Melanotic neuroectodermal tumor of infancy (pigmented melanoameloblastoma). The paper describes a large pigmented maxillary tumor in a 46-day-old infant. The histological examination showed the tumor to be a melanotic neuroectodermal tumor of infancy (MNTI). No raised urinary levels of vanilmandelic acid could be shown. The tumor was surgically removed and during a 5 1/2-year follow-up period there was no clinical evidence of recurrence. The origin, behavior and treatment of these rare tumors are discussed.


Exchangeable sodium in angiotensinogenic and nonangiotensinogenic renovascular hypertension. Previous studies have suggested that angiotensin II and sodium can act as alternative mechanisms in maintaining high blood pressure in chronic renovascular hypertension, In the present study, exchangeable sodium was measured in rats in which angiotensin II has been confirmed or excluded as the main cause of the hypertension. To determine the degree of participation of angiotensin II in the maintenance of the high blood pressure, we studied the mean blood pressure response to an angiotensin antagonist (1-Sar-8-Ala-angiotensin II) and to a converting enzyme inhibitor (SQ20,881). Rats with a decrease in blood pressure of less than 20 mm Hg, in response to both inhibitors, were classified as nonresponders; those with a decrease of 20 mm Hg or more, as responders. Fifty percent of the rats with two-kidney hypertension were nonresponders, and they had lower blood pressure and plasma renin activity than the responders. Further, these two-kidney, hypertensive, nonresponder rats had normal exchangeable sodium. The two-kidney hypertensive responders, on the other hand, had significantly higher exchangeable sodium than both the two-kidney, hypertensive nonresponders and the two-kidney control rats. These results suggest that angiotensin II and exchangeable sodium do not play a major role in the maintenance of the high blood pressure in the two-kidney hypertensive nonresponders. However, there appears to be an abnormal relationship between renin and exchangeable sodium in the two-kidney hypertensive responders that could contribute to the maintenance of the hypertension.


Hemodynamic and antihypertensive effects of captopril, an orally active angiotensin converting enzyme inhibitor. Captopril inhibits angiotensin II formation and bradykinin degradation in vivo. Eleven patients with essential hypertension (EH) and four patients with renovascular hypertension (RVH) were treated with captopril for periods ranging from 3 days to 12 months. All patients had a diastolic blood pressure (DBP) over 95 mm Hg after receiving a placebo for 3 days. Captopril given in ascending doses (10-1000 mg/day) caused normalization of blood pressure in all but three patients, one with severe RVH whose pressure fell 11%, one patient with severe EH, whose pressure fell 27%, and one with EH whose blood pressure fell 8.5%. The average control DBP in patients with EH was 113.7 +/- 5.5 (SE) mm Hg and fell to 89.9 +/- 3.6 mm Hg (p less than 0.001), while DBP in patients with RVH fell from 110.7 +/- 7.6 mm Hg to 94.5 +/- 8.2 (p less than 0.005). All patients were studied in balance on a 100 mEq sodium (Na) diet. Plasma renin activity (PRA) versus 24-hour urinary Na excretion increased sevenfold during therapy while converting enzyme activity fell by about one half. The magnitude of the blood pressure response was not related to control PRA. Cardiac output was estimated by echocardiography during placebo administration and during maintenance therapy with captopril. A significant change was not observed. Total peripheral resistance fell an average of 18.9% (p less than 0.05) in 11 of the 13 patients in whom the measurement could be made. It is concluded that captopril effectively lowers blood pressure in patients with EH or RHV by reducing total peripheral resistance.


Regional odontodysplasia.

Regional odontodysplasia. Regional odontodysplasia was diagnosed in a girl aged 6 with lesions of a number of deciduous and permanent teeth in the right lower and upper quadrants. The literature comprises reports on some 50 cases of regional odontodysplasia, including only one patient who also showed involvement of both the maxilla and the mandible.


