bluuebunny/crossref_metadata_2025_split huggingface dataset:
Adaptation of Sindbis Virus to BHK Cells Selects for Use of Heparan Sulfate as an Attachment Receptor
Authors: Klimstra William B. , Ryman Kate D. , Johnston Robert E. 
Published: December 2019

ABSTRACTAttachment of Sindbis virus to the cell surface glycosaminoglycan heparan sulfate (HS) and the selection of this phenotype by cell culture adaptation were investigated. Virus (TR339) was derived from a cDNA clone representing the consensus sequence of strain AR339 (K. L. McKnight, D. A. Simpson, S. C. Lin, T. A. Knott, J. M. Polo, D. F. Pence, D. B. Johannsen, H. W. Heidner, N. L. Davis, and R. E. Johnston, J. Virol. 70:1981–1989, 1996) and from mutant clones containing either one or two dominant cell culture adaptations in the E2 structural glycoprotein (Arg instead of Ser at E2 position 1 [designated TRSB]) or this mutation plus Arg for Ser at E2 114 [designated TRSB-R114]). The consensus virus, TR339, bound to baby hamster kidney (BHK) cells very poorly. The mutation in TRSB increased binding 10- to 50-fold, and the additional mutation in TRSB-R114 increased binding 3- to 5-fold over TRSB. The magnitude of binding was positively correlated with the degree of cell culture adaptation and with attenuation of these viruses in neonatal mice. HS was identified as the attachment receptor for the mutant viruses by the following experimental results. (i) Low concentrations of soluble heparin inhibited plaque formation on and binding of mutant viruses to BHK cells by >95%. In contrast, TR339 showed minimal inhibition at high concentrations. (ii) Binding and infectivity of TRSB-R114 was sensitive to digestion of cell surface HS with heparinase III, and TRSB was sensitive to both heparinase I and heparinase III. TR339 infectivity was only slightly affected by either digestion. (iii) Radiolabeled TRSB and TRSB-R114 attached efficiently to heparin-agarose beads in binding assays, while TR339 showed virtually no binding. (iv) Binding and infectivity of TRSB and TRSB-R114, but not TR339, were greatly reduced on Chinese hamster ovary cells deficient in HS specifically or all glycosaminoglycans. (v) High-multiplicity-of-infection passage of TR339 on BHK cell cultures resulted in rapid coselection of high-affinity binding to BHK cells and attachment to heparin-agarose beads. Sequencing of the passaged virus population revealed a mutation from Glu to Lys at E2 70, a mutation common to many laboratory strains of Sindbis virus. These results suggest that TR339, the most virulent virus tested, attaches to cells through a low-affinity, primarily HS-independent mechanism. Adaptive mutations, selected during cell culture growth of Sindbis virus, enhance binding and infectivity by allowing the virus to attach by an alternative mechanism that is dependent on the presence of cell surface HS.
Utilization of Chimeras between Human (HM-175) and Simian (AGM-27) Strains of Hepatitis A Virus To Study the Molecular Basis of Virulence
Authors: Raychaudhuri Gopa , Govindarajan Sugantha , Shapiro Max , Purcell Robert H. , Emerson and Suzanne U. 
Published: December 2019

ABSTRACT

Chimeras between human (HM-175) and simian (AGM-27) strains of hepatitis A virus (HAV) were constructed to evaluate the effect of the 2C gene of AGM-27 on HAV replication in cell culture and virulence in tamarins (
Saguinus mystax
) and chimpanzees (
Pan troglodytes
). Kinetic studies and radioimmunofocus assays demonstrated that replacement of the 2C gene of HAV/7, a cell culture-adapted strain of HM-175, with that of AGM-27 drastically reduced the ability of the virus to replicate in cultured cells. Intragenic chimeras containing AGM-27 sequences in either the 5′ or 3′ half of the 2C gene replicated in cell culture at an intermediate level. Whereas HAV/7 is attenuated for tamarins, a chimera containing the simian virus 2C gene in the HAV/7 background was virulent in tamarins, demonstrating that the simian virus 2C gene alone can confer the phenotype of virulence to an otherwise attenuated virus. Clusters of AGM-27-specific residues near both ends of the 2C protein were required for virulence since a chimera containing AGM-27 sequences in the carboxy-terminal half of 2C was partially attenuated for tamarins while one containing AGM-27 sequences only in the amino-terminal half of 2C was even more attenuated. Chimeras containing either the entire or only the 3′ half of the simian virus 2C gene in the HAV/7 background were attenuated for chimpanzees.
Recognition of Herpes Simplex Virus Type 2 Tegument Proteins by CD4 T Cells Infiltrating Human Genital Herpes Lesions
Authors: Koelle David M. , Frank Jeannine M. , Johnson Matthew L. , Kwok William W. 
Published: December 2019

