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Antigenic variants of rabies virus
Comp Immun Mi~rahiol. inli'(t. Di~. Vol. 5. Nos. I 3, pp. 101 107, 1982. Printed in Great Britain
0147 9571 <82/0101(]1 07 $03 00,'0 ( 1982 Pcrgamon Press Ltd.
A N T I G E N I C V A R I A N T S OF RABIES VIRUS LOTHAR G . SCHNEIDER Federal Research Institute for Animal Virus Diseases, P.O. Box 1149, D-7400 Tuebingen, Federal Republic of Germany Monoclonal antibodies directed either against the nucleocapsid or the glycoprotein of rabies virus were reacted by indirect immunofluorescence and mouse neutralization tests with vaccine strains and rabies field isolates of various origin. Major antigenic determinants allow the grouping of viruses and minor ones the differentiation of virus strains. A protection study showed the existence in nature of certain antigenic variants against which conventional vaccines do not fully protect. The isolation of two such variants from "imported' bats in the Federal Republic of Germany is reported here for the first time. Key words." Monoclonal antibodies, nucleocapsid, glycoprotein, rabies virus, vaccine strains, rabies field isolates, grouping of viruses, protection study, antigenic virus variants Abstract
VARIANTS
ANTIGENIQUES
DU VIRUS
RABIQUE
Resum~Les anticorps monoclonaux dirig~s soit contre la nucl6ocapside soit contre la glycoprot6ine du virus rabique ont reagi avec les virus rabiques de diff6rentes provenances au cours d'une epreuve indirecte d'immunofluorescence et d'une epreuve de neutralisation sur souris. Les d&erminants antigeniques majeurs permettent de grouper les virus, les d6terminants mineurs permettant la diff6rentiation des souches de virus. L'epreuve de protection crois(~e a prouv+ l'existence sur le terrain de quelques variants abtigeniques du virus rabique contre lesquels les vaccins conventionnels ne prot6gent pas compl&ement. C'est la premi6re fois qu'est rapport6e en Republique Fbd6rale Allemagne l'existence de tels variants isol6s de chauve-souris "importees'. Mots-cle/Z~': Anticorps monoclonaux, nucl~ocapside, glycoprot~ine, virus rabique, souches vaccinales, souches sauvages, groupement des virus, protection, varients antig6niques
INTRODUCTION A n t i r a b i e s m o n o c l o n a l a n t i b o d i e s w e r e u s e d to s t u d y the d e g r e e o f a n t i g e n i c v a r i a t i o n a m o n g v a c c i n e strains a n d field isolates o f rabies virus. O f p r i m a r y i n t e r e s t was the e x i s t e n c e o f v a r i a n t s a g a i n s t w h i c h c o n v e n t i o n a l rabies v a c c i n e m a y n o t p r o t e c t .
MATERIALS AND METHODS M o n o c l o n a l n u c l e o c a p s i d ( N C ) a n d g l y c o p r o t e i n (G) a n t i b o d i e s w e r e p r e p a r e d , c l o n e d a n d selected as d e s c r i b e d b e f o r e [1, 2]. A p a n e l o f t w e n t y N C a n t i b o d y t y p e s was o b t a i n e d f r o m T. J. W i k t o r [3]. R a b i e s v i r u s isolates u s e d w e r e f r o m the i n s t i t u t e ' s s t r a i n c o l l e c t i o n , o r o b t a i n e d f r o m r a b i e s d i a g n o s t i c o r r e s e a r c h facilities t h r o u g h o u t the w o r l d . 101
102
LOTHARG. SCHNEIDER Table 1. Geographic distribution of rabies virus strains (1)
(2)
Europe
31
(3)
-
CSSR
2
North
-
England
2
-
-
France
1 16"
--
10
Switzerland
Asia
America
USA
Latin
- Germany (FRG)
15
Americas
10 America
-
Mexico
3
-
Chile
1
-
Argentina
1
4
--
Pakistan
1
-
Turkey
1
--
USSR
2
(4)
(5)
Totals:
--
Botswana
-
Nigeria
-
South
Laboratory
35
Total:
81
20
Africa 4 2 Africa
strains
14
11
46
Virus
strains
*Two bat strains of African origin included. RESULTS
Origin ~)[" virus isolates The geographic distribution of the virus isolates (Table 1) is worldwide. Thirty-one strains were obtained from Europe, four from Asia, fifteen from the Americas, twenty from the African continent, and eleven were attenuated laboratory and vaccine strains. In addition to the eleven laboratory strains, ten virus isolates were from human rabies deaths, and fourteen originated from domestic animals (8 dogs, 1 cat, 5 cattle), (Table 2). A total of forty-six viruses were derived from wildlife animals (16 red foxes, 2 polar foxes, 1 bobcat, 4 jackals, 1 raccoon, 2 mongooses, 2 antilopes, 8 rodents, 2 insectivores, and 8 bats).
