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Tumor-associated macrophages (TAMs) promote tumor progression and inhibit anti-tumor immune response by producing various mediators and preferentially express CD163, CD204, and CD206. However, the role of these TAM subsets in oral squamous cell carcinoma (OSCC) remains unclear. Here we investigated the expression and function of TAM subsets in OSCC, especially in cancer cell proliferation. Biopsy sample from 44 patients with OSCC were examined for the expression of TAM markers and EGF by immunohistochemistry. EGF production of TAM subsets isolated from OSCC patients was assessed by flow cytometry. We also examined the effect of conditioned medium from TAM subsets on the proliferation of OSCC cells. CD163(+) cells were detected diffusely all over the tumor and connective tissue area, while CD204(+) and CD206(+) cells were mainly detected in/around the tumors. Flow cytometric analysis found that CD206(+) TAMs strongly produced EGF compared with CD163(+) and CD204(+) TAMs. Cell proliferation and invasion of OSCC cells cultured with conditioned medium of CD206(+) TAMs were strongly enhanced and inhibited by anti-EGFR. The number of CD206(+) TAMs positively correlated with worse clinical prognosis. Our results revealed differences in localization and EGF production among these TAM subsets. CD206(+) TAMs might play a critical role in the proliferation of OSCC via EGF production. |
Point-of-care tests are needed for the screening of head and neck squamous cell carcinoma (HNSCC) and other malignancies. Luciferase immunoprecipitation systems (LIPS), employing light-emitting proteins, were used to examine serum antibodies against several cancer-associated targets in blood donor controls and subjects with colon cancer (CC) and HNSCC. The assessment of antibodies against the wild type p53 tumor antigen showed that approximately 25% of the CC and 20% of the HNSCC patients were seropositive. In addition, humoral responses against two p53 mutants, p53-R175H and p53-R273H, generally tracked the antibody responses seen against wild type p53. Analysis of antibodies against highly specific biomarkers of HPV-16-associated malignancy, E2, E6, and E7 oncoproteins, revealed no seropositivity in blood donors and CC patients. However, 45% (9/20) of the HNSCC patients showed E6 seropositivity, which overlapped all the detectable E2 (40%; 8/20) and E7 seropositive subjects (35%; 7/20). Using neodymium magnets, ultrarapid LIPSTICKS testing of HPV-16 E6 antibodies in <60 s per HNSCC sample demonstrated almost the same diagnostic performance (40% sensitivity and 100% specificity) as LIPS testing in 2.5 h. While additional improvements and standardization are needed, these results highlight the possibility of using these approaches for the diagnosis of HPV-16-associated HNSCC. |
Interplay between Cymbidium mosaic virus (CymMV)/Odontoglossum ringspot virus (ORSV) and its host plant Phalaenopsis equestris remain largely unknown, which led to deficiency of effective measures to control disease of P. equestris caused by infecting viruses. In this study, for the first time, we characterized viral small interfering RNAs (vsiRNAs) profiles in P. equestris co-infected with CymMV and ORSV through small RNA sequencing technology. CymMV and ORSV small interfering RNAs (siRNAs) demonstrated several general and specific/new characteristics. vsiRNAs, with A/U bias at the first nucleotide, were predominantly 21-nt long and they were derived predominantly (90%) from viral positive-strand RNA. 21-nt siRNA duplexes with 0-nt overhangs were the most abundant 21-nt duplexes, followed by 2-nt overhangs and then 1-nt overhangs 21-nt duplexes in infected P. equestris. Continuous but heterogeneous distribution and secondary structures prediction implied that vsiRNAs originate predominantly by direct Dicer-like enzymes cleavage of imperfect duplexes in the most folded regions of the positive strand of both viruses RNA molecular. Furthermore, we totally predicted 54 target genes by vsiRNAs with psRNATarget server, including disease/stress response–related genes, RNA interference core components, cytoskeleton-related genes, photosynthesis or energy supply related genes. Gene Ontology classification showed that a majority of the predicted targets were related to cellular components and cellular processes and performed a certain function. All target genes were down-regulated with different degree by vsiRNAs as shown by real-time reverse transcription polymerase chain reaction. Taken together, CymMV and ORSV siRNAs played important roles in interplay with P. equestris by down modulating the expression levels of endogenous genes in host plant. |
Zika virus is an emergent pathogen that gained significant importance during the epidemic in South and Central America as unusual and alarming complications of infection were recognized. Although initially considered a self-limited benign infection, a panoply of neurologic complications were recognized including a Guillain–Barré-like syndrome and in-utero fetal infection causing microcephaly, blindness, and other congenital neurologic complications. Numerous Zika virus vaccines were developed, with nine different vaccines representing five different platforms entered into clinical trials, one progressing to Phase II. Here we review the current landscape and challenges confronting Zika virus vaccine development. |
Even though transitivity is a central structural feature of social networks, its influence on epidemic spread on coevolving networks has remained relatively unexplored. Here we introduce and study an adaptive susceptible-infected-susceptible (SIS) epidemic model wherein the infection and network coevolve with nontrivial probability to close triangles during edge rewiring, leading to substantial reinforcement of network transitivity. This model provides an opportunity to study the role of transitivity in altering the SIS dynamics on a coevolving network. Using numerical simulations and approximate master equations (AMEs), we identify and examine a rich set of dynamical features in the model. In many cases, AMEs including transitivity reinforcement provide accurate predictions of stationary-state disease prevalence and network degree distributions. Furthermore, for some parameter settings, the AMEs accurately trace the temporal evolution of the system. We show that higher transitivity reinforcement in the model leads to lower levels of infective individuals in the population, when closing a triangle is the dominant rewiring mechanism. These methods and results may be useful in developing ideas and modeling strategies for controlling SIS-type epidemics. |
OBJECTIVES: There are numerous point‐of‐care tests (POCTs) available on the market, but many of these are not used. This study reviewed literature pertaining to the evaluation/usage of POCTs in primary care, to investigate whether outcomes being reported reflect aspects previously demonstrated to be important for general practitioners (GPs) in the decision to implement a POCT in practice. METHODS: Scopus and Medline were searched to identify studies that evaluated a POCT in primary care. We identified abstracts and full‐texts consisting of applied studies (eg trials, simulations, observational studies) and qualitative studies (eg interviews, surveys). Data were extracted from the included studies, such as the type of study, the extent to which manufacturers were involved in the study, and the biomarker/assay measured by the test(s). Studies were evaluated to summarise the extent to which they reported on, amongst others, clinical utility, user‐friendliness, turnaround‐time and technical performance (aspects previously identified as important). RESULTS: The initial search resulted in 1398 publications, of which 125 met the inclusion criteria. From these studies, 83 POCTs across several disease areas (including cardiovascular disease, venous thromboembolism and respiratory‐tract‐infections) were identified. There was an inconsistency between what is reported in the studies and what GPs consider important. GPs perceive clinical utility as the most important aspect, yet this was rarely included explicitly in test evaluations in the literature, with only 8% of evaluations incorporating it in their analysis/discussion. CONCLUSIONS: This review showed that, despite the growing market and development of new POCTs, studies evaluating such tests fail to report on aspects that GPs find important. To ensure that an evaluation of a POCT is useful to primary care clinicians, future evaluations should not only focus on the technical performance aspects of a test, but also report on the aspects relating to the clinical utility and risks. |
Guertu virus (GTV) is a tick-borne phleboviruses (TBPVs) which belongs to the genus Banyangvirus in the family of Phenuiviridae. In vitro and in vivo studies of GTV demonstrated that it was able to infect animal and human cell lines and could cause pathological lesions in mice. Glycoproteins (GP, including Gn and Gc) on the surface of Guertu virus (GTV) could bind to receptors on host cells and induce protective immunity in the host, but knowledge is now lacking on the information of B cell epitopes (BCEs) present on GTV-GP protein. The aim of this study was to identify all BCEs on Gn of the GTV DXM strain using rabbit pAbs against GTV-Gn. Seven fine BCEs and two antigenic peptides (APs) from nine reactive 16mer-peptides were identified, which are E(Gn)1 ((2)PIICEGLTHS(11)), E(Gn)2 ((135)CSQDSGT(141)), E(Gn)3 ((165)IP EDVF(170)), E(Gn)4 ((169)VFQEL K(174)), E(Gn)5 ((187)IDGILFN(193)), E(Gn)6 ((223)QTKWIQ(228)), E(Gn)7 ((237)CHKDGIGPC(245)), AP-8 ((299)GVRVRPKCYGFSRMMA(314)) and AP-9 ((355)CASH FCSSAESGKKNT(370)), of which six of mapped BCEs were recognized by the IgG-positive sheep serum obtained from sheep GTV-infected naturally. Multiple sequence alignments (MSA) based on each mapped BCE motif identified that the most of identified BCEs and APs are highly conserved among 10 SFTSV strains from different countries and lineages that share relatively close evolutionary relationships with GTV. The fine epitope mapping of the GTV-Gn would provide basic data with which to explore the GTV-Gn antigen structure and pathogenic mechanisms, and it could lay the foundation for the design and development of a GTV multi-epitope peptide vaccine and detection antigen. |
We report a hereditary leukodystrophy in Standard Schnauzer puppies. Clinical signs occurred shortly after birth or started at an age of under 4 weeks and included apathy, dysphoric vocalization, hypermetric ataxia, intension tremor, head tilt, circling, proprioceptive deficits, seizures and ventral strabismus consistent with a diffuse intracranial lesion. Magnetic resonance imaging revealed a diffuse white matter disease without mass effect. Macroscopically, the cerebral white matter showed a gelatinous texture in the centrum semiovale. A mild hydrocephalus internus was noted. Histopathologically, a severe multifocal reduction of myelin formation and moderate diffuse edema without inflammation was detected leading to the diagnosis of leukodystrophy. Combined linkage analysis and homozygosity mapping in two related families delineated critical intervals of approximately 29 Mb. The comparison of whole genome sequence data of one affected Standard Schnauzer to 221 control genomes revealed a single private homozygous protein changing variant in the critical intervals, TSEN54:c.371G>A or p.(Gly124Asp). TSEN54 encodes the tRNA splicing endonuclease subunit 54. In humans, several variants in TSEN54 were reported to cause different types of pontocerebellar hypoplasia. The genotypes at the c.371G>A variant were perfectly associated with the leukodystrophy phenotype in 12 affected Standard Schnauzers and almost 1000 control dogs from different breeds. These results suggest that TSEN54:c.371G>A causes the leukodystrophy. The identification of a candidate causative variant enables genetic testing so that the unintentional breeding of affected Standard Schnauzers can be avoided in the future. Our findings extend the known genotype-phenotype correlation for TSEN54 variants. |
BACKGROUND: Respiratory viral (RV) outbreaks in rehabilitation facilities can jeopardise patient safety, interfere with patient rehabilitation goals and cause unit closures that impede patient flow in referring facilities. PROBLEM: Despite education about infection prevention practices, frequent RV outbreaks were declared each year at our rehabilitation facility. METHODS: Before and after study design. The primary outcome was the number of bed closure days due to outbreak per overall bed days. Process measures included delays in initiation of transmission-based precautions, RV testing and reporting of staff to occupational health and safety (OHS). Balancing measures included the number of isolation days and staff missed work hours. INTERVENTIONS: Based on comprehensive analysis of prior outbreaks, the following changes were implemented: (1) clear criteria for initiation of transmission-based precautions, (2) communication to visitors to avoid visitation if infectious symptoms were present, (3) exemption of staff absences if documented due to infectious illness, (4) development of an electronic programme providing guidance to staff about whether they should be excluded from work due to infectious illness. RESULTS: The number of bed closure days due to outbreak per overall bed days dropped from 2.8% to 0.5% during the intervention season and sustained at 0.6% during the postintervention season (p<0.001). There were fewer delays in initiation of droplet and contact precautions (28.8% to 15.5%, p=0.005) and collection of RV testing (42.9% to 20.3%, p<0.001), better reporting to OHS (9 vs 28.8 reports per 100 employees; p<0.001) and fewer isolation days (7.8% vs 7.3%; p=0.02) without a significant increase in missed work hours per 100 hours worked (4.0 vs 3.9; p=0.12). CONCLUSION: This Quality Improvement study highlights the process changes that can prevent respiratory outbreaks in the rehabilitation setting. |
As a response to a diverse array of external stimuli, early growth response protein 1 (Egr-1) plays important roles in the transcriptional regulation of inflammation and the cellular immune response. However, a number of intracellular pathogens colonize immune cells and the implication of Egr-1 in the host-pathogen interplay has remained elusive. Here, we have characterized the Egr-1 responses of primary murine and human dendritic cells (DCs) upon challenge with the obligate intracellular parasite Toxoplasma gondii. We report that live intracellular parasites induce a sustained high expression of Egr-1 in DCs, different from the immediate-early Egr-1 response to parasite lysates, inactivated parasites or LPS. Moreover, a distinct nuclear localization of elevated amounts of Egr-1 protein was detected in infected DCs, but not in by-stander DCs. The ERK1/2 MAPK signaling pathway mediated the canonical immediate-early Egr-1 response to soluble antigens in a MyD88/TLR-dependent fashion. In contrast, a non-canonical extended Egr-1 response that relied primarily on p38 MAPK signaling was induced by intracellular parasites and was exhibited similarly by MyD88-deficient and wildtype DCs. The extended phase Egr-1 response was dramatically reduced upon challenge of DCs with T. gondii parasites deficient in GRA24, a secreted p38-interacting protein. Further, Egr-1-silenced primary DCs maintained their migratory responses upon T. gondii challenge. Importantly, Egr-1 silencing led to elevated expression of co-stimulatory molecules (CD40, CD80) in Toxoplasma-infected DCs and in LPS-challenged immature DCs, indicating that Egr-1 responses suppressed maturation of DCs. Moreover, the IL-12 and IL-2 responses of Toxoplasma-challenged DCs were modulated in a GRA24-dependent fashion. Jointly, the data show that the Egr-1 responses of DCs to microbial external stimuli and intracellular stimuli can be selectively mediated by ERK1/2 or p38 MAPK signaling, and that Egr-1 can act as an intrinsic negative modulator of maturation in primary DCs. |
BACKGROUND: Arbaeenia mass gathering (MG) in Karbala, Iraq, is becoming one of the largest MGs in the world. The health care infrastructure in Iraq is inadequately prepared to serve the health needs of the millions of pilgrims. OBJECTIVE: This study aimed to describe the temporary health care facilities installed and run by the local community to provide health care services to Arbaeenia pilgrims in Karbala, Iraq. METHODS: A survey was conducted in all community-based health care facilities located along part of Najaf to Karbala road within Karbala governorate. A structured questionnaire was answered through an interview with the workers and direct observation. Data were collected on staff profile, type of services provided, use of basic infection control measures, medical equipment, drugs and supplies, and the most commonly encountered medical problems. RESULTS: The total number of health care facilities was 120, staffed by 659 workers. Only 18 (15.0%, 18/120) facilities were licensed, and 44.1% (53/120) of the workers were health professionals. The health care workers provided different services including dispensing drugs (370/1692, 21.87%), measuring blood pressure and blood sugar (350/1692, 20.69%), and caring for wounds and injuries (319/1692, 18.85%). Around 97% (116/120) health facilities provided services for musculoskeletal disorders and only 16.7% (20/120) provided services for injuries. The drugs available in the clinic were analgesics, drugs for gastrointestinal and respiratory diseases, and antibiotics, with an availability range of 13.3% to 100.0%. Infection control practices for individual protection, environmental sanitation, and medical waste disposal were available in a range of 18.1% to 100.0%. CONCLUSIONS: Community-based health care facilities experienced a profound shortage of trained human resources and medical supplies. They can significantly contribute to health services if they are adequately equipped and follow standardized operation procedures. |
