Progressive multifocal leukoencephalopathy. A case report with special reference to SV40 etiology

Clinical, light- and electron microscopic, and immunohistochemical findings of a 44-year-old woman with progressive multifocal leukoencephalopathy were presented. Autopsy revealed a wide distribution of the demyelinating lesion in the cerebrum, cerebellum, brain stem and spinal cord, and intranuclear inclusion bodies and papova-like virions in transmission electron microscopy in the nuclei of oligodendrocytes. SV40 antigen was immunohistochemically detected in these inclusion bodies. The widespread extension of the lesions seemed to correlate with the duration of the patient's illness. The prolongation of the clinical course in this case may be dependent upon the lack of serious underlying diseases except for a small nodule of thyroid carcinoma, SV40 infection rather than JC virus infection and/or improved care of that kind of patient.     

Electron microscopic evidence for the cruciform structure in intracellular SV40 DNA

The conformation of intracellular SV40 DNA during lytic infection of CV-1 cells was studied by the psoralen crosslinking technique. Analysis of the crosslinked SV40 DNA in the electron microscope revealed a rare population (0.1%) with a cruciform structure at coordinates 0.62 +/- 0.05 or at 0.37 +/- 0.05 of the SV40 genome. The implication of this observation in relation to SV40 DNA replication is discussed.     

Electron Microscopic Demonstration of Viral Particles in Sudden Infant Death Syndrome

Viral particles consistent with adenovirus were demonstrated in the intranuclear inclusion bodies of the intestinal epithelial cells in a 3¹/₂month-old girl who died of sudden infant death syndrome (SIDS). The particles were demonstrated by electron microscopy using a routine hematoxylin-eosin-stained section. Although viral infection is known to contribute to the pathogenesis of SIDS, this is the first case of SIDS in which viral inclusion bodies have been demonstrated in the intestinal epithelial cells by light and electron microscopy.     

Ultrastructural study of intranuclear inclusion bodies of pulmonary adenocarcinoma

Intranuclear inclusion bodies are sometimes observed in pulmonary adenocarcinoma by light microscopy. Electron microscopic characteristics of lung cancer cells with intranuclear inclusion bodies were studied. In addition, polymerase chain reaction (PCR) was performed using primers coding for human papillomavirus (HPV) types 16, 18, and 33. Eosinophilic intranuclear inclusion bodies were observed in 22 out of 285 cases by light microscopy. Immunohistochemically, cancer cell nuclei stained with PE-10. Three types of intranuclear inclusion bodies were classified electron microscopically. Type A showed aggregation of electron dense particles (30-40 nm) with an electron-dense core and was most frequently observed. Type B consisted of a mass of branching and whirling tubular structures. Type B intranuclear inclusions had a relationship with inner nuclear membrane. In type C, several spherical inclusions were observed in one nucleus. HPV DNA was detected using PCR and type-specific probes in a case with type A inclusion bodies. This study suggests that intranuclear inclusion bodies in pulmonary adenocarcinoma are formed by several different mechanisms.     

Mechanism of interferon action. Differential effect of interferon on the synthesis of simian virus 40 and reovirus polypeptides in monkey kidney cells

The effect of interferon (IFN) treatment on the synthesis of viral proteins was examined in BSC-1 monkey kidney cells singly and doubly infected with simian virus 40 (SV40) and reovirus. The pattern and amount of SV40 T antigen and reovirus λ, μ, and σ polypeptide synthesis were comparable in the singly and doubly infected cells in the absence of IFN treatment. As determined by Yakobson et al. (Cell12: 73–81, 1977) and Kingsman and Samuel (Virology101: 458–465, 1980) SV40-specific RNA synthesis is inhibited by treatment of cells with IFN before infection, but is not inhibited by IFN treatment after infection. IFN treatment initiated before infection inhibited the synthesis of both SV40 T antigen and reovirus polypeptides. However, SV40 T antigen synthesis was not inhibited in SV40 wt or tsA mutant-infected cells when treated with monkey kidney or human leukocyte IFN after infection. By contrast, reovirus polypeptide synthesis was markedly inhibited in BSC-1 cells treated with IFN after SV40 infection and superinfected with reovirus. SV40 infection did not rescue reovirus polypeptide synthesis from the inhibitory action of IFN, and reovirus infection did not alter the resistance of SV40 T antigen synthesis to IFN in cells IFN-treated after SV40 infection. Total cellular protein synthesis was not significantly affected by IFN treatment. These results suggest that the interferon-induced inhibitor(s) of protein synthesis discriminate between SV40 and reovirus mRNAs; the translation of SV40 mRNA, like most cellular mRNAs, is not affected under conditions where the IFN-induced inhibitor(s) significantly affect the translation of reovirus mRNA.     

