嗜神经性疱疹病毒：神经科学研究的有力工具

病毒是研究现代神经科学的有力工具。对神经元的连接方式及功能研究大都是利用重组病毒完成的,嗜神经性疱疹病毒便是其中一种重要工具。随着基因工程学以及分子生物学技术的不断发展,多种嗜神经性疱疹病毒被改造为不同的重组病毒工具应用于神经科学研究。本文基于几种常见且应用较为广泛的嗜神经性疱疹病毒作为神经传导示踪工具、治疗神经性疾病的病毒载体和溶瘤病毒治疗神经肿瘤等应用进行阐述及讨论,为进一步开发嗜神经性疱疹病毒的功能提供参考。     

Cellular Inflammatory Response to Flaviviruses in the Central Nervous System of a Primate Host

Flaviviruses such as tick-borne encephalitis virus, Japanese encephalitis virus, West Nile virus, and St. Louis encephalitis virus are important neurotropic human pathogens, typically causing a devastating and often fatal neuroinfection. Flaviviruses induce neuroinflammation with typical features of viral encephalitides, including inflammatory cell infiltration, activation of microglia, and neuronal degeneration. Development of safe and effective live-virus vaccines against neurotropic flavivirus infections demands a detailed knowledge of their neuropathogenesis in a primate host that is evolutionarily close to humans. Here, we used computerized morphometric analysis to quantitatively assess the cellular inflammatory responses in the central nervous system (CNS) of rhesus monkeys infected with three antigenically divergent attenuated flaviviruses. The kinetics, spatial pattern, and magnitude of microglial activation, trafficking of T and B cells, and changes in T cell subsets within the CNS define unique phenotypic signatures for each of the three viruses. Our results provide a benchmark for investigation of cellular inflammatory responses induced by attenuated flaviviruses in the CNS of primate hosts and provide insight into the neuropathogenesis of flavivirus encephalitis that might guide the development of safe and effective live-virus vaccines. (J Histochem Cytochem 57:973–989, 2009)     

CRISPR/Cas9, a powerful tool to target human herpesviruses

Over 90% of the adult population is infected with one or multiple herpesviruses. These viruses are characterized by their ability to establish latency, where the host is unable to clear the invader from infected cells resulting in a lifelong infection. Herpesviruses cause a wide variety of (recurrent) diseases such as cold sores, shingles, congenital defects and several malignancies. Although the productive phase of a herpesvirus infection can often be efficiently limited by nucleoside analogs, these drugs are ineffective during a latent herpesvirus infection and are therefore unable to clear herpesviruses from the human host. Advances in genome engineering using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 facilitates virus research and may hold potential to treat or cure previously incurable herpesvirus infections by directly targeting these viruses within infected cells. Here, we review recent applications of the CRISPR/Cas9 system for herpesviral research and discuss the therapeutic potential of the system to treat, or even cure, productive and latent herpesviral infections.     

Diva vaccines that reduce virus transmission.

This brief review deals with the effect of diva (Differentiating Infected from VAccinated individuals) vaccines (also termed marker vaccines) on transmission of herpesviruses and pestiviruses in swine and cattle. Pseudorabies and bovine herpesvirus 1 diva vaccines have been demonstrated to reduce transmission of wild-type virus in populations of pigs and cattle in the laboratory as well as in the field. A subunit diva vaccine based on the immunodominant E2 protein of classical swine fever virus that is expressed in the baculovirus system may reduce transmission of wild-type virus among pigs and also transmission from mother to foetuses. A similar diva vaccine against bovine virus diarrhoea infections protected sheep against transplacental transmission of antigenically homologous wild-type virus. Diva vaccines along with their companion diagnostic tests can play a role in control of infections, ultimately leading to eradication of viruses.     

Bovine cells expressing bovine herpesvirus 1 (BHV-1) glycoprotein IV resist infection by BHV-1, herpes simplex virus, and pseudorabies virus.