Among the various nano/biomaterials used in cancer treatment, the beauty and benefits of DNA nanocomposites are outstanding. The specificity and programmability of the base pairing of DNA strands, together with their ability to conjugate with different types of functionalities have realized unsurpassed potential for the production of two- and three-dimensional nano-sized structures in any shape, size, surface chemistry and functionality. This review aims to provide an insight into the diversity of static DNA nanodevices, including DNA origami, DNA polyhedra, DNA origami arrays and bioreactors, DNA nanoswitch, DNA nanoflower, hydrogel and dendrimer as young but promising platforms for cancer theranostics. The utility and potential of the individual formats in biomedical science and especially in cancer therapy will be discussed. |
Rift Valley fever virus (RVFV) is a zoonotic mosquito-borne virus that was first discovered in Kenya in 1930 and has since spread to become endemic in much of Africa and the Arabian Peninsula. Rift Valley fever (RVF) causes recurrent outbreaks of febrile illness associated with high levels of mortality and poor outcomes during pregnancy—including foetal malformations, spontaneous abortion and stillbirths—in livestock, and associated with miscarriage in humans. No vaccines are available for human use and those licensed for veterinary use have potential drawbacks, including residual virulence that may contraindicate their use in pregnancy. To address this gap, we previously developed a simian adenovirus vectored vaccine, ChAdOx1 RVF, that encodes RVFV envelope glycoproteins. ChAdOx1 RVF is fully protective against RVF in non-pregnant livestock and is also under development for human use. Here, we now demonstrate that when administered to pregnant sheep and goats, ChAdOx1 RVF is safe, elicits high titre RVFV neutralizing antibody, and provides protection against viraemia and foetal loss, although this protection is not as robust for the goats. In addition, we provide a description of RVFV challenge in pregnant goats and contrast this to the pathology observed in pregnant sheep. Together, our data further support the ongoing development of ChAdOx1 RVF vaccine for use in livestock and humans. |
BACKGROUND: Viral diseases are a matter of great concern for poultry farmers in Pakistan. Multiple common viral respiratory diseases (CVRDs) cause huge economic losses in the poultry industry. The prevalence of CVRDs in many countries, including Pakistan, is not clearly understood. RESULTS: Incidences of 5 chicken respiratory viruses: avian influenza virus (AIV), Newcastle disease virus (NDV/AAVV-1), infectious bronchitis virus (IBV), avian metapneumovirus (aMPV) and infectious laryngotracheitis virus (ILTV) were assessed on commercial Pakistani farms with respiratory problems from 2014 through to 2016. While AIV and AAVV-1 were frequently detected (16 to 17% of farms), IBV and aMPV were rarely detected (in 3 to 5% of farms) and ILTV was not detected. We characterized H9 AIV of the G1 lineage, genotype VII AAVV-1, GI-13 IBV, and type B aMPV strains with very little genetic variability in the 2-year study period. Co-infections with AIV and AAVV-1 were common and wild type AAVV-1 was detected despite the use of vaccines. Control measures to limit the virus burden in chicken flocks are discussed. CONCLUSIONS: Our data shows that AIV (H9), AAVV-1, IBV and aMPV are prevalent in commercial poultry in Pakistan. Further studies are necessary to assess circulating strains, economic losses caused by infections and coinfections of these pathogens, and the costs and benefits of countermeasures. Furthermore, veterinarians and farmers should be informed of the pathogens circulating in the field and hence advised on the use of vaccines. |
INTRODUCTION: Epidemic forecasting and prediction tools have the potential to provide actionable information in the midst of emerging epidemics. While numerous predictive studies were published during the 2016–2017 Zika Virus (ZIKV) pandemic, it remains unknown how timely, reproducible, and actionable the information produced by these studies was. METHODS: To improve the functional use of mathematical modeling in support of future infectious disease outbreaks, we conducted a systematic review of all ZIKV prediction studies published during the recent ZIKV pandemic using the PRISMA guidelines. Using MEDLINE, EMBASE, and grey literature review, we identified studies that forecasted, predicted, or simulated ecological or epidemiological phenomena related to the Zika pandemic that were published as of March 01, 2017. Eligible studies underwent evaluation of objectives, data sources, methods, timeliness, reproducibility, accessibility, and clarity by independent reviewers. RESULTS: 2034 studies were identified, of which n = 73 met the eligibility criteria. Spatial spread, R(0) (basic reproductive number), and epidemic dynamics were most commonly predicted, with few studies predicting Guillain-Barré Syndrome burden (4%), sexual transmission risk (4%), and intervention impact (4%). Most studies specifically examined populations in the Americas (52%), with few African-specific studies (4%). Case count (67%), vector (41%), and demographic data (37%) were the most common data sources. Real-time internet data and pathogen genomic information were used in 7% and 0% of studies, respectively, and social science and behavioral data were typically absent in modeling efforts. Deterministic models were favored over stochastic approaches. Forty percent of studies made model data entirely available, 29% provided all relevant model code, 43% presented uncertainty in all predictions, and 54% provided sufficient methodological detail to allow complete reproducibility. Fifty-one percent of predictions were published after the epidemic peak in the Americas. While the use of preprints improved the accessibility of ZIKV predictions by a median of 119 days sooner than journal publication dates, they were used in only 30% of studies. CONCLUSIONS: Many ZIKV predictions were published during the 2016–2017 pandemic. The accessibility, reproducibility, timeliness, and incorporation of uncertainty in these published predictions varied and indicates there is substantial room for improvement. To enhance the utility of analytical tools for outbreak response it is essential to improve the sharing of model data, code, and preprints for future outbreaks, epidemics, and pandemics. |
Respiratory syncytial virus (RSV) infection is a leading cause of severe acute lower respiratory tract infection in infants and children worldwide. Vitamin A deficiency (VAD) is one of the most prevalent nutrition-related health problems in the world and is a significant risk factor in the development of severe respiratory infections in infants and young children. Bovine RSV (BRSV) is a primary cause of lower respiratory tract disease in young cattle. The calf model of BRSV infection is useful to understand the immune response to human RSV infection. We have previously developed an amphiphilic polyanhydride nanoparticle (NP)-based vaccine (i.e., nanovaccine) encapsulating the fusion and attachment proteins from BRSV (BRSV-NP). Calves receiving a single, intranasal dose of the BRSV-NP vaccine are partially protected from BRSV challenge. Here, we evaluated the impact of VAD on the immune response to the BRSV-NP vaccine and subsequent challenge with BRSV. Our results show that VAD calves are unable to respond to the mucosal BRSV-NP vaccine, are afforded no protection from BRSV challenge and have significant abnormalities in the inflammatory response in the infected lung. We further show that acute BRSV infection negatively impacts serum and liver retinol, rendering even well-nourished individuals susceptible to VAD. Our results support the use of the calf model for elucidating the impact of nutritional status on mucosal immunity and respiratory viral infection in infants and underline the importance of VA in regulating immunity in the respiratory mucosa. |
BACKGROUND: Nucleic acid amplification tests have allowed simultaneous detection of multiple respiratory viruses. METHODS: We compared the results of a liquid bead array xTAG Respiratory Virus Panel (RVP; (Luminex Corporation, Toronto, Canada) and a solid microarray Verigene Respiratory Virus Plus (RV+; Nanosphere, Northbrook, IL) for the detection of influenza A virus (INF A), influenza B virus (INF B), and respiratory syncytial virus (RSV) in 170 respiratory specimens from hospitalized patients. RESULTS: Overall, xTAG RVP demonstrated sensitivities and specificities of 97.6 and 100% for INF A, 100 and 99.4% for INF B, and 100 and 100% for RSV, while the Verigene RV+ test sensitivities and specificities were 95.1 and 98.5%, 100.0 and 99.4%, and 97.1 and 100%, respectively. There were no significant differences in the area under the curves between the two assays for each virus (P = 0.364 for INF A, P = 1.000 for INF B, P = 0.317 for RSV). Comparing the results of two assays, discordant results were present mostly due to subtype assignments and identification of coinfections. The detection of viruses was not significantly different (P = 1.000) and the virus/subtype assignment showed good agreement with kappa coefficients of 0.908. CONCLUSION: The xTAG RVP and Verigene RV+ showed high sensitivities and specificities, and good overall agreement in detection and identification of INF and RSV. These assays can be used in clinical settings for a reliable detection of respiratory viruses found commonly in hospitalized patients. |
BACKGROUND: The Eastern Mediterranean Public Health Network, supported by the Biosecurity Engagement Program, contributed significantly to strengthening the preparedness and response to the emerging and re-emerging infections in the region. OBJECTIVE: This study aimed to determine the gaps, challenges, and priorities for preventing the emerging and re-emerging infections, with a focus on biosafety and biosecurity in four countries of the region, namely, Egypt, Iraq, Jordan, and Morocco. METHODS: A total of two different methods were used to determine the gaps and priorities for preventing the emerging and re-emerging infections. The first method was a rapid assessment for the preparedness and response to the emerging and re-emerging infections in four countries of the region, with a focus on biosafety and biosecurity. The second method was a face-to-face round table meeting of the participating teams for two days, where the teams from all countries presented their countries’ profiles, findings, priorities, and gaps based on the countries’ assessments. RESULTS: The assessment and meeting resulted in several priorities and recommendations for each of the countries in the areas of legislation and coordination, biosafety and biosecurity, surveillance and human resources, case management and response, infection control and prevention, and risk communication and laboratory capacity. CONCLUSIONS: Many recommendations were relatively consistent throughout, including improving communication or building collaborations to improve the overall health of the country. |
Little is known about the impact of infectious diseases on large carnivores. We investigated factors structuring the helminth and protozoan infections of wolves (Canis lupus) by using coprological analyses. Faecal samples (n = 342) were analysed from 11 wolf packs belonging to three different geographical and ecological settings in Italy (Abruzzo, Lazio e Molise National Park, PNALM: 4 packs, 88 samples), in France (Mercantour National Park, PNM: 4 packs, 68 samples) and in the U.S.A. (Yellowstone National Park, YNP: 3 packs, 186 samples). Parasites were found in 29.4%–88.6% of the samples and parasite taxa ranged from four to ten in each study area. Taeniidae (Taenia/Echinococcus), Sarcocystis spp. and Toxascaris leonina were most common in faecal samples from YNP, whereas Capillaria spp., Taeniidae and Uncinaria stenocephala were predominant in PNALM. We used generalised linear mixed models to assess the relationship between parasite infection or the number of parasite taxa and selected ecological drivers across study areas. Significant effects illustrated the importance of the ecological factors such as occurrence of free-ranging dogs, diet composition and wolf density, as well as the ancestry of the wolf populations, in shaping parasite-wolf communities. Additional investigations are needed to elucidate the impact of parasitic infections on wolf populations, as well as the role of anthropogenic factors in facilitating parasitic diffusion to apex predators. |
BACKGROUND AND AIM: Determining reference values for hematological and biochemical parameters of Asian house shrew (Suncus murinus) is important for wildlife research to protect human health in surrounding communities. This study aimed to establish the reference values for selected hematology and serum clinical chemistry analyses that may contribute to research on shrew in future. MATERIALS AND METHODS: Blood samples (n=51) were collected from shrews between July and December 2015, Bangladesh, to estimate the levels of hemoglobin (Hb), packed cell volume (PCV), total leukocyte count (TLC), total erythrocyte count (TEC), lymphocyte, monocyte, neutrophil, eosinophil, basophil, calcium, phosphorus (P), sodium (Na), chloride (Cl), urea, glucose, total protein (TP), creatinine, and alanine transaminase (ALT). RESULTS: Although the values did not differ significantly among sexes, age was found to be a significant factor. Hb, PCV, TEC, glucose, and P were higher in males; eosinophil, Na, Cl, TP, and ALT were higher among females. Adults had significantly greater urea and glucose (p<0.05) while juveniles had insignificantly higher values for TLC, PCV, neutrophil, P, and TP. CONCLUSION: This study provides the first reference values for this species in Bangladesh and can be used to guide wildlife research studies. |
The lung plays a vital role in maintaining homeostasis, as it is responsible for the exchange of oxygen and carbon dioxide. Pulmonary homeostasis is maintained by a network of tissue-resident cells, including epithelial cells, endothelial cells and leukocytes. Myeloid cells of the innate immune system and epithelial cells form a critical barrier in the lung. Recently developed unbiased next generation sequencing (NGS) has revealed cell heterogeneity in the lung with respect to physiology and pathology and has reshaped our knowledge. New phenotypes and distinct gene signatures have been identified, and these new findings enhance the diagnosis and treatment of lung diseases. Here, we present a review of the new NGS findings on myeloid cells in lung development, homeostasis, and lung diseases, including acute lung injury (ALI), lung fibrosis, chronic obstructive pulmonary disease (COPD), and lung cancer. |
Overlap extension polymerase chain reaction (PCR) is a powerful technology for DNA assembly. Based on this technology, we synthesized DNA templates, which were transcribed into sgRNA in vitro, and further detected their efficiency of purified sgRNAs with Cas9 nuclease. The sgRNAs synthesized by this approach can effectively cleave the DNA fragments of interest in vitro and in vivo. Compared with the conventional method for generating sgRNA, it does not require construction of recombinant plasmids and design of primers to amplify sgRNA core fragment. Only several short primers with overlapped sequences are needed to assemble a DNA fragment as the template of sgRNA. This modified and simplified method is highly applicable and less time-consuming. |
The delivery of biologic cargoes to airway epithelial cells is challenging due to the formidable barriers imposed by its specialized and differentiated cells. Among cargoes, recombinant proteins offer therapeutic promise but the lack of effective delivery methods limits their development. Here, we achieve protein and SpCas9 or AsCas12a ribonucleoprotein (RNP) delivery to cultured human well-differentiated airway epithelial cells and mouse lungs with engineered amphiphilic peptides. These shuttle peptides, non-covalently combined with GFP protein or CRISPR-associated nuclease (Cas) RNP, allow rapid entry into cultured human ciliated and non-ciliated epithelial cells and mouse airway epithelia. Instillation of shuttle peptides combined with SpCas9 or AsCas12a RNP achieves editing of loxP sites in airway epithelia of ROSA(mT/mG) mice. We observe no evidence of short-term toxicity with a widespread distribution restricted to the respiratory tract. This peptide-based technology advances potential therapeutic avenues for protein and Cas RNP delivery to refractory airway epithelial cells. |
As one of many nonstructural proteins of porcine reproductive and respiratory syndrome virus (PRRSV), nonstructural protein 12 (Nsp12) has received relatively little attention, and its role in virus replication, if any, is essentially unknown. By the application of reverse genetic manipulation of an infectious PRRSV clone, the current study is the first to demonstrate that Nsp12 is a key component of PRRSV replication. In addition, the biochemical properties of Nsp12 were evaluated, revealing that Nsp12 forms dimers when exposed to oxidative conditions. Furthermore, we systemically analyzed the function of Nsp12 in PRRSV RNA synthesis using a strand-specific PCR method. To our surprise, Nsp12 was not found to be involved in minus-strand genomic RNA (-gRNA) synthesis; importantly, our results indicate that Nsp12 is involved in the synthesis of both plus- and minus-strand subgenomic mRNAs (+sgmRNA and -sgmRNA). Finally, we found that the combination of cysteine 35 and cysteine 79 in Nsp12 is required for sgmRNA synthesis. To our knowledge, we are the first to report the biological role of Nsp12 in the PRRSV lifecycle, and we conclude that Nsp12 is involved in the synthesis of both + sgRNA and -sgRNA. |