Large granular nuclear bodies (karyosphaeridia) in experimental Borna virus infection

The inoculation of Borna virus produces florid encephalomyelitis in rabbits and a persistent symptomless infection in Syrian hamsters. Notwithstanding the diverse course of the infectious process, nuclear bodies are abundant and sometimes unusually large in the central nervous tissues of both species. It is possible that the very large bodies shown under the electron microscope correspond to the Joest-Degen inclusion bodies seen with the light microscope. The relevance of the nuclear bodies to viral synthesis or maturation would require further investigation.     

Investigation of the SV40 – Human Cancer Association: Look for the Full Signature of the Virus

In the controversy about the association of simian virus 40 with human cancers, the greatest problem is the ascertainment of SV40 exposure. This difficulty would be resolved if one were to look for all components of SV40 infection. How does SV 40 circulate in the human community? Do cancer patients with SV40-positive tumors have serological correlates of SV 40 infection and of SV40-induced cancer? SV40 association with a cancer should be studied in the context of the known risk factors for that cancer. The tumor cell-virus relationship should be characterized with respect to viral integration and viral localization to the tumor cell. Specimens should be masked and the assays should include panels of specimens to estimate analytic sensitivity and specificity. In view of the rarity of some of the tumors reported to be associated with SV40, a multi-institutional investigation initiated and coordinated by the NIH would be most effective.     

Adjuvant role of p53 immunostaining in detecting BK viral infection in renal allograft biopsies

BK virus infection is a significant threat to renal transplant outcome. Detecting viral infection in renal transplant biopsies using SV40 staining is less than ideal. SV40 antibody reacts with the large T-antigen of BK virus only at the early phases of infection and can miss cells in later stages of infection. As p53 is upregulated during both early and late phases of infection, this study set out to determine whether p53 staining could improve detection of BK virus infection in renal transplant patients. The control group consisted of 16 renal allograft biopsies without histologic evidence of BK virus infection, while the BK group consisted of 15 renal allograft biopsies with histologic evidence of BK virus infection. The biopsies from both groups were immunohistochemically stained with both SV40 and p53 antibodies. Dual staining with both markers was also performed to identify their nuclear co-localization. In the BK group, the percent of p53 staining (16.6 ± 4.8 %) was significantly higher than the percent of SV40 staining (5.4 ± 2.7%). BK virus infected cells revealed a unique p53 immunostaining pattern (strong nuclear staining with a central halo). Co-localization of SV40 and p53 was identified in cells that had characteristic nuclear features of BK virus infection by histology. The sensitivity and specificity for using p53 staining to identify BK infected cells was 92% and 86 %, respectively. In conclusion, p53 staining detects a higher percentage of BK virus infected cells than SV40 staining alone. Thus, for diagnosis of BK virus infection in renal allograft biopsies, p53 staining is a sensitive and specific method when used along with SV40 staining.     

Elektronenoptische Untersuchungen an Nierenzellen von Erythrocebus patas nach Infektion mit SV40 und Adenovirus Typ 12 1