Type 1 diabetes and Coxsackie virus infection. The role played by viruses in the aetiopathogenesis of type 1 diabetes mellitus has been studied by several authors; in particular the importance of Coxsackie virus B4 infection has been stated by some authors and not confirmed by others. 43 diabetic children were studied at the time of the diagnosis of the disease. No viruses could be isolated from stools; the titres of anti-Coxsackie viruses B1 to 6 complement fixing antibodies and anti-Coxsackie virus B4 neutralizing antibodies, compared to controls, indicated that Coxsackie virus infection was not associated with the onset of diabetes in these children. A cross-reaction with anti-Coxsackie viruses sera and a human pancreas demonstrated that there are not antigens in common between these viruses and the human pancreas.


Extrarenal Wilms' tumour: a review and case report. A case of extrarenal nephroblastoma in a girl aged 5 1/2 years is reported. From a review of the few other published instances, it is apparent that extrarenal Wilms' tumours are rare occurrences which present a number of unanswered questions of great theoretical interest, particularly as far as their relation to teratomas is concerned.


Potassium fluxes and ouabain binding in growing, density-inhibited and Rous sarcoma virus-transformed chicken embryo cells. Potassium fluxes, ouabain binding, and Na+ and K+ intracellular concentrations were determined for cultures of growing normal, density-inhibited and Rous sarcoma virus-transformed chicken embryo fibroblasts. No significant differences in K+ influx or ouabain binding were detected between growing normal cells and Rous sarcoma virus-transformed cells; however, ouabain binding and ouabain-sensitive K+ influx were 1.5- to 1.8-fold lower in density-inhibited cells. Thus, potassium influx in this system can be classified as a growth-related, but not transformation-specific change. As determined by both flame photometry and radioisotopic (42K) equilibration, growing normal and density-inhibited cells had similar potassium contents, whereas transformed cells exhibited 1.4-fold higher potassium levels. Sodium ion levels, as measured by flame photometry, were also 2- to 4.5-fold higher in transformed than normal or density-inhibited cells. Complementary studies of potassium efflux showed a 1.3- to 1.5-fold higher rate (based on the percentage of pool exiting the cell) in growing normal versus density-inhibited or transformed fibroblasts. Because of the larger potassium pool in transformed cells, efflux based on absolute number of potassium ions is similar in normal and transformed chicken embryo fibroblasts.


Periodate oxidation analysis of carbohydrates. XIII. Simultaneous gas chromatographic determination of the aldehydes in the periodate oxidation products of non-dialyzable urinary carbohydrate materials as diethyl dithioacetals. The aldehydes in the periodate oxidation products of non-dialyzable urinary carbohydrate materials were determined simultaneously by gas chromatography of their diethyl dithioacetal derivatives. The yields of aldehydes for normal male urine varied considerably among subjects, but their molar ratios were almost constant. The average values for glyceraldehyde/glyoxal and lactaldehyde/glyoxal molar ratios were 0.41 and 0.27, respectively. The average amount of L-fucose at the non-reducing terminals of carbohydrate chains, as estimated from the yield of lactaldehyde, was about 5 mg/day.


Histochemical studies on Raillietina (Raillietina) johri (Cestoda: Davaineidae). II. Nucleoside diphosphatase and thiamine pyrophosphatase. Thiamine pyrophosphatase and nucleoside diphosphatase have been studied histochemically in Raillietina (Raillietina) johri. Thiamine pyrophosphatase activity has been observed in the tegument, subtegumental muscle, subtegumental cells, medullary parenchyma, excretory canal and various reproductive structures like testes, ovary, vas deferens, spermatozoa and vitellaria. Eggs exhibit moderate enzyme activity. Various nucleoside diphosphates have been found to be hydrolyzed by thiamine pyrophosphatase. CaCl2, MgCl2 and MnCl2 each activated the enzyme at a final concentration of 6 mM whereas cysteine, reduced glutathione and PCMB inhibited the enzyme activity at a final concentration of 10 mM, 10 mM and 20 mM, respectively. KCN and NaF had no effect on the enzyme staining at concentration as high as 50 mM and 30 mM, respectively. Possible roles of the enzyme in the parasite have been discussed.


