Whole blood transcriptome analysis in dairy calves experimentally challenged with bovine herpesvirus 1 (BoHV-1) and comparison to a bovine respiratory syncytial virus (BRSV) challenge.

Bovine herpesvirus 1 (BoHV-1), is associated with several clinical syndromes in cattle, among which bovine respiratory disease (BRD) is of particular significance. Despite the importance of the disease, there is a lack of information on the molecular response to infection via experimental challenge with BoHV-1. The objective of this study was to investigate the whole-blood transcriptome of dairy calves experimentally challenged with BoHV-1. A secondary objective was to compare the gene expression results between two separate BRD pathogens using data from a similar challenge study with BRSV. Holstein-Friesian calves (mean age (SD) = 149.2 (23.8) days; mean weight (SD) = 174.6 (21.3) kg) were either administered BoHV-1 inoculate (1 × 107/mL × 8.5 mL) (n = 12) or were mock challenged with sterile phosphate buffered saline (n = 6). Clinical signs were recorded daily from day (d) -1 to d 6 (post-challenge), and whole blood was collected in Tempus RNA tubes on d six post-challenge for RNA-sequencing. There were 488 differentially expressed (DE) genes (p < 0.05, False Discovery rate (FDR) < 0.10, fold change ≥2) between the two treatments. Enriched KEGG pathways (p < 0.05, FDR <0.05); included Influenza A, Cytokine-cytokine receptor interaction and NOD-like receptor signalling. Significant gene ontology terms (p < 0.05, FDR <0.05) included defence response to virus and inflammatory response. Genes that are highly DE in key pathways are potential therapeutic targets for the treatment of BoHV-1 infection. A comparison to data from a similar study with BRSV identified both similarities and differences in the immune response to differing BRD pathogens.

Assessment of Rapid MinION Nanopore DNA Virus Meta-Genomics Using Calves Experimentally Infected with Bovine Herpes Virus-1.

Bovine respiratory disease (BRD), which is the leading cause of morbidity and mortality in cattle, is caused by numerous known and unknown viruses and is responsible for the widespread use of broad-spectrum antibiotics despite the use of polymicrobial BRD vaccines. Viral metagenomics sequencing on the portable, inexpensive Oxford Nanopore Technologies MinION sequencer and sequence analysis with its associated user-friendly point-and-click Epi2ME cloud-based pathogen identification software has the potential for point-of-care/same-day/sample-to-result metagenomic sequence diagnostics of known and unknown BRD pathogens to inform a rapid response and vaccine design. We assessed this potential using in vitro viral cell cultures and nasal swabs taken from calves that were experimentally challenged with a single known BRD-associated DNA virus, namely, bovine herpes virus 1. Extensive optimisation of the standard Oxford Nanopore library preparation protocols, particularly a reduction in the PCR bias of library amplification, was required before BoHV-1 could be identified as the main virus in the in vitro cell cultures and nasal swab samples within approximately 7 h from sample to result. In addition, we observed incorrect assignment of the bovine sequence to bacterial and viral taxa due to the presence of poor-quality bacterial and viral genome assemblies in the RefSeq database used by the EpiME Fastq WIMP pathogen identification software.

Efficacy of single pass UVC air treatment for the inactivation of coronavirus, MS2 coliphage and Staphylococcus aureus bioaerosols.