Zusammenfassung Die Ultrastruktur von Pataszellen, die mit SV40 und 48 Stunden später mit Adenovirus 12 (Ad12) doppelt infiziert wurden, wurde in verschiedenen Zeitabständen zwischen 12 und 144 Stunden nach der Zweitinfektion untersucht. 12 Stunden nach der Ad 12-Zugabe waren im Kern lange Faserbündel zu erkennen, die offenbar mit dem Early-Protein und dem T-Antigen von Ad12 identisch sind. Zwischen 16 und 48 Stunden erschienen 4 Arten von Kerneinschlüssen, wobei die Ad 12-Synthese die langsam voranschreitende SV40-Neubildung zu überdecken vermochte. Es wurde vermutet, daß dem Nukleolus gewisse Aufgaben im Virussyntheseprozeß zuzuordnen sind. Schließlich wurde ein in späten Stadien der Ad 12-Vermehrung auftretendes Netzwerk aus strangförmigen Elementen, das schon von anderen Autoren in menschlichen Zellen nach Ad 3-Infektion beobachtet wurde, sowie den Kern durchziehende myelinartige Figuren aus Doppelmembranen beschrieben und diskutiert. Diese Ergebnisse wurden mit den Kernumbildungen verglichen, die als Folge des inkompletten Vermehrungszyklus von Ad 12 nach einfacher Infektion derselben Zellen auftreten. Aus all den Versuchen ging hervor, daß das Ad 12 nach doppelter Infektion in primären Affenzellen weitgehend dieselben Kernveränderungen hervorruft, wie nach einfacher Infektion in etablierten menschlichen Tumorzellen.

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Electron microscopic investigations on monkey kidney cells after double infection with SV40 and adenovirus 12

Summary The ultrastructure of kidney cells fromErythrocebus patas, double infected with SV 40 and 48 hours later with adenovirus type 12 (Ad12), was studied at various intervals from 12 to 144 hours after the second infection. Twelve hours after addition of Ad 12, bundles of long fibers appeared in the nucleus, which were obviously identical with an early-protein and the Ad 12 T-antigen. Between 16 and 48 hours, four types of inclusion bodies were seen, whereby the synthesis of Ad 12 could cover up the slowly progressing formation of SV 40. It was supposed that the nucleolus which showes fundamental alterations in its architecture, takes some active participation in the process of viral synthesis. A network of cord-like elements, already seen in human cells after infection with Ad 3 by others, as well as extensive myeline-like figures appearing in infected nuclei during late stages, were described and discussed. These results were compared with the nuclear lesions occurring in cells of the same origin after single infection with Ad 12. It appeared from all these observations that double infection of primary monkey cells provokes almost the same nuclear changes as single infection of established human tumor cells with Ad 12.

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Diese Arbeit wurde vom schweizerischen Nationalfonds unterstützt. Nummer des Forschungsprojektes: 5262.3.     

Pathological studies on chlamydiosis in parakeets (Psittacula krameri manillensis)

Histological, immunohistochemical and ultrastructural findings are described in 20 parakeets (Psittacula krameri ntanillensis) affected with chlamydiosis. The main histological lesions consisted of focal necrosis in the liver and adrenal gland, lymphocytic depletion with fibrin exudation in the splenic sinuses and follicles and fibrinopurulent airsacculitis. In these lesions basophilic chlamydial inclusion bodies were found. Macrophages and plasma cells increased mainly in the liver and spleen. Immunohistochemically. more chlamydial inclusion bodies were observed in cells of various organs and tissues including epithelial cells, capillary endothelium and proliferated macrophages. With an electron microscope, the chlamydial inclusion bodies were shown to consist of chlamydial organisms in developmental stages. Concurrent lesions of pulmonary herpesvirus infection appeared frequently in the present cases and seemed to have a close relationship with the chlamydiosis onset.     

Association of simian virus 40 with a central nervous system lesion distinct from progressive multifocal leukoencephalopathy in macaques with AIDS

The primate polyomavirus SV40 is known to cause interstitial nephritis in primary infections and progressive multifocal leukoencephalopathy (PML) upon reactivation of a latent infection in SIV-infected macaques. We now describe a second central nervous system manifestation of SV40: a meningoencephalitis affecting cerebral gray matter, without demyelination, distinct from PML. Meningoencephalitis appears also to be a primary manifestation of SV40 infection and can be seen in conjunction with SV40-induced interstitial nephritis and pneumonitis. The difference in the lesions of meningoencephalitis and PML does not appear to be due to cellular tropism, as both oligodendrocytes and astrocytes are infected in PML and meningoencephalitis, as determined by in situ hybridization or immunohistochemistry for SV40 coupled with immunohistochemistry for cellular determinants. This is further supported by examination of SV40 nucleic acid sequences from the ori-enhancer and large-T-antigen regions, which reveals no tissue-or lesion-specific variation in SV40 sequences.     

