- Email: [email protected]
Fc receptors do not mediate african swine fever virus replication in macrophages
181,
VIROLOGY
756-759(1991)
Fc Receptors
Do Not Mediate
African
ANTONIO Centro
de Biologia
Molecular
(CSIC-UAM), Received
Facultad August
Swine Fever Virus Replication
ALCAMI
AND
de Ciencias, 8, 1990;
in Macrophages
ELADIO WJUELA’ Universidad
accepted
November
Aut&oma, 30,
Canto
Blanco,
28049
Madrid,
Spain
1990
Titration experiments in swine macrophages have shown that African swine fever virus infectivity was not enhanced in the presence of antiviral antibodies. The early viral protein synthesis and the viral DNA replication in swine macrophages infected with virus-antibody complexes were inhibited in the presence of high doses of uv-inactivated virus, which saturated specific virus receptors, but not when Fc receptors were saturated with antibodies. These results indicate that African swine fever virus does not infect swine macrophages through Fc receptors and that the normal entry pathway through virus receptors is not bypassed by the virus-antibody complexes. o 19% Academic press, I~C.
In certain virus-ceil interactions, the presence of antiviral antibodies increases infectivity, a phenomenon which is known as antibody-dependent enhancement of viral infectivity (for reviews, see Refs. (1-3)). This type of infection is not mediated by specific virus receptors, but by receptors for the Fc portion of immunoglobulins (FcR) that function as accessory virus receptors for antibody-coated virus particles. ASF virus, an enveloped icosahedral deoxyvirus (46) responsible for an important disease of swine (7)) replicates mainly in mononuclear phagocytes (8) and in a small fraction of polymorphonuclear leukocytes (9). The sera from pigs and other virus-resistant animal species inoculated with ASF virus do not neutralize the virus, although they contain virus-specific antibodies ( IO). We have reported that ASF virus attachment to specific receptors in susceptible Vero cells and swine macrophages is necessary for a productive infection ( 11, 12), suggesting that ASF virus particles could be neutralized with antibodies directed against the viral protein involved in the interaction with the virus receptor on the target cell. Since swine macrophages are the primary target of ASF virus, it is possible that the virus receptor is not the sole binding site for virus entry, since FcR might mediate the entry of virus-antibody complexes into macrophages. If this alternative mechanism of virus entry takes place in ASF virus infection of macrophages, it could play an important role in ASF virus pathology, since antiviral antibodies would facilitate virus infection instead of protecting the host. For
’ To whom addressed.
0042-6822/91
correspondence
and
$3.00
Copyright 0 1991 by Academic Press. Inc. All rtghts of reproduction I” any form resewed.
requests
for reprints
should
this reason, we have examined the infection of swine macrophages with virus-antibody complexes after attachment to either specific virus receptors or FcR. To study antibody-FcR interaction, we first tested culture conditions of swine alveolar macrophages, obtained by broncho-alveolar lavage from miniature pigs ( 13), according to Carrascosa et a/. ( 14). A rosette assay with rabbit antibody-coated swine erythrocytes ( 15) showed that 97.6% of rosetted macrophages were observed when cultured in Dulbecco’s modified Eagle medium supplemented with 10% heat-inactivated fetal calf serum (FCS), and this value decreased to 1.7% in the presence of 30% swine serum, the standard conditions. Since the FCS did not saturate FcR and did not affect virus production (not shown), the experiments described in this report were performed under these conditions. To investigate a possible antibody-mediated enhancement of ASF viral infectivity, different amounts of the ASF virus isolate BA7 1 ( 16) ( 1 02-1 0’ hemadsorption units) were preincubated with different concentrations of anti-ASF virus IgG (1 O-6-1O-12 pg/ml) for 1 hr at 4”, conditions that favor enhancement (2). The immunoglobulins were purified from a rabbit antiserum against the Vero-adapted virus strain BA7 1V ( 17) by affinity chromatography on protein A-Sepharose CL4B (Pharmacia) ( 18). The purified IgG had a titer against BA7 1V, determined by an ELISA test ( 19), of 7 X 1 O4 at a concentration of 1 mg/ml. After the incubation with IgG at 4”, the virus was titrated in a hemadsorption assay in swine macrophages (20), in the presence of the same quantities of IgG. No differences were found in the virus titer (not shown), suggesting
