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Evaluation of the antigenic variation within type A foot and mouth disease virus isolates from Asia
Evaluation of the an lgenic variation within type A foot and mouth disease V "l r u s I " s o l a t e s fi-om A s i "a
M. M. Ru:o+emamu, ~ E beth J . Om~5"ia~e,T M. Head, ~ and Fm ~a Pm:se~
~ e ~roiogicM in~erce|ationshlps among |7 +y|x: A FMD virt+s Sirs*ins G-ore eight Asian ¢ountfi~ ~ r c ~¢~di¢~ b F +he t~:o+dimen,ion~| m~c¢~me~+t,~i~z~th&~ ~¢s~+ Comple× d~rec~ ~nd indirect re|~+ionship~ were obve~+ed+ Oi%+r'a||+ t+ow¢~ver~lhc virus strains studicxi ~ l t | d ~" c|~.s~fi¢~d as ~ I o n ~ i n g to die A22 g¢~+P on !|+e txtsls o f ++vM~e diffca+n+i~tlon +,+ P < O+OI
INTRODUCTION In 1 9 ~ a n e w e p i z ~ t i c o f f+~t a n d m o u t h di ( F M D ) d u e t o t y p e A virus s t a r t ~ J in t h e Persian G u | f a n d w i t h i n t w o y e a ~ it had s p ~ a d t h r o u g h o u t t h e M i d d l e East as 6at as T u r k e y and Russia+ Since t h e virus woo s h o w n t o ~ cliff%rent ~ o m all p r e v i o u s l y isolated t y p e A s t ~ i n s + it wets ~ssigne,+| a n e w s u b t y 22+ D u r i n g t h e e n s u i n g ~ r i e M t h e v i r u s ~*came established a n d xv~ e n z c o t i c in m a n y parts o f t h e M i d d l e Ea:sr w i t h c~casional incursions i n t o T h r a c e G a n d F~rts o f Turkey+ + A mith 2 studi~J t h e a n t i g e n i c v a r i a t i o n a m o n g t y ~ A virus s t = i n s f r o m t h e M i d d l e East ~ t w e e n 1964 a n d I 9 7 2 u s i n g a c o m p l e m e n t fixation test s y s t e m a n d a p p l y i n g t h e criteria for F M D virus subty:ping d e s c r i ~ J b y B r ~ k s b y . ~n she d e m o n s t r a t e d a d i v e r s i t y o f relationships + a m o n g the s t ~ i n s studied+ N o n e r h e | e s s she obs t h a t t h e s e virus strains related m o r e to t h e A22 s u b t y p e t h a n t o t h e E u n A s s u b t y ~ a n d c o n c l u d e d th-at t h e y s h o u l d c o n t i n u e ro be t e g a r d ~ J ~ ~mlonging t o t h e A22 s u b t y ~ . P e r e i ~ 4 c o n s i d e r - ~ + Received for publicatlon 15 June t983 * \re||come B$otc~hnology Lira|co-d. The E~cl|come FMDV L~.r~,or~,+ Pi~rlgh¢+ S~.~rrcy+ U K * T o wi~m all c o r r ~ n d e n c ¢ should ~" a&t + ( ~ 2 - | i5 019 t +O~ $0~ 1~}(0 ~ 1 9 ~ ~¢+ |nt¢+rna~ior~|A~:iatk>n of Bi~ogicM S~P~{ardi~at~oa 191
M, M, RWIYEMAML~ I 7 ' A I . that t h e c o n t i n u e d classification o f t y l x - A viruses from the Mi&tle East as AZ2 subtyjx. was b a ~ d more on epizt~tiologi¢al grounds~ i~e place o f o r i g i n , rather t h a n on a p # i c a t i o n o f t h e B m o k s b y classification system~ Adherence m these criteria w o u l d have iustifi~3 g r o u p i n g M i d d l e Eastern Type A viruses as b e l o n g i n g to d i s t i n c t subtypes. Such compromis¢'s have h-d several authors to question the subtyping