Hyperacetylation and differential deacetylation of histones H4 and H3 define two distinct classes of acetylated SV40 chromosomes early in infection

SV40 chromosomes undergoing encapsidation late in infection and SV40 chromatin in virions are hyperacetylated on histones H4 and H3. However, the fate of the SV40 chromosomes containing hyperacetylated histones in a subsequent round of infection has not been determined. In order to determine if SV40 chromosomes undergo changes in the extent of histone acetylation during early infection, we have analyzed SV40 chromosomes isolated 30 min and 3 h postinfection by quantitative ChIP assays, depletion ChIP assays, competitive ChIP assays, and ChIP assays combined with restriction endonuclease sensitivity using antibodies to hyperacetylated histones H4 and H3. We have shown that at 30 min postinfection, the hyperacetylated histones are associated with two distinct classes of SV40 chromosomes. One form is hyperacetylated specifically on histone H4 while a second form is hyperacetylated on both H4 and H3. Both forms of chromosomes appear to contain a nucleosome-free promoter region. Over the course of the next few hours of infection, the class of SV40 chromosomes hyperacetylated on only H4 is reduced or completely eliminated through deacetylation.     

Biology of simian virus 40 (SV40) transplantation antigen (TrAg). V In vitro demonstration of SV40 TrAg in SV40 infected nonpermissive mouse cells by the lymphocyte mediated cytotoxicity assay.

The development of SV40-specific transplantation antigen (TrAg) on the surface of nonpermissive mouse cells infected with SV40 was demonstrated using a sensitive in vitro lymphocyte mediated cytotoxicity assay. The in vitro lymphocyte mediated ⁵¹Cr release assay was shown to be specific for the detection of SV40 TrAg. SV40 TrAg was detected 24 hr after infection of mouse cells with SV40 and high levels of TrAg expression persisted for as long as 96 to 120 hr after virus infection. The development of TrAg on the surface of SV40-infected mouse cells correlated with the synthesis of tumor or T antigen in the nucleus of infected cells. The synthesis and the expression of TrAg at the surface of SV40-infected mouse cells may be an important step in the development of immunological resistance of the cell mediated type in an SV40-inoculated host leading to the elimination of stably transformed cells.     

Recombination between simian virus 40 and adeno-associated virus: virion coinfection compared to DNA cotransfection

Recombination between simian virus 40 (SV40) and adeno-associated virus (AAV) has been detected, by infectious center in situ plaque hybridization procedures, after both DNA contransfection and virion coinfection of monkey BSC-1 cells. The number of cells producing recombinants (1 in a 1000) was the same irrespective of the way in which the SV40 and AAV genomes were delivered to the cell, despite the fact that 5-10 times more cells were infected after virion coinfection. Several other dosage-response parameters of the recombination process consequent to virion coinfection were comparable to those after DNA cotransfection. The sole difference observed between the two infection systems was that the SV40/AAV recombinants formed after virion coinfection contained an inordinately high proportion of AAV terminal DNA sequences. By separating the SV40 and AAV infections in time, such that the AAV infection was delayed until after certain events in the SV40 cycle had taken place, an optimum phase for recombination in the SV40 cycle was identified. This phase occurs a few hours after infection, well before the onset of SV40 DNA replication and the synthesis of SV40-specific early proteins.     

