ERA STRAIN® RABIES VACCINE: ORAL VACCINATION OF NONHUMAN PRIMATES K. F. Lawson, M. J. Walcroft and J. F. Crawley*
intracerebrally into mice at a dosage of 0.03 ml. The animals were observed over a 21 day period and the results recorded as the reciprocal of the dilution of serum which protected 50% of the mice against 50-100 LD50 of CVS virus. Virus titrations were carried out by inoculating 0.03 ml of the tenfold dilutions intracerebrally into mice weighing 10-12 grams. The street virus used for challenging the monkeys was prepared as a 20% suspension of finely ground, salivary glands from rabiespositive foxes received from Agriculture Canada. The virus was dispensed into glass ampoules which were sealed and frozen at -200C until used. The challenge pool contained approximately 107 mouse LD50 per 2 ml. Following an initial titration, the pool was diluted to contain approximately 105.6 mouse LD50 per 2 ml for the challenge of vaccinated monkeys. This was administered by multiple injections into the neck muscles. The diluent for the virus and serum dilutions was distilled water to which 2% normal horse serum, 500 units of penicillin and 1 mg of streptomycin per 1 ml were added.
INTRODUCTION
THERE HAS BEEN A GREAT DEAL of interest during the past few years in the vaccination of animals against rabies, using the oral route of administration (1, 2, 3, 4, 5, 7, 11). These publications have dealt primarily with the immunization of wildlife. Wiktor and others (6, 9, 10) have shown that primates can be successfully immunized against rabies by various vaccines when inoculated by the parenteral route. This paper reports on the susceptibility of the rhesus monkey to street rabies virus and the immunization of this species against rabies using a modified live virus vaccine ERA Strain® administered by the oral route. MATERIALS AND METHODS
Animals
Thirty-six rhesus monkeys (M. mulatta) were obtained from a commercial supplier' and stabilized for a period of 56 days before being allocated to the experiment. Only tuberculin negative monkeys were used. White swiss mice (Connaught strain), 1012 grams, approximately four weeks of age were used in the serum neutralization test and virus titrations.
Inoculation of Animals In preparation for vaccination and challenge the monkeys were immobilized with phencyclidine hydrochloride2 administered at a dosage of 0.2 mg per kg. For vaccination, the animal's mouth was held open and 1 ml of the vaccine was administered on to the tongue. The animal was maintained in an upright position until the vaccine had been swallowed. The challenge of 2 ml was injected into the lateral neck muscles, 0.4 ml into each of five sites within a circumscribed area measuring approximately 3 cm. In Experiment 1, three groups, containing four monkeys in each, received various dilutions of street rabies virus. The challenge was diluted from 10-1 to 10-3 and the dose was 2 ml administered as described above in the lateral neck muscles. In Experiment 2, an oral vaccination trial
Viruses
The rabies vaccine used in the trial was a regular production serial of ERA Strain® rabies vaccine produced in primary porcine kidney cell cultures. The regular diluent supplied with the vaccine was used for reconstitution. The challenge virus strain (CVS) used in the serum neutralization test was originally received from the National Institutes of Health, United States Public Health Service and has been propagated in mouse brains. The test consisted of preparing fivefold dilutions of serum to which rabies virus containing 50-100 LD50 was added. After incubation at 37°C for 90 minutes, this mixture was inoculated 'Connaught Laboratories Limited, 1755 Steeles
Avenue West, Willowdale, Ontario (Lawson and Walcroft) and R.R. #6, Guelph, Ontario (Craw-
2Sernylan, Bio-Ceutic Laboratories, Inc., St. Joseph, Missouri.
ley). lPrimate Imports Corp., New York.
t55 CAN. VET. JOUR., vol. 17, no. 10, October, 1976
CANADIAN VETERINARY JOURNAL
was carried out, in which 30 monkeys were assigned at random to five groups of six. Groups I and II received ERA vaccine containing 106.3 mouse LD50 while Groups III and IV received 1082 mouse LD50 of vaccine. Groups II and IV received, three weeks later, a second dose of vaccine which contained 106.7 and 1084 mouse LD50 respectively. The vaccine in all cases was administered by the oral route, as described above. Group V did not receive any vaccine and acted as control animals for the trial. The animals were challenged 194 days after vaccination with virulent rabies virus containing 105.6 mouse LD50 per 2 ml.
