Br. vet. J. (1976), 13ll,
I 12
SEROLOGICAL RESPONSE TO THE INDIRECT FLUORESCENT ANTIBODY TEST OF CATTLE INFECTED WITH THEILERIA SERGENT/ BY
K.
TAKAHASHI,*
S. Y.
YAMASHITA,*
Y.
lsAYAMAt
AND
SHIMIZU:j:
*Department tif Veterinary Medicine, College of Dairying, Nishinopporo, Ebetsu, Japan, t Hokkaido Branch Laboratory, National Institute of Animal Health, Hitsujigaoka, Toyohiraku, Sapporo, Japan, :j:National Institute tif Animal Health, Kodaira, Tokyo, Japan SUMMARY
Sera from two calves experimentally infected with Theileria sergenti were tested for antibody by the indirect fluorescent antibody (IFA) test. Antibody appeared on the 5th day after infection and reached a peak on the 2oth day. A decrease in severity ofparasitaemia and an increase in packed cell volume (PCV) were observed when antibody titre reached a peak and when it was maintained at a high level after recovery. There was a rapid rise in antibody titre of the lgM fractions with the intensification of parasitaemia after infection. The titre declined in parallel with the number of parasites detected and was maintained at a low level until 6o days after infection. On the contrary, IgG antibody was first detected on the 1oth day, and its high titre persisted for more than 6o days, as well as that of total antibody. The experimental results obtained from naturally infected cows corresponded to those from the experimentally infected calves. The role of humoral antibody detected by IFA test in the inhibition of parasitaemia in cattle infected with T. sergenti is discussed. INTRODUCTION
In a previous report the application of the indirect fluorescent antibody (IFA) test to bovine piroplasmosis (Theileria sergenti infection of cattle) was described (Takahashi, Yamashita & Shimizu, 1972). A significant antibody titre was demonstrated in the serum of cattle during the 2-3 months after recovery. The recovered cattle resisted challenge infection with the homologous strain. This paper reports studies on the serological activities of the immunoglobulin fractions of sera collected from cattle infected with T. sergenti by using the IFA test. MATERIALS AND METHODS
Experimental animals Two male Holstein-Friesian calves, 15 days old (No. 86 and No. 88), were used in this experiment. To ensure that the calves were free from infection with
SEROLOGICAL RESPONSE IN BOVINE THEILERIOSIS
113
T. sergenti or any other blood parasite, smears were collected and stained with Giemsa stain for examination. Sera collected before infection were negative for antibody in the IFA test. The calves were infected by the intravenous inoculation of 50 ml of blood obtained from a splenectomized donor calf which had been suffering from severe parasitaemia after experimental infection. The donor's blood had been confirmed to be free from parasites other than T. sergenti and contained 1 ·5 X I0 10 parasites of this species. Blood samples were collected from both experimentally inoculated calves at 5-day intervals until 6o days after infection, and tested for parasitaemia, antibody level, and packed cell volume (PCV). Another five pastured cows which had recovered more than 1oo days following natural infection of T. sergenti were examined by the same method. Estimation of parasitaemia and PCV Parasitaemia was estimated by the microscopic examination of blood films stained with Giemsa stain. The degree of parasitaemia was expressed as a percentage of parasitized erythrocytes. PCV was examined in a capillary tube by centrifugation at I2 ooo rev/min for 5 min. Fractionation of serum Two ml samples of serum were fractionated by gel filtration through a 2·5 X 45 em column of Sephadex G-200 (Pharmacia, Uppsala, Sweden) equilibrated with o·I5 M- NaCl in o·oi M phosphate buffer solution (PBS). The effluent fraction was collected in a fraction collector at a flow rate of 2 ml per IO min and examined for optical density at 280 ml-' by a spectrophotometer. Fluorescent antibody technique Rabbit anti-bovine serum globulin was conjugated with fluorescein isothiocyanate using the method of Marshall, Eveland & Smith (I958). The