criteria~ ~-~ More recently we have advocated t h e use of ~he virus n e u t ~ l i z a t i o n reaction as dae reference m vitro ~est tbr F M D virus strain differentiation, since neumllizauion is unaffected by a n t i g e n - a n t i t × ~ y reactions involving tt~e n o n - i m m u n ~ e n i c a m i g e n s . ~ T h e present c o m m u n i c a t i o n descri~-s a s t u d y o f t y f ~ A virus isolates from the M i d d l e East, t h e Indian s u b c o n t i n e n t a n d B u r t o n Vires i~olates from t h e Philippines were not included in this s t u d y since previous analysis had shown t h e m m ix: related to t h e A24 s u b t y w : a n d probably associated w i t h intr~×tuetion o f disease from SOutil America,;* T h e test system e m p l o y e d was t h e two~dimensiona| m i c r o n e u t t a l i z a t i o n test re'It p ~ v i o u s l y s h o w n to h a v e n p~1¢~3 rest ~aaNance o f O - | 0 6 log SNN~. ~2 Hornotogouso-het¢'rologo~s c o m p a r i s o n x~s by t h e r vaI~e relationship, i t . ~a~ion o f serum [itre ~gainst t h e heterologous a n d homologous viruses. TaMe I s u m m a r i z e s m e a n r values for replicate comparisons. Tlmse values w h i c h were significantly less t h a n r = I "00 a t P < 0q) l are starred. In our experience this level o f differentiation in the t w o - d i m e n s i o n a l micronuetraliz~tion test iS appropriate tbr ~oth F M D virus classification and the selection o f v a c d n e viruses tbr r e ) u l a r i m m u n o prophylactic programmes~ ~ T.~m.~ 1~~-m~o~ical variathm within tyw¢ A FMD virus strains from Asia Anti~ra
Virus
l~dia 57/79
smfim lraq Turkey
k*banon JoMaa Pakistan
India ~ u t h c m Centra| Nort~ BangMdesh
Burma
24f64 Mahmzdi ~Turk~T 73} 9/78 0-08" M80 O,O~~ td78 O" 1~~ 8(78 O.41 I0/78 O*OI~ ~80 0.0| ~ 5W79 1-OO 54/79 0-32 8G~79 O~O2* 25/81 <002* 85/79 0-02* ~'78 O" 14~ 9/80 0"03 ~ 31/80 0"02" ~'78
O* 11"
Mahma~/i {Turkey 7~) O. t9
Daq 24164
Pakista,~ 7/8(}
0q}3" U"~7 i-OO
0-28
~'9~ > I~O 0.38 > tqg) > 1-00 0Q3
Bangladesh 9180
0,01"
>I~
O,O7" O-~* 0"03* 0"02* O-OI* O" 15~ O'O | " 0~03*
0*65 0-45 0~{0 0"t19* 0" 17" > 1-00 0%9 > 1"00 > I "O0
I>34
0-25
0"28 0-25 > lql0 0"08"
I'O0
O*I ~*
O~05*
O~O7i O~17"
O" ~6 ~
0"08 ~ 0"2 I*
0" 17"
* Va|u~ of r ate significantly dif|~tent from l.~J at P < DuOt.. Value, of r obtained in ~wo-dlmen,~onzl mkmneumdi~a¢ion ~¢~t~ 192
India 25/81
bOO
Iq~ Oq~l" 0-O 1~ 0~) ~ 0-20 ~ O"TO"
0"05*
Turkey 2/80
0~65 0,81
0~/8 *
O IO~ 0"O4" O'58 t'~ O*16" O"12~ 0"31 0"22"
0"56 O'(K)* 0.42 O"36 1 *00 0~8 l
AN"~ | G I N ~ C V A R I A T ~ ( ) N
IN FMI)V ISOI.ATES
l
L e D o ~ ~178
Fig, i, Intefc~-Iatio~si~ips a m o n g s t | : g l D ¢FD" A ~it~s $~rains gf~}m A$ia ~]~asrxt of~, x~a~am ~*ot dif}k et'~t
t~om I-0 a~ P = ()'OD, The interrelationship groupings are illustrated in Fig. I t¥oln which the following inl;erences are apparent. The A22 Mahmatii virus had the narrowest serological sptvtrum o f all e~,e strains studied~ This strain {here~re appears to l~z no longer a re as a vaccine virus ~or the M i d d l e Easr~ \Xeith Turkey 2180 antiserum it ~'as ~ s s i b l e u~ group viruses t h e East Mediterranean basin (Turkey, Lebanon and Jordan)° The interrelationship amot~g strains l?om the Indian sub¢ontinen~ was more ¢oroplex. Those froro Southern India were e)txsely related to each other bur tlae isolare~ from Central and N o r t h e r n India were relawd either to strai~~s £rotn Pakistan or those G-on Bangladesh..Ar~tisem to A22 lraq 20164 show~t this strain to have the brwadest serological spectrum covering viruses froro the whole region seLld~ed. Ttlis broad spectrum v;.a~ detected w i t h differem A22 Iwaq 24/64 ant|sets and had beet, observed by Arruwsmith. 