Peptides are secreted by different cell types and are trendy therapeutic agents that have attracted attention for the treatment of several diseases such as infections. Antimicrobial peptides exert various mechanisms such as changing cell membrane permeability which leads to inhibition or death of bacterial cells. mesenchymal stem cells (MSCs) are key to produce antimicrobial peptides and to inhibit the growth of pathogens. These cells have been shown to be capable of producing antimicrobial peptides upon exposure to different bacteria. As a result, antimicrobial peptides can be considered as novel agents for the treatment of infectious diseases. The purpose of this review was to investigate the targets and mechanisms of antimicrobial peptides secreted by MSCs. |
Asthma exacerbation can be a major life threatening event. Viruses have been pinned as the cause behind the vast majority of these exacerbations. The purpose of this short review is to explore the mechanisms behind these exacerbations, focusing mostly on viral infections as triggers. We will also be discussing the phenotypes prone to asthma exacerbation, the pathophysiology of viral induced asthma and ventilation patterns of asthmatic lungs. This manuscript will assist primary care physicians in delineating the proper pathophysiology of the disease as well as the management. |
Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine that increasingly is being studied in cancers and inflammatory diseases. Though murine models have been instrumental in understanding the functional role of MIF in different pathological conditions, the information obtained from these models is biased towards a specific species. In experimental science, results obtained from multiple clinically relevant animal models always provide convincing data that might recapitulate in humans. Syrian golden hamster (Mesocricetus auratus), is a clinically relevant animal model for multiple human diseases. Hence, the major objectives of this study were to characterize the structure and function of Mesocricetus auratus MIF (MaMIF) and finally evaluate its effect on pancreatic tumor growth in vivo. Initially, the recombinant MaMIF was cloned, expressed and purified in a bacterial expression system. The MaMIF primary sequence, biochemical properties, and crystal structure analysis showed greater similarity with human MIF. The crystal structure of MaMIF illustrates that it forms a homotrimer as known in human and mouse. However, MaMIF exhibits some minor structural variations when compared to human and mouse MIF. The in vitro functional studies show that MaMIF has tautomerase activity and enhances activation and migration of hamster peripheral blood mononuclear cells (PBMCs). Interestingly, injection of MaMIF into HapT1 pancreatic tumor-bearing hamsters significantly enhanced the tumor growth and tumor-associated angiogenesis. Together, the current study shows a structural and functional similarity between the hamster and human MIF. Moreover, it has demonstrated that a high level of circulating MIF originating from non-tumor cells might also promote pancreatic tumor growth in vivo. |
BACKGROUND: Pertussis causes severe disease in young unvaccinated infants, with preterms potentially at highest risk. We studied pertussis in hospitalized infants as related to gestational age (GA) and vaccination history. METHODS: Medical record data of 0-2y old patients hospitalized for pertussis during 2005–2014 were linked to vaccination data. Multivariable logistic regression was used to study the association between GA and vaccination history on the clinical disease course. We compared vaccine effectiveness (VE) against hospitalization for pertussis between term and preterm infants (i.e., <37w GA) using the screening method as developed by Farrington. RESULTS: Of 1187 records, medical data from 676 were retrieved. Of these, 12% concerned preterms, whereas they are 8% of Dutch birth cohorts. Median age at admission was 3 m for preterms and 2 m for terms (p < 0.001). Preterms more often had received pertussis vaccination (62% vs 44%; p = 0.01) and more often had coinfections (37% vs 21%; p = 0.01). Preterms tended more often to have complications, to require artificial respiration or to need admittance to the intensive care unit (ICU). Preterms had longer ICU stays (15d vs 9d; p = 0.004). Vaccinated preterms and terms had a lower median length of hospital stay and lower crude risks of apneas and the need for artificial respiration, additional oxygen, and ICU admittance than those not vaccinated. After adjustment for presence of coinfections and age at admittance, these differences were not significant, except the lower need of oxygen treatment in vaccinated terms. Effectiveness of the first vaccination against pertussis hospitalizations was 95% (95% CI 93–96%) and 73% (95% CI 20–91%) in terms and preterms, respectively. Effectiveness of the second dose of the primary vaccination series was comparable in both groups (86 and 99%, respectively). CONCLUSIONS: Infants hospitalized for pertussis suffer from severe disease. Preterms were overrepresented, with higher need for intensive treatment and less VE of first vaccination. These findings stress the need for alternative prevention, in particular prenatal vaccination of mothers, to reduce pertussis in both groups. |
Non-structural protein 1 (NS1) of influenza virus has been shown to inhibit the innate immune response by blocking the induction of interferon (IFN). In this study, we isolated two single-stranded RNA aptamers specific to NS1 with K(d) values of 1.62 ± 0.30 nM and 1.97 ± 0.27 nM, respectively, using a systematic evolution of ligand by exponential enrichment (SELEX) procedure. The selected aptamers were able to inhibit the interaction of NS1 with tripartite motif-containing protein 25 (TRIM25), and suppression of NS1 enabled retinoic acid inducible gene I (RIG-I) to be ubiquitinated regularly by TRIM25. Additional luciferase reporter assay and quantitative real-time PCR (RT-PCR) experiments demonstrated that suppression of NS1 by the selected aptamers induced IFN production. It is noted that viral replication was also inhibited through IFN induction in the presence of the selected aptamers. These results suggest that the isolated aptamers are strongly expected to be new therapeutic agents against influenza infection. |
[Image: see text] Nucleoside analogues are among the most common medications given for the treatment of viral infections and cancers. The therapeutic effectiveness of nucleoside analogues can be dramatically improved by phosphorylation. The ProTide approach was developed using a phosphorylated nucleoside that is masked by esterification with an amino acid and phenol forming a chiral phosphorus center. The biological activity of the ProTides depends, in part, on the stereochemistry at phosphorus, and thus, it is imperative that efficient methods be developed for the chemical synthesis and isolation of diastereomerically pure ProTides. Chiral ProTides are often synthesized by direct displacement of a labile phenol (p-nitrophenol or pentafluorophenol) from a chiral phosphoramidate precursor with the appropriate nucleoside analogue. The ability to produce these chiral products is dictated by the synthesis of the chiral phosphoramidate precursors. The enzyme phosphotriesterase (PTE) from Pseudomonas diminuta is well-known for its high stereoselectivity and broad substrate profile. Screening PTE variants from enzyme evolution libraries enabled the identification of variants of PTE that can stereoselectively hydrolyze the chiral phosphoramidate precursors. The variant G60A-PTE exhibits a 165-fold preference for hydrolysis of the R(P) isomer, while the variant In1W-PTE has a 1400-fold preference for hydrolysis of the S(P) isomer. Using these mutants of PTE, the S(P) and R(P) isomers were isolated on a preparative scale with no detectable contamination of the opposite isomer. Combining the simplicity of the enzymatic resolution of the precursor with the latest synthetic strategy will facilitate the production of diastereometrically pure nucleotide phosphoramidate prodrugs. |
BACKGROUND: Interleukin-35 (IL-35) is a recently discovered cytokine that plays a role in immune suppression and has therefore been the subject of a great deal of research. A bibliometric analysis of the global research concerning IL-35, however, is rare. OBJECTIVES: The aim of this research was to assess the international scientific output of IL-35 research and explore its hotspots and frontiers from 2009 to 2018 by bibliometric analysis. METHODS: Publications about IL-35 research from 2009 to 2018 were retrieved from the Web of Science Core Collection (WoSCC). Citespace V was used to analyze years, journals, countries, research institutions, areas of exploration, research hotspots, and trends of publication. RESULTS: We retrieved a total of 416 publications and observed a trend of publications increasing over the past decade. Original articles (351) were the most frequently occurring document type. The largest number of publications belonging to one country and one institution, respectively, was China (202) and Tianjin Medical University (17). Trending keywords may indicate frontier topics, including “infectious tolerance,” “autoimmune,” and “central nervous system.” CONCLUSION: This study provides valuable information on the study of IL-35 so that researchers may identify new research fields. |
AIM: Newcastle disease virus (NDV) is a member of genus Avulavirus within the family Paramyxoviridae. Interest of using NDV as an anticancer agent has arisen from its ability to kill tumor cells with limited toxicity to normal cells. METHODS: In this investigation, the proliferation of brain tumor cell line, glioblastoma multiform (DBTRG.05MG) induced by NDV strain AF2240 was evaluated in-vitro, by using MTT proliferation assay. Furthermore, Cytological observations were studied using fluorescence microscopy and transmission electron microscopy, DNA laddering in agarose gel electrophoresis assay used to detect the mode of cell death and analysis of the cellular DNA content by flowcytometery. RESULTS: MTT proliferation assay, Cytological observations using fluorescence microscopy and transmission electron microscopy show the anti-proliferation effect and apoptogenic features of NDV on DBTRG.05MG. Furthermore, analysis of the cellular DNA content showed that there was a loss of treated cells in all cell cycle phases (G1, S and G2/M) accompanied with increasing in sub-G1 region (apoptosis peak). CONCLUSION: It could be concluded that NDV strain AF2240 is a potent antitumor agent that induce apoptosis and its cytotoxicity increasing while increasing of time and virus titer. |
OBJECTIVE: Spontaneous intracerebral hemorrhage (ICH) is caused by the rupture of small blood vessels and other health problems. In ICH patients, hematoma enlargement is the most critical risk factor for poor outcomes. Tranexamic acid, an anti-fibrinolytic agent, has been used to reduce hematoma expansion. We analyzed the risk factors for hematoma expansion in ICH patients and compared the predictability of hematoma expansion in ICH patients with the use of tranexamic acid. METHODS: We performed retrospective analysis of ICH patients who underwent follow-up computed tomography scans from October 2008 to October 2018. Of the 329 included patients, 67 who received tranexamic acid and 262 who did not receive tranexamic acid were compared. We also analyzed the risk factors of 45 and 284 patients who did and did not experience hematoma expansion, respectively. RESULTS: Hematoma expansion was observed in 7 (10.4%) of 67 patients in the tranexamic acid group and 38 (14.5%) of the 262 patients who did not receive tranexamic acid. There was no statistically significant difference between patients who did and did not received tranexamic acid (p=0.389). In the multivariate logistic regression analysis of risk factors for hematoma expansion, spot sign and a maximal diameter of 40 mm were identified as risk factors. CONCLUSION: We could not confirm the effect of tranexamic acid on hematoma expansion in ICH patients. Spot sign and the maximal diameter of hematomas were confirmed as risk factors of hematoma expansion. If the maximal diameter is greater than 40 mm, the hematoma should be closely monitored. |
SIMPLE SUMMARY: Recent research has debated the effects of milk and forage feeding regimes in the first weeks of life on the future performance of dairy calves. However, little is known about how feeding regime can affect behavioural and physiological responses, which have the potential to impact on calf health and well-being. Traditional methods of assessing calf health and welfare such as behavioural observations and blood sampling can be time consuming and impractical for producers and invasive for animals involved. Developments in technology have increased the availability of on-farm non-invasive devices which allow automatic and remote collection of behavioural and physiological data linked to animal health and welfare. This study aimed to use devices to measure lying behaviour, heart rate, heart rate variability and infrared temperature of calves offered high or low levels of milk replacer and different types of forage throughout the first ten weeks of life. Calves displayed changes in lying behaviour and heart rate variability as a result of changes in milk replacer feeding frequency. Additionally, infrared temperature changes were detected during periods of vaccination which corresponded with a rise in core body temperature. Results have highlighted that these sensors can provide important and useable data regarding overall calf well-being on commercial farms. ABSTRACT: This study aimed to examine the use of non-invasive monitoring technologies as a means of capturing behavioural, physiological and health responses of calves allocated to different nutritional regimes. Seventy-four Holstein Friesian calves were individually penned and allocated to receive either high (HML) or conventional (CML) milk replacer (MR) levels between 5–70 days of age. Additionally calves were allocated to one of four forage treatments: (i) chopped straw offered between 14–70 days of age (CS14), (ii) chopped straw offered between 56–70 days of age (CS56), (iii) grass silage offered between 56–70 days of age (GS56), and (iv) no forage in the pre-wean period (NF). A representative sample of calves from each treatment were fitted with activity sensors and heart rate monitors throughout the experimental period to examine lying behaviour and heart rate variability, respectively. Thermal images of the eye and rectal area of each calf were taken 5 days/week between 5–77 days of age. Faecal and respiratory scoring of each individual calf was carried out on a daily basis throughout the experimental period. Milk replacer feeding level had limited effects on measures of calf health, although HML calves tended to have an increased likelihood for receiving treatment for scour than CML calves. Daily lying time (min/d) was lower in HML calves following reduction in MR feeding frequency at 43 days of age and weaning at 71 days of age when compared with CML calves. Additionally, HML calves displayed a lower heart rate variability following weaning, this suggestive of increased stress load. There were limited effects of forage treatment, however, CS14 calves displayed a greater daily lying time following MR step-down at 68 days of age, this potentially indicating increased rumination. Results of the present study highlight the benefits of using remote monitoring technologies as a means of detecting behavioural and physiological changes as a result of nutritional management strategy in individually housed dairy calves. |
The experimental infection of rhesus macaques (rh) with simian immunodeficiency virus (SIV) is an important model for human immunodeficiency virus (HIV) infection of humans. The interferon-induced transmembrane protein 3 (IFITM3) inhibits HIV and SIV infection at the stage of host cell entry. However, it is still unclear to what extent the antiviral activity of IFITM3 observed in cell culture translates into inhibition of HIV/SIV spread in the infected host. We have shown previously that although rhIFITM3 inhibits SIV entry into cultured cells, polymorphisms in the rhIFITM3 gene are not strongly associated with viral load or disease progression in SIV infected macaques. Here, we examined whether rhIFITM3(2), which is closely related to rhIFITM3 at the sequence level, exerts antiviral activity and whether polymorphisms in the rhIFITM3(2) gene impact the course of SIV infection. We show that expression of rhIFITM3(2) is interferon-inducible and inhibits SIV entry into cells, although with reduced efficiency as compared to rhIFITM3. We further report the identification of 19 polymorphisms in the rhIFITM3(2) gene. However, analysis of a well characterized cohort of SIV infected macaques revealed that none of the polymorphisms had a significant impact upon the course of SIV infection. These results and our previous work suggest that polymorphisms in the rhIFITM3 and rhIFITM3(2) genes do not strongly modulate the course of SIV infection in macaques. |
We prospectively analyzed outcomes of haploidentical hematopoietic stem cell transplantation using αβ(+) T-cell receptor/CD19(+) depleted grafts. Sixty-three transplantations were performed in 60 patients. Twenty-eight patients were diagnosed with acute lymphoblastic leukemia (ALL), 27 patients were diagnosed with acute myelogenous leukemia, and in eight other hematological malignancies were diagnosed. Twenty-three were in first complete remission (CR), 20 in second CR, 20 beyond second CR. Four patients developed graft failure. Median time to neutrophil and platelet recovery was 14 (range 9–25) and 10 days (range 7–30), respectively. The probability of non-relapse mortality (NRM) by day +100 after transplantation was 10 ± 4%. With a median follow-up of 28 months, the probability of relapse was 32 ± 6% and disease-free survival was 52 ± 6%. Immune reconstitution was leaded by NK cells. As such, a high CD56(dim/)CD56(bright) NK cell ratio early after transplantation was associated with better disease-free survival (DFS) (≥3.5; 77 ± 8% vs. <3.5; 28 ± 5%; p = 0.001) due to lower relapse incidence (≥3.5; 15 ± 7% vs. <3.5; 37 ± 9%; p = 0.04). T-cell reconstitution was delayed and associated with severe infections after transplant. Viral reactivation/disease and presence of venooclusive disease of liver in the non-caucasian population had a significant impact on NRM. αβ(+) T-cell receptor/CD19(+) cell-depleted haploidentical transplant is associated with good outcomes especially in patients in early phase of disease. A rapid expansion of “mature” natural killer cells early after transplantation resulted on lower probability of relapse, suggesting a graft vs. leukemia effect independent from graft-vs.-host reactions. |
