scholarly journals Infectious Molecular Clones from a Simian Immunodeficiency Virus-Infected Rapid-Progressor (RP) Macaque: Evidence of Differential Selection of RP-Specific Envelope Mutations In Vitro and In Vivo

2006 ◽  
Vol 80 (3) ◽  
pp. 1463-1475 ◽  
Author(s):  
Takeo Kuwata ◽  
Houman Dehghani ◽  
Charles R. Brown ◽  
Ronald Plishka ◽  
Alicia Buckler-White ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Simian Immunodeficiency Virus ◽  
Mononuclear Cells ◽  
Rapid Progression ◽  
Peripheral Blood Mononuclear ◽  
Human T Cell ◽  
Immunodeficiency Virus ◽  
Infectious Molecular Clones

ABSTRACT A minor fraction of simian immunodeficiency virus (SIV)-infected macaques progress rapidly to AIDS in the absence of SIV-specific immune responses. Common mutations in conserved residues of env in three SIVsmE543-3-infected rapid-progressor (RP) macaques suggest the evolution of a common viral variant in RP macaques. The goal of the present study was to analyze the biological properties of these variants in vitro and in vivo through the derivation of infectious molecular clones. Virus isolated from a SIVsmE543-3-infected RP macaque, H445 was used to inoculate six naive rhesus macaques. Although RP-specific mutations dominated in H445 tissues, they represented only 10% of the population of the virus stock, suggesting a selective disadvantage in vitro. Only one of these macaques (H635) progressed rapidly to AIDS. Plasma virus during primary infection of H635 was similar to the inoculum. However, RP-specific mutations were apparently rapidly reselected by 4 to 9 weeks postinfection. Terminal plasma from H635 was used as a source of viral RNA to generate seven full-length, infectious molecular clones. With the exception of one clone, which was similar to SIVsmE543-3, clones with RP-specific mutations replicated with delayed kinetics in rhesus peripheral blood mononuclear cells and human T-cell lines. None of the clones replicated in monocyte-derived or alveolar macrophages, and all used CCR5 as their major coreceptor. RP variants appear to be well adapted to replicate in vivo in RP macaques but are at a disadvantage in tissue culture compared to their parent, SIVsmE543-3. Therefore, tissue culture may not provide a good surrogate for replication of RP variants in macaques. These infectious clones will provide a valuable reagent to study the roles of specific viral variants in rapid progression in vivo.

scholarly journals Novel, Live Attenuated Simian Immunodeficiency Virus Constructs Containing Major Deletions in Leader RNA Sequences

2001 ◽  
Vol 75 (6) ◽  
pp. 2776-2785 ◽  
Author(s):  
Yongjun Guan ◽  
James B. Whitney ◽  
Chen Liang ◽  
Mark A. Wainberg
Keyword(s):  
Tissue Culture ◽  
Simian Immunodeficiency Virus ◽  
Mononuclear Cells ◽  
Polyacrylamide Gel Electrophoresis ◽  
Wild Type Virus ◽  
Replication Capacity ◽  
Peripheral Blood Mononuclear ◽  
Immunodeficiency Virus

ABSTRACT We have constructed a series of simian immunodeficiency virus (SIV) mutants containing deletions within a 97-nucleotide (nt) region of the leader sequence. Deletions in this region markedly decreased the replication capacity in tissue culture, i.e., in both the C8166 and CEMx174 cell lines, as well as in rhesus macaque peripheral blood mononuclear cells. In addition, these deletions adversely affected the packaging of viral genomic RNA into virions, the processing of Gag precursor proteins, and patterns of viral proteins in virions, as assessed by biochemical labeling and polyacrylamide gel electrophoresis. Different levels of attenuation were achieved by varying the size and position of deletions within this 97-nt region, and among a series of constructs that were generated, it was possible to rank in vitro virulence relative to that of wild-type virus. In all of these cases, the most severe impact on viral replication was observed when the deletions that were made were located at the 3′ rather than 5′ end of the leader region. The potential of viral reversion over protracted periods was investigated by repeated viral passage in CEMx174 cells. The results showed that several of these constructs showed no signs of reversion after more than 6 months in tissue culture. Thus, a series of novel, attenuated SIV constructs have been developed that are significantly impaired in replication capacity yet retain all viral genes. One of these viruses, termed SD4, may be appropriate for study with rhesus macaques, in order to determine whether reversions will occur in vivo and to further study this virus as a candidate for attenuated vaccination.