conjugated globulin was passed through a gel filtration column and forced through a DEAE cellulose column. The conjugate obtained was stored at -20°C. The IFA test was carried out essentially by the same method as that described in the previous report (Takahashi et al., I972). The antigen was prepared from whole blood from a splenectomized calf experimentally infected with 10 per cent T. sergenti to induce parasitaemia. Erythrocytes were collected from the blood and washed three times in PBS. Thin films were made on coverslips 18 X I8 mm in size and dried, fixed in cold acetone for IO min and washed with o·oi M PBS (pH 7·2) for 5 min. Sera from infected calves were stored at -20°C until ready for use. Two-fold dilutions ranging from I: IO to I: 10 240 were prepared in tubes, and o·I ml of each dilution was placed on the antigen-coated coverslip. The coverslips were placed in a moist chamber and incubated at 37°C for 30 min and then washed with PBS for I5 min. The coverslip antigen was stained with conjugated rabbit anti-globulin at 37°0 for 30 min, rinsed in several changes of
BRITISH VETERINARY JOURNAL,
132, 1
PBS, mounted in 50 per cent buffered glycerin (pH g·o), and examined by the fluorescent microscope. The end-point was recorded as the highest dilution of serum giving specific fluorescence of the piroplasma, a titre of I : 20 or higher was considered significant.
Antibody activities of lgM and lgG Reading of absorbance at 280 mf' gave three peaks. The antibody titre of each fraction was measured by the IFA test. The antibody titres of the IgM and lgG fractions were calculated from the total of antibody titres ofthe first and second peak, respectively. RESULTS
The degree of parasitaemia, PCV, and antibody level in the two calves, are illustrated in Fig. I. The parasite began to appear in the blood 5 days after infection, and a marked rise in numbers occurred between IO and 20 days. The maximum rate of increase of infected erythrocytes during this period was I I per cent. The PCV fell to I3-I6 per cent when infected erythrocytes reached a maximum level. The increase of PCV coincided with the drop in number of infected erythrocytes thereafter. The antibody titre rose rapidly with the appearance of protozoa in the peripheral blood. When it reached a peak, parasitaemia tended to decrease gradually until 6o days after infection. An elevated plateau was maintained for more than 40 days. The markedly high antibody titre of IgM accompanied by the severest parasitaemia was observed 20 days after infection. Then IgM began to show a gradual decrease in level, but no shift to IgG antibody was observed until 6o days after infection. The pattern of IgG antibody production was parallel to that of serum antibody titres in all the serum samples. The number of parasites and the lgG antibody level increased more rapidly in calf No. 88 than in calf No. 86. Although there was a slight difference in the antibody response or parasitaemia between the two calves, the overall picture was essentially the same in both calves. Table I records the antibody titres, parasitaemia, and PCV obtained with serum or blood samples from cows naturally infected with T. sergenti. It is clear that the serum samples gave high antibody titres, while the value of parasitaemia ranged from 2·I to g·o. The antibody titre of the IgM fraction was higher in these cows than in experimentally infected calves in the convalescent stage. DISCUSSION
Calves experimentally infected with T. sergenti produced IgM and IgG antibodies detectable by the IFA test, but no evidence of a shift from IgM to IgG synthesis was demonstrated. Similar results were obtained with Babesia microti (Cox & Turner, I970a) and Plasmodium berghei yoelii in mice (Cox & Turner, I97ob). IgM antibody appeared earlier than IgG antibody in calves
SEROLOGICAL RESPONSE IN BOVINE THEILERIOSIS
115
~----(>---0
-,
·------·--..:----·
3 Calf No. 86
~·
30
~
'
..
' __ ',
0
> 0. 20
"'
-.
', ______ .._ _____
___....-l(~
x_____x
0 _J
.
--"'..............-
12 10
8
~ .Q
i
6 .e -~
4
10
&.
2
0
0
3
30
12
f
~ 0
> 20 0.