Nucleocapsid reaction patterns The nucleocapsid reaction patterns of eighty-one rabies virus strains were determined by reacting the viruses in indirect immunofluorescent tests with twenty-five selected monoclonal nucleocapsid antibody types. The nucleocapsid reaction patterns of the individual viruses allowed all strains tested to be arranged in either one of ten nucleocapsid reaction groups (Table 3). The viruses of NC-groups 1 6 originated from Europe, Asia and the Americas, those
Antigenic variants of rabies virus
103
Table 2. Origin of rabies virus strains according to animal species (1)
(2)
(3}
Domestic -
canine
-
Animals
14
(4)
Wild
Animals
46
8
-
Fox,
red
feline
1
-
Fox,
polar
bovine
5
-
Bobcat
1
-
Jackal
4
Racoon
1
-
Mongoose
2
-
Antilopes
2
-
Rodents
8
-
Insectivores
2
-
Bats
8
Humans
Laboratory
10
Strains
11
Totals:
35
Total:
81
16 2
46
Virus
strains
from groups 7 10 from Africa. Some of the groups were tentatively named according to the animal species most frequently represented therein. For instance, all eighteen red and polar fox origin viruses could be placed into either group 3 or 4. All attenuated rabies vaccine viruses except the Flury HEP strains are found in group 1, which in addition contains field isolates from three humans, two dogs, two rodents, and one mongoose. The viruses placed into groups 1-4 represented almost two-thirds of all strains tested. Most characteristic for the viruses obtained from the African continent is the reduced degree of cross-reactivity with the monclonal NC antibody panel. This feature is most prominent in viruses of group 7, which almost exclusively contains virus isolates from African bats. Of the seven viruses shown only two (SA-2, SA-3) gave identical reactions, indicating a common--possibly geographic--heterogeneity among rabies viruses of African bats.
Glycoprotein reaction patterns and protective activio' Rabies viruses assayed in mouse neutralization tests [4] against a panel of eighteen selected monoclonal G antibody types were grouped according to their individual reaction patterns (Table 4). In contrast to the viruses of groups A-E, those from groups F and G (horizontal panels of Table 4) are characterized by a significant reduction of their cross-reactivity with the panel of virus-neutralizing monoclonal antibodies. A correclation with NC-reaction groups could not be shown since one virus each was derived from NC-groups 1, 4 and 8, three from group 2 and five from the African group 7. The African viruses Lagos Bat (LBV), Mokola and Duvenhage (DUV) were shown previously [5] to be distinct serotypes within the genus rabies virus and are known to give
104
LOTHAR G. SCHNEIDER Table 3. Monoclonal rabies nucleocapsid antibodies W 1
O
NC R e a c t i o n
W 2
W 3
r-.
~
~
~
1
Type
2
Wild type
None
3
Wild l y p e
Fox I
•
4
Wild
type
F o x II
~E)
s
Flory
HEP
6
Wild
type H u m a n
USA
7
W,ld
type
Bat
African
AI .....
Shre .
.
.
DEU'70
Bat
Bat
76 Bat SA2 Bat
SA3
8
W.dtyp~
9
W . d t y p e Af . . . . 2
10
Wild
type
~
Af,+~a
Afr=ca
r-, N
(~
B~t
7 5 Hamburg
77
"-
Bat
Duvenhage
7 4 Stade
(''
"
(~ (~ E)
+ •
7 2 Mokola 7 3
r", ~
V,rus
tago~
~
3
®
~ ~,~
7~
Tu 6
Type
+a
Fixed
W5
W 4
DEU68
•
*
*
- • •
0000
• + + •
• • • • 0 0 0 0
+
+ •
•
•
•
•
+
+
*
+ •
•
•
•
•
+
*
+ + * • + + * •
• •
• 00 000
• •
* • • + 00
* + +
+
.
