Assessment of an Enzyme-linked Immunosorbent Assay Using a Taenia hydatigena Fraction Antigen in the Diagnosis of Cysticercosis in Cattle H.J. Smith, K.E. Snowdon, D. Gregory and G.G. Finley
ABSTRACT
presence d'anticorps dans le serum des bovins porteurs ne semble pas donner Enzyme linked immunosorbent des resultats fiables, du moins chez des assay (ELISA) using a fraction of animaux faiblement infestes. larval Taenia hydatigena cyst fluid antigen was carried out on 469 bovine sera collected at slaughter from feedlot Early applications of enzyme-linked cattle for the presence of immunosorbent assay (ELISA) to anticysticercosis antibodies. diagnose cysticercosis in cattle gave Cysticerci, in low numbers, were disappointing results due to a lack of found in the heart, tongue and/or the necessary sensitivity and/or masseter muscles of 84 of the 469 cattle specificity of the antigenic reagents at postmortem inspection. Only nine available (1-3). This is due in part to sera gave positive ELISA reactions the extensive antigenic overlap that and in only one of these nine animals exists not only between cestodes but were cysticerci found. Within the also between cestodes and other limitations of this study, the high rate parasites and, in part, to the moderate of false negative and false positive antibody response of infected animals reactions suggests that the ELISA to cestodes. Rhoads and colleagues with the antigen used is not a (4,5) recently showed that a fraction of satisfactory procedure to diagnose larval Taenia hydatigena cyst fluid cysticercosis in cattle, at least in (ThFAS antigen) has sensitivity and animals with light infections. specificity in the ELISA for the detection of anti-Taenia saginata antibodies. In experimental calves this test RESUME detected IgG and IgM antibody levels by the third week after infection until Une epreuve serologique (a l'aide all were slaughtered 13 to 26 weeks d'une technique ELISA), mise au after infection (5). Late in 1985, an point afin de deceler les bovins ongoing outbreak of cysticercosis was porteurs de la cysticercose bovine, a diagnosed in a feedlot in Ontario ete veriflee sur 469 serums de bovins which provided an opportunity to provenant de parquets d'engrais- evaluate an ELISA under field sement. Un faible nombre de conditions. From May to November 1987, sera cysticerques a ete retrouve dans le coeur, la langue et/ou les muscles were collected from 469 beef cattle at masseters chez 84 des 469 bovins slaughter. The carcasses were rouexamines. Seulement neuf serums ont tinely inspected for the presence of reagi positivement a l'epreuve ELISA cysticerci by incising masseter, tongue et seulement un seul de ces neuf and heart musculature. Sera were animaux etait porteur de la maladie. examined in duplicate using a tripleLes resultats de cette etude suggerent antibody ELISA with the ThFAS que l'antigene utilise afin de detecter la antigen and rabbit antibovine IgG
(heavy and light chain specific) antiserum as described by Rhoads and colleagues (4,5). Three positive and three negative sera were examined on each plate. Positive control sera from experimentally infected calves were provided by the Animal Parasitology Institute, USDA, Beltsville, Maryland, while negative control sera were collected from cysticercosis-free cattle of approximately the same age as the animals under study. An optical density (OD) reading at 405 nm > 5 times the mean of the negative sera was considered to be positive. At slaughter cysticercosis lesions confirmed by parasitological and histological examinations were found in the heart, tongue and/or masseter muscles of 84 of the 469 animals examined. The number of lesions found in individual animals was low, usually in the range of one to three cysts. Serologically, only 9 of the 469 animals gave OD readings that were considered positive for anticysticercosis antibodies. Cysticerci were only found in one of the nine animals that reacted positively. Even though lesions were found in approximately 18% of the cattle at slaughter, it is obvious that the infections were light. The fact that no lesions were found in eight of the nine cattle that gave a positive serological reaction suggests that a high proportion of the cattle had been infected at one time, or that they were infected but the cysticerci were not found. The positive control sera gave consistently high OD readings while the negative sera gave low readings indicating that the ELISA detected
Agriculture Canada, Health of Animals Laboratory, P.O. Box 1410, Sackville, New Brunswick EOA Canada, Animal Health Division, 2255 Carling Avenue, Ottawa, Ontario KIA 0Y9 (Gregory). Submitted September 8, 1988.
