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Using complementary RNA of human papilloma virus type 1 (HPV1) and in situ hybridization techniques the localization of viral DNA replication was studied in sections of 38 human virus acanthomata from 31 different patients. In five cases significant labeling was detected by autoradiography. Labeling started always in the first or the second suprabasal cell layer and was strongly limited to the nuclei. A remarkable early beginning of the vacuolated process seemed to be correlated with the visible DNA replication. No labeling could be detected in the basal cell layer. This suggests that these cells contain at the most only a small number of viral genomes. Our findings represent only the situation in human warts definitely caused by HPV 1. We are not able to say, whether our negative hybridization results in the remaining warts indicate either the scarcity of viral genomes within these sections or their infection by a different agent.

From skin papillomas of the chaffinch (Fringilla coelebs), a virus has been purified and studied by physicochemical techniques and electron microscopy. The virions measure 52 nm in diameter and are composed of 72 morphological units arranged in a skew T = 7d surface lattice. A sedimentation coefficient of about 300S and a buoyant density of 1.34 g/ml in CsCl were determined for the particle. Its protein composition resembles that of human papillomavirus, and the circular double-stranded genome measures 2.6 micronm. This is the first demonstration of a member of the Papovaviridae family that affects a nonmammalian host.

A routine cervical smear from a 56-year-old postmenopausal woman showed appearances indicative of papilloma virus infection with atypia. In an attempt to prove the diagnosis, the smear was reprocessed for electron microscopic examination. The reprocessing technique is described. Ultrastructural preservation was poor, and significant shrinkage was observed. In spite of these technical difficulties it was possible to prove that the selected cells in the smear contained papilloma virus particles. The virus particles seen were intranuclear in location and spherical in shape with a diameter of 40 to 50 nm. As was demonstrated in this case, the technique described will prove useful in evaluating the reliability of various diagnostic criteria for identifying viral infections in routine smears. This will apply particularly in cases where cultural confirmation of a reported viral infection is not possible.

Naturally occurring cutaneous fibromas affecting white-tailed deer (Odocoileus virginianus) and mule deer (O hemionus), and cutaneous fibropapillomas of domestic cattle were tested for papillomavirus using indirect immunofluorescence (IF), peroxidase-antiperoxidase (PAP), and negative-stain electron microscopic techniques. Papillomavirus was consistently detected using rabbit antiserum against papillomavirus group-specific antigen in all mule deer fibromas and bovine fibropapillomas; only 16 of 28 white-tailed deer fibromas tested by IF and 9 of 15 tested by PAP were detected. Normal skin from white-tailed deer or cattle was consistently negative for virus. Similar results were obtained by negative-stain electron microscopic examination of partially purified tumor homogenates. Using deer fibroma virus or bovine papillomavirus type 1-specific antisera, viruses were typed by IF, PAP, and immunoelectron microscopy.

A spectrum of proliferative cutaneous lesions occurred in 12 dogs at the injection site of live canine oral papillomavirus (COP) vaccine, suggesting a viral etiology for the masses. Lesions included epidermal hyperplasia, epidermal cysts, squamous papilloma, basal cell epithelioma, and squamous cell carcinoma. Peroxidase-antiperoxidase staining of tumor sections revealed nuclei which stained for group-specific papillomavirus antigen in five of 12 masses. Electron microscopic examination of tumor sections did not reveal virions. In transmission studies, macerated tumor tissue did not produce oral papillomas on the scarified mucosa of puppies; this procedure did protect the puppies from development of lesions when challenged with infectious papilloma material. These findings are evidence that COP can induce hyperplastic and neoplastic lesions in sites other than oral, pharyngeal, and ocular mucosa.