*
+
@
@
•
•
•
•
+
.
•
"
*
*
•
,
• •
+
~
÷ •
•
•
* •
+
N m ~
~) @ (~ @
•
~
=
f--
+
*
~
14
+
+ •
11
•
•
6
+ •
•
20
•
@ @ •
,-, N
+
•
•
•
,
•
•
•
@ @ @ (~
•
.
*
•
4
•
4
+ @ ~) •
•
7
00•
00
000
+ • • • 0 0 0 0
00
• • 00
• •
+ •
•
*
•
•
•
+ •
•
+ •
•
• • • + ÷ * ÷ •
•
+ @
•
•
+ •
*
* 000 + 00•
•0 • 000
"
+ •
•
"
•
•
•
.
•
•
•
+
•
•
+ •
* •
•
•
+ •
+ •
•
•
*
+
+
+
÷ 00 + 000
•
+
'
•
*
* • •
+ •
*
+ •
+ •
*
z •
•
7
Tolal
81
Nucleocapsid reaction groups as obtained from reacting 81 virus strains with 25 types of antibody. + , Positive reaction; O, no reaction.
only poor or no protection at all against a challenge infection by classical rabies viruses. We therefore selected four viruses from group F (rodent CS46, polar fox Alo 280, and two bat isolates Stade and Hamburg (SVF) from the Federal Republic of Germany) which were titrated intracerebrally in immunized mice and in non-vaccinated controls. The vaccine was prepared in suckling mice [6] with the Pitman-Moore strain of rabies virus which is widely used for the vaccine production and rabies prophylaxis in man. The results of the protection study are shown in Fig. I. Complete protection of vaccinated mice was obtained against the challenge infection with the Pitman -Moore virus (PM), the rodent isolate W239 and the polar fox virus ALO 280. The rodent strain CS 46 showed some breakthrough at virus dilutions - 1 and - 2 . Vaccinated mice exposed to Hamburg (SVF) and Stade bat viruses showed practically no protection. DISCUSSION The present study clearly indicates the worldwide existence of minor and major antigenic variants of rabies virus. As measured by monoclonal nucleocapsid antibodies, old and new
Antigenic variants of rabies virus
105
Table 4. Monoclonal rabies glycoprotein antibodies
W
00
• 0 0 • 0 0 . . 0 0 ÷ ÷ 0 0 0
00
÷ * 0 0 0 0 0
184
. . . .
W 187
. . . .
W
246
.
W
229
K 41
.
.
239
0 ÷ ÷ + 0 0 0 0 0
W
876
W
238
0 ÷ ÷ ÷ 0 0 0 0 00 ÷ ÷ 0 0 0 0
W
218
.
.
.
•
.
00
W
56
W 131
.
.
.
÷0
0 0 ÷ ÷ 0 0 0 ÷ * 0 0
÷
l l
S 91
•
+
l .
.
.
svM1
00
KELEV
• ÷ • • • •
+
Moko,~
LBv
.
.
.
l
.
.
.
.
.
.
.
.
i
.
.
l l
.
.
l
÷
•
•
• •
0 0 0 0 0 0 0 0
00
O ÷ +00 00 ÷ 00
0 0 0 0 0 0 0 0
.
,
s
e
789
•
÷
•
÷
÷
•
÷
+ +
.
+ ÷
•
÷
÷ 0 0
+
÷
.
0 .
+
÷
.
0 ÷
+
÷
+
+
÷
+
.
.
÷
. .
.
.
.
.
. ,
÷ i l l
.
0 0 ÷
÷ l ÷ . .
. .
0 0
÷ i t .
I
•
.
÷00
÷ ÷
÷
÷ ÷0 + ÷00 ÷ . . . . 000÷ ÷ ÷ 0 0 0 0 0 ÷ + 0 0 0 0 ÷ ÷ 0 0 0 0 ÷ 00 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
0 0 0 0 0 0 0 • 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 • 0 0 0 0 0 0 0 ~ 23
+
+
÷
+ ÷00
0 0 ÷ 0 0 0 • 0 0 0 • 0 0 0 0 0 0 • 0 0 0 0 0 0 0 • 0 0 0
÷
÷
+
+
. .
