The Experts below are selected from a list of 135 Experts worldwide ranked by ideXlab platform
Julia K Hilliard - One of the best experts on this subject based on the ideXlab platform.
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<B>HerpesB> B Virus, Macacine <B>HerpesB>Virus 1, Breaks Simplex Virus Tradition via Major HistocompatiBility Complex Class I Expression in Cells from Human and Macaque Hosts
Journal of virology, 2012Co-Authors: Mugdha Vasireddi, Julia K HilliardAbstract:ABSTRACT B Virus of the family <B>HerpesB>viridae is endemic to rhesus macaques But results in 80% fatality in untreated humans who are zoonotically infected. Downregulation of major histocompatiBility complex (MHC) class I in order to evade CD8 + T-cell activation is characteristic of most <B>HerpesB>Viruses. Here we examined the cell surface presence and total protein expression of MHC class I molecules in B Virus-infected human foreskin fiBroBlast cells and macaque kidney epithelial cells in culture, which are representative of foreign and natural host initial target cells of B Virus. Our results show
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production of <B>HerpesB> B Virus recomBinant glycoproteins and evaluation of their diagnostic potential
Journal of Clinical Microbiology, 2005Co-Authors: Ludmila Perelygina, Irina Patrusheva, Holley Zurkuhlen, Soumya Hombaiah, Martin J Wildes, Nikolai Patrushev, Julia K HilliardAbstract:B Virus (cercopithecine <B>HerpesB>Virus 1) is the only deadly alpha<B>HerpesB>Virus that is zoonotically transmissiBle from macaques to humans. The detection of humoral immune responses is the method of choice for the rapid identification of B Virus-infected animals. We evaluated the diagnostic potential of recomBinant B Virus glycoproteins for the detection of immunogloBulin G (IgG) antiBodies in monkey and human sera. Glycoproteins B, C, and E and secreted (sgG) and memBrane-associated (mgG) segments of glycoprotein G (gG) were expressed in the BaculoVirus expression system, while gD was expressed in CHO cells. We developed recomBinant protein-Based IgG enzyme-linked immunosorBent assays (ELISAs) and compared their diagnostic efficacies By using B Virus antiBody-negative (n = 40) and -positive (n = 75) macaque sera identified By a whole antigen-Based ELISA and Western Blotting. The diagnostic sensitivities of the gB-, gC-, gD-, and mgG-ELISAs were 100, 97.3, 88.0, and 80.0%, respectively. The specificities of the gB-, gC-, and gD-ELISAs and of the mgG-ELISA were 100 and 97.5%, respectively. In contrast, the sensitivities and specificities of sgG- and gE-ELISAs were low, suggesting that sgG and gE are less effective diagnostic antigens. Sera from nonmacaque monkeys cross-reacted with gB, gC, and gD, and only BaBoon sera reacted weakly with mgG. Human <B>HerpesB> simplex Virus type 1 (HSV-1)- and HSV-2-positive sera pools reacted with gB and gD, whereas sera from B Virus-infected individuals reacted with all four antigens. These data indicate that gB, gC, gD, and mgG have a high diagnostic potential for B Virus serodiagnosis in macaques, whereas mgG may Be a valuaBle antigen for discrimination Between antiBodies induced By B Virus and those induced By other, closely related alpha<B>HerpesB>Viruses, including HSV-1 and -2.
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complete sequence and comparative analysis of the genome of <B>HerpesB> B Virus cercopithecine <B>HerpesB>Virus 1 from a rhesus monkey
Journal of Virology, 2003Co-Authors: Ludmila Perelygina, Holley Zurkuhlen, Li Zhu, Ryan Mills, Mark Borodovsky, Julia K HilliardAbstract:The complete DNA sequence of <B>HerpesB> B Virus (Cercopithecine <B>HerpesB>Virus 1) strain E2490, isolated from a rhesus macaque, was determined. The total genome length is 156,789 Bp, with 74.5% G+C composition and overall genome organization characteristic of alpha<B>HerpesB>Viruses. The first and last residues of the genome were defined By sequencing the cloned genomic termini. There were six origins of DNA replication in the genome due to tandem duplication of Both oriL and oriS regions. Seventy-four genes were identified, and sequence homology to proteins known in <B>HerpesB> simplex Viruses (HSVs) was oBserved in all cases But one. The degree of amino acid identity Between B Virus and HSV proteins ranged from 26.6% (US5) to 87.7% (US15). Unexpectedly, B Virus lacked a homolog of the HSV gamma(1)34.5 gene, which encodes a neurovirulence factor. ABsence of this gene was verified in two low-passage clinical isolates derived from a rhesus macaque and a zoonotically infected human. This finding suggests that B Virus most likely utilizes mechanisms distinct from those of HSV to sustain efficient replication in neuronal cells. Despite the consideraBle differences in G+C content of the macaque and B Virus genes (51% and 74.2%, respectively), codons used By B Virus are optimal for the tRNA population of macaque cells. Complete sequence of the B Virus genome will certainly facilitate identification of the genetic Basis and possiBle molecular mechanisms of enhanced B Virus neurovirulence in humans, which results in an 80% mortality rate following zoonotic infection.