Recent biochemical and serological studies have shown the existence of at least four distinct types of human papillomaviruses (HPVs) causing benign skin lesions. These viruses show hardly no antigenic relationships; their DNAs differ by their sensitivity to restriction endonucleases, and show little, if any, sequence homology, as detected by molecular hybridization using complementary RNAs transcribed in vitro. Data on the pathogenicity of HPVs are still incomplete but indicate that some types of benign skin lesions (plantar warts, common warts, flat warts) may be preferentially associated with some types of HPV. Most interesting is that epidermodysplasia verruciformis has been found associated with two types of virus, and that malignant conversion of some lesions has been observed in all the patients infected with one of them. This suggests that at least a HPV may have a higher oncogenic potential, as do rabbit (Shope) papillomavirus and bovine alimentary tract papillomavirus. Much remains to be known on human papilloma-viruses and further studies may lead to the characterization of additional types of HPVs, especially in genital condylomata acuminata and laryngeal papillomas whose malignant conversion, although rare, may be observed. Progress in this field has been and remains hampered by the lack of cell culture systems allowing replication of these highly host and tissue specific viruses, and by the widely variable virus content of the different human lesions known to be associated with a papillomavirus. Further studies are warranted by the possible role of these widespread and epitheliotropic viruses in the origin of some carcinomas in man.

In rare cases of progressive multifocal leukoencephalopathy (PML) the lesions are atypical in that the characteristic alteration of oligodendrocytic nuclei is infrequent or absent, and the demyelinated foci contain mononuclear inflammatory infiltrates including numerous plasma cells. Otherwise these cases, both as to the topographic features of the lesions, and the background of chronic lymphoproliferative or myeloproliferative disease or immunosuppressive treatment, conform to the usual pattern. In the case here reported, that of a 54-year-old man receiving immunosuppressive drugs for chronic polymyositis, electron microscopy showed papovavirions in astrocytic nuclei and fluorescent antibody studies indicated that these represented JC virus. In this and the 3 previously reported similar cases of atypical PML, the neurologic illness was less devastating than generally occurs in classic PML. Possibly these cases represent instances of unusually strong host-resistance against the disease.

HPV-induced changes in the portio vaginalis uteri can and have lead to misinterpretations since they frequently occur in combination with or resemble precancerous or cancerous changes. The morphological peculiarities and growth patterns of koilocytotic changes in the portio vaginalis uteri are described. Among 684 conizations performed, 169 (24.7%) were found to have features of an HPV-infection. Of these, 75.7% also had preneoplastic or neoplastic changes. Review of the cytological findings revealed that in 66.7% of the cases with histologically verified koilocytosis a false positive finding of malignancy had been recorded.

In a series of 774 women with pregnancies ranging from 7 weeks to term, virus particles representing five different groups have been detected in urine specimens by negative staining electron microscopy. Of the virus-positive women, 0,39% excreted papillomavirus (in the absence of genital warts), 3,1% polyomavirus, 0,26% adenovirus type II, 0,65% an antibody-coated unidentified virus particle and 7,7% a membrane-associated virus-like particle. The implications of these findings are discussed in the light of the apparently increased susceptibility of pregnant women to virus infection.

54 Tumors of Mastomys natalensis (GRA Giessen), described in the literature as "Keratoacanthomas" were investigated by light and electron microscopy. They are induced by a virus of the Papova group. Their histogenesis and their course are very different from human keratoacanthoma. The results reveal problems related to experimental animal tumour models and the similarities of different malignancies in the nomenclatures of several disciplines.

Virus induced changes occur not so rarely in combination with precancerous and cancerous lesions. We compared the cytological and histological findings. The increasing frequency of this event is demonstrated by an analysis of the material seen in our laboratory, smears from more than 120,000 patients per year. It was possible to verify the viral etiology by the electron microscopical demonstration of virus particles.

