Génétique des Virus, Institut Cochin (INSERM U567 CNRS UMR 8104), 22 rue Méchain, 75014 Paris, France1
Immunopathologie Cellulaire et Moléculaire, Institut National de la Recherche Agronomique (INRA)2 and UMR INRA ENVA AFSSA 1161 de Virologie, Ecole Nationale Vétérinaire dAlfort (ENVA)3, Maisons-Alfort, France
Author for correspondence: Pierre Sonigo. Fax +33 1 40 51 64 30. e-mail sonigo{at}cochin.inserm.fr
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Abstract |
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Numerous studies have shown that certain antibodies directed against HIV type 1 (HIV-1) can enhance virus replication in various cell-culture models (Eaton et al., 1994 ; Homsy et al., 1989
; Jouault et al., 1989
; Kostrikis et al., 1996
; Robinson et al., 1987
, 1990
, 1991
; Schutten et al., 1995
; Sullivan et al., 1995
, 1998
; Takeda et al., 1988
) or are associated with disease progression (Fust et al., 1994
; Homsy et al., 1990
; Montefiori et al., 1991
, 1996
; Toth et al., 1991
) or maternal transmission (Lallemant et al., 1994
; Pancino et al., 1998
). Nevertheless, not all instances of immune-mediated enhancement can be attributed to enhancing antibodies. In our laboratory, plasma virus RNA was detected substantially earlier in cats immunized with env gene DNA than in control cats (Richardson et al., 1997
), though measurable levels of FIV-specific antibodies were not elicited prior to challenge, raising the possibility that the cell-mediated immune response and, in particular, concomitant lymphoid activation had accelerated the course of primary infection.
High-level replication of immunodeficiency viruses in T lymphocytes requires lymphoid activation (Ascher & Sheppard, 1990 ; Bukrinsky et al., 1991
; Zack et al., 1990
), and such activation may limit virus production during natural infection. Indeed, in the chronic stage of HIV-1 infection, exogenous factors that induce immune activation, such as contemporaneous infections (Bentwich et al., 2000
; Bush et al., 1996
; Goletti et al., 1996
), immunization (Brichacek et al., 1996
; OBrien et al., 1995
; Ortigao-de-Sampaio et al., 1998
; Ostrowski et al., 1998
; Stanley et al., 1996
; Staprans et al., 1995
) and administration of cytokines (Davey et al., 1997
; Kovacs et al., 1995
), create bursts of virus replication. Should the level of lymphoid activation also be limiting during the initial stages of natural infection, early virus dissemination could be accelerated under conditions where activation is enhanced.
In our previous DNA vaccination trial, cats were challenged shortly after a series of closely spaced DNA injections and thus presumably during the effector phase of the immune response, when the number of activated lymphocytes, both antigen-specific and non-specific, ought to have been substantially elevated. In the absence of potent antiviral effector functions, amplification of target cells may have given rise to the accelerated infection observed in immunized animals. In the present FIV vaccination trial, we have attempted to decrease the potentially deleterious effect of vaccine-related polyclonal activation by prolonging the intervals between DNA injections and between the final DNA injection and challenge. The vaccine vectors and vaccination schedule are represented schematically in Fig. 1. In the plasmid vector pCMV-env, expression of the env gene of the 34TF10 molecular clone of FIV (Talbott et al., 1989
) was placed under the transcriptional control of the CMV-IE promoter, by PCR amplification of the sequence encoding both the surface (SU) and transmembrane glycoproteins and insertion in the PstI and BamHI sites of the VR1012 plasmid (Vical Inc.). In the vector pCMV-S (Davis et al., 1993
), transcription of the S region of the hepatitis B virus (HBV) envelope gene was placed under the control of the CMV-IE promoter, permitting expression of a heterologous antigen, the S antigen of HBV (HBVsAg). Plasmid DNA was purified by using commercially available materials (Plasmid megakit, Qiagen) and endotoxin was removed by extraction with Triton X-114 (Aida & Pabst, 1990
).
