Growth of Lion and Puma Lentiviruses in Domestic Cat Cells and Comparisons with FIV

AbstractFeline immunodeficiency virus (FIV-Fca) is a lentivirus that causes gradual immunological deterioration in domestic cats. Lentiviruses related to FIV have been detected in several nondomestic feline species; the biologic significance of these viruses remains to be defined. To examine thein vitrocell tropism of these nondomestic cat lentiviruses, prototypical puma and lion lentiviruses (FIV-Pco and FIV-Ple) were cultured in a variety of feline cell cultures. A domestic cat T lymphoma cell line, 3201, best supported the replication of both FIV-Pco and FIV-Ple. Moreover, FIV-Ple was lytic for these cells. RT-PCR amplification of a conservedpolgene region demonstrated species-specific primer homology. Sequence and phylogenetic analyses of this amplification product confirmed the identity of the replicating viruses and classified two previously uncharacterized viruses within predictable lion and puma clades. Sequence analysis of a conservedpolregion demonstrated homology with previously characterized FIV-Ple and FIV-Pco. Western blot analysis using domestic cat anti-FIV-Fca sera showed that both FIV-Pco and FIV-Ple were antigenically related, to differing degrees, to three serotypes of FIV-Fca. These studies demonstrate that though nondomestic cat lentiviruses differ significantly from FIV-Fca and that a viral-specific protocol may be necessary for sensitive viral detection, these viruses can replicate in cells of domestic cats, suggesting the potential for cross-species transmission

An extended discrete element method for the estimation of contact pressure at the ankle joint during stance phase

Abnormalities in the ankle contact pressure are related to the onset of osteoarthritis. In vivo measurements are not possible with currently available techniques, so computational methods such as the finite element analysis (FEA) are often used instead. The discrete element method (DEM), a computationally efficient alternative to time-consuming FEA, has also been used to predict the joint contact pressure. It describes the articular cartilage as a bed of independent springs, assuming a linearly elastic behaviour and absence of relative motion between the bones. In this study, we present the extended DEM (EDEM) which is able to track the motion of talus over time. The method was used, with input data from a subject-specific musculoskeletal model, to predict the contact pressure in the ankle joint during gait. Results from EDEM were also compared with outputs from conventional DEM. Predicted values of contact area were larger in EDEM than they were in DEM (4.67 and 4.18 cm2, respectively). Peak values of contact pressure, attained at the toe-off, were 7.3 MPa for EDEM and 6.92 MPa for DEM. Values predicted from EDEM fell well within the ranges reported in the literature. Overall, the motion of the talus had more effect on the extension and shape of the pressure distribution than it had on the magnitude of the pressure. The results indicated that EDEM is a valid methodology for the prediction of ankle contact pressure during daily activities

Metagenomic characterization of swine slurry in a North American swine farm operation

Abstract Modern day large-scale, high-density farming environments are inherently susceptible to viral outbreaks, inadvertently creating conditions that favor increased pathogen transmission and potential zoonotic spread. Metagenomic sequencing has proven to be a useful tool for characterizing the microbial burden in both people, livestock, and environmental samples. International efforts have been successful at characterizing pathogens in commercial farming environments, especially swine farms, however it is unclear whether the full extent of microbial agents have been adequately captured or is representative of farms elsewhere. To augment international efforts we performed metagenomic next-generation sequencing on nine swine slurry and three environmental samples from a United States of America (U.S.A.) farm operation, characterized the microbial composition of slurry, and identified novel viruses. We assembled a remarkable total of 1792 viral genomes, of which 554 were novel/divergent. We assembled 1637 Picobirnavirus genome segments, of which 538 are novel. In addition, we discovered 10 new viruses belonging to a novel taxon: porcine Statoviruses; which have only been previously reported in human, macaques, mouse, and cows. We assembled 3 divergent Posaviruses and 3 swine Picornaviruses. In addition to viruses described, we found other eukaryotic genera such as Entamoeba and Blastocystis, and bacterial genera such as Listeria, Treponema, Peptoclostridium and Bordetella in the slurry. Of these, two species Entamoeba histolytica and Listeria monocytogenes known to cause human disease were detected. Further, antimicrobial resistance genes such as tetracycline and MLS (macrolide, lincosamide, streptogramin) were also identified. Metagenomic surveillance in swine fecal slurry has great potential for novel and antimicrobial resistant pathogen detection

The sputum transcriptome better predicts COPD exacerbations after the withdrawal of inhaled corticosteroids than sputum eosinophils.