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anticysticercosis antibodies if present. The findings further suggest that development of anticysticercosis antibodies is dependent, at least, on the magnitude of the infection established and perhaps also on the age of the infection when the animal is tested. Subsequent studies carried out with experimentally infected cattle confirmed that level of antibody is dose related and develops more slowly in lightly infected animals (Smith, unpublished observations). The high rate of false negative (serologically negative animals known to have cysticerci in the musculature) and false positive (serologically positive animals in which cysticerci were not demonstrated) reactions indicate that the ELISA using the ThFAS antigen has limited usefulness in the diagnosis of light cysticercosis infections in cattle.
300
Fewer false positives might have been found in this study had the carcasses been thoroughly dissected rather than subjected to routine meat inspection procedures. ACKNOWLEDGMENTS The authors gratefully acknowledge the cooperation of Ms. Marcia Rhoads, API, USDA, Beltsville, Maryland in providing ThFAS antigen and positive control sera to carry out this study. The assistance of Dr. K.L. Easton, Meat Hygiene Division, Food Production and Inspection Branch, Agriculture Canada, Toronto, Ontario and his staff in the inspection of the cattle at slaughter and the collection of tissue and serum samples is sincerely appreciated. The technical assistance of Ms. JoAnn Daniels in the histological preparation of tissues is much
appreciated.
REFERENCES 1. HARRISON LJS, SEWELL MMH. Antibody levels in cattle naturally infected with Taenia saginata metacestodes in Britain. Res Vet Sci 1981; 31: 62-64. 2. HARRISON LJS, HOLT K, SEWELL MMH. Serum antibody levels to Taenia saginata in cattle grazed on Scottish pastures. Res Vet Sci 1986; 40: 344-346. 3. GEERT S, KUMAR V, AERTS N, CEULEMANS F. Comparative evaluation of immunoelectrophoresis, counterimmunoelectrophoresis and enzyme linked immunosorbent assay for the diagnosis of Taenia saginata cysticercosis. Vet Parasitol 1981; 8: 299-307. 4. RHOADS ML, MURRELL KD, DILLING GW, WONG MM, BAKER NF. A potential diagnostic reagent for bovine cysticercosis. J Parasitol 1985; 71: 779-787. 5. KAMANGA-SOLLO EIF, RHOADS ML, MURRELL KD. Evaluation of an antigenic fraction of Taenia hydatigena metacestode cyst fluid for immunodiagnosis of bovine cysticercosis. Am J Vet Res 1987; 48: 12061210.
ABSTRACT
presence d'anticorps dans le serum des bovins porteurs ne semble pas donner Enzyme linked immunosorbent des resultats fiables, du moins chez des assay (ELISA) using a fraction of animaux faiblement infestes. larval Taenia hydatigena cyst fluid antigen was carried out on 469 bovine sera collected at slaughter from feedlot Early applications of enzyme-linked cattle for the presence of immunosorbent assay (ELISA) to anticysticercosis antibodies. diagnose cysticercosis in cattle gave Cysticerci, in low numbers, were disappointing results due to a lack of found in the heart, tongue and/or the necessary sensitivity and/or masseter muscles of 84 of the 469 cattle specificity of the antigenic reagents at postmortem inspection. Only nine available (1-3). This is due in part to sera gave positive ELISA reactions the extensive antigenic overlap that and in only one of these nine animals exists not only between cestodes but were cysticerci found. Within the also between cestodes and other limitations of this study, the high rate parasites and, in part, to the moderate of false negative and false positive antibody response of infected animals reactions suggests that the ELISA to cestodes. Rhoads and colleagues with the antigen used is not a (4,5) recently showed that a fraction of satisfactory procedure to diagnose larval Taenia hydatigena cyst fluid cysticercosis in cattle, at least in (ThFAS antigen) has sensitivity and animals with light infections. specificity in the ELISA for the detection of anti-Taenia saginata antibodies. In experimental calves this test RESUME detected IgG and IgM antibody levels by the third week after infection until Une epreuve serologique (a l'aide all were slaughtered 13 to 26 weeks d'une technique ELISA), mise au after infection (5). Late in 1985, an point afin de deceler les bovins ongoing outbreak of cysticercosis was porteurs de la cysticercose bovine, a diagnosed in a feedlot in Ontario ete veriflee sur 469 serums de bovins which provided an opportunity to provenant de parquets d'engrais- evaluate an ELISA under field sement. Un faible nombre de conditions. From May to November 1987, sera cysticerques a ete retrouve dans le coeur, la langue et/ou les muscles were collected from 469 beef cattle at masseters chez 84 des 469 bovins slaughter. The carcasses were rouexamines. Seulement neuf serums ont tinely inspected for the presence of reagi positivement a l'epreuve ELISA cysticerci by incising masseter, tongue et seulement un seul de ces neuf and heart musculature. Sera were animaux etait porteur de la maladie. examined in duplicate using a tripleLes resultats de cette etude suggerent antibody ELISA with the ThFAS que l'antigene utilise afin de detecter la antigen and rabbit antibovine IgG