There is strong evidence that SARS-CoV-2 is spread predominantly by airborne transmission, with high viral loads released into the air as respiratory droplets and aerosols from the infected subject. The spread and persistence of SARS-CoV-2 in diverse indoor environments reinforces the urgent need to supplement distancing and PPE based approaches with effective engineering measures for microbial decontamination - thereby addressing the significant risk posed by aerosols. We hypothesized that a portable, single-pass UVC air treatment device (air flow 1254 L/min) could effectively inactivate bioaerosols containing bacterial and viral indicator organisms, and coronavirus without reliance on filtration technology, at reasonable scale. Robust experiments demonstrated UVC dose dependent inactivation of Staphylococcus aureus (UV rate constant (k) = 0.098 m2/J) and bacteriophage MS2, with up to 6-log MS2 reduction achieved in a single pass through the system (k = 0.119 m2/J). The inclusion of a PTFE diffuse reflector increased the effective UVC dose by up to 34% in comparison to a standard Al foil reflector (with identical lamp output), resulting in significant additional pathogen inactivation (1-log S. aureus and MS2, p < 0.001). Complete inactivation of bovine coronavirus bioaerosols was demonstrated through tissue culture infectivity (2.4-log reduction) and RT-qPCR analysis - confirming single pass UVC treatment to effectively deactivate coronavirus to the limit of detection of the culture-based method. Scenario-based modelling was used to investigate the reduction in risk of airborne person-to-person transmission based upon a single infected subject within the small room. Use of the system providing 5 air changes per hour was shown to significantly reduce airborne viral load and maintain low numbers of RNA copies when the infected subject remained in the room, reducing the risk of airborne pathogen transmission to other room users. We conclude that the application of single-pass UVC systems (without reliance on HEPA filtration) could play a critical role in reducing the risk of airborne pathogen transfer, including SARS-CoV2, in locations where adequate fresh air ventilation cannot be implemented.

Small Non-coding RNA Expression Following Respiratory Syncytial Virus or Measles Virus Infection of Neuronal Cells.

Respiratory syncytial virus (RSV) or measles virus (MeV) infection modifies host responses through small non-coding RNA (sncRNA) expression. We show that RSV or MeV infection of neuronal cells induces sncRNAs including various microRNAs and transfer RNA fragments (tRFs). We show that these tRFs originate from select tRNAs (GCC and CAC for glycine, CTT and AAC for Valine, and CCC and TTT for Lysine). Some of the tRNAs are rarely used by RSV or MeV as indicated by relative synonymous codon usage indices suggesting selective cleavage of the tRNAs occurs in infected neuronal cells. The data implies that differentially expressed sncRNAs may regulate host gene expression via multiple mechanisms in neuronal cells.

Diagnosis of sheep fasciolosis caused by Fasciola hepatica using cathepsin L enzyme-linked immunosorbent assays (ELISA).

Fasciolosis, a global parasitic disease of agricultural livestock, is caused by the liver fluke Fasciola hepatica. Management and strategic control of fasciolosis on farms depends on early assessment of the extent of disease so that control measures can be implemented quickly. Traditionally, this has relied on the detection of eggs in the faeces of animals, a laborious method that lacks sensitivity, especially for sub-clinical infections, and identifies chronic infections only. Enzyme linked immunosorbent assays (ELISA) offer a quicker and more sensitive serological means of diagnosis that could detect early acute infection before significant liver damage occurs. The performance of three functionally-active recombinant forms of the major F. hepatica secreted cathepsins L, rFhCL1, rFhCL2, rFhCL3, and a cathepsin B, rFhCB3, were evaluated as antigens in an indirect ELISA to serologically diagnose liver fluke infection in experimentally and naturally infected sheep. rFhCL1 and rFhCL3 were the most effective of the four antigens detecting fasciolosis in sheep as early as three weeks after experimental infection, at least five weeks earlier than both coproantigen and faecal egg tests. In addition, the rFhCL1 and rFhCL3 ELISAs had a very low detection limit for liver fluke in lambs exposed to natural infection on pastures and thus could play a major role in the surveillance of farms and a 'test and treat' approach to disease management. Finally, antibodies to all three cathepsin L proteases remain high throughout chronic infection but decline rapidly after drug treatment with the flukicide, triclabendazole, implying that the test may be adapted to trace the effectiveness of drug treatment.

Elucidation of the Host Bronchial Lymph Node miRNA Transcriptome Response to Bovine Respiratory Syncytial Virus.