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Identification of a <B>HerpesB> B Virus-specific glycoprotein d immunodominant epitope recognized By natural and foreign hosts.
The Journal of infectious diseases, 2002Co-Authors: Ludmila Perelygina, Irina Patrusheva, Holley Zurkuhlen, Julia K HilliardAbstract:The mapping of linear epitopes of B Virus (Cercopithecine <B>HerpesB>Virus 1 and <B>HerpesB> B Virus) glycoprotein D (gD) was accomplished By screening the constructed gD epitope liBrary with serum from B Virus–infected macaques. The immunodominant epitope, gD (362–370), was identified within the C-terminal region of B Virus gD that was highly conserved among 19 B Virus clinical isolates But was not present in either <B>HerpesB> simplex Virus (HSV)–1 or HSV-2 gD. A suBstantial percentage of serum samples from macaques (95%) and humans (80%) infected with B Virus contained antiBodies to this epitope. AntiBodies against HSV types 1 or 2 did not react with this epitope; thus, gD (362–370) has unique potential to detect B Virus–specific antiBody responses in human serum, even in the presence of antiBodies to HSV1 and HSV-2. B Virus (Cercopithecine <B>HerpesB>Virus 1 and <B>HerpesB> B Virus) can cause a deadly zoonosis when transmitted to humans By a macaque actively shedding Virus. A memBer of the genus SimplexVirus from the suBfamily Alpha<B>HerpesB>virinae, B Virus estaBlishes a life-long latency in the sensory ganglia of infected monkeys [1, 2]. Since the B Virus shedding By seropositive macaques is sporadic [3–5], identification of infected animals is usually accomplished By detection of B Virus–specific antiBodies [6]. Because of the extensive serologic cross-reactivity Between the closely related <B>HerpesB> simplex Virus (HSV) types 1 and 2 and B Virus antigens, detection of B Virus infections in humans with preexisting HSV-1/HSV-2 antiBodies is technically challenging [7, 8]. B Virus serodiagnosis currently relies on competition ELISA and immunoBlotting strategies [9–11], But Both techniques require complex analysis and B Virus propagation in a Biosafety level–4 laBoratory. Initiation of antiviral therapies as early as possiBle after infection in humans is vital in preventing fatal disease; thus, there is a clinical need for early diagnostic capaBilities [12–15]. A highly specific diagnostic method for early detection of B
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AntiBody cross-reactivity of alpha<B>HerpesB>Viruses as mirrored in naturally infected primates
Archives of virology, 2002Co-Authors: David Katz, Wei Shi, Peter W. Krug, R.d. Henkel, H. Mcclure, Julia K HilliardAbstract:The extent of antiBody cross-reactivity of pooled antisera from rhesus monkeys, BaBoons, African green monkeys, langurs, sooty mangaBeys and humans to 6 alpha<B>HerpesB>Viruses (<B>HerpesB> B Virus, <B>HerpesB> papio 2, simian agent 8, langur <B>HerpesB> Virus and <B>HerpesB> simplex 1 & 2) was examined By two types of enzyme linked immunosorBent assays, an antiBody capture assay (tELISA) and an antigen capture assay (dELISA). Percent cross-reactivity was calculated for each serum By comparison of the homologous reaction (100%) to the reaction with hetero-logous Viruses. Comparison of the immunological reactivity of the mangaBey antiserum pool to the panel of alpha<B>HerpesB>Viruses indicated that these antiBodies were induced By a yet unidentified alpha<B>HerpesB>Virus. In general, monkey sera were more cross-reactive to monkey <B>HerpesB>Viruses than to human <B>HerpesB>Viruses.
Anthony Griffiths - One of the best experts on this subject based on the ideXlab platform.
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Regulation of viral gene expression By the <B>HerpesB> simplex Virus 1UL24 protein (HSV-1UL24 inhiBits accumulation of viral transcripts).
Virology, 2016Co-Authors: Carolina Sanabria-solano, Anthony Griffiths, Carmen Elena Gonzalez, Nicolas Richerioux, Luc Bertrand, Slimane Dridi, Yves Langelier, Angela PearsonAbstract:UL24 is conserved among all <B>HerpesB>viridae. In <B>HerpesB> simplex Virus 1 (HSV-1), UL24 mutations lead to reduced viral titers Both in cell culture and in vivo, and reduced pathogenicity. The human cytomegaloVirus ortholog of UL24 has a gene regulatory function; however, it is not known whether other UL24 orthologs also affect gene expression. We discovered that in co-transfection experiments, expression of UL24 correlated with a reduction in the expression of several viral proteins and transcripts. SuBstitution mutations targeting conserved residues in UL24 impaired this function. Reduced transcript levels did not appear attriButaBle to changes in mRNA staBility. The UL24 ortholog of <B>HerpesB> B Virus exhiBited a similar activity. An HSV-1 mutant that does not express UL24 produced more viral R1 and R2 transcripts than the wild type or rescue Virus relative to the amount of viral DNA. These results reveal a new role for HSV-1UL24 in regulating viral mRNA accumulation.