be
756
SHORT
COMMUNICATIONS
that ASF virus does not show, under these conditions, an enhancement of viral infectivity mediated by antibodies. To study in more detail whether virus-antibody complexes were able to initiate the infection after the interaction with the cell through FcR, we obtained preparations of opsonized virus particles. With this purpose, purified BA71V was incubated with rabbit IgG antiBA71V for 1 hr at 4’, at a concentration of 200 pg of IgG/pg of virus protein. Under these conditions most of virus particles were bound to antibodies, since 859/o of 3H-labeled virus particles were immunoprecipitated with Staphylococcus aureus, and virus titer decreased to about lo%, determined by a plaque assay in Vero cell monolayers after an overnight incubation at 37” with the antibodies (20)) conditions that favor neutralization of infectivity (2). To avoid the interaction of free antibodies with Fc receptors virus-antibody complexes were separated from unbound antibodies by centrifugation on 10 and 80% sucrose in phosphatebuffered saline for 30 min at 87,000 g. Swine macrophages were infected with antibodycoated BA7 1V, obtained as described, in the presence of either high doses of uv-inactivated BA71V, to saturate specific ASF virus receptors ( 12)) or IgG, to saturate FcR. Recently, we have reported that the specific, saturable binding of tritiated ASF virus to swine macrophages was competed by high doses of unlabeled virus, and viral protein and DNA syntheses were not detected in the presence of similar doses of uv-inactivated virus ( 12). Virus replication took place in the presence of inactivated virus when it was added at later times of infection, indicating that the inhibitory effect is not due to a cellular toxic effect of the inactivated virus but to the blocking of an early step of the infection, probably the interaction of virus particles with the specific cellular receptors. In the presence of the concentration of antibody or inactivated virus used, we observed 2.3 or 90.5% respectively, of rosetted macrophages in an assay with antibody-coated erythrocytes, indicating that IgG, but not inactivated virus, saturated FcR. Swine macrophages infected under the indicated conditions were then assayed for viral protein and DNA syntheses. The synthesis of the early viral protein p34 in swine macrophages infected with virus-antibody complexes was similar to that obtained in BA71V-infected cells (Fig. 1) In both cases, the saturation of specific virus receptors and not FcR, inhibited the early viral protein synthesis, indicating that the entry of virus-antibody complexes that led to a productive infection was not mediated directly by the attached anti-
757 MOCK -- ab
BA71V ---abc
100
BA71V-Ab ---abc
14.7 90.9
54.8
7.3
40.5
FIG. 1. Early viral protein synthesis in swine macrophages infected with antibody-opsonized virus particles. Swine macrophages were mock-infected or infected either with 6 PFU of purified BA71V per cell or with a similar amount of protein of opsonized virus particles (BA71V-Ab). The infection was carried out in the absence (a) or in the presence of either UV-BA7 1 V (3 X 10 5 virus particles per cell) (b) or 2.5 mg of nonimmune purified swine IgG (Cappel Laboratories) per milliliter (c). Cell monolayers were pulse-labeled with 500 pCi of [%]methionine (1200 Ci/mmol; Amersham, UK) per milliliter from 2 to 4 hr postadsorption and immunoprecipitated by anti-ASF virus-infected Vero cell rabbit antiserum, without removing the nuclei, as described (2 1). The electrophoretic analysis of the immunoprecipitated proteins and the quantitation of the viral protein ~34, performed as described (22), are shown. The percentages refer to the synthesis of p34 obtained in BA71Vinfected macrophages in the absence of competitor UV-BA7 1 V or IgG.
