screening is more likely to succeed if integrated into the electronic record., Objective: To implement an electronic health record-based child abuse screen in a diverse hospital system and to evaluate the screening rate, rate of positive screens, and number of reports to Child Protective Services and assess whether hospital and patient characteristics are associated with these rates., Participants and Setting: Children <13 years of age evaluated at one of 13 Emergency Departments within University of Pittsburgh Medical Center Health System., Methods: A previously validated child abuse screen was slightly modified and integrated into Cerner. Multivariable logistic regression models were used to estimate the odds of the outcomes of interest, controlling for key covariates., Results: Of 17,163 eligible children: 68% received the screen of which 1.9% were positive. The rate of reports to Child Protective Services was higher among children who were screened (p < 0.0001). Younger children were more likely to be screened, have a positive screen, and have a report filed. There was no difference in the odds of being screened according to hospital teaching status, size or urban vs rural location., Conclusions: A child abuse screening tool can be integrated into the electronic health record in a large health-care network. The increased number of reports among children who were screened suggests that screening facilitates detection of suspected maltreatment., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

A metastatic cancer diagnosis is associated with high levels of distress in patients and caregivers. Mindfulness interventions can reduce distress and improve quality of life in cancer patients. However, standard mindfulness training relies on in-person instruction, which is often not practical for either patients receiving chemotherapy or their caregivers. In the Being Present single arm pilot study, we designed and tested an 8-week audio-based mindfulness meditation program for patients with metastatic colorectal cancer receiving chemotherapy with or without a participating caregiver. The study accrued 33 of 74 (45%) eligible patients consenting together with 20 family caregivers (53 participants total) within nine months. Forty-one participants were evaluable (77%); 10 of 12 cases of attrition were attributable to hospitalization or death. Median participant age was 51 (range 21-78 years); 38% were men. Baseline levels of distress were similar in patients and caregivers. The top reasons for participation cited in pre-intervention interviews were to increase relaxation/calm, improve mood/emotions, and reduce stress/anxiety. In measures of adherence, 59% of responses to weekly texts asking: "Have you practiced today?" were "Yes" and 59% of interviewees reported practicing >50% of the time. Compared to baseline, post-intervention surveys demonstrated significantly reduced distress (p = 0.01) and anxiety (p = 0.03); as well as increased non-reactivity (p<0.01), and feeling at peace (p<0.01). Post-intervention qualitative interviews, where 71% of participants reported benefit, were consistent with quantitative findings. In the interviews, participants spontaneously described reduced stress/anxiety and increased relaxation/calm. Benefits appeared to be accentuated in patient-caregiver pairs as compared to unpaired patients. Seventy-nine percent of participants reported plans for continued practice after study completion. We conclude that the Being Present audio-based mindfulness meditation program is of interest to, feasible, and acceptable for patients with metastatic colorectal cancer and caregivers, with initial evidence of efficacy. These results will guide plans for a follow-up study., Trial Registration: ClinicalTrials.gov NCT02423720., Competing Interests: The authors have declared that no competing interests exist.

We previously developed the technique of conditional reprogramming (CR), which allows primary epithelial cells from fresh or cryopreserved specimens to be propagated long-term in vitro, while maintaining their genetic stability and differentiation potential. This method requires a combination of irradiated fibroblast feeder cells and a Rho-associated kinase (ROCK) inhibitor. In the present study, we demonstrate increased levels of full-length p53 and its natural isoform, Δ133p53α, in conditionally reprogrammed epithelial cells from primary prostate, foreskin, ectocervical, and mammary tissues. Increased Δ133p53α expression is critical for CR since cell proliferation is rapidly inhibited following siRNA knockdown of endogenous Δ133p53α. Importantly, overexpression of Δ133p53α consistently delays the onset of cellular senescence of primary cells when cultured under non-CR conditions in normal keratinocyte growth medium (KGM). More significantly, the combination of Δ133p53α overexpression and ROCK inhibitor, without feeder cells, enables primary epithelial cells to be propagated long-term in vitro. We also show that Δ133p53α overexpression induces hTERT expression and telomerase activity and that siRNA knockdown of hTERT causes rapid inhibition of cell proliferation, indicating a critical role of hTERT for mediating the effects of Δ133p53α. Altogether, these data demonstrate a functional and regulatory link between p53 pathways and hTERT expression during the conditional reprogramming of primary epithelial cells.

