THE JOURNAL OF INFECTIOUS DISEASES. VOL. 134, © 1976 by the University of Chicago. All rights reserved.
xo.
4 • OCTOBER 1976
Effect of Serum and Nasal Neutralizing Antibodies on Bovine Respiratory Syncytial Virus Infection in Calves From the Department of Veterinary Science, University of Maryland, College Park, Maryland
Sashi B. Mohanty, Mary G. Lillie, and Allen L. Ingling
Eagle's minimal essential medium with Earles' salts, 10% fetal calf serum (free of adventitious viruses), 0.5% lactalbumin hydrolysate, and 0.11 mg of sodium pyruvate/mi. The maintenance medium for BT cells was the same as the growth medium, except 2% fetal calf serum was used. Penicillin and streptomycin were added to all media. Test animals and inoculation procedure. Nine six- to eight-week-old bull calves were alloted to four groups and confined to separate isolation stalls (table 1). Calves were inoculated intranasally with 10 ml of virus (titer, I()3·5 TCID 5o / m l) and another 10 ml was given intratracheally. These procedures have been described [6]. Control calves were similarly inoculated with normal tissue culture fluid. In group A three calves were controls, in groups Band C two calves in each group were infected with the virus, and in group D one calf was infected and another was a contact control. Challenge with bovine RSV. Five weeks after the initial exposure, all calves except no. 9, which died from salmonellosis on day 15, were challenged with bovine RSV for study of the effect of serum and nasal neutralizing antibodies in this virus infection (table 2). The calves were inoculated intranasally and intratracheally with the virus. Clinical specimens. Procedures used in clinical observation, collection of sera and nasal swabs, and viral identification have been described [10]. Nasal secretions were collected with small sanitary tampons (Tampax Jr., Tampax, Inc., Palmer, Mass.) and processed as described
Isolation of bovine respiratory syncytial virus (RSV) has been reported from several countries [1-5]. The bovine RSV is capable of producing respiratory infection in calves [5-7]. we isolated a chloroform-sensitive RNA virus from the trachea of a calf; this virus produced a syncytial type of CPE and was subsequently identified as a RSV serologically related to the human RSV. Evidence for the possible enhancement of RSV pathogenesis in infants by immunologic factors has been observed [8, 9]. The purpose of this study was to induce RSV infection experimentally in calves and to determine the role of serum and nasal neutralizing antibodies on the outcome of the disease. Materials and Methods
Virus and cell cultures. Bovine RSV strain no. A51908 underwent five passages in primary and/or secondary bovine embryonic kidney cells before use. Media for growth and maintenance of these cell cultures have been described [10]. A bovine turbinate (BT) cell line (Dr. A. ,tV. MeClurkin, Ames, Iowa) was also used in this study. The growth medium for BT cells consisted of Received for publication February 19, 1976, and in revised form March 25, 1976. This paper was approved as scientific article no. A2190, contribution no. 5163, from the Maryland Agricultural Experiment Station, University of Maryland, College Park, Md. Please address requests for reprints to Dr. Sashi B. Mohanty, Department of Veterinary Science, University of Maryland, College Park, Maryland 20740.
409
Downloaded from http://jid.oxfordjournals.org/ at Emory University on August 24, 2015
Calves exposed to a bovine respiratory syncytial virus showed mild clinical signs of respiratory illness and responded serologically. The disease occurred in the presence or absence of circulating antibodies, but there was no evidence of exacerbation of the disease due to preexisting serum antibody. Nasal secretory antibody appeared to protect the calves against the disease. Calves previously exposed to the virus were immune to challenge. The virus was recovered at a high frequency when specimens of nasal secretions (not subjected to freezing and thawing) were inoculated into susceptible cells within I hr after collection.
Mohanty, Lillie, and Ingling
410
Results
Clinical response. Three control calves (no. 1-3) of group A remained normal throughout the experiment. The contact control calf (no. 9) remained normal until day 13 and then had
diarrhea for two days; this calf died from salmonellosis on day 15. Of the inoculated calves, two (no. 4 and 7) had a transient increase in body temperature (39.5 C and 40.5 C) on days 8 and 9. The temperature returned to normal (39 C) on day 10. Calves no. 5 and 8 had a temperature of 39.5 C on day 9. Calves no. 4 and 7 had slight serous nasal discharge on day 10, mild cough on day 9, and also slight hyperpnea on day 11. Calf no. 5 had slight nasal discharge, mild cough, and hyperpnea only on day 11. Calf no. 6 had no clinical signs. Recovery of virus. Results of recovery of virus from calves are presented in table 1. No virus was isolated from calves at the start of the experiment. Bovine RSV was isolated once from the contact control calf (day 7), but no virus was isolated from this calf at necropsy on day 15. No virus was isolated from control calves. All calves inoculated with the virus shed virus in nasal secretions for various lengths of time. Recovery of virus was much higher when material from nasal swabs was inoculated into susceptible BT cells within 1 hr after collection, without previ-
Table 1. Recovery of virus and antibody response in calves after initial exposure to bovine respiratory syncytial virus strain no. A51908. Titers of neutralizing antibodies at indicated rime f Virus recovered on indicated days after inoculation
Group, calf no. *
Inoculum'[
Group A (controls) 1 2 3
Uninoculated NTCF NTCF
Group B 4 5
Virus Virus
2,9, 11, 18,21 2,4, 7, 11, 18,21
Group C 6 7
Virus Virus
2,4,9, 11, 18 2,4, 7, 11, 18
Virus Uninoculated (contact control)
2,7, 11
7
Group D 8 9
Day O§ S
NS
Day 7 S
NS
Day 14 S
NS
Day 35
Day 28 S
NS
S
NS
16 2
xo.