ABSTRACTThe local cellular immune response to herpes simplex virus (HSV) is important in the control of recurrent HSV infection. The antiviral functions of infiltrating CD4-bearing T cells may include cytotoxicity, inhibition of viral growth, lymphokine secretion, and support of humoral and CD8 responses. The antigens recognized by many HSV-specific CD4 T cells localizing to genital HSV-2 lesions are unknown. T cells recognizing antigens encoded within map units 0.67 to 0.73 of HSV DNA are frequently recovered from herpetic lesions. Expression cloning with this region of DNA now shows that tegument protein VP22 and the viral dUTPase, encoded by genes UL49 and UL50, respectively, are T-cell antigens. Separate epitopes in VP22 were defined for T-cell clones from each of three patients. Reactivity with the tegument protein encoded by UL21 was identified for an additional patient. Three new epitopes were identified in VP16, a tegument protein associated with VP22. Some tegument-specific CD4 T-cell clones exhibited cytotoxic activity against HSV-infected cells. These results suggest that herpes simplex tegument proteins are processed for antigen presentation in vivo and are possible candidate compounds for herpes simplex vaccines.
Shortening of the Symptom-Free Period in Rhesus Macaques Is Associated with Decreasing Nonsynonymous Variation in the
env
Variable Regions of Simian Immunodeficiency Virus SIVsm during Passage
Authors: Valli P. J. Spencer , Lukashov Vladimir V. , Heeney Jonathan L. , Goudsmit Jaap 
Published: December 2019

ABSTRACT

During six blood passages of simian immunodeficiency virus SIVsm in rhesus macaques, the asymptomatic period shortened from 18 months to 1 month. To study SIVsm envelope gene (
env
) evolution during passage in rhesus macaques, the C1 to CD4 binding regions of multiple clones were sequenced at seroconversion and again at death. The
env
variation found during adaptation was almost completely confined to the variable regions. Intrasample sequence variation among clones at seroconversion was lower than the variation among clones at death. Intrasample variation among clones from a single time point as well as intersample variation decreased during the passage. In the variable regions, the mean number of intrasample nonsynonymous nucleotide substitutions decreased from the first passage (5.26 × 10
−2
± 0.6 × 10
−2
per site) to the fifth passage (2.24 × 10
−2
± 0.4 × 10
−2
per site), whereas in the constant regions, the mean number of intrasample nonsynonymous nucleotide substitutions differed less between the first and fifth passages (1.14 × 10
−2
± 0.27 × 10
−2
and 0.80 × 10
−2
± 0.24 × 10
−2
per site). Shortening of the asymptomatic period coincided with a rise in the
Ks/Ka
ratio (ratio between the number of synonymous [
Ks
] and the number of nonsynonymous [
Ka
] substitutions) from 1.080 in passage one to 1.428 in passage five and mimicked the difference seen in the intrahost evolution between asymptomatic and fast-progressing individuals infected with human immunodeficiency virus type 1. The distribution of nonsynonymous substitutions was biphasic, with most of the adaptation of
env
variable regions occurring in the first three passages. This phase, in which the symptom-free period fell to 4 months, was followed by a plateau phase of apparently reduced adaptation. Analysis of codon usage revealed decreased codon redundancy in the variable regions. Overall, the results suggested a biphasic pattern of adaptation and evolution, with extremely rapid selection in the first three passages followed by an equilibrium or stabilization of the variation between
env
clones at different time points in passages four to six.
Replacement of Loop II of VP1 of the DA Strain with Loop II of the GDVII Strain of Theiler’s Murine Encephalomyelitis Virus Alters Neurovirulence, Viral Persistence, and Demyelination
Authors: Wada Yoshiaki , McCright Ingeborg J. , Whitby Frank G. , Tsunoda Ikuo , Fujinami Robert S. 
Published: December 2019

ABSTRACT
Theiler’s murine encephalomyelitis viruses, which are murine picornaviruses, can cause central nervous system inflammatory disease. To study the role of loop II in capsid protein VP1, two mutant viruses of strain DA in which DA loop II amino acids were replaced with strain GDVII amino acids were constructed. Infection of mice with the two mutant viruses led to dramatically different patterns of disease.
Complete Bovine Leukemia Virus (BLV) Provirus Is Conserved in BLV-Infected Cattle throughout the Course of B-Cell Lymphosarcoma Development
Authors: Tajima Shigeru , Ikawa Yoji , Aida Yoko 
Published: December 2019