body. The results obtained in the analysis of the viral DNA replication were similar (Fig. 2) and confirmed that the opsonized virus particles used the same receptor as the virus to infect swine macrophages. The decrease of p34 synthesis and the delay of viral DNA replication when macrophages were infected with opsonized virus particles might reflect the partial neutralization effect of antibodies on viral infectivity, observed when assayed in Vero cell monolayers (see above). In other systems, it is not clear whether FcR mediate directly virus entry into macrophages or whether, by concentrating the virus on the cell surface, they increase infection through receptors for the virus (23). The entry of Dengue virus-antibody complexes takes place only through FcR (24,25). In the case of human immunodeficiencyvirus, antibody-dependentenhancement is mediated by FcR (26-28)) although it has been reported recently that CD4 interaction is also required (29). However, for Yellow fever virus (30) and influenza virus (31) it has been suggested that the complexes enter the cell also through the virus receptors. We show here that ASF virus-antibody complexes are able to infect the cell only through specific virus receptors, since the infection was inhibited in the presence of uv-inactivated virus. The results presented in this report indicate that there is not an enhancement of ASF virus infectivity mediated by antiviral antibodies. The effect of monoclo-
758
SHORT
COMMUNICATIONS
dent virus entry via FcR is not a mechanism that facilitates the progression of ASF virus infection. The involvement of complement and complement receptors in a similar enhancement mechanism, described for West Nile virus (35) and human immunodeficiency virus type 1 (37), could also play a role in ASF virus pathology. 9
12
15
6
9
12
15
ACKNOWLEDGMENTS
BAllV
+UV-BAHV
ii ‘3 d
We thank Dr. Angel L. Carrascosa for critical reading of the manuscript. This work was supported by grants from Comisidn Interministerial de Ciencia y Tecnologia, Junta de Extremadura, European Economic Community, and by an institutional grant of Fundation Ram&r Areces. I
I
1
a lS-
-
104
BA71V-Ab
_b
9--9 BA71V-Ab
I c
1
I
I
BA71V-Ab + IgG
+UV-BA7lV
REFERENCES
P 10 s-
A ?( 6
9
,
;
Q++v
12
15
6
J
9
12
1s
6
TIME AFTER ADSORPTION,
1 9
1 12
1. IS
h
FIG. 2. Viral DNA synthesis in swine macrophages infected with antibody-opsonized virus particles. Swine macrophages were mockinfected or infected either with purified BA7 1 V (6 pfu per cell) or with a similar amount of protein of antibody-coated virus particles (BA7lV-Ab). The infection was performed in the absence (a) or in the presence of either UV-BA7 1 V (3 X 1 O5 virus particles per cell) (b ) or 2.5 mg of nonimmune swine IgG per milliliter (c). The cells were pulse-labeled with 30 &i of [3H]thymidine (40-60 Ci/mmol; Amersham, UK) per milliliter for 2 hr at different times after adsorption. After the pulse period, macrophages were washed twice with phosphate-buffered saline and solubilized with 1% Nonidet P-40 in phosphate-buffered saline, and the radioactivity incorporated into trichloroacetic acid-insoluble material was determined.