Recurrent self-mating can result in nearly clonal propagation of biological lineages, but even occasional outcrossing can serve to redistribute variation in future generations, providing cohesion among regional populations. The zoonotic parasite Trichinella spiralis has been suspected to undergo frequent inbreeding, resulting in genetically uniform larval cohorts which differ markedly from one another. Here, we explored the extent of inbreeding for this parasite by determining how genetic variation (at variable microsatellite markers) is distributed among 1379 larvae derived from 41 wild boars in Extremadura, Spain. In particular, we sought to determine how much of the genetic variation in this region's parasites occurs among the larvae of any given wild boar, and whether each derives from one, or more, parental lineages. We found strong evidence for inbreeding, resulting in genetically distinct parasite subpopulations among the parasites derived from many pairs of wild boar. Fully two-thirds of these parasite cohorts appear to derive from inbred parents; in 10% of the wild boars, parasites were so inbred as to become absolutely fixed in all of the assayed genetic loci. In spite of this, more than one pair of parents appear to have given rise to the infections in one-third of the sampled wild boars, resulting in mixed infections. These mixed infections should slow losses of heterozygosity and multi-locus polymorphism in any given parasite lineage. Such outcrossing should limit distinctions that would otherwise accumulate among transmission chains, thereby enforcing cohesion through the region's population in spite of its marked departure from panmixia. Conditions of transmission may differ in other regions, where such epidemiological features may engender different evolutionary outcomes., (Copyright © 2018 Australian Society for Parasitology. All rights reserved.)

Objective: We sought to determine under what conditions brighter lighting improves reading performance., Method: Thirteen participants with typical sight and 9 participants with age-related macular degeneration (AMD) read sentences ranging from 0.0 to 1.3 logMAR under luminance levels ranging from 3.5 to 696 cd/m²., Results: At the dimmest luminance level (3.5 cd/m²), reading speeds were slowest at the smaller letter sizes and reached an asymptote for larger sizes. When luminance was increased to 30 cd/m², reading speed increased only for the smaller letter sizes. Additional lighting did not increase reading speeds for any letter size. Similar size-related effects of luminance were observed in participants with AMD., Conclusion: In some instances, performance on acuity-limited tasks might be improved by brighter lighting. However, brighter lighting does not always improve reading; the magnitude of the effect depends on the text size and the relative changes in light level., (Copyright © 2018 by the American Occupational Therapy Association, Inc.)

Here we report a new species of Sarcocystis with a barred owl ( Strix varia) as the natural definitive host and interferon gamma gene knockout (KO) mice as an experimental intermediate host. A barred owl submitted to the Carolina Raptor Center, Huntersville, North Carolina, was euthanized because of paralysis. Fully sporulated 12.5 × 9.9 μm sporocysts were found in intestinal scrapings from the owl. Sporocysts from the barred owl were orally fed to 4 laboratory-reared outbred Swiss Webster (SW) ( Mus musculus) and 8 KO mice. All mice remained asymptomatic. Microscopic sarcocysts were found in all 5 KO mice euthanized on day 32, 59, 120, 154, and 206 post-inoculation (PI), not in KO mice euthanized on day 4, 8, and 14 PI. Sarcocysts were not found in any SW mice euthanized on day 72, 120, 206, and 210 PI. Sarcocysts were microscopic, up to 70 μm wide. By light microscopy, the sarcocyst wall < 2 μm thick had undulating, flat to conical, protrusions of varying dimensions. Numerous sarcocysts were seen in the histological sections of tongue and skeletal muscles from the abdomen, limbs, and eye but not in the heart. By transmission electron microscopy, the sarcocyst wall was "type 1j." The ground substance layer (gs) was homogenous, up to 2 μm thick, with very fine granules, and a few vesicles concentrated toward the villar projections. No microtubules were seen in the gs. Longitudinally cut bradyzoites at 206 days PI were 7.8 × 2.2 μm. Based on molecular characterization using 18S rRNA, 28S rRNA, and cox1 genes and morphology of sarcocysts, the parasite in the present study was biologically and structurally different from species so far described, and we therefore propose a new species name, Sarcocystis strixi n. sp.

