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PubMed2003

Antigenic variations in the CD4 induced sites of the CCR5-tropic, pathogenic SHIVsf162p3 gp120 variants.

In vivo passage of non-pathogenic, CCR5-tropic simian/human immunodeficiency virus (SHIV) - SHIVsf162 resulted in a pathogenic isolate, SHIVsf162p3. In an attempt to characterize envelope (Env)-mediated properties that may contribute to its pathogenicity, major (P3 major) and minor (P3 minor) Env gp120 variants were cloned from the plasma of a SHIVsf162p3-infected animal, and expressed in the context of luciferase reporter viruses. Entry mediated by these envelopes and susceptibility to neutralization by CD4 induced-site (CD4i) antibodies (MAbs) was analyzed in comparison to parental SF162. Sequence analysis revealed that the P3 major and minor variant Envs contained 14 and 17 amino acid changes, respectively, compared with SF162. The rank order of entry mediated by the three envelopes was P3 major > SF162 > P3 minor, whereas the reverse order was observed for susceptibility to neutralization by CD4i MAbs. Since CD4i epitopes overlap the coreceptor (CoR) binding site, these findings suggest that the amino acid changes accumulated upon in vivo passage of SHIVsf162 result in Env gp120 structural rearrangements that modulate the exposure and/or conformation of the CoR binding site. This, in turn, led to increased entry and infectivity of the P3 major variant and may be responsible, in part, for the enhanced pathogenicity of SHIVsf162p3.

Amino Acid Sequence↗

PubMed2001

Regulation of human immunodeficiency virus type 1 infection, beta-chemokine production, and CCR5 expression in CD40L-stimulated macrophages: immune control of viral entry.

Mononuclear phagocytes (MP) and T lymphocytes play a pivotal role in the host immune response to human immunodeficiency virus type 1 (HIV-1) infection. Regulation of such immune responses can be mediated, in part, through the interaction of the T-lymphocyte-expressed molecule CD40 ligand (CD40L) with its receptor on MP, CD40. Upregulation of CD40L on CD4+ peripheral blood mononuclear cells during advanced HIV-1 disease has previously been reported. Based on this observation, we studied the influence of CD40L-CD40 interactions on MP effector function and viral regulation in vitro. We monitored productive viral infection, cytokine and beta-chemokine production, and beta-chemokine receptor expression in monocyte-derived macrophages (MDM) after treatment with soluble CD40L. Beginning 1 day after infection and continuing at 3-day intervals, treatment with CD40L inhibited productive HIV-1 infection in MDM in a dose-dependent manner. A concomitant and marked upregulation of beta-chemokines (macrophage inhibitory proteins 1alpha and 1beta and RANTES [regulated upon activation normal T-cell expressed and secreted]) and the proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) was observed in HIV-1-infected and CD40L-treated MDM relative to either infected or activated MDM alone. The addition of antibodies to RANTES or TNF-alpha led to a partial reversal of the CD40L-mediated inhibition of HIV-1 infection. Surface expression of CD4 and the beta-chemokine receptor CCR5 was reduced on MDM in response to treatment with CD40L. In addition, treatment of CCR5- and CD4-transfected 293T cells with secretory products from CD40L-stimulated MDM prior to infection with a CCR5-tropic HIV-1 reporter virus led to inhibition of viral entry. In conclusion, we demonstrate that CD40L-mediated inhibition of viral entry coincides with a broad range of MDM immune effector responses and the down-modulation of CCR5 and CD4 expression.

CD40 Ligand↗

PubMed2001

Replacement of the V3 region of gp120 with SDF-1 preserves the infectivity of T-cell line-tropic human immunodeficiency virus type 1.