nal antibodies and sera from different sources should be tested to support these observations. As far as we are aware, the failure to detect antibody-mediated enhancement of viral infectivity has been reported only for Mengo virus and herpes simplex virus (32). A negative result for these viruses was obtained in an antibodydependent plaque enhancement assay, but there are no reports of further investigations. Antibody-mediated enhancement of viral growth has been indicated as a factor in the pathogenesis of Dengue virus (33), rabies virus (34), human immunodeficiency virus type 1 (27), and Visna-maedi virus (35). The results presented here show that antibody-depen-
1. HALSTEAD, S. B. Prog. Allergy 31, 301-364 (1982). 2. PORTERFIELD, 1. S., and CARDOSA, M. 1. In “Concepts in Viral Pathogenesis” (A. L. Notkins and M. B.A. Oldfield, Eds.), pp. 117-l 22. Springer-Verlag, New York (1984). 3. PORTERFIELD, 1. S. Adv. Virus Res. 31, 335-355 (1986). 4. ALMEIDA, J. C., WATERSON, A. P., and PLOWRIGHT, W. Arch. Gesam. Virusforsch. 20, 392-396 (1967). 5. ENJUANES. L., CARRASCOSA, A. L., and WJUELA, E. 1. Gen. Viral. 32,479-492 (1976). 6. CARRASCOSA, J. L., CARAZO, J. M., CARRASCOSA, A. L., GARCIA, N., SANTISTEBAN, A., and VI~IJELA, E. virology 132, 160-172 (1984). 7. HESS, W. R. Adv. Vet. SC;. Comp. Med. 25, 39-69 (1981). 8. MAURER, F. D., GRIESEMER, R. A., and JONES, T. C. Amer. J. Vet. Res. 19, 517-539 (1958). 9. CABAL, I., ENJUANES, L., and WWELA. E. J. Viral. 52, 37-46 (1984). 10. DE BOER, C. J. Arch. Gesam. Virusforsch. 20, 164-l 79 (1967). 11. ALCAMI, A., CARRASCOSA, A. L., and VI~ELA, E. virology 168, 393-398 (1989). 12. ALCAM/, A., CARRASCOSA, A. L., and VI~ELA, E. virus Res. 17, 93-104 (1990). 13. SACHS, D. M., LEIGHT, G., CONE, J., SCHWARZ, S., STUART, L., and ROSENBERG, S. Transplantation 22, 559-567 (1976). 14. CARRASCOSA, A. L., SANTAR~N, J. F., and VIQJELA, E. 1. Viral. Methods 3, 303-3 10 ( 1982). 15. STEWART, J., GLASS, E. J., WEIR, D. M., and DAHA, M. R. In “Handbook of Experimental Immunology. Cellular Immunology” (D. M. Weir, Ed), Vol. 2, pp. 48.1-48.17. Blackwell Scientific Publications, London (1986). 16. GARCIA-BARRENO, B., SANZ, A., NOGAL, M. L., VI~JELA, E., and ENJUANES, L. J. Viol. 58, 385-392 (1986). 17. CAR~SCOSA, A. L., DEL VAL, M., SANTAR~N, 1. F., and VIRUELA, E.
J. Viral. 54, 337-344
( 1985).
KRONVALL, G., GREIG, H. M., and WILLIAMS, R. C., JR. J. Immunol. 105, 1116-1122(1970). 19. VOLLER, A., BIDWELL, D., and BARTLETT, A. ln “Manual of Clinical Immunology” (N. Rose and H. Friedman, Eds.), 2nd ed.. p. 359. Amer. Sot. Microbial., Washington D.C. (1980).
18.
SHORT 20. 21. 22. 23. 24. 25. 26. 27. 28.
COMMUNICATIONS
ENJUANES, L., CARRAXOSA, A. L., MORENO, M. A., and VI&JELA, E. 1. Gen. Viral. 32, 471-477 (1976). DELVAL, M., and VI~ELA, E. Virus Res. 7,297-30&l (1987). ALCAM{, A., CARRASCOSA, A. L., and VI~~UELA, E. Virology 171, 68-75 (1989). PEIRIS, J. S. M., GORDON, S., UNKELESS, J. C., and PORTERFIELD, J. S. Nature (London) 289, 189-191 (1981). DAUGHADAY, C. C., BFIANDT, W. E., MCCOWN, J. M., and RUSSELL, P. K. Infect. Immun. 32, 469-473 (1981). BRANDT, W. E., MCCOWN, J. M., GENTRY, M. K., and RUSSELL, P. K. Infect. Immun. 36, 1036-1041 (1982). HOMSY, J., TATENO. M., and LEW, J. A. Lancer 1, 1285-1286 (1988). TAKEDA, A., TUAZON, C. U., and ENNIS, F. A. Science 242, 580583(1988). HOMSY, J., MEYER, M., TATENO, M., CLARKSON, S., and LEW, 1. A.
