Vol. 28, No. 1

JOURNAL OF VIROLOGY, Oct. 1978, p. 337-343 0022-538X/78/0028-0337$02.00/0 Copyright © 1978 American Society for Microbiology

Printed in U.S.A.

Characterization of K Virus and Its Comparison with Polyoma Virus SHEILA B.

BOND,`*

PETER M. HOWLEY,2

AND

KENNETH K. TAKEMOTO'

Laboratory of Viral Diseases, National Institute ofAllergy and Infectious Diseases,' and Laboratory of Pathology, National Cancer Institute,2 Bethesda, Maryland 20014 Received for publication 28 March 1978

The antigenic relationship between the two murine papovaviruses, K virus and polyoma virus, was examined by serological techniques to determine whether they shared any antigenic components. No cross-reactivity was found associated with the viral (V) antigens by the indirect immunofluorescence, neutralization, or hemagglutination-inhibition tests. The tumor (T) antigens expressed in transformed cells or cells productively infected by either K or polyoma virus did not cross-react by indirect immunofluorescence. An antigenic relationship was detected, however, among the late proteins of K virus, polyoma virus, simian virus 40, and the human papovavirus BKV, when tested with either hyperimmune sera prepared against polyoma virus and simian virus 40 or sera prepared against disrupted virions. The nucleic acids of K and polyoma viruses were compared by agarose gel electrophoresis and restriction endonuclease analysis. No nucleotide sequence homology between the genomes of these two viruses was detectable by DNA-DNA hybridization techniques under stringent conditions. The genome of K virus was found to be slightly smaller than that of polyoma virus, and the cleavage patterns of the viral DNAs with six restriction endonucleases were different. These findings indicate that there is little relationship between these two murine papovaviruses.

The murine papovavirus K virus was first identified by Kilham in 1952 (17). On the basis of its physical structure and the demonstration that its nucleic acid was double-stranded DNA (21, 23), it was assigned to the papovavirus family in 1963. To date, K virus and polyoma virus are the only recognized murine members of the polyoma genus of papovaviruses (6). Biological and biochemical characterization of K virus has been limited primarily by lack of a cell culture system for its propagation. Inoculation of newborn mice with K virus produces a fatal pneumonia (19) with a well-characterized pathogenesis (8, 20), and it has been found to be endemic in many wild mouse populations (4, 12, 26). Although K virus differs from polyoma virus and other well-characterized papovaviruses by its inability to induce tumors, it has been shown to transform mouse cells in culture (29). Because of these somewhat unusual properties, we decided to characterize K virus more fully and to examine its relationship to polyoma virus and the other known papovaviruses. In this study we have examined the serological relationship between the virion (V) antigens, the early or tumor (T) antigens, and also antigens exposed by disruption of the viral particles. In addition, we have examined the K virus genome

and compared it with that of polyoma virus by use of restriction endonucleases and DNA-DNA

hybridization techniques. MATERIALS AND METHODS Viruses. The large-plaque strain of polyoma virus was obtained from Tom Benjamin (Harvard Medical School, Cambridge, Mass.) and was grown in primary cultures of baby mouse kidney cells. The MM strain of BK virus [BKV(MM)] was grown in primary cultures of human embryonic kidney. Simian virus 40 (SV40) (small-plaque strain) was grown in CV-1 cells. Purification of virus. K virus (obtained from J.