We expressed the bovine herpesvirus 1 (BHV-1) glycoprotein IV (gIV) in bovine cells. The protein expressed was identical in molecular mass and antigenic reactivity to the native gIV protein but was localized in the cytoplasm. Expressing cells were partially resistant to BHV-1, herpes simplex virus, and pseudorabies virus, as shown by a 10- to 1,000-fold-lower number of plaques forming on these cells than on control cells. The level of resistance depended on the level of gIV expression and the type and amount of challenge virus. These data are consistent with previous reports by others that cellular expression of the BHV-1 gIV homologs, herpes simplex virus glycoprotein D, and pseudorabies virus glycoprotein gp50 provide partial resistance against infection with these viruses. We have extended these findings by showing that once BHV-1 enters gIV-expressing cells, it replicates and spreads normally, as shown by the normal size of BHV-1 plaques and the delayed but vigorous synthesis of viral proteins. Our data are consistent with the binding of BHV-1 gIV to a cellular receptor required for initial penetration by all three herpesviruses and interference with the function of that receptor molecule.     

Herpes virus replication

Herpesviruses are large double stranded DNA animal viruses with the distinguishing ability to establish latent, life-long infections. To date, eight human herpesviruses that exhibit distinct biological and corresponding pathological/clinical properties have been identified. During their life cycles, herpesviruses execute an intricate chain of events geared towards optimizing their replication. This sets an interesting paradigm to study fundamental biological processes. This review summarizes recent developments in herpesvirus research with emphasis on genome transactions, particularly with respect to the prototypic herpes simplex virus type-1.     

Exploiting human herpesvirus immune evasion for therapeutic gain: potential and pitfalls

Herpesviruses stand out for their capacity to establish lifelong infections of immunocompetent hosts, generally without causing overt symptoms. Herpesviruses are equipped with sophisticated immune evasion strategies, allowing these viruses to persist for life despite the presence of a strong antiviral immune response. Although viral evasion tactics appear to target virtually any stage of the innate and adaptive host immune response, detailed knowledge is now available on the molecular mechanisms underlying herpesvirus obstruction of MHC class I-restricted antigen presentation to T cells. This opens the way for clinical application. Here, we review and discuss recent efforts to exploit human herpesvirus MHC class I evasion strategies for the rational design of novel strategies for vaccine development, cancer treatment, transplant protection and gene therapy.     

Cross-Reactivity of Neutralizing Antibodies among Malignant Catarrhal Fever Viruses

Some members of the gamma herpesvirus genus Macavirus are maintained in nature as subclinical infections in well-adapted ungulate hosts. Transmission of these viruses to poorly adapted hosts, such as American bison and cattle, can result in the frequently fatal disease malignant catarrhal fever (MCF). Based on phylogenetic analysis, the MCF viruses (MCFV) cluster into two subgroups corresponding to the reservoir hosts’ subfamilies: Alcelaphinae/Hippotraginae and Caprinae. Antibody cross-reactivity among MCFVs has been demonstrated using techniques such as enzyme linked immunosorbent and immunofluorescence assays. However, minimal information is available as to whether virus neutralizing antibodies generated against one MCFV cross react with other members of the genus. This study tested the neutralizing activity of serum and plasma from select MCFV-infected reservoir hosts against alcelaphine herpesvirus 1 (AlHV-1) and ovine herpesvirus 2 (OvHV-2). Neutralizing antibody activity against AlHV-1 was detected in samples from infected hosts in the Alcelaphinae and Hippotraginae subfamilies, but not from hosts in the Caprinae subfamily. OvHV-2 neutralizing activity was demonstrated in samples from goats (Caprinae) but not from wildebeest (Alcelaphinae). These results show that neutralizing antibody cross reactivity is present to MCFVs within a virus subgroup but not between subgroups. This information is important for diagnosing infection with MCFVs and in the development of vaccines against MCF.     

Specificity of cleavage in replicative-form DNA of bovine herpesvirus 1.

The linear double-stranded DNA genome of herpesvirus as it is present in infectious virions needs to be circularized after infection of host cells and before DNA replication. Replicative-form genomes have to be cleaved into linear unit-length molecules during virion maturation and are most probably the substrate for inversion of the short segment relative to the long segment of the bovine herpesvirus 1 (BHV-1) genome. Those regions of the BHV-1 genome which are functionally involved in these processes have been analyzed at the molecular level by cloning and sequencing the genomic termini, the fusion of both termini from replicative-form molecules, and the junction between the short and the long genome segment. On the basis of the simple genome arrangement of BHV-1, it was inferable that the cleavage of replicative-form genomes by a hypothetical BHV-1 terminase activity may be specified by a sequence at the left end of UL (An element), which is located proximal to a reiterated beta element that makes up the cleavage site itself. The relationship of those elements in BHV-1 and the comparison to similar regions of other herpesviruses indicate consensus sequence elements which are functionally important for cleavage and isomerization of viral DNA during maturation of virions.     