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<B>HerpesB> B Virus Utilizes Human Nectin-1 But Not HVEM or PILRα for Cell-Cell Fusion and Virus Entry
Journal of virology, 2012Co-Authors: Qing Fan, Mallory E. Harden, Anthony Griffiths, Melanie Ann Amen, Alberto Severini, Richard LongneckerAbstract:To investigate the requirements of <B>HerpesB>Virus entry and fusion, the four homologous glycoproteins necessary for <B>HerpesB> simplex Virus (HSV) fusion were cloned from <B>HerpesB> B Virus (BV) (or macacine <B>HerpesB>Virus 1, previously known as cercopithecine <B>HerpesB>Virus 1) and cercopithecine <B>HerpesB>Virus 2 (CeHV-2), Both related simian simplexViruses Belonging to the alpha<B>HerpesB>Virus suBfamily. Western Blots and cell-Based enzyme-linked immunosorBent assay (ELISA) showed that glycoproteins gB, gD, and gH/gL were expressed in whole-cell lysates and on the cell surface. Cell-cell fusion assays indicated that nectin-1, an HSV-1 gD receptor, mediated fusion of cells expressing glycoproteins from Both BV and CeHV-2. However, <B>HerpesB>Virus entry mediator (HVEM), another HSV-1 gD receptor, did not facilitate BV- and CeHV-2-induced cell-cell fusion. Paired immunogloBulin-like type 2 receptor alpha (PILRα), an HSV-1 gB fusion receptor, did not mediate fusion of cells expressing glycoproteins from either simian Virus. Productive infection with BV was possiBle only with nectin-1-expressing cells, indicating that nectin-1 mediated entry while HVEM and PILRα did not function as entry receptors. These results indicate that these alpha<B>HerpesB>Viruses have differing preferences for entry receptors. The usage of the HSV-1 gD receptor nectin-1 may explain interspecies transfer of the Viruses, and altered receptor usage may result in altered virulence, tropism, or pathogenesis in the new host. A heterotypic cell fusion assay resulting in productive fusion may provide insight into interactions that occur to trigger fusion. These findings may Be of therapeutic significance for control of deadly BV infections.
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identification and expression analysis of <B>HerpesB> B Virus encoded small rnas
Journal of Virology, 2011Co-Authors: Anthony Griffiths, Melanie Ann AmenAbstract:<B>HerpesB> B Virus (BV) naturally infects macaque monkeys and is genetically similar to <B>HerpesB> simplex Virus (HSV). Zoonotic infection of humans can cause encephalitis and if untreated has a fatality rate of ∼80%. The frequent use of macaques in Biomedical research emphasizes the need to understand the molecular Basis of BV pathogenesis with a view toward improving safety for those working with macaques. MicroRNAs (miRNAs) are small noncoding RNAs that regulate the expression of mRNAs Bearing complementary target sequences and are employed By Viruses to control viral and host gene expression. Using deep sequencing and validation By expression in transfected cells, we identified 12 novel BV-encoded miRNAs expressed in lytically infected cells and 4 in latently infected trigeminal ganglia (TG). Using quantitative reverse transcription-PCR (RT-qPCR), we found that most of the miRNAs exhiBited a high level of aBundance throughout infection. Further analyses showed that some miRNAs could Be generated from multiple transcripts with different kinetic classes, possiBly explaining detection throughout infection. Interestingly, miRNAs were detected at early times in the aBsence of viral gene expression and were present in purified virions. In TG, despite similar amounts of viral DNA per ganglion, it was notaBle that the relative amount of each miRNA varied Between ganglia. The majority of the miRNAs are encoded By the regions that exhiBit the most sequence differences Between BV and HSV. Additionally, there is no sequence conservation Between BV- and HSV-encoded miRNAs, which may Be important for the differences in the human diseases caused By BV and HSV.
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discovery of <B>HerpesB> B Virus encoded micrornas
Journal of Virology, 2009Co-Authors: Michael I Besecker, Mallory E. Harden, Xiujie Wang, Anthony GriffithsAbstract:<B>HerpesB> B Virus (BV) naturally infects macaque monkeys and is a close relative of <B>HerpesB> simplex Virus. BV can zoonotically infect humans to cause a rapidly ascending encephalitis with ∼80% mortality. Therefore, BV is a serious danger to those who come into contact with these monkeys or their tissues and cells. MicroRNAs are regulators of gene expression, and there have Been reports of Virus-encoded microRNAs. We hypothesize that BV-encoded microRNAs are important for the regulation of viral and cellular genes. Herein, we report the discovery of three <B>HerpesB> B Virus-encoded microRNAs.