Science244,1357-1360(1989).
29. 30. 31. 32. 33. 34. 35. 36. 37.
759 TAKEDA, A., SWEET. R. W., and ENNIS, F. A. J. Viral. 64, 56055619(1990). BRANDRISS. M. W., and SCHLESINGER, J. J. J. Gen. Viral. 65, 791794(1984). OCHIAI, H., KUROKAWA, M., HAYASHI, K., and NIMAYAMA, S. 1. Viral. 62, 20-26 (1988). PEIRIS, J. S. M., and PORTERFIELD, J. S. J. Gen. Viral. 57, 1 19-125 (1981). HALSTEAD, S. B. Science 239, 476-481 (1988). PRABHAKAR, B. S., and NATHANSON, N. Nature (London) 290, 590-591(1981). JOLLY, P. E., Huso, D. L.. SCHEFFER, D., and NARAYAN. 0.1. Viral, 63, 1811-1813 (1989). CARDOSA, M. J., PORTERFIELD, J. S., and GORDON S. 1. hp. Med. 158,258-263( 1983). ROBINSON, W. E., JR., MONTEFIORI, D. C., and MITCHELL, W. M. Lancet 1,790-794(1988).
VIROLOGY
756-759(1991)
Fc Receptors
Do Not Mediate
African
ANTONIO Centro
de Biologia
Molecular
(CSIC-UAM), Received
Facultad August
Swine Fever Virus Replication
ALCAMI
AND
de Ciencias, 8, 1990;
in Macrophages
ELADIO WJUELA’ Universidad
accepted
November
Aut&oma, 30,
Canto
Blanco,
28049
Madrid,
Spain
1990
Titration experiments in swine macrophages have shown that African swine fever virus infectivity was not enhanced in the presence of antiviral antibodies. The early viral protein synthesis and the viral DNA replication in swine macrophages infected with virus-antibody complexes were inhibited in the presence of high doses of uv-inactivated virus, which saturated specific virus receptors, but not when Fc receptors were saturated with antibodies. These results indicate that African swine fever virus does not infect swine macrophages through Fc receptors and that the normal entry pathway through virus receptors is not bypassed by the virus-antibody complexes. o 19% Academic press, I~C.
In certain virus-ceil interactions, the presence of antiviral antibodies increases infectivity, a phenomenon which is known as antibody-dependent enhancement of viral infectivity (for reviews, see Refs. (1-3)). This type of infection is not mediated by specific virus receptors, but by receptors for the Fc portion of immunoglobulins (FcR) that function as accessory virus receptors for antibody-coated virus particles. ASF virus, an enveloped icosahedral deoxyvirus (46) responsible for an important disease of swine (7)) replicates mainly in mononuclear phagocytes (8) and in a small fraction of polymorphonuclear leukocytes (9). The sera from pigs and other virus-resistant animal species inoculated with ASF virus do not neutralize the virus, although they contain virus-specific antibodies ( IO). We have reported that ASF virus attachment to specific receptors in susceptible Vero cells and swine macrophages is necessary for a productive infection ( 11, 12), suggesting that ASF virus particles could be neutralized with antibodies directed against the viral protein involved in the interaction with the virus receptor on the target cell. Since swine macrophages are the primary target of ASF virus, it is possible that the virus receptor is not the sole binding site for virus entry, since FcR might mediate the entry of virus-antibody complexes into macrophages. If this alternative mechanism of virus entry takes place in ASF virus infection of macrophages, it could play an important role in ASF virus pathology, since antiviral antibodies would facilitate virus infection instead of protecting the host. For
’ To whom addressed.