Forward with BACs: new tools for herpesvirus genomics

The large, complex genomes of herpesviruses document the high degree of adaptation of these viruses to their hosts. Not surprisingly, the methods developed over the past 30 years to analyse herpesvirus genomes have paralleled those used to investigate the genetics of eukaryotic cells. The recent use of bacterial artificial chromosome (BAC) technology in herpesvirus genetics has made their genomes accessible to the tools of bacterial genetics. This has opened up new avenues for reverse and forward genetics of this virus family in basic research, and also for vector and vaccine development.     

Simian homologues of human gamma-2 and betaherpesviruses in mandrill and drill monkeys

Recent serological and molecular surveys of different primate species allowed the characterization of several Kaposi's sarcoma-associated herpesvirus (KSHV) homologues in macaques, African green monkeys, chimpanzees, and gorillas. Identification of these new primate rhadinoviruses revealed the existence of two distinct genogroups, called RV1 and RV2. Using a degenerate consensus primer PCR method for the herpesvirus DNA polymerase gene, the presence of KSHV homologues has been investigated in two semi-free-ranging colonies of eight drill (Mandrillus leucophaeus), five mandrill (Mandrillus sphinx), and two hybrid (Mandrillus leucophaeus-Mandrillus sphinx) monkeys, living in Cameroon and Gabon, Central Africa. This search revealed the existence of not only two distinct KSHV homologues, each one belonging to one of the two rhadinovirus genogroups, but also of two new betaherpesvirus sequences, one being close to cytomegaloviruses and the other being related to human herpesviruses 6 and 7 (HHV-6 and -7). The latter viruses are the first simian HHV-6 and -7 homologues identified to date. These data show that mandrill and drill monkeys are the hosts of at least four novel distinct herpesviruses. Moreover, mandrills, like macaques and African green monkeys, harbor also two distinct gamma-2 herpesviruses, thus strongly suggesting that a second gamma-2 herpesvirus, belonging to the RV2 genogroup, may exist in humans.     

Bovine herpesvirus 5 glycoprotein E is important for neuroinvasiveness and neurovirulence in the olfactory pathway of the rabbit

Glycoprotein E (gE) is important for full virulence potential of the alphaherpesviruses in both natural and laboratory hosts. The gE sequence of the neurovirulent bovine herpesvirus 5 (BHV-5) was determined and compared with that of the nonneurovirulent BHV-1. Alignment of the predicted amino acid sequences of BHV-1 and BHV-5 gE open reading frames showed that they had 72% identity and 77% similarity. To determine the role of gE in the differential neuropathogenesis of BHV-1 and BHV-5, we have constructed BHV-1 and BHV-5 recombinants: gE-deleted BHV-5 (BHV-5gEDelta), BHV-5 expressing BHV-1 gE (BHV-5gE1), and BHV-1 expressing BHV-5 gE (BHV-1gE5). Neurovirulence properties of these recombinant viruses were analyzed using a rabbit seizure model (S. I. Chowdhury et al., J. Comp. Pathol. 117:295-310, 1997) that distinguished wild-type BHV-1 and -5 based on their differential neuropathogenesis. Intranasal inoculation of BHV-5 gEDelta and BHV-5gE1 produced significantly reduced neurological signs that affected only 10% of the infected rabbits. The recombinant BHV-1gE5 did not invade the central nervous system (CNS). Virus isolation and immunohistochemistry data suggest that these recombinants replicate and spread significantly less efficiently in the brain than BHV-5 gE revertant or wild-type BHV-5, which produced severe neurological signs in 70 to 80% rabbits. Taken together, the results of neurological signs, brain lesions, virus isolation, and immunohistochemistry indicate that BHV-5 gE is important for efficient neural spread and neurovirulence within the CNS and could not be replaced by BHV-1 gE. However, BHV-5 gE is not required for initial viral entry into olfactory pathway.     