scholarly journals Characteristics of a Pathogenic Molecular Clone of an End-Stage Serum-Derived Variant of Simian Immunodeficiency Virus (SIVF359)

2001 ◽  
Vol 75 (19) ◽  
pp. 9328-9338 ◽  
Author(s):  
Lennart Holterman ◽  
Rob Dubbes ◽  
James Mullins ◽  
Gerald Learn ◽  
Henk Niphuis ◽  
...  
Keyword(s):  
Simian Immunodeficiency Virus ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Peripheral Blood Lymphocytes ◽  
Peripheral Blood Mononuclear ◽  
Molecular Clone ◽  
Immunodeficiency Virus ◽  
End Stage

ABSTRACT End-stage simian immunodeficiency virus (SIV) isolates are suggested to be the most fit of the evolved virulent variants that precipitate the progression to AIDS. To determine if there were common characteristics of end-stage variants which emerge from accelerated cases of AIDS, a molecular clone was derived directly from serum following in vivo selection of a highly virulent SIV isolate obtained by serial end-stage passage in rhesus monkeys (Macaca mulatta). This dominant variant caused a marked cytopathic effect and replicated to very high levels in activated but not resting peripheral blood lymphocytes. Furthermore, although this clone infected but did not replicate to detectable levels in rhesus monocyte-derived macrophages, these cells were able to transmit infection to autologous T cells upon contact. Interestingly, although at low doses this end-stage variant did not use any of the known coreceptors except CCR5, it was able to infect and replicate in human peripheral blood mononuclear cells homozygous for the Δ32 deletion of CCR5, suggesting the use of a novel coreceptor. It represents the first pathogenic molecular clone of SIV derived from viral RNA in serum and provides evidence that not only the genetic but also the biological characteristics acquired by highly fit late-stage disease variants may be distinct in different hosts.

scholarly journals Simian Immunodeficiency Virus SIVagm Efficiently Utilizes Non-CCR5 Entry Pathways in African Green Monkey Lymphocytes: Potential Role for GPR15 and CXCR6 as Viral Coreceptors

2015 ◽  
Vol 90 (5) ◽  
pp. 2316-2331 ◽  
Author(s):  
Nadeene E. Riddick ◽  
Fan Wu ◽  
Kenta Matsuda ◽  
Sonya Whitted ◽  
Ilnour Ourmanov ◽  
...  
Keyword(s):  
T Cells ◽  
Mononuclear Cells ◽  
Rhesus Macaques ◽  
Target Cells ◽  
Peripheral Blood Mononuclear ◽  
Immunodeficiency Virus ◽  
Sooty Mangabeys ◽  
Natural Hosts