Here, we show that the US Food and Drug Administration-approved oral drug nitazoxanide (NTZ) broadly amplifies the host innate immune response to viruses and inhibits Ebola virus (EBOV) replication. We find that NTZ enhances retinoic-acid-inducible protein I (RIG-I)-like-receptor, mitochondrial antiviral signaling protein, interferon regulatory factor 3, and interferon activities and induces transcription of the antiviral phosphatase GADD34. NTZ significantly inhibits EBOV replication in human cells through its effects on RIG-I and protein kinase R (PKR), suggesting that it counteracts EBOV VP35 protein's ability to block RIG-I and PKR sensing of EBOV. NTZ also inhibits a second negative-strand RNA virus, vesicular stomatitis virus (VSV), through RIG-I and GADD34, but not PKR, consistent with VSV's distinct host innate immune evasion mechanisms. Thus, NTZ counteracts varied virus-specific immune evasion strategies by generally enhancing the RNA sensing and interferon axis that is triggered by foreign cytoplasmic RNA exposure, and holds promise as an oral therapy against EBOV. |
Hepatitis E virus (HEV) infection is a major cause of acute hepatitis worldwide. It is transmitted enterically but replicates in the liver. Recent studies indicate that HEV exists in two forms: naked, nonenveloped virions that are shed into feces to mediate inter-host transmission, and membrane-cloaked, quasienveloped virions that circulate in the bloodstream to mediate virus spread within a host. Both virion types are infectious, but differ in the way they infect cells. Elucidating the entry mechanism for both virion types is essential to understand HEV biology and pathogenesis, and is relevant to the development of treatments and preventions for HEV. This review summarizes the current understanding of the cell entry mechanism for these two HEV virion types. |
The papain-like cysteine protease 2 (PLP2) within the N-terminus of the porcine reproductive and respiratory syndrome virus (PRRSV) nsp2 replicase protein specifies a deubiquitinating enzyme (DUB), but its biochemical properties and the role in infection have remained poorly defined. By using in vitro assays, we found that the purified PLP2 could efficiently cleave K63 and K48 linked polyubiquitin chains Ub3-7 in vitro although displaying a differential activity in converting the respective ubiquitin dimers to monomer. The subsequent mutagenesis analyses revealed that the requirement for PLP2 DUB activity surprisingly resembled that for cis-cleavage activity, as several mutations (e.g., D91R, D85R, etc.) that largely ablated the DUB function also blocked the cis- but not trans-proteolytic cleavage of nsp2/3 polyprotein. Moreover, the analyses identified key mutations that could differentiate DUB from PLP2 cis- and trans-cleavage activities. Further reverse genetics analyses revealed the following findings: (i) mutations that largely blocked the DUB activity were all lethal to the virus, (ii) a point mutation T88G that selectively blocked the cis-cleavage activity of PLP2 did not affect viral viability in cell culture, and (iii) an E90Q mutation that did not affect either of the PLP2 activities led to rescue of WT-like virus but displayed significantly reduced ability to induce TNF-α production. Our findings support the possibility that the PLP2 DUB activity, but not cis-cleavage activity, is essential for PRRSV replication. The data also establish a strong link of nsp2 to pro-inflammatory cytokine induction during infection that operates in a manner independent of PLP2 DUB activity. |
Non-alcoholic fatty liver disease (NAFLD) is a chronic liver disease found worldwide. The present study aimed to evaluate the mechanisms of inhibiting lipid accumulation in free fatty acid (FFA)-treated HepG2 cells caused by bark and fruit extracts of Toona sinensis (TSB and TSF). FFA induced lipid and triglyceride (TG) accumulation, which was attenuated by TSB and TSF. TSB and/or TSF promoted phosphorylation of AMP-activated protein kinase (AMPK) and acetyl-coA carboxylase and peroxisome proliferator-activated receptor alpha upregulation. Furthermore, TSB and TSF suppressed FFA-induced liver X receptor, sterol regulatory element-binding transcription protein 1, fatty acid synthase, and stearoyl-CoA desaturase 1 protein expression. Moreover, TSB and/or TSF induced phosphorylation of Unc-51 like autophagy-activating kinase and microtubule-associated protein 1A/1B-light chain 3 expressions. Therefore, TSB and TSF relieve lipid accumulation by attenuating lipogenic protein expression, activating the AMPK pathway, and upregulating the autophagic flux to enhance lipid metabolism. Moreover, TSB and TSF reduced TG contents, implying the therapeutic use of TSB and TSF in NAFLD. |
In recent years, affective computing has been actively researched to provide a higher level of emotion-awareness. Numerous studies have been conducted to detect the user’s emotions from physiological data. Among a myriad of target emotions, boredom, in particular, has been suggested to cause not only medical issues but also challenges in various facets of daily life. However, to the best of our knowledge, no previous studies have used electroencephalography (EEG) and galvanic skin response (GSR) together for boredom classification, although these data have potential features for emotion classification. To investigate the combined effect of these features on boredom classification, we collected EEG and GSR data from 28 participants using off-the-shelf sensors. During data acquisition, we used a set of stimuli comprising a video clip designed to elicit boredom and two other video clips of entertaining content. The collected samples were labeled based on the participants’ questionnaire-based testimonies on experienced boredom levels. Using the collected data, we initially trained 30 models with 19 machine learning algorithms and selected the top three candidate classifiers. After tuning the hyperparameters, we validated the final models through 1000 iterations of 10-fold cross validation to increase the robustness of the test results. Our results indicated that a Multilayer Perceptron model performed the best with a mean accuracy of 79.98% (AUC: 0.781). It also revealed the correlation between boredom and the combined features of EEG and GSR. These results can be useful for building accurate affective computing systems and understanding the physiological properties of boredom. |
BACKGROUND: The sepsis-induced immunodepression contributes to impaired clinical outcomes of various stress conditions. This syndrome is well documented and characterized by attenuated function of innate and adaptive immune cells. Several pharmacological interventions aimed to restore the immune response are emerging of which interferon-gamma (IFNγ) is one. It is of paramount relevance to obtain clinical information on optimal timing of the IFNγ-treatment, −tolerance, −effectiveness and outcome before performing a RCT. We describe the effects of IFNγ in a cohort of 18 adult and 2 pediatric sepsis patients. METHODS: In this open-label prospective multi-center case-series, IFNγ treatment was initiated in patients selected on clinical and immunological criteria early (< 4 days) or late (> 7 days) following the onset of sepsis. The data collected in 18 adults and 2 liver transplanted pediatric patients were: clinical scores, monocyte expression of HLA-DR (flow cytometry), lymphocyte immune-phenotyping (flow cytometry), IL-6 and IL-10 plasma levels (ELISA), bacterial cultures, disease severity, and mortality. RESULTS: In 15 out of 18 patients IFNγ treatment was associated with an increase of median HLA-DR expression from 2666 [IQ 1547; 4991] to 12,451 [IQ 4166; 19,707], while the absolute number of lymphocyte subpopulations were not affected, except for the decrease number of NK cells 94.5 [23; 136] to 32.5 [13; 90.8] (0.0625)]. Plasma levels of IL-6 464 [201–770] to 108 (89–140) ng/mL (p = 0.04) and IL-10 from IL-10 from 29 [12–59] to 9 [1–15] pg/mL decreased significantly. Three patients who received IFNγ early after ICU admission (<4 days) died. The other patients had a rapid clinical improvement assessed by the SOFA score and bacterial cultures that were repeatedly positive became negative. The 2 pediatric cases improved rapidly, but 1 died for hemorrhagic complication. CONCLUSION: Guided by clinical and immunological monitoring, adjunctive immunotherapy with IFNγ appears well-tolerated in our cases and improves immune host defense in sepsis induced immuno suppression. Randomized clinical studies to assess its potential clinical benefit are warranted. |
Metabolomics aims to measure and characterise the complex composition of metabolites in a biological system. Metabolomics studies involve sophisticated analytical techniques such as mass spectrometry and nuclear magnetic resonance spectroscopy, and generate large amounts of high-dimensional and complex experimental data. Open source processing and analysis tools are of major interest in light of innovative, open and reproducible science. The scientific community has developed a wide range of open source software, providing freely available advanced processing and analysis approaches. The programming and statistics environment R has emerged as one of the most popular environments to process and analyse Metabolomics datasets. A major benefit of such an environment is the possibility of connecting different tools into more complex workflows. Combining reusable data processing R scripts with the experimental data thus allows for open, reproducible research. This review provides an extensive overview of existing packages in R for different steps in a typical computational metabolomics workflow, including data processing, biostatistics, metabolite annotation and identification, and biochemical network and pathway analysis. Multifunctional workflows, possible user interfaces and integration into workflow management systems are also reviewed. In total, this review summarises more than two hundred metabolomics specific packages primarily available on CRAN, Bioconductor and GitHub. |
Cynomolgus macaques (Macaca fascicularis, Mafa) have been used as important experimental animal models for carrying out biomedical researches. The results of biomedical experiments strongly depend on the immunogenetic background of animals, especially on the diversity of major histocompatibility complex (MHC) alleles. However, there is much less information available on the polymorphism of MHC class I genes in cynomolgus macaques, than is currently available for humans. In this study, we have identified 40 Mafa-A and 60 Mafa-B exons 2 and 3 sequences from 30 unrelated cynomolgus macaques of Vietnamese origin. Among these alleles, 28 are novel. As for the remaining 72 known alleles, 15 alleles are shared with other cynomolgus macaque populations and 32 are identical to alleles previously reported in other macaque species. A potential recombination event was observed between Mafa-A1*091:02 and Mafa-A1*057:01. In addition, the Mafa-A1 genes were found to be more diverse than human HLA-A and the functional residues for peptide binding sites (PBS) or TCR binding sites (TBS) in Mafa-A1 have greater variability than that for non-PBS or non-TBS regions. Overall, this study provides important information on the diversity of Mafa-A and Mafa-B alleles from Vietnamese origin, which may help researchers to choose the most appropriate animals for their studies. |
BACKGROUND: The Hindu Kush and Karakoram mountain ranges in Pakistan’s northern areas are a natural habitat of the snow leopard (Panthera uncia syn. Uncia uncia) but the ecological studies on this animal are scarce since it is human shy by nature and lives in difficult mountainous tracts. The pilot study is conducted to exploit the genetic diversity and population structure of the snow leopard in this selected natural habitat of the member of the wildcat family in Pakistan. METHOD: About 50 putative scat samples of snow leopard from five localities of Gilgit-Baltistan (Pakistan) along with a control sample of zoo maintained male snow leopard were collected for comparison. Significant quality and quantity of genomic DNA was extracted from scat samples using combined Zhang–phenol–chloroform method and successful amplification of cytochrome c oxidase I gene (190 bp) using mini-barcode primers, seven simple sequence repeats (SSR) markers and Y-linked AMELY gene (200 bp) was done. RESULTS: Cytochrome c oxidase I gene sequencing suggested that 33/50 (66%) scat samples were of snow leopard. AMELY primer suggested that out of 33 amplified samples, 21 (63.63%) scats were from male and 12 (36.36%) from female leopards. Through successful amplification of DNA of 25 out of 33 (75.75%) scat samples using SSR markers, a total of 68 alleles on seven SSR loci were identified, showing low heterozygosity, while high gene flow between population. DISCUSSION: The low gene flow rate among the population results in low genetic diversity causing decreased diversification. This affects the adaptability to climatic changes, thus ultimately resulting in decreased population size of the species. |
Lower respiratory tract infections (LRTIs) refer to the inflammation of the trachea, bronchi, bronchioles, and lung tissue. Old people have an increased risk of developing LRTIs compared to young adults. The prevalence of LRTIs in the elderly population is not only related to underlying diseases and aging itself, but also to a variety of clinical issues, such as history of hospitalization, previous antibacterial therapy, mechanical ventilation, antibiotic resistance. These factors mentioned above have led to an increase in the prevalence and mortality of LRTIs in the elderly, and new medical strategies targeting LRTIs in this population are urgently needed. After a systematic review of the current randomized controlled trials and related studies, we recommend novel pharmacotherapies that demonstrate advantages for the management of LRTIs in people over the age of 65. We also briefly reviewed current medications for respiratory communicable diseases in the elderly. Various sources of information were used to ensure all relevant studies were included. We searched Pubmed, MEDLINE (OvidSP), EMBASE (OvidSP), and ClinicalTrials.gov. Strengths and limitations of these drugs were evaluated based on whether they have novelty of mechanism, favorable pharmacokinetic/pharmacodynamic profiles, avoidance of interactions and intolerance, simplicity of dosing, and their ability to cope with challenges which was mainly evaluated by the primary and secondary endpoints. The purpose of this review is to recommend the most promising antibiotics for treatment of LRTIs in the elderly (both in hospital and in the outpatient setting) based on the existing results of clinical studies with the novel antibiotics, and to briefly review current medications for respiratory communicable diseases in the elderly, aiming to a better management of LRTIs in clinical practice. |
Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) is the most important endemic pathogen in the U.S. swine industry. Despite control efforts involving improved biosecurity and different vaccination protocols, the virus continues to circulate and evolve. One of the foremost challenges in its control is high levels of genetic and antigenic diversity. Here, we quantify the co-circulation, emergence and sequential turnover of multiple PRRSV lineages in a single swine-producing region in the United States over a span of 9 years (2009–2017). By classifying over 4,000 PRRSV sequences (open-reading frame 5) into phylogenetic lineages and sub-lineages, we document the ongoing diversification and temporal dynamics of the PRRSV population, including the rapid emergence of a novel sub-lineage that appeared to be absent globally pre-2008. In addition, lineage 9 was the most prevalent lineage from 2009 to 2010, but its occurrence fell to 0.5% of all sequences identified per year after 2014, coinciding with the emergence or re-emergence of lineage 1 as the dominant lineage. The sequential dominance of different lineages, as well as three different sub-lineages within lineage 1, is consistent with the immune-mediated selection hypothesis for the sequential turnover in the dominant lineage. As host populations build immunity through natural infection or vaccination toward the most common variant, this dominant (sub-) lineage may be replaced by an emerging variant to which the population is more susceptible. An analysis of patterns of non- synonymous and synonymous mutations revealed evidence of positive selection on immunologically important regions of the genome, further supporting the potential that immune-mediated selection shapes the evolutionary and epidemiological dynamics for this virus. This has important implications for patterns of emergence and re-emergence of genetic variants of PRRSV that have negative impacts on the swine industry. Constant surveillance on PRRSV occurrence is crucial to a better understanding of the epidemiological and evolutionary dynamics of co-circulating viral lineages. Further studies utilizing whole genome sequencing and exploring the extent of cross-immunity between heterologous PRRS viruses could shed further light on PRRSV immunological response and aid in developing strategies that might be able to diminish disease impact. |
