RESUMEN
Recently, numerous large-scale mumps outbreaks have occurred in vaccinated populations. Clinical isolates sequenced from these outbreaks have invariably been of genotypes distinct from those of vaccine viruses, raising concern that certain mumps virus strains may escape vaccine-induced immunity. To investigate this concern, sera obtained from children 6 weeks after receipt of measles, mumps, and rubella (MMR) vaccine were tested for the ability to neutralize a carefully selected group of genetically diverse mumps virus strains. Although the geometric mean neutralizing antibody titer of the sera was lower against some virus strains than others, all viruses were readily neutralized, arguing against immune escape.
Asunto(s)
Brotes de Enfermedades , Vacuna contra el Sarampión-Parotiditis-Rubéola/inmunología , Virus de la Parotiditis/inmunología , Paperas/epidemiología , Paperas/inmunología , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Niño , Preescolar , Femenino , Humanos , Masculino , Vacuna contra el Sarampión-Parotiditis-Rubéola/administración & dosificación , Datos de Secuencia Molecular , Paperas/virología , Virus de la Parotiditis/clasificación , Virus de la Parotiditis/genética , Filogenia , Estados Unidos/epidemiología , VacunaciónRESUMEN
In cell culture experiments, phosphorylation appears to be a critical regulator of the herpes simplex virus 1 (HSV-1) immediate-early (IE) protein, ICP0, which is an E3 ubiquitin ligase that transactivates viral gene expression. Three major regions of phosphorylation in ICP0 (amino acids 224 to 232, 365 to 371, and 508 to 518) have been identified, and mutant viruses that block phosphorylation sites within each region (termed Phos 1, 2, and 3, respectively) have been constructed. Previous studies indicated that replication of Phos 1 is significantly reduced compared to that of wild-type virus in cell culture (C. Boutell, et al., J. Virol. 82:10647-10656, 2008). To determine the effects these phosphorylation site mutations have on the viral life cycle in vivo, mice were ocularly infected with wild-type HSV-1, the Phos mutants, or their marker rescue counterparts. Subsequently, viral replication, establishment of latency, and viral explant-induced reactivation of these viruses were examined. Relative to wild-type virus, Phos 1 eye titers were reduced as much as 7- and 18-fold on days 1 and 5 postinfection, respectively. Phos 2 eye titers showed a decrease of 6-fold on day 1 postinfection. Titers of Phos 1 and 2 trigeminal ganglia were reduced as much as 16- and 20-fold, respectively, on day 5 postinfection. Additionally, the reactivation efficiencies of Phos 1 and 2 were impaired relative to wild-type HSV-1, although both viruses established wild-type levels of latency in vivo. The acute replication, latency, and reactivation phenotypes of Phos 3 were similar to those of wild-type HSV-1. We conclude from these studies that phosphorylation is likely a key modulator of ICP0's biological activities in a mouse ocular model of HSV-1 infection.
Asunto(s)
Oftalmopatías/virología , Herpes Simple/virología , Herpesvirus Humano 1/patogenicidad , Proteínas Inmediatas-Precoces/genética , Mutación/genética , Ganglio del Trigémino/virología , Ubiquitina-Proteína Ligasas/genética , Activación Viral , Replicación Viral , Secuencia de Aminoácidos , Animales , Chlorocebus aethiops , Oftalmopatías/metabolismo , Femenino , Genoma Viral , Herpes Simple/metabolismo , Proteínas Inmediatas-Precoces/metabolismo , Ratones , Datos de Secuencia Molecular , Fosforilación , Homología de Secuencia de Aminoácido , Transcripción Genética , Ganglio del Trigémino/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Células Vero , Latencia del VirusRESUMEN
The public health crisis created by the COVID-19 pandemic has spurred a deluge of scientific research aimed at informing the public health and medical response to the pandemic. However, early in the pandemic, those working in frontline public health and clinical care had insufficient time to parse the rapidly evolving evidence and use it for decision-making. Academics in public health and medicine were well-placed to translate the evidence for use by frontline clinicians and public health practitioners. The Novel Coronavirus Research Compendium (NCRC), a group of >60 faculty and trainees across the United States, formed in March 2020 with the goal to quickly triage and review the large volume of preprints and peer-reviewed publications on SARS-CoV-2 and COVID-19 and summarize the most important, novel evidence to inform pandemic response. From April 6 through December 31, 2020, NCRC teams screened 54 192 peer-reviewed articles and preprints, of which 527 were selected for review and uploaded to the NCRC website for public consumption. Most articles were peer-reviewed publications (n = 395, 75.0%), published in 102 journals; 25.1% (n = 132) of articles reviewed were preprints. The NCRC is a successful model of how academics translate scientific knowledge for practitioners and help build capacity for this work among students. This approach could be used for health problems beyond COVID-19, but the effort is resource intensive and may not be sustainable in the long term.