ABSTRACTAfrican green monkeys (AGM) are natural hosts of simian immunodeficiency virus (SIV), and infection in these animals is generally nonpathogenic, whereas infection of nonnatural hosts, such as rhesus macaques (RM), is commonly pathogenic. CCR5 has been described as the primary entry coreceptor for SIVin vivo, while human-derived CXCR6 and GPR15 also appear to be usedin vitro. However, sooty mangabeys that are genetically deficient in CCR5 due to an out-of-frame deletion are infectible with SIVsmm, indicating that SIVsmm can use alternative coreceptorsin vivo. In this study, we examined the CCR5 dependence of SIV strains derived from vervet AGM (SIVagmVer) and the ability of AGM-derived GPR15 and CXCR6 to serve as potential entry coreceptors. We found that SIVagmVer replicated efficiently in AGM and RM peripheral blood mononuclear cells (PBMC) in the presence of the CCR5 antagonist maraviroc, despite the fact that maraviroc was capable of blocking the CCR5-tropic strains SIVmac239, SIVsmE543-3, and simian-human immunodeficiency virus SHIV-AD8 in RM PBMC. We also found that AGM CXCR6 and AGM GPR15, to a lesser extent, supported entry of pseudotype viruses bearing SIVagm envelopes, including SIVagm transmitted/founder envelopes. Lastly, we found that CCR5, GPR15, and CXCR6 mRNAs were detected in AGM and RM memory CD4+T cells. These results suggest that GPR15 and CXCR6 are expressed on AGM CD4+T cells and are potential alternative coreceptors for SIVagm usein vivo. These data suggest that the use of non-CCR5 entry pathways may be a common feature of SIV replication in natural host species, with the potential to contribute to nonpathogenicity in these animals.IMPORTANCEAfrican green monkeys (AGM) are natural hosts of SIV, and infection in these animals generally does not cause AIDS, whereas SIV-infected rhesus macaques (RM) typically develop AIDS. Although it has been reported that SIV generally uses CD4 and CCR5 to enter target cellsin vivo, other molecules, such as GPR15 and CXCR6, also function as SIV coreceptorsin vitro. In this study, we investigated whether SIV from vervet AGM can use non-CCR5 entry pathways, as has been observed in sooty mangabeys. We found that SIVagmVer efficiently replicated in AGM and RM peripheral blood mononuclear cells in the presence of the CCR5 antagonist maraviroc, suggesting that non-CCR5 entry pathways can support SIVagm entry. We found that AGM-derived GPR15 and CXCR6 support SIVagmVer entryin vitroand may serve as entry coreceptors for SIVagmin vivo, since their mRNAs were detected in AGM memory CD4+T cells, the preferred target cells of SIV.

scholarly journals Response of Simian Immunodeficiency Virus to the Novel Nucleoside Reverse Transcriptase Inhibitor 4′-Ethynyl-2-Fluoro-2′-DeoxyadenosineIn VitroandIn Vivo

2012 ◽  
Vol 56 (9) ◽  
pp. 4707-4712 ◽  
Author(s):  
Michael Murphey-Corb ◽  
Premeela Rajakumar ◽  
Heather Michael ◽  
Julia Nyaundi ◽  
Peter J. Didier ◽  
...  
Keyword(s):  
Drug Treatment ◽  
Reverse Transcriptase ◽  
Simian Immunodeficiency Virus ◽  
Mononuclear Cells ◽  
Clinical Signs ◽  
Long Term Treatment ◽  
Essential Components ◽  
Immunodeficiency Virus

ABSTRACTNucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) are essential components in first-line therapy for human immunodeficiency virus (HIV) infection. However, long-term treatment with existing NRTIs can be associated with significant toxic side effects and the emergence of drug-resistant strains. The identification of new NRTIs for the continued management of HIV-infected people therefore is paramount. In this report, we describe the response of a primary isolate of simian immunodeficiency virus (SIV) to 4′-ethynyl-2-fluoro-2′-deoxyadenosine (EFdA) bothin vitroandin vivo. EFdA was 3 orders of magnitude better than tenofovir (TFV), zidovudine (AZT), and emtricitabine (FTC) in blocking replication of SIV in monkey peripheral blood mononuclear cells (PBMCs)in vitro, and in a preliminary study using two SIV-infected macaques with advanced AIDS, it was highly effective at treating SIV infection and AIDS symptomsin vivo. Both animals had 3- to 4-log decreases in plasma virus burden within 1 week of EFdA therapy (0.4 mg/kg of body weight, delivered subcutaneously twice a day) that eventually became undetectable. Clinical signs of disease (diarrhea, weight loss, and poor activity) also resolved within the first month of treatment. No detectable clinical or pathological signs of drug toxicity were observed within 6 months of continuous therapy. Virus suppression was sustained until drug treatment was discontinued, at which time virus levels rebounded. Although the rebound virus contained the M184V/I mutation in the viral reverse transcriptase, EFdA was fully effective in maintaining suppression of mutant virus throughout the drug treatment period. These results suggest that expanded studies with EFdA are warranted.