Background: The genus Orthobornavirus comprises non-segmented, negative-stranded RNA viruses able to infect humans, mammals, reptiles and various birds. Parrot bornavirus 1 to 8 (PaBV-1 to 8) causes neurological and/or gastrointestinal syndromes and death on psittacines. We aimed to identify and to produce epidemiologic knowledge about the etiologic agent associated with a death of two female Psittacus erithacus (grey parrot). Methods and Results: Both parrots were submitted for a complete standardised necropsy. Tissue samples were analysed by PCR. The findings in necropsy were compatible with bornavirus infection. Analysis revealed PaBV-4 related with genotypes detected in captive and in wild birds. The N and X proteins of PaBV-4 were more related to avian bornaviruses, while phosphoprotein was more related to variegated squirrel bornavirus 1 (VSBV-1). Within the P gene/phosphoprotein a highly conserved region between and within bornavirus species was found. Conclusions: Portugal is on the routes of the intensive world trade of psittacines. Broad screening studies are required to help understanding the role of wild birds in the emergence and spread of pathogenic bornaviruses. PaBV-4 phosphoprotein is closer to VSBV-1 associated with lethal encephalitis in humans than with some of the avian bornaviruses. The highly conserved P gene/phosphoprotein region is a good target for molecular diagnostics screenings. |
BACKGROUND: Infection by multidrug-resistant (MDR) pathogens leads to poor patient outcomes in intensive care units (ICUs). Contact precautions are necessary to reduce the transmission of MDR pathogens. However, the importance of the surrounding environment is not well known. We studied the effects of ICU relocation on MDR respiratory pathogen detection rates and patient outcomes. METHODS: Patients admitted to the ICU before and after the relocation were retrospectively analyzed. Baseline patient characteristics, types of respiratory pathogens detected, antibiotics used, and patient outcomes were measured. RESULTS: A total of 463 adult patients admitted to the ICU, 4 months before and after the relocation, were included. Of them, 234 were admitted to the ICU before the relocation and 229 afterward. Baseline characteristics, including age, sex, and underlying comorbidities, did not differ between the two groups. After the relocation, the incidence rate of MDR respiratory pathogen detection decreased from 90.0 to 68.8 cases per 1,000 patient-days, but that difference was statistically insignificant. The use of colistin was significantly reduced from 53.5 days (95% confidence interval [CI], 20.3 to 86.7 days) to 18.7 days (95% CI, 5.6 to 31.7 days). Furthermore, the duration of hospital stay was significantly reduced from a median of 29 days (interquartile range [IQR], 14 to 50 days) to 21 days (IQR, 11 to 39 days). CONCLUSIONS: Incidence rates of MDR respiratory pathogen detection were not significantly different before and after ICU relocation. However, ICU relocation could be helpful in reducing the use of antibiotics against MDR pathogens and improving patient outcomes. |
AIM: Evaluation of the thermal and physical conditions for inactivation of adenovirus (AdV), porcine sapelovirus 1 (PSV1) and the economically important viruses porcine epidemic diarrhoea virus (PEDV) and porcine circovirus 2 (PCV2) in the production of spray‐dried porcine plasma (SDPP). METHODS AND RESULTS: Citrate‐treated porcine plasma of pH 7·5, 9·8 and 10·2 (8·5% dry‐matter) was spiked with PEDV, PSV1, PCV2 and AdV and incubated at 3°C for maximum 24 h, and at 44 or 48°C for maximum 10 min (Experiment 1). Spiked citrate‐treated concentrated plasma of pH 7·5 and 9·8 (24% dry‐matter) was spray dried in a laboratory scale apparatus (Experiment 2). Aliquots of SDPP were stored over a period of 0–10 weeks at 11 and 20°C (Experiment 3). Reverse transcription(RT)‐quantitative PCR detected no notable reduction in viral genomes in treated plasma and SDPP samples. No infectious PSV1 was re‐isolated from plasma and SDPP samples in cell culture. At pH 10·2 and 3°C, infectivity of PEDV in plasma was reduced with a reduction factor of 4·2 log 10 (LRF) at 10 h contact time, whereas heating to 44°C for at least 1 min at alkali pH was needed to achieve a LRF of 4·2 for AdV. Spray drying at an outlet temperature of 80°C reduced AdV infectivity effectively (LRF = 5·2) and PEDV infectivity for 95% (LRF = 1·4). After storage at 20°C for 2 weeks no infectious PEDV was re‐isolated from SDPP anymore (LRF ≥4·0). Due to growth of antibiotic‐resistant bacteria from plasma in cell cultures used for PCV2 isolation, no data regarding inactivation of PCV2 were obtained. CONCLUSIONS: Five percent of PEDV stayed infectious after our spray drying conditions. Spray drying in combination with storage for ≥2 weeks at 20°C eliminated infectivity of PEDV effectively. SIGNIFICANCE AND IMPACT OF THE STUDY: The conditions for inactivation of virus in plasma and SDPP determined are important for producers to inactivate PEDV during production of SDPP. |
For more than 50 years there has been an ongoing effort to combat transfusion‐transmitted infections and provide patients with the safest possible blood. This initiative has driven much of the research within the transfusion community. Initial methods included screening donors for travel histories to banned areas and for high‐risk behaviors, but pathogen‐specific assays performed at the collection and manufacturing sites also have become key factors in assuring blood safety. Many of these have focused on donor and laboratory‐based screening for transfusion‐transmitted diseases, as evidenced by the hepatitis and human immunodeficiency virus screening in the 1970s, 1980s, and 1990s. More recently, this effort has expanded to develop donor screening assays to identify other blood‐borne pathogens, such as Zika and West Nile viruses and Babesia. Bacterial contamination of units of platelets (PLTs), however, remains a significant concern. In recent years, the Food and Drug Administration has approved rapid tests to identify bacterially contaminated PLT units in the blood bank before transfusion. Other supplemental methods have been developed, however, that aim to inactivate blood‐borne pathogen(s) present in the blood product, rather than to rely on our ability to identify and interdict contaminated and infected components. Pathogen reduction technology, as this is referred to, provides a proactive way to further reduce the risk posed by transfusion‐transmitted infections. |
In this work, we study the environmental and operational factors that influence airborne transmission of nosocomial infections. We link a deterministic zonal ventilation model for the airborne distribution of infectious material in a hospital ward, with a Markovian multicompartment SIS model for the infection of individuals within this ward, in order to conduct a parametric study on ventilation rates and their effect on the epidemic dynamics. Our stochastic model includes arrival and discharge of patients, as well as the detection of the outbreak by screening events or due to symptoms being shown by infective patients. For each ventilation setting, we measure the infectious potential of a nosocomial outbreak in the hospital ward by means of a summary statistic: the number of infections occurred within the hospital ward until end or declaration of the outbreak. We analytically compute the distribution of this summary statistic, and carry out local and global sensitivity analysis in order to identify the particular characteristics of each ventilation regime with the largest impact on the epidemic spread. Our results show that ward ventilation can have a significant impact on the infection spread, especially under slow detection scenarios or in overoccupied wards, and that decreasing the infection risk for the whole hospital ward might increase the risk in specific areas of the health‐care facility. Moreover, the location of the initial infective individual and the protocol in place for outbreak declaration both form an interplay with ventilation of the ward. |
Transmissible gastroenteritis (TGE), caused by transmissible gastroenteritis virus (TGEV), is one many gastrointestinal inflections in piglets, characterized by diarrhea, and high mortality. Probiotics are ubiquitous bacteria in animal intestines, which have many functions, such as promoting intestinal peristalsis and maintaining the intestinal balance. We found that the supernatant of the Lp-1 strain of Lactobacillus plantarum, isolated in our laboratory, and named Lp-1s had marked anti-TGEV effect on IPEC-J2 cells. Lp-1s could induce large amounts of interferon-β in IPEC-J2 cells in the early stage (6 h) of infection with TGEV, and increased the level of phosphorylated signal transducer and activator of transcription and its nuclear translocation in the late stage (24–48 h) of infection. This resulted in upregulated expression of interferon-stimulated genes, and increased the transcription and protein expression of antiviral proteins, resulting in an anti-TGEV effect. |
OBJECTIVE: To develop a method to perform multiple tests on single nasopharyngeal (NP) swab. STUDY DESIGN: We collected a NP swab on children aged 2–12 years with acute sinusitis and processed it for bacterial culture, viruses, cytokine expression, and 16S ribosomal RNA gene sequencing analysis. During the course of the study, we expand the scope of evaluation to include RNA sequencing, which we accomplished by cutting the tip of the swab. RESULTS: Of the 174 children enrolled, 126 (72.4%) had a positive bacterial culture and 121(69.5%) tested positive for a virus. Cytokine measurement, as judged by the adequate levels of a housekeeping enzyme (GAPDH), appeared successful. From the samples used for 16S ribosomal sequencing we recovered, on average, 16,000 sequences per sample, accounting for a total of 2,646 operational taxonomic units across all samples sequenced. Samples used for RNA sequencing had a mean RNA Integrity number of 6.0. Cutting the tip of the swab did not affect the recovery yield for viruses or bacteria, nor did it affect species richness in microbiome analysis. CONCLUSION: We describe a minimally invasive sample collection protocol that allows for multiple diagnostic and research investigations in young children. |
Glucose-6-phosphate dehydrogenase (G6PD) is the rate-limiting enzyme of the pentose phosphate pathway that modulates cellular redox homeostasis via the regeneration of NADPH. G6PD-deficient cells have a reduced ability to induce the innate immune response, thus increasing host susceptibility to pathogen infections. An important part of the immune response is the activation of the inflammasome. G6PD-deficient peripheral blood mononuclear cells (PBMCs) from patients and human monocytic (THP-1) cells were used as models to investigate whether G6PD modulates inflammasome activation. A decreased expression of IL-1β was observed in both G6PD-deficient PBMCs and PMA-primed G6PD-knockdown (G6PD-kd) THP-1 cells upon lipopolysaccharide (LPS)/adenosine triphosphate (ATP) or LPS/nigericin stimulation. The pro-IL-1β expression of THP-1 cells was decreased by G6PD knockdown at the transcriptional and translational levels in an investigation of the expression of the inflammasome subunits. The phosphorylation of p38 MAPK and downstream c-Fos expression were decreased upon G6PD knockdown, accompanied by decreased AP-1 translocation into the nucleus. Impaired inflammasome activation in G6PD-kd THP-1 cells was mediated by a decrease in the production of reactive oxygen species (ROS) by NOX signaling, while treatment with hydrogen peroxide (H(2)O(2)) enhanced inflammasome activation in G6PD-kd THP-1 cells. G6PD knockdown decreased Staphylococcus aureus and Escherichia coli clearance in G6PD-kd THP-1 cells and G6PD-deficient PBMCs following inflammasome activation. These findings support the notion that enhanced pathogen susceptibility in G6PD deficiency is, in part, due to an altered redox signaling, which adversely affects inflammasome activation and the bactericidal response. |
Lassa virus (LASV) is the causative agent of a fatal hemorrhagic fever in humans. The glycoprotein (GP) of LASV mediates viral entry into host cells, and correct processing and modification of GP by host factors is a prerequisite for virus replication. Here, using an affinity purification-coupled mass spectrometry (AP-MS) strategy, 591 host proteins were identified as interactors of LASV GP. Gene ontology analysis was performed to functionally annotate these proteins, and the oligosaccharyltransferase (OST) complex was highly enriched. Functional studies conducted by using CRISPR-Cas9-mediated knockouts showed that STT3A and STT3B, the two catalytically active isoforms of the OST complex, are essential for the propagation of the recombinant arenavirus rLCMV/LASV glycoprotein precursor, mainly via affecting virus infectivity. Knockout of STT3B, but not STT3A, caused hypoglycosylation of LASV GP, indicating a preferential requirement of LASV for the STT3B-OST isoform. Furthermore, double knockout of magnesium transporter 1 (MAGT1) and tumor suppressor candidate 3 (TUSC3), two specific subunits of STT3B-OST, also caused hypoglycosylation of LASV GP and affected virus propagation. Site-directed mutagenesis analysis revealed that the oxidoreductase CXXC active-site motif of MAGT1 or TUSC3 is essential for the glycosylation of LASV GP. NGI-1, a small-molecule OST inhibitor, can effectively reduce virus infectivity without affecting cell viability. The STT3B-dependent N-glycosylation of GP is conserved among other arenaviruses, including both the Old World and New World groups. Our study provided a systematic view of LASV GP-host interactions and revealed the preferential requirement of STT3B for LASV GP N-glycosylation. IMPORTANCE Glycoproteins play vital roles in the arenavirus life cycle by facilitating virus entry and participating in the virus budding process. N-glycosylation of GPs is responsible for their proper functioning; however, little is known about the host factors on which the virus depends for this process. In this study, a comprehensive LASV GP interactome was characterized, and further study revealed that STT3B-dependent N-glycosylation was preferentially required by arenavirus GPs and critical for virus infectivity. The two specific thioredoxin subunits of STT3B-OST MAGT1 and TUSC3 were found to be essential for the N-glycosylation of viral GP. NGI-1, a small-molecule inhibitor of OST, also showed a robust inhibitory effect on arenavirus. Our study provides new insights into LASV GP-host interactions and extends the potential targets for the development of novel therapeutics against Lassa fever in the future. |
Zika virus (ZIKV) infection during human pregnancy may lead to severe fetal pathology and debilitating impairments in offspring. However, the majority of infections are subclinical and not associated with evident birth defects. Potentially detrimental life-long health outcomes in asymptomatic offspring evoke high concerns. Thus, animal models addressing sequelae in offspring may provide valuable information. To induce subclinical infection, we inoculated selected porcine fetuses at the mid-stage of development. Inoculation resulted in trans-fetal virus spread and persistent infection in the placenta and fetal membranes for two months. Offspring did not show congenital Zika syndrome (e.g., microcephaly, brain calcifications, congenital clubfoot, arthrogryposis, seizures) or other visible birth defects. However, a month after birth, a portion of offspring exhibited excessive interferon alpha (IFN-α) levels in blood plasma in a regular environment. Most affected offspring also showed dramatic IFN-α shutdown during social stress providing the first evidence for the cumulative impact of prenatal ZIKV exposure and postnatal environmental insult. Other eleven cytokines tested before and after stress were not altered suggesting the specific IFN-α pathology. While brains from offspring did not have histopathology, lesions, and ZIKV, the whole genome expression analysis of the prefrontal cortex revealed profound sex-specific transcriptional changes that most probably was the result of subclinical in utero infection. RNA-seq analysis in the placenta persistently infected with ZIKV provided independent support for the sex-specific pattern of in utero-acquired transcriptional responses. Collectively, our results provide strong evidence that two hallmarks of fetal ZIKV infection, altered type I IFN response and molecular brain pathology can persist after birth in offspring in the absence of congenital Zika syndrome. |
Since the progression of cirrhosis is accelerated each time a complication recurs, the management and treatment of the complication is critical in enhancement of the quality of life and expectation of life in patients. The use of model for end-stage liver disease with incorporation of serum-sodium (MELD-Na) with physiological indicators can be used to assess severity and differentiate therapeutic interventions. This study is aimed to determine the mean survival period and cumulative survival rate by classifying patients into high-risk and low-risk groups based on MELD-Na, a predictor of mortality in liver disease, and to investigate the mortality prognostic factors. A retrospective cohort study, which follows the STROBE checklist, was performed. 263 patients who were diagnosed with liver cirrhosis complications for the first time and hospitalized were selected as the subjects of this study. The collected data were analyzed based on the survival package provided by the statistical program R version 3.4.2. Subjects were classified into high-risk and low-risk groups using MELD-Na 14 points where sensitivity and specificity crossed the cut-off point. Gender, age, and primary caregiver were significant variables in the mortality high-risk group, and AST, albumin, and primary caregiver were significant variables in the mortality low-risk group. Based on these mortality prognostic factors, it is possible to present the factors affecting mortality in patients who were diagnosed with liver cirrhosis complications for the first time. The classification of patients by risk level could be the foundation to provide accurate guidelines for management and it is necessary to modify prognostic factors and apply nursing interventions to manage complications. |