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Similar to the previous DNA vaccination trial, humoral responses directed against envelope glycoproteins were virtually undetectable prior to challenge, while antibodies directed against the HBVsAg, assayed using commercially available reagents (MONOLISA Anti-HBs 3.0, Sanofi Diagnostics Pasteur), were detected in only one of four cats (Mexique) immunized with pCMV-S (data not shown). In an attempt to identify cell-mediated immune correlates of enhancement, T-helper (Th) responses directed against the SU glycoprotein were assayed in PBMC isolated 2 weeks after the fourth DNA injection.
For metabolic labelling with [3H]thymidine, PBMC were resuspended in RPMI 1640 medium containing 10% foetal calf serum, 10 mM HEPES, 2 mM glutamine, antibiotics and 50 µM -mercaptoethanol (complete RPMI) supplemented with 1% feline plasma. Cells (4x105 cells per well) were cultured in triplicate wells of transfer tubes (Costar) in the presence of immunoaffinity-purified FIV SU glycoprotein or medium alone, as well as in the presence of 10 µM concanavalin A, as a control for cell viability, in a final volume of 200 µl. Fifty µl volumes were removed after 24 and 48 h for assay of IL-2. After 72 h, the cells were pulsed with [3H]thymidine (1 µCi per well) and cells were collected onto glass filter paper with a semi-automated cell harvester (Scatron) after overnight incubation. Incorporation of [3H]thymidine was assessed by liquid scintillation counting.
For biological assay of IL-2, the IL-2-dependent CTL-L cell line was adjusted in complete RPMI to 2x105 cells per well and 50 µl volumes were added to the PBMC supernatants and cultivated for 1624 h. The cells were then pulsed with [3H]thymidine as described above. Although proliferative responses were not discernible in any of the cats, antigenic stimulation gave rise to a substantial production of IL-2 in PBMC of two cats (Michelle and Mauritanie) inoculated l.n. with env DNA (Table 1).
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Feline PBMC were resuspended in complete RPMI containing 1% feline plasma. Cells (4x105 cells per well) were cultured in triplicate wells of transfer tubes in the presence of immunoaffinity-purified SU glycoprotein or medium alone, in a final volume of 200 µl. After 24 h, the PBMC were infected by addition of 25 TCID50 of the Wo strain of FIV (Moraillon et al., 1992 ). After a further 24 h, the virus inoculum was removed by washing the cells twice with 0·5 ml RPMI 1640 medium. The PBMC were then resuspended in 200 µl complete RPMI containing 5 µg/ml concanavalin A and 100 U/ml recombinant IL-2 and transferred to wells of 96-well microtitre plates. Half the medium was replaced 4 days after infection. At 4 and 7 days post-infection, aliquots of 10 µl were removed for assay of reverse transcriptase activity. Quantitative densitometry of autoradiography film was performed by using the program NIH Image version 1.54.
For cats immunized with env gene DNA, the extent of virus replication in the absence of antigen-specific stimulation was particularly high in PBMC isolated from two cats (Michelle and Mauritanie) of group 2 and, notably, the only two cats for which Th responses were detected (Table 1). Moreover, even higher levels of replication were obtained after incubation of PBMC from these two cats with SU glycoprotein. For cats of the two control groups, virus replication was particularly efficient in the PBMC of one cat (Mexique) of group 4, immunized with DNA encoding HBsAg, possibly because of immune activation elicited against this heterologous antigen.
Following homologous challenge by intraperitoneal injection of 10 median cat infectious doses (CID50) of the Petaluma strain (Pedersen et al., 1987 ) of FIV (stock provided by M. Hosie, University of Glasgow, UK), prepared from the supernatant of the chronically infected FL-4 cell line (Yamamoto et al., 1991
), virus load in cats was assessed as plasma virus RNA by competitive RTPCR (Richardson et al., 1998
). Virus RNA was detected in the plasma of all but two cats, Moto and Mado, respectively from groups 1 and 3. Peak viraemia occurred between 32 and 36 days for control group 3 or between 25 and 46 days for both control group 4 and immunized group 1 (Table 2
). No statistically significant differences were observed in plasma virus load upon pairwise comparison of these groups at any time point examined.