Parker, Microbiological Associates, Bethesda, Md.) was prepared by intraperitoneal inoculation of newborn mice with 0.05 ml of a stock virus pool. At 8 to 10 days after infection, the livers of dead and moribund mice were removed, minced with scissors, and dispersed by trypsinization with the use of a magnetic stirrer. The cells were lysed by incubating at 37°C for 30 min in 1% sodium deoxycholate. The virus-containing suspension was clarified by centrifugation at 8,000 rpm for 15 min; the sediment was reextracted with trypsin and sodium deoxycholate three or four times, and the pooled clarified supematant fluids were layered over a cesium chloride cushion (1.34 g/cm3) and centrifuged in an SW25 rotor for 2 h at 23,000 rpm. The virus band which formed below the interface was collected, and the virus was purified by isopycnic 337

338

BOND, HOWLEY, ANI) TAKEMOTO

banding in cesium chloride in an SW56 rotor at 35,000 rpm for 18 h. The virus yield from 10 litters of mice varied from 3 to 7 mg. Antisera. Mouse antiserum to K virus was obtained from the Office of Program Resources and Logistics, National Cancer Institute. This serum contained antibodies to both T and V antigens of K virus (29). By the fluorescent-antibody (FA) test it had a titer of 1:2,560 against infected cells and 1:640 against K virus-transformed cells. Polyoma T antiserum was obtained from serum of hamsters carrying transplantable polyoma tumors. This serum pool had a titer of 1:1,280 by the FA technique. Antipolyoma serum for various serological tests was prepared in rabbits by intravenous inoculation of purified polyoma virus followed by intramuscular inoculation of polyoma virus mixed with complete Freund's adjuvant at 3-week intervals. Hyperimmune polyoma and SV40 antiviral sera were prepared by the intraperitoneal inoculation of hamsters with purified preparations of the virus followed by subcutaneous inoculation of a mixture of virus with complete Freund's adjuvant at 3-week intervals. The hamsters were bled by intraorbital bleeding 2 weeks after the second immunization. The polyoma hyperimmune serum had a hemagglutination-inhibition (HI) titer of 1:50,000. Antisera to disrupted virus particles were prepared by the following procedure. Purified virus was suspended in glycine buffer, pH 10.5, containing 1% sodium dodecyl sulfate (SDS) and 0.1% mercaptoethanol at 37°C for 30 min. The disrupted virus was then dialyzed against phosphate-buffered saline and inoculated subcutaneously into hamsters. The animals were boosted twice at 3-week intervals with the same material mixed with complete Freund's adjuvant. Serological tests. HI tests for polyoma virus were carried out at 4°C with guinea pig erythrocytes. The indirect FA test was employed on air-dried and acetone-fixed cover slips of cells which had been infected 24 to 48 h previously with various viruses or of cells prepared from lungs of baby mice infected with K virus. The cover slips were stained by use of goat anti-mouse and goat anti-hamster globulin conjugated with fluorescein isothiocyanate. The neutralization test was performed by mixing equal volumes of polyoma virus of known infectious titer with diluted preparations of both anti-polyoma rabbit serum and anti-K virus mouse serum. After the reactants had been left at room temperature for 30 min, the infectivity was determined by plaque assay on monolayer cultures of baby mouse kidney cells. Viral DNA. Unlabeled K virus DNA was prepared directly from virions by incubation with 1% Sarkosyl at 50°C for 30 min, phenol extraction, and isopycnic centrifugation in cesium chloride-ethidium bromide as described for SV40 DNA (11). Unlabeled polyoma DNA, unlabeled SV40 DNA, and 32P-labeled polyoma DNA were similarly prepared. Unlabeled adenovirus 2 DNA was purchased from Bethesda Research Laboratories (Bethesda, Md.). Viral DNAs were mechanically sheared at 50,000 lb/in2 in a Ribi cell fractionator (Ivan Sorvall, Inc.) to a molecular size of 3.1 x 105 daltons prior to DNA-DNA reassociation experiments.

J. VIROL.

DNA-DNA reassociation. Denatured, mechaniwere allowed to reassociate in 0.14 M sodium phosphate buffer at 60°C, and samples removed at various times were analyzed for single- and double-stranded DNA by hydroxyapatite column

cally sheared DNAs

chromatography.