Human papillomaviruses (HPVs) found in lesions of 11 patients suffering from epidermodysplasia verruciformis were compared to HPV type 1 (HPV-1) and HPV type 2 (HPV-2) previously characterized in plantar and common warts, respectively. Complementary RNAs (cRNAs) to HPV-1, HPV-2, and viruses obtained from two patients with epidermodysplasia verruciformis (J.D. HPV and J.K. HPV) were used in cRNA.DNA filter hybridization experiments. No sequence homology was detected between HPV-1 or HPV-2 DNAs and DNAs obtained from the 11 epidermodysplasia verruciformis HPV isolates. Furthermore, with J.D. and J.K. HPV cRNAs, epidermodysplasia verruciformis HPV DNAs fell into two groups showing little, if any, sequence homology. A lower extent of annealing was observed for the DNAs of some isolates showing a genetic heterogeneity within each of the two groups. Almost no antigenic crossreaction was detected by immunodiffusion and indirect immunofluorescence tests, either between epidermodysplasia verruciformis HPVs and HPV-1 or HPV-2 or between J.D. and J.K. HPVs. Viruses belonging to the same group have common antigenic properties, but antigenic differences were observed when two of the viruses sharing only partial DNA sequence homology were compared. Viruses related to J.D. HPV were preferentially associated with flat wart-like lesions of epidermodysplasia verruciformis and were further found in the lesions of five patients bearing multiple flat warts. Viruses related to J.K. HPV were found in morphologically distinct lesions (red spots) present in some patients with epidermodysplasia verruciformis. Thus, we propose to distinguish two other types of HPVs designated provisionally as HPV type 3 (HPV-3) and HPV type 4 (HPV-4), with J.D. and J.K. HPVs as prototypes, respectively. Malignant conversion of some epidermodysplasia verruciformis lesions is more frequently associated with HPV-4 than with HPV-3 infection.

Papillomaviruses are ubiquitous DNA viruses that are epitheliotropic and produce a range of epithelial neoplasms, both benign and malignant, in animals and man. Human papillomaviruses are associated with a variety of rare and uncommon oral lesions, and there has been increasing suspicion that they may be implicated also in some premalignant and malignant oral lesions.

SA12 virus, originally isolated from an uninoculated South African vervet monkey kidney culture, was identified as a new member of the simian virus 40 (SV40)-polyoma subgroup of papovaviruses. The virus produced a cytopathic effect with nuclear enlargement in primary rhesus kidney cells. The virion had papovavirus morphology and a diameter of 44 to 45 nm. The DNA of the virus was a circular, double-stranded, superhelical molecule with a mean length 101% that of SV40 DNA and an estimated molecular weight of 3.3 X 10(6). The virus was found to be unrelated to other papovaviruses by neutralization, immune electron microscopy, and immunofluorescence tests with antiviral sera. SA12 virus-infected cells exhibited a capsid antigen, which has recently been found to be common to viruses of the SV40-polyoma subgroup. The virus readily transformed kideny cells from 10-day-old hamsters. Inoculation of transformed cells produced tumors in 3- to 4-week-old hamsters. The T antigens of SA12 and SV40 viruses were strongly and reciprocally cross-reactive. A high proportion of the sera of chacma baboons, Papio ursinus, and a comparatively lower proportion of the sera of vervet monkeys, Cercopithecus pygerythrus, had neutralizing antibodies to SA12 virus

Negative staining electron microscopy was used to identify viruses in 157 normal and 29 diarrhoeal faecal samples collected from 156 dogs admitted to an animal shelter during an 8 month period (March to October) in 1982. Seven distinct viral types were detected: 21-26 nm parvovirus-like particles, 28-31 nm astrovirus-like particles, a previously undescribed 34-35 nm "round" virus particle, coronavirus, coronavirus-like particles ( CVLP ), rotavirus and papova-like virus. Parvovirus-like particles alone were detected in 14 diarrhoeal and 50 normal faeces, astrovirus-like particles in 3 normal faeces, "round" viruses in 4 normal faeces, coronavirus in 2 diarrhoeal and 5 normal faeces, CVLP in one diarrhoeal and one normal faeces, rotavirus in 2 normal faeces, papova-like virus in one normal faeces, both parvovirus-like particles and coronavirus in 2 diarrhoeal and 2 normal faeces, parvovirus-like particles and rotavirus in one normal faeces and parvovirus-like and papova-like virus in one normal faeces. The significance of these findings in canine and human disease is discussed.