Evidence of simian virus 40 exposure in a colony of captive baboons

Simian virus 40 (SV40) is a polyomavirus for which non-human primates are the permissive host. The baboon (Papio spp.) is an old world monkey that is used in a variety of research investigations; however, natural infection of SV40 among baboons has not been thoroughly examined or reported. Initially, we were interested in determining the prevalence of SV40 infection among a captive colony of baboons based on the presence of antibodies to SV40 large T-antigen (Tag). An overall seroprevalence rate of >50% was found after screening sera from 142 baboons in the colony based on ELISA. Endpoint titer values for serum antibody binding to SV40 Tag reached as high as 1280 for 5 out of 142 baboons. Peptide binding assays revealed that a range of SV40 Tag epitopes are immunogenic in the baboon, and that individual animals differ in their humoral immune responses to SV40 Tag based on epitope recognition. Specificity to SV40 Tag and not some other primate polyomavirus encoded large Tag was further examined by serologic reactivity to peptide epitopes unique to SV40 Tag. Additional serology was performed to assess SV40 Tag reactivity by Western blot and whether antibodies were capable of neutralizing SV40 infectivity in vitro. Although antibodies with high levels of SV40 neutralization were observed in a number of the baboons, there was a lack of correlation between viral neutralization and antibodies to SV40 Tag. Further examination using molecular-based diagnosis and SV40 Tag specific real-time quantitative PCR determined that some of the baboons appeared to be exposed to SV40. DNA sequence analysis of the PCR products confirmed that SV40 Tag specific sequences were detected in baboons.     

Polyribosomes of cells abortively or productively infected with adenovirus,papovavirus, or their hybrid

Summary The polyribosomes of African green monkey kidney (GMK) cells have been characterized during productive and abortive infections with adenovirus type 2, simian papovavirus SV40, and the PARA (defective SV40)-adenovirus type 2 hybrid. An early increase in uptake of H³-uridine was followed by a progressive decrease in both the amount of polyribosomes and in uptake of the label in all three systems that involved an adenovirus. Thus, the polyribosome distribution patterns obtained from GMK cells abortively infected with adenovirus type 2 did not differ significantly in optical density profile or incorporation of H³-uridine from the distributions obtained from GMK cells productively infected with either adenovirus 2 and SV40 or PARA-adenovirus type 2.A decrease in the amount of polyribosomes was not evident when GMK cells were infected with SV40. During the productive cycle of SV40 in GMK cells, the fifth (pentamer) polysome peak enlarged as the infection progressed. This increase was reflected both in increased optical density and incorporated H³-uridine. Material from this peak reacted with antibody to SV40 capsid antigen in the complement-fixation test. Arabinofurano-sylcytosine, a DNA inhibitor that blocks SV40 replication at a step prior to capsid formation, inhibited the formation of this peak. Treatment of the cellular extracts with EDTA or ribonuclease did not shift this peak to a lighter part of the gradient, along with the other polyribosomes. Virus particles, some of which proved to be infective for monkey cells, were present in electron micrographs of the pentamer peak.     

Light and electron microscopic observation of intracytoplasmic inclusion bodies in the locus coeruleus of the hamster

The authors previously demonstrated that intracytoplasmic inclusion bodies (1-3 microm) in the mouse locus coeruleus under light and electron microscopy are characteristically stained using the Holmes modified method. We reported that one inclusion body existed in almost all neurons of the locus coeruleus. The present study examined whether similar inclusion bodies are present in the Syrian hamster (weight, about 60 g). Paraffin sections stained with the modified Holmes' method dis played numerous small inclusion bodies in the cytoplasm of cells in the locus coeruleus. Epon sections (1 microm thick) stained using toluidine blue were observed under light microscopy, and numerous small inclusion bodies were again observed. Under electron microscopy observation, inclusion bodies (<1 microm in diameter) predominantly comprised small granular materials, similar to those described by previous investigators. Although inclusion bodies were devoid of a limiting membrane, the relation ship to cytoplasmic organelles was unclear. However, free and polyribosomes were occasionally noted in close proximity to inclusion bodies. Inclusion bodies may thus be formed from ribosomes. Intracytoplasmic inclusion bodies in the hamster locus coeruleus differed in appearance compared with inclusion bodies in the mouse locus coeruleus.     

Simian virus 40-Chinese hamster kidney cell interaction II. The semipermissivity of the cell system

Summary Throughoutin vitro passages, Chinese hamster kidney (CHK) cells progressively lost susceptibility to SV 40 virus infection while remaining continuously susceptible to viral DNA infection. Upon infection with SV 40 virus or viral DNA, the CHK cell line supported viral DNA and virus replication at a low level. SV 40-transformed CHK cell lines spontaneously produced small amounts of viral DNA and virions. The percentage of virus-producing cells was low. Various clones derived from each of these lines behaved as the parental cell population, leading to the conclusion that each CHK cell, whether transformed or not with SV 40, is potentially permissive for this virus.With 4 Figures