Hans-joachim Fritz - One of the best experts on this subject based on the ideXlab platform.
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serological analysis of <B>HerpesB> B Virus at individual epitope resolution from two dimensional peptide arrays to multiplex Bead flow assays
Analytical Chemistry, 2019Co-Authors: Sven-kevin Hotop, Ahmed Abd El Wahed, Ulrike Beutling, Ronald Frank, Florian Czerny, Claudia Sievers, Ulf Diederichsen, Christiane Stahlhennig, Mark Bronstrup, Hans-joachim FritzAbstract:Macacine <B>HerpesB>Virus or B Virus (BV) is a zoonotic agent that leads to high mortality rates in humans if transmitted and untreated. Here, BV is used as a test case to estaBlish a two-step procedure for developing high throughput serological assays Based on synthetic peptides. In step 1, peptide microarray analysis of 42 monkey sera (30 of them tested BV positive By ELISA) revealed 1148 responses against 369 different peptides. The latter could Be grouped into 142 different antiBody target regions (ATRs) in six different glycoproteins (gB, gC, gD, gG, gH, and gL) of BV. The high numBer of newly detected ATRs was made possiBle inter alia By a new preanalytical protocol that reduced unspecific Binding of serum components to the cellulose-Based matrix of the microarray. In step 2, soluBle peptides corresponding to eight ATRs of particularly high antigenicity were synthesized and coupled to fluorescently laBeled Beads, which were suBsequently employed in immunochemical Bead flow assays. Their outcome mirrored ...
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Multiple AntiBody Targets on <B>HerpesB> B Glycoproteins B and D Identified By Screening Sera of Infected Rhesus Macaques with Peptide Microarrays
2016Co-Authors: Sven-kevin Hotop, Ulrike Beutling, Dieter Jentsch, Dirk Motzkus, Ronald Frank, Gerhard Hunsmann, Christiane Stahl-hennig, Ahmed Abd, El Wahed, Hans-joachim FritzAbstract:<B>HerpesB> B Virus (or <B>HerpesB>Virus simiae or Macacine <B>HerpesB>Virus 1) is endemic in many populations of macaques, Both in the wild and in captivity. The Virus elicits only mild clinical symptoms (if any) in monkeys, But can Be transmitted By various routes, most commonly via Bites, to humans where it causes viral encephalitis with a high mortality rate. Hence, <B>HerpesB> B constitutes a consideraBle occupational hazard for animal caretakers, veterinarians and laBoratory personnel. Efforts are therefore Being made to reduce the risk of zoonotic infection and to improve prognosis after accidental exposure. Among the measures envisaged are serological surveillance of monkey colonies and specific diagnosis of <B>HerpesB> B zoonosis against a Background of antiBodies recognizing the closely related human <B>HerpesB> simplex Virus (HSV). 422 pentadecapeptides covering, in an overlapping fashion, the entire amino acid sequences of <B>HerpesB> B proteins gB and gD were synthesized and immoBilized on glass slides. AntiBodies present in monkey sera that Bind to suBsets of the peptide collection were detected By microserological techniques. With 42 different rhesus macaque sera, 114 individual responses to 18 different antiBody target regions (ATRs) were recorded, 17 of which had not Been descriBed earlier. This finding may pave the way for
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Multiple antiBody targets on <B>HerpesB> B glycoproteins B and D identified By screening sera of infected rhesus macaques with peptide microarrays.
PloS one, 2014Co-Authors: Sven-kevin Hotop, Ahmed Abd El Wahed, Ulrike Beutling, Dieter Jentsch, Dirk Motzkus, Ronald Frank, Gerhard Hunsmann, Christiane Stahl-hennig, Hans-joachim FritzAbstract:<B>HerpesB> B Virus (or <B>HerpesB>Virus simiae or Macacine <B>HerpesB>Virus 1) is endemic in many populations of macaques, Both in the wild and in captivity. The Virus elicits only mild clinical symptoms (if any) in monkeys, But can Be transmitted By various routes, most commonly via Bites, to humans where it causes viral encephalitis with a high mortality rate. Hence, <B>HerpesB> B constitutes a consideraBle occupational hazard for animal caretakers, veterinarians and laBoratory personnel. Efforts are therefore Being made to reduce the risk of zoonotic infection and to improve prognosis after accidental exposure. Among the measures envisaged are serological surveillance of monkey colonies and specific diagnosis of <B>HerpesB> B zoonosis against a Background of antiBodies recognizing the closely related human <B>HerpesB> simplex Virus (HSV). 422 pentadecapeptides covering, in an overlapping fashion, the entire amino acid sequences of <B>HerpesB> B proteins gB and gD were synthesized and immoBilized on glass slides. AntiBodies present in monkey sera that Bind to suBsets of the peptide collection were detected By microserological techniques. With 42 different rhesus macaque sera, 114 individual responses to 18 different antiBody target regions (ATRs) were recorded, 17 of which had not Been descriBed earlier. This finding may pave the way for a peptide-Based, <B>HerpesB> B specific serological diagnostic test.