÷
•
• •
÷
.
÷ +
•
.
.
+ +
.
I
.
÷
. . . . . . . .
.
.
+
+
•
. .
.
.
+
+
• •
.
.
÷
+
.
.
.
.
•
.
+ ÷
+
00 O ÷ O
.
.
. . . .
.
÷
.
+ +
00
.
.
.
•
. 0 0 0 . ÷ ÷ 0 0 0 • ' 0
l
.
.
.
.
.
l
.
. . . . . .
÷
+
00
237
. . . . .
.
÷
svF
B. . . . .
.
0 0 0 ÷ 0
ALO2eO
DUV
.
.
.
.
÷ I
0 0 0 ÷ • • • + • • • • ÷ • •
STADE
.
l
+
E
1
.
t
.
.
.
÷ l
Apip,
.
. . . .
. . . .
ERA
cs*e
.
00 00 00 00
. . . . . . . .
.
.
.
0 ÷ . ÷ 0
CVS 11
8obc,,
.
*
LEP
z
.
. . . . . . . . . . . . . . . ÷ ÷ • ÷ • . . . . .
Pitm Moore
svM
.
.
÷0 I I •
HEP
.
.
. . . . . . .
.
.
.
.
0+
BF 297
CVS 24
.
.
.
.
0 0 •
W
.
.
.
. . . . . .
t . ÷ ÷ l l l t ,
37
.
.
.
Nig H o r s e
156
.
÷ 00
W
MIC
G
0 0 0 0
~o.12131415161718
+
+
+
+
+
+
+
÷
÷
÷
+
÷
÷
÷
+
÷
+
+
+
*
÷
+
•
÷
•. ÷
+
•÷ 19
20
Neutralization patterns of 35 virus strains with 18 types of virus neutralizing antibodies. +, Virus neutralized; O, virus not neutralized.
106
LOTHAR G. SCHNEIDER 100%
100 %
50 %
50%
,
,
,
0
-2
0
-2
,
,
-4
,
,
-6
o
-2
-4
-6
-4
-6
100 %
50%
100%
i 00! -4
-6
0
-2
" 50 %
0
-2
-t,
-6
0
-2
,DE
-4
'
-'6
Fig. I. Protective activity of Pitman Moore (PM) suckling mouse brain vaccine. Per cent mortality of nonvaccinated ( O - - O ) versus that of vaccinated mice (© O) after challenge with homologous (PM) and hetorologous viruses (rodent viruses W 239 and CS 46); polar fox virus ALO 280: bat viruses Hamburg (SVF) and Stade.
world rabies viruses as well as those from Asia may differ slightly but significantly from each other. Substantial differences of major NC-determinants exist however between the latter viruses and those obtained from the African continent. All African viruses tested so far have at least one major determinant in common. In fact, two rabies viruses isolated from unidentified bats found in the Northern part of the Federal Republic of Germany were retrospectively proven for the first time to be true African viruses. When evaluated by monoclonal virus neutralizing antibodies, rabies strains can be grouped into similar reaction pattern groups. From NC- and glycoprotein reaction groups, viruses can be selected which by subsequent analysis of their protective activity may be classified as true variants of the rabies virus or as newly recognized serotypes against which conventional vaccines may not fully protect. This study further indicates that a panel of nucleocapsid and neutralizing monoclonal antibodies representing major antigenic determinants may selectively identify such variants of field virus strains, thus being extremely helpful in epidemiological investigations. Acknowledgements--The technical assistance and excellent performance of Mrs. Susanne Meyer and Mrs. Dagmar Lindner in cloning and testing of monoclonal antibodies is gratefully acknowledged.
