Difficulty in detecting in vivo extracellular infective virus in cattle naturally infected with bovine leukemia virus.

A search was made for the presence in vivo of infective bovine leukemia virus (BLV) in secretions and excretions as well as in several tissues from five bulls, three cows and one calf. Two of the bulls and three of the cows were natural tumour cases of enzootic bovine leukosis, another three of the bulls were natural cases of persistent lymphocytosis and the calf was a natural tumour case of juvenile bovine leukosis. BLV infection was confirmed for all cattle, except the juvenile leukosis case, by detection of BLV-specific agar gel immunodiffusion (AGID) antibodies. Cell-free preparations were made from homogenates of lymphocytes, lymph nodes, spleens, livers, intestines, urinary bladders, salivary glands, mammary glands, pools of prostate glands and testicles and feces as well as from plasma, urine and milk, by passing them through 5 micrometers membrane filters. BLV infectivity in these cell-free preparations was examined by syncytia infectivity assay using susceptible cell cultures of bovine or ovine origin. Infective BLV could not be isolated from any of these cell-free preparations of plasma, secretions, excretions and tissues, although it was isolated consistently from the in vitro cultures of viable lymphocytes obtained from BLV-infected cattle. There was no indication of BLV involvement in the case of juvenile bovine leukosis.

Deformities of chick embryos in experimental Akabane virus infection.

A material containing 10(3.0) approximately 10(5.0) TCID50 of Akabane virus was inoculated into 6-day-old chick embryos by the yolk sac route. Death of embryos did not increase in the course of embryonic development till 18 days of age, as compared with control groups. Later than 18 days of age, however, the numbers of dead and peeping but unhatched embryos increased, making the hatching rate significantly low. Deformities, such as arthrogryposis and hydranencephaly, appeared in almost all the dead and unhatched embryos. They were severe in dead embryos, considerably severe in peeping unhatched embryos, and comparatively mild, though highly frequent, in affected hatched chicks. Many of the hatched chicks manifested ataxia, abnormal gait, astasia, or tremor of body or legs separately or together. Virus growth was demonstrated in chick embryos inoculated at 7 days of age. The virus titer was the highest (10(3.25 approximately 10(3.75) TCID50/O.1g) in head, trunk, and muscle, and the second highest (10(2.0) approximately 10(2.5) in brain, heart, and other visceral organs. It was the highest (10(4.0) approximately 10(4.5)) in muscle and a mixture of cerebellum and brain stem in embryos inoculated at 8 days of age.

Susceptibility of chickens to avian encephalomyelitis virus. II. Behavior of the virus in day-old chicks.

The VR strain of avian encephalomyelitis virus, which had been adapted to embryonated hen's eggs, was inoculated into 2-day-old chicks by the subcutaneous route (10(2.5) approximately 10(3.0) EID50) or by the oral route (10(4.8) EID50). The chicks were examined chronologically for the distribution of the virus in the body. As a result, minute amounts of the virus were detected from the liver, spleen, pancreas, and muscle at the site of inoculation one day after inoculation and various amounts from almost all the organs 3 days and more after inoculation. The virus titer could nearly reach a maximum 7 to 9 days after inoculation. Above all, such high virus titers as ranging from 10(4.3) to 10(5.8) EID50/0.1 g were demonstrated in the brain, heart, liver, spleen, and pancreas. After that, there was a tendency for virus titer to decrease in most organs and for virus to multiply persistently in the pancreas, brain, and eyeball. Virus titer was maintained at a level of 10(2.3) approximately 10(2.8) EID50/0.1 g in these three organs even 21 days after inoculation. In the group of subcutaneous inoculation, all the chicks manifested clinical signs of infection 5 to 10 days after inoculation. On the other hand, no chicks were involved in clinical infection in the group of oral inoculation. Multiplication of the virus was delayed in the body of these chicks. Small amounts of the virus were detected from the spleen and pancreas 11 days after inoculation. Low titers (10(2.7) EID50/0.1 g at the highest) of the virus were only detected from the brain, spinal cord, spleen, pancreas, esophagus, and other organs 14 and 21 days after inoculation.

Susceptibility of chickens to avian encephalomyelitis virus. III. Behavior of the virus in growing chicks.

A chick embryo-adapted strain of avian encephalomyelitis virus was inoculated subcutaneously and orally into 40-day-old (middle-aged) and 110-day-old (advanced-aged) chicks to examine the behavior of the virus in the chick body. In the middle-aged chicks, the virus appeared in the muscle at the site of inoculation, liver, spleen, pancreas, lumbar and cervical portions of the spinal cord, and brain 1 approximately 9 days after subcutaneous inoculation, and remained mostly in the central nervous system up to 17 days after the inoculation. The virus was found in large amounts in the muscle at the site of inoculation (10(3.1)), lumbar portion (10(2.5)) and cervical portion (10(2.1)) of the spinal cord, brain (10(1.9)), and in minute amounts in the other organs examined. It appeared in 11 of 21 organs examined. In the middle-aged chicks inoculated by the oral route, the virus was detected transiently in small amounts from esophagus, pancreas, and rectum 4 approximately 14 days after inoculation. In the advanced-aged chicks inoculated by the subcutaneous route, the virus was detected in titer of 10(2.1) approximately 10(3.0) from the muscle at the site of inoculation 2 approximately 7 days after inoculation. The virus was also found sporadically in several organs up to 17 days after inoculation. In the advanced-aged chicks inoculated by the oral route, no virus appeared in any organ, but these chicks turned to be weakly positive for neutralizing antibody in the 4th or later week after inoculation.