scholarly journals Kinetics of Replication of a Partially Attenuated Virus and of the Challenge Virus during a Three-Year Intersubtype Feline Immunodeficiency Virus Superinfection Experiment in Cats

1999 ◽  
Vol 73 (2) ◽  
pp. 1518-1527 ◽  
Author(s):  
Mauro Pistello ◽  
Donatella Matteucci ◽  
Giancarlo Cammarota ◽  
Paola Mazzetti ◽  
Simone Giannecchini ◽  
...  
Keyword(s):  
Feline Immunodeficiency Virus ◽  
Mononuclear Cells ◽  
Acute Infection ◽  
Low Grade ◽  
Peripheral Blood Mononuclear ◽  
Subtype B ◽  
Immunodeficiency Virus ◽  
Attenuated Virus

ABSTRACT The effects of preinfecting cats with a partially attenuated feline immunodeficiency virus (FIV) on subsequent infection with a fully virulent FIV belonging to a different subtype were investigated. Eight specific-pathogen-free cats were preinfected with graded doses of a long-term in vitro-cultured cell-free preparation of FIV Petaluma (FIV-P, subtype A). FIV-P established a low-grade or a silent infection in the inoculated animals. Seven months later, the eight preinfected cats and two uninfected cats were challenged with in vivo-grown FIV-M2 (subtype B) and periodically monitored for immunological and virological status. FIV-P-preinfected cats were not protected from acute infection by FIV-M2, and the sustained replication of this virus was accompanied by a reduction of FIV-P viral loads in the peripheral blood mononuclear cells and plasma. However, from 2 years postchallenge (p.c.) until 3 years p.c., when the experiment was terminated, preinfected cats exhibited reduced total viral burdens, and some also exhibited a diminished decline of circulating CD4+ T lymphocytes relative to control cats infected with FIV-M2 alone. Interestingly, most of the virus detected in challenged cats at late times p.c. was of FIV-P origin, indicating that the preinfecting, attenuated virus had become largely predominant. By the end of follow-up, two challenged cats had no FIV-M2 detectable in the tissues examined. The possible mechanisms underlying the interplay between the two viral populations are discussed.

scholarly journals The Liver Is a Major Organ for Clearing Simian Immunodeficiency Virus in Rhesus Monkeys

2002 ◽  
Vol 76 (10) ◽  
pp. 5271-5273 ◽  
Author(s):  
Linqi Zhang ◽  
Peter J. Dailey ◽  
Agegnehu Gettie ◽  
Jim Blanchard ◽  
David D. Ho
Keyword(s):  
Simian Immunodeficiency Virus ◽  
Viral Vectors ◽  
Mononuclear Cells ◽  
Significant Proportion ◽  
Viral Clearance ◽  
High Rate ◽  
Peripheral Blood Mononuclear ◽  
Immunodeficiency Virus ◽  
Rapid Clearance

ABSTRACT Infection with human or simian immunodeficiency virus (SIV) is characterized by the rapid turnover of both viral particles and productively infected cells. It has recently been reported that the clearance of SIV in vivo is exceedingly fast, with half-lives on the order of minutes. The underlying mechanism or site responsible for this rapid clearance, however, remains unknown. To investigate this issue, we chose to infuse infectious SIVmac239 grown from autologous peripheral blood mononuclear cells that were radioactively labeled by [35S]methionine and [35S]cysteine. This approach eliminates from the viral membrane alloantigens that may have a significant impact on viral clearance. In addition, this approach also permits identification of the sites of viral clearance by measuring the radioactive intensity, even if degradation of SIV RNA occurs in tissues. We now report that the half-life of infused SIV in blood is extremely close to estimates from a previous study, in which unlabeled SIV grown in a heterologous cell line was used. The allogeneic effect due to the presence of human antigens on the surfaces of virions may, therefore, play a minimal role in the high rate of virion clearance. Moreover, close to 30% of infused radioactivity was found in the liver and measureable amounts were detected in the lungs (5.4%), lymph nodes (3.0%), and spleen (0.4%). The detection of a significant proportion of infused virus in the liver suggests that viral clearance from circulation is mediated by a common, nonspecific mechanism, such as the phagocytic functions of the reticuloendothelial system. The rapid clearance and degradation of exogenously infused virions may pose a major obstacle for gene therapy with viral vectors, unless strategies to overcome the rapid in vivo elimination of these particles are developed.