Introduction: Continuing inhaled corticosteroid (ICS) use does not benefit all patients with COPD, yet it is difficult to determine which patients may safely sustain ICS withdrawal. Although eosinophil levels can facilitate this decision, better biomarkers could improve personalised treatment decisions. Methods: We performed transcriptional profiling of sputum to explore the molecular biology and compared the predictive value of an unbiased gene signature versus sputum eosinophils for exacerbations after ICS withdrawal in COPD patients. RNA-sequencing data of induced sputum samples from 43 COPD patients were associated with the time to exacerbation after ICS withdrawal. Expression profiles of differentially expressed genes were summarised to create gene signatures. In addition, we built a Bayesian network model to determine coregulatory networks related to the onset of COPD exacerbations after ICS withdrawal. Results: In multivariate analyses, we identified a gene signature (LGALS12, ALOX15, CLC, IL1RL1, CD24, EMR4P) associated with the time to first exacerbation after ICS withdrawal. The addition of this gene signature to a multiple Cox regression model explained more variance of time to exacerbations compared to a model using sputum eosinophils. The gene signature correlated with sputum eosinophil as well as macrophage cell counts. The Bayesian network model identified three coregulatory gene networks as well as sex to be related to an early versus late/nonexacerbation phenotype. Conclusion: We identified a sputum gene expression signature that exhibited a higher predictive value for predicting COPD exacerbations after ICS withdrawal than sputum eosinophilia. Future studies should investigate the utility of this signature, which might enhance personalised ICS treatment in COPD patients

Natural coagulates for wastewater treatment; a review for application and mechanism

The increase of water demand and wastewater generation is among the global concerns in the world. The less effective management of water sources leads to serious consequences, the direct disposal of untreated wastewater is associated with the environmental pollution, elimination of aquatic life and the spread of deadly epidemics. The flocculation process is one of the most important stages in water and wastewater treatment plants, wherein this phase the plankton, colloidal particles, and pollutants are precipitated and removed. Two major types of coagulants are used in the flocculation process included the chemical and natural coagulants. Many studies have been performed to optimize the flocculation process while most of these studies have confirmed the hazardous effects of chemical coagulants utilization on the ecosystem. This chapter reviews a summary of the coagulation/flocculation processes using natural coagulants as well as reviews one of the most effective natural methods of water and wastewater treatment

Clinical and biological heterogeneity of multisystem inflammatory syndrome in adults following SARS-CoV-2 infection: a case series

ImportanceMultisystem inflammatory syndrome in adults (MIS-A) is a poorly understood complication of SARS-CoV-2 infection with significant morbidity and mortality.ObjectiveIdentify clinical, immunological, and histopathologic features of MIS-A to improve understanding of the pathophysiology and approach to treatment.DesignThree cases of MIS-A following SARS-CoV-2 infection were clinically identified between October 2021 – March 2022 using the U.S. Centers for Disease Control and Prevention diagnostic criteria. Clinical, laboratory, imaging, and tissue data were assessed.FindingsAll three patients developed acute onset cardiogenic shock and demonstrated elevated inflammatory biomarkers at the time of hospital admission that resolved over time. One case co-occurred with new onset Type 1 diabetes and sepsis. Retrospective analysis of myocardial tissue from one case identified SARS-CoV-2 RNA. All three patients fully recovered with standard of care interventions plus immunomodulatory therapy that included intravenous immunoglobulin, corticosteroids, and in two cases, anakinra.ConclusionMIS-A is a severe post-acute sequela of COVID-19 characterized by systemic elevation of inflammatory biomarkers. In this series of three cases, we find that although clinical courses and co-existent diseases vary, even severe presentations have potential for full recovery with prompt recognition and treatment. In addition to cardiogenic shock, glucose intolerance, unmasking of autoimmune disease, and sepsis can be features of MIS-A, and SARS-CoV-2 myocarditis can lead to a similar clinical syndrome

Pulmonary Metagenomic Sequencing Suggests Missed Infections in Immunocompromised Children

This article is made available for unrestricted re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for the duration of the COVID-19 pandemic or until permissions are revoked in writing.BACKGROUND: Despite improved diagnostics, pulmonary pathogens in immunocompromised children frequently evade detection, leading to significant mortality. Therefore, we aimed to develop a highly sensitive metagenomic next-generation sequencing (mNGS) assay capable of evaluating the pulmonary microbiome and identifying diverse pathogens in the lungs of immunocompromised children. METHODS: We collected 41 lower respiratory specimens from 34 immunocompromised children undergoing evaluation for pulmonary disease at 3 children's hospitals from 2014-2016. Samples underwent mechanical homogenization, parallel RNA/DNA extraction, and metagenomic sequencing. Sequencing reads were aligned to the National Center for Biotechnology Information nucleotide reference database to determine taxonomic identities. Statistical outliers were determined based on abundance within each sample and relative to other samples in the cohort. RESULTS: We identified a rich cross-domain pulmonary microbiome that contained bacteria, fungi, RNA viruses, and DNA viruses in each patient. Potentially pathogenic bacteria were ubiquitous among samples but could be distinguished as possible causes of disease by parsing for outlier organisms. Samples with bacterial outliers had significantly depressed alpha-diversity (median, 0.61; interquartile range [IQR], 0.33-0.72 vs median, 0.96; IQR, 0.94-0.96; P < .001). Potential pathogens were detected in half of samples previously negative by clinical diagnostics, demonstrating increased sensitivity for missed pulmonary pathogens (P < .001). CONCLUSIONS: An optimized mNGS assay for pulmonary microbes demonstrates significant inoculation of the lower airways of immunocompromised children with diverse bacteria, fungi, and viruses. Potential pathogens can be identified based on absolute and relative abundance. Ongoing investigation is needed to determine the pathogenic significance of outlier microbes in the lungs of immunocompromised children with pulmonary disease