Concluding overview: looking back, looking forward

The gamma-herpesviruses are a group of related agents which share the same broad strategy for infection of and persistence within the lymphoid tissues of their hosts. Yet in evolutionary terms these agents are sufficiently diverse to display multiple different molecular mechanisms whereby that strategy can be achieved. Attempts are made to relate the different in vitro growth transforming capacities of the gamma1-herpesviruses, the T-lymphotropic gamma2-herpesviruses and the B-lymphotropic gamma2-herpesviruses to what is known about the biology of these virus infections in their natural or in experimental hosts. The review then summarizes the evidence linking gamma-herpesviruses with oncogenesis and proposes that the diverse spectrum of Epstein-Barr virus and human herpesvirus 8-associated human tumours falls into three pathogenetically distinct categories. Many questions remain unanswered in the areas of gamma-herpesvirus biology and disease pathogenesis: resolving these questions will require a broadening of our experimental approaches and a willingness to relinquish 'single-model' panaceas.     

Serological survey of herpesvirus infections in wild ruminants of France and Belgium

The presence of antibodies against bovine herpesvirus 1 (BHV-1), bovid herpesvirus 6 (BHV-6), herpesvirus of Cervidae type 1 (HVC-1), reindeer herpesvirus, bovine herpesvirus 2 (BHV-2) and bovid herpesvirus 4 (BHV-4) was investigated in wild ruminants of France and Belgium between 1981 and 1986. There were no animals serologically positive for BHV-4. Antibodies against BHV-2 were demonstrated in roe deer (Cervus capreolus) (less than 1%) and chamois (Rupicapra rupicapra) (1%) in France. Animals seropositive to the four related viruses (BHV-1, BHV-6, HVC-1, reindeer herpesvirus) were detected in red deer (Cervus elaphus) in France and Belgium (1% and 11%, respectively), in roe deer (less than 1%) from France, in chamois (4%) in France and in ibex (Capra ibex) (4%) from France. The presence of antibodies against HVC-1, especially in red deer from Belgium, may suggest that wild ruminants in continental Europe are now infected with this virus, which previously has been isolated only in Scotland.     

The gE and gI homologs from two alphaherpesviruses have conserved and divergent neuroinvasive properties.

The membrane glycoproteins gE and gI are encoded by pseudorabies virus (PRV), a neurotropic, broad-host-range alphaherpesvirus of swine. PRV gE and gI are required for anterograde spread to a restricted set of retinorecipient neurons in the brain after infection of the rat retina. A related alphaherpesvirus, encoding gE and gI homologs, is called bovine herpesvirus 1.1 (BHV-1.1). BHV-1.1 is a respiratory pathogen of highly restricted host range and, in contrast to PRV, is unable to propagate in or cause disease in rodents. We have shown previously that the BHV-1.1 gE and gI proteins are capable of complementing the virulence functions of PRV gE and gI in a rodent model (A. C. Knapp and L. W. Enquist, J. Virol. 71:2731-2739, 1997). We examined the ability of the BHV-1.1 gE and gI homologs to direct circuit-specific invasion of the rat central nervous system by PRV. Both complete open reading frames were cloned into a PRV mutant lacking the PRV gE and gI genes. Recombinant viruses were analyzed for the ability to invade the rat brain after infection of the retina. Surprisingly, in a portion of the animals tested, the BHV-1.1 gE and gI proteins functioned autonomously to promote spread of PRV to a subset of retinorecipient regions of the brain. First, the presence of BHV-1.1 gI alone, but not PRV gI alone, promoted viral invasion of the optic tectum. Second, expression of BHV-1.1 gE alone facilitated PRV infection of a subset of neurons in the hippocampus not normally infected by PRV. When both BHV-1.1 proteins were expressed in a coinfection, all retinorecipient regions of the rat brain were infected. Therefore, depending on the viral source, homologs of gE and gI differentially affect spread between synaptically connected neurons in the rat.     