0042-6822/91
correspondence
and
$3.00
Copyright 0 1991 by Academic Press. Inc. All rtghts of reproduction I” any form resewed.
requests
for reprints
should
this reason, we have examined the infection of swine macrophages with virus-antibody complexes after attachment to either specific virus receptors or FcR. To study antibody-FcR interaction, we first tested culture conditions of swine alveolar macrophages, obtained by broncho-alveolar lavage from miniature pigs ( 13), according to Carrascosa et a/. ( 14). A rosette assay with rabbit antibody-coated swine erythrocytes ( 15) showed that 97.6% of rosetted macrophages were observed when cultured in Dulbecco’s modified Eagle medium supplemented with 10% heat-inactivated fetal calf serum (FCS), and this value decreased to 1.7% in the presence of 30% swine serum, the standard conditions. Since the FCS did not saturate FcR and did not affect virus production (not shown), the experiments described in this report were performed under these conditions. To investigate a possible antibody-mediated enhancement of ASF viral infectivity, different amounts of the ASF virus isolate BA7 1 ( 16) ( 1 02-1 0’ hemadsorption units) were preincubated with different concentrations of anti-ASF virus IgG (1 O-6-1O-12 pg/ml) for 1 hr at 4”, conditions that favor enhancement (2). The immunoglobulins were purified from a rabbit antiserum against the Vero-adapted virus strain BA7 1V ( 17) by affinity chromatography on protein A-Sepharose CL4B (Pharmacia) ( 18). The purified IgG had a titer against BA7 1V, determined by an ELISA test ( 19), of 7 X 1 O4 at a concentration of 1 mg/ml. After the incubation with IgG at 4”, the virus was titrated in a hemadsorption assay in swine macrophages (20), in the presence of the same quantities of IgG. No differences were found in the virus titer (not shown), suggesting
be
756
SHORT
COMMUNICATIONS
that ASF virus does not show, under these conditions, an enhancement of viral infectivity mediated by antibodies. To study in more detail whether virus-antibody complexes were able to initiate the infection after the interaction with the cell through FcR, we obtained preparations of opsonized virus particles. With this purpose, purified BA71V was incubated with rabbit IgG antiBA71V for 1 hr at 4’, at a concentration of 200 pg of IgG/pg of virus protein. Under these conditions most of virus particles were bound to antibodies, since 859/o of 3H-labeled virus particles were immunoprecipitated with Staphylococcus aureus, and virus titer decreased to about lo%, determined by a plaque assay in Vero cell monolayers after an overnight incubation at 37” with the antibodies (20)) conditions that favor neutralization of infectivity (2). To avoid the interaction of free antibodies with Fc receptors virus-antibody complexes were separated from unbound antibodies by centrifugation on 10 and 80% sucrose in phosphatebuffered saline for 30 min at 87,000 g. Swine macrophages were infected with antibodycoated BA7 1V, obtained as described, in the presence of either high doses of uv-inactivated BA71V, to saturate specific ASF virus receptors ( 12)) or IgG, to saturate FcR. Recently, we have reported that the specific, saturable binding of tritiated ASF virus to swine macrophages was competed by high doses of unlabeled virus, and viral protein and DNA syntheses were not detected in the presence of similar doses of uv-inactivated virus ( 12). Virus replication took place in the presence of inactivated virus when it was added at later times of infection, indicating that the inhibitory effect is not due to a cellular toxic effect of the inactivated virus but to the blocking of an early step of the infection, probably the interaction of virus particles with the specific cellular receptors. In the presence of the concentration of antibody or inactivated virus used, we observed 2.3 or 90.5% respectively, of rosetted macrophages in an assay with antibody-coated erythrocytes, indicating that IgG, but not inactivated virus, saturated FcR. Swine macrophages infected under the indicated conditions were then assayed for viral protein and DNA syntheses. The synthesis of the early viral protein p34 in swine macrophages infected with virus-antibody complexes was similar to that obtained in BA71V-infected cells (Fig. 1) In both cases, the saturation of specific virus receptors and not FcR, inhibited the early viral protein synthesis, indicating that the entry of virus-antibody complexes that led to a productive infection was not mediated directly by the attached anti-