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In the present study, challenge was performed 10 weeks after the final DNA injection, so as to avoid the effector stage of the immune response. Under these conditions, and in contrast with the results of our previous trial, virus dissemination was not accelerated in cats vaccinated with env gene DNA by the i.d./i.m. routes. Immune-mediated enhancement was, however, observed in cats immunized by l.n. inoculation. Intriguingly, Th activity was restricted to the two cats of this immunization group with particularly rapid kinetics of infection. Moreover, rapid virus dissemination in vivo was linked to increased susceptibility of lymphocytes to ex vivo infection, as would be expected if cellular immunity underlies immune-mediated enhancement. While, given the small number of animals in question, far-reaching conclusions cannot be drawn, these results are consistent with the hypothesis, already formulated (Schwartz, 1994 ), that induction of a virus-specific Th response might provide a pool of highly susceptible target cells upon lentivirus infection and, paradoxically, enhance infection.
These results underscore an apparent paradox in vaccination against lentiviruses: namely, that effective vaccination requires lymphoid activation, but lentiviruses cannot replicate efficiently in lymphocytes unless they are activated. Immune activation may therefore be a confounding factor in vaccination against lentivirus infection, diminishing vaccine efficacy and giving rise to immune-mediated enhancement of infection in the absence of potent counteracting protective responses. At present, the consequences of such activation for susceptibility to infection and disease progression are unknown. A better understanding of immune enhancement of infection is therefore necessary not only to improve upon current vaccines, but also to appreciate the risk that this phenomenon represents in human AIDS trials.
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Acknowledgments |
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Footnotes |
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c Present address: UMR 956 INRA ENVA AFSSA de Biologie Moléculaire et Immunologie Parasitaires et Fongiques, Maisons-Alfort, France.
d Present address: Génopole, Evry, France.
e Present address: Unité de Biologie des Rétrovirus, Institut Pasteur, Paris, France.
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References |
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Ascher, M. S. & Sheppard, H. W. (1990). AIDS as immune system activation. II. The panergic imnesia hypothesis. Journal of Acquired Immune Deficiency Syndromes 3, 177-191.[Medline]
Bentwich, Z., Maartens, G., Torten, D., Lal, A. A. & Lal, R. B. (2000). Concurrent infections and HIV pathogenesis. AIDS 14, 2071-2081.[Medline]
Brichacek, B., Swindells, S., Janoff, E. N., Pirruccello, S. & Stevenson, M. (1996). Increased plasma human immunodeficiency virus type 1 burden following antigenic challenge with pneumococcal vaccine. Journal of Infectious Diseases 174, 1191-1199.[Medline]
Bukrinsky, M. I., Stanwick, T. L., Dempsey, M. P. & Stevenson, M. (1991). Quiescent T lymphocytes as an inducible virus reservoir in HIV-1 infection. Science 254, 423-427.[Medline]
Bush, C. E., Donovan, R. M., Markowitz, N. P., Kvale, P. & Saravolatz, L. D. (1996). A study of HIV RNA viral load in AIDS patients with bacterial pneumonia. Journal of Acquired Immune Deficiency Syndromes and Human Retrovirology 13, 23-26.[Medline]
Davey, R. T.Jr, Chaitt, D. G., Piscitelli, S. C., Wells, M., Kovacs, J. A., Walker, R. E., Falloon, J., Polis, M. A., Metcalf, J. A., Masur, H., Fyfe, G. & Lane, H. C. (1997). Subcutaneous administration of interleukin-2 in human immunodeficiency virus type 1-infected persons. Journal of Infectious Diseases 175, 781-789.[Medline]
Davis, H. L., Michel, M. L. & Whalen, R. G. (1993). DNA-based immunization induces continuous secretion of hepatitis B surface antigen and high levels of circulating antibody. Human Molecular Genetics 2, 1847-1851.[Abstract]
Dawson, J. D. (1994). Comparing treatment groups on the basis of slopes, areas-under-the-curve, and other summary measures. Drug Information Journal 28, 723-732.