Antigenic variants of rabies virus
107
REFERENCES 1. Schneider, L. G., Antigenic determinants of rabies vaccine, Proceedings ~tf Cell Culture Rabies Vaccines and their Protective Effect in Man, Essen, Germany, March 1980, in press. 2. Schneider, L, G. and Meyer, S., Antigenic determinants of rabies virus as demonstrated by monoclonal antibody, Proceedings ¢~[the 4th lnternation~d Symposia ¢~fNegative Strand Viruses. Elsevier/North Holland, in press. 3. Wiktor, T. J. and Koprowski, H., Monclonal antibodies against rabies virus produced by somatic cell hybridization. Detection of antigenic variants, Proc. ham. Acad. Sci. U.S.A. 75, 3938 3942 (1978). 4. Atanasiu, P., Quantitative assay and potency test of antirabies serum and immunoglobulin, in Laboratory Techniques in Rabies (Eds. Kaplan, M. M. and Koprowski, H.), pp. 314~318. World Health Organization. Geneva (1973). 5. Schneider, L. G., Dietzaschold, B., Dierks, R. E., Matthaeus, W., Enzmann, P. J. and Strohmaier, K., Rabies group-specific ribonucleoprotein antigen and a test system for grouping and typing of rhabdoviruses, J. Virol. II, 748 755 (1973). 6. Fuenzalida, E., Suckling mouse brain vaccine, in Laboratory Techniques in Rabies (Eds. Kaplan, M. M. and Koprowski, H.), pp. 216 220. World Health Organization, Geneva (1973).
CtMIO
5~1-3
- H
0147 9571 <82/0101(]1 07 $03 00,'0 ( 1982 Pcrgamon Press Ltd.
A N T I G E N I C V A R I A N T S OF RABIES VIRUS LOTHAR G . SCHNEIDER Federal Research Institute for Animal Virus Diseases, P.O. Box 1149, D-7400 Tuebingen, Federal Republic of Germany Monoclonal antibodies directed either against the nucleocapsid or the glycoprotein of rabies virus were reacted by indirect immunofluorescence and mouse neutralization tests with vaccine strains and rabies field isolates of various origin. Major antigenic determinants allow the grouping of viruses and minor ones the differentiation of virus strains. A protection study showed the existence in nature of certain antigenic variants against which conventional vaccines do not fully protect. The isolation of two such variants from "imported' bats in the Federal Republic of Germany is reported here for the first time. Key words." Monoclonal antibodies, nucleocapsid, glycoprotein, rabies virus, vaccine strains, rabies field isolates, grouping of viruses, protection study, antigenic virus variants Abstract
VARIANTS
ANTIGENIQUES
DU VIRUS
RABIQUE
Resum~Les anticorps monoclonaux dirig~s soit contre la nucl6ocapside soit contre la glycoprot6ine du virus rabique ont reagi avec les virus rabiques de diff6rentes provenances au cours d'une epreuve indirecte d'immunofluorescence et d'une epreuve de neutralisation sur souris. Les d&erminants antigeniques majeurs permettent de grouper les virus, les d6terminants mineurs permettant la diff6rentiation des souches de virus. L'epreuve de protection crois(~e a prouv+ l'existence sur le terrain de quelques variants abtigeniques du virus rabique contre lesquels les vaccins conventionnels ne prot6gent pas compl&ement. C'est la premi6re fois qu'est rapport6e en Republique Fbd6rale Allemagne l'existence de tels variants isol6s de chauve-souris "importees'. Mots-cle/Z~': Anticorps monoclonaux, nucl~ocapside, glycoprot~ine, virus rabique, souches vaccinales, souches sauvages, groupement des virus, protection, varients antig6niques
INTRODUCTION A n t i r a b i e s m o n o c l o n a l a n t i b o d i e s w e r e u s e d to s t u d y the d e g r e e o f a n t i g e n i c v a r i a t i o n a m o n g v a c c i n e strains a n d field isolates o f rabies virus. O f p r i m a r y i n t e r e s t was the e x i s t e n c e o f v a r i a n t s a g a i n s t w h i c h c o n v e n t i o n a l rabies v a c c i n e m a y n o t p r o t e c t .