10~
E 0
"'0
a
0 .E
6
]
J
10
~
4 2
0
0
20
30
40
50
60
.o
Days after nfection
Fig. I. Antibody levels, parasitaemia and packed cell volume of calves experimentally infected with T. Sergenti. 0 - - 0 whole serum; .A----.A IgG; 0----0 parasitaemia; e ----e IgM; x- - x PCV. TABLE I ANTIBODY LEVELS, PARASITAEMIA AND PACKED CELL VOLUME OF COWS RECOVERED FROM NATURAL INFECTION WITH T. sergenti
Antibody titre Cow No.
Whole serum
lgM
lgG
Parasitaemia (%)
PCV* ( %)
I 2 3 4 5
640 2560 640 I280 I280
230 I30
670 3480 790 I500 II 50
3•2 2"I s·6 4"7 g·o
27 28 22 Ig 22
* Packed cell volume.
IOO
I8o 8o
116
BRITISH VETERINARY JOURNAL, 132, r
experimentally infected with T. sergenti. These findings were almost parallel with those from the observation on Plasmodium rynomologi and P. vivax in man (Abele et al., 1965). IgM and IgG antibodies produced by B. microti (Cox & Turner, 1970a) and P. berghei yoelii (Cox & Turner, 197ob), however were detected from mice at the same time after infection. In mice experimentally infected with B. microti, the protozoa disappeared completely from the peripheral blood when the IgG antibody level reached a peak (Cox & Turner, 1970a). In calves experimentally infected with T . sergenti and showing an lgG antibody level having reached a peak, low parasitaemia was observed until 6o days after infection. This suggested that there might be differences in IgG antibody activity among the species of erythrocytic protozoa. A decrease in the numbers of protozoa in erythrocytes and remarkable recovery from anaemia were seen with a rise of IgG antibody level in calves experimentally or naturally infected with T. sergenti. It seemed that humoral antibody might have inhibited the parasitaemia caused by T. sergenti infection. The investigation of P. berghei and B. rodhaini (Phillips, 1969) also indicated the inhibitory effect ofhumoral antibody on parasitaemia. Phillips &Jones (1972) described a transfer of resistance toP. berghei by passively administered antisera. This transfer resulted in the inhibition ofparasitaemia and brought about early recovery from anaemia. Since then, several workers have demonstrated a passive transfer of immunity with serum collected from infected animals. ACKNOWLEDGEMENT
We are particularly grateful to Dr W. J. Brinley Morgan, Ministry of Agriculture, Fisheries and Food, Central Veterinary Laboratory, Weybridge, England, for his advice. REFERENCES
ABELE, D. C., ToBIE,]. E ., HILL, G.]., CoNTAcos, P. G. & EvANs, C. B. (1965). Am.]. trop. Med. Hyg. 14, 191. Cox, F. E. G. & TuRNER, S. A. (1970a). Ann. trop. Med. Parasit. 64> 167. Cox, F. E. G. & TuRNER, S. A. (1970b). Ann. trop. Med. Parasit. 64, 175. MARSHALL, ]. D., EVELAND, W. C. & SMITH, C. W. (1958). Proc. Soc. exp. Biol. Med. !JS, 898. PHILLIPS, R . S. {1969). Parasitology, 59• 35 7. PHILLIPS, R. S. & joNEs, V. E. (I972). Parasitology, 64, 117. TAKAHASHI, K., YAMASHITA, S. & SHIMIZU, Y. (1972). Jap. J. Vet. Sci. 34> 275·
(Accepted for publication IB March I975)
Response serologique du betail infecte par Theileria serge11ti au test de detection des anticorps par inununofluorescence indirecte
(Takahashi et a/.) ResUDle. Des serums de deux veaux infectes experimentalement par Theileria sergenti ant ete analyses par immunofluorescence indirecte (IFI). Les anticorps apparurent au 5e jour apres l'infection et atteignirent un niveau maximum au 20e jour. On a note une diminution de Ia
SEROLOGICAL RESPONSE IN BOVINE THEILERIOSIS
; 117