757 MOCK -- ab
BA71V ---abc
100
BA71V-Ab ---abc
14.7 90.9
54.8
7.3
40.5
FIG. 1. Early viral protein synthesis in swine macrophages infected with antibody-opsonized virus particles. Swine macrophages were mock-infected or infected either with 6 PFU of purified BA71V per cell or with a similar amount of protein of opsonized virus particles (BA71V-Ab). The infection was carried out in the absence (a) or in the presence of either UV-BA7 1 V (3 X 10 5 virus particles per cell) (b) or 2.5 mg of nonimmune purified swine IgG (Cappel Laboratories) per milliliter (c). Cell monolayers were pulse-labeled with 500 pCi of [%]methionine (1200 Ci/mmol; Amersham, UK) per milliliter from 2 to 4 hr postadsorption and immunoprecipitated by anti-ASF virus-infected Vero cell rabbit antiserum, without removing the nuclei, as described (2 1). The electrophoretic analysis of the immunoprecipitated proteins and the quantitation of the viral protein ~34, performed as described (22), are shown. The percentages refer to the synthesis of p34 obtained in BA71Vinfected macrophages in the absence of competitor UV-BA7 1 V or IgG.
body. The results obtained in the analysis of the viral DNA replication were similar (Fig. 2) and confirmed that the opsonized virus particles used the same receptor as the virus to infect swine macrophages. The decrease of p34 synthesis and the delay of viral DNA replication when macrophages were infected with opsonized virus particles might reflect the partial neutralization effect of antibodies on viral infectivity, observed when assayed in Vero cell monolayers (see above). In other systems, it is not clear whether FcR mediate directly virus entry into macrophages or whether, by concentrating the virus on the cell surface, they increase infection through receptors for the virus (23). The entry of Dengue virus-antibody complexes takes place only through FcR (24,25). In the case of human immunodeficiencyvirus, antibody-dependentenhancement is mediated by FcR (26-28)) although it has been reported recently that CD4 interaction is also required (29). However, for Yellow fever virus (30) and influenza virus (31) it has been suggested that the complexes enter the cell also through the virus receptors. We show here that ASF virus-antibody complexes are able to infect the cell only through specific virus receptors, since the infection was inhibited in the presence of uv-inactivated virus. The results presented in this report indicate that there is not an enhancement of ASF virus infectivity mediated by antiviral antibodies. The effect of monoclo-
758
SHORT
COMMUNICATIONS
dent virus entry via FcR is not a mechanism that facilitates the progression of ASF virus infection. The involvement of complement and complement receptors in a similar enhancement mechanism, described for West Nile virus (35) and human immunodeficiency virus type 1 (37), could also play a role in ASF virus pathology. 9
12
15
6
9
12
15
ACKNOWLEDGMENTS
BAllV
+UV-BAHV
ii ‘3 d
We thank Dr. Angel L. Carrascosa for critical reading of the manuscript. This work was supported by grants from Comisidn Interministerial de Ciencia y Tecnologia, Junta de Extremadura, European Economic Community, and by an institutional grant of Fundation Ram&r Areces. I
I
1
a lS-
-
104
BA71V-Ab
_b
9--9 BA71V-Ab
I c
1
I
I
BA71V-Ab + IgG
+UV-BA7lV
REFERENCES
P 10 s-
A ?( 6
9
,
;
Q++v
12
15
6
J
9
12
1s
6
TIME AFTER ADSORPTION,
1 9
1 12
1. IS
h
FIG. 2. Viral DNA synthesis in swine macrophages infected with antibody-opsonized virus particles. Swine macrophages were mockinfected or infected either with purified BA7 1 V (6 pfu per cell) or with a similar amount of protein of antibody-coated virus particles (BA7lV-Ab). The infection was performed in the absence (a) or in the presence of either UV-BA7 1 V (3 X 1 O5 virus particles per cell) (b ) or 2.5 mg of nonimmune swine IgG per milliliter (c). The cells were pulse-labeled with 30 &i of [3H]thymidine (40-60 Ci/mmol; Amersham, UK) per milliliter for 2 hr at different times after adsorption. After the pulse period, macrophages were washed twice with phosphate-buffered saline and solubilized with 1% Nonidet P-40 in phosphate-buffered saline, and the radioactivity incorporated into trichloroacetic acid-insoluble material was determined.