Samples Blood samples were taken before vaccination and at three, seven and 22 weeks after vaccination. The serum was removed and stored at -200C until required and then inactivated at 56°C for 30 minutes prior to use. The 50% end points for titration and serum neutralization tests were calculated according to the method of Reed and Muench (8). RESULTS
Table I shows the results obtained in Experiment 1 when the challenge virus was titrated in monkeys. The 10-3 dilution of the rabies challenge virus, which contained 1046 mouse LD50 per 2 ml killed three of the four monkeys on test. The results of the serum neutralization tests carried out on samples taken from the monkeys in Experiment 2 are shown in Table II. It should be noted that although the titre of the vaccine dose influenced the number of animals showing a serum conversion, the second dose of vaccine did not increase the antibody response. Table III shows the results obtained when the vaccinated and unvaccinated monkeys were challenged. While 67% of the control monkeys died on challenge, only two of 24 (8.3%) vaccinated monkeys died of rabies. These animals, numbers 621 and 690, died 22 and 48 days respectively, post-challenge. They had received the lower vaccine dosage and did not develop an antibody titre following vaccination. DISCUSSION
results of this investigation and similar ones carried out at these laboratories, with the same challenge pool, it appears that monkeys are more susceptible to rabies than dogs, cats, cattle and horses. Whether or not this is due to the administration of multiple injections of the challenge virus in the lateral neck muscles is not known at this time. Deaths occurred between 12 and 47 days with a mean of 20.3 days. Primates can be immunized successfully by the administration of ERA Strain® of rabies vaccine by the oral route as was shown by serum neutralization and challenge tests. The amount of virus in the vaccine at the time of administration influenced the antibody response, whereas a second vaccination did not increase the antibody levels in these groups. Two monkeys died during the investigation. One, number 622, died 22 days after the second dose of vaccine. The cause of death was attributed to hemorrhage from a ruptured aorta. The brain was removed and found to be negative by the fluorescent antibody test. Monkey number 608 died two days after challenge without showing any symptoms of rabies. Death was attributed to acute gastric dilation. Unfortunately, the brain tissue collected from this animal was not suitable for examination. SUMMARY
The investigations carried out showed that rhesus monkeys (Macaca mulatta) were very susceptible to a challenge with street rabies virus administered by multiple injections in the neck muscles. Monkeys may be successfully immunized by a single dose of the ERA Strain® of rabies vaccine given by the oral route.
RESUM'E Les experiences decrites dans le present article ont demontre, de la part du singe rhesus (Macaca mulatta), une tres grande susceptibilite a la rage, suite a des injections multiples du virus des rues dans la musculature du cou. Une seule dose orale du vaccin antirabique prepar6 avec la souche ERA suffit pour immuniser des singes avec succes. ACKNOWLEDGMENTS
Rhesus monkeys (M. mulatta) are very susThe authors wish to express their appreciation ceptible to street rabies virus when challenged to Dr. R. J. Avery, Director of the Animal Diseases by the method described. A dilution contain- Research Institute of Agriculture Canada, who ing 104.6 mouse LD50 was capable of killing provided the fox salivary glands for preparing three of four monkeys in the titration. From the stock challenge virus. We also gratefully ac256
ERA
STRAINOf
TABLE I TITRATION OF STREET RABIES VIRUS IN MONKEYS
Brain Examination F.A.b Mouse" 106.6 702 Died day 12 + 2.7 703 Died day 14 + 3.5 704 Died day 13 + 3 705 Died day 16 + 3 10-2 105.6 647 Died day 22 + >4 698 Died day 19 + 3.4 699 Died day 19 + 3.9 700 Died day 16 >4 + 104.6 10-3 640 Died day 25 + 2.5 672 Survived N.D. N.D. 711 Died day 47 + >4 712 Died day 20 + 3.2 in Challenge: Administered 2 ml amounts by multiple injection in the neck muscles. "Recorded as mouse LD5o per 2 ml (logio). bF.A. + = fluorescent antibody-positive for rabies. "Recorded as mouse LD50 per 0.03 ml intracerebral inoculation into mice weighing 10 to 12 grams (logio). N.D. - Not Done. Virus Dilution 10-1
Virus Titrea
Monkey Number
Results
TABLE II RESULTS OF RABIES SERUM NEUTRALIZATION TESTS OF MONKEYS VACCINATED BY THE ORAL ROUTE WITH ERA RABIES VACCINE Serum Neutralization Titreb
Vaccine Titrea Group I
1st Dose
2nd Dose
6.3
None
II
6.3
6.7
III
8.2
None
IV
8.2
8.4
None
None
V
Monkey Number
Pre Vacc.