Interaction between the human immunodeficiency virus type 1 (HIV-1) envelope and the relevant chemokine receptors is crucial for subsequent membrane fusion and viral entry. Although the V3 region of gp120 is known to determine the cell tropism as well as the coreceptor usage, the significance of the binding of the V3 region to the chemokine receptor has not been fully understood. To address this issue, we adopted the pseudotyped virus infection assay in which the V3 region of the T-cell line-tropic (T-tropic) NL4-3 envelope was replaced with a portion of stromal cell-derived factor 1 (SDF-1), the ligand of CXCR4. The V3 region of the NL4-3 envelope expression vector was replaced with three different stretches of SDF-1 cDNA. Expression of each chimeric envelope protein was confirmed by immunoprecipitation and Western blotting. Luciferase reporter viruses were prepared by cotransfection of the pNL4-3.Luc.E(-)R(-) vector and each chimeric envelope expression vector, and the infection assay was then carried out. We showed that pseudotyped viruses with one of the chimeric envelopes, NL4-3/SDF1-51, could infect U87.CD4.CXCR4 but not U87.CD4 or U87.CXCR4 cells and that this infection was inhibited by the ligand of CXCR4, SDF-1beta, by anti-human SDF-1 antibody, or by an anti-CD4 antibody, Leu3a, in a dose-dependent manner. Furthermore, chimeric NL4-3/SDF1-51 gp120 significantly inhibited binding of labeled SDF-1 to CXCR4. It was suggested that replacement of the V3 region of the NL4-3 envelope with SDF-1 preserved the CD4-dependent infectivity of T-tropic HIV-1. These results indicate that binding between the V3 region and the relevant coreceptor is important for viral entry, whether its amino acid sequence is indigenous to the virus or not.

Amino Acid Sequence↗

PubMed1998

Social stress and the reactivation of latent herpes simplex virus type 1.

Psychological stress is thought to contribute to reactivation of latent herpes simplex virus (HSV). Although several animal models have been developed in an effort to reproduce different pathogenic aspects of HSV keratitis or labialis, until now, no good animal model existed in which application of a psychological laboratory stressor results in reliable reactivation of the virus. Reported herein, disruption of the social hierarchy within colonies of mice increased aggression among cohorts, activated the hypothalamic-pituitary-adrenal axis, and caused reactivation of latent HSV type 1 in greater than 40% of latently infected animals. However, activation of the hypothalamic-pituitary-adrenal axis using restraint stress did not activate the latent virus. Thus, the use of social stress in mice provides a good model in which to investigate the neuroendocrine mechanisms that underlie behaviorally mediated reactivation of latent herpesviruses.

Animals↗

PubMed2002

Nef-mediated downregulation of CD4 enhances human immunodeficiency virus type 1 replication in primary T lymphocytes.

The accessory protein Nef plays a crucial role in primate lentivirus pathogenesis. Nef enhances human immunodeficiency virus type 1 (HIV-1) infectivity in culture and stimulates viral replication in primary T cells. In this study, we investigated the relationship between HIV-1 replication efficiency in CD4(+) T cells purified from human blood and two various known activities of Nef, CD4 downregulation and single-cycle infectivity enhancement. Using a battery of reporter viruses containing point mutations in nef, we observed a strong genetic correlation between CD4 downregulation by Nef during acute HIV-1 infection of activated T cells and HIV-1 replication efficiency in T cells. In contrast, HIV-1 replication ability was not significantly correlated with the ability of Nef to enhance single-cycle virion infectivity, as determined by using viruses produced in cells lacking CD4. These results demonstrate that CD4 downregulation by Nef plays a crucial role in HIV-1 replication in activated T cells and underscore the potential for the development of therapies targeting this conserved activity of Nef.

CD4 Antigens↗

PubMed2000

Molecular function of the CD4 D1 domain in coreceptor-mediated entry by HIV type 1.

The surface molecule CD4 plays a key role in initiating cellular entry by the human immunodeficiency virus type 1 (HIV-1), and it is now recognized as acting synergistically with select chemokine receptors (coreceptors) in the infection process. The present study was undertaken to determine whether the extracellular region of CD4 is sufficient to induce fusion of HIV-1 virions with target cells in the absence of its anchoring function. Using pseudotype reporter viruses to quantitate infection, soluble CD4 (sCD4) was tested for its ability to induce fusion by viruses utilizing CCR5 as their coreceptor. We found that sCD4 was competent to replace membrane-bound CD4 to trigger infection mediated by several HIV-1 envelopes. Furthermore, in a comparison of the envelopes of HIV-1 NL4-3 and a chimera containing the gp120 V3 loop of Ba-L, the V3 region was found to be one factor affecting susceptibility to induction by sCD4. In addition, using truncated and mutant derivatives of sCD4, the amino-terminal D1 domain of CD4 was found to be necessary and sufficient for induction of fusion and to require an intact gp120-binding site for this activity. These results delineate determinants on CD4 and gp120 required for fusion induction in collaboration with a coreceptor, and suggest a mechanism whereby CD4 may contribute to viral infection in trans.