Bovine respiratory disease (BRD) causes substantial morbidity and mortality, affecting cattle of all ages. One of the main causes of BRD is an initial inflammatory response to bovine respiratory syncytial virus (BRSV). MicroRNAs are novel and emerging non-coding small RNAs that regulate many biological processes and are implicated in various inflammatory diseases. The objective of the present study was to elucidate the changes in the bovine bronchial lymph node miRNA transcriptome in response to BRSV following an experimental viral challenge. Holstein-Friesian calves were either administered a challenge dose of BRSV (103.5 TCID50/ml × 15 ml) (n = 12) or were mock inoculated with sterile phosphate buffered saline (n = 6). Daily scoring of clinical signs was performed and calves were euthanized at day 7 post-challenge. Bronchial lymph nodes were collected for subsequent RNA extraction and sequencing (75 bp). Read counts for known miRNAs were generated using the miRDeep2 package using the UMD3.1 reference genome and the bovine mature miRNA sequences from the miRBase database (release 22). EdgeR was used for differential expression analysis and Targetscan was used to identify target genes for the differentially expressed (DE) miRNAs. Target genes were examined for enriched pathways and gene ontologies using Ingenuity Pathway Analysis (Qiagen). Multi-dimensional scaling (MDS) based on miRNA gene expression changes, revealed a clearly defined separation between the BRSV challenged and control calves, although the clinical manifestation of disease was only mild. One hundred and nineteen DE miRNAs (P < 0.05, FDR < 0.1, fold change > 1.5) were detected between the BRSV challenged and control calves. The DE miRNAs were predicted to target 465 genes which were previously found to be DE in bronchial lymph node tissue, between these BRSV challenged and control calves. Of the DE predicted target genes, 455 had fold changes that were inverse to the corresponding DE miRNAs. There were eight enriched pathways among the DE predicted target genes with inverse fold changes to their corresponding DE miRNA including: granulocyte and agranulocyte adhesion and diapedesis, interferon signalling and role of pathogen recognition receptors in recognition of bacteria and viruses. Functions predicted to be increased included: T cell response, apoptosis of leukocytes, immune response of cells and stimulation of cells. Pathogen recognition and proliferation of cytotoxic T cells are vital for the recognition of the virus and its subsequent elimination.

Messenger RNA biomarkers of Bovine Respiratory Syncytial Virus infection in the whole blood of dairy calves.

Bovine Respiratory Syncytial Virus (BRSV) is a primary viral cause of Bovine Respiratory Disease (BRD) in young calves, which is responsible for substantial morbidity and mortality. Infection with BRSV induces global gene expression changes in respiratory tissues. If these changes are observed in tissues which are more accessible in live animals, such as whole blood, they may be used as biomarkers for diagnosis of the disease. Therefore, the objective of the current study was to elucidate the whole blood transcriptomic response of dairy calves to an experimental challenge with BRSV. Calves (Holstein-Friesian) were either administered BRSV inoculate (103.5 TCID50/ml × 15 ml) (n = 12) or sterile phosphate buffered saline (n = 6). Clinical signs were scored daily and whole blood was collected in Tempus RNA tubes immediately prior to euthanasia, at day 7 post-challenge. RNA was extracted from blood and sequenced (150 bp paired-end). The sequence reads were aligned to the bovine reference genome (UMD3.1) and EdgeR was subsequently employed for differential gene expression analysis. Multidimensional scaling showed that samples from BRSV challenged and control calves segregated based on whole blood gene expression changes, despite the BRSV challenged calves only displaying mild clinical symptoms of the disease. There were 281 differentially expressed (DE) genes (p < 0.05, FDR < 0.1, fold change > 2) between the BRSV challenged and control calves. The top enriched KEGG pathways and gene ontology terms were associated with viral infection and included "Influenza A", "defense response to virus", "regulation of viral life cycle" and "innate immune response". Highly DE genes involved in these pathways may be beneficial for the diagnosis of subclinical BRD from blood samples.

ATAC-Seq identifies regions of open chromatin in the bronchial lymph nodes of dairy calves experimentally challenged with bovine respiratory syncytial virus.