Recent patents involving virus nucleotide sequences; host defense, RNA silencing and expression vector strategies

Improved knowledge of the molecular biology of viruses, including recent gains in virus sequence data analysis, has greatly contributed to recent innovations in medical diagnostics, therapeutics, drug development and other related areas. Virus sequences have been used for the development of vaccines and antiviral agents to block the spread of viral infections, as well as to target and battle chronic diseases such as cancer. Virus sequences are now routinely employed in a wide array of RNA silencing technologies. Viruses can also be engineered into expression vectors which in turn can be used as protein production platforms as well as delivery vehicles for gene therapies. This review article outlines a number of patents that have been recently issued with respect to virus sequence data and describes some of their biotechnological applications.     

Abduction of Chemokine Elements by Herpesviruses

Chemokines play a key role in orchestrating leukocytic recruitment during inflammatory responses, including those to viral infections. Chemokines are soluble cytokines which mediate their effects through specific G protein-coupled, seven-transmembrane receptors which are expressed on a wide range of cells, including monocytes, T-cells, dendritic cells, and NK cells. Analyses of herpesvirus genomes have revealed that these viral pathogens encode their own versions of both chemokines and chemokine receptors. Viral genes encoding chemokine elements were likely to have been acquired from the host genome and have been remodeled during virus evolution to presumably optimize function or acquire new properties not displayed by their cellular homologues. Virus-encoded chemokines and chemokine receptors are important players in the continuing confrontation between viruses and their mammalian hosts. Detailed characterization of these elements will provide a better understanding of how the immune system responds to viral infection and may suggest new antiviral drug targets and new avenues for the development of antiviral therapies. We will review here the chemokine elements encoded by herpesviruses and how they may aid viral infection and propagation.     

Ocular herpes:the pathophysiology,management and treatment of herpetic eye diseases

Herpesviruses are a prominent cause of human viral disease, second only to the cold and influenza viruses. Most herpesvirus infections are mild or asymptomatic. However, when the virus invades the eye, a number of pathologies can develop and its associated sequelae have become a considerable source of ocular morbidity. The most common culprits of herpetic eye disease are the herpes simplex virus(HSV), varicella zoster virus(VZV), and cytomegalovirus(CMV). While primary infection can produce ocular disease, the most destructive manifestations tend to arise from recurrent infection. These recurrent infections can wreck devastating effects and lead to irreversible vision loss accompanied by a decreased quality of life, increased healthcare usage, and significant cost burden. Unfortunately, no method currently exists to eradicate herpesviruses from the body after infection. Treatment and management of herpes-related eye conditions continue to revolve around antiviral drugs, although corticosteroids, interferons, and other newer therapies may also be appropriate depending on the disease presentation. Ultimately, the advent of effective vaccines will be crucial to preventing herpesvirus diseases altogether and cutting the incidence of ocular complications.     

Ocular herpes: the pathophysiology,management and treatment of herpetic eye diseases

Herpesviruses are a prominent cause of human viral disease, second only to the cold and influenza viruses. Most herpesvirus infections are mild or asymptomatic. However, when the virus invades the eye, a number of pathologies can develop and its associated sequelae have become a considerable source of ocular morbidity. The most common culprits of herpetic eye disease are the herpes simplex virus (HSV), varicella zoster virus (VZV), and cytomegalovirus (CMV). While primary infection can produce ocular disease, the most destructive manifestations tend to arise from recurrent infection. These recurrent infections can wreck devastating effects and lead to irreversible vision loss accompanied by a decreased quality of life, increased healthcare usage, and significant cost burden. Unfortunately, no method currently exists to eradicate herpesviruses from the body after infection. Treatment and management of herpes-related eye conditions continue to revolve around antiviral drugs, although corticosteroids, interferons, and other newer therapies may also be appropriate depending on the disease presentation. Ultimately, the advent of effective vaccines will be crucial to preventing herpesvirus diseases altogether and cutting the incidence of ocular complications.     

Progress towards rabies control.

Although safe and efficacious tissue-culture-derived rabies vaccines are available in developed countries, much of the world still depends on vaccines derived from neural tissue which were introduced half a century ago. Considerable advances have been made in our understanding of the molecular biology of rabies virus, and genetically engineered recombinant viruses (vaccinia-rabies virus glycoprotein) have been developed. These may facilitate the control of rabies in some species by oral vaccination campaigns.