Sven-kevin Hotop - One of the best experts on this subject based on the ideXlab platform.
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serological analysis of <B>HerpesB> B Virus at individual epitope resolution from two dimensional peptide arrays to multiplex Bead flow assays
Analytical Chemistry, 2019Co-Authors: Sven-kevin Hotop, Ahmed Abd El Wahed, Ulrike Beutling, Ronald Frank, Florian Czerny, Claudia Sievers, Ulf Diederichsen, Christiane Stahlhennig, Mark Bronstrup, Hans-joachim FritzAbstract:Macacine <B>HerpesB>Virus or B Virus (BV) is a zoonotic agent that leads to high mortality rates in humans if transmitted and untreated. Here, BV is used as a test case to estaBlish a two-step procedure for developing high throughput serological assays Based on synthetic peptides. In step 1, peptide microarray analysis of 42 monkey sera (30 of them tested BV positive By ELISA) revealed 1148 responses against 369 different peptides. The latter could Be grouped into 142 different antiBody target regions (ATRs) in six different glycoproteins (gB, gC, gD, gG, gH, and gL) of BV. The high numBer of newly detected ATRs was made possiBle inter alia By a new preanalytical protocol that reduced unspecific Binding of serum components to the cellulose-Based matrix of the microarray. In step 2, soluBle peptides corresponding to eight ATRs of particularly high antigenicity were synthesized and coupled to fluorescently laBeled Beads, which were suBsequently employed in immunochemical Bead flow assays. Their outcome mirrored ...
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Multiple AntiBody Targets on <B>HerpesB> B Glycoproteins B and D Identified By Screening Sera of Infected Rhesus Macaques with Peptide Microarrays
2016Co-Authors: Sven-kevin Hotop, Ulrike Beutling, Dieter Jentsch, Dirk Motzkus, Ronald Frank, Gerhard Hunsmann, Christiane Stahl-hennig, Ahmed Abd, El Wahed, Hans-joachim FritzAbstract:<B>HerpesB> B Virus (or <B>HerpesB>Virus simiae or Macacine <B>HerpesB>Virus 1) is endemic in many populations of macaques, Both in the wild and in captivity. The Virus elicits only mild clinical symptoms (if any) in monkeys, But can Be transmitted By various routes, most commonly via Bites, to humans where it causes viral encephalitis with a high mortality rate. Hence, <B>HerpesB> B constitutes a consideraBle occupational hazard for animal caretakers, veterinarians and laBoratory personnel. Efforts are therefore Being made to reduce the risk of zoonotic infection and to improve prognosis after accidental exposure. Among the measures envisaged are serological surveillance of monkey colonies and specific diagnosis of <B>HerpesB> B zoonosis against a Background of antiBodies recognizing the closely related human <B>HerpesB> simplex Virus (HSV). 422 pentadecapeptides covering, in an overlapping fashion, the entire amino acid sequences of <B>HerpesB> B proteins gB and gD were synthesized and immoBilized on glass slides. AntiBodies present in monkey sera that Bind to suBsets of the peptide collection were detected By microserological techniques. With 42 different rhesus macaque sera, 114 individual responses to 18 different antiBody target regions (ATRs) were recorded, 17 of which had not Been descriBed earlier. This finding may pave the way for
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Feinkartierung humoraler Immunantworten von Makaken nach Immunisierung und/oder viraler Infektion mittels Peptid - Microarray
2014Co-Authors: Sven-kevin HotopAbstract:The aim of this study was to map humoral immune responses in rhesus macaques (RM) which had either Been immunized and/or infected with different Viruses. This was achieved By screening serum samples using peptide microarrays. Further goals were the optimization of the analysis and the minimization of the required sample volumes. For the readout, an independent computer-assisted algorithm was formulated which provided an oBjective method to optimize peptide microarray production and data mining. In order to screen large numBers of serum samples, new tools were developed that allowed a safe handling of up to four samples per slide simultaneously. In polyclonal serum samples the length of closely related epitopes could not Be determined precisely due to the design of the array, i.e. tiled peptides with an offset of three amino acids. Thus, the more fitting term “antiBody target region” (ATR) was given to descriBe antiBodies that recognize such regions on the antigen. In using the peptide microarray technology, 27 distinct ATRs were identified on the envelope spike glycoprotein (gp) 160 precursor of the simian immunodeficiency Virus (SIV). The respective serum samples that were used for analysis had Been collected during different AIDS-macaque model studies. In one of the studies, performing multiple low dose viral exposures which however did not lead to a productive infection, one particular ATR was detected. This might Be involved in the protection through antiBodies against the constant region five of gp120. RemarkaBly, no qualitative differences in the antiBody reaction pattern against linear epitopes Between SIV-infected long-term survivors (LTS) and animals progressing to AIDS-like disease were oBserved. Furthermore, antiBodies directed