2 As discussed elsewhere8 the differen¢iatio= of FMD virus stnlins on the basis o f r v a ! u ~ in a matmer illustrated in Fig. 1 is a system which allows for complex. relationships to k~. evaluatc~. For e:,:ample two viruses froro Indian t2518 ~ and 85[79) which were p~0rly neatraliz~u] by the lraq antiserum are related to the I~mq 24164 strain t h r o u g h their reaction wi~h other Indian and Ban,g|adesh isolates~ In this India 25/81 antiserum r~aicted we|l with the isolates Pakistan, In~dia and ~ang~ ladesh, which were also readily neutralized by Iraq 24]64 antiserum. Bangladesh antiserum showe~J India 85/79 to be closely related ro India 8~¢79 and the Bangladesh isolates all o f which were related to the l ~ q 24164. Similarly, th e relationship of~he Mahmatli strain to the other strains is t h r o u g h its relationship with lraq Consequently, it is now possible to regaM all the virus strains s t u d i M as beiorlging co the A m g r o u p as a result o f serological data rather than simply on field epizootiMogicat considerations. Ir would ~ useftll to extend these studies by application of Mochemical analytical tevlmiques such as ol u c l ~ t i d e T 1 m a p p i n g and even g e n o m e s ~ u e n c i n g since2 Robson et al, t3 also strated similarities between T t maps o f three provisionMly d e s i g n a t M A22 strains. Such an approach wo u ld help to study the naature of the evolution of antigenic variation in F/~{D. I¢ is interesting that the vir~as w i t h the b ~ d e s t serologicM spectrum should ~" the iso{ate ~s¢~ciated w i t h the originM e p i a ~ t i c in I964. This would to
9~
M M, RWEYEMAMU ET AL. -t D compatible with the concept that the maior source of antigenic change in F~,~ endemic areas is antigenic drift, Over a ~ r i c d of time new variants ari~. inde~ndently which, although different from one another, continue to ~ a r some relationship with the original virus isolate. Finally, a virus like A22 lraq 24f64 seems to be the prime candidate for a baseline A22 vaccine strain for application throughout Asia. In isolated circumstances it could t~ supplemented with local vir~s i~lates in order to contain particular troublesome outbreaks ~ f o r e they develop into epizo~ti¢s, It is apparent from the data pre~ented in this communication that it should no longer ~ considered appropriate to select va~ine strains merely on the b ~ i s oftheir subtype h u m o r designation or ~ representari~e of the most recent isolate ~ has be~,n traditional f.or selecting FMD vaccine viruses. REFERENCES 1. Boldrini GM~ Vaccination and the control of f ~ t and mouth dis~ase. Vet R ~ 1978; t02: 194-I98. 