scholarly journals Limited Breadth of the Protective Immunity Elicited by Simian Immunodeficiency Virus SIVmne gp160 Vaccines in a Combination Immunization Regimen

1999 ◽  
Vol 73 (1) ◽  
pp. 618-630 ◽  
Author(s):  
Patricia Polacino ◽  
Virginia Stallard ◽  
James E. Klaniecki ◽  
David C. Montefiori ◽  
Alphonse J. Langlois ◽  
...  
Keyword(s):  
Simian Immunodeficiency Virus ◽  
Neutralizing Antibodies ◽  
Protective Immunity ◽  
Mononuclear Cells ◽  
Protective Mechanism ◽  
Peripheral Blood Mononuclear ◽  
Challenge Virus ◽  
Human T Cell ◽  
Homologous Virus ◽  
Immunodeficiency Virus

ABSTRACT We previously reported that immunization with recombinant simian immunodeficiency virus SIVmne envelope (gp160) vaccines protected macaques against an intravenous challenge by the cloned homologous virus, E11S. In this study, we confirmed this observation and found that the vaccines were effective not only against virus grown on human T-cell lines but also against virus grown on macaque peripheral blood mononuclear cells (PBMC). The breadth of protection, however, was limited. In three experiments, 3 of 10 animals challenged with the parental uncloned SIVmne were completely protected. Of the remaining animals, three were transiently virus positive and four were persistently positive after challenge, as were 10 nonimmunized control animals. Protection was not correlated with levels of serum-neutralizing antibodies against the homologous SIVmne or a related virus, SIVmac251. To gain further insight into the protective mechanism, we analyzed nucleotide sequences in the envelope region of the uncloned challenge virus and compared them with those present in the PBMC of infected animals. The majority (85%) of the uncloned challenge virus was homologous to the molecular clone from which the vaccines were made (E11S type). The remaining 15% contained conserved changes in the V1 region (variant types). Control animals infected with this uncloned virus had different proportions of the two genotypes, whereas three of four immunized but persistently infected animals had >99% of the variant types early after infection. These results indicate that the protective immunity elicited by recombinant gp160 vaccines is restricted primarily to the homologous virus and suggest the possibility that immune responses directed to the V1 region of the envelope protein play a role in protection.

scholarly journals Isolation and Characterization of a Neuropathogenic Simian Immunodeficiency Virus Derived from a Sooty Mangabey

1998 ◽  
Vol 72 (11) ◽  
pp. 8841-8851 ◽  
Author(s):  
Francis J. Novembre ◽  
Juliette De Rosayro ◽  
Shawn P. O’Neil ◽  
Daniel C. Anderson ◽  
Sherry A. Klumpp ◽  
...  
Keyword(s):  
Rhesus Macaque ◽  
Simian Immunodeficiency Virus ◽  
Mononuclear Cells ◽  
Giant Cells ◽  
Sooty Mangabey ◽  
Neurologic Disease ◽  
Peripheral Blood Mononuclear ◽  
Isolation And Characterization ◽  
Immunodeficiency Virus