BACKGROUND: Bovine Respiratory Syncytial Virus (BRSV) is a cause of Bovine Respiratory Disease (BRD). DNA-based biomarkers contributing to BRD resistance are potentially present in non-protein-coding regulatory regions of the genome, which can be determined using ATAC-Seq. The objectives of this study were to: (i) identify regions of open chromatin in DNA extracted from bronchial lymph nodes (BLN) of healthy dairy calves experimentally challenged with BRSV and compare them with those from non-challenged healthy control calves, (ii) elucidate the chromatin regions that were differentially or uniquely open in the BRSV challenged relative to control calves, and (iii) compare the genes found in regions proximal to the differentially open regions to the genes previously found to be differentially expressed in the BLN in response to BRSV and to previously identified BRD susceptibility loci. This was achieved by challenging clinically healthy Holstein-Friesian calves (mean age 143 ± 14 days) with either BRSV inoculum (n = 12) or with sterile phosphate buffered saline (PBS) (n = 6) and preparing and sequencing ATAC-Seq libraries from fresh BLN tissues. RESULTS: Using Diffbind, 9,144 and 5,096 differentially accessible regions (P < 0.05, FDR < 0.05) were identified between BRSV challenged and control calves employing DeSeq2 and EdgeR, respectively. Additionally, 8,791 chromatin regions were found to be uniquely open in BRSV challenged calves. Seventy-six and 150 of the genes that were previously found to be differentially expressed using RNA-Seq, were located within 2 kb downstream of the differentially accessible regions, and of the regions uniquely open in BRSV challenged calves, respectively. Pathway analyses within ClusterProfiler indicated that these genes were involved in immune responses to infection and participated in the Th1 and Th2 pathways, pathogen recognition and the anti-viral response. There were 237 differentially accessible regions positioned within 40 previously identified BRD susceptibility loci. CONCLUSIONS: The identified open chromatin regions are likely to be involved in the regulatory response of gene transcription induced by infection with BRSV. Consequently, they may contain variants which impact resistance to BRD that could be used in breeding programmes to select healthier, more robust cattle.

Experimental challenge with bovine respiratory syncytial virus in dairy calves: bronchial lymph node transcriptome response.

Bovine Respiratory Disease (BRD) is the leading cause of mortality in calves. The objective of this study was to examine the response of the host's bronchial lymph node transcriptome to Bovine Respiratory Syncytial Virus (BRSV) in a controlled viral challenge. Holstein-Friesian calves were either inoculated with virus (103.5 TCID50/ml × 15 ml) (n = 12) or mock challenged with phosphate buffered saline (n = 6). Clinical signs were scored daily and blood was collected for haematology counts, until euthanasia at day 7 post-challenge. RNA was extracted and sequenced (75 bp paired-end) from bronchial lymph nodes. Sequence reads were aligned to the UMD3.1 bovine reference genome and differential gene expression analysis was performed using EdgeR. There was a clear separation between BRSV challenged and control calves based on gene expression changes, despite an observed mild clinical manifestation of the disease. Therefore, measuring host gene expression levels may be beneficial for the diagnosis of subclinical BRD. There were 934 differentially expressed genes (DEG) (p < 0.05, FDR <0.1, fold change >2) between the BRSV challenged and control calves. Over-represented gene ontology terms, pathways and molecular functions, among the DEG, were associated with immune responses. The top enriched pathways included interferon signaling, granzyme B signaling and pathogen pattern recognition receptors, which are responsible for the cytotoxic responses necessary to eliminate the virus.

Canine and Phocine Distemper Viruses: Global Spread and Genetic Basis of Jumping Species Barriers.

Canine distemper virus (CDV) and phocine distemper (PDV) are closely-related members of the Paramyxoviridae family, genus morbillivirus, in the order Mononegavirales. CDV has a broad host range among carnivores. PDV is thought to be derived from CDV through contact between terrestrial carnivores and seals. PDV has caused extensive mortality in Atlantic seals and other marine mammals, and more recently has spread to the North Pacific Ocean. CDV also infects marine carnivores, and there is evidence of morbillivirus infection of seals and other species in Antarctica. Recently, CDV has spread to felines and other wildlife species in the Serengeti and South Africa. Some CDV vaccines may also have caused wildlife disease. Changes in the virus haemagglutinin (H) protein, particularly the signaling lymphocyte activation molecule (SLAM) receptor binding site, correlate with adaptation to non-canine hosts. Differences in the phosphoprotein (P) gene sequences between disease and non-disease causing CDV strains may relate to pathogenicity in domestic dogs and wildlife. Of most concern are reports of CDV infection and disease in non-human primates raising the possibility of zoonosis. In this article we review the global occurrence of CDV and PDV, and present both historical and genetic information relating to these viruses crossing species barriers.