Restriction endonuclease cleavage of viral DNAs. Restriction endonuclease R HincII, R HindIII, R Hind-II + III, R HpaI, and R EcoRI were purchased from New England Biolabs (Lowell, Mass.). R KpnI was purchased from Bethesda Research Laboratories. In general, analytical reaction mixtures (0.03 ml) containing from 0.25 to 0.50 jug of viral DNA in the appropriate buffer and 1 U of enzyme were incubated at 37'C for 1 h. Cleavage products were determined by electrophoresis in either 1.4% agarose or composite 3.0% acrylamide-0.5% agarose

gels.

Slab gel electrophoresis. Analytical electrophoresis through 1.4% (wt/vol) agarose (Seakem) slab gels (17 by 12 by 0.3 cm) at 60 V for 20 h was carried out in a buffer containing 40 mM Tris, pH 7.8, 5 mM sodium acetate, and 1 mM EDTA. Analytical electrophoretic analysis of restriction endonuclease cleavage fragments of viral DNA in slab gels consisting of 3.0% acrylamide-0.5% agarose was under conditions previously described (15). The DNA bands were visualized after staining for 15 min with ethidium bromide (0.5 tig/ml) under UV light (28).

RESULTS

Lack of antigenic relationship between polyoma and K viruses. Tests to evaluate the possible antigenic relationship between polyoma virus and K virus were conducted by three different techniques: neutralization, HI, and FA. In the neutralization test, equal volumes of polyoma virus (5 x 106 PFU/ml) were mixed with mouse anti-K virus serum and rabbit antipolyoma serum (1:5 dilution) for 30 min at room temperature. Titration of the mixtures on monolayer cultures of baby mouse kidney cells by plaque assay showed that there was no reduction in the titer of polyoma virus which had been allowed to react with the K virus antiserum, whereas complete neutralization was observed with the polyoma antiserum. Since no cell culture system is available in which to evaluate the infectivity of K virus, similar neutralization tests could not be performed with K virus. An HI test of polyoma virus by K virus and polyoma antisera was performed as follows. Serial twofold dilutions of K virus antiserum were prepared and used in an HI test with 8 hemagglutinating units of polyoma virus and guinea pig erythrocytes. There was no inhibition of the reaction even at the lowest dilution (1:10) tested. Rabbit antipolyoma serum, on the other hand, inhibited agglutination with polyoma virus at a dilution of 1:2,560. Although K virus has been reported to agglutinate sheep erythrocytes (18),

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COMPARISON OF K VIRUS AND POLYOMA VIRUS

high concentrations of purified virus are necessary for agglutination; therefore, the reciprocal test with K virus and polyoma antiserum was not performed. In FA tests for viral antigens, polyoma and K virus-infected cells on cover slips were examined for reactivity with both polyoma rabbit antiserun and K virus mouse antiserum. In each case the infected cells reacted only with the homologous antisera. T antigens of polyoma and K viruses. The T antigens of polyoma and K viruses were examined by FA tests on transformed as well as infected cells. The polyoma-transformed mouse line (Py-T-54) and the K virus-transformed mouse line (K-C57-C2) were examined for the presence of T antigens with polyoma anti-T and mouse anti-K virus sera. The Py-T-54 cells were positive only with the polyoma anti-T serum, and the K-C57-C2 cell line was positive only with anti-K virus serum. Similar results were obtained in tests on polyoma and K virus-infected cells; positive staining was observed only in cells reacted with homologous antiserum. The T antigens of K and polyoma viruses are thus immunologically distinct and unrelated. These serological tests revealed no cross-reactivity between K virus and polyoma virus with respect to their infectivity, hemagglutinins, and viral or T antigens. Common papovavirus antigens. Shah et al. (27), using antiserum prepared against either disrupted viral particles or purified VP-1, have recently reported on the existence of an antigenic determinant which appears to be shared by all the members of the polyoma virus genus. Since we were unable to show antigenic relatedness between polyoma virus and K virus by standard serological methods, experiments similar to those of Shah et al. (27) were performed with antisera prepared by two different methods. (i) Cover slips of polyoma virus-, K virus-, SV40-, or BKV(MM)-infected cells were allowed