CD4 Antigens↗

PubMed1991

CD4: its structure, role in immune function and AIDS pathogenesis, and potential as a pharmacological target.

CD4 is critical for the development and function of the CD4+ subset of T cells and also subserves as the receptor for the human immunodeficiency viruses. Reports in the past year clarify the role and the molecular interactions of CD4 in these events. Determination of the structure of an extracellular fragment of CD4 reveals novel variations of the immunoglobulin fold and provides an atomic framework for interpretation of its interactions with MHC class II molecules and with gp120, the external envelope glycoprotein of the human immunodeficiency virus.

Acquired Immunodeficiency Syndrome↗

PubMed1995

Stringent chemical and thermal regulation of recombinant gene expression by vaccinia virus vectors in mammalian cells.

We developed a stringently regulated expression system for mammalian cells that uses (i) the RNA polymerase, phi 10 promoter, and T phi transcriptional terminator of bacteriophage T7; (ii) the lac repressor, lac operator, rho-independent transcriptional terminators and the gpt gene of Escherichia coli; (iii) the RNA translational enhancer of encephalomyocarditis virus; and (iv) the genetic background of vaccinia virus. In cells infected with the recombinant vaccinia virus, reporter beta-galactosidase synthesis was not detected in the absence of inducer. An induction of at least 10,000- to 20,000-fold occurred upon addition of isopropyl beta-D-thiogalactopyranoside or by temperature elevation from 30 to 37 degrees C using a temperature-sensitive lac repressor. Regulated synthesis of the secreted and highly glycosylated human immunodeficiency virus 1 envelope protein gp120 was also demonstrated. Yields of both proteins were approximately 2 mg per 10(8) cells in 24 hr. Plasmid transfer vectors for cloning and expression of complete or incomplete open reading frames in recombinant vaccinia viruses are described.

Animals↗

PubMed1997

Macrophage tropism of human immunodeficiency virus type 1 and utilization of the CC-CKR5 coreceptor.

The recent identification of the CC-CKR5 beta chemokine receptor as a major cofactor for entry of macrophage-tropic isolates of human immunodeficiency virus type 1 (HIV-1) raises the question of whether macrophage tropism is determined by utilization of this chemokine receptor. We observe that in addition to macrophage-tropic isolates of clades A, B, and E, macrophage-tropic isolates of clade F also utilize the CC-CKR5 molecule for entry. However, using single-round replication-competent reporter viruses carrying the envelope genes of T-cell line-tropic or macrophage-tropic phenotypic recombinant and mutant HIV-1 strains in infection of stable cell lines that coexpress the CD4 and chemokine receptors, we were unable to establish a strict correlation between macrophage tropism and utilization of the CC-CKR5 chemokine receptor. This latter finding suggests that a cofactor other than CC-CKR5 serves to determine entry into primary macrophages.

Amino Acid Sequence↗

PubMed1979

Chorioretinitis of the newborn with herpes simplex virus type 1. Report of a case.

A disseminated herpes simplex virus (HSV) infection involving the central nervous system and accompanied by chrioretinitis in a a 3-week-old girl is described. The aetiologic diagnosis was established on the basis of vrius isolated from skin vesicles, and a significant rise in complement fixing antibodies to HSV type 1. The mode of transmission of the virus to the infant apparently was direct contact with an oral lesion in the mother, that was present at the time of delivery. The patient survived but became blind and microcephalic, with severe neurological sequelae. The virus isolated was identified as HSV type 1, which is an infrequent finding in herpetic chorioretinitis of the newborn.

Adult↗

PubMed2004

Differential regulation of human immunodeficiency virus type 2 and simian immunodeficiency virus promoter activity.