2, Arrowsmirh AEM~ Variation among st~ins of tyr~ A f~at and mouth disarm virus in the Eastern Mediterranean region J Hyg 1975; 7 t: 387. 3- Bm~ksby lB. Variants a ~ immunity~efinitions riot serol~kal inx~.~tigarion Syrup lmm Stand 1968; 8: I~I0. 4. Perelra HG, Antigenic variation in relation to epidemioMgy and control of f~)t and mouth disease. Br Vet J 1978; | M : 58-62° 5. Norman AJ. The subty~ classification of strains of f~ot and mouth dir~uc vires. J Hyg 1975; 74: 227-232. 6. Perei~ HG. Subtyping of' fi¢~t and mouth dis~se virus. D~cv Biol Stand 1977; 35: I6%-!74. 7. R area MM, Pay TV4F, Parker MJ, ~rological differentiation of ~oot and mouth disuse virus strains in relation to selection of suitaMe vaccine viruses, Dev Biol Stand 1977; 35: 205-214. 8~ Rweyemamu MM~ Ant igenk ~riation ofti~)t and mouth disease virus. J BiN Stand 1984; 12: in pr~s. Rweyemamu MM, Pay T~'F. Studies on fi~3t and mouth dise~e virus strains A Morrocco 9,, 5/77o Bull Inr Epiz~3t 1978; 89: 861~875. 10. B ~ t h J C , R~.eyemamu MM, Pay TWF, ~ s e resg~nse relationshi~ in a mktoneum~lisa~ tion tt~t for f~y~tand mouth disease v i r u s . J Hyg I978; 80:31-42. M, Pay T~/F. Mkroacut~Iisarion tests for ~-rotogical II. Rweyemamu MM, ~ o t h J C , typing ~nd subtyping o f f ~ x and mouth d i ~ s c vires srraies~ ) Hyg 1978; 8 t: 107-123. 12. Rweyemmmu MM, Hingk T P.I. F ~ r and mouth disease virus strains differentiation: AnNysis d the semifinal data. J Biol Stand 1984; 12: 225-229. ~ H , CrowtherJR, King AMQ, Brown Fo Comparative bi~hemical and seroI3. R logical analysis of five isolates of a single seroty~ a r t i s t and mouth d i s e ~ virus. ) Gen Virol I979; 45: 579-590o
M. M. Ru:o+emamu, ~ E beth J . Om~5"ia~e,T M. Head, ~ and Fm ~a Pm:se~
~ e ~roiogicM in~erce|ationshlps among |7 +y|x: A FMD virt+s Sirs*ins G-ore eight Asian ¢ountfi~ ~ r c ~¢~di¢~ b F +he t~:o+dimen,ion~| m~c¢~me~+t,~i~z~th&~ ~¢s~+ Comple× d~rec~ ~nd indirect re|~+ionship~ were obve~+ed+ Oi%+r'a||+ t+ow¢~ver~lhc virus strains studicxi ~ l t | d ~" c|~.s~fi¢~d as ~ I o n ~ i n g to die A22 g¢~+P on !|+e txtsls o f ++vM~e diffca+n+i~tlon +,+ P < O+OI
INTRODUCTION In 1 9 ~ a n e w e p i z ~ t i c o f f+~t a n d m o u t h di ( F M D ) d u e t o t y p e A virus s t a r t ~ J in t h e Persian G u | f a n d w i t h i n t w o y e a ~ it had s p ~ a d t h r o u g h o u t t h e M i d d l e East as 6at as T u r k e y and Russia+ Since t h e virus woo s h o w n t o ~ cliff%rent ~ o m all p r e v i o u s l y isolated t y p e A s t ~ i n s + it wets ~ssigne,+| a n e w s u b t y 22+ D u r i n g t h e e n s u i n g ~ r i e M t h e v i r u s ~*came established a n d xv~ e n z c o t i c in m a n y parts o f t h e M i d d l e Ea:sr w i t h c~casional incursions i n t o T h r a c e G a n d F~rts o f Turkey+ + A mith 2 studi~J t h e a n t i g e n i c v a r i a t i o n a m o n g t y ~ A virus s t = i n s f r o m t h e M i d d l e East ~ t w e e n 1964 a n d I 9 7 2 u s i n g a c o m p l e m e n t fixation test s y s t e m a n d a p p l y i n g t h e criteria for F M D virus subty:ping d e s c r i ~ J b