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The Journal of General Virology May 2024Porcine reproductive and respiratory syndrome (PRRSV) is an enveloped single-stranded positive-sense RNA virus and one of the main pathogens that causes the most...
Porcine reproductive and respiratory syndrome (PRRSV) is an enveloped single-stranded positive-sense RNA virus and one of the main pathogens that causes the most significant economical losses in the swine-producing countries. PRRSV is currently divided into two distinct species, PRRSV-1 and PRRSV-2. The PRRSV virion envelope is composed of four glycosylated membrane proteins and three non-glycosylated envelope proteins. Previous work has suggested that PRRSV-linked glycans are critical structural components for virus assembly. In addition, it has been proposed that PRRSV glycans are implicated in the interaction with host cells and critical for virus infection. In contrast, recent findings showed that removal of N-glycans from PRRSV does not influence virus infection of permissive cells. Thus, there are not sufficient evidences to indicate compellingly that N-glycans present in the PRRSV envelope play a direct function in viral infection. To gain insights into the role of N-glycosylation in PRRSV infection, we analysed the specific contribution of the envelope protein-linked N-glycans to infection of permissive cells. For this purpose, we used a novel strategy to modify envelope protein-linked N-glycans that consists of production of monoglycosylated PRRSV and viral glycoproteins with different glycan states. Our results showed that removal or alteration of N-glycans from PRRSV affected virus infection. Specifically, we found that complex N-glycans are required for an efficient infection in cell cultures. Furthermore, we found that presence of high mannose type glycans on PRRSV surface is the minimal requirement for a productive viral infection. Our findings also show that PRRSV-1 and PRRSV-2 have different requirements of N-glycan structure for an optimal infection. In addition, we demonstrated that removal of N-glycans from PRRSV does not affect viral attachment, suggesting that these carbohydrates played a major role in regulating viral entry. In agreement with these findings, by performing immunoprecipitation assays and colocalization experiments, we found that N-glycans present in the viral envelope glycoproteins are not required to bind to the essential viral receptor CD163. Finally, we found that the presence of N-glycans in CD163 is not required for PRRSV infection.
Topics: Porcine respiratory and reproductive syndrome virus; Glycosylation; Animals; Swine; Polysaccharides; Porcine Reproductive and Respiratory Syndrome; Viral Envelope Proteins; Cell Line; Receptors, Cell Surface; Antigens, Differentiation, Myelomonocytic; Antigens, CD; Viral Envelope
PubMed: 38776134
DOI: 10.1099/jgv.0.001994 -
Nature Communications May 2024The genome of Pseudomonas aeruginosa encodes three type VI secretion systems, each comprising a dozen distinct proteins, which deliver toxins upon T6SS sheath...
The genome of Pseudomonas aeruginosa encodes three type VI secretion systems, each comprising a dozen distinct proteins, which deliver toxins upon T6SS sheath contraction. The least conserved T6SS component, TssA, has variations in size which influence domain organisation and structure. Here we show that the TssA Nt1 domain interacts directly with the sheath in a specific manner, while the C-terminus is essential for oligomerisation. We built chimeric TssA proteins by swapping C-termini and showed that these can be functional even when made of domains from different TssA sub-groups. Functional specificity requires the Nt1 domain, while the origin of the C-terminal domain is more permissive for T6SS function. We identify two regions in short TssA proteins, loop and hairpin, that contribute to sheath binding. We propose a docking mechanism of TssA proteins with the sheath, and a model for how sheath assembly is coordinated by TssA proteins from this position.
Topics: Type VI Secretion Systems; Pseudomonas aeruginosa; Bacterial Proteins; Protein Domains; Protein Binding; Recombinant Fusion Proteins
PubMed: 38769318
DOI: 10.1038/s41467-024-48487-8 -
PLoS Computational Biology May 2024Recent pandemics like COVID-19 highlighted the importance of rapidly developing diagnostics to detect evolving pathogens. CRISPR-Cas technology has recently been used to...