Here, we report a new species, Sarcocystis pantherophisi n. sp., with the Eastern rat snake (Pantherophis alleghaniensis) as natural definitive host and the interferon gamma gene knockout (KO) mouse as the experimental intermediate host. Sporocysts (n = 15) from intestinal contents of the snake were 10.8 × 8.9 μm. Sporocysts were orally infective to KO mice but not to laboratory-raised albino outbred house mice (Mus musculus). The interferon gamma KO mice developed schizont-associated neurological signs, and schizonts were cultivated in vitro from the brain. Mature sarcocysts were found in skeletal muscles of KO mice examined 41 days postinoculation (PI). Sarcocysts were slender, up to 70 μm wide and up to 3.5 mm long. By light microscopy, sarcocysts appeared thin-walled (<1 μm) without projections. By transmission electron microscopy, the sarcocyst wall was a variant of "type 1" (type 1i, new designation). The parasitophorous vacuolar membrane (pvm) had approximately 100-nm-wide × 100-nm-long bleb-like evaginations interspersed with 100-nm-wide × 650-nm-long elongated protrusions at irregular distances, and invaginations into the ground substance layer (gs) for a very short distance (6 nm). The gs was smooth, up to 500 nm thick, without tubules, and contained a few vesicles. Longitudinally cut bradyzoites at 54 days PI were banana-shaped, 7.8 × 2.2 μm (n = 5). Molecular characterization using 18S rRNA, 28S rRNA, ITS-1, and cox1 genes indicated a close relationship with other Sarcocystis parasites that have snake-rodent life cycles. The parasite in the present study was molecularly and biologically similar to a previously reported isolate (designated earlier as Sarcocystis sp. ex Pantherophis alleghaniensis) from P. alleghaniensis, and it was structurally different from other Sarcocystis species so far described.

Here, we report a new species of Sarcocystis with red-tailed hawk (RTH, Buteo jamaicensis) as the natural definitive host and IFN-γ gene knockout (KO) mice as an experimental intermediate host in which sarcocysts form in muscle. Two RTHs submitted to the Carolina Raptor Center, Huntersville, North Carolina, were euthanized because they could not be rehabilitated and released. Fully sporulated 12.5 × 9.9-μm sized sporocysts were found in intestinal scrapings of both hawks. Sporocysts were orally fed to laboratory-reared outbred Swiss Webster mice (SW, Mus musculus) and also to KO mice. The sporocysts were infective for KO mice but not for SW mice. All SW mice remained asymptomatic, and neither schizonts nor sarcocysts were found in any SW mice euthanized on days 54, 77, 103 (n = 2) or 137 post-inoculation (PI). The KO mice developed neurological signs and were necropsied between 52 to 68 days PI. Schizonts/merozoites were found in all KO mice euthanized on days 52, 55 (n = 3), 59, 61 (n = 2), 66, and 68 PI and they were confined to the brain. The predominant lesion was meningoencephalitis characterized by perivascular cuffs, granulomas, and necrosis of the neural tissue. The schizonts/merozoites were located in neural tissue and were apparently extravascular. Brain homogenates from infected KO mice were infective to KO mice by subcutaneous inoculation and when seeded on to CV-1 cells. Microscopic sarcocysts were found in skeletal muscles of 5 of 8 KO mice euthanized between 55-61 days PI. Only a few sarcocysts were detected. Sarcocysts were microscopic, up to 3.5 mm long. When viewed with light microscopy, the sarcocyst wall appeared thin (<1 μm thick) and smooth. By transmission electron microscopy, the sarcocyst wall classified as "type 1j" (new designation). Molecular characterization using 18S rRNA, 28S rRNA, ITS-1, and cox1 genes revealed a close relationship with Sarcocystis microti and Sarcocystis glareoli; both species infect birds as definitive hosts. The parasite in the present study was biologically and molecularly different from species so far described in RTHs and we therefore propose a new species name, Sarcocystis jamaicensis n. sp.