612 655 688 689 690 710 608 619 621 622 654 709 602 603 606 662 664 708 615 616 618 623 648 656 682 691 692 694 695
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
3 wks. 160 625 10 1 0 187 0 224 0 238 98 0 625 625 625 25 84 187 1
84 178 328 84 625 N.D. N.D. N.D. N.D. N.D. N.D.
Postvaccination 7 wks. 22 wks. 440 56 41 3 0 71 56 203 0 48 35 0 625 229 625 49 187 625 3 285 625 328 153 625 N.D. N.D. N.D. N.D. N.D.
625 153 1 1 0 625 280 625 0 N.D. 66 0 488 625 625 70 328 625 0 379 625 245 280 178 N.D. N.D. N.D. N.D. N.D.
N.D. N.D. 697 N.D. Not Done. "Recorded as the number of mouse LD50 administered to each animal (logio). bRecorded as reciprocal of the dilution which protects 50% of mice against 100 LD50 of CVS virus. 257
CANADIAN VETERINARY JOURNAL
TABLE III CHALLENGE RESULTS OF MONKEYS VACCINATED 6 MONTHS PREVIOUSLY WITH ERA RABIES VACCINE BY THE ORAL ROUTE
Group
Results
Monkey
I
F.A. ResultSa
612 Survived Survived 655 Survived 688 Survived 689 690 Died day 48 postchallenge Survived 710 II 608 Died day 2 postchallenge 619 Survived 621 Died day 22 postchallenge 622 Died prechallenge Survived 654 Survived 709 III 602 Survived 603 Survived 606 Survived 662 Survived 664 Survived Survived 708 Survived IV 615 Survived 616 Survived 618 Survived 623 Survived 648 Survived 656 V Died day 53 postchallenge 682 Died day 21 postchallenge 691 Died day 30 postchallenge 692 Survived 694 Died day 13 postchallenge 695 Survived 697 N.D. Not Done. Challenge: 106-2 mouse LD50 per 2 ml inoculated by multiple in the neck muscles. aF.A. + = fluorescent-antibody test positive for rabies.
knowledge the technical assistance of Miss E. Gaynor, and Messrs. D. Flemming and R. Gertus.
1. 2.
3.
4.
5.
REFERENCES BAER, G. M., M. K. ABELSETH and J. G. DEBBIE. Oral vaccination of foxes against rabies. Am. J. Epidem. 93: 487-490. 1971. BAER, G. M., J. R. BRODERSON and P. A. YAGER. Determination of the site of oral rabies vaccination. Am. J. Epidem. 101: 160164. 1975. BLACK, J. G. and K. F. LAWSON. Sylvatic rabies studies in the silver fox (Vulpes vulpes). Susceptibility and immune response. Can. J. comp. Med. 34: 309-311. 1970. BLACK, J. G. and K. F. LAWSON. Further studies of sylvatic rabies in the fox (Vulpes vulpes). Vaccination by the oral route. Can. vet. J. 14: 206-211. 1973. DEBBIE, J. G., M. K. ABELSETH and G. M. BAER. The use of commercially available vaccines for the oral vaccination of foxes against rabies. Am. J. Epidem. 96: 231-235.
1972.
258
N.D. N.D. N.D. N.D. + N.D. N.D. N.D. +
N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. + + + N.D. + N.D.
injection
6. LAVENDER, J. F. Immune response in primates vaccinated with duck embryo cell culture rabies vaccine. Appl. Microbiol. 25: 327331. 1973.
7. MAYR, A., H. KRAFT, 0. JAEGER and J. HAACKE. Orale Immunisierung von Fuchsen gegen Tollwut. Zentbl. VetMed. [B] 19: 615625. 1972. 8. REED, L.S. and H. MUENCH. A simple method of estimating 50 per cent endpoints. Am. J. Hyg. 27: 493-497. 1938. 9. SIKEs, R. K., W. F. CLEARY, H. KOPROWSKI, T. J. WIKTOR and M. M. KAPLAN. Effective protection of monkeys against death from street virus by post-exposure administration of tissue-culture rabies vaccine. Bull. Wld Hlth Org. 45: 1-11. 1971. 10. WIKTOR, T. J. and H. KoPRowsiu. Successful immunization of primates with rabies vaccine prepared in human diploid cell strain WI-38. Proc. Soc. exp. Biol. Med. 118: 10691073. 1965. 11. WINKLER, W. G., R. G. McLEAN and J. C. COWART. Vaccination of foxes against rabies using ingested baits. J. Wildl. Dis. 11:
382-388. 1975.