Recent pandemics like COVID-19 highlighted the importance of rapidly developing diagnostics to detect evolving pathogens. CRISPR-Cas technology has recently been used to develop diagnostic assays for sequence-specific recognition of DNA or RNA. These assays have similar sensitivity to the gold standard qPCR but can be deployed as easy to use and inexpensive test strips. However, the discovery of diagnostic regions of a genome flanked by conserved regions where primers can be designed requires extensive bioinformatic analyses of genome sequences. We developed the Python package krisp to aid in the discovery of primers and diagnostic sequences that differentiate groups of samples from each other, using either unaligned genome sequences or a variant call format (VCF) file as input. Krisp has been optimized to handle large datasets by using efficient algorithms that run in near linear time, use minimal RAM, and leverage parallel processing when available. The validity of krisp results has been demonstrated in the laboratory with the successful design of a CRISPR diagnostic assay to distinguish the sudden oak death pathogen Phytophthora ramorum from closely related Phytophthora species. Krisp is released open source under a permissive license with all the documentation needed to quickly design CRISPR-Cas diagnostic assays.
Topics: CRISPR-Cas Systems; Software; Humans; Whole Genome Sequencing; SARS-CoV-2; Computational Biology; COVID-19; Algorithms
PubMed: 38768250
DOI: 10.1371/journal.pcbi.1012139 -
Cancer Research Communications Jun 2024Hemangiosarcoma and angiosarcoma are soft-tissue sarcomas of blood vessel-forming cells in dogs and humans, respectively. These vasoformative sarcomas are aggressive and...
UNLABELLED
Hemangiosarcoma and angiosarcoma are soft-tissue sarcomas of blood vessel-forming cells in dogs and humans, respectively. These vasoformative sarcomas are aggressive and highly metastatic, with disorganized, irregular blood-filled vascular spaces. Our objective was to define molecular programs which support the niche that enables progression of canine hemangiosarcoma and human angiosarcoma. Dog-in-mouse hemangiosarcoma xenografts recapitulated the vasoformative and highly angiogenic morphology and molecular characteristics of primary tumors. Blood vessels in the tumors were complex and disorganized, and they were lined by both donor and host cells. In a series of xenografts, we observed that the transplanted hemangiosarcoma cells created exuberant myeloid hyperplasia and gave rise to lymphoproliferative tumors of mouse origin. Our functional analyses indicate that hemangiosarcoma cells generate a microenvironment that supports expansion and differentiation of hematopoietic progenitor populations. Furthermore, gene expression profiling data revealed hemangiosarcoma cells expressed a repertoire of hematopoietic cytokines capable of regulating the surrounding stromal cells. We conclude that canine hemangiosarcomas, and possibly human angiosarcomas, maintain molecular properties that provide hematopoietic support and facilitate stromal reactions, suggesting their potential involvement in promoting the growth of hematopoietic tumors.
SIGNIFICANCE
We demonstrate that hemangiosarcomas regulate molecular programs supporting hematopoietic expansion and differentiation, providing insights into their potential roles in creating a permissive stromal-immune environment for tumor progression.
Topics: Hemangiosarcoma; Dogs; Animals; Humans; Mice; Tumor Microenvironment; Hematopoietic Stem Cells; Hematopoiesis; Cell Differentiation
PubMed: 38757809
DOI: 10.1158/2767-9764.CRC-23-0441 -
BioRxiv : the Preprint Server For... May 2024elements are non-autonomous Short INterspersed Elements (SINEs) derived from the gene that are present at over one million copies in human genomic DNA. mobilizes by a...
elements are non-autonomous Short INterspersed Elements (SINEs) derived from the gene that are present at over one million copies in human genomic DNA. mobilizes by a mechanism known as retrotransposition, which requires the Long INterspersed Element-1 (LINE-1 or L1) -encoded protein (ORF2p). Here, we demonstrate that HeLa strains differ in their capacity to support retrotransposition. Human elements retrotranspose efficiently in HeLa-HA and HeLa-CCL2 ( -permissive) strains, but not in HeLa-JVM or HeLa-H1 ( -nonpermissive) strains. A similar pattern of retrotransposition was observed for other -derived SINEs and -derived SINEs. In contrast, mammalian LINE-1s, a zebrafish LINE, a human - ( ) element, and an -containing messenger RNA can retrotranspose in all four HeLa strains. Using an reverse transcriptase-based assay, we show that RNAs associate with ORF2p and are converted into cDNAs in both -permissive and -nonpermissive HeLa strains, suggesting that - and -derived SINE RNAs use strategies to 'hijack' L1 ORF2p that are distinct from those used by elements and -containing mRNAs. These data further suggest ORF2p associates with the RNA poly(A) tract in both -permissive and -nonpermissive HeLa strains, but that retrotransposition is blocked after this critical step in -nonpermissive HeLa strains.