ABSTRACT

Bovine leukemia virus (BLV) and human T-cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2) belong to the same subfamily of oncoviruses. Defective HTLV-1 proviral genomes have been found in more than half of all patients with adult T-cell leukemia examined. We have characterized the genomic structure of integrated BLV proviruses in peripheral blood lymphocytes and tumor tissue taken from animals with lymphomas at various stages. Genomic Southern hybridization with
Sac
I, which generates two major fragments of BLV proviral DNA, yielded only bands that corresponded to a full-size provirus in all of 23 cattle at the lymphoma stage and in 7 BLV-infected but healthy cattle. Long PCR with primers located in long terminal repeats clearly demonstrated that almost the complete provirus was retained in all of 27 cattle with lymphomas and in 19 infected but healthy cattle. However, in addition to a PCR product that corresponded to a full-size provirus, a fragment shorter than that of the complete virus was produced in only one of the 27 animals with lymphomas. Moreover, when we performed conventional PCR with a variety of primers that spanned the entire BLV genome to detect even small defects, PCR products were produced that specifically covered the entire BLV genome in all of the 40 BLV-infected cattle tested. Therefore, it appears that at least one copy of the full-length BLV proviral genome was maintained in each animal throughout the course of the disease and, in addition, that either large or small deletions of proviral genomes may be very rare events in BLV-infected cattle.
Migration of Mitochondria to Viral Assembly Sites in African Swine Fever Virus-Infected Cells
Authors: Rojo Gema , Chamorro Margarita , Salas María L. , Viñuela Eladio , Cuezva JoséM. , Salas José 
Published: December 2019

ABSTRACT
An examination by electron microscopy of the viral assembly sites in Vero cells infected with African swine fever virus showed the presence of large clusters of mitochondria located in their proximity. These clusters surround viral factories that contain assembling particles but not factories where only precursor membranes are seen. Immunofluorescence microscopy revealed that these accumulations of mitochondria are originated by a massive migration of the organelle to the virus assembly sites. Virus infection also promoted the induction of the mitochondrial stress-responsive proteins p74 and cpn 60 together with a dramatic shift in the ultrastructural morphology of the mitochondria toward that characteristic of actively respiring organelles. The clustering of mitochondria around the viral factory was blocked in the presence of the microtubule-disassembling drug nocodazole, indicating that these filaments are implicated in the transport of the mitochondria to the virus assembly sites. The results presented are consistent with a role for the mitochondria in supplying the energy that the virus morphogenetic processes may require and make of the African swine fever virus-infected cell a paradigm to investigate the mechanisms involved in the sorting of mitochondria within the cell.
Analysis of Functional Conservation in the Surface and Transmembrane Glycoprotein Subunits of Human T-Cell Leukemia Virus Type 1 (HTLV-1) and HTLV-2
Authors: Rosenberg Arielle R. , Delamarre Lélia , Preira Anna , Dokhélar Marie-Christine 
Published: December 2019

ABSTRACTHuman T-cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2) are closely related retroviruses with nucleotide sequences that are 65% identical. To determine whether their envelope glycoproteins function similarly and to define the molecular determinants of HTLV-2 envelope-mediated functions, we have used pseudotyped viruses and have introduced mutations into regions of the HTLV-2 glycoproteins homologous to those known to be important for HTLV-1 glycoprotein functions. The envelopes of the two viruses could be exchanged with no loss of infectivity, suggesting that the glycoproteins function in broadly similar ways. However, comparative analysis of the HTLV-1 and HTLV-2 glycoproteins showed subtle differences in the structure-function relationships of the two surface glycoprotein (SU) subunits, even though they recognize the same receptor. Indeed, mutations introduced at equivalent positions in the two SU glycoproteins resulted in different phenotypes in the two viruses. The scenario is the opposite for the transmembrane glycoprotein (TM) subunits, in which the functional domains of the two viruses are strictly conserved, confirming the involvement of the TM ectodomain in postfusion events required for full infectivity of the HTLVs. Thus, although they recognize the same receptor, the HTLV-1 and HTLV-2 SU subunits have slightly different ways of transducing the conformational information that primes a common fusion mechanism effected by similar TM subunits.
Stabilization of Human Immunodeficiency Virus Type 1 Envelope Glycoprotein Trimers by Disulfide Bonds Introduced into the gp41 Glycoprotein Ectodomain
Authors: Farzan Michael , Choe Hyeryun , Desjardins Elizabeth , Sun Ying , Kuhn Jens , Cao Jie , Archambault Danielle , Kolchinsky Peter , Koch Markus , Wyatt Richard , Sodroski Joseph 
Published: December 2019

