In Borna Disease Virus Infected Rabbit Neurons 100 nm Particle Structures Accumulate at Areas of Joest‐Degen Inclusion Bodies
Authors: Sasaki S. , Ludwig H. 
Published: May 2010

Borna disease virus infected rabbits were chosen to search for electronmicroscopic structures. Intensively investigated hippocampal neurons showed intranuclear inclusions; 100 nm particle‐like structures surrounded by 20 nm granular forms were prominent. In connection with elsewhere reported in situ hybridization studies of virus‐specific RNA to areas of the Joest‐Degen inclusions we suggest that these particle structures may represent Borna virus.

Jost‐Degen (8) found intranuclear inclusion bodies in neurons to be pathognomonic for Borna disease (BD) in the horse. Half a century later these structures were suggested to represent BD virus (BDV)‐specific antigen aggregates (15). A century later we characterized the virus to contain a single and negative stranded RNA of 8.5 kb, which transcribes in the nucleus (5) and could show that virus complementary RNA seems to hybridize spot‐like to nuclear areas, probably representing the Jost‐Degen inclusions (7).

Electron microscopic (EM) findings on structures in BDV infected brain cells and about particle‐like structures obtained from infected tissue culture cells have been reported by different groups (1, 2, 3, 4, 5, 6, 9, 11, 12, 13). The demonstration of crystalline aggregates and filament bundels in the cytoplasma and karyosphaeridia (nuclear bodies) were prominent. Such structures were seen in infected rabbits, hamsters, rats mice and naturally infected horses. The phaenotypic description of filamentous structures, crystalline aggregates in the cytoplasma and large karyo‐spheridia were prominent.

Based on our experience with BDV infections in rabbits we selected this species for ultra‐structural studies. From a series of intranasally infected rabbits (BDV strain V), which were typically sick 6–8 weeks later, and showed the classical neuropathology (14), animal No. 5/37 was depicted. It showed the characteristic multifocal retinopathy (10) and had BDV antibodies of 1: 200 in the serum and 1: 10 in the CSF. The animal was perfused with a glutaraldehyde/PBS solution and small blocks preferentially from the hippocampus were processed according to standard procedures for EM. Two uninfected rabbits served as controls.

In the large pyramidal neurons of the hippocampus frequently nuclear bodies with sizes of 0.5 üm were found, which never were seen in control brains. Two representative series from hundreds of sections present our findings about inclusion bodies and other structures (Figs. 1 and 2).

Densely stained particle‐like structures of approximately 100 nm with granules of approx. 20 nm on their surface were numerously detected. They were occasionally surrounded by a capsule of low density. These “particles” could in a more transitional form be suspected also at other locations of the nucleus. They concentrated, however, close to dense granular material. The smaller, ca. 20 nm structures, seemed to fill the capsular ball. None of such 100 nm structures were detected in control sections. Additionally to the nuclear structures in BDV infected neurons we detected the intracytoplasmic crystalline areas, shown in Fig. 3. Such aggregates are considered to be of nonspecific character (1).

Blinzinger and Anzil (4) extensively discussed and showed evidence for the larger nuclear inclusion bodies to represent Jost‐Degen inclusions of BDV infected neurons in rabbits and persistently infected hamsters. We and others came to the same conclusion, partially based on immunhistological studies (3, 6, 7a, 12).

Recently we found by in situ hybridization of BDV plus‐strand RNA (recognizing viral RNA) a strong reaction with nuclear areas which represent Jost‐Degen bodies (7). Our molecular biological data on this intranuclearly transcribed RNA virus (BDV) together with the EM observations presented here leave the conclusion that the 100 nm structures may represent the virus. ‐ We are aware of the discrepancies concerning the 50–60 nm particles obtained from infected green monkey kidney cells after affinity chromatography (11), and recently discussed similar sized particles, released from MDCK cells (5). Particles released from infected human oligodendroglia cells, however, (Zimmermann, W. & Ludwig, H., unpublished) which correlate with BDV infectivity and can be neutralized, seem to reinforce findings presented here for infected rabbit neuron structures.