parasitemie et une augmentation de la concentration globulaire en hemoglobine (CGH), lorsque le titre de l'anticorps atteignit un niveau maximum et il devait rester eleve durant la periode de guerison. II y eut une rapide elevation de la fraction IgM des anticorps avec une intensification de la parasitemie apres !'infection. Le titre devait decliner parallelement au nombre de parasites detectes et reste bas jusqu'a 6o jours apres !'infection. Par contre, les anticorps IgG detectes pour la lere fois au we jour avaient un titre eleve qui persista pendant 6o jours comme celui de !'ensemble des anticorps. Les resultats experimentaux obtenus sur des vaches naturellement infectees ont ete comparables a ceux des veaux experimentalement infectes. Le role de l'anticorps humoral detecte en IFI dans le freinage de la parasitemie du betail infecte au T. sergenti est discute. Serumsreaktionen bei Rindern, die mit Thei/erill sergenti infiziert worden waren und mit deDl indirekten Fluoreszenztest auf Antikorper untersucht wurden (Takahashi et a/.) Zusanunenfassung. Von zwei Kalbem, die experimentell mit Theileria sergenti infiziert worden waren, wurden die Sera mit dem indirekten Antikorperfluoreszenztest (IFA) auf Vorhandensein von Antikorpern untersucht. Antikorper zeigten sich am 5· Tag nach der Infektion und erreichten einen Hohepunkt am 20. Tag. Wenn der Antikorpertiter einen Hohepunkt erreichte und nach der Wiederstellung hoch blieb, wurde eine Verminderung der Parasitaemie und eine Vermehrung des Zellvolumen nach Sedimentation (PVC) festgestellt. Mit der erhohten Parasitaemie nach der Infektion ging ein rapider Anstieg des Antikorpertiters der lgM-Anteile einher. Parallel zu der Anzahl der gefundenen Parasiten nahm der Titer ab und hielt sich in verminderter Hohe his zu 6o Tagen nach der lnfektion. Im Gegensatz hierzu wurde der lgG-Antikorper zuerst am 1o.Tage entdeckt, und sein hoher Titer hielt 6o Tage lang an ebenso wie der des gesamten Antikorpers. Bei unter natiirlichen Umstanden infizierten Kiihen entsprachen die Resultate den bei experimentell infizierten Kalbern beobachteten. Es wird erortert, welche Rolle der durch den !FA-Test entdeckte humorale Antikorper bei lnhibierung der bei mit T. sergenti infizierten Rind ern auftretende Parasitaemie spielt. Respuesta serol6gica a Ia prueba indirecta de anticuerpos fluorescentes en el ganado infectado con Theileria sergenti (Takahashi et a/.) Resmnen. Se comprobaron los anticuerpos del suero de dos terneros, infectados experimentalmente con Theileria sergenti, mediante la prueba indirecta de anticuperos fluorescentes (IAF). Los anticuerpos aparecieron el 5° dia despues de la infecci6n y alcanzaron un maximo en el 20° dia. Se observaron una disminuci6n en la gravedad de la parasitemia y un aumento en el volumen de celulas concentradas cuando el valor de anticuerpos alcanz6 un maximo y cuando se mantuvo a un nivel elevado despues de la recuperaci6n. Se di6 una r:ipida subida del valor de anticuerpos de las fracciones IgM con la intensificaci6n de parasitemia despues de la infecci6n. El valor declin6 en paralelo con el numero de par:isitos descubierto y se mantuvo a un nivel bajo hasta 6o dias despues de la infecci6n. Por el contrario, el anticuerpo lgG se descubri6 por primera vez el 10° dia, y su valor elevado persisti6 por 6o dias, ademas del de los anticuerpos totales. Los resultados experimentales obtenidos de vacas infectadas de forma natural correspondieron con los obtenidos de temeros infectados experimentalmente. Se discute la funci6n del anticuerpo humoral, descubierto por la prueba IAF, en la inhibici6n de parasitemia en el ganado infectado con T. sergenti.