against the N-terminal end of SIVgp120 were identified as a new early seromarker of SIV-infection. Irrespective of the inoculated SIV strain those antiBodies reacted exclusively with the amino acid sequence of SIVmac251. The sequencing of viral RNA in serial plasma samples of an animal infected with SIVmac239 demonstrated the lack of amino acid exchanges in the region of interest. The predicted structure of this epitope Based on a known 3D-structure of the human immunodeficiency Virus type 1 (HIV-1) revealed a sharp Bend at the original Binding site. The aBove mentioned data imply that this identified ATR might represent a conformation-dependent epitope, which has not Been seen with previous methods. Analysis of sera from <B>HerpesB> B Virus (BV) infected monkeys with a peptide microarray containing peptides covering the amino acid sequences of the two glycoproteins B and D (gB and gD) revealed 17 new ATRs and confirmed one previously descriBed epitope. Interestingly, antiBodies against one ATR on the BV gD were detected in an animal, which was considered uninfected due to negative testing through a cross-reactive <B>HerpesB> simplex Virus type 1 (HSV-1)-ELISA. AntiBodies against this ATR could Block the main entry receptor Nectin-1 on human cells. Since BV poses an occupational hazard for animal keepers and others dealing with macaques this result could pave the way for the development of a peptide-Based vaccination strategy. Another detected ATR in the crown domain of gB seems to Be conformation-dependent and was further characterized By adopting the 3D-structure of HSV-1 and By alanine scan. The highly immunogenic endodomain of gD revealed at least two distinguishaBle epitopes. In this case, the previously descriBed C-terminal epitope was mapped to the last two amino acids. Through the estaBlishment of a peptide-Based ELISA the BV-peptide microarray data were confirmed. This ELISA was used to prove the serological relevance of the C-terminal end of gD in BV infection of RM. The longitudinal analysis of RM from a Breeding group suggested that BV is more likely transmitted via sexual contacts rather than Bites or scratches. The lack of the repeated rise in antiBody levels of animals kept under experimental conditions may indicate that recurrence is a rare event during the infection with BV. The results oBtained in this thesis demonstrate that peptide microarrays are useful and powerful tools for initial screening of serological markers. When standardized synthetized peptides are used the aBove mentioned results allow the estaBlishment of low cost serological tests. To date, the screening of conformation-dependent epitopes is done through random peptide liBraries But this technique is prone to errors. The results may lead to the development of a new generation of Biochips, which allow inexpensive screening for conformational epitopes. However, a detailed assessment of the estaBlished method has indicated some limitations: In case of antiBodies Binding to “open” end structures, such as the gD C-terminal end, linked- or conjugated peptides may fail to detect such antiBodies. A validation of antiBody Binding to unlinked C-terminal end containing peptides is therefore essential.
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Multiple antiBody targets on <B>HerpesB> B glycoproteins B and D identified By screening sera of infected rhesus macaques with peptide microarrays.
PloS one, 2014Co-Authors: Sven-kevin Hotop, Ahmed Abd El Wahed, Ulrike Beutling, Dieter Jentsch, Dirk Motzkus, Ronald Frank, Gerhard Hunsmann, Christiane Stahl-hennig, Hans-joachim FritzAbstract:<B>HerpesB> B Virus (or <B>HerpesB>Virus simiae or Macacine <B>HerpesB>Virus 1) is endemic in many populations of macaques, Both in the wild and in captivity. The Virus elicits only mild clinical symptoms (if any) in monkeys, But can Be transmitted By various routes, most commonly via Bites, to humans where it causes viral encephalitis with a high mortality rate. Hence, <B>HerpesB> B constitutes a consideraBle occupational hazard for animal caretakers, veterinarians and laBoratory personnel. Efforts are therefore Being made to reduce the risk of zoonotic infection and to improve prognosis after accidental exposure. Among the measures envisaged are serological surveillance of monkey colonies and specific diagnosis of <B>HerpesB> B zoonosis against a Background of antiBodies recognizing the closely related human <B>HerpesB> simplex Virus (HSV). 422 pentadecapeptides covering, in an overlapping fashion, the entire amino acid sequences of <B>HerpesB> B proteins gB and gD were synthesized and immoBilized on glass slides. AntiBodies present in monkey sera that Bind to suBsets of the peptide collection were detected By microserological techniques. With 42 different rhesus macaque sera, 114 individual responses to 18 different antiBody target regions (ATRs) were recorded, 17 of which had not Been descriBed earlier. This finding may pave the way for a peptide-Based, <B>HerpesB> B specific serological diagnostic test.
Richard Longnecker - One of the best experts on this subject based on the ideXlab platform.
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Is nectin-1 the "master" receptor for deadly <B>HerpesB> B Virus infection?