339

TABLE 1. Cross-reactivity among some papovaviruses as detected by FA with antisera to disrupted virus and hyperimmune antiviral sera Antibody to:

Antigen (infected cells)

Intact virusa SDS-disrupted virus Poly- PolyBKV oma vi- oma vi- SV40 K virus (MM) rus rus

40 80 10 80 K virus 160b 40 640 10 320 160 BKV(MM) 160 80 1,280 20 Polyoma vi- 40 rus 40 5,120 40 80 40 SV40 a Hyperimmune serum prepared with purified virus. b Reciprocal of the highest dilution giving a positive reaction.

ing differs from those of Shah et al. (27), who reported the titers of homologous and heterologous reactions to be about the same. Characterization of K virus DNA. Viral DNA extracted from virions of the other wellstudied papovaviruses, polyoma, SV40, BKV, and JCV, consists predominantly of closed circular, supercoiled molecules (form I), with small amounts of nicked, open, circular molecules (form II) (2, 3, 5, 15, 22). The majority of the DNA purified from K virions and examined by isopycnic centrifugation in cesium chlorideethidium bromide gradients cobanded with supercoiled SV40 DNA at 1.60 g/cm3, indicating that K virus DNA is also supercoiled. The DNA was further examined by agarose gel electrophoresis and found to consist of a mixture of form I and form II molecules (Fig. 1, lane e). The DNA appeared to be quite homogeneous with regard to size, and, based on the greater electrophoretic mobilities of both the form I and form II molecules, the K virus genome was slightly smaller than either SV40 DNA (Fig. 1, lane c) or polyoma DNA (Fig. 1, lane d). The smaller size of the K virus genome was also reflected in the greater electrophoretic mobility of the fullto react with hamster sera obtained after im- length linear K virus DNA than of linear SV40 munization with SDS-disrupted virions of poly- DNA (Fig. 1, lanes a and b). The molecular size of the K virus DNA calculated from the electrooma virus, K virus, or BKV(MM). Considerable cross-reactivity among the different papovavi- phoretic mobility of the full-length linear molecule in a composite 2.2% acrylamide-0.5% agaruses was detected with these sera, in confirmation of the observations of Shah et al. (27) rose gel with the use of the R EcoRI fragments of adenovirus 2 and the full-length linear SV40 (Table 1). (ii) Cover slips of cells prepared as in the genome as markers was 3.25 x 106 daltons based preceding experiment were allowed to react with on a size of 3.6 x 106 daltons for SV40 DNA hyperimmune hamster sera to polyoma virus or (data not shown). Restriction endonuclease analysis of K SV40. Cross-reactivity among these viruses could also be detected with hyperimmune sera virus DNA. To further characterize the genome of K virus and to compare it with that of polyprepared against intact virus (Table 1). The results presented in Table 1 show that oma virus, the cleavage patterns obtained with the homologous titers were higher than those a variety of restriction endonucleases were exfound with the heterologous reactions. This find- amined. R EcoRI, an enzyme which cleaves the

340

BOND, HOWLEY, AND TAKEMOTO

J. VIROL,.

FIG. 1. Agarose gel electrophoresis of K virus, polyoma virus, and SV40 DNAs. Samples (30 ,ul) containing

approximately 0.25 ,ig of unlabeled (a) full-length linear SV40 DNA cleaved with R EcoRI, (b) full-length linear K virus DNA cleaved with R KpnI, (c) form I SV40 DNA, (d) form I polyoma DNA, and (e) K virus DNA purified from virions were subjected to electrophoresis for 20 h at 60 V in 1.4% agarose as described. The

DNA was visualized and photographed under UV excitation after staining for 30 mmn with ethidium bromide

(0.5 Hg/ml).

