MJ. Welsh and A.E. Smith, RAC Application 
Results 
Examination. Both monkeys tolerated the procedure well. Visual inspection of the nasal 
epithelium revealed no inflammatory response. Daily examination revealed no evidence of 
coryza or conjunctivitis. There was no cough, sneezing, or diarrhea. For both monkeys, 
the nasal mucosa was mildly erythematous in both the infection side and the control side on 
the first day after infection; we interpret this as a change due to the instrumentation. This 
decreased in subsequent studies as our techniques improved. Appetites and weights were 
not affected by virus administration in either monkey. Physical examination revealed no 
evidence of lymphadenopathy, tachypnea, or tachycardia. On day 21, monkey B had a 
temperature 39.1° C (normal for Rhesus monkey 38.8°C) but had no other abnormalities on 
physical exam or in laboratory data. 
Blood counts and serology. Monkey A had a slight leukocytosis on day 1 post infection 
which returned to normal by day 4: the WBC was 4,920 on the day of infection, 8,070 on 
day 1, and 5,200 on day 4. The ESR, serum electrolytes, transaminases, BUN and 
creatinine were normal throughout . 
Cytology of the nasal epithelium. We assessed epithelial inflammation by cytological 
examination of Wright-stained cells (cytospin) obtained from brushings of the nasal 
epithelium. We compared the percentage of neutrophils and lymphocytes to that of the 
control nostril and to the normal values from four control monkeys. Wright stains of cells 
from nasal brushing were performed on each of the evaluation days. They revealed less 
than 5% neutrophils and lymphocytes. There was no difference between the infected and 
the control side. 
Presence of virus and virus replication on the airway epithelium. We tested for the 
presence of virus in the supernatant of the cell suspension from swabs and brushes from 
each nostril, the pharynx, and trachea of both monkeys. Each supernatant was used to 
infect the virus-sensitive 293 cell line. Cytopathic changes in the 293 cells were monitored 
for 1 week and then the cells were fixed and stained for 6-galactosidase activity. 
Cytopathic effects and blue-stained cells indicated the presence of live virus. In Monkey 
A, live virus was detected in both nostrils on day 3 after infection; no live virus was 
detected at either one or two weeks post-infection. In Monkey B, live virus was detected in 
both nostrils, pharynx, and trachea on day 3, and only in the infected nostril on day 7 after 
infection. No live virus was detected at 2 weeks or after. 
Stool culture. Stools were collected every week and cultured for virus; all were negative. 
Adenovirus antibody titers. Adenovirus antibody titers were measured by ELISA at the 
Iowa State Hygienic Laboratory. Adenovirus 2 antibody titers were measured by ELISA at 
Genzyme. Both monkeys had an increase in adenovirus 2 antibody titers. 
Conclusions 
The results of these safety studies are reassuring in several respects. Neither monkey 
developed any signs associated with wild-type adenoviral infection. Although live virus 
was present in the nasal washings for at least 1 week (in one of the monkeys), there was no 
evidence of live virus at day 14 or beyond. Virus was present in the contralateral nostril 
and pharynx; we think the virus most likely reached those locations via drainage by gravity 
after we released the posterior occlusion or it may have been caused by the monkey putting 
its finger from one nostril to another after recovery from anesthesia. The monkeys received 
approximately 16-80 times the highest proposed human dose (on a PFU/cell basis), there 
were no adverse consequences and live virus rapidly disappeared. Finally, Ad2/BGal-l was 
Recombinant DNA Research, Volume 16 
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