Eaton, A. M., Ugen, K. E., Weiner, D. B., Wildes, T. & Levy, J. A. (1994). An anti-gp41 human monoclonal antibody that enhances HIV-1 infection in the absence of complement. AIDS Research and Human Retroviruses 10, 13-18.[Medline]
Fust, G., Toth, F. D., Kiss, J., Ujhelyi, E., Nagy, I. & Banhegyi, D. (1994). Neutralizing and enhancing antibodies measured in complement-restored serum samples from HIV-1-infected individuals correlate with immunosuppression and disease. AIDS 8, 603-609.[Medline]
Goletti, D., Weissman, D., Jackson, R. W., Graham, N. M., Vlahov, D., Klein, R. S., Munsiff, S. S., Ortona, L., Cauda, R. & Fauci, A. S. (1996). Effect of Mycobacterium tuberculosis on HIV replication. Role of immune activation. Journal of Immunology 157, 1271-1278.[Abstract]
Hesselink, W., Sondermeijer, P., Pouwels, H., Verblakt, E. & Dhore, C. (1999). Vaccination of cats against feline immunodeficiency virus (FIV): a matter of challenge. Veterinary Microbiology 69, 109-110.[Medline]
Homsy, J., Meyer, M., Tateno, M., Clarkson, S. & Levy, J. A. (1989). The Fc and not CD4 receptor mediates antibody enhancement of HIV infection in human cells. Science 244, 1357-1360.[Medline]
Homsy, J., Meyer, M. & Levy, J. A. (1990). Serum enhancement of human immunodeficiency virus (HIV) infection correlates with disease in HIV-infected individuals. Journal of Virology 64, 1437-1440.[Medline]
Hosie, M. J., Osborne, R., Reid, G., Neil, J. C. & Jarrett, O. (1992). Enhancement after feline immunodeficiency virus vaccination. Veterinary Immunology and Immunopathology 35, 191-197.[Medline]
Huisman, W., Karlas, J. A., Siebelink, K. H., Huisman, R. C., de Ronde, A., Francis, M. J., Rimmelzwaan, G. F. & Osterhaus, A. D. (1998). Feline immunodeficiency virus subunit vaccines that induce virus neutralising antibodies but no protection against challenge infection. Vaccine 16, 181-187.[Medline]
Johnston, M. I. (2000). The role of nonhuman primate models in AIDS vaccine development. Molecular Medicine Today 6, 267-270.[Medline]
Jouault, T., Chapuis, F., Olivier, R., Parravicini, C., Bahraoui, E. & Gluckman, J.-C. (1989). HIV infection of monocytic cells: role of antibody-mediated virus binding to Fc-gamma receptors. AIDS 3, 125-133.[Medline]
Karlas, J. A., Siebelink, K. H., van Peer, M. A., Huisman, W., Rimmelzwaan, G. F. & Osterhaus, A. D. (1998). Accelerated viraemia in cats vaccinated with fixed autologous FIV-infected cells. Veterinary Immunology and Immunopathology 65, 353-365.[Medline]
Karlas, J. A., Siebelink, K. H. J., van Peer, M. A., Huisman, W., Cuisinier, A. M., Rimmelzwaan, G. F. & Osterhaus, A. D. M. E. (1999). Vaccination with experimental feline immunodeficiency virus vaccines, based on autologous infected cells, elicits enhancement of homologous challenge infection. Journal of General Virology 80, 761-765.[Abstract]
Kostrikis, L. G., Cao, Y., Ngai, H., Moore, J. P. & Ho, D. D. (1996). Quantitative analysis of serum neutralization of human immunodeficiency virus type 1 from subtypes A, B, C, D, E, F, and I: lack of direct correlation between neutralization serotypes and genetic subtypes and evidence for prevalent serum-dependent infectivity enhancement. Journal of Virology 70, 445-458.[Abstract]
Kovacs, J. A., Baseler, M., Dewar, R. J., Vogel, S., Davey, R. T., Jr, Falloon, J., Polis, M. A., Walker, R. E., Stevens, R., Salzman, N. P. and others (1995). Increases in CD4 T lymphocytes with intermittent courses of interleukin-2 in patients with human immunodeficiency virus infection. A preliminary study. New England Journal of Medicine 332, 567575.