MATERIALS AND METHODS M o n o c l o n a l n u c l e o c a p s i d ( N C ) a n d g l y c o p r o t e i n (G) a n t i b o d i e s w e r e p r e p a r e d , c l o n e d a n d selected as d e s c r i b e d b e f o r e [1, 2]. A p a n e l o f t w e n t y N C a n t i b o d y t y p e s was o b t a i n e d f r o m T. J. W i k t o r [3]. R a b i e s v i r u s isolates u s e d w e r e f r o m the i n s t i t u t e ' s s t r a i n c o l l e c t i o n , o r o b t a i n e d f r o m r a b i e s d i a g n o s t i c o r r e s e a r c h facilities t h r o u g h o u t the w o r l d . 101
102
LOTHARG. SCHNEIDER Table 1. Geographic distribution of rabies virus strains (1)
(2)
Europe
31
(3)
-
CSSR
2
North
-
England
2
-
-
France
1 16"
--
10
Switzerland
Asia
America
USA
Latin
- Germany (FRG)
15
Americas
10 America
-
Mexico
3
-
Chile
1
-
Argentina
1
4
--
Pakistan
1
-
Turkey
1
--
USSR
2
(4)
(5)
Totals:
--
Botswana
-
Nigeria
-
South
Laboratory
35
Total:
81
20
Africa 4 2 Africa
strains
14
11
46
Virus
strains
*Two bat strains of African origin included. RESULTS
Origin ~)[" virus isolates The geographic distribution of the virus isolates (Table 1) is worldwide. Thirty-one strains were obtained from Europe, four from Asia, fifteen from the Americas, twenty from the African continent, and eleven were attenuated laboratory and vaccine strains. In addition to the eleven laboratory strains, ten virus isolates were from human rabies deaths, and fourteen originated from domestic animals (8 dogs, 1 cat, 5 cattle), (Table 2). A total of forty-six viruses were derived from wildlife animals (16 red foxes, 2 polar foxes, 1 bobcat, 4 jackals, 1 raccoon, 2 mongooses, 2 antilopes, 8 rodents, 2 insectivores, and 8 bats).
Nucleocapsid reaction patterns The nucleocapsid reaction patterns of eighty-one rabies virus strains were determined by reacting the viruses in indirect immunofluorescent tests with twenty-five selected monoclonal nucleocapsid antibody types. The nucleocapsid reaction patterns of the individual viruses allowed all strains tested to be arranged in either one of ten nucleocapsid reaction groups (Table 3). The viruses of NC-groups 1 6 originated from Europe, Asia and the Americas, those
Antigenic variants of rabies virus
103
Table 2. Origin of rabies virus strains according to animal species (1)
(2)
(3}
Domestic -
canine
-
Animals
14
(4)
Wild
Animals
46
8
-
Fox,
red
feline
1
-
Fox,
polar
bovine
5
-
Bobcat
1
-
Jackal
4
Racoon
1
-
Mongoose
2
-
Antilopes
2
-
Rodents
8
-
Insectivores
2
-
Bats
8
Humans
Laboratory
10
Strains
11
Totals:
35
Total:
81
16 2
46
Virus
strains
from groups 7 10 from Africa. Some of the groups were tentatively named according to the animal species most frequently represented therein. For instance, all eighteen red and polar fox origin viruses could be placed into either group 3 or 4. All attenuated rabies vaccine viruses except the Flury HEP strains are found in group 1, which in addition contains field isolates from three humans, two dogs, two rodents, and one mongoose. The viruses placed into groups 1-4 represented almost two-thirds of all strains tested. Most characteristic for the viruses obtained from the African continent is the reduced degree of cross-reactivity with the monclonal NC antibody panel. This feature is most prominent in viruses of group 7, which almost exclusively contains virus isolates from African bats. Of the seven viruses shown only two (SA-2, SA-3) gave identical reactions, indicating a common--possibly geographic--heterogeneity among rabies viruses of African bats.
Glycoprotein reaction patterns and protective activio' Rabies viruses assayed in mouse neutralization tests [4] against a panel of eighteen selected monoclonal G antibody types were grouped according to their individual reaction patterns (Table 4). In contrast to the viruses of groups A-E, those from groups F and G (horizontal panels of Table 4) are characterized by a significant reduction of their cross-reactivity with the panel of virus-neutralizing monoclonal antibodies. A correclation with NC-reaction groups could not be shown since one virus each was derived from NC-groups 1, 4 and 8, three from group 2 and five from the African group 7. The African viruses Lagos Bat (LBV), Mokola and Duvenhage (DUV) were shown previously [5] to be distinct serotypes within the genus rabies virus and are known to give
104
LOTHAR G. SCHNEIDER Table 3. Monoclonal rabies nucleocapsid antibodies W 1
O
NC R e a c t i o n
W 2
W 3
r-.