Forty-one Feulgen-stained cervical imprint smears were analyzed by means of the SAMBA 200 cell image processor in order to quantitatively score human papillomavirus (HPV) 16-18-induced morphonuclear modifications as assessed by morphometric, densitometric, and textural parameters. Molecular hybridization technology using 16 and 18 type specific genetic probes made it possible to divide our series into three groups: Group 1, containing noninfected smears; Group 2, containing "suspicious", i.e., borderline positive, smears; and Group 3, those related to infected patients. Our results show that nuclei from infected smears are much more hyperchromatic and bigger than those arising from noninfected smears. This quantitative description of HPV 16-18-induced chromatin modifications enabled us to create preliminary data banks which could lead to an objective and reproducible grading of unknown cases. This approach is now being prospectively assessed on a large series of cases because the value of the current study is limited until the data bank is tested against unknown specimens with a broader spectrum of HPV infection.

Human papillomavirus (HPV) is an agent responsible for squamous cell tumors (verrucae, condylomata and papillomas) at various sites of the body, the oral cavity included. Due to the recently pointed-out association between these HPV lesions and the squamous cell dysplasias and malignancies in the uterine cervix, in the bronchus and in the larynx, the present work was carried out to assess, whether morphological signs (cytopathic effects of HPV) or HPV antigens could be found in lesions of oral squamous cell carcinomas. Biopsies from 40 cases of oral squamous cell carcinomas were surveyed by light microscopy with special emphasis on the presence of the histopathological features suggesting an HPV lesion, i.e. whether flat, inverted or papillomatous condylomas are present concomitantly with the malignancy. All specimens were also subjected to immunoperoxidase staining with anti-HPV serum to disclose the possible HPV antigens in the lesions. Morphological signs of the flat-type HPV lesion were found in 4 cases (10%), those of an inverted type in 3 cases (7.5%), and those of a papillomatous type in 9 cases (22.5%). Epithelial cells (mostly koilocytes) showing HPV-positive nuclei were disclosed in 5 papillomatous lesions, in 2 inverted lesions and in 1 flat lesion. The results suggest that HPV might be the agent involved in the development of at least certain special types of oral squamous cell carcinomas; albeit further confirmatory evidence with other techniques (DNA hybridization) is still needed.

Case report of a 21 years old woman with cervical, vaginal and vulvar condyloma. We have demonstrated the presence of human papilloma viruses (HPV) with help of in-situ-hybridisation.

The male sexual partners of 156 women with human papillomavirus infection of the cervix uteri were examined. In 120 men (77 per cent) penile lesions were found on examination of the penis via a colposcope (peniscopy) the most common of which were flat acetowhite lesions (53 per cent). Predilection sites of lesions were the urethral meatus and the corona glandis (23 and 19 per cent of the lesions, respectively). Using filter in situ hybridization human papillomavirus-deoxyribonucleic acid was found in penile smears of 61 men (39 per cent). The cancer-associated viral types (human papillomavirus 16 and 18) were identified in 75 per cent of the human papillomavirus positive men. Viral types of sexual partners were identical in 87 per cent. The correlation between infections with human papillomavirus 16 and 18, and the severity of the cervical lesion was significant in corresponding sexual partners. Our results support the hypothesis that male sexual partners represent a risk factor by acting as a reservoir for genital infections with papillomaviruses. The majority of human papillomavirus infections are of subclinical character. They require sensitive diagnostic techniques, such as peniscopy and hybridization for their identification. Detection and treatment of subclinical human papillomavirus infection in men may be important for the prevention of genital cancer in women.

Correlation averaging was used to enhance the three-dimensional spherical harmonic expansion of human wart virus particles from randomly-oriented negatively-stained electron microscopic images in holey grids. The reconstruction reveals the coat protein arrangement and variations in stain distribution inside the virus core. Selection rules imposed by the spherical symmetry on the harmonic expansion component are obtained indeed for angular momenta, l, lower than 11, but cannot be shown for l greater than or equal to 11, due to non-spherical distortions of about 1/11th of the radius of the virus. This prevents us from resolving fine details of packing of the protomers on the surface lattice. Distribution of stain inside the virus core, and its spherical symmetry, is reconstructed and may be related to nucleic acids and core protein structure.

We studied 89 seborrheic keratoses. In four cases, light microscopy and electron microscopy demonstrated that seborrheic keratoses may be associated with human papillomavirus (HPV) infection.