Lallemant, M., Baillou, A., Lallemant-Le Coeur, S., Nzingoula, S., Mampaka, M., MPelé, P., Barin, F. & Essex, M. (1994). Maternal antibody response at delivery and perinatal transmission of human immunodeficiency virus type 1 in African women. Lancet 343, 1001-1005.[Medline]
Lombardi, S., Garzelli, C., Pistello, M., Massi, C., Matteucci, D., Baldinotti, F., Cammarota, G., Da Prato, L., Bandecchi, P., Tozzini, F. & Bendinelli, M. (1994). A neutralizing antibody-inducing peptide of the V3 domain of feline immunodeficiency virus envelope glycoprotein does not induce protective immunity. Journal of Virology 68, 8374-8379.[Abstract]
Montefiori, D. C., Lefkowitz, L. B.Jr, Keller, R. E., Holmberg, V., Sandstrom, E. & Phair, J. P. (1991). Absence of a clinical correlation for complement-mediated, infection-enhancing antibodies in plasma or sera from HIV-1-infected individuals. Multicenter AIDS Cohort Study Group. AIDS 5, 513-517.[Medline]
Montefiori, D. C., Pantaleo, G., Fink, L. M., Zhou, J. T., Zhou, J. Y., Bilska, M., Miralles, G. D. & Fauci, A. S. (1996). Neutralizing and infection-enhancing antibody responses to human immunodeficiency virus type 1 in long-term nonprogressors. Journal of Infectious Diseases 173, 60-67.[Medline]
Moraillon, A., Barre-Sinoussi, F., Parodi, A., Moraillon, R. & Dauguet, C. (1992). In vitro properties and experimental pathogenic effect of three feline immunodeficiency viruses (FIV) isolated from cats with terminal disease. Veterinary Microbiology 31, 41-54.[Medline]
OBrien, W. A., Grovit-Ferbas, K., Namazi, A., Ovcak-Derzic, S., Wang, H.-J., Park, J., Yeramian, C., Mao, S.-H. & Zack, J. A. (1995). Human immunodeficiency virus-type 1 replication can be increased in peripheral blood of seropositive patients after influenza vaccination. Blood 86, 1082-1089.
Ortigao-de-Sampaio, M. B., Shattock, R. J., Hayes, P., Griffin, G. E., Linhares-de-Carvalho, M. I., Ponce de Leon, A., Lewis, D. J. & Castello-Branco, L. R. (1998). Increase in plasma viral load after oral cholera immunization of HIV-infected subjects. AIDS 12, F145-F150.[Medline]
Ostrowski, M. A., Krakauer, D. C., Li, Y., Justement, S. J., Learn, G., Ehler, L. A., Stanley, S. K., Nowak, M. & Fauci, A. S. (1998). Effect of immune activation on the dynamics of human immunodeficiency virus replication and on the distribution of viral quasispecies. Journal of Virology 72, 7772-7784.
Pancino, G., Leste-Lasserre, T., Burgard, M., Costagliola, D., Ivanoff, S., Blanche, S., Rouzioux, C. & Sonigo, P. (1998). Apparent enhancement of perinatal transmission of human immunodeficiency virus type 1 by high maternal anti-gp160 antibody titer. Journal of Infectious Diseases 177, 1737-1741.[Medline]
Pedersen, N. C., Ho, E. W., Brown, M. L. & Yamamoto, J. K. (1987). Isolation of T-lymphotropic virus from domestic cats with an immunodeficiency-like syndrome. Science 235, 790-793.[Medline]
Richardson, J., Moraillon, A., Baud, S., Cuisinier, A. M., Sonigo, P. & Pancino, G. (1997). Enhancement of feline immunodeficiency virus (FIV) infection after DNA vaccination with the FIV envelope. Journal of Virology 71, 9640-9649.[Abstract]
Richardson, J., Moraillon, A., Crespeau, F., Baud, S., Sonigo, P. & Pancino, G. (1998). Delayed infection after immunization with a peptide from the transmembrane glycoprotein of the feline immunodeficiency virus. Journal of Virology 72, 2406-2415.