TRPA1 activation in a human sensory neuronal model: relevance to cough hypersensitivity?

The cough reflex becomes hyperresponsive in acute and chronic respiratory diseases, but understanding the underlying mechanism is hampered by difficulty accessing human tissue containing both nerve endings and neuronal cell bodies. We refined an adult stem cell sensory neuronal model to overcome the limited availability of human neurones and applied the model to study transient receptor potential ankyrin 1 (TRPA1) channel expression and activation.Human dental pulp stem cells (hDPSCs) were differentiated towards a neuronal phenotype, termed peripheral neuronal equivalents (PNEs). Using molecular and immunohistochemical techniques, together with Ca2+ microfluorimetry and whole cell patch clamping, we investigated roles for nerve growth factor (NGF) and the viral mimic poly I:C in TRPA1 activation.PNEs exhibited morphological, molecular and functional characteristics of sensory neurons and expressed functional TRPA1 channels. PNE treatment with NGF for 20 min generated significantly larger inward and outward currents compared to untreated PNEs in response to the TRPA1 agonist cinnamaldehyde (p<0.05). PNE treatment with poly I:C caused similar transient heightened responses to TRPA1 activation compared to untreated cells.Using the PNE neuronal model we observed both NGF and poly I:C mediated sensory neuronal hyperresponsiveness, representing potential neuro-inflammatory mechanisms associated with heightened nociceptive responses recognised in cough hypersensitivity syndrome.

Respiratory virus infection up-regulates TRPV1, TRPA1 and ASICS3 receptors on airway cells.

Receptors implicated in cough hypersensitivity are transient receptor potential vanilloid 1 (TRPV1), transient receptor potential cation channel, Subfamily A, Member 1 (TRPA1) and acid sensing ion channel receptor 3 (ASIC3). Respiratory viruses, such as respiratory syncytial virus (RSV) and measles virus (MV) may interact directly and/or indirectly with these receptors on sensory nerves and epithelial cells in the airways. We used in vitro models of sensory neurones (SHSY5Y or differentiated IMR-32 cells) and human bronchial epithelium (BEAS-2B cells) as well as primary human bronchial epithelial cells (PBEC) to study the effect of MV and RSV infection on receptor expression. Receptor mRNA and protein levels were examined by qPCR and flow cytometry, respectively, following infection or treatment with UV inactivated virus, virus-induced soluble factors or pelleted virus. Concentrations of a range of cytokines in resultant BEAS-2B and PBEC supernatants were determined by ELISA. Up-regulation of TRPV1, TRPA1 and ASICS3 expression occurred by 12 hours post-infection in each cell type. This was independent of replicating virus, within the same cell, as virus-induced soluble factors alone were sufficient to increase channel expression. IL-8 and IL-6 increased in infected cell supernatants. Antibodies against these factors inhibited TRP receptor up-regulation. Capsazepine treatment inhibited virus induced up-regulation of TRPV1 indicating that these receptors are targets for treating virus-induced cough.

Phocine distemper virus in seals, east coast, United States, 2006.

In 2006 and 2007, elevated numbers of deaths among seals, constituting an unusual mortality event, occurred off the coasts of Maine and Massachusetts, United States. We isolated a virus from seal tissue and confirmed it as phocine distemper virus (PDV). We compared the viral hemagglutinin, phosphoprotein, and fusion (F) and matrix (M) protein gene sequences with those of viruses from the 1988 and 2002 PDV epizootics. The virus showed highest similarity with a PDV 1988 Netherlands virus, which raises the possibility that the 2006 isolate from the United States might have emerged independently from 2002 PDVs and that multiple lineages of PDV might be circulating among enzootically infected North American seals. Evidence from comparison of sequences derived from different tissues suggested that mutations in the F and M genes occur in brain tissue that are not present in lung, liver, or blood, which suggests virus persistence in the central nervous system.

Transcriptional upregulation of SOCS 1 and suppressors of cytokine signaling 3 mRNA in the absence of suppressors of cytokine signaling 2 mRNA after infection with West Nile virus or tick-borne encephalitis virus.