A 24-year-old male drug-dependent patient with perianal exophytic condylomata acuminata is reported. An increased production of papillomaviruses could be demonstrated by means of electron microscopy and immunocytochemistry (peroxidase-antiperoxidase technique). This is in contrast to earlier investigations of exophytic condylomata acuminata, which revealed characteristically small amounts of mature papillomavirus particles. The basis for the different form of papillomavirus infection of the anogenital region in this drug-dependent patient is discussed.

Papillomaviruses have been identified as the causative agents of benign epithelial proliferation in many animals including man. Recent evidence has shown that each viral type will only infect a specific species and tissue. Furthermore, certain papillomavirus types have been found associated with lesions capable of malignant conversion, particularly in response to secondary physical or chemical factors. Many advances have been made in identifying particular papillomavirus types inducing papillomas capable of progression and in the identification of the virally encoded functions involved in viral replication and cellular transformation.

A papovavirus, CCL 33 PV, isolated from a porcine cell line, CCL 33 (GT), was characterized. Based on a comparison of four isoenzyme systems, CCL 33 (GT) and CCL 33 (ATCC), obtained directly from the American Type Culture Collection, appeared to be the same. In addition to the previously characterized C-type virus of CCL 33 cultures, CCL 33 (GT) produced CCL 33 PV in high quantity, but CCL 33 (ATCC) produces a papovalike virus, presumably the same as CCL 33 PV, in barely detectable amounts. Serological results showed that CCL 33 PV is apparently identical to a papovavirus (SPV) isolated by Newman and Smith after inoculation of CCL 33 with concentrated porcine trypsin. These studies extend the characterization of this papovavirus, demonstrating that CCL 33 PV is weakly hemagglutinating after neuraminidase treatment, has a high affinity for a component of fetal bovine serum and is highly infectious in appropriate porcine cell systems rather than very defective as reported previously. Moreover, it was concluded from the data that CCL 33 PV is probably indigenous to the CCL 33 porcine cell line.

DNA-in situ hybridisation on epithelial cells taken from cervical swabs of 101 different patients cytologically diagnosed as III D to V (Munich classification 1975) were performed using 32P-labelled DNA of the human papilloma virus (HPV) types 6, 11, 16 and 18 as smears. The following correlations between the cytological classification and DNA hybridisation were obtained: 55 out of 58 women (94.8%) having a normal Pap smear (Pap II) were negative by hybridisation as well, three cases contained HPV 6 or 11.59.3% of patients revealing abnormalities characteristic of a papillomavirus infection reacted with the 32P-labelled DNA, while HPV 6/11 or 16/18, respectively, occurred at approximately the same rate of frequency. In cases with severe dysplasia, carcinoma in situ or invasive cervical carcinoma (IVa, IVb, V) HPV 16 or 18 were more frequently found (67%) than in materials with mild dysplasia (IIID; 32% positive). HPV 6 or 11, on the other hand, was present only once in nine IVb/V-cases (11%), in 17% of IVa-cases, but in 32% of patients with mild dysplasia. Infections with all different virus types were found in 9% of the IVa-und 16% of the IIID-cases. The high prevalence of HPV 16 or 18 in CIS as well as in invasive carcinomas is in line with the biological correlation between both types of lesions. HPV 6 or 11 is more frequently associated with the usually reversible mild dysplastic epithelial alterations. The DNA-in situ swab hybridisation should, therefore, be of diagnostic value and might help to optimise the therapy for the individual patient.

High mortality rates have been reported in budgerigars between one and 15 days of age in 19 aviaries in the Province of Quebec. The most consistent signs of disease were abdominal distention, lack of down feathers on the back and abdomen, lack of filoplumes on the head and neck, and retarded growth of the tail and contour feathers in birds that either survived or died later. Internal gross lesions were hydropericardium, enlarged heart and liver with multiple pinpoint white spots or large, yellow foci, pale or congested kidneys, congested lungs, and ascites. Histologic examination revealed large, slightly basophilic inclusion bodies in the enlarged nuclei of many different cells. These inclusion bodies were composed of viral particles. Multiple foci of coagulation necrosis were scattered in the myocardium and liver parenchyma, and granulovacuolar degeneration was common in renal tubular epithelial cells. Ballooning degeneration was multifocal in the epidermis and very extensive in the epithelial cells of developing feather follicles, and this led to their partial or complete destruction. Viral particles 50 to 55 nm in diameter were observed in negatively stained preparations from different organs of affected birds. These particles had the size and morphology of a papovavirus. In experimentally infected 25-day-old budgerigars, histologic examinations revealed the presence of intranuclear inclusions in hepatocytes, epithelial cells of the kidney tubules, and reticular cells of the spleen, despite the absence of clinical signs. We feel that this disease is caused by a papovavirus-like agent that can replicate in many tissues of the body, causing widespread lesions responsible for the high mortality rate of very young budgerigars and for the absence and/or incomplete development of feathers.