intracerebrally into mice at a dosage of 0.03 ml. The animals were observed over a 21 day period and the results recorded as the reciprocal of the dilution of serum which protected 50% of the mice against 50-100 LD50 of CVS virus. Virus titrations were carried out by inoculating 0.03 ml of the tenfold dilutions intracerebrally into mice weighing 10-12 grams. The street virus used for challenging the monkeys was prepared as a 20% suspension of finely ground, salivary glands from rabiespositive foxes received from Agriculture Canada. The virus was dispensed into glass ampoules which were sealed and frozen at -200C until used. The challenge pool contained approximately 107 mouse LD50 per 2 ml. Following an initial titration, the pool was diluted to contain approximately 105.6 mouse LD50 per 2 ml for the challenge of vaccinated monkeys. This was administered by multiple injections into the neck muscles. The diluent for the virus and serum dilutions was distilled water to which 2% normal horse serum, 500 units of penicillin and 1 mg of streptomycin per 1 ml were added.
INTRODUCTION
THERE HAS BEEN A GREAT DEAL of interest during the past few years in the vaccination of animals against rabies, using the oral route of administration (1, 2, 3, 4, 5, 7, 11). These publications have dealt primarily with the immunization of wildlife. Wiktor and others (6, 9, 10) have shown that primates can be successfully immunized against rabies by various vaccines when inoculated by the parenteral route. This paper reports on the susceptibility of the rhesus monkey to street rabies virus and the immunization of this species against rabies using a modified live virus vaccine ERA Strain® administered by the oral route. MATERIALS AND METHODS
Animals
Thirty-six rhesus monkeys (M. mulatta) were obtained from a commercial supplier' and stabilized for a period of 56 days before being allocated to the experiment. Only tuberculin negative monkeys were used. White swiss mice (Connaught strain), 1012 grams, approximately four weeks of age were used in the serum neutralization test and virus titrations.
Inoculation of Animals In preparation for vaccination and challenge the monkeys were immobilized with phencyclidine hydrochloride2 administered at a dosage of 0.2 mg per kg. For vaccination, the animal's mouth was held open and 1 ml of the vaccine was administered on to the tongue. The animal was maintained in an upright position until the vaccine had been swallowed. The challenge of 2 ml was injected into the lateral neck muscles, 0.4 ml into each of five sites within a circumscribed area measuring approximately 3 cm. In Experiment 1, three groups, containing four monkeys in each, received various dilutions of street rabies virus. The challenge was diluted from 10-1 to 10-3 and the dose was 2 ml administered as described above in the lateral neck muscles. In Experiment 2, an oral vaccination trial
Viruses
The rabies vaccine used in the trial was a regular production serial of ERA Strain® rabies vaccine produced in primary porcine kidney cell cultures. The regular diluent supplied with the vaccine was used for reconstitution. The challenge virus strain (CVS) used in the serum neutralization test was originally received from the National Institutes of Health, United States Public Health Service and has been propagated in mouse brains. The test consisted of preparing fivefold dilutions of serum to which rabies virus containing 50-100 LD50 was added. After incubation at 37°C for 90 minutes, this mixture was inoculated 'Connaught Laboratories Limited, 1755 Steeles
Avenue West, Willowdale, Ontario (Lawson and Walcroft) and R.R. #6, Guelph, Ontario (Craw-
2Sernylan, Bio-Ceutic Laboratories, Inc., St. Joseph, Missouri.
ley). lPrimate Imports Corp., New York.
t55 CAN. VET. JOUR., vol. 17, no. 10, October, 1976
CANADIAN VETERINARY JOURNAL
was carried out, in which 30 monkeys were assigned at random to five groups of six. Groups I and II received ERA vaccine containing 106.3 mouse LD50 while Groups III and IV received 1082 mouse LD50 of vaccine. Groups II and IV received, three weeks later, a second dose of vaccine which contained 106.7 and 1084 mouse LD50 respectively. The vaccine in all cases was administered by the oral route, as described above. Group V did not receive any vaccine and acted as control animals for the trial. The animals were challenged 194 days after vaccination with virulent rabies virus containing 105.6 mouse LD50 per 2 ml.