y B r ~ k s b y . ~n she d e m o n s t r a t e d a d i v e r s i t y o f relationships + a m o n g the s t ~ i n s studied+ N o n e r h e | e s s she obs t h a t t h e s e virus strains related m o r e to t h e A22 s u b t y p e t h a n t o t h e E u n A s s u b t y ~ a n d c o n c l u d e d th-at t h e y s h o u l d c o n t i n u e ro be t e g a r d ~ J ~ ~mlonging t o t h e A22 s u b t y ~ . P e r e i ~ 4 c o n s i d e r - ~ + Received for publicatlon 15 June t983 * \re||come B$otc~hnology Lira|co-d. The E~cl|come FMDV L~.r~,or~,+ Pi~rlgh¢+ S~.~rrcy+ U K * T o wi~m all c o r r ~ n d e n c ¢ should ~" a&t + ( ~ 2 - | i5 019 t +O~ $0~ 1~}(0 ~ 1 9 ~ ~¢+ |nt¢+rna~ior~|A~:iatk>n of Bi~ogicM S~P~{ardi~at~oa 191
M, M, RWIYEMAML~ I 7 ' A I . that t h e c o n t i n u e d classification o f t y l x - A viruses from the Mi&tle East as AZ2 subtyjx. was b a ~ d more on epizt~tiologi¢al grounds~ i~e place o f o r i g i n , rather t h a n on a p # i c a t i o n o f t h e B m o k s b y classification system~ Adherence m these criteria w o u l d have iustifi~3 g r o u p i n g M i d d l e Eastern Type A viruses as b e l o n g i n g to d i s t i n c t subtypes. Such compromis¢'s have h-d several authors to question the subtyping
criteria~ ~-~ More recently we have advocated t h e use of ~he virus n e u t ~ l i z a t i o n reaction as dae reference m vitro ~est tbr F M D virus strain differentiation, since neumllizauion is unaffected by a n t i g e n - a n t i t × ~ y reactions involving tt~e n o n - i m m u n ~ e n i c a m i g e n s . ~ T h e present c o m m u n i c a t i o n descri~-s a s t u d y o f t y f ~ A virus isolates from the M i d d l e East, t h e Indian s u b c o n t i n e n t a n d B u r t o n Vires i~olates from t h e Philippines were not included in this s t u d y since previous analysis had shown t h e m m ix: related to t h e A24 s u b t y w : a n d probably associated w i t h intr~×tuetion o f disease from SOutil America,;* T h e test system e m p l o y e d was t h e two~dimensiona| m i c r o n e u t t a l i z a t i o n test re'It p ~ v i o u s l y s h o w n to h a v e n p~1¢~3 rest ~aaNance o f O - | 0 6 log SNN~. ~2 Hornotogouso-het¢'rologo~s c o m p a r i s o n x~s by t h e r vaI~e relationship, i t . ~a~ion o f serum [itre ~gainst t h e heterologous a n d homologous viruses. TaMe I s u m m a r i z e s m e a n r values for replicate comparisons. Tlmse values w h i c h were significantly less t h a n r = I "00 a t P < 0q) l are starred. In our experience this level o f differentiation in the t w o - d i m e n s i o n a l micronuetraliz~tion test iS appropriate tbr ~oth F M D virus classification and the selection o f v a c d n e viruses tbr r e ) u l a r i m m u n o prophylactic programmes~ ~ T.~m.~ 1~~-m~o~ical variathm within tyw¢ A FMD virus strains from Asia Anti~ra
Virus
l~dia 57/79
smfim lraq Turkey
k*banon JoMaa Pakistan
India ~ u t h c m Centra| Nort~ BangMdesh
Burma
24f64 Mahmzdi ~Turk~T 73} 9/78 0-08" M80 O,O~~ td78 O" 1~~ 8(78 O.41 I0/78 O*OI~ ~80 0.0| ~ 5W79 1-OO 54/79 0-32 8G~79 O~O2* 25/81 <002* 85/79 0-02* ~'78 O" 14~ 9/80 0"03 ~ 31/80 0"02" ~'78