Rhesus TRIM5α disrupts the HIV-1 capsid at the inter-hexamer interfaces

TRIM proteins play important roles in the innate immune defense against retroviral infection, including human immunodeficiency virus type-1 (HIV-1). Rhesus macaque TRIM5α (TRIM5αrh) targets the HIV-1 capsid and blocks infection at an early post-entry stage, prior to reverse transcription. Studies have shown that binding of TRIM5α to the assembled capsid is essential for restriction and requires the coiled-coil and B30.2/SPRY domains, but the molecular mechanism of restriction is not fully understood. In this study, we investigated, by cryoEM combined with mutagenesis and chemical cross-linking, the direct interactions between HIV-1 capsid protein (CA) assemblies and purified TRIM5αrh containing coiled-coil and SPRY domains (CC-SPRYrh). Concentration-dependent binding of CC-SPRYrh to CA assemblies was observed, while under equivalent conditions the human protein did not bind. Importantly, CC-SPRYrh, but not its human counterpart, disrupted CA tubes in a non-random fashion, releasing fragments of protofilaments consisting of CA hexamers without dissociation into monomers. Furthermore, such structural destruction was prevented by inter-hexamer crosslinking using P207C/T216C mutant CA with disulfide bonds at the CTD-CTD trimer interface of capsid assemblies, but not by intra-hexamer crosslinking via A14C/E45C at the NTD-NTD interface. The same disruption effect by TRIM5αrh on the inter-hexamer interfaces also occurred with purified intact HIV-1 cores. These results provide insights concerning how TRIM5α disrupts the virion core and demonstrate that structural damage of the viral capsid by TRIM5α is likely one of the important components of the mechanism of TRIM5α-mediated HIV-1 restriction. © 2011 Zhao et al

Genomic and serologic characterization of enterovirus A71 brainstem encephalitis

OBJECTIVE: In 2016, Catalonia experienced a pediatric brainstem encephalitis outbreak caused by enterovirus A71 (EV-A71). Conventional testing identified EV in the periphery but rarely in CSF. Metagenomic next-generation sequencing (mNGS) and CSF pan-viral serology (VirScan) were deployed to enhance viral detection and characterization. METHODS: RNA was extracted from the CSF (n = 20), plasma (n = 9), stool (n = 15), and nasopharyngeal samples (n = 16) from 10 children with brainstem encephalitis and 10 children with meningitis or encephalitis. Pathogens were identified using mNGS. Available CSF from cases (n = 12) and pediatric other neurologic disease controls (n = 54) were analyzed with VirScan with a subset (n = 9 and n = 50) validated by ELISA. RESULTS: mNGS detected EV in all samples positive by quantitative reverse transcription polymerase chain reaction (qRT-PCR) (n = 25). In qRT-PCR-negative samples (n = 35), mNGS found virus in 23% (n = 8, 3 CSF samples). Overall, mNGS enhanced EV detection from 42% (25/60) to 57% (33/60) (p-value = 0.013). VirScan and ELISA increased detection to 92% (11/12) compared with 46% (4/12) for CSF mNGS and qRT-PCR (p-value = 0.023). Phylogenetic analysis confirmed the EV-A71 strain clustered with a neurovirulent German EV-A71. A single amino acid substitution (S241P) in the EVA71 VP1 protein was exclusive to the CNS in one subject. CONCLUSION: mNGS with VirScan significantly increased the CNS detection of EVs relative to qRT-PCR, and the latter generated an antigenic profile of the acute EV-A71 immune response. Genomic analysis confirmed the close relation of the outbreak EV-A71 and neuroinvasive German EV-A71. A S241P substitution in VP1 was found exclusively in the CSF.Grants supporting this project include the National Multiple Sclerosis Society and the American Academy of Neurology award FAN-1608-25607 (R.D.S.), Clinical Research Training Scholarship P0534134 (P.S.R.), Sandler and William K. Bowes Jr Foundations (M.R.W., J.L.D., L.M.K., H.A.S., K.C.Z.), Rachleff Family Foundation (M.R.W.), and NINDS of the NIH under award K08NS096117 (M.R.W.) and F31NS113432 (K.E.L.). This study was partially supported by a grant from the Spanish National Health Institute [grant number PI15CIII-00020] and the European Regional Development Fund (FEDER funds). UCSF Biomedical Sciences Program (I.A.H., K.E.L.), UCSF Medical Scientist Training Program (K.E.L.), and the Chan Zuckerberg Biohub (J.E.P., W.W., C.K.C., J.L.D., E.D.C.) also supported this project.S