Samples Blood samples were taken before vaccination and at three, seven and 22 weeks after vaccination. The serum was removed and stored at -200C until required and then inactivated at 56°C for 30 minutes prior to use. The 50% end points for titration and serum neutralization tests were calculated according to the method of Reed and Muench (8). RESULTS
Table I shows the results obtained in Experiment 1 when the challenge virus was titrated in monkeys. The 10-3 dilution of the rabies challenge virus, which contained 1046 mouse LD50 per 2 ml killed three of the four monkeys on test. The results of the serum neutralization tests carried out on samples taken from the monkeys in Experiment 2 are shown in Table II. It should be noted that although the titre of the vaccine dose influenced the number of animals showing a serum conversion, the second dose of vaccine did not increase the antibody response. Table III shows the results obtained when the vaccinated and unvaccinated monkeys were challenged. While 67% of the control monkeys died on challenge, only two of 24 (8.3%) vaccinated monkeys died of rabies. These animals, numbers 621 and 690, died 22 and 48 days respectively, post-challenge. They had received the lower vaccine dosage and did not develop an antibody titre following vaccination. DISCUSSION
results of this investigation and similar ones carried out at these laboratories, with the same challenge pool, it appears that monkeys are more susceptible to rabies than dogs, cats, cattle and horses. Whether or not this is due to the administration of multiple injections of the challenge virus in the lateral neck muscles is not known at this time. Deaths occurred between 12 and 47 days with a mean of 20.3 days. Primates can be immunized successfully by the administration of ERA Strain® of rabies vaccine by the oral route as was shown by serum neutralization and challenge tests. The amount of virus in the vaccine at the time of administration influenced the antibody response, whereas a second vaccination did not increase the antibody levels in these groups. Two monkeys died during the investigation. One, number 622, died 22 days after the second dose of vaccine. The cause of death was attributed to hemorrhage from a ruptured aorta. The brain was removed and found to be negative by the fluorescent antibody test. Monkey number 608 died two days after challenge without showing any symptoms of rabies. Death was attributed to acute gastric dilation. Unfortunately, the brain tissue collected from this animal was not suitable for examination. SUMMARY
The investigations carried out showed that rhesus monkeys (Macaca mulatta) were very susceptible to a challenge with street rabies virus administered by multiple injections in the neck muscles. Monkeys may be successfully immunized by a single dose of the ERA Strain® of rabies vaccine given by the oral route.
RESUM'E Les experiences decrites dans le present article ont demontre, de la part du singe rhesus (Macaca mulatta), une tres grande susceptibilite a la rage, suite a des injections multiples du virus des rues dans la musculature du cou. Une seule dose orale du vaccin antirabique prepar6 avec la souche ERA suffit pour immuniser des singes avec succes. ACKNOWLEDGMENTS
Rhesus monkeys (M. mulatta) are very susThe authors wish to express their appreciation ceptible to street rabies virus when challenged to Dr. R. J. Avery, Director of the Animal Diseases by the method described. A dilution contain- Research Institute of Agriculture Canada, who ing 104.6 mouse LD50 was capable of killing provided the fox salivary glands for preparing three of four monkeys in the titration. From the stock challenge virus. We also gratefully ac256
ERA
STRAINOf
TABLE I TITRATION OF STREET RABIES VIRUS IN MONKEYS
Brain Examination F.A.b Mouse" 106.6 702 Died day 12 + 2.7 703 Died day 14 + 3.5 704 Died day 13 + 3 705 Died day 16 + 3 10-2 105.6 647 Died day 22 + >4 698 Died day 19 + 3.4 699 Died day 19 + 3.9 700 Died day 16 >4 + 104.6 10-3 640 Died day 25 + 2.5 672 Survived N.D. N.D. 711 Died day 47 + >4 712 Died day 20 + 3.2 in Challenge: Administered 2 ml amounts by multiple injection in the neck muscles. "Recorded as mouse LD5o per 2 ml (logio). bF.A. + = fluorescent antibody-positive for rabies. "Recorded as mouse LD50 per 0.03 ml intracerebral inoculation into mice weighing 10 to 12 grams (logio). N.D. - Not Done. Virus Dilution 10-1
Virus Titrea
Monkey Number
Results
TABLE II RESULTS OF RABIES SERUM NEUTRALIZATION TESTS OF MONKEYS VACCINATED BY THE ORAL ROUTE WITH ERA RABIES VACCINE Serum Neutralization Titreb
Vaccine Titrea Group I
1st Dose
2nd Dose
6.3
None
II
6.3
6.7
III
8.2
None
IV
8.2
8.4
None
None
V
Monkey Number
Pre Vacc.