nal antibodies and sera from different sources should be tested to support these observations. As far as we are aware, the failure to detect antibody-mediated enhancement of viral infectivity has been reported only for Mengo virus and herpes simplex virus (32). A negative result for these viruses was obtained in an antibodydependent plaque enhancement assay, but there are no reports of further investigations. Antibody-mediated enhancement of viral growth has been indicated as a factor in the pathogenesis of Dengue virus (33), rabies virus (34), human immunodeficiency virus type 1 (27), and Visna-maedi virus (35). The results presented here show that antibody-depen-
1. HALSTEAD, S. B. Prog. Allergy 31, 301-364 (1982). 2. PORTERFIELD, 1. S., and CARDOSA, M. 1. In “Concepts in Viral Pathogenesis” (A. L. Notkins and M. B.A. Oldfield, Eds.), pp. 117-l 22. Springer-Verlag, New York (1984). 3. PORTERFIELD, 1. S. Adv. Virus Res. 31, 335-355 (1986). 4. ALMEIDA, J. C., WATERSON, A. P., and PLOWRIGHT, W. Arch. Gesam. Virusforsch. 20, 392-396 (1967). 5. ENJUANES. L., CARRASCOSA, A. L., and WJUELA, E. 1. Gen. Viral. 32,479-492 (1976). 6. CARRASCOSA, J. L., CARAZO, J. M., CARRASCOSA, A. L., GARCIA, N., SANTISTEBAN, A., and VI~IJELA, E. virology 132, 160-172 (1984). 7. HESS, W. R. Adv. Vet. SC;. Comp. Med. 25, 39-69 (1981). 8. MAURER, F. D., GRIESEMER, R. A., and JONES, T. C. Amer. J. Vet. Res. 19, 517-539 (1958). 9. CABAL, I., ENJUANES, L., and WWELA. E. J. Viral. 52, 37-46 (1984). 10. DE BOER, C. J. Arch. Gesam. Virusforsch. 20, 164-l 79 (1967). 11. ALCAMI, A., CARRASCOSA, A. L., and VI~ELA, E. virology 168, 393-398 (1989). 12. ALCAM/, A., CARRASCOSA, A. L., and VI~ELA, E. virus Res. 17, 93-104 (1990). 13. SACHS, D. M., LEIGHT, G., CONE, J., SCHWARZ, S., STUART, L., and ROSENBERG, S. Transplantation 22, 559-567 (1976). 14. CARRASCOSA, A. L., SANTAR~N, J. F., and VIQJELA, E. 1. Viral. Methods 3, 303-3 10 ( 1982). 15. STEWART, J., GLASS, E. J., WEIR, D. M., and DAHA, M. R. In “Handbook of Experimental Immunology. Cellular Immunology” (D. M. Weir, Ed), Vol. 2, pp. 48.1-48.17. Blackwell Scientific Publications, London (1986). 16. GARCIA-BARRENO, B., SANZ, A., NOGAL, M. L., VI~JELA, E., and ENJUANES, L. J. Viol. 58, 385-392 (1986). 17. CAR~SCOSA, A. L., DEL VAL, M., SANTAR~N, 1. F., and VIRUELA, E.
J. Viral. 54, 337-344
( 1985).
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