Unlike most species in the genus Sarcocystis, Sarcocystis canis has a broad intermediate host range. Its life cycle is incompletely known and most reports are from the USA. Here we report fatal hepatitis in a 4year old male Indo-Pacific bottlenose dolphin (Tursiops aduncus) from Hong Kong associated with a S. canis-like infection. Diagnosis was made based on clinical presentation, histopathology, transmission electron microscopy (TEM), and molecular characterization. Microscopically, S. canis-like like infection was confined to the liver. Immature and mature schizonts were found in hepatocytes and the parasite was associated with generalized hepatic necrosis. By TEM, schizonts divided by endopolygeny, and merozoites lacked rhoptries. Molecular characterization of parasites present in liver and brain tissues at the cox1 gene showed a high degree of identity (97-98%) and clustered together with Sarcocystis canis, S. lutrae, S. arctica, S. speeri, S. turdusi, and S. rileyi in a phylogenetic study. This is the first report of S. canis-like infection from Asia., (Published by Elsevier B.V.)

The excretory-secretory products (ESPs) released by the muscle-larvae (ML) stage of Trichinella spiralis have been suggested to be involved in nurse cell formation. However, the molecular mechanisms by which ML-ESPs modulate nurse cell formation remain unclear. Macrophages exert either beneficial or deleterious effects on tissue repair, depending on their activation/polarization state. They are crucial for skeletal muscle repair, notably, via their actions on myogenic precursor cells. However, these interactions during T. spiralis infection have not been characterized. In the present study, the ability of conditioned medium (CM) from J774A.1 macrophages treated with ML-ESPs to influence the differentiation of murine myoblasts, and the mechanisms of this influence, were investigated in vitro. The results showed that the expression of Myogenic Regulatory Factors (MRFs) MyoD and myogenin, myosin heavy chain (MyHC), and the p21 cyclin-dependent kinase inhibitor were reduced in CM treated cells compared to their expression in the control group. These findings indicated that CM inhibited myoblast differentiation. Conversely, CM promoted myoblast proliferation and increased cyclin D1 levels. Taken together, results of our study suggested that CM can indirectly influence myoblast differentiation and proliferation, which provides a new method for the elucidation of the complex mechanisms involved in cell-parasite and cell-cell interactions during T. spiralis infection., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Trichinella spiralis can cause immunosuppression during the intestinal phase of early infection. However, changes in the peripheral blood during T. spiralis early infection remain unclear. Here, select immune cells in mice infected with 500 muscle larvae (ML) of T. spiralis during the intestinal phase of infection were studied. First, the recovery rates of the intestinal worms and female fecundity were determined, and the results showed that the intestinal worms were completely eliminated at 17 days post-infection (dpi) and that large numbers of new-born larvae (NBL) were generated from 5 to 9dpi. Using flow cytometry, it was shown that the number of CD4+ T cells and CD8+ T cells increased over the entire intestinal phase, except on 7dpi when CD4+ T cells decreased significantly compared to the control groups. Although both CD4+ and CD8+ T cells increased, CD8+ T cells increased more than CD4+ T cells, leading to a lower CD4+/CD8+ ratio compared to the control group. Subsequently, a depression of the proliferative response of T cells to concanavalin A (Con A) was noticed at 7 and 11dpi. Although the proliferative response of B cells to LPS was enhanced, the number of B cells from mouse peripheral blood stimulated by T. spiralis antigens showed no differences with the control group prior to 11dpi. The expression of CD14 on monocyte-macrophages decreased during the same period, which meant that the antigen-presenting response was reduced in the immune system of the infected mice. Moreover, the alternatively activated macrophages were induced in T. spiralis early infection. These data provide a better understanding of the development of the intestinal immune response in mice infected with T. spiralis., (Copyright © 2016 Elsevier B.V. All rights reserved.)

To advance scientific knowledge about human diseases and effective therapeutic treatments, investigators need access to human biospecimens and associated data. However, regulatory and procedural requirements may impede investigators' efforts to share biospecimens and data within and across institutions. Although a number of studies have explored experiences and attitudes of study participants and others about biospecimen and data sharing, less is known about investigators' perspectives. We conducted an electronic survey to learn about investigators' experiences and attitudes about research with biospecimens and associated data. A total of 114 practicing scientists from a pool of 60 university medical schools with Clinical and Translational Science Awards (CTSAs) funded by the National Institutes of Health (NIH) participated. We found a high degree of variability in investigators' experiences with institutional review boards (IRBs) when seeking approval to conduct biospecimen research, as well as differences in approaches to informed consent for the collection of specimens. Participants also expressed concerns that the risks of biospecimen research may be overestimated by IRBs. This research suggests that the current regulatory environment for human research protections may require reconsideration with regard to standards for collection, use, and sharing of biospecimens and data., (© The Author(s) 2015.)