I 12
SEROLOGICAL RESPONSE TO THE INDIRECT FLUORESCENT ANTIBODY TEST OF CATTLE INFECTED WITH THEILERIA SERGENT/ BY
K.
TAKAHASHI,*
S. Y.
YAMASHITA,*
Y.
lsAYAMAt
AND
SHIMIZU:j:
*Department tif Veterinary Medicine, College of Dairying, Nishinopporo, Ebetsu, Japan, t Hokkaido Branch Laboratory, National Institute of Animal Health, Hitsujigaoka, Toyohiraku, Sapporo, Japan, :j:National Institute tif Animal Health, Kodaira, Tokyo, Japan SUMMARY
Sera from two calves experimentally infected with Theileria sergenti were tested for antibody by the indirect fluorescent antibody (IFA) test. Antibody appeared on the 5th day after infection and reached a peak on the 2oth day. A decrease in severity ofparasitaemia and an increase in packed cell volume (PCV) were observed when antibody titre reached a peak and when it was maintained at a high level after recovery. There was a rapid rise in antibody titre of the lgM fractions with the intensification of parasitaemia after infection. The titre declined in parallel with the number of parasites detected and was maintained at a low level until 6o days after infection. On the contrary, IgG antibody was first detected on the 1oth day, and its high titre persisted for more than 6o days, as well as that of total antibody. The experimental results obtained from naturally infected cows corresponded to those from the experimentally infected calves. The role of humoral antibody detected by IFA test in the inhibition of parasitaemia in cattle infected with T. sergenti is discussed. INTRODUCTION
In a previous report the application of the indirect fluorescent antibody (IFA) test to bovine piroplasmosis (Theileria sergenti infection of cattle) was described (Takahashi, Yamashita & Shimizu, 1972). A significant antibody titre was demonstrated in the serum of cattle during the 2-3 months after recovery. The recovered cattle resisted challenge infection with the homologous strain. This paper reports studies on the serological activities of the immunoglobulin fractions of sera collected from cattle infected with T. sergenti by using the IFA test. MATERIALS AND METHODS
Experimental animals Two male Holstein-Friesian calves, 15 days old (No. 86 and No. 88), were used in this experiment. To ensure that the calves were free from infection with
SEROLOGICAL RESPONSE IN BOVINE THEILERIOSIS
113
T. sergenti or any other blood parasite, smears were collected and stained with Giemsa stain for examination. Sera collected before infection were negative for antibody in the IFA test. The calves were infected by the intravenous inoculation of 50 ml of blood obtained from a splenectomized donor calf which had been suffering from severe parasitaemia after experimental infection. The donor's blood had been confirmed to be free from parasites other than T. sergenti and contained 1 ·5 X I0 10 parasites of this species. Blood samples were collected from both experimentally inoculated calves at 5-day intervals until 6o days after infection, and tested for parasitaemia, antibody level, and packed cell volume (PCV). Another five pastured cows which had recovered more than 1oo days following natural infection of T. sergenti were examined by the same method. Estimation of parasitaemia and PCV Parasitaemia was estimated by the microscopic examination of blood films stained with Giemsa stain. The degree of parasitaemia was expressed as a percentage of parasitized erythrocytes. PCV was examined in a capillary tube by centrifugation at I2 ooo rev/min for 5 min. Fractionation of serum Two ml samples of serum were fractionated by gel filtration through a 2·5 X 45 em column of Sephadex G-200 (Pharmacia, Uppsala, Sweden) equilibrated with o·I5 M- NaCl in o·oi M phosphate buffer solution (PBS). The effluent fraction was collected in a fraction collector at a flow rate of 2 ml per IO min and examined for optical density at 280 ml-' by a spectrophotometer. Fluorescent antibody technique Rabbit anti-bovine serum globulin was conjugated with fluorescein isothiocyanate using the method of Marshall, Eveland & Smith (I958). The conjugated