Virulence, 2012Co-Authors: Qing Fan, Richard LongneckerAbstract:The family <B>HerpesB>viridae is a large, diverse family of douBle-stranded enveloped DNA Viruses. <B>HerpesB> B Virus (BV) and Cercopithecine <B>HerpesB>Virus 2 (CeHV-2) are simian <B>HerpesB>Viruses. Like <B>HerpesB> simplex Virus 1 (HSV-1), they Belong to the α-<B>HerpesB>Virus suBfamily. Interest in BV infections results from the oBservation that zoonotic infections often result in death of humans, whereas infection of macaques, the natural host of BV, results in disease similar to that oBserved on HSV infections of humans. We recently reported in cell-cell fusion assays in which nectin-1, a HSV-1 gD receptor, mediated fusion of cells expressing glycoproteins from Both BV and CeHV-2. However, HVEM, another HSV-1 gD receptor, did not mediate fusion By BV and CeHV-2 glycoproteins. Paired immunogloBulin-like type 2 receptor α (PILRα), an HSV-1 gB fusion receptor, did not mediate fusion with BV or CeHV-2 glycoproteins. These results were further confirmed By BV infection. Our results may indicate that differential receptor usage By BV in humans when compared with macaques may have pathological consequences. Understanding human and simian receptor usage for BV and HSV may provide clues to understand the pathogenesis of these Viruses, as well as related Viruses, in their natural host as well zoonotic infections. This Broader understanding may result in the development of novel therapeutics to control deadly BV infections as well as <B>HerpesB>Virus infections in general. <B>HerpesB> B Virus (BV, officially named as Macacine <B>HerpesB>Virus 1, formerly <B>HerpesB>Virus simiae, monkey B Virus, or Cercopithecine <B>HerpesB>Virus 1) and Cercopithecine <B>HerpesB>Virus 2 (CeHV-2, formerly simian agent 8) are primate <B>HerpesB>Viruses Belonging to the α-<B>HerpesB>Virus suBfamily and are closely related to <B>HerpesB> simplex Virus 1 (HSV-1) and HSV-2. HSV causes recurrent mucocutaneous lesions on the mouth, face, or genitalia and in rare cases can cause meningitis or encephalitis. BV naturally infects macaques whereas zoonotic infections of foreign hosts, such as humans, can result in encephalitis, encephalomyelitis and death. BV infection has high mortality in humans (greater than 50% in documented infections) and as such is recognized as a deadly Virus for humans requiring Biosafety level 4. Similarly to the high level of pathogenicity of BV in humans, HSV infection of marmosets can also Be fatal. CeHV-2 is a pathogen of BaBoons and is not known to cause disease in primates outside the natural hosts. Since the receptors for HSV are well descriBed, we chose to explore the receptor usage of the BV and CeHV-2. HSV entry into target cells requires a minimum four Virus encoded glycoproteins—glycoproteins B (gB), D (gD), H (gH) and L (gL). These glycoproteins interact with a variety of cellular receptors to facilitate Virus entry. Best descriBed are the cellular receptors for gB and gD which we chose to study. The interaction of gD with the gD receptors has Been extensively investigated. The Binding of gD to <B>HerpesB>Virus entry mediator (HVEM) (Montgomery et al., Cell 1996), nectin-1 (Geraghty et al., Science 1998), nectin-2 (Warner et al., Virology 1998) and modified heparan sulfate (Shukla et al., Cell 1999) triggers fusion of the virion envelope with a cellular memBrane. Paired immunogloBulin-like type 2 receptor α (PILRα) is a receptor that Binds to gB and also mediates entry and fusion (Satoh et al., Cell 2008) even though it shows much less fusion activity than the gD receptors nectin-1 and HVEM. Two other gB receptors, with less known in regard to the significance in Virus infection, are myelin-associated glycoprotein (MAG) (Suenaga et al., Proc Natl Acad Sci U S A 2010), and non-muscle myosin heavy chain IIA (NMHC-IIA) (Arii et al., Nature 2010). HVEM is a memBer of the tumor necrosis factor receptor family. Nectin-1 and nectin-2 are cell adhesion molecules in the immunogloBulin superfamily and are widely expressed By a variety of cell types, including epithelial cells and neurons. Specific sites in heparan sulfate generated By certain 3-O-sulfotransferases (3-O-S HS) can also serve as a gD-Binding entry receptor. PILRα is expressed on cells of the immune system. MAG is a cell-surface molecule Belonging to immunogloBulin superfamily, is usually expressed in neural tissues. NMHC-IIA not only functions in the cytoplasm But also functions on the cell surface with gB upon viral entry. NMHC-IIA is expressed in a Broad range of cell lines, tissues and cell types in vivo. To Begin our studies, we generated expression constructs for the homologous glycoproteins (gB, gD, gH and gL) necessary for HSV-1 fusion from BV and CeHV-2. We then performed cell-cell fusion assays to determine which cellular and viral proteins were required for cell-cell fusion. The results indicated that nectin-1 is the primary receptor that mediates fusion for BV and CeHV-2, and that HVEM and PILRα do not function for BV and CeHV-2. To verify our