612 655 688 689 690 710 608 619 621 622 654 709 602 603 606 662 664 708 615 616 618 623 648 656 682 691 692 694 695
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
3 wks. 160 625 10 1 0 187 0 224 0 238 98 0 625 625 625 25 84 187 1
84 178 328 84 625 N.D. N.D. N.D. N.D. N.D. N.D.
Postvaccination 7 wks. 22 wks. 440 56 41 3 0 71 56 203 0 48 35 0 625 229 625 49 187 625 3 285 625 328 153 625 N.D. N.D. N.D. N.D. N.D.
625 153 1 1 0 625 280 625 0 N.D. 66 0 488 625 625 70 328 625 0 379 625 245 280 178 N.D. N.D. N.D. N.D. N.D.
N.D. N.D. 697 N.D. Not Done. "Recorded as the number of mouse LD50 administered to each animal (logio). bRecorded as reciprocal of the dilution which protects 50% of mice against 100 LD50 of CVS virus. 257
CANADIAN VETERINARY JOURNAL
TABLE III CHALLENGE RESULTS OF MONKEYS VACCINATED 6 MONTHS PREVIOUSLY WITH ERA RABIES VACCINE BY THE ORAL ROUTE
Group
Results
Monkey
I
F.A. ResultSa
612 Survived Survived 655 Survived 688 Survived 689 690 Died day 48 postchallenge Survived 710 II 608 Died day 2 postchallenge 619 Survived 621 Died day 22 postchallenge 622 Died prechallenge Survived 654 Survived 709 III 602 Survived 603 Survived 606 Survived 662 Survived 664 Survived Survived 708 Survived IV 615 Survived 616 Survived 618 Survived 623 Survived 648 Survived 656 V Died day 53 postchallenge 682 Died day 21 postchallenge 691 Died day 30 postchallenge 692 Survived 694 Died day 13 postchallenge 695 Survived 697 N.D. Not Done. Challenge: 106-2 mouse LD50 per 2 ml inoculated by multiple in the neck muscles. aF.A. + = fluorescent-antibody test positive for rabies.
knowledge the technical assistance of Miss E. Gaynor, and Messrs. D. Flemming and R. Gertus.
1. 2.
3.
4.
5.
REFERENCES BAER, G. M., M. K. ABELSETH and J. G. DEBBIE. Oral vaccination of foxes against rabies. Am. J. Epidem. 93: 487-490. 1971. BAER, G. M., J. R. BRODERSON and P. A. YAGER. Determination of the site of oral rabies vaccination. Am. J. Epidem. 101: 160164. 1975. BLACK, J. G. and K. F. LAWSON. Sylvatic rabies studies in the silver fox (Vulpes vulpes). Susceptibility and immune response. Can. J. comp. Med. 34: 309-311. 1970. BLACK, J. G. and K. F. LAWSON. Further studies of sylvatic rabies in the fox (Vulpes vulpes). Vaccination by the oral route. Can. vet. J. 14: 206-211. 1973. DEBBIE, J. G., M. K. ABELSETH and G. M. BAER. The use of commercially available vaccines for the oral vaccination of foxes against rabies. Am. J. Epidem. 96: 231-235.
1972.
258
N.D. N.D. N.D. N.D. + N.D. N.D. N.D. +
N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. N.D. + + + N.D. + N.D.
injection
6. LAVENDER, J. F. Immune response in primates vaccinated with duck embryo cell culture rabies vaccine. Appl. Microbiol. 25: 327331. 1973.
7. MAYR, A., H. KRAFT, 0. JAEGER and J. HAACKE. Orale Immunisierung von Fuchsen gegen Tollwut. Zentbl. VetMed. [B] 19: 615625. 1972. 8. REED, L.S. and H. MUENCH. A simple method of estimating 50 per cent endpoints. Am. J. Hyg. 27: 493-497. 1938. 9. SIKEs, R. K., W. F. CLEARY, H. KOPROWSKI, T. J. WIKTOR and M. M. KAPLAN. Effective protection of monkeys against death from street virus by post-exposure administration of tissue-culture rabies vaccine. Bull. Wld Hlth Org. 45: 1-11. 1971. 10. WIKTOR, T. J. and H. KoPRowsiu. Successful immunization of primates with rabies vaccine prepared in human diploid cell strain WI-38. Proc. Soc. exp. Biol. Med. 118: 10691073. 1965. 11. WINKLER, W. G., R. G. McLEAN and J. C. COWART. Vaccination of foxes against rabies using ingested baits. J. Wildl. Dis. 11:
382-388. 1975.