globulin was passed through a gel filtration column and forced through a DEAE cellulose column. The conjugate obtained was stored at -20°C. The IFA test was carried out essentially by the same method as that described in the previous report (Takahashi et al., I972). The antigen was prepared from whole blood from a splenectomized calf experimentally infected with 10 per cent T. sergenti to induce parasitaemia. Erythrocytes were collected from the blood and washed three times in PBS. Thin films were made on coverslips 18 X I8 mm in size and dried, fixed in cold acetone for IO min and washed with o·oi M PBS (pH 7·2) for 5 min. Sera from infected calves were stored at -20°C until ready for use. Two-fold dilutions ranging from I: IO to I: 10 240 were prepared in tubes, and o·I ml of each dilution was placed on the antigen-coated coverslip. The coverslips were placed in a moist chamber and incubated at 37°C for 30 min and then washed with PBS for I5 min. The coverslip antigen was stained with conjugated rabbit anti-globulin at 37°0 for 30 min, rinsed in several changes of
BRITISH VETERINARY JOURNAL,
132, 1
PBS, mounted in 50 per cent buffered glycerin (pH g·o), and examined by the fluorescent microscope. The end-point was recorded as the highest dilution of serum giving specific fluorescence of the piroplasma, a titre of I : 20 or higher was considered significant.
Antibody activities of lgM and lgG Reading of absorbance at 280 mf' gave three peaks. The antibody titre of each fraction was measured by the IFA test. The antibody titres of the IgM and lgG fractions were calculated from the total of antibody titres ofthe first and second peak, respectively. RESULTS
The degree of parasitaemia, PCV, and antibody level in the two calves, are illustrated in Fig. I. The parasite began to appear in the blood 5 days after infection, and a marked rise in numbers occurred between IO and 20 days. The maximum rate of increase of infected erythrocytes during this period was I I per cent. The PCV fell to I3-I6 per cent when infected erythrocytes reached a maximum level. The increase of PCV coincided with the drop in number of infected erythrocytes thereafter. The antibody titre rose rapidly with the appearance of protozoa in the peripheral blood. When it reached a peak, parasitaemia tended to decrease gradually until 6o days after infection. An elevated plateau was maintained for more than 40 days. The markedly high antibody titre of IgM accompanied by the severest parasitaemia was observed 20 days after infection. Then IgM began to show a gradual decrease in level, but no shift to IgG antibody was observed until 6o days after infection. The pattern of IgG antibody production was parallel to that of serum antibody titres in all the serum samples. The number of parasites and the lgG antibody level increased more rapidly in calf No. 88 than in calf No. 86. Although there was a slight difference in the antibody response or parasitaemia between the two calves, the overall picture was essentially the same in both calves. Table I records the antibody titres, parasitaemia, and PCV obtained with serum or blood samples from cows naturally infected with T. sergenti. It is clear that the serum samples gave high antibody titres, while the value of parasitaemia ranged from 2·I to g·o. The antibody titre of the IgM fraction was higher in these cows than in experimentally infected calves in the convalescent stage. DISCUSSION
Calves experimentally infected with T. sergenti produced IgM and IgG antibodies detectable by the IFA test, but no evidence of a shift from IgM to IgG synthesis was demonstrated. Similar results were obtained with Babesia microti (Cox & Turner, I970a) and Plasmodium berghei yoelii in mice (Cox & Turner, I97ob). IgM antibody appeared earlier than IgG antibody in calves
SEROLOGICAL RESPONSE IN BOVINE THEILERIOSIS
115
~----(>---0
-,
·------·--..:----·
3 Calf No. 86
~·
30
~
'
..
' __ ',
0
> 0. 20
"'
-.
', ______ .._ _____
___....-l(~
x_____x
0 _J
.
--"'..............-
12 10
8
~ .Q
i
6 .e -~
4
10
&.
2
0
0
3
30
12
f
~ 0
> 20 0.