polyoma genome at one unique site, cleaved K virus DNA into four fragments varying in size from 0.07 x 106 to 1.35 x 106 daltons (Table 2). R .KpnI recognized a set of sequences present twice in the polyoma genome and cleaved K virus DNA only once (Table 2). This enzyme converted the form I and form II molecules of K virus DNA to full-length linear (form III) molecules (Fig. 1, lane b). The patterns of fragments generated by cleavage of K virus DNA by the

restriction enzymes R HpaI, R HincII, R HindIII and R HindII + III are shown in Fig. 2, and these patterns are totally different from those obtained with cleavage of polyoma DNA (Table 2). Examination with six different restriction endonucleases therefore revealed no similarities between the digest patterns of K and polyoma virus DNAs, thus establishing biochemically that these two murine papovaviruses are distinct. The smaller size of the K virus

TABLE 2.

Cleavage of K and polyoma virus DNAs by restriction endonucleases No. of sites

Enzyme KpnI HinII HindIII HindII + III

HpaI

Polyoma' 2 2 2 4

K virus

0 1

2

1 3 3 6

Mol wt of K vi- SuxIuof K fragDNA frag virus rus ment mol mentab (x ' Wt (X 106) 3.25 3.25 3.23 1.69, 1.41, 0.13 3.34 2.00, 1.16, 0.18 1.41, 1.15, 0.27, 3.24 0.18, 0.13, 0.10

106;

1.82,1.41

3.23

1.35, 1.30, 0.66, 3.38 0.08 a The number of cleavage sites in polyoma virus DNA for each of the enzymes was obtained from the review by Fried and Griffin (9). 'Molecular weights determined from electrophoretic mobilities of fragments in composite 2.2% acrylamide-0.5% agarose gels, 3% acrylamide-0.5% agarose gels, or 5.0% acrylamide-0.5% agarose gels.

EcoRI

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COMPARISON OF K VIRUS AND POLYOMA VIRUS

VOL. 28, 1978

4

a

b

c

d

e

f

0 1

2

3 4

H

5Z

6

m

7

C)

8 > m 9 1 10 3

genome was confirmed by summing the molecular weights of fragments generated by each of the five restriction endonucleases used in ana11 lyzing K virus DNA. The molecular weight determinations made in this manner ranged from 12 3.23 x 106 to 3.38 x 106, with the molecular weight of 3.60 x 106 of SV40 used as a standard 13 (Table 2). Reassociation of polyoma DNA in the presence of K virus DNA. In view of the crossFIG. 2. Restriction endonuclease cleavage patreactivity detected with antisera prepared terns of K virus DNA. Samples (50 ,ul) containing 0.5 against disrupted viral particles, we decided to pg of K virus DNA cleaved with (a) R HpaI, (b) examine whether there was any polynucleotide R HincII, (c) R HindIII and (d) R HindII + III were composite 3.0% sequence homology between the DNAs of K and analyzed by electrophoresis ingela for 12 h as deacrylamide-0.5% agarose slab polyoma viruses. The homology was assessed by scribed. + III fragments of SV40 (lane R The HindII examining the effect of unlabeled K virus DNA e) and the R EcoRI fragments of adenovirus 2 (lane on the reassociation of 32P-labeled polyoma t) are included as markers. DNA. This technique has been used previously in examining the polynucleotide sequences shared by the DNAs of the primate papovavi- the primate papovavirus BKV, JCV, and SV40 ruses BKV, JCV, and SV40 (13-15). In the pres- genomes, no homology could be detected beence of a 200-fold excess of unlabeled K virus tween the DNAs of the murine papovaviruses DNA, no effect on the rate of reassociation of polyoma virus and K virus. Experiments examthe 32P-labeled polyoma DNA probe could be ining the relationship of these two viral genomes detected when the reaction was monitored by under less stringent conditions are in progress. hydroxyapatite column chromatography (Fig. DISCUSSION 3). In the presence of a threefold excess of unIn this report we have examined the relationlabeled polyoma DNA, the reassociation of the radiolabeled probe corresponded closely to the ship between the two known murine members curve for 100% homology as a positive control of the polyomavirus genus, K virus and polyoma (Fig. 3). In addition, neither a 200-fold excess of virus. No cross-reactivity was detected between the unlabeled SV40 DNA nor BKV DNA af- their viral antigens by HI, indirect FA, or neufected the reassociation of the polyoma DNA tralization tests. The T antigens of K and polyprobe, confirming earlier reports that homology oma viruses in transformed or infected cells were cannot be detected between polyoma and SV40 also shown to be unrelated when examined by DNAs or polyoma and BKV DNAs under these the FA technique. By immunoprecipitation methods, the K and polyoma virus T antigens stringent conditions (3, 16). Therefore, under conditions which demon- were also found to be immunologically distinct strate common polynucleotide sequences among (D. T. Simmons and K. K. Takemoto, unpub-