4 • OCTOBER 1976
Effect of Serum and Nasal Neutralizing Antibodies on Bovine Respiratory Syncytial Virus Infection in Calves From the Department of Veterinary Science, University of Maryland, College Park, Maryland
Sashi B. Mohanty, Mary G. Lillie, and Allen L. Ingling
Eagle's minimal essential medium with Earles' salts, 10% fetal calf serum (free of adventitious viruses), 0.5% lactalbumin hydrolysate, and 0.11 mg of sodium pyruvate/mi. The maintenance medium for BT cells was the same as the growth medium, except 2% fetal calf serum was used. Penicillin and streptomycin were added to all media. Test animals and inoculation procedure. Nine six- to eight-week-old bull calves were alloted to four groups and confined to separate isolation stalls (table 1). Calves were inoculated intranasally with 10 ml of virus (titer, I()3·5 TCID 5o / m l) and another 10 ml was given intratracheally. These procedures have been described [6]. Control calves were similarly inoculated with normal tissue culture fluid. In group A three calves were controls, in groups Band C two calves in each group were infected with the virus, and in group D one calf was infected and another was a contact control. Challenge with bovine RSV. Five weeks after the initial exposure, all calves except no. 9, which died from salmonellosis on day 15, were challenged with bovine RSV for study of the effect of serum and nasal neutralizing antibodies in this virus infection (table 2). The calves were inoculated intranasally and intratracheally with the virus. Clinical specimens. Procedures used in clinical observation, collection of sera and nasal swabs, and viral identification have been described [10]. Nasal secretions were collected with small sanitary tampons (Tampax Jr., Tampax, Inc., Palmer, Mass.) and processed as described
Isolation of bovine respiratory syncytial virus (RSV) has been reported from several countries [1-5]. The bovine RSV is capable of producing respiratory infection in calves [5-7]. we isolated a chloroform-sensitive RNA virus from the trachea of a calf; this virus produced a syncytial type of CPE and was subsequently identified as a RSV serologically related to the human RSV. Evidence for the possible enhancement of RSV pathogenesis in infants by immunologic factors has been observed [8, 9]. The purpose of this study was to induce RSV infection experimentally in calves and to determine the role of serum and nasal neutralizing antibodies on the outcome of the disease. Materials and Methods
Virus and cell cultures. Bovine RSV strain no. A51908 underwent five passages in primary and/or secondary bovine embryonic kidney cells before use. Media for growth and maintenance of these cell cultures have been described [10]. A bovine turbinate (BT) cell line (Dr. A. ,tV. MeClurkin, Ames, Iowa) was also used in this study. The growth medium for BT cells consisted of Received for publication February 19, 1976, and in revised form March 25, 1976. This paper was approved as scientific article no. A2190, contribution no. 5163, from the Maryland Agricultural Experiment Station, University of Maryland, College Park, Md. Please address requests for reprints to Dr. Sashi B. Mohanty, Department of Veterinary Science, University of Maryland, College Park, Maryland 20740.
409
Downloaded from http://jid.oxfordjournals.org/ at Emory University on August 24, 2015
Calves exposed to a bovine respiratory syncytial virus showed mild clinical signs of respiratory illness and responded serologically. The disease occurred in the presence or absence of circulating antibodies, but there was no evidence of exacerbation of the disease due to preexisting serum antibody. Nasal secretory antibody appeared to protect the calves against the disease. Calves previously exposed to the virus were immune to challenge. The virus was recovered at a high frequency when specimens of nasal secretions (not subjected to freezing and thawing) were inoculated into susceptible cells within I hr after collection.
Mohanty, Lillie, and Ingling
410
Results
Clinical response. Three control calves (no. 1-3) of group A remained normal throughout the experiment. The contact control calf (no. 9) remained normal until day 13 and then had
diarrhea for two days; this calf died from salmonellosis on day 15. Of the inoculated calves, two (no. 4 and 7) had a transient increase in body temperature (39.5 C and 40.5 C) on days 8 and 9. The temperature returned to normal (39 C) on day 10. Calves no. 5 and 8 had a temperature of 39.5 C on day 9. Calves no. 4 and 7 had slight serous nasal discharge on day 10, mild cough on day 9, and also slight hyperpnea on day 11. Calf no. 5 had slight nasal discharge, mild cough, and hyperpnea only on day 11. Calf no. 6 had no clinical signs. Recovery of virus. Results of recovery of virus from calves are presented in table 1. No virus was isolated from calves at the start of the experiment. Bovine RSV was isolated once from the contact control calf (day 7), but no virus was isolated from this calf at necropsy on day 15. No virus was isolated from control calves. All calves inoculated with the virus shed virus in nasal secretions for various lengths of time. Recovery of virus was much higher when material from nasal swabs was inoculated into susceptible BT cells within 1 hr after collection, without previ-
Table 1. Recovery of virus and antibody response in calves after initial exposure to bovine respiratory syncytial virus strain no. A51908. Titers of neutralizing antibodies at indicated rime f Virus recovered on indicated days after inoculation
Group, calf no. *
Inoculum'[
Group A (controls) 1 2 3
Uninoculated NTCF NTCF
Group B 4 5
Virus Virus
2,9, 11, 18,21 2,4, 7, 11, 18,21
Group C 6 7
Virus Virus
2,4,9, 11, 18 2,4, 7, 11, 18
Virus Uninoculated (contact control)
2,7, 11
7
Group D 8 9
Day O§ S
NS
Day 7 S
NS
Day 14 S
NS
Day 35
Day 28 S
NS
S
NS
16 2