Promoter activity of the HIV-1 long terminal repeat (LTR) is largely dependent on intact NF-kB and SpI binding sites in the U3 region. In contrast, upstream LTR sequences allow efficient simian immunodeficiency virus (SIVmac) transcription in the absence of the core enhancer promoter region. In the present study, we investigated whether the regulation of HIV-2 Rod LTR activity is more reminiscent of HIV-1 having the same host or of SIVmac239 belonging to the same phylogenetic group. Viral promoter activity was studied in the context of the integrated provirus using both single cycle assays with pseudotyped luciferase reporter viruses and replication-competent HIV-2 LTR mutants. Our results demonstrate that intact SpI binding sites are important for both HIV-2 and SIVmac LTR activity in T cells and monocyte-derived macrophages. In contrast, deletion of the NF-kB binding site or of upstream regulatory sequences impaired HIV-2 Rod LTR activity but had little effect on SIVmac239 promoter function. Thus, similar to HIV-1, regulation of HIV-2 LTR promoter activity shows a low degree of functional redundancy possibly suggesting a specific adaptation to the human host.

Binding Sites↗

PubMed1986

Detection of an atypical rotavirus associated with diarrhoea in Chaco, Argentina.

Recently some unusual human and animal rotaviruses have been described which, although indistinguishable from standard rotaviruses by electron microscopy (EM), fail to react with antibody directed against the rotavirus group antigen. The genome of these viruses is composed of 11 double-stranded (ds) RNA segments; these RNA segments display different patterns when analysed by means of polyacrylamide gel electrophoresis (PAGE). The electrophoretic profile of the ds RNA segments of those human, antigenically different, viruses reported up to now appear to fit into two different patterns. Faecal samples from children with diarrhoea from different geographical areas of Argentina were evaluated for the presence of rotaviruses, using EM, ELISA and PAGE. During this survey a rotavirus-like agent was detected in the faeces of a child with diarrhoea from Chaco province, North-East Argentine. The pattern of the ds RNA segment of this virus on PAGE analysis appeared to be related to one of the atypical rotaviruses. This is the first description of such a virus strain in Argentina.

Antigens, Viral↗

PubMed2004

[Construction of human phage-displayed scFv library and selection of the ScFv against rabies virus].

AIM: To construct a human phage displayed single chain Fv antibody (scFv) library and screen specific scFv against rabies virus. METHODS: Using a phage-display technique, IgVH and IgVL genes were cloned from peripheral blood lymphocytes from three donors immunized with the WISTAR PM strain vaccine. Genes encoding scFv fragments were constructed by random linkage of VH gene and VL gene by SOE PCR, and then the constructed scFv genes were cloned into phagemid vector and transfected into E. coli TG1. The recombinant phages were screened by four rounds of panning with rabies virus vaccine. The positive recombinant phages were sequenced and antigen-binding activity of recombinant scFv was detected by competition ELISA. RESULTS: A human phage-displayed scFv library with about 7 x 10(8) sink size was constructed. A new human scFv was selected, which bound specifically to rabies virus and had higher affinity. CONCLUSION: The results lay a solid foundation for preparation of human engineering antibody to rabies virus reported herein with higher affinity.

Amino Acid Sequence↗

PubMed1999

[Detection rate of TTV-DNA in healthy medical workers].

We examined the positive rate in healthy medical workers about TT virus (TTV) which was a new hepatitis virus reported in 1997. The healthy medical workers showed a positive rate of 24%. Because there was no significant difference of the positive rate between the medical workers and general healthy persons we could not conclude that the positive rate was influenced by their profession. Generally it is known that a positive rate changes according to PCR primers used for the measurement. As for TTV as well, it is reported that a positive rate is greatly dependent on the selected region of primer. Therefore, we must pay attention to the evaluation of the positive rate for each assay system, especially using different primers.

Adult↗

PubMed1996

Experience of European collaboration in influenza surveillance in the winter of 1993-1994.