Asunto(s)
COVID-19 , Curaduría de Datos/métodos , Difusión de la Información/métodos , Investigación Interdisciplinaria/organización & administración , Revisión de la Investigación por Pares , Preimpresos como Asunto , SARS-CoV-2 , Humanos , Salud Pública , Estados UnidosRESUMEN
The public health crisis created by the SARS-CoV-2 pandemic has spurred a deluge of scientific research aimed at informing public health and medical response to the COVID-19 pandemic. However, those working in frontline public health and clinical care had insufficient time to parse the rapidly evolving evidence and use it for decision making. Academics in public health and medicine were well-placed to translate the evidence for use by frontline clinicians and public health practitioners. The Novel Coronavirus Research Compendium (NCRC), a group of >50 faculty and trainees, began in March 2020 with the goal to quickly triage and review the large volume of preprints and peer-reviewed publications on SARS-CoV-2 and COVID-19, and to summarize the most important, novel evidence to inform pandemic response. From April 6, 2020 through January 1, 2021, 54,192 papers and preprints were screened by NCRC teams and 527 were selected for review and uploaded to the NCRC website for public consumption. The majority of papers reviewed were peer-reviewed publications (n=395, 75%), published in 102 journals; 25% (n=132) of papers reviewed were of preprints. The NCRC is a successful model of how academics can support practitioners by translating scientific knowledge into action and help to build capacity among students for this work. This approach could be used for health problems beyond COVID-19, but the effort is resource intensive and may not be sustainable over the long term.
RESUMEN
Although several effective mumps virus vaccines have been developed, almost nothing is known about the genetic changes responsible for loss of virulence. One vaccine, Urabe AM9, was withdrawn from the market because of insufficient attenuation. The vaccine was found to contain a mixture of viruses that could be distinguished based on the sequence of the hemagglutinin-neuraminidase gene (HN). Viruses containing lysine at HN amino acid position 335 were isolated from cases of post-vaccination parotitis or meningitis whereas viruses containing glutamic acid at this position were not associated with post-vaccination disease. Using a rat based model of mumps neurovirulence, we demonstrate that this latter virus is significantly attenuated compared to a virus isolated from a patient with post-vaccination meningitis. Complete sequence analysis of the genomes of the two viruses identified sixteen genetic differences, some or all of which must be responsible for differences in virulence. These same genetic differences also account for changes in tropism in cell culture.
Asunto(s)
Proteína HN/genética , Virus de la Parotiditis/genética , Virus de la Parotiditis/patogenicidad , Mutación Missense , Sustitución de Aminoácidos/genética , Animales , Humanos , Meningitis Viral/virología , Vacuna contra la Parotiditis/efectos adversos , Virus de la Parotiditis/aislamiento & purificación , Parotiditis/virología , Ratas , Análisis de Secuencia de ADN , Vacunas Atenuadas/efectos adversos , VirulenciaRESUMEN
The recent global resurgence of mumps has drawn attention to the continued need for robust mumps immunization programs. Unfortunately, some vaccines derived from inadequately attenuated vaccine strains of mumps virus have caused meningitis in vaccinees, leading to withdrawal of certain vaccine strains from the market, public resistance to vaccination, or in some cases, cessation of national mumps vaccination programs. The most widely implicated mumps vaccine in cases of postvaccination meningitis is derived from the Urabe AM9 strain, which remains in use in some countries. The Urabe AM9 vaccine virus has been shown to exhibit a considerable degree of nucleotide and amino acid heterogeneity. Some studies have specifically implicated variants containing a lysine residue at amino acid position 335 in the hemagglutinin-neuraminidase (HN) protein with neurotoxicity, whereas a glutamic acid residue at this position was associated with attenuation. To test this hypothesis we generated two modified Urabe AM9 cDNA clones coding either for a lysine or a glutamic acid at position 335 in the HN gene. The two viruses were rescued by reverse genetics and characterized in vitro and in vivo. Both viruses exhibited similar growth kinetics in neuronal and non-neuronal cell lines and were of similar neurotoxicity when tested in rats, suggesting that amino acid 335 is not a crucial determinant of Urabe AM9 growth or neurovirulence.