Suppressors of cytokine signaling (SOCS) proteins are a family of proteins that are able to act in a classic negative feedback loop to regulate cytokine signal transduction. The regulation of the immune response by SOCS proteins may contribute to persistent infection or even a fatal outcome. In this study, we have investigated the induction of SOCS 1-3 after peripheral infection with West Nile virus (WNV) or tick-borne encephalitis virus (TBEV) in the murine model. We have shown that the cytokine response after infection of mice with WNV or TBEV induces an upregulation in the brain of mRNA transcripts for SOCS 1 and SOCS 3, but not SOCS 2. We hypothesize that SOCS proteins may play a role in limiting cytokine responses in the brain as a neuroprotective mechanism, which may actually enhance the ability of neuroinvasive viruses such as WNV and TBEV to spread and cause disease.

Adrenomedullin gene delivery is cardio-protective in a model of chronic nitric oxide deficiency combining pressure overload, oxidative stress and cardiomyocyte hypertrophy.

BACKGROUND/AIMS: Chronic inhibition of nitric oxide (NO) synthesis is associated with hypertension, myocardial ischemia, oxidative stress and hypertrophy; expression of the vasodilator peptide, adrenomedullin (AM) and its receptors is augmented in cardiomyocytes, indicating that the myocardial AM system may be activated in response to pressure loading and ischemic insult to serve a counter-regulatory, cardio-protective role. The study examined the hypothesis that oxidative stress and hypertrophic remodeling in NO-deficient cardiomyocytes are attenuated by adenoviral vector-mediated delivery of the human adrenomedullin (hAM) gene in vivo. METHODS: The NO synthesis inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME, 15mg . kg(-1) . day(-1)) was given to rats for 4 weeks following systemic administration via the tail vein of a single injection of either adenovirus harbouring hAM cDNA under the control of the cytomegalovirus promoter-enhancer (Ad.CMV-hAM-4F2), or for comparison, adenovirus alone (Ad.Null) or saline. Cardiomyocytes were subsequently isolated for assessment of the influence of each intervention on parameters of oxidative stress and hypertrophic remodelling. RESULTS: Cardiomyocyte expression of the transgene persisted for > or =4 weeks following systemic administration of adenoviral vector. In L-NAME treated rats, relative to Ad.Null or saline administration, Ad.CMV-hAM-4F2 (i) reduced augmented cardiomyocyte membrane protein oxidation and mRNA expression of pro-oxidant (p22phox) and anti-oxidant (SOD-3, GPx) genes; (ii) attenuated increased cardiomyocyte width and mRNA expression of hypertrophic (sk-alpha-actin) and cardio-endocrine (ANP) genes; (iii) did not attenuate hypertension. CONCLUSIONS: Adenoviral vector mediated delivery of hAM resulted in attenuation of myocardial oxidative stress and hypertrophic remodelling in the absence of blood pressure reduction in this model of chronic NO-deficiency. These findings are consistent with a direct cardio-protective action in the myocardium of locally-derived hAM which is not dependant on NO generation.

Sildenafil citrate (Viagra) impairs fertilization and early embryo development in mice.

OBJECTIVE: To determine the effects of sildenafil citrate, a cyclic monophosphate-specific type 5 phosphodiesterase inhibitor known to affect sperm function, on fertilization and early embryo cleavage. DESIGN: This acute mammal study included male and female mice assigned randomly, the females sacrificed after mating and their oocytes/embryos evaluated at four time periods after treatment. SETTING: Academic research environment. ANIMAL(S): Male and female CBAB(6) mice. INTERVENTION(S): Female mice were injected intraperitoneally with 5 IU gonadotropin (hCG) to stimulate follicular growth and induce ovulation. They were each caged with a male that had been gavaged with sildenafil citrate (0.06 mg/0.05 mL) and allowed to mate. After 12, 36, 60, and 84 h, females were killed, their oviducts were dissected out, and retrieved embryos were assessed for blastomere number and quality. MAIN OUTCOME MEASURE(S): Fertilization rates and numbers of embryos were evaluated after treatment. RESULT(S): Fertilization rates (day 1) were markedly reduced (-33%) in matings where the male had taken sildenafil citrate. Over days 2-4, the numbers of embryos developing in the treated group were significantly fewer than in the control group. There was also a trend for impaired cleavage rates within those embryos, although this did not reach significance. CONCLUSION(S): The impairments to fertility caused by sildenafil citrate have important implications for infertility centers and for couples who are using this drug precoitally while attempting to conceive.