ABSTRACT Transfusion of blood from a simian immunodeficiency virus (SIV)- and simian T-cell lymphotropic virus-infected sooty mangabey (designated FGb) to rhesus and pig-tailed macaques resulted in the development of neurologic disease in addition to AIDS. To investigate the role of SIV in neurologic disease, virus was isolated from a lymph node of a pig-tailed macaque (designated PGm) and the cerebrospinal fluid of a rhesus macaque (designated ROn2) and passaged to additional macaques. SIV-related neuropathogenic effects were observed in 100% of the pig-tailed macaques inoculated with either virus. Lesions in these animals included extensive formation of SIV RNA-positive giant cells in the brain parenchyma and meninges. Based upon morphology, the majority of infected cells in both lymphoid and brain tissue appeared to be of macrophage lineage. The virus isolates replicated very well in pig-tailed and rhesus macaque peripheral blood mononuclear cells (PBMC) with rapid kinetics. Differential replicative abilities were observed in both PBMC and macrophage populations, with viruses growing to higher titers in pig-tailed macaque cells than in rhesus macaque cells. An infectious molecular clone of virus derived from the isolate from macaque PGm (PGm5.3) was generated and was shown to have in vitro replication characteristics similar to those of the uncloned virus stock. While molecular analyses of this virus revealed its similarity to SIV isolates from sooty mangabeys, significant amino acid differences in Env and Nef were observed. This virus should provide an excellent system for investigating the mechanism of lentivirus-induced neurologic disease.

scholarly journals Drug Combination of AZT and ddl: Synergism of Action and Prevention of Appearance of AZT-Resistance

1994 ◽  
Vol 5 (1) ◽  
pp. 51-55 ◽  
Author(s):  
G. Antonelli ◽  
F. Dianzani ◽  
D. Bellarosa ◽  
O. Turriziani ◽  
E. Riva ◽  
...  
Keyword(s):  
In Vitro Selection ◽  
Mononuclear Cells ◽  
Peripheral Blood Mononuclear ◽  
Immunodeficiency Virus ◽  
Resistant Strains ◽  
Azt Resistance ◽  
Hiv 1

Both 3′-azido-3′-deoxythymidine (AZT) and 2′,3′-dideoxynosine (ddl) strongly inhibit the replication of human immunodeficiency virus type 1 (HIV-1). Here, it is shown that combination of AZT and ddl at concentrations that are readily achievable in vivo synergistically inhibit HIV-1 replication in C8166 cells and peripheral blood mononuclear cells. The synergism is significant even when the effect of AZT and ddl alone was negligible. Our findings show that AZT-resistance is less likely to occur when a combination of AZT and ddl is used. Particularly, generation of AZT-resistant strains by in vitro selection is prevented, or delayed, by the combination of AZT plus ddl. Taken together these observations provide a rationale for combination of AZT and ddl in the therapy of AIDS patients.

Constitutive Activation of STATs Upon In Vivo Human Immunodeficiency Virus Infection

Blood ◽  
1999 ◽  
Vol 94 (12) ◽  
pp. 4202-4209 ◽  
Author(s):  
Chiara Bovolenta ◽  
Laura Camorali ◽  
Alessandro L. Lorini ◽  
Silvia Ghezzi ◽  
Elisa Vicenzi ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Mononuclear Cells ◽  
Janus Kinase ◽  
Peripheral Blood Mononuclear ◽  
Constitutive Activation ◽  
Protein Levels ◽  
Immunodeficiency Virus ◽  
Stat Pathway

Abstract Infection by the human immunodeficiency virus (HIV) either upregulates or downregulates the expression of several cytokines and interferons (IFNs) that use the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway for signal transduction. However, very little is known on the state of activation of the JAK/STAT pathway after HIV infection either in vivo or in vitro. In this regard, we report here that a constitutive activation of a C-terminal truncated STAT5 (STAT5▵) and of STAT1 occurs in the majority (∼75%) of individuals with progressive HIV disease. We have further demonstrated that, among peripheral blood mononuclear cells (PBMCs), STAT5▵ is activated preferentially in CD4+ T cells. In contrast to a published report, expression of STATs from PBMCs of infected individuals was comparable with that of seronegative donors. In addition, in vitro infection of mitogen-activated PBMCs with a panel of laboratory-adapted and primary HIV strains characterized by differential usage of chemokine coreceptors did not affect STAT protein levels. However, enhanced activation of STAT was observed after in vitro infection of resting PBMCs and nonadherent PBMCs by different viral strains. Thus, constitutive STAT activation in CD4+T lymphocytes represents a novel finding of interest also as a potential new marker of immunological reconstitution of HIV-infected individuals.

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