Asunto(s)
Sustitución de Aminoácidos , Proteína HN/genética , Vacuna contra la Parotiditis/genética , Virus de la Parotiditis/genética , Animales , Chlorocebus aethiops , ADN Complementario/genética , Humanos , Lisina/genética , Virus de la Parotiditis/patogenicidad , Virus de la Parotiditis/fisiología , Mutación , ARN Viral/genética , Ratas , Ratas Endogámicas Lew , Análisis de Secuencia de ADN , Vacunas Atenuadas/genética , Células Vero , Virulencia , Replicación ViralRESUMEN
Two in-frame, C-terminal isoforms of the herpes simplex virus type 1 (HSV-1) origin binding protein (OBP), OBPC-1 and OBPC-2, and a unique C-terminal transcript, UL8.5, are specified by HSV-1 DNA. As the first isoform identified, OBPC-1 was initially assumed to be the product of the UL8.5 transcript. Recent evidence has demonstrated, however, that OBPC-1 is a cathepsin B-mediated cleavage product of OBP, suggesting that OBPC-2 is the product of the UL8.5 transcript. Because both OBPC-1 and -2 contain the majority of the OBP DNA binding domain, we hypothesized that both may be involved in regulating origin-dependent, OBP-mediated viral DNA replication. In this paper, we demonstrate that OBPC-2 is, indeed, the product of the UL8.5 transcript. The translational start site of OBPC-2 was mapped, and a virus (M571A) that does not express this protein efficiently was constructed. Using M571A, we have shown that OBPC-2 is able to bind origin DNA, even though it lacks seven N-terminal amino acid residues of the previously mapped OBP DNA binding domain, resulting in a revision of the limits of the OBP DNA binding domain. Consistent with their proposed roles in regulating viral DNA replication, OBPC-1 and -2 act together to down-regulate viral DNA replication in vitro. During functional studies in vivo, OBPC-2 was identified as a factor that increases mortality in the mouse ocular model of HSV-1 infection.
Asunto(s)
Replicación del ADN , Herpes Simple/virología , Herpesvirus Humano 1/fisiología , Complejo de Reconocimiento del Origen/metabolismo , Replicación Viral/genética , Animales , ADN Viral/biosíntesis , Herpesvirus Humano 1/genética , Ratones , Mutación , Complejo de Reconocimiento del Origen/genética , Biosíntesis de Proteínas , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estructura Terciaria de Proteína , Origen de Réplica , Transcripción Genética , Latencia del Virus/genéticaRESUMEN
Although the seven viral proteins required for herpes simplex virus type 1 (HSV-1) DNA replication have been identified, the mechanism by which viral DNA synthesis is regulated is unclear. HSV-1 DNA replication is thought to occur in two stages: origin-dependent DNA replication (stage I) mediated by the origin binding protein (OBP), followed by origin- and OBP-independent DNA replication (stage II). The mechanism that facilitates the switch from stage I to stage II is unknown; however, it must involve the loss of OBP function or OBP itself from the replication initiation complex. Previous studies from this laboratory identified a transcript (UL8.5) and protein (OBPC) that are in frame with and comprise the C terminus of the gene specifying OBP. Because of its DNA binding ability, OBPC has been hypothesized to mediate the switch from stage I to stage II. Here, we identify a second protein (OBPC-2) that is also in frame with the C terminus of OBP but comprises a smaller portion of the protein. We demonstrate that the protein originally identified (OBPC-1) is a cathepsin B-mediated cleavage product of OBP, while OBPC-2 may be the product of the UL8.5 transcript. We further demonstrate that the cleavage of OBP to yield OBPC-1 is dependent upon viral DNA replication. These results suggest that cleavage may be a mechanism by which OBP levels and/or activity are regulated during infection.