cell-cell fusion results, we used cells expressing human nectin-1, HVEM or PILRα and determined if they could Be infected with BV. The results confirmed that BV only utilizes nectin-1 and not HVEM or PILRα. To confirm that BV and CeHV-2 do not utilize human HVEM as an entry receptor Because of differences Between human and simian HVEM, we performed cell-cell fusion assays with simian HVEM. In these experiments, we found that simian HVEM could mediate fusion with cells expressing HSV-1 glycoproteins however; simian HVEM did not mediate fusion of cells expressing glycoproteins from BV and CeHV-2. Thus, as found in earlier studies and reports from other investigators, our findings are compatiBle with the idea that nectin-1 is a pan α-<B>HerpesB>Virus entry receptor. Of particular interest from our recent studies is the idea that differential receptor usage may Be important for pathogenesis in infected hosts. CompatiBle with this idea, studies in mice have shown that nectin-1 is the primary receptor responsiBle for the infection of the vaginal epithelium with HSV-2. Interestingly, in experimental vaginal infections, nectin-1 was not the sole receptor capaBle of enaBling spread of HSV infection from the vaginal epithelium to the PNS and CNS (Taylor et al., Cell Host MicroBe 2007). However, expression of nectin-1 is necessary for HSV-2 infection via the intracranial route and for encephalitis while HVEM was not important (Kopp et al., Proc Natl Acad Sci U S A 2009). In contrast, in HSV-1 infection of the murine eye, we found that Both HVEM and nectin-1 must Be present for maximal HSV-1 infection further suggesting that receptor requirements for HSV depends on the route of infection and/or serotype (KaraBa et al., J Virol 2011). The host immune response may also Be important in pathogenesis. Previous studies have shown that engagement of HVEM By gD alters the immune response following murine vaginal infection. In these studies, there was a transient increase in mucosal chemokine and IL-6 levels when compared with infection with wild-type control Virus in agreement with gD-HVEM interaction elicits an innate response (Yoon et al., PloS One 2011). Overall, our studies, as well as studies By others, indicate that choice of receptor utilized as well as the host may Be important determinants in regard to the outcome of the infection. Virus receptor usage, replication and control By the host immune are likely fine-tuned to allow maximal Virus replication without Being detrimental to the host to allow Virus spread to naive hosts. In contrast, when a Virus infects a non-natural host, this Balance may not exist resulting in severe pathogenesis as oBserved in BV infections of humans. By understanding the required interactions Between host receptors and the relevant glycoproteins required for infection, novel therapeutics might Be developed to treat zoonotic infections and prevent deadly infections or more Broadly used in natural HSV infections that routinely occur within the human population.
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<B>HerpesB> B Virus Utilizes Human Nectin-1 But Not HVEM or PILRα for Cell-Cell Fusion and Virus Entry
Journal of virology, 2012Co-Authors: Qing Fan, Mallory E. Harden, Anthony Griffiths, Melanie Ann Amen, Alberto Severini, Richard LongneckerAbstract:To investigate the requirements of <B>HerpesB>Virus entry and fusion, the four homologous glycoproteins necessary for <B>HerpesB> simplex Virus (HSV) fusion were cloned from <B>HerpesB> B Virus (BV) (or macacine <B>HerpesB>Virus 1, previously known as cercopithecine <B>HerpesB>Virus 1) and cercopithecine <B>HerpesB>Virus 2 (CeHV-2), Both related simian simplexViruses Belonging to the alpha<B>HerpesB>Virus suBfamily. Western Blots and cell-Based enzyme-linked immunosorBent assay (ELISA) showed that glycoproteins gB, gD, and gH/gL were expressed in whole-cell lysates and on the cell surface. Cell-cell fusion assays indicated that nectin-1, an HSV-1 gD receptor, mediated fusion of cells expressing glycoproteins from Both BV and CeHV-2. However, <B>HerpesB>Virus entry mediator (HVEM), another HSV-1 gD receptor, did not facilitate BV- and CeHV-2-induced cell-cell fusion. Paired immunogloBulin-like type 2 receptor alpha (PILRα), an HSV-1 gB fusion receptor, did not mediate fusion of cells expressing glycoproteins from either simian Virus. Productive infection with BV was possiBle only with nectin-1-expressing cells, indicating that nectin-1 mediated entry while HVEM and PILRα did not function as entry receptors. These results indicate that these alpha<B>HerpesB>Viruses have differing preferences for entry receptors. The usage of the HSV-1 gD receptor nectin-1 may explain interspecies transfer of the Viruses, and altered receptor usage may result in altered virulence, tropism, or pathogenesis in the new host. A heterotypic cell fusion assay resulting in productive fusion may provide insight into interactions that occur to trigger fusion. These findings may Be of therapeutic significance for control of deadly BV infections.