O* 11"
Mahma~/i {Turkey 7~) O. t9
Daq 24164
Pakista,~ 7/8(}
0q}3" U"~7 i-OO
0-28
~'9~ > I~O 0.38 > tqg) > 1-00 0Q3
Bangladesh 9180
0,01"
>I~
O,O7" O-~* 0"03* 0"02* O-OI* O" 15~ O'O | " 0~03*
0*65 0-45 0~{0 0"t19* 0" 17" > 1-00 0%9 > 1"00 > I "O0
I>34
0-25
0"28 0-25 > lql0 0"08"
I'O0
O*I ~*
O~05*
O~O7i O~17"
O" ~6 ~
0"08 ~ 0"2 I*
0" 17"
* Va|u~ of r ate significantly dif|~tent from l.~J at P < DuOt.. Value, of r obtained in ~wo-dlmen,~onzl mkmneumdi~a¢ion ~¢~t~ 192
India 25/81
bOO
Iq~ Oq~l" 0-O 1~ 0~) ~ 0-20 ~ O"TO"
0"05*
Turkey 2/80
0~65 0,81
0~/8 *
O IO~ 0"O4" O'58 t'~ O*16" O"12~ 0"31 0"22"
0"56 O'(K)* 0.42 O"36 1 *00 0~8 l
AN"~ | G I N ~ C V A R I A T ~ ( ) N
IN FMI)V ISOI.ATES
l
L e D o ~ ~178
Fig, i, Intefc~-Iatio~si~ips a m o n g s t | : g l D ¢FD" A ~it~s $~rains gf~}m A$ia ~]~asrxt of~, x~a~am ~*ot dif}k et'~t
t~om I-0 a~ P = ()'OD, The interrelationship groupings are illustrated in Fig. I t¥oln which the following inl;erences are apparent. The A22 Mahmatii virus had the narrowest serological sptvtrum o f all e~,e strains studied~ This strain {here~re appears to l~z no longer a re as a vaccine virus ~or the M i d d l e Easr~ \Xeith Turkey 2180 antiserum it ~'as ~ s s i b l e u~ group viruses t h e East Mediterranean basin (Turkey, Lebanon and Jordan)° The interrelationship amot~g strains l?om the Indian sub¢ontinen~ was more ¢oroplex. Those froro Southern India were e)txsely related to each other bur tlae isolare~ from Central and N o r t h e r n India were relawd either to strai~~s £rotn Pakistan or those G-on Bangladesh..Ar~tisem to A22 lraq 20164 show~t this strain to have the brwadest serological spectrum covering viruses froro the whole region seLld~ed. Ttlis broad spectrum v;.a~ detected w i t h differem A22 Iwaq 24/64 ant|sets and had beet, observed by Arruwsmith. 2 As discussed elsewhere8 the differen¢iatio= of FMD virus stnlins on the basis o f r v a ! u ~ in a matmer illustrated in Fig. 1 is a system which allows for complex. relationships to k~. evaluatc~. For e:,:ample two viruses froro Indian t2518 ~ and 85[79) which were p~0rly neatraliz~u] by the lraq antiserum are related to the I~mq 24164 strain t h r o u g h their reaction wi~h other Indian and Ban,g|adesh isolates~ In this India 25/81 antiserum r~aicted we|l with the isolates Pakistan, In~dia and ~ang~ ladesh, which were also readily neutralized by Iraq 24]64 antiserum. Bangladesh antiserum showe~J India 85/79 to be closely related ro India 8~¢79 and the Bangladesh isolates all o f which were related to the l ~ q 24164. Similarly, th e relationship of~he Mahmatli strain to the other strains is t h r o u g h its relationship with lraq Consequently, it is now possible to regaM all the virus strains s t u d i M as beiorlging co the A m g r o u p as a result o f serological data rather than simply on field epizootiMogicat considerations. Ir would ~ useftll to extend these studies by application of Mochemical analytical tevlmiques such as ol u c l ~ t i d e T 1 m a p p i n g and even g e n o m e s ~ u e n c i n g since2 Robson et al, t3 also strated similarities between T t maps o f three provisionMly d e s i g n a t M A22 strains. Such an approach wo u ld help to study the naature of the evolution of antigenic variation in F/~{D. I¢ is interesting that the vir~as w i t h the b ~ d e s t serologicM spectrum should ~" the iso{ate ~s¢~ciated w i t h the originM e p i a ~ t i c in I964. This would to