10~
E 0
"'0
a
0 .E
6
]
J
10
~
4 2
0
0
20
30
40
50
60
.o
Days after nfection
Fig. I. Antibody levels, parasitaemia and packed cell volume of calves experimentally infected with T. Sergenti. 0 - - 0 whole serum; .A----.A IgG; 0----0 parasitaemia; e ----e IgM; x- - x PCV. TABLE I ANTIBODY LEVELS, PARASITAEMIA AND PACKED CELL VOLUME OF COWS RECOVERED FROM NATURAL INFECTION WITH T. sergenti
Antibody titre Cow No.
Whole serum
lgM
lgG
Parasitaemia (%)
PCV* ( %)
I 2 3 4 5
640 2560 640 I280 I280
230 I30
670 3480 790 I500 II 50
3•2 2"I s·6 4"7 g·o
27 28 22 Ig 22
* Packed cell volume.
IOO
I8o 8o
116
BRITISH VETERINARY JOURNAL, 132, r
experimentally infected with T. sergenti. These findings were almost parallel with those from the observation on Plasmodium rynomologi and P. vivax in man (Abele et al., 1965). IgM and IgG antibodies produced by B. microti (Cox & Turner, 1970a) and P. berghei yoelii (Cox & Turner, 197ob), however were detected from mice at the same time after infection. In mice experimentally infected with B. microti, the protozoa disappeared completely from the peripheral blood when the IgG antibody level reached a peak (Cox & Turner, 1970a). In calves experimentally infected with T . sergenti and showing an lgG antibody level having reached a peak, low parasitaemia was observed until 6o days after infection. This suggested that there might be differences in IgG antibody activity among the species of erythrocytic protozoa. A decrease in the numbers of protozoa in erythrocytes and remarkable recovery from anaemia were seen with a rise of IgG antibody level in calves experimentally or naturally infected with T. sergenti. It seemed that humoral antibody might have inhibited the parasitaemia caused by T. sergenti infection. The investigation of P. berghei and B. rodhaini (Phillips, 1969) also indicated the inhibitory effect ofhumoral antibody on parasitaemia. Phillips &Jones (1972) described a transfer of resistance toP. berghei by passively administered antisera. This transfer resulted in the inhibition ofparasitaemia and brought about early recovery from anaemia. Since then, several workers have demonstrated a passive transfer of immunity with serum collected from infected animals. ACKNOWLEDGEMENT
We are particularly grateful to Dr W. J. Brinley Morgan, Ministry of Agriculture, Fisheries and Food, Central Veterinary Laboratory, Weybridge, England, for his advice. REFERENCES
ABELE, D. C., ToBIE,]. E ., HILL, G.]., CoNTAcos, P. G. & EvANs, C. B. (1965). Am.]. trop. Med. Hyg. 14, 191. Cox, F. E. G. & TuRNER, S. A. (1970a). Ann. trop. Med. Parasit. 64> 167. Cox, F. E. G. & TuRNER, S. A. (1970b). Ann. trop. Med. Parasit. 64, 175. MARSHALL, ]. D., EVELAND, W. C. & SMITH, C. W. (1958). Proc. Soc. exp. Biol. Med. !JS, 898. PHILLIPS, R . S. {1969). Parasitology, 59• 35 7. PHILLIPS, R. S. & joNEs, V. E. (I972). Parasitology, 64, 117. TAKAHASHI, K., YAMASHITA, S. & SHIMIZU, Y. (1972). Jap. J. Vet. Sci. 34> 275·
(Accepted for publication IB March I975)
Response serologique du betail infecte par Theileria serge11ti au test de detection des anticorps par inununofluorescence indirecte
(Takahashi et a/.) ResUDle. Des serums de deux veaux infectes experimentalement par Theileria sergenti ant ete analyses par immunofluorescence indirecte (IFI). Les anticorps apparurent au 5e jour apres l'infection et atteignirent un niveau maximum au 20e jour. On a note une diminution de Ia
SEROLOGICAL RESPONSE IN BOVINE THEILERIOSIS
; 117