Lectins are glycan-binding proteins with no catalytic activity and ubiquitously expressed in nature. Numerous bacteria use lectins to efficiently bind to epithelia, thus facilitating tissue colonisation. Wounded skin is one of the preferred niches for Pseudomonas aeruginosa, which has developed diverse strategies to impair tissue repair processes and promote infection. Here, we analyse the effect of the P. aeruginosa fucose-binding lectin LecB on human keratinocytes and demonstrate that it triggers events in the host, upon binding to fucosylated residues on cell membrane receptors, which extend beyond its role as an adhesion molecule. We found that LecB associates with insulin-like growth factor-1 receptor and dampens its signalling, leading to the arrest of cell cycle. In addition, we describe a novel LecB-triggered mechanism to down-regulate host cell receptors by showing that LecB leads to insulin-like growth factor-1 receptor internalisation and subsequent missorting towards intracellular endosomal compartments, without receptor activation. Overall, these data highlight that LecB is a multitask virulence factor that, through subversion of several host pathways, has a profound impact on keratinocyte proliferation and survival. |
PURPOSE: This study evaluated the specificity of different avian secondary antibodies used in Western blot and dot-blot ELISA to detect avian bornavirus antibodies in bird plasma. METHODS: Plasma samples were collected from: two Blue and gold macaws, one positive and one negative for avian bornavirus by RT-PCR; a Cockatiel and a Monk parakeet prior to and following experimental infection; and, two Mallards, one positive and one negative for avian bornavirus by RT-PCR Samples were analyzed by Western blot and dot-blot ELISA that incorporated recombinant avian bornavirus nucleoprotein as the target analyte. Four species-specific anti-IgY secondary antibodies were used in the assays: goat anti-macaw IgY, goat anti-bird IgY, goat anti-duck IgY, and rabbit anti-chicken IgY. RESULTS: In the Western blot, anti-macaw IgY secondary antibody produced strong signals with Blue and gold macaw and Cockatiel positive plasma, but no signal with Mallard positive plasma. Anti-bird IgY secondary antibody produced strong signals with Blue and gold macaw, Cockatiel, and Mallard positive plasma. Anti-duck and anti-chicken IgY secondary antibody produced a strong and moderate signal, respectively, only with Mallard positive plasma. In the dot-blot ELISA, there was a distinct and significant difference (P<0.05) in the signal intensity between the different secondary antibodies within a bird species. Anti-macaw IgY secondary antibody produced significantly (P<0.05) stronger signals than the other secondary antibodies in Blue and gold macaw, Cockatiel, and Monk parakeet positive plasma, while anti-duck IgY secondary antibody produced significantly (P<0.05) stronger signals than the other secondary antibodies in Mallard positive plasma. CONCLUSION: In testing psittacines with immunoassays, and especially in assays that incorporate short incubation reaction times such as a dot-blot ELISA, species-specific anti-IgY secondary antibodies provided more accurate results. |
The Crandell-Rees Feline Kidney Cell (CRFK) is an immortalised cell line derived from the feline kidney that is utilised for the growth of certain vaccinal viruses. Confusion exists as to whether CRFK are epithelial or mesenchymal in phenotype. The aim of this study was to characterise CRFK cells via immunofluorescence, enzyme cytochemistry, western blotting, RT-qPCR for S100A4 and comparison to primary feline proximal tubular epithelial cells (FPTEC) and feline cortical fibroblasts (FCF). CRFK cells were of fusiform morphology and appeared similar to FCF. CRFK expressed the mesenchymal intermediate filament (IF) protein vimentin together with two cell adhesion molecules associated with feline fibroblasts (CD29 and CD44), and lacked expression of the epithelial IF cytokeratin, myogenic IF desmin and endothelial marker von Willebrand factor (vWF). In addition, CRFK did not demonstrate brush border enzyme activity typical of FPTEC. S100A4 gene expression, implicated in both neoplastic transformation and epithelial to mesenchymal transition, was highly upregulated in CRFK in comparison to the primary feline renal cells. CRFK appear phenotypically similar to fibroblasts, rather than tubular epithelial cells, and may have undergone neoplastic transformation or epithelial-to-mesenchymal transition after extensive passaging. This finding may have potential implications for future research utilising this cell line. |
Viral encephalitis is a rare but clinically serious consequence of viral invasion of the brain and insight into its pathogenesis is urgently needed. Important research questions concern the involvement of the host innate immune response in pathogenesis, key to which is the role played by microglia, resident macrophages of the brain parenchyma. Do microglia have a protective function, by coordinating the innate immune response to viral infection, or do they drive pathogenic neuroinflammation? Here we synthesize recent data from mouse models of acute viral encephalitis, which reveal an unambiguously protective role for microglia. Depletion of microglia, via blockade of colony-stimulating factor 1 receptor (CSF1R) signaling, led to increased viral replication accompanied by more severe neurological disease and heightened mortality. Whilst the underlying mechanism(s) remain to be defined, microglial interactions with T cells and phagocytosis of infected neurones appear to play a role. Paradoxically, the production of inflammatory cytokines was increased in several instances following viral infection in microglia-depleted brains, suggesting that: (i) cells other than microglia mediate inflammatory responses and/or (ii) microglia may exert a regulatory function. Under certain circumstances the microglial antiviral response might contribute negatively to longer-term neurological sequelae, although fewer studies have focused on this aspect in encephalitis models. Understanding regulation of the microglial response, and how it contributes to disease is therefore a priority for future studies. Collectively, these findings demonstrate the central role of microglia in pathogenesis, suggesting the exciting possibility that defects of microglial function might contribute to encephalitis susceptibility and/or outcome in humans. |
Artificial blood is an innovative concept of transfusion medicine where specifically designed compounds perform the task of transport and delivery of oxygen in the body to replace this function of allogenic human blood transfusion. Several molecules have been developed in the past few decades to achieve this objective and continous refinements are being continuously made in the quest of the ideal blood substitute. Currently, available technology manufactures artificial blood from haemoglobin obtained from outdated human/bovine blood (Haemoglobin Based Oxygen Carriers) or utilizing Perfluorocarbons. These synthetic blood substitutes are advantageous in that they do not require compatibility testing, are free from blood borne infections, have prolonged shelf life and do not require refrigeration. Artificial blood is projected to have a significant impact on the development of medical care in the future. It can complement the current blood products for transfusion and create a stable supply of safe and effective products. It is likely to reduce the requirements of blood transfusions drastically especially in settings of trauma and surgery thereby reducing the reliance on banked donated blood. |
The synthesis of novel pyrazolylnucleosides 3a–e, 4a–e, 5a–e, and 6a–e are described. The structures of the regioisomers were elucidated by using extensive NMR studies. The pyrazolylnucleosides 5a–e and 6a–e were screened for anticancer activities on sixty human tumor cell lines. The compound 6e showed good activity against 39 cancer cell lines. In particular, it showed significant inhibition against the lung cancer cell line Hop-92 (GI(50) 9.3 µM) and breast cancer cell line HS 578T (GI(50) 3.0 µM). |
IMPORTANCE: Measles cases and outbreaks continue to occur in the United States after the introduction of measles from endemic settings. OBJECTIVE: To discern the factors associated with measles transmission in the United States after measles had been eliminated. DESIGN, SETTING, AND PARTICIPANTS: This cross-sectional study was conducted from January 1, 2001, to December 31, 2017, in the United States among US residents and international visitors with confirmed measles. A maximum likelihood algorithm that uses the observed dates of rash onset and the known distribution of the serial interval (time between symptom onset in related consecutive cases) was applied to outbreak notification data to estimate the effective reproduction number (R), or the mean number of new infections generated per case. Transmissibility was assessed by comparing R based on the characteristics of primary and secondary cases of measles. EXPOSURES: Measles virus. MAIN OUTCOMES AND MEASURES: Effective reproduction number (R), or the mean number of successful transmission events per case of measles (ie, the mean number of persons to whom each patient with measles spreads measles). RESULTS: A total of 2218 individuals with confirmed measles cases (1025 female, 1176 male, and 17 sex not reported; median age, 15 years [range, 0-89 years]) reported from 2001 to 2017 were evaluated. Among patients who received no doses of measles vaccine, R was 0.76 (95% CI, 0.71-0.81); among patients who received 1 dose of measles vaccine, R was 0.17 (95% CI, 0.11-0.26); among patients who received 2 doses or more of measles vaccine, R was 0.27 (95% CI, 0.17-0.39); and among patients with unknown vaccination status, R was 0.52 (95% CI, 0.44-0.60). Among patients born before 1957, R was 0.35 (95% CI, 0.20-0.58), and among those born on or after 1957, R was 0.64 (95% CI, 0.61-0.68). R was higher when primary and secondary cases of measles were patients aged 5 to 17 years (0.36 [95% CI, 0.31-0.42]) compared with assortative transmission in other age groups (<1 year, 0.14 [95% CI, 0.10-0.20]; 1-4 years, 0.25 [95% CI, 0.20-0.30]; 18-29 years, 0.19 [95% CI, 0.15-0.24]; 30-49 years, 0.15 [95% CI, 0.11-0.20]; ≥50 years, 0.04 [95% CI, 0.01-0.10]). CONCLUSIONS AND RELEVANCE: The findings of this study support having high targets for 2-dose measles vaccine coverage, particularly among school-aged children in the United States. |
Human viral hepatitis, a major cause of morbidity and mortality worldwide, is caused by highly diverse viruses with different genetic, ecological, and pathogenetic features. Technological advances that allow throughput sequencing of viral genomes, as well as the development of computational tools to analyze such genome data, have largely expanded our knowledge on the host range and evolutionary history of human hepatitis viruses. Thus, with the exclusion of hepatitis D virus, close or distant relatives of these human pathogens were identified in a number of domestic and wild mammals. Also, sequences of human viral strains isolated from different geographic locations and over different time-spans have allowed the application of phylogeographic and molecular dating approaches to large viral phylogenies. In this review, we summarize the most recent insights into our understanding of the evolutionary events and ecological contexts that determined the origin and spread of human hepatitis viruses. |
BACKGROUND: Aflatoxins are fungal metabolites associated with contaminated food products. Intake of aflatoxin-contaminated food results in serious health hazards and even death. Therefore, the aim of this study is to evaluate the global scientific output of research of aflatoxin by using bibliometric techniques. METHODS: This bibliometric study was conducted using Scopus database and classified the retrieved publications were classified from different aspects, including the countries/region of focus, journals, authors, institutes, citations, and content analysis to discover any hot and emerging topics. In addition, the bibliometric analysis of the international collaborative network and hot research topics were generated by VOSviewer© software version 1.6.10. The publication period was restricted in the search for two decades (1998–2017). RESULTS: The search engine of the Scopus database found 9845 documents published in the field of aflatoxin. The USA is the top publishing source in the world (22.85%), followed by China (11.85%), India (9.32%), and Italy (5.25%). In earlier years, researchers focused on terms related to the topics of “sources and biosynthesis of aflatoxin”, “health effects by aflatoxin”, and “detoxification and treatment of aflatoxin”. However, in recent years, researchers pay more attention to the topic of detection and quantification of aflatoxin. CONCLUSIONS: The quantity of research in global aflatoxin has substantially increased over the past two decades. The evaluation of the historical status and development trend in aflatoxin scientific research can guide future research, and ultimately provide the basis for improving management procedures for governmental decisions, healthcare, industries, and educational institutions. |
BACKGROUND: There is little descriptive data on Stenotrophomonas maltophilia hospital-acquired pneumonia (HAP) in critically ill patients. The optimal modalities of antimicrobial therapy remain to be determined. Our objective was to describe the epidemiology and prognostic factors associated with S. maltophilia pneumonia, focusing on antimicrobial therapy. METHODS: This nationwide retrospective study included all patients admitted to 25 French mixed intensive care units between 2012 and 2017 with hospital-acquired S. maltophilia HAP during intensive care unit stay. Primary endpoint was time to in-hospital death. Secondary endpoints included microbiologic effectiveness and antimicrobial therapeutic modalities such as delay to appropriate antimicrobial treatment, mono versus combination therapy, and duration of antimicrobial therapy. RESULTS: Of the 282 patients included, 84% were intubated at S. maltophilia HAP diagnosis for duration of 11 [5–18] days. The Simplified Acute Physiology Score II was 47 [36–63], and the in-hospital mortality was 49.7%. Underlying chronic pulmonary comorbidities were present in 14.1% of cases. Empirical antimicrobial therapy was considered effective on S. maltophilia according to susceptibility patterns in only 30% of cases. Delay to appropriate antimicrobial treatment had, however, no significant impact on the primary endpoint. Survival analysis did not show any benefit from combination antimicrobial therapy (HR = 1.27, 95%CI [0.88; 1.83], p = 0.20) or prolonged antimicrobial therapy for more than 7 days (HR = 1.06, 95%CI [0.6; 1.86], p = 0.84). No differences were noted in in-hospital death irrespective of an appropriate and timely empiric antimicrobial therapy between mono- versus polymicrobial S. maltophilia HAP (p = 0.273). The duration of ventilation prior to S. maltophilia HAP diagnosis and ICU length of stay were shorter in patients with monomicrobial S. maltophilia HAP (p = 0.031 and p = 0.034 respectively). CONCLUSIONS: S. maltophilia HAP occurred in severe, long-stay intensive care patients who mainly required prolonged invasive ventilation. Empirical antimicrobial therapy was barely effective while antimicrobial treatment modalities had no significant impact on hospital survival. TRIAL REGISTRATION: clinicaltrials.gov, NCT03506191 |
Viral late domains are used by many viruses to recruit the cellular endosomal sorting complex required for transport (ESCRT) to mediate membrane scission during viral budding. Unlike the P(S/T)AP and YPX((1–3))L late domains, which interact directly with the ESCRT proteins Tsg101 and ALIX, the molecular linkage connecting the PPXY late domain to ESCRT proteins is unclear. The mammarenavirus lymphocytic choriomeningitis virus (LCMV) matrix protein, Z, contains only one late domain, PPXY. We previously found that this domain in LCMV Z, as well as the ESCRT pathway, are required for the release of defective interfering (DI) particles but not infectious virus. To better understand the molecular mechanism of ESCRT recruitment by the PPXY late domain, affinity purification-mass spectrometry was used to identify host proteins that interact with the Z proteins of the Old World mammarenaviruses LCMV and Lassa virus. Several Nedd4 family E3 ubiquitin ligases interact with these matrix proteins and in the case of LCMV Z, the interaction was PPXY-dependent. We demonstrated that these ligases directly ubiquitinate LCMV Z and mapped the specific lysine residues modified. A recombinant LCMV containing a Z that cannot be ubiquitinated maintained its ability to produce both infectious virus and DI particles, suggesting that direct ubiquitination of LCMV Z alone is insufficient for recruiting ESCRT proteins to mediate virus release. However, Nedd4 ligases appear to be important for DI particle release suggesting that ubiquitination of targets other than the Z protein itself is required for efficient viral ESCRT recruitment. |
African swine fever virus is transmissible through animal consumption of contaminated feed. To determine virus survival during transoceanic shipping, we calculated the half-life of the virus in 9 feed ingredients exposed to 30-day shipment conditions. Half-lives ranged from 9.6 to 14.2 days, indicating that the feed matrix environment promotes virus stability. |