(heavy and light chain specific) antiserum as described by Rhoads and colleagues (4,5). Three positive and three negative sera were examined on each plate. Positive control sera from experimentally infected calves were provided by the Animal Parasitology Institute, USDA, Beltsville, Maryland, while negative control sera were collected from cysticercosis-free cattle of approximately the same age as the animals under study. An optical density (OD) reading at 405 nm > 5 times the mean of the negative sera was considered to be positive. At slaughter cysticercosis lesions confirmed by parasitological and histological examinations were found in the heart, tongue and/or masseter muscles of 84 of the 469 animals examined. The number of lesions found in individual animals was low, usually in the range of one to three cysts. Serologically, only 9 of the 469 animals gave OD readings that were considered positive for anticysticercosis antibodies. Cysticerci were only found in one of the nine animals that reacted positively. Even though lesions were found in approximately 18% of the cattle at slaughter, it is obvious that the infections were light. The fact that no lesions were found in eight of the nine cattle that gave a positive serological reaction suggests that a high proportion of the cattle had been infected at one time, or that they were infected but the cysticerci were not found. The positive control sera gave consistently high OD readings while the negative sera gave low readings indicating that the ELISA detected
Agriculture Canada, Health of Animals Laboratory, P.O. Box 1410, Sackville, New Brunswick EOA Canada, Animal Health Division, 2255 Carling Avenue, Ottawa, Ontario KIA 0Y9 (Gregory). Submitted September 8, 1988.
3C0 (Smith, Snowdon, Finley) and Agriculture
Can J Vet Res 1990; 54: 299-300
299
anticysticercosis antibodies if present. The findings further suggest that development of anticysticercosis antibodies is dependent, at least, on the magnitude of the infection established and perhaps also on the age of the infection when the animal is tested. Subsequent studies carried out with experimentally infected cattle confirmed that level of antibody is dose related and develops more slowly in lightly infected animals (Smith, unpublished observations). The high rate of false negative (serologically negative animals known to have cysticerci in the musculature) and false positive (serologically positive animals in which cysticerci were not demonstrated) reactions indicate that the ELISA using the ThFAS antigen has limited usefulness in the diagnosis of light cysticercosis infections in cattle.
300
Fewer false positives might have been found in this study had the carcasses been thoroughly dissected rather than subjected to routine meat inspection procedures. ACKNOWLEDGMENTS The authors gratefully acknowledge the cooperation of Ms. Marcia Rhoads, API, USDA, Beltsville, Maryland in providing ThFAS antigen and positive control sera to carry out this study. The assistance of Dr. K.L. Easton, Meat Hygiene Division, Food Production and Inspection Branch, Agriculture Canada, Toronto, Ontario and his staff in the inspection of the cattle at slaughter and the collection of tissue and serum samples is sincerely appreciated. The technical assistance of Ms. JoAnn Daniels in the histological preparation of tissues is much
appreciated.
REFERENCES 1. HARRISON LJS, SEWELL MMH. Antibody levels in cattle naturally infected with Taenia saginata metacestodes in Britain. Res Vet Sci 1981; 31: 62-64. 2. HARRISON LJS, HOLT K, SEWELL MMH. Serum antibody levels to Taenia saginata in cattle grazed on Scottish pastures. Res Vet Sci 1986; 40: 344-346. 3. GEERT S, KUMAR V, AERTS N, CEULEMANS F. Comparative evaluation of immunoelectrophoresis, counterimmunoelectrophoresis and enzyme linked immunosorbent assay for the diagnosis of Taenia saginata cysticercosis. Vet Parasitol 1981; 8: 299-307. 4. RHOADS ML, MURRELL KD, DILLING GW, WONG MM, BAKER NF. A potential diagnostic reagent for bovine cysticercosis. J Parasitol 1985; 71: 779-787. 5. KAMANGA-SOLLO EIF, RHOADS ML, MURRELL KD. Evaluation of an antigenic fraction of Taenia hydatigena metacestode cyst fluid for immunodiagnosis of bovine cysticercosis. Am J Vet Res 1987; 48: 12061210.