parasitemie et une augmentation de la concentration globulaire en hemoglobine (CGH), lorsque le titre de l'anticorps atteignit un niveau maximum et il devait rester eleve durant la periode de guerison. II y eut une rapide elevation de la fraction IgM des anticorps avec une intensification de la parasitemie apres !'infection. Le titre devait decliner parallelement au nombre de parasites detectes et reste bas jusqu'a 6o jours apres !'infection. Par contre, les anticorps IgG detectes pour la lere fois au we jour avaient un titre eleve qui persista pendant 6o jours comme celui de !'ensemble des anticorps. Les resultats experimentaux obtenus sur des vaches naturellement infectees ont ete comparables a ceux des veaux experimentalement infectes. Le role de l'anticorps humoral detecte en IFI dans le freinage de la parasitemie du betail infecte au T. sergenti est discute. Serumsreaktionen bei Rindern, die mit Thei/erill sergenti infiziert worden waren und mit deDl indirekten Fluoreszenztest auf Antikorper untersucht wurden (Takahashi et a/.) Zusanunenfassung. Von zwei Kalbem, die experimentell mit Theileria sergenti infiziert worden waren, wurden die Sera mit dem indirekten Antikorperfluoreszenztest (IFA) auf Vorhandensein von Antikorpern untersucht. Antikorper zeigten sich am 5· Tag nach der Infektion und erreichten einen Hohepunkt am 20. Tag. Wenn der Antikorpertiter einen Hohepunkt erreichte und nach der Wiederstellung hoch blieb, wurde eine Verminderung der Parasitaemie und eine Vermehrung des Zellvolumen nach Sedimentation (PVC) festgestellt. Mit der erhohten Parasitaemie nach der Infektion ging ein rapider Anstieg des Antikorpertiters der lgM-Anteile einher. Parallel zu der Anzahl der gefundenen Parasiten nahm der Titer ab und hielt sich in verminderter Hohe his zu 6o Tagen nach der lnfektion. Im Gegensatz hierzu wurde der lgG-Antikorper zuerst am 1o.Tage entdeckt, und sein hoher Titer hielt 6o Tage lang an ebenso wie der des gesamten Antikorpers. Bei unter natiirlichen Umstanden infizierten Kiihen entsprachen die Resultate den bei experimentell infizierten Kalbern beobachteten. Es wird erortert, welche Rolle der durch den !FA-Test entdeckte humorale Antikorper bei lnhibierung der bei mit T. sergenti infizierten Rind ern auftretende Parasitaemie spielt. Respuesta serol6gica a Ia prueba indirecta de anticuerpos fluorescentes en el ganado infectado con Theileria sergenti (Takahashi et a/.) Resmnen. Se comprobaron los anticuerpos del suero de dos terneros, infectados experimentalmente con Theileria sergenti, mediante la prueba indirecta de anticuperos fluorescentes (IAF). Los anticuerpos aparecieron el 5° dia despues de la infecci6n y alcanzaron un maximo en el 20° dia. Se observaron una disminuci6n en la gravedad de la parasitemia y un aumento en el volumen de celulas concentradas cuando el valor de anticuerpos alcanz6 un maximo y cuando se mantuvo a un nivel elevado despues de la recuperaci6n. Se di6 una r:ipida subida del valor de anticuerpos de las fracciones IgM con la intensificaci6n de parasitemia despues de la infecci6n. El valor declin6 en paralelo con el numero de par:isitos descubierto y se mantuvo a un nivel bajo hasta 6o dias despues de la infecci6n. Por el contrario, el anticuerpo lgG se descubri6 por primera vez el 10° dia, y su valor elevado persisti6 por 6o dias, ademas del de los anticuerpos totales. Los resultados experimentales obtenidos de vacas infectadas de forma natural correspondieron con los obtenidos de temeros infectados experimentalmente. Se discute la funci6n del anticuerpo humoral, descubierto por la prueba IAF, en la inhibici6n de parasitemia en el ganado infectado con T. sergenti.