Robinson, W. E.Jr, Montefiori, D. C. & Mitchell, W. M. (1987). A human immunodeficiency virus type 1 (HIV-1) infection-enhancing factor in seropositive sera. Biochemical and Biophysical Research Communications 149, 693-699.[Medline]
Robinson, W. E.Jr, Kawamura, T., Lake, D., Masuho, Y., Mitchell, W. M. & Hersh, E. M. (1990). Antibodies to the primary immunodominant domain of human immunodeficiency virus type 1 (HIV-1) glycoprotein gp41 enhance HIV-1 infection in vitro. Journal of Virology 64, 5301-5305.[Medline]
Robinson, W. E.Jr, Gorny, M. K., Xu, J. Y., Mitchell, W. M. & Zolla-Pazner, S. (1991). Two immunodominant domains of gp41 bind antibodies which enhance human immunodeficiency virus type 1 infection in vitro. Journal of Virology 65, 4169-4176.[Medline]
Schutten, M., Andeweg, A. C., Bosch, M. L. & Osterhaus, A. D. M. E. (1995). Enhancement of infectivity of a non-syncytium inducing HIV-1 by sCD4 and by human antibodies that neutralize syncytium inducing HIV-1. Scandinavian Journal of Immunology 41, 18-22.[Medline]
Schwartz, D. H. (1994). Potential pitfalls on the road to an effective HIV vaccine. Immunology Today 15, 54-57.[Medline]
Siebelink, K. H. J., Tijhaar, E., Huisman, R. C., Huisman, W., de Ronde, A., Darby, I. H., Francis, M. J., Rimmelzwaan, G. F. & Osterhaus, A. D. M. E. (1995). Enhancement of feline immunodeficiency virus infection after immunization with envelope glycoprotein subunit vaccines. Journal of Virology 69, 3704-3711.[Abstract]
Stanley, S., Ostrowski, M. A., Justement, J. S., Gantt, K., Hedayati, S., Mannix, M., Roche, K., Schwartzentruber, D. J., Fox, C. H. & Fauci, A. S. (1996). Effect of immunization with a common recall antigen on viral expression in patients infected with human immunodeficiency virus type 1. New England Journal of Medicine 334, 1222-1230.
Staprans, S. I., Hamilton, B. L., Follansbee, S. E., Elbeik, T., Barbosa, P., Grant, R. M. & Feinberg, M. B. (1995). Activation of virus replication after vaccination of HIV-1-infected individuals. Journal of Experimental Medicine 182, 1727-1737.[Abstract]
Sullivan, N., Sun, Y., Li, J., Hofmann, W. & Sodroski, J. (1995). Replicative function and neutralization sensitivity of envelope glycoproteins from primary and T-cell line-passaged human immunodeficiency virus type 1 isolates. Journal of Virology 69, 4413-4422.[Abstract]
Sullivan, N., Sun, Y., Binley, J., Lee, J., Barbas, C. F.III, Parren, P. W. H. I., Burton, D. R. & Sodroski, J. (1998). Determinants of human immunodeficiency virus type 1 envelope glycoprotein activation by soluble CD4 and monoclonal antibodies. Journal of Virology 72, 6332-6338.
Takeda, A., Tuazon, C. U. & Ennis, F. A. (1988). Antibody-enhanced infection by HIV-1 via Fc receptor-mediated entry. Science 242, 580-583.[Medline]
Talbott, R. L., Sparger, E. E., Lovelace, K. M., Fitch, W. M., Pedersen, N. C., Luciw, P. A. & Elder, J. H. (1989). Nucleotide sequence and genomic organization of feline immunodeficiency virus. Proceedings of the National Academy of Sciences, USA 86, 5743-5747.[Abstract]
Toth, F. D., Szabo, B., Ujhelyi, E., Paloczi, K., Horvath, A., Fust, G., Kiss, J., Banhegyi, D. & Hollan, S. R. (1991). Neutralizing and complement-dependent enhancing antibodies in different stages of HIV infection. AIDS 5, 263-268.[Medline]
Yamamoto, J. K., Ackley, C. D., Zochlinski, H., Louie, H., Pembroke, E., Torten, M., Hansen, H., Munn, R. & Okuda, T. (1991). Development of IL-2-independent feline lymphoid cell lines chronically infected with feline immunodeficiency virus: importance for diagnostic reagents and vaccines. Intervirology 32, 361-375.[Medline]
Zack, J. A., Arrigo, S. J., Weitsman, S. R., Go, A. S., Haislip, A. & Chen, I. S. Y. (1990). HIV-1 entry into quiescent primary lymphocytes: molecular analysis reveals a labile, latent viral structure. Cell 61, 213-222.[Medline]
Received 2 January 2002;
accepted 4 June 2002.