Trichinellosis is a zoonotic disease caused by Trichinella muscle larvae (ML) through ingestion of raw or undercooked meat. To date, 12 taxa are recognized in this genus, of which four are circulating in Europe (Trichinella spiralis, Trichinella nativa, Trichinella britovi and Trichinella pseudospiralis). T. spiralis and T. britovi circulate in European wildlife and occur simultaneously in the same host species. The possibility of hybrid formation between T. britovi and T. spiralis has hardly been addressed and so far, results of experimental hybridisation attempts between T. britovi and T. spiralis are inconclusive. The aim of the present study was to analyse molecular polymorphisms of single T. spiralis and T. britovi ML from natural infections based on nuclear 5S rDNA intergenic spacer region (5S rDNA-ISR) and mitochondrial cytochrome c oxidase 1 (CO1) gene sequences. Six haplotypes of the 5S rDNA intergenic spacer region (5S rDNA-ISR) and 14 of the cytochrome c oxidase 1 (CO1) gene were demonstrated in 89 individual T. britovi ML from Latvia and Poland. In contrast, only two haplotypes were observed at both 5S rDNA-ISR and CO1 of 57 individual T. spiralis ML from Polish wild boar and red foxes. Moreover, this study demonstrates hybridisation in eight individual ML between T. britovi and T. spiralis under natural conditions in four Polish wild boar and two red foxes, revealed by combining 5S rDNA-ISR and CO1 sequence information of individual Trichinella ML. To our knowledge, this is the first report of interspecies hybridisation between T. spiralis and T. britovi under field conditions., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The North American opossum (Didelphis virginiana) is the definitive host for at least three named species of Sarcocystis: Sarcocystis falcatula, Sarcocystis neurona and Sarcocystis speeri. The South American opossums (Didelphis albiventris, Didelphis marsupialis and Didelphis aurita) are definitive hosts for S. falcatula and S. lindsayi. The sporocysts of these Sarcocystis species are similar morphologically. They are also not easily distinguished genetically because of the difficulties of DNA extraction from sporocysts and availability of distinguishing genetic markers. Some of these species can be distinguished by bioassay; S. neurona and S. speeri are infective to gamma interferon gene knockout (KO) mice, but not to budgerigars (Melopsittacus undulatus); whereas S. falcatula and S. lindsayi are infective to budgerigars but not to KO mice. The natural intermediate host of S. speeri is unknown. In the present study, development of sarcocysts of S. speeri in the KO mice is described. Sarcocysts were first seen at 12 days post-inoculation (p.i.), and they became macroscopic (up to 4 mm long) by 25 days p.i. The structure of the sarcocyst wall did not change from the time bradyzoites had formed at 50-220 days p.i. Sarcocysts contained unique villar protrusions, 'type 38'. The polymerase chain reaction amplifications and sequences analysis of three nuclear loci (18S rRNA, 28S rRNA and ITS1) and two mitochondrial loci (cox1 and cytb) of S. speeri isolate from an Argentinean opossum (D. albiventris) confirmed its membership among species of Sarcocystis and indicated an especially close relationship to another parasite in this genus that employs opossums as its definitive host, S. neurona. These results should be useful in finding natural intermediate host of S. speeri.