ABSTRACT
Biochemical and structural studies of fragments of the ectodomain of the human immunodeficiency virus type 1 (HIV-1) gp41 transmembrane envelope glycoprotein have demonstrated that the molecular contacts between alpha helices allow the formation of a trimeric coiled coil. By introducing cysteine residues into specific locations along these alpha helices, the normally labile HIV-1 gp160 envelope glycoprotein was converted into a stable disulfide-linked oligomer. Although proteolytic cleavage into gp120 and gp41 glycoproteins was largely blocked, the disulfide-linked oligomer was efficiently transported to the cell surface and was recognized by a series of conformationally dependent antibodies. The pattern of hetero-oligomer formation between this construct and an analogous construct lacking portions of the gp120 variable loops and of the gp41 cytoplasmic tail demonstrates that these oligomers are trimers. These results support the relevance of the proposed gp41 structure and intersubunit contacts to the native, complete HIV-1 envelope glycoprotein. Disulfide-mediated stabilization of the labile HIV-1 envelope glycoprotein oligomer, which has been suggested to possess advantages as an immunogen, may assist attempts to develop vaccines.
Resistance-Associated Loss of Viral Fitness in Human Immunodeficiency Virus Type 1: Phenotypic Analysis of Protease and
gag
Coevolution in Protease Inhibitor-Treated Patients
Authors: Mammano Fabrizio , Petit Caroline , Clavel François 
Published: December 2019

ABSTRACT
We have studied the phenotypic impact of adaptative Gag cleavage site mutations in patient-derived human immunodeficiency virus type 1 (HIV-1) variants having developed resistance to the protease inhibitor ritonavir or saquinavir. We found that Gag mutations occurred in a minority of resistant viruses, regardless of the duration of the treatment and of the protease mutation profile. Gag mutations exerted only a partial corrective effect on resistance-associated loss of viral fitness. Reconstructed viruses with resistant proteases displayed multiple Gag cleavage defects, and in spite of Gag adaptation, several of these defects remained, explaining the limited corrective effect of cleavage site mutations on fitness. Our data provide clear evidence of the interplay between resistance and fitness in HIV-1 evolution in patients treated with protease inhibitors.
Human T-Lymphotropic Virus Type 2 (HTLV-2) Provirus in Circulating Cells of the Monocyte/Macrophage Lineage in Patients Dually Infected with Human Immunodeficiency Virus Type 1 and HTLV-2 and Having Predominantly Sensory Polyneuropathy
Authors: Zehender Gianguglielmo , Meroni Luca , Varchetta Stefania , De Maddalena Chiara , Cavalli Barbara , Gianotto Monica , Bianchi Bosisio Anna , Colasante Chiara , Rizzardini Giuliano , Moroni Mauro , Galli Massimo 
Published: December 2019

ABSTRACT
We investigated the presence of human T-lymphotropic virus type 2 (HTLV-2) DNA in the peripheral blood mononuclear cell subsets obtained from 18 patients coinfected with human immunodeficiency virus type 1 and HTLV-2, 6 of whom also had predominantly sensory polyneuropathy (PSP). HTLV-2 DNA and RNA were found in CD8- and CD19-positive cells, and, for patients with PSP, in CD14-positive cells as well. Furthermore, the patients with PSP had higher proviral loads than those without PSP.
AIDS Vaccination Studies Using an Ex Vivo Feline Immunodeficiency Virus Model: Detailed Analysis of the Humoral Immune Response to a Protective Vaccine
Authors: Mazzetti Paola , Giannecchini Simone , Del Mauro Daniela , Matteucci Donatella , Portincasa Pietro , Merico Antonio , Chezzi Carlo , Bendinelli Mauro 
Published: December 2019