PubMed: 38746229
DOI: 10.1101/2024.05.03.592410 -
MedRxiv : the Preprint Server For... Apr 2024To develop the Mexico Smoking and Vaping Model (Mexico SAVM) to estimate cigarette and electronic nicotine delivery systems (ENDS) prevalence and the public health...
OBJECTIVE
To develop the Mexico Smoking and Vaping Model (Mexico SAVM) to estimate cigarette and electronic nicotine delivery systems (ENDS) prevalence and the public health impact of legalizing ENDS use.
METHODS
SAVM, a cohort-based discrete-time simulation model, compares two scenarios. The estimates smoking prevalence and associated mortality outcomes under the current policy of an ENDS ban, using Mexico-specific population projections, death rates, life expectancy, and smoking and e-cigarette prevalence. projects smoking and vaping prevalence under a hypothetical scenario where ENDS use is allowed. The impact of legalizing ENDS use is estimated as the difference in smoking- and vaping-attributable deaths (SVADs) and life-years lost (LYLs) between the ENDS-Restricted and Unrestricted scenarios.
RESULTS
Compared to a national ENDS ban, The Mexico SAVM projects that legalizing ENDS use could decrease smoking prevalence by 40.1% in males and 30.9% in females by 2049 compared to continuing the national ENDS ban. This reduction in prevalence would save 2.9 (2.5 males and 0.4 females) million life-years and avert almost 106 (91.0 males and 15.5 females) thousand deaths between 2025 and 2049. Public health gains decline by 43% to 59,748 SVADs averted when the switching rate is reduced by half and by 24.3% (92,806 SVADs averted) with a 25% ENDS risk level from that of cigarettes but increased by 24.3% (121,375 SVADs averted) with the 5% ENDS risk.
CONCLUSIONS
Mexico SAVM suggests that greater access to ENDS and a more permissive ENDS regulation, simultaneous with strong cigarette policies, would reduce smoking prevalence and decrease smoking-related mortality. The unanticipated effects of an ENDS ban merit closer scrutiny, with further consideration of how specific ENDS restrictions may maximize public health benefits.
PubMed: 38746147
DOI: 10.1101/2024.04.28.24306511 -
Research Square May 2024The rs6265 single nucleotide polymorphism (SNP) in the gene for brain-derived neurotrophic factor is a common variant that alters therapeutic outcomes for individuals...
The rs6265 single nucleotide polymorphism (SNP) in the gene for brain-derived neurotrophic factor is a common variant that alters therapeutic outcomes for individuals with Parkinson's disease (PD). We previously investigated the effects of this SNP on the experimental therapeutic approach of neural grafting, demonstrating that young adult parkinsonian rats carrying the variant Met allele exhibited enhanced graft function compared to wild-type rats, and also exclusively developed aberrant graft-induced dyskinesias (GID). Aging is the primary risk factor for PD and reduces graft efficacy. Here we investigated whether aging interacts with this SNP to further alter cell transplantation outcomes. We hypothesized that aging would dampen enhancement of graft function associated with this genetic variant and exacerbate GID in all grafted subjects. Unexpectedly, beneficial graft function was maintained in aged rs6265 subjects. However, aging was permissive to GID induction, regardless of genotype, with the greatest incidence and severity found in rs6265 expressing animals.
PubMed: 38746088
DOI: 10.21203/rs.3.rs-4243843/v1 -
PloS One 2024Non-human primate (NHP)-based model systems are highly relevant for biomedical research. However, only few NHP cell lines are available and the generation of additional...
Non-human primate (NHP)-based model systems are highly relevant for biomedical research. However, only few NHP cell lines are available and the generation of additional cell lines is an urgent need to help in the refinement and replacement of these models. Using lentiviral transduction of c-Fos, we established cell lines from the brain of rhesus macaques (Macaca mulatta). Transcriptome analysis revealed that these cell lines are closely related to astrocytes, which was confirmed by immunoblot and immunofluorescence microscopy detecting expression of the astrocyte marker glial fibrillary acidic protein (GFAP). Quantitative real-time PCR (qRT-PCR) demonstrated that major pathways of the interferon (IFN) system are intact. Using retroviral pseudotypes we found that the cell lines are susceptible to entry driven by the glycoproteins of vesicular stomatitis virus (VSV), lymphocytic choriomeningitis virus (LCMV) and to a lesser extent influenza A virus (IAV). Finally, these cells supported growth of Zika virus (ZIKV) and Papiine alphaherpesvirus 2 (PaHV2). In summary, we developed IFN-responsive cell lines from the rhesus macaque brain that allowed entry driven by several viral glycoproteins and were permissive to infection with ZIKV and a primate simplexvirus. These cell lines will be useful for efforts to analyze neurotropic viral infections in rhesus macaque models.