Transmission of pathogens between domestic and wild life animals plays an important role in epidemiology. Feral pig populations are increasing and expanding in the USA, and may constitute a risk to non-biosecure domestic pig facilities by serving as reservoirs for pathogens. We surveyed, for Sarcocystis infection, the myocardium of 1006 feral pigs (Sus scrofa) trapped or hunted in 29 states during the Comprehensive Feral Swine Disease Surveillance Program of the USDA's Animal and Plant Health Inspection Service, Wildlife Services unit during 2012-2014. Sarcocysts were detected in histological sections of 25% (251/1006) of myocardium with an average parasitic load/intensity of infection of 3.03 sarcocysts/section (1.5×0.7 cm), and higher prevalence of myocarditis in severe infections. Microscopic examination of pepsin digests of 147 hearts revealed a higher prevalence of Sarcocystis bradyzoites (49%, 72/147) than when diagnosed by histology. A fragment of Sarcocystis 18S rRNA was amplified and digested with a restriction endonuclease, revealing a pattern consistent with Sarcocystis miescheriana in all 44 selected samples. Sequencing 31 of these 44 isolates confirmed their correspondence to S. miescheriana. Thus, S. miescheriana infection, but not the zoonotic parasite Sarcocystis suihominis, appears to be prevalent and widespread in feral pigs in the USA.

The distribution, abundance, and diversity of life on Earth have been greatly shaped by human activities. This includes the geographic expansion of parasites; however, measuring the extent to which humans have influenced the dissemination and population structure of parasites has been challenging. In-depth comparisons among parasite populations extending to landscape-level processes affecting disease emergence have remained elusive. New research methods have enhanced our capacity to discern human impact, where the tools of population genetics and molecular epidemiology have begun to shed light on our historical and ongoing influence. Only since the 1990s have parasitologists coupled morphological diagnosis, long considered the basis of surveillance and biodiversity studies, with state-of-the-art tools enabling variation to be examined among, and within, parasite populations. Prior to this time, populations were characterized only by phenotypic attributes such as virulence, infectivity, host range, and geographical location. The advent of genetic/molecular methodologies (multilocus allozyme electrophoresis, polymerase chain reaction-DNA [PCR-DNA] fragments analysis, DNA sequencing, DNA microsatellites, single nucleotide polymorphisms, etc.) have transformed our abilities to reveal variation among, and within, populations at local, regional, landscape, and global scales, and thereby enhanced our understanding of the biosphere. Numerous factors can affect population structure among parasites, e.g., evolutionary and ecological history, mode of reproduction and transmission, host dispersal, and life-cycle complexity. Although such influences can vary considerably among parasite taxa, anthropogenic factors are demonstrably perturbing parasite fauna. Minimal genetic structure among many geographically distinct (isolated) populations is a hallmark of human activity, hastened by geographic introductions, environmental perturbation, and global warming. Accelerating environmental change now plays a primary role in defining where hosts, parasites, and other pathogens occur. This review examines how anthropogenic factors serve as drivers of globalization and genetic homogenization of parasite populations and demonstrates the impact that human intervention has had on the global dissemination of parasites and the accompanying diseases.

Sarcocystis infections have been reported from the African buffalo ( Syncerus caffer ), but the species have not been named. Here we propose a new name Sarcocystis cafferi from the African buffalo. Histological examination of heart (92), skeletal muscle (36), and tongue (2) sections from 94 buffalos from the Greater Kruger National Park, South Africa, and a review of the literature revealed only 1 species of Sarcocystis in the African buffalo. Macrocysts were up to 12 mm long and 6 mm wide and were located in the neck muscles and overlying connective tissue. They were pale yellow; shaped like a lychee fruit stone or cashew nut; turgid or flaccid and oval to round (not fusiform). By light microscopy (LM) the sarcocyst wall was relatively thin. By scanning electron microscopy (SEM), the sarcocyst wall had a mesh-like structure with irregularly shaped villar protrusions (vp) that were of different sizes and folded over the sarcocyst wall. The entire surfaces of vp were covered with papillomatous structures. By transmission electron microscopy (TEM), the sarcocyst wall was up to 3.6 μm thick and had highly branched villar protrusions that were up to 3 μm long. The villar projections contained filamentous tubular structures, most of which were parallel to the long axis of the projections, but some tubules criss-crossed, especially at the base. Granules were absent from these tubules. Longitudinally cut bradyzoites were 12.1 × 2.7 μm in size, had a long convoluted mitochondrion, and only 2 rhoptries. Phylogenetic analysis of 18S rRNA and cytochrome C oxidase subunit 1 (cox1) gene sequences indicated that this Sarcocystis species is very closely related to, but distinct from, Sarcocystis fusiformis and Sarcocystis hirsuta. Thus, morphological findings by LM, SEM, and TEM together with molecular phylogenetic data (from 18S rRNA and cox1) confirm that the Sarcocystis species in the African buffalo is distinct from S. fusiformis and has therefore been named Sarcocystis cafferi.