ABSTRACTThe feline immunodeficiency virus (FIV) cat model is extensively used to investigate possible vaccination approaches against AIDS in humans. Although consistent levels of protection have been achieved with FIV, as with other model systems, by immunizing with whole inactivated virus or fixed infected cells, the mechanisms responsible for protection are elusive. In previous studies we showed that cats immunized with a vaccine consisting of fixed infected cells were protected or unprotected against cell-free or cell-associated FIV challenge depending on the time interval between completion of vaccination and challenge. In an attempt to define possible humoral immune correlates of protection, selected sera harvested at the times of challenge from such cats were examined for anti-FIV-antibody titers and properties by using binding and functional immunological assays. Binding assays included quantitative Western blotting, enzyme-linked tests for antibodies to FIV glycoproteins and immunodominant linear epitopes, and tests for measuring conformation dependence and avidity of anti-viral-envelope antibodies. Functional assays included virus neutralization performed with two different cell substrates, complement- and antibody-dependent virolysis, blocking of reverse transcriptase, and an assay that measured the ability of sera to prevent FIV growth in cocultures of infected and uninfected cells. Despite the wide spectrum of parameters investigated, no correlation between vaccine-induced protection and the humoral parameters measured was noted.
Full-Length Human Immunodeficiency Virus Type 1 Genomes from Subtype C-Infected Seroconverters in India, with Evidence of Intersubtype Recombination
Authors: Lole Kavita S. , Bollinger Robert C. , Paranjape Ramesh S. , Gadkari Deepak , Kulkarni Smita S. , Novak Nicole G. , Ingersoll Roxann , Sheppard Haynes W. , Ray Stuart C. 
Published: December 2019

ABSTRACT

The development of an effective human immunodeficiency virus type 1 (HIV-1) vaccine is likely to depend on knowledge of circulating variants of genes other than the commonly sequenced
gag
and
env
genes. In addition, full-genome data are particularly limited for HIV-1 subtype C, currently the most commonly transmitted subtype in India and worldwide. Likewise, little is known about sequence variation of HIV-1 in India, the country facing the largest burden of HIV worldwide. Therefore, the objective of this study was to clone and characterize the complete genome of HIV-1 from seroconverters infected with subtype C variants in India. Cocultured HIV-1 isolates were obtained from six seroincident individuals from Pune, India, and virtually full-length HIV-1 genomes were amplified, cloned, and sequenced from each. Sequence analysis revealed that five of the six genomes were of subtype C, while one was a mosaic of subtypes A and C, with multiple breakpoints in
env
,
nef
, and the 3′ long terminal repeat as determined by both maximal χ
2
analysis and phylogenetic bootstrapping. Sequences were compared for preservation of known cytotoxic T lymphocyte (CTL) epitopes. Compared with those of the HIV-1
LAI
sequence, 38% of well-defined CTL epitopes were identical. The proportion of nonconservative substitutions for Env, at 61%, was higher (
P
< 0.001) than those for Gag (24%), Pol (18%), and Nef (32%). Therefore, characterized CTL epitopes demonstrated substantial differences from subtype B laboratory strains, which were most pronounced in Env. Because these clones were obtained from Indian seroconverters, they are likely to facilitate vaccine-related efforts in India by providing potential antigens for vaccine candidates as well as for assays of vaccine responsiveness.
Chimeric Lyssavirus Glycoproteins with Increased Immunological Potential
Authors: Jallet Corinne , Jacob Yves , Bahloul Chokri , Drings Astrid , Desmezieres Emmanuel , Tordo Noël , Perrin Pierre 
Published: December 2019

ABSTRACTThe rabies virus glycoprotein molecule (G) can be divided into two parts separated by a flexible hinge: the NH2half (site II part) containing antigenic site II up to the linear region (amino acids [aa] 253 to 275 encompassing epitope VI [aa 264]) and the COOH half (site III part) containing antigenic site III and the transmembrane and cytoplasmic domains. The structural and immunological roles of each part were investigated by cell transfection and mouse DNA-based immunization with homogeneous and chimeric G genes formed by fusion of the site II part of one genotype (GT) with the site III part of the same or another GT. Various site II-site III combinations between G genes of PV (Pasteur virus strain) rabies (GT1), Mokola (GT3), and EBL1 (European bat lyssavirus 1 [GT5]) viruses were tested. Plasmids pGPV-PV, pGMok-Mok, pGMok-PV, and pGEBL1-PV induced transient expression of correctly transported and folded antigens in neuroblastoma cells and virus-neutralizing antibodies against parental viruses in mice, whereas, pG-PVIII (site III part only) and pGPV-Mok did not. The site III part of PV (GT1) was a strong inducer of T helper cells and was very effective at presenting the site II part of various GTs. Both parts are required for correct folding and transport of chimeric G proteins which have a strong potential value for immunological studies and development of multivalent vaccines. Chimeric plasmid pGEBL1-PV broadens the spectrum of protection against European lyssavirus genotypes (GT1, GT5, and GT6).