342

J. VIROL.

BOND, HOWLEY, AND TAKEMOTO

t

FIG. 3. Reassociation of radiolabeled polyoma DNA in the presence of unlabeled K virus DNA. Denatured, mechanically fragmented, 0P-labeled polyoma DNA (8.3 x 104 cpm/lg) was allowed to reassociate at a concentration of 1.56 x 10 2 ,ug/ml alone (0), in the presence of a 200-fold mass excess of unlabeled, mechanically fragmented, denatured K virus DNA (A), or in the presence of a threefold excess ofpolyoma virus DNA (0) for various periods of time in 0.14 M sodium phosphate buffer at 600C. The fraction of radiolabeled DNA remaining single stranded (f,,) was determined by hydroxyapatite column chromatography. The t112 for 32P-labeled polyoma DNA reassociating alone was 12.1 h. The dashed line with a slope of four is the theoretical curve for the radiolabeled polyoma DNA reassociating in the presence of a threefold excess of an unlabeled homologous DNA.

lished data). This confirms the earlier findings that the T antigens induced by these two viruses are immunologically unrelated as determined by complement-fixation tests (29). Antigenic relatedness could be detected between the structural proteins of K and polyoma viruses as well as between those of K virus and the primate papovaviruses, BKV(MM) and SV40, when antisera which were raised against SDS-disrupted virions were used. This confirms the observations of Shah et al. (27), who have established that there is a common antigenic determinant shared by members of the polyoma virus genus. Disruption of virions by either SDS or alkaline pH exposes antigenic determinants present in the capsid proteins which are not exposed on the surface of the intact virions. The

common papovavirus antigen apparently resides in VP-1 since Shah et al. (27) showed that antibody to VP-1 of SV40 reacted with cells infected with other papovaviruses. In this report we have shown that hyperimmunization with purified virus also results in the development of "broadreacting" antibody. Presumably, this is due to disruption of virus particles in vivo, thereby "exposing" or "releasing" the common antigen. The serological studies thus indicate that K virus has in common with polyoma virus only those antigenic determinants that are shared by all members of the polyoma virus genus. This is surprising since there is significant cross-relatedness of the V and T antigens among the primate papovaviruses BKV, JCV, and SV40 (1, 10, 24, 30). Two other recently described primate papovaviruses, SA12 and the stump-tailed macaque virus, also have cross-reacting T antigens with BKV, JCV, and SV40; however, the V antigens appear to be unrelated (25, 31). Thus, although significant relatedness can be demonstrated among the five primate papovaviruses, no such relationship can be found between the two murine papovaviruses. The genome of K virus is a supercoiled, double-stranded, closed, circular DNA molecule, as are the genomes of all the papovaviruses. No similarities were noted between the DNAs of polyoma virus and K virus when they were analyzed by six different restriction endonucleases, establishing biochemically that polyoma virus and K virus are distinct viruses. The lack of a strong antigenic relatedness between polyoma virus and K virus was supported by the hybridization studies, which detected no sequence homology between their genomes under stringent conditions. We would predict that under less stringent conditions, similar to those used by Ferguson and Davis (7), homology will be detected between K virus and polyoma virus because of the antigenic cross-reactivity as revealed by the studies with disrupted virus. The serological and biochemical studies presented in this paper indicate that, despite the fact that K virus and polyoma virus share a common host, K virus is as distantly related to polyoma virus as polyoma virus is to the primate papovaviruses, BKV, JCV, and SV40. Further studies examining the polynucleotide sequence relationships between K virus and other members of the polyoma virus genus with the use of less stringent hybridization conditions are in progress. LITERATURE CITED Tsai, and D. C. Gajdusek. 1975. Seroepidemiology of human papovaviruses. Am. J. Epide-