The authors describe the electron microscope findings in a case of papilloma of the uterine cervix. In the nuclei of single koilocytes spherical inclusions measuring 47-56 nm were found. The ultrastructural pattern of the particles is entirely identical with one of the human papilloma virus particles.

Histological investigations of verrucae vulgares of the hand and of plantar warts show striking differences between HPV-1-induced warts and those caused by the virus type HPV-4. In warts caused by HPV-1 massive vacuolization and disturbance of epidermal cell differentiation through the entire thickness of the epidermis is predominant whereas in HPV-4 stimulated warts cell differentiation is impaired to a much lesser degree. Despite the presence of inclusions and of vacuolized cells a thick stratum granulosum and a compact horny layer are formed in the latter type.

The complete genome of the hamster papovavirus (HaPV) which was isolated from virions found in multiple skin tumors of the Syrian hamsters was measured by electron microscopy and cloned in Escherichia coli using the certified plasmid vector pBR322. The cloned viral DNA were characterized by digestion of the recombinant DNA with various restriction enzymes followed by comparison of their electrophoretic mobilities in agarose gels with that of similarly digested uncloned DNA and by electron microscopy to determine the genome size of cloned HaPV DNA. The restriction enzyme analysis of the cloned HaPV DNA showed the same cleavage pattern as the corresponding fragments from the uncloned DNA. No major insertions or deletions could be detected by heteroduplex analysis between cloned HaPV DNA and the starting material. The estimated genome size of 5.52 kb for HaPV DNA is approx. 300 bases larger than those determined for other known papovaviruses as SV40 or polyoma.

A large number of different papillomavirus types have been identified. Several viruses often infect the same species and each virus is associated with a defined tissue. Recent evidence has shown that certain benign lesions can undergo malignant transformation in both animals and humans in response to genetic or environmental factors. Fine mapping of the structure and function of the viral genome may enhance our understanding of the interaction between the virus and the cofactors involved in malignant transformation.

A technique is described whereby inclusion-bearing cells identified by light microscopy in stained smears of urinary sediment were reprocessed for examination in the electron microscope. The nuclei of the abnormal cells were found to contain numerous virus particles, 35 nm in diameter, which morphologically resembled papovaviruses. The technique was applied in this case to identify further the virus producing the cytopathic changes in the Papanicolaou smear. It could be particularly valuable for retrospective studies of mounted cytological or histological material when suitable specimens are no longer available for virological investigation.

The finding of virus particles by transmission electron microscopy (TEM) in fixed cervical and vaginal scrape material and their identification as papilloma viruses by negative staining is described. The colposcopic, cytological, and histological appearances in a group of 10 patients in whom virus particles were seen are discussed and the possibly considered of an association between papilloma virus infection and the development of cervical intraepithelial neoplasia (CIN) and multifocal intraepithelial neoplasia of the female lower genital tract.

In 202 women with koilocytotic atypia in cervical smears, 136 had predominantly small condylomata of the uterine cervix, and 66 had cervical intraepithelial neoplasia (CIN) of varying degree either with koilocytosis of the neoplasia or associated with condylomata. Koilocytosis correlated well with the histological diagnosis of condylomata, but occasionally it obscured the cytological evidence of CIN. Human papilloma virus particles were found in the cells of condylomata in 10 cases and in those of CIN II with koilocytosis in two cases of 21 examined ultrastructurally. There was evidence that the condyloma of the uterine cervix is a well-defined morphological entity and also that cytopathie changes similar to those seen in condylomata are present in some cases of CIN.