Asunto(s)
Catepsina B/metabolismo , Proteínas de Unión al ADN/metabolismo , Herpesvirus Humano 1/fisiología , Proteínas Virales/metabolismo , Replicación Viral/fisiología , Animales , Chlorocebus aethiops , Modelos Biológicos , Isoformas de Proteínas , Procesamiento Proteico-Postraduccional , Células Vero , Proteínas Virales/genéticaRESUMEN
Many acyclovir-resistant herpes simplex virus isolates from patients contain insertions or deletions in homopolymeric sequences in the thymidine kinase (TK) gene (tk). Viruses that have one (G8) or two (G9) base insertions in a run of seven G's (G string) synthesize low levels of active TK (TK-low phenotype), evidently via ribosomal frameshifting. These levels of TK can suffice to permit reactivation from latently infected mouse ganglia, but in a majority of ganglia, especially with the G9 virus, reactivation of virus that has reverted to the TK-positive phenotype predominates. To help address the relative contributions of translational mechanisms and reversion in reactivation, we generated viruses with a base either inserted or deleted just downstream of the G string. Both of these viruses had a TK-low phenotype similar to that of the G8 and G9 viruses but with less reversion. Both of these viruses reactivated from latently infected trigeminal ganglia, albeit inefficiently, and most viruses that reactivated had a uniformly TK-low phenotype. We also generated viruses that have one insertion in a run of six C's or one deletion in a run of five C's. These viruses lack measurable TK activity. However, they reactivated from latently infected ganglia, albeit inefficiently, with the reactivating viruses having reverted to the wild-type TK phenotype. Therefore, for G-string mutants, levels of active TK as low as 0.25% generated by translational mechanisms can suffice for reactivation, but reversion can also contribute. For viruses that lack TK activity due to mutations on other homopolymeric sequences, reactivation can occur via reversion.
Asunto(s)
Secuencia de Bases/genética , Farmacorresistencia Viral/genética , Herpesvirus Humano 1/genética , Eliminación de Secuencia , Timidina Quinasa/genética , Proteínas Virales/genética , Aciclovir/farmacología , Animales , Antivirales/farmacología , Chlorocebus aethiops , Farmacorresistencia Viral/efectos de los fármacos , Herpes Simple/enzimología , Herpes Simple/genética , Herpesvirus Humano 1/enzimología , Humanos , Ratones , Mutagénesis Insercional , Biosíntesis de Proteínas/efectos de los fármacos , Biosíntesis de Proteínas/genética , Ganglio del Trigémino/virología , Células Vero , Activación Viral/efectos de los fármacos , Activación Viral/genéticaRESUMEN
Arcanobacterium pyogenes is an opportunistic pathogen associated with suppurative diseases in economically important food animals such as cattle, pigs, and turkeys. A. pyogenes adheres to host epithelial cells, and adhesion is promoted by the action of neuraminidase, which is expressed by this organism. However, a neuraminidase-deficient mutant of A. pyogenes only had a reduced ability to adhere to host epithelial cells, indicating that other factors are involved in adhesion. Far Western blotting revealed the presence of an approximately 120-kDa A. pyogenes cell wall protein that binds collagen type I. The 3.5-kb gene that encodes the 124.7-kDa CbpA protein was cloned, and sequence analysis indicated that CbpA contains a typical MSCRAMM protein domain structure. Recombinant, six-His-tagged CbpA (HIS-CbpA) was capable of binding collagen types I, II, and IV but not fibronectin. In addition, CbpA was involved in the ability of A. pyogenes to adhere to HeLa and 3T6 cells, as a cbpA knockout strain had 38.2 and 57.0% of wild-type adhesion, respectively. This defect could be complemented by providing cbpA on a multicopy plasmid. Furthermore, HIS-CbpA blocked A. pyogenes adhesion to HeLa or 3T6 cells in a dose-dependent manner. cbpA was only present in 48% of the A. pyogenes strains tested (n = 75), and introduction of plasmid-encoded cbpA into a naturally cbpA-deficient strain increased the ability of this strain to bind to HeLa and 3T6 cells 2.9- and 5.7-fold, respectively. These data indicate that CbpA, a collagen-binding protein of A. pyogenes, plays a role in the adhesion of this organism to host cells.