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miol. 102:331-340. 2. Crawford, L. V., and P. H. Black. 1964. The nucleic

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COMPARISON OF K VIRUS AND POLYOMA VIRUS

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C3H mice harboring Bittner milk agent. Science 116:391-392. 18. Kilham, L. 1961. Hemagglutination by K-virus. Virology 15:384-385. 19. Kilham, L., and H. W. Murphy. 1953. A pneumotropic virus isolated from C3H mice carrying the Bittner milk agent. Proc. Soc. Exp. Biol. Med. 82:133-137. 20. Margolis, G., L. R. Jacobs, and L. Kilham. 1976. Oxygen tension and the selective tropism of K virus for mouse pulmonary endothelium. Am. Rev. Respir. Dis. 114:45-51. 21. Mattern, C. F. T., A. C. Allison, and W. P. Rowe. 1963. Structure and composition of K virus and its relation to the "papovavirus" group. Virology 20:413-419. 22. Osborn, J. E., S. M. Robertson, B. L. Padgett, G. M. Zu Rhein, D. L. Walker, and B. Weisblum. 1974. Comparison of JC and BK human papovaviruses with simian virus 40: restriction endonuclease digestion and gel electrophoresis of resultant fragments. J. Virol. 13:614-622. 23. Parsons, D. F. 1963. Morphology of K virus and its relation to the papova group of viruses. Virology 20:385-388. 24. Penney, J. B., and 0. Narayan. 1973. Studies of the antigenic relationships of the new human papovaviruses by electron microscopy agglutination. Infect. Immun. 8:299-300. 25. Reissig, M., T. J. Kelly, Jr., R. W. Daniel, S. R. S. Rangan, and K. V. Shah. 1976. Identification of the stump tailed macaque virus as new papovavirus. Infect. Immun. 14:225-231. 26. Rowe, W. P., J. W. Hartley, and R. J. Huebner. 1961. Polyoma and other indigenous mouse viruses, p. 131-142. In R. J. C. Harris (ed.), Problems of laboratory animals diseases. Academic Press Inc., New York. 27. Shah, K. V., H. L. Ozer, H. N. Ghazey, and T. J. Kelly. 1977. Common structural antigen of papovaviruses of the simian virus 40-polyoma subgroup. J. Virol. 21:179-186. 28. Sharp, P. A., B. Sugden, and J. Sambrook. 1973. Detection of two restriction endonucleases in Haemophilus parainfluenzae using analytical agarose-ethidium bromide electrophoresis. Biochemistry 12:3055-3063. 29. Takemoto, K. K., and P. Fabisch. 1970. Transformation of mouse cells by K-papovavirus. Virology 40:135-143. 30. Takemoto, K. K., and M. F. Mullarkey. 1973. Human papovavirus, BK strain: biological studies including antigenic relationship to SV40. J. Virol. 12:625-631. 31. Valis, J. D., N. Newell, M. Reissig, H. Malherbe, V. R. Kaschula, and K. V. Shah. 1977. Characterization of SA12 as a simian virus 40-related papovavirus of chacma baboons. Infect. Immun. 18:247-252.

Characterization of K virus and its comparison with polyoma virus.

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