~
~
~
1
Type
2
Wild type
None
3
Wild l y p e
Fox I
•
4
Wild
type
F o x II
~E)
s
Flory
HEP
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only poor or no protection at all against a challenge infection by classical rabies viruses. We therefore selected four viruses from group F (rodent CS46, polar fox Alo 280, and two bat isolates Stade and Hamburg (SVF) from the Federal Republic of Germany) which were titrated intracerebrally in immunized mice and in non-vaccinated controls. The vaccine was prepared in suckling mice [6] with the Pitman-Moore strain of rabies virus which is widely used for the vaccine production and rabies prophylaxis in man. The results of the protection study are shown in Fig. I. Complete protection of vaccinated mice was obtained against the challenge infection with the Pitman -Moore virus (PM), the rodent isolate W239 and the polar fox virus ALO 280. The rodent strain CS 46 showed some breakthrough at virus dilutions - 1 and - 2 . Vaccinated mice exposed to Hamburg (SVF) and Stade bat viruses showed practically no protection. DISCUSSION The present study clearly indicates the worldwide existence of minor and major antigenic variants of rabies virus. As measured by monoclonal nucleocapsid antibodies, old and new
Antigenic variants of rabies virus
105
Table 4. Monoclonal rabies glycoprotein antibodies
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106
LOTHAR G. SCHNEIDER 100%
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Fig. I. Protective activity of Pitman Moore (PM) suckling mouse brain vaccine. Per cent mortality of nonvaccinated ( O - - O ) versus that of vaccinated mice (© O) after challenge with homologous (PM) and hetorologous viruses (rodent viruses W 239 and CS 46); polar fox virus ALO 280: bat viruses Hamburg (SVF) and Stade.
world rabies viruses as well as those from Asia may differ slightly but significantly from each other. Substantial differences of major NC-determinants exist however between the latter viruses and those obtained from the African continent. All African viruses tested so far have at least one major determinant in common. In fact, two rabies viruses isolated from unidentified bats found in the Northern part of the Federal Republic of Germany were retrospectively proven for the first time to be true African viruses. When evaluated by monoclonal virus neutralizing antibodies, rabies strains can be grouped into similar reaction pattern groups. From NC- and glycoprotein reaction groups, viruses can be selected which by subsequent analysis of their protective activity may be classified as true variants of the rabies virus or as newly recognized serotypes against which conventional vaccines may not fully protect. This study further indicates that a panel of nucleocapsid and neutralizing monoclonal antibodies representing major antigenic determinants may selectively identify such variants of field virus strains, thus being extremely helpful in epidemiological investigations. Acknowledgements--The technical assistance and excellent performance of Mrs. Susanne Meyer and Mrs. Dagmar Lindner in cloning and testing of monoclonal antibodies is gratefully acknowledged.
Antigenic variants of rabies virus
107
REFERENCES 1. Schneider, L. G., Antigenic determinants of rabies vaccine, Proceedings ~tf Cell Culture Rabies Vaccines and their Protective Effect in Man, Essen, Germany, March 1980, in press. 2. Schneider, L, G. and Meyer, S., Antigenic determinants of rabies virus as demonstrated by monoclonal antibody, Proceedings ¢~[the 4th lnternation~d Symposia ¢~fNegative Strand Viruses. Elsevier/North Holland, in press. 3. Wiktor, T. J. and Koprowski, H., Monclonal antibodies against rabies virus produced by somatic cell hybridization. Detection of antigenic variants, Proc. ham. Acad. Sci. U.S.A. 75, 3938 3942 (1978). 4. Atanasiu, P., Quantitative assay and potency test of antirabies serum and immunoglobulin, in Laboratory Techniques in Rabies (Eds. Kaplan, M. M. and Koprowski, H.), pp. 314~318. World Health Organization. Geneva (1973). 5. Schneider, L. G., Dietzaschold, B., Dierks, R. E., Matthaeus, W., Enzmann, P. J. and Strohmaier, K., Rabies group-specific ribonucleoprotein antigen and a test system for grouping and typing of rhabdoviruses, J. Virol. II, 748 755 (1973). 6. Fuenzalida, E., Suckling mouse brain vaccine, in Laboratory Techniques in Rabies (Eds. Kaplan, M. M. and Koprowski, H.), pp. 216 220. World Health Organization, Geneva (1973).
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