9~
M M, RWEYEMAMU ET AL. -t D compatible with the concept that the maior source of antigenic change in F~,~ endemic areas is antigenic drift, Over a ~ r i c d of time new variants ari~. inde~ndently which, although different from one another, continue to ~ a r some relationship with the original virus isolate. Finally, a virus like A22 lraq 24f64 seems to be the prime candidate for a baseline A22 vaccine strain for application throughout Asia. In isolated circumstances it could t~ supplemented with local vir~s i~lates in order to contain particular troublesome outbreaks ~ f o r e they develop into epizo~ti¢s, It is apparent from the data pre~ented in this communication that it should no longer ~ considered appropriate to select va~ine strains merely on the b ~ i s oftheir subtype h u m o r designation or ~ representari~e of the most recent isolate ~ has be~,n traditional f.or selecting FMD vaccine viruses. REFERENCES 1. Boldrini GM~ Vaccination and the control of f ~ t and mouth dis~ase. Vet R ~ 1978; t02: 194-I98. 2, Arrowsmirh AEM~ Variation among st~ins of tyr~ A f~at and mouth disarm virus in the Eastern Mediterranean region J Hyg 1975; 7 t: 387. 3- Bm~ksby lB. Variants a ~ immunity~efinitions riot serol~kal inx~.~tigarion Syrup lmm Stand 1968; 8: I~I0. 4. Perelra HG, Antigenic variation in relation to epidemioMgy and control of f~)t and mouth disease. Br Vet J 1978; | M : 58-62° 5. Norman AJ. The subty~ classification of strains of f~ot and mouth dir~uc vires. J Hyg 1975; 74: 227-232. 6. Perei~ HG. Subtyping of' fi¢~t and mouth dis~se virus. D~cv Biol Stand 1977; 35: I6%-!74. 7. R area MM, Pay TV4F, Parker MJ, ~rological differentiation of ~oot and mouth disuse virus strains in relation to selection of suitaMe vaccine viruses, Dev Biol Stand 1977; 35: 205-214. 8~ Rweyemamu MM~ Ant igenk ~riation ofti~)t and mouth disease virus. J BiN Stand 1984; 12: in pr~s. Rweyemamu MM, Pay T~'F. Studies on fi~3t and mouth dise~e virus strains A Morrocco 9,, 5/77o Bull Inr Epiz~3t 1978; 89: 861~875. 10. B ~ t h J C , R~.eyemamu MM, Pay TWF, ~ s e resg~nse relationshi~ in a mktoneum~lisa~ tion tt~t for f~y~tand mouth disease v i r u s . J Hyg I978; 80:31-42. M, Pay T~/F. Mkroacut~Iisarion tests for ~-rotogical II. Rweyemamu MM, ~ o t h J C , typing ~nd subtyping o f f ~ x and mouth d i ~ s c vires srraies~ ) Hyg 1978; 8 t: 107-123. 12. Rweyemmmu MM, Hingk T P.I. F ~ r and mouth disease virus strains differentiation: AnNysis d the semifinal data. J Biol Stand 1984; 12: 225-229. ~ H , CrowtherJR, King AMQ, Brown Fo Comparative bi~hemical and seroI3. R logical analysis of five isolates of a single seroty~ a r t i s t and mouth d i s e ~ virus. ) Gen Virol I979; 45: 579-590o