There are uncertainties concerning the identity and host species specificity of Sarcocystis species of the water buffalo (Bubalus bubalis) and cattle (Bos taurus). Currently, in cattle three species are recognized with known endogenous stages, viz.: S. cruzi (with canine definitive host), S. hirsuta (feline definitive host), and S. hominis (primate definitive host). Recently, a fourth Sarcocystis species with an unknown life cycle has been reported from cattle. In the water buffalo, four species of Sarcocystis have been described: S. fusiformis (feline definitive host), S. buffalonis (feline definitive host), S. levinei (canine definitive host), and S. dubeyi (definitive host unknown but not cat or dog). Besides, there are studies of Sarcocystis infections in buffalo and cattle from China with results that are difficult to interpret and validate. For example, some of the studies report transmission of Sarcocystis species between cattle and buffalo, but steps to preclude exogenous exposures were not reported. A species of the water buffalo, 'S. sinensis', was proposed at a Chinese national conference in 1990, and published as an abstract without figures and with no archived type specimens for verification. The International Code of Zoological Nomenclature Articles 9 and 10 state that "abstracts of articles, papers, posters, text of lectures, and similar material when issued primarily to participants at meetings, symposia, colloquia or congress does not constitute published work"; therefore, S. sinensis is a nomen nudum., (Published by Elsevier B.V.)

Background: Substantial attention has recently been placed on fractures of the posterior malleolus. Fracture extension to the posteromedial rim ("posterior pilon variant") may result in articular incongruity and talar subluxation. Current classification systems fail to account for these fractures. The relative frequency of this fracture, its associated patient characteristics, and the reliability of its diagnosis have never been reported in such a large series., Methods: We retrospectively identified 270 patients who met our inclusion criteria. Basic demographic data were collected. The fractures were classified according to Lauge-Hansen and AO/OTA. Additional radiographic data included whether the fracture involved the posterior malleolus and whether the fracture represented a posterior pilon variant. Univariate statistical methods, chi-square analysis, and interobserver reliability were assessed., Results: The relative frequency of posterior malleolus fracture was 50%. The relative frequency of the posterior pilon variant was 20%. No significant difference was noted with respect to the frequency of posterior malleolar or posterior pilon variant between the subgroups of the AO/OTA and Lauge-Hansen classification systems when compared to the overall fracture distribution. Patients with posterior malleolar fractures and posterior pilon variants were significantly older. Females were significantly more likely than men to sustain posterior malleolar fractures and posterior pilon variants. Patients with diabetes trended toward a greater risk of both types of fractures. Interobserver reliability data revealed substantial agreement for posterior malleolar fractures and posterior pilon variants., Conclusion: These data represent the highest reported rate of posterior malleolar involvement in operatively treated ankle fractures and is the first to describe the percentage of the posterior pilon variant in such a large series. The interobserver reliability data demonstrate substantial agreement in identification of posterior malleolar fractures and the posterior pilon variant based on plain radiographs. Certain patient characteristics such as age, sex, and diabetes may be associated with these fractures., Level of Evidence: Level III, retrospective cohort study., (© The Author(s) 2014.)

Objective: To assess the association of insurance status on infant rehospitalization in a population-based setting., Methods: In this longitudinal retrospective study, hospitalizations were tracked for 1 year after birth discharge for 203 031 infants born in hospitals during 2008 using data from the New York State Inpatient Database. Relative risk was estimated using multivariable negative binomial regression models., Results: Rehospitalization occurred in 9010 infants (4.4%). Medicaid coverage and being uninsured were strong predictors of rehospitalizations after adjustment for birth weight and other factors. Medicaid also bears a disproportionate share of the economic burden. Normal birth weight infants have the lowest risk, but comprise the majority of costs. Jaundice and acute bronchiolitis were the leading causes of rehospitalization within 30 days and 1 year, respectively., Discussion: Future research can explore the preventability of rehospitalizations, and evaluate novel strategies for discharge and postnatal care coordination especially for uninsured and Medicaid-enrolled infants., (© The Author(s) 2014.)