Control chart methods have been received much attentions in biosurvillance studies. The correlation between charting statistics or regions could be considerably important in monitoring the states of multiple outcomes or regions. In addition, the process variable distribution is unknown in most situations. In this paper, we propose a new nonparametric strategy for multivariate process monitoring when the distribution of a process variable is unknown. We discuss the EWMA control chart based on rank methods for a multivariate process, and the approach is completely nonparametric. A simulation study demonstrates that the proposed method is efficient in detecting shifts for multivariate processes. A real Japanese influenza data example is given to illustrate the performance of the proposed method. |
Newcastle disease virus (NDV) causes severe infectious disease in poultry and selectively kills tumor cells, by inducing apoptosis and cytokines secretion. In this report, we study the mechanisms underlying NDV-induced apoptosis by investigating the unfolded protein response (UPR). We found that NDV infection activated all three branches of the UPR signaling (PERK-eIF2α, ATF6, and IRE1α) and triggered apoptosis, in avian cells (DF-1 and CEF) and in various human cancer cell types (HeLa, Cal27, HN13, A549, H1299, Huh7, and HepG2). Interestingly, the suppression of either apoptosis or UPR led to impaired NDV proliferation. Meanwhile, the inhibition of UPR by 4-PBA protected cells from NDV-induced apoptosis. Further study revealed that activation of PERK-eIF2α induced the expression of transcription factor CHOP, which subsequently promoted apoptosis by downregulating BCL-2/MCL-1, promoting JNK signaling and suppressing AKT signaling. In parallel, IRE1α mediated the splicing of XBP1 mRNA and resulted in the translation and nuclear translocation of XBP1s, thereby promoting the transcription of ER chaperones and components of ER-associated degradation (ERAD). Furthermore, IRE1α promoted apoptosis and cytokines secretion via the activation of JNK signaling. Knock down and overexpression studies showed that CHOP, IRE1α, XBP1, and JNK supported efficient virus proliferation. Our study demonstrates that the induction of eIF2α-CHOP-BCL-2/JNK and IRE1α-XBP1/JNK signaling cascades promote apoptosis and cytokines secretion, and these signaling cascades support NDV proliferation. |
ISG15 is an interferon-stimulated, ubiquitin-like protein, with anti-viral and anti-bacterial activity. Here, we map the endogenous in vivo ISGylome in the liver following Listeria monocytogenes infection by combining murine models of reduced or enhanced ISGylation with quantitative proteomics. Our method identifies 930 ISG15 sites in 434 proteins and also detects changes in the host ubiquitylome. The ISGylated targets are enriched in proteins which alter cellular metabolic processes, including upstream modulators of the catabolic and antibacterial pathway of autophagy. Computational analysis of substrate structures reveals that a number of ISG15 modifications occur at catalytic sites or dimerization interfaces of enzymes. Finally, we demonstrate that animals and cells with enhanced ISGylation have increased basal and infection-induced autophagy through the modification of mTOR, WIPI2, AMBRA1, and RAB7. Taken together, these findings ascribe a role of ISGylation to temporally reprogram organismal metabolism following infection through direct modification of a subset of enzymes in the liver. |
Socio-economic factors are widely believed to have been an important driver of the transmission of Ebola Virus Disease (EVD) during the West African outbreak of 2014–16, however, studies that have investigated the relationship between socio-economic status (SES) and EVD have found inconsistent results. Using nationally representative household survey data on whether respondents knew a close friend or family member with Ebola, we explore the SES determinants of EVD exposure along individual, household, and community lines in Liberia and Sierra Leone. While we find no overall association between household wealth and EVD exposure, we find that pooled data mask important differences observed within countries with higher wealth households more likely to have been exposed to EVD in Sierra Leone and the opposite relationship in Liberia. Finally, we also generally find a positive association between education and EVD exposure both at the individual and the community levels in the full sample. There is an urgent need to better understand these relationships to examine both why the outbreak spread and to help prepare for future outbreaks. |
BACKGROUND: Pneumocystis jirovecii pneumonia (PJP) remains a severe disease associated with high rates of invasive mechanical ventilation (MV) and mortality. The objectives of this study were to assess early risk factors for severe PJP and 90-day mortality, including the broncho-alveolar lavage fluid cytology profiles at diagnosis. METHODS: We prospectively enrolled all patients meeting pre-defined diagnostic criteria for PJP admitted at Nantes university hospital, France, from January 2012 to January 2017. Diagnostic criteria for PJP were typical clinical features with microbiological confirmation of P. jirovecii cysts by direct examination or a positive specific quantitative real-time polymerase chain reaction (PCR) assay. Severe PJP was defined as hypoxemic acute respiratory failure requiring high-flow nasal oxygen with at least 50% FiO(2), non-invasive ventilation, or MV. RESULTS: Of 2446 respiratory samples investigated during the study period, 514 from 430 patients were positive for P. jirovecii. Of these 430 patients, 107 met criteria for PJP and were included in the study, 53 (49.5%) patients had severe PJP, including 30 who required MV. All patients were immunocompromised with haematological malignancy ranking first (n = 37, 35%), followed by solid organ transplantation (n = 27, 25%), HIV-infection (n = 21, 20%), systemic diseases (n = 13, 12%), solid tumors (n = 12, 11%) and primary immunodeficiency (n = 6, 8%). By multivariate analysis, factors independently associated with severity were older age (OR, 3.36; 95% CI 1.4–8.5; p < 0.05), a P. jirovecii microscopy-positive result from bronchoalveolar lavage (BAL) (OR, 1.3; 95% CI 1.54–9.3; p < 0.05); and absence of a BAL fluid alveolitis profile (OR, 3.2; 95% CI 1.27–8.8; p < 0.04). The 90-day mortality rate was 27%, increasing to 50% in the severe PJP group. Factors independently associated with 90-day mortality were worse SOFA score on day 1 (OR, 1.05; 95% CI 1.02–1.09; p < 0.001) whereas alveolitis at BAL was protective (OR, 0.79; 95% CI 0.65–0.96; p < 0.05). In the subgroup of HIV-negative patients, similar findings were obtained, then viral co-infection were independently associated with higher 90-day mortality (OR, 1.25; 95% CI 1.02–1.55; p < 0.05). CONCLUSIONS: Older age and P. jirovecii oocysts at microscopic examination of BAL were independently associated with severe PJP. Both initial PJP severity as evaluated by the SOFA score and viral co-infection predicted 90-day mortality. Alveolitis at BAL examination was associated with less severe PJP. The pathophysiological mechanism underlying this observation deserves further investigation. |
[Image: see text] Recombinase polymerase amplification (RPA) is an isothermal DNA amplification method with broad applications as a point-of-care test and in molecular biology techniques. Currently, most of the applications are focused on target-specific amplification. Because RPA has the advantage of amplifying DNA under isothermal conditions, we utilized RPA as a DNA library amplification tool. In this study, we used a sheared genomic DNA library and an oligonucleotide (oligo) library for the comparison of polymerase chain reaction and RPA. For the sheared DNA library, we observed biased amplification after RPA was conducted. Thus, to amplify the size-variable DNA library uniformly, we introduced a linear amplification strategy with RPA and successfully improved the uniformity. On the other hand, using the same-sized oligo library, we confirmed that RPA amplified this library uniformly without modification of the protocol. These results demonstrate that RPA can be applied not only to amplify a specific target as previously demonstrated but also to amplify a complex DNA library composed of a large number of different DNA molecules. |
OBJECTIVE: Endoplasmic reticulum (ER) stress and mitochondrial function affected intramuscular fat accumulation. However, there is no clear evident on the effect of the regulation of ER stress and mitochondrial function by Angiotensin-converting enzyme 2 (ACE2) on the prevention of intramuscular fat metabolism. We investigated the effects of ACE2 on ER stress and mitochondrial function in skeletal muscle lipid metabolism. METHODS: The triglyceride (TG) content in skeletal muscle of ACE2 knockout mice and Ad-ACE2-treated db/db mice were detected by assay kits. Meanwhile, the expression of lipogenic genes (ACCα, SREBP-1c, LXRα, CPT-1α, PGC-1α and PPARα), ER stress and mitochondrial function related genes (GRP78, eIF2α, ATF4, BCL-2, and SDH6) were analyzed by RT-PCR. Lipid metabolism, ER stress and mitochondrial function related genes were analyzed by RT-PCR in ACE2-overexpression C2C12 cell. Moreover, the IKKβ/NFκB/IRS-1 pathway was determined using lysate sample from skeletal muscle of ACE2 knockout mice. RESULTS: ACE2 deficiency in vivo is associated with increased lipid accumulation in skeletal muscle. The ACE2 knockout mice displayed an elevated level of ER stress and mitochondrial dysfunctions in skeletal muscle. In contrast, activation of ACE2 can ameliorate ER stress and mitochondrial function, which slightly accompanied by reduced TG content and down-regulated the expression of skeletal muscle lipogenic proteins in the db/db mice. Additionally, ACE2 improved skeletal muscle lipid metabolism and ER stress genes in the C2C12 cells. Mechanistically, endogenous ACE2 improved lipid metabolism through the IKKβ/NFκB/IRS-1 pathway in skeletal muscle. CONCLUSIONS: ACE2 was first reported to play a notable role on intramuscular fat regulation by improving endoplasmic reticulum and mitochondrial function. This study may provide a strategy for treating insulin resistance in skeletal muscle. |
Host immune response may be involved in the pathogenesis of children Mycoplasma pneumoniae pneumonia (MPP). In the current study, we investigated the alterations of cytokines levels among control, mild MPP and severe MPP children to determine whether cytokine signatures associate with MPP and correlate with disease severity. We measured 13 cytokines in bronchoalveolar lavage fluid (BALF) of 88 children with MPP and 26 children with foreign body aspiration (FB) using a Luminex system. Linear discriminant analyses were performed to develop predictive models of mild MPP and severe MPP on these children. We observed nearly complete separations of severe MPP group, mild MPP group and control group in linear discriminant analyses. Eleven cytokines significantly increased in children with MPP, and seven cytokines had statistically significant upward linear trends correlated with MPP severity. In addition, compared to control group, both IFNγ/IL4 ratio and IFNγ/IL13 ratio increased in mild MPP and severe MPP groups. Our results suggest that children MPP can alter BALF cytokines signatures which associate with disease severity and can be characterized by a distinct airway molecular phenotype that has elevated Th1/Th2 ratios. |
Lack of disease surveillance in small companion animals worldwide has contributed to a deficit in our ability to detect and respond to outbreaks. In this paper we describe the first real-time syndromic surveillance system that conducts integrated spatio-temporal analysis of data from a national network of veterinary premises for the early detection of disease outbreaks in small animals. We illustrate the system’s performance using data relating to gastrointestinal disease in dogs and cats. The data consist of approximately one million electronic health records for dogs and cats, collected from 458 UK veterinary premises between March 2014 and 2016. For this illustration, the system predicts the relative reporting rate of gastrointestinal disease amongst all presentations, and updates its predictions as new data accrue. The system was able to detect simulated outbreaks of varying spatial geometry, extent and severity. The system is flexible: it generates outcomes that are easily interpretable; the user can set their own outbreak detection thresholds. The system provides the foundation for prompt detection and control of health threats in companion animals. |
The 2019 hepatitis A outbreak has become increasingly prevalent among adults in Korea and is the largest outbreak since that in 2009-2010. The incidence in the current outbreak is highest among adults aged 35-44 years, corresponding to the peak incidence among those aged 25-34 years 10 years ago. This may indicate a cohort effect in the corresponding age group. Causes of these repeated outbreaks of hepatitis A in Korea are low level of immunity among adults, Korean food culture that consumes raw seafood such as salted clam and inadequate public health system. Among countermeasures, along with general infectious disease control measures including control of the infectious agent, infection spread, and host, urgent actions are needed to review the vaccination policy and establish an adequate public health system. |
Porcine epidemic diarrhea (PED) is a highly contagious enteric swine disease. The large economic impact of PED on the swine industry worldwide has made the development of an effective PED vaccine a necessity. S0, a truncated region of the porcine epidemic diarrhea virus (PEDV) spike protein, has been suggested as a candidate antigen for PED subunit vaccines; however, poor solubility problems when the protein is expressed in Escherichia coli, and the inherent problems of subunit vaccines, such as low immunogenicity, remain. Flagellin has been widely used as a fusion partner to enhance the immunogenicity and solubility of many difficult-to-express proteins; however, the conjugation effect of flagellin varies depending on the target antigen or the position of the fusion placement. Here, we conjugated flagellin, Vibrio vulnificus FlaB, to the N- and C-termini of S0 and evaluated the ability of the fusion to enhance the solubility and immunogenicity of S0. Flagellin conjugation in the presence of the trigger factor chaperone tig greatly improved the solubility of the fusion protein (up to 99%) regardless of its conjugation position. Of importance, flagellin conjugated to the N-terminus of S0 significantly enhanced S0-specific humoral immune responses compared to other recombinant antigens in Balb/c mice. The mechanism of this phenomenon was investigated through in vitro and in vivo studies. These findings provide important information for the development of a novel PED vaccine and flagellin-based immunotherapeutics. |
Pet owners frequently administer antimicrobials to their pets and therefore have an important role to play in promoting antimicrobial stewardship in veterinary medicine. However, best methods of educating pet owners about antimicrobial stewardship have yet to be defined. While visual materials such as brochures and posters are often used in health promotion campaigns, their effectiveness in veterinary medicine is unknown. The objective of this study was to determine whether pet owners noticed and retained the message of a poster with an antimicrobial stewardship message placed in veterinary clinic exam rooms. A total of 111 pet owners from five veterinary clinics (three general practices, two low-cost clinics) in the greater Philadelphia area participated in the study. Participants completed a survey asking whether they noticed the poster and if they could paraphrase its message. In a follow-up survey, an antibiotic knowledge score was calculated from answers to questions assessing their knowledge of the poster message. Baseline knowledge was assessed by asking participants to define antibiotic resistance. At the end of the study, veterinarians at participating clinics were interviewed about their experiences with the poster. Only 51 (46.4%) participants noticed the poster, and only 11 (9.9%) could partially or completely reproduce its message. No demographic or clinic-level factors were significantly associated with noticing the poster or recalling its message. Antibiotic knowledge scores were highly correlated (ρ = 0.87, p < 0.001) with baseline knowledge and not affected by viewing the poster (p = 0.955). Veterinarians expressed skepticism that the poster was effective in conveying a message of judicious antibiotic use to clients and noted no difference in the frequency with which they discussed antibiotic resistance or felt pressured to prescribe antibiotics by their clients. Posters alone will likely have limited impact in conveying a message of judicious antibiotic use to pet owners. However, they might be useful as part of an active, multi-modal education strategy, especially if complemented by veterinarian actions. |
BACKGROUND: Since the Sepsis-3 criteria, change in Sequential Organ Failure Assessment (SOFA) score has become a key component of sepsis identification. Thus, it could be argued that reversal of this change (Δ(SOFA)) may reflect sepsis response and could be used as measure of efficacy in interventional trials. We aimed to assess the predictive performance of Δ(SOFA) for 28-day mortality. METHODS: Data from two previously published randomized controlled trials were studied: the first reporting on patients with severe Gram-negative infections as a derivation cohort and the second reporting on patients with ventilator-associated pneumonia as a validation cohort. Only patients with sepsis according to the Sepsis-3 definition were included in this analysis. SOFA scores were calculated on days 1, 2, 3, 5, 7, 14, and 28. RESULTS: We included 448 patients within the derivation cohort and 199 within the validation cohort. Mean SOFA scores on day 1 were 6.06 ± 4.07 and 7.84 ± 3.39, and 28 day mortality 22.8% and 29.6%, respectively. In the derivation cohort, the earliest time point where Δ(SOFA) score predicted mortality was day 7 (AUROC (95% CI) 0.84 (0.80–0.89); p < 0.001). The best tradeoff for prediction was found with 25% changes (78% sensitivity, 80% specificity); less than 25% decrease of admission SOFA was associated with increased mortality (odds ratio for death 14.87). This finding was confirmed in the validation cohort. CONCLUSIONS: Δ(SOFA) on day 7 is a useful early prognostic marker of 28-day mortality and could serve as an endpoint in future sepsis trials alongside mortality. TRIAL REGISTRATION: ClinicalTrials.gov numbers NCT01223690 and NCT00297674 |