BACKGROUND: The spread of influenza is of major public health concern. Surveillance programmes exist in many countries. We here describe an international collaboration sharing information from Belgium, France, The Netherlands, Portugal and the United Kingdom during the influenza A(H3N2) epidemic occurring in Europe at the end of 1993. METHODS: Procedures for clinical and virological surveillance in each country are described. National data were assembled at one point in each country and entered on a computerized data base in Paris for integration. A background rate of influenza was identified from the two previous winters (1991-1992 and 1992-1993) using the measurement of incidence routinely used in each country. The epidemic in late 1993 was described in relation to the background rate. The proportion of cases in each of three age groups (0-14, 15-44 and 65 years and over) was compared in different stages of the epidemic. RESULTS: All national systems effectively identified the onset and monitored the course of the epidemic. Standardization of incidence data gave only limited assistance to the interpretation of the data. The distribution of cases by age group was similar in the early and late periods of the epidemic in all countries. Increased clinical incidence was matched by a simultaneous increase in positive virus identifications in all countries except England and Wales, where the interval between the week of peak clinical incidence and of peak virus reports is mainly due to a reporting procedure based on date of report received at the Central Public Health Laboratory rather than date of specimen capture. CONCLUSION: An international early warning system for monitoring influenza has been established.

Adolescent↗

PubMed1990

Characteristically distinct isolates of the nuclear polyhedrosis virus from Spodoptera litura.

More than 100 isolates were plaque-purified to examine the genetic variations in four wild stocks of Spodoptera litura nuclear polyhedrosis virus (NPV) collected in Japan. These isolates were characterized by their in vitro host range in three established insect cell lines, growth characteristics, polyhedral protein, DNA restriction endonuclease pattern and DNA hybridization. The isolates were separated into four distinct groups: (I) isolates corresponding to Autographa californica NPV, (II and IV) two different groups of isolates of S. littoralis NPV which had been previously characterized and (III) isolates with no correspondence to any reported virus group. Of the S. litura NPV wild stocks, two were mixtures of more than two different groups of NPVs. We have discussed the advantage of having a mixture of different NPV groups in the same wild virus stocks.

Animals↗

PubMed2004

Influenza A viruses of migrating wild aquatic birds in North America.

Surveillance of North America's wild ducks and shorebirds for 26 and 16 years, respectively, revealed differences in the prevalence of orthomyxoviruses between these hosts. Shorebirds had a high frequency of influenza A virus isolation during their northern migration, while wild ducks had high virus isolation frequencies during their southern migration. Some subtypes of influenza occurred regularly in both hosts with a 2-year periodicity, whereas others rarely occurred. Hemagglutinin subtypes H1 through H12 occurred in both hosts; H13 occurred only in shorebirds; and H14, H15, and influenza B and C never were detected. Shorebirds manifested a broader range of subtypes suggesting that shorebirds are the leading source of some viruses (such as H5) which are isolated less frequently from wild ducks. The viruses reported in this study are available for genomic study to determine whether prediction of host range or pandemic potential is possible.

Animal Migration↗

PubMed2006

Virus-bacterium interactions in water and sediment of West African inland aquatic systems.

The ecology of virioplankton in tropical aquatic ecosystems is poorly documented, and in particular, there are no references concerning African continental waters in the literature. In this study, we examined virus-bacterium interactions in the pelagic and benthic zones of seven contrasting shallow inland waters in Senegal, including one hypersaline lake. SYBR Gold-stained samples revealed that in the surface layers of the sites, the numbers of viruses were in the same range as the numbers of viruses reported previously for productive temperate systems. Despite high bacterial production rates, the percentages of visibly infected cells (as determined by transmission electron microscopy) were similar to the lowest percentages (range, 0.3 to 1.1%; mean, 0.5%) found previously at pelagic freshwater or marine sites, presumably because of the local environmental and climatic conditions. Since the percentages of lysogenic bacteria were consistently less than 8% for pelagic and benthic samples, lysogeny did not appear to be a dominant strategy for virus propagation at these sites. In the benthic samples, viruses were highly concentrated, but paradoxically, no bacteria were visibly infected. This suggests that sediment provides good conditions for virus preservation but ironically is an unfavorable environment for proliferation. In addition, given the comparable size distributions of viruses in the water and sediment samples, our results support the paradigm that aquatic viruses are ubiquitous and may have moved between the two compartments of the shallow systems examined. Overall, this study provides additional information about the relevance of viruses in tropical areas and indicates that the intensity of virus-bacterium interactions in benthic habitats may lower than the intensity in the adjacent bodies of water.

Bacteria↗