Topics: Animals; Macaca mulatta; Astrocytes; Cell Line; Brain; Humans
PubMed: 38743751
DOI: 10.1371/journal.pone.0303059 -
Cureus Apr 2024Nutritional support is a critical component of care for critically ill patients, impacting their recovery and overall prognosis. Traditional approaches to feeding in the... (Review)
Review
Nutritional support is a critical component of care for critically ill patients, impacting their recovery and overall prognosis. Traditional approaches to feeding in the intensive care unit (ICU) have focused on meeting estimated energy requirements, often resulting in unintended consequences such as overfeeding and associated complications. Permissive underfeeding, a concept gaining attention recently, offers a more controlled approach by intentionally providing fewer calories than traditionally recommended. This comprehensive review explores the rationale, evidence, and practical considerations surrounding permissive underfeeding in critically ill patients. We discuss the physiological basis of permissive underfeeding, its potential benefits in mitigating the risks of overfeeding, and the challenges associated with implementation in clinical practice. Through an analysis of critical studies and clinical trials, we evaluate the comparative effectiveness of permissive underfeeding versus traditional feeding methods and examine its impact on patient outcomes. Recommendations for patient selection, monitoring, and future research directions are provided to guide clinicians in optimizing nutritional support strategies for critically ill individuals. By considering the role of permissive underfeeding alongside traditional feeding approaches, healthcare professionals can tailor nutritional interventions to individual patient needs, ultimately improving outcomes in the ICU.
PubMed: 38741818
DOI: 10.7759/cureus.58083 -
Journal of Translational Medicine May 2024Facioscapulohumeral muscular dystrophy (FSHD) is a high-prevalence autosomal dominant neuromuscular disease characterized by significant clinical and genetic...
BACKGROUND
Facioscapulohumeral muscular dystrophy (FSHD) is a high-prevalence autosomal dominant neuromuscular disease characterized by significant clinical and genetic heterogeneity. Genetic diagnosis of FSHD remains a challenge because it cannot be detected by standard sequencing methods and requires a complex diagnosis workflow.
METHODS
We developed a comprehensive genetic FSHD detection method based on Oxford Nanopore Technologies (ONT) whole-genome sequencing. Using a case-control design, we applied this procedure to 29 samples and compared the results with those from optical genome mapping (OGM), bisulfite sequencing (BSS), and whole-exome sequencing (WES).
RESULTS
Using our ONT-based method, we identified 59 haplotypes (35 4qA and 24 4qB) among the 29 samples (including a mosaic sample), as well as the number of D4Z4 repeat units (RUs). The pathogenetic D4Z4 RU contraction identified by our ONT-based method showed 100% concordance with OGM results. The methylation levels of the most distal D4Z4 RU and the double homeobox 4 gene (DUX4) detected by ONT sequencing are highly consistent with the BSS results and showed excellent diagnostic efficiency. Additionally, our ONT-based method provided an independent methylation profile analysis of two permissive 4qA alleles, reflecting a more accurate scenario than traditional BSS. The ONT-based method detected 17 variations in three FSHD2-related genes from nine samples, showing 100% concordance with WES.
CONCLUSIONS
Our ONT-based FSHD detection method is a comprehensive method for identifying pathogenetic D4Z4 RU contractions, methylation level alterations, allele-specific methylation of two 4qA haplotypes, and variations in FSHD2-related genes, which will all greatly improve genetic testing for FSHD.
Topics: Muscular Dystrophy, Facioscapulohumeral; Humans; Whole Genome Sequencing; DNA Methylation; Haplotypes; Male; Case-Control Studies; Homeodomain Proteins; Female; Nanopore Sequencing; Adult
PubMed: 38741136
DOI: 10.1186/s12967-024-05259-8