Resident memory T cells (T(RM) cells) are an important first-line defense against respiratory pathogens, but the unique contributions of lung T(RM) cell populations to protective immunity and the factors that govern their localization to different compartments of the lung are not well understood. Here, we show that airway and interstitial T(RM) cells have distinct effector functions and that CXCR6 controls the partitioning of T(RM) cells within the lung by recruiting CD8 T(RM) cells to the airways. The absence of CXCR6 significantly decreases airway CD8 T(RM) cells due to altered trafficking of CXCR6(−/−) cells within the lung, and not decreased survival in the airways. CXCL16, the ligand for CXCR6, is localized primarily at the respiratory epithelium, and mice lacking CXCL16 also had decreased CD8 T(RM) cells in the airways. Finally, blocking CXCL16 inhibited the steady-state maintenance of airway T(RM) cells. Thus, the CXCR6/CXCL16 signaling axis controls the localization of T(RM) cells to different compartments of the lung and maintains airway T(RM) cells. |
Severe influenza infections are often associated with the excessive induction of pro-inflammatory cytokines, which is also referred to as “cytokine storms”. Several studies have shown that cytokine storms are directly associated with influenza-induced fatal acute lung injury and acute respiratory distress syndrome. Due to the narrow administration window, current antiviral therapies are often inadequate. The efforts to use immunomodulatory agents alone or in combination with antiviral agents in the treatment of influenza in animal models have resulted in the achievement of protective effects accompanied with reduced cytokine production. Currently, there are no immunomodulatory drugs for influenza available for clinical use. Animal models, despite being ideal to study the anti-inflammatory responses to influenza virus infection, are very costly and time-consuming. Therefore, there is an urgent need to establish fast and economical screening methods using cell-based models to screen and develop novel immunomodulatory agents. In this study, we screened seven human cell lines and found that the human monocytic cell U937 supports the replication of different subtypes of influenza viruses as well as the production of the important pro-inflammatory cytokines and was selected to develop the cell-based model. The U937 cell model was validated by testing a panel of known antiviral and immunomodulatory agents and screening a drug library consisting of 1280 compounds comprised mostly of FDA-approved drugs. We demonstrated that the U937 cell model is robust and suitable for the high-throughput screening of immunomodulators and antivirals against influenza infection. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12250-019-00145-w) contains supplementary material, which is available to authorized users. |
The aim of the present study was to screen the anti-dengue potential of crude leaf extracts of two plants from Pavetta tomentosa and Tarenna asiatica. For larvicidal assay, the acetone extract of both plants showed maximum effects, with the least LC(50) and LC(90) values (P. tomentosa (5.968 and 7.493 μg/ml) and T. asiatica (1.288 and 1.992 μg/ml)) and the same extract of both plants exhibited better pupicidal potency. The adulticidal activity of both plants (0–60 min interval periods) recorded best results in acetone extracts and the LC(50) and LC(90) values were recorded as P. tomentosa (32.105 and 41.001 μg/ml) and T. asiatica (09.012 and 11.854 μg/ml). Among the two plants P. tomentosa acetone leaf extract have good antiviral property against Dengue viral cell line. In addition, the phytochemical nature of the plant reveals the presence of saponins, flavonoids and alkaloids in all the tested extracts of both plants. GC-MS analysis revealed Hexanedioic acid, Bis(2-Ethylhexyl) Ester (22.54) and 2,6,10,14,18,22- Tetracosahexane, 2,6,10, 15, 19,15,19,23- Hexamethyl-(ALL-E)- (25.33) identified as two major phytoconstitutents in P. tomentosa and Tetracontane (23.580) is a major compound identified from T. asiatica acetone extracts. The functional groups of chemical compounds (aromatis, alkanes, alkyls and carboxylic acids) from P. tomentosa and T. asiatica were analyzed by FT-IR spectrum. |
As infectious disease outbreaks emerge, public health agencies often enact vaccination and social distancing measures to slow transmission. Their success depends on not only strategies and resources, but also public adherence. Individual willingness to take precautions may be influenced by global factors, such as news media, or local factors, such as infected family members or friends. Here, we compare three modes of epidemiological decision-making in the midst of a growing outbreak using network-based mathematical models that capture plausible heterogeneity in human contact patterns. Individuals decide whether to adopt a recommended intervention based on overall disease prevalence, the proportion of social contacts infected, or the number of social contacts infected. While all strategies can substantially mitigate transmission, vaccinating (or self isolating) based on the number of infected acquaintances is expected to prevent the most infections while requiring the fewest intervention resources. Unlike the other strategies, it has a substantial herd effect, providing indirect protection to a large fraction of the population. |
AIMS/HYPOTHESIS: Diabetes is associated with an increased risk of dementia. We estimated the potential impact of trends in diabetes prevalence upon mortality and the future burden of dementia and disability in England and Wales. METHODS: We used a probabilistic multi-state, open cohort Markov model to integrate observed trends in diabetes, cardiovascular disease and dementia to forecast the occurrence of disability and dementia up to the year 2060. Model input data were taken from the English Longitudinal Study of Ageing, Office for National Statistics vital data and published effect estimates for health-state transition probabilities. The baseline scenario corresponded to recent trends in obesity: a 26% increase in the number of people with diabetes by 2060. This scenario was evaluated against three alternative projected trends in diabetes: increases of 49%, 20% and 7%. RESULTS: Our results suggest that changes in the trend in diabetes prevalence will lead to changes in mortality and incidence of dementia and disability, which will become visible after 10–15 years. If the relative prevalence of diabetes increases 49% by 2060, expected additional deaths would be approximately 255,000 (95% uncertainty interval [UI] 236,000–272,200), with 85,900 (71,500–101,600) cumulative additional cases of dementia and 104,900 (85,900–125,400) additional cases of disability. With a smaller relative increase in diabetes prevalence (7% increase by 2060), we estimated 222,200 (205,700–237,300) fewer deaths, and 77,000 (64,300–90,800) and 93,300 (76,700–111,400) fewer additional cases of dementia and disability, respectively, than the baseline case of a 26% increase in diabetes. CONCLUSIONS/INTERPRETATION: Reducing the burden of diabetes could result in substantial reductions in the incidence of dementia and disability over the medium to long term. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s00125-019-05015-4) contains peer-reviewed but unedited supplementary supplementary material, which is available to authorised users. |
This work presents a simple one-pot protocol to achieve core-doped shell nanohybrids comprising silver nanoparticles, curcumin and thermoresponsive polymeric shell taking advantage of the reducing properties of phenolic curcumin substance and its ability to decorate metallic surfaces. Silver nanoparticles were synthesized, via sodium citrate and silver nitrate addition into a boiling aqueous solution of curcumin, monomers and surfactant. Curcumin and sodium citrate promoted silver nucleation, acting as reducing and stabilizing agents. These curcumin-capped AgNPs enabled, after adding the radical polymerization initiator, the assembling of the growing polymer chains around the hydrophobic AgNP surface. The resultant core-doped shell nanohybrids exhibit plasmonic, luminescent and volume thermoresponsive properties, with improved possibilities to be used as successful therapeutic platforms. In fact, the possibility to nanoconfine the synergistic antioxidant, antiviral, antibacterial features of silver and curcumin in one bioavailable hybrid paves the way to promising applications in the biomedical field. |
Advocates for a One Health approach recognize that global health challenges require multidisciplinary collaborative efforts. While past publications have looked at interdisciplinary competency training for collaboration, few have identified the factors and conditions that enable operational One Health. Through a scoping review of the literature, a multidisciplinary team of researchers analyzed peer-reviewed publications describing multisectoral collaborations around infectious disease-related health events. The review identified 12 factors that support successful One Health collaborations and a coordinated response to health events across three levels: two individual factors (education & training and prior experience & existing relationships), four organizational factors (organizational structures, culture, human resources and, communication), and six network factors (network structures, relationships, leadership, management, available & accessible resources, political environment). The researchers also identified the stage of collaboration during which these factors were most critical, further organizing into starting condition or process-based factors. The research found that publications on multisectoral collaboration for health events do not uniformly report on successes or challenges of collaboration and rarely identify outputs or outcomes of the collaborative process. This paper proposes a common language and framework to enable more uniform reporting, implementation, and evaluation of future One Health collaborations. |
Influenza A viruses (IAVs) are important animal and human emerging and re-emerging pathogens that are responsible for yearly seasonal epidemics and sporadic pandemics. IAVs cause a wide range of clinical illnesses, from relatively mild infections by seasonal strains, to acute respiratory distress during infections with highly pathogenic avian IAVs (HPAI). For this study, we infected A549 human lung cells with lab prototype A/PR/8/34 (H1N1) (PR8), a seasonal H1N1 (RV733), the 2009 pandemic H1N1 (pdm09), or with two avian strains, an H5N1 HPAI strain or an H7N9 strain that has low pathogenicity in birds but high pathogenicity in humans. We used a newly-developed aptamer-based multiplexed technique (SOMAscan(®)) to examine >1300 human lung cell proteins affected by the different IAV strains, and identified more than 500 significantly dysregulated cellular proteins. Our analyses indicated that the avian strains induced more profound changes in the A549 global proteome compared to all tested low-pathogenicity H1N1 strains. The PR8 strain induced a general activation, primarily by upregulating many immune molecules, the seasonal RV733 and pdm09 strains had minimal effect upon assayed molecules, and the avian strains induced significant downregulation, primarily in antimicrobial response, cardiovascular and post-translational modification systems. |
Background: Rhinovirus C is an important pathogen of asthmatic and non-asthmatic children hospitalised with episodic wheeze. Previous studies on other respiratory viruses have shown that several host cytokines correlate with duration of hospitalisation, but this has yet to be investigated in children with RV-C infection. We determined the nasal cytokine profiles of these children and investigated their relationship with RV-C load and clinical outcome. Flocked nasal swabs were collected from children aged 24–72 months presenting to the Emergency Department at Princess Margaret Hospital with a clinical diagnosis of acute wheeze and an acute upper respiratory tract viral infection. RV-C load was determined by quantitative RT-PCR and cytokine profiles were characterised by a commercial human cytokine 34-plex panel. RV-C was the most commonly detected virus in pre-school-aged children hospitalised with an episodic wheeze. RV-C load did not significantly differ between asthmatic and non-asthmatic patients. Both groups showed a Th2-based cytokine profile. However, Th17 response cytokines IL-17 and IL-1β were only elevated in RV-C-infected children with pre-existing asthma. Neither RV-C load nor any specific cytokines were associated illness severity in this study. Medically attended RV-C-induced wheeze is characterised by a Th2 inflammatory pattern, independent of viral load. Any therapeutic interventions should be aimed at modulating the host response following infection. |
Zika virus (ZIKV) infection during pregnancy leads to severe congenital Zika syndrome, which includes microcephaly and other neurological malformations. No therapeutic agents have, so far, been approved for the treatment of ZIKV infection in humans; as such, there is a need for a continuous effort to develop effective and safe antiviral drugs to treat ZIKV-caused diseases. After screening a natural product library, we have herein identified four natural products with anti-ZIKV activity in Vero E6 cells, including gossypol, curcumin, digitonin, and conessine. Except for curcumin, the other three natural products have not been reported before to have anti-ZIKV activity. Among them, gossypol exhibited the strongest inhibitory activity against almost all 10 ZIKV strains tested, including six recent epidemic human strains. The mechanistic study indicated that gossypol could neutralize ZIKV infection by targeting the envelope protein domain III (EDIII) of ZIKV. In contrast, the other natural products inhibited ZIKV infection by targeting the host cell or cell-associated entry and replication stages of ZIKV. A combination of gossypol with any of the three natural products identified in this study, as well as with bortezomib, a previously reported anti-ZIKV compound, exhibited significant combinatorial inhibitory effects against three ZIKV human strains tested. Importantly, gossypol also demonstrated marked potency against all four serotypes of dengue virus (DENV) human strains in vitro. Taken together, this study indicates the potential for further development of these natural products, particularly gossypol, as the lead compound or broad-spectrum inhibitors against ZIKV and other flaviviruses, such as DENV. |
Human immunodeficiency virus (HIV) is a global health concern affecting millions of individuals with a wide variety of currently circulating subtypes affecting various regions of the globe. HIV relies on multiple regulatory proteins to modify the host cell to promote replication in infected T cells, and these regulatory proteins can have subtle phenotypic differences between subtypes. One of these proteins, HIV-1 Trans-Activator of Transcription (Tat), is capable of RNA interference (RNAi) Silencing Suppressor (RSS) activity and induction of cell death in T cells. However, the subtype-specific RSS activity and induction of cell death have not been explored. We investigated the ability of Tat subtypes and variants to induce RSS activity and cell death. TatB, from HIV-1 subtype B, was found to be a potent RSS activator by 40% whereas TatC, from HIV-1 subtype C, showed 15% RSS activity while subtype TatC variants exhibited varying levels. A high level of cell death (50–53%) was induced by subtype TatB when compared to subtype TatC (25–28%) and varying levels were observed with subtype TatC variants. These differential activities could be due to variations in the functional domains of Tat. These observations further our understanding of subtype-specific augmentation of Tat in HIV-1 replication and pathogenesis. |
Zika virus (ZIKV) is transmitted by Aedes mosquitoes and exhibits genetic variation with African and Asian lineages. ZIKV Natal RGN strain, an Asian-lineage virus, has been identified in brain tissues from fetal autopsy cases with microcephaly and is suggested to be a neurotropic variant. However, ZIKV Natal RGN strain has not been isolated; its biological features are not yet illustrated. This study rescued and characterized recombinant, single-round infectious particles (SRIPs) of the ZIKV Natal RGN strain using reverse genetic and synthetic biology techniques. The DNA-launched replicon of ZIKV Natal RGN was constructed and contains the EGFP reporter, lacks prM-E genes, and replicates under CMV promoter control. The peak in the ZIKV Natal RGN SRIP titer reached 6.25 × 10(6) TCID50/mL in the supernatant of prM-E-expressing packaging cells 72 h post-transfection with a ZIKV Natal RGN replicon. The infectivity of ZIKV Natal RGN SRIPs has been demonstrated to correlate with the green florescence intensity of the EGFP reporter, the SRIP-induced cytopathic effect, and ZIKV’s non-structural protein expression. Moreover, ZIKV Natal RGN SRIPs effectively self-replicated in rhabdomyosarcoma/muscle, glioblastoma/astrocytoma, and retinal pigmented epithelial cells, displaying unique cell susceptibility with differential attachment activity. Therefore, the recombinant ZIKV Natal RGN strain was rescued as SRIPs that could be used to elucidate the biological features of a neurotropic strain regarding cell tropism and pathogenic components, apply for antiviral agent screening, and develop vaccine candidates. |
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