Herpes simplex virus type 1 and Alzheimer's disease: the autophagy connection.

The causes of Alzheimer's disease (AD) and of the characteristic pathological features - amyloid plaques and neurofibrillary tangles - of AD brain are unknown, despite the enormous resources provided over the years for their investigation. Indeed, the only generally accepted risk factors are age, Down syndrome, carriage of the type 4 allele of the apolipoprotein E gene (APOE-epsilon 4), and possibly brain injury. Following the authors' previous studies implicating herpes simplex virus type 1 (HSV1) in brain of APOE-epsilon 4 carriers as a major cause of AD, the authors propose here, on the basis of their and others' recent studies, that not only does HSV1 generate the main components of amyloid plaques and neurofibrillary tangles (NFTs) - beta-amyloid (A beta) and abnormally phosphorylated tau but also, by disrupting autophagy, it prevents degradation of these aberrant proteins, leading to their accumulation and deposition, and eventually to AD.

Measles virus superinfection immunity and receptor redistribution in persistently infected NT2 cells.

A recombinant measles virus (MV) expressing red fluorescent protein (MVDsRed1) was used to produce a persistently infected cell line (piNT2-MVDsRed1) from human neural precursor (NT2) cells. A similar cell line (piNT2-MVeGFP) was generated using a virus that expresses enhanced green fluorescent protein. Intracytoplasmic inclusions containing the viral nucleocapsid protein were evident in all cells and viral glycoproteins were present at the cell surface. Nevertheless, the cells did not release infectious virus nor did they fuse to generate syncytia. Uninfected NT2 cells express the MV receptor CD46 uniformly over their surface, whereas CD46 was present in cell surface aggregates in the piNT2 cells. There was no decrease in the overall amount of CD46 in piNT2 compared to NT2 cells. Cell-to-cell fusion was observed when piNT2 cells were overlaid onto confluent monolayers of MV receptor-positive cells, indicating that the viral glycoproteins were correctly folded and processed. Infectious virus was released from the underlying cells, indicating that persistence was not due to gross mutations in the virus genome. Persistently infected cells were superinfected with MV or canine distemper virus and cytopathic effects were not observed. However, mumps virus could readily infect the cells, indicating that superinfection immunity is not caused by general soluble antiviral factors. As MVeGFP and MVDsRed1 are antigenically indistinguishable but phenotypically distinct it was possible to use them to measure the degree of superinfection immunity in the absence of any cytopathic effect. Only small numbers of non-fusing green fluorescent piNT2-MVDsRed1 cells (1 : 300 000) were identified in which superinfecting MVeGFP entered, replicated and expressed its genes.

Is an improved measles-mumps-rubella vaccine necessary or feasible?

The measles-mumps-rubella (MMR) vaccine has been very effective in the elimination of disease and has high biosafety. However, it has been associated with several adverse effects and has recently caused controversy with regard to its possible association with inflammatory bowel disease and autism. This has been postulated to be a property of the measles component of the vaccine, and a "new variant" autism has recently been described and suggested to be associated with vaccine virus. Although one study has reported the presence of measles RNA in inflammatory bowel disease associated with autism, this has not been independently confirmed. This and most of the other demonstrated or perceived adverse effects of the MMR vaccine could theoretically be ascribed to its composition as a mixture of three live replicating viruses, one of which (measles) can induce immunosuppression, although this hypothesis is speculative. It may nonetheless be desirable to improve the biosafety of the MMR vaccine by the development of a nonreplicating vaccine that will stimulate efficient immunity and protection. DNA vaccines for measles, mumps, and rubella viruses have been constructed and tested in animal models but are poorly immunogenic. Several other prototype candidate vaccines are possible, including those based on the rubella virus component of the vaccine as a vector.