Cytomegalovirus infection in guinea pigs. III. Persistent viruria, blood transmission, and viral interference. Chronic persistent infection with cytomegalovirus (CMV) was studied in random-bred Hartley and inbred strain 2 guinea pigs. Infectious virus was isolated from the urine, kidney, spleen, pancreas, salivary gland, and cervix, but not from buffy coat of persistently infected guinea pigs. Strain 2 animals developed a high rate of chronic viruria, which was not related to isolation of CMV from renal tissue. In female strain 2 guinea pigs viruria was more than twice as prevalent as in males (56% vs. 24%). Transfusion of buffy coat from persistently infected strain 2 animals resulted in CMV infection in both isogenic and allogenic blood recipients, but buffy coat from uninfected donors did not activate CMV in persistently infected isogenic and allogenic blood recipients. Experimental CMV infection of young strain 2 guinea pigs interfered with the expression of guinea pig herpes-like virus, a common endogenous virus in strain 2 animals.


Quantitative determination of delta 9-tetrahydrocannabinol in cadaver blood. We have described a highly sensitive, accurate, and selective method for the determination of delta 9-THC in postmortem blood. Although the method requires the use of fairly complex and sophisticated equipment, the procedure is straightforward and has been reproduced many times in our laboratories. It is thought that this procedure will serve as a useful adjunct to the forensic scientist in his armamentarium for determining physiological levels of important, abused drugs in postmortem analysis.


The value of the Duquénois test for cannabis--a survey. The Duquénois test is employed throughout the world as part of the identification procedure for cannabis. A survey of results published for more than 400 herbally derived and botanical materials originating from some 270 different plant species is presented. A survey of results from some 200 organic compounds is also presented. The data show that the D-L modification is the most specific. There is no published report of an obviously botanical material, apart from cannabis, that gives a positive D-L test. Apart from cannabinoids, some resorcinol derivatives give a positive test. All of the materials are easily distinguished from cannabis by TLC.


The core technique in the determination of age at death of skeletons. This study proposed an histological method of estimating age at death in skeletons that uses a 0.4-cm-diameter core of cortical bone. Age-estimating regression equations were generated from data obtained from the analysis of bone cores taken from femurs, tibiae, humeri, and ulnas of cadavers. When the regression equations generated in this study were applied to eight forensic science cases, accurate ages at death were estimated.


Variation in the virulence of strains of Mycoplasma pulmonis related to susceptibility to killing by macrophages in vivo. The virulence of five strains of Mycoplasma pulmonis, as judged by their ability to survive in the respiratory tract and induce pneumonia in CBA mice, was related to the ability of viable organisms to persist in the peritoneal cavity. This appeared to be the result of differences in the ability of the strains to resist killing by peritoneal macrophages in vivo. It is suggested that resistance to phagocytosis by macrophages is an important determinant of virulence for M. pulmonis.


Isolation and partial characterization of four plasmids from antibiotic-resistant thermophilic bacilli. Twenty-nine antibiotic-resistant isolates of thermophilic bacilli were examined for the presence of covalently closed circular duplex DNA molecules by agarose-gel electrophoresis and caesium chloride-ethidium bromide density gradient centrifugation. Five of the 29 strains tested contained covalently closed circular molecules. Two of the streptomycin-resistant strains contained the same two plasmids: pAB118A of molecular weight 4.9 X 10(6) (7.0 kilobases) and pAB118B of molecular weight 3.0 X 10(6) (4.3 kilobases). Two of the tetracycline-resistant strains each contained a plasmid (pAB124) of molecular weight 2.9 X 10(6) (4.14 kilobases), while a third harboured a small plasmid (pAB128) of molecular weight 2.5 X 10(6) (3.57 kilobases). These plasmids were digested with 19 different restriction endonucleases and the numbers of cleavage sites were determined. Transformation of Bacillus subtilis (168 (Trp-) with purified plasmid DNA indicated that pAB124 conferred tetracycline resistance on the host.


The ecology of gonococcal plasmids.

The ecology of gonococcal plasmids. Of 261 strains of Neisseria gonorrhoeae examined for plasmids, 6 were plasmid-free, 217 contained only a small multicopy 2.6 X 10(6) dalton plasmid and 38 carried a large 24.5 X 10(6) dalton plasmid. Restriction enzyme digests and DNA-DNA hybridization studies revealed that the large plasmids isolated between 1940 and 1978 share a common core of DNA sequences (70 to 100%) and represent a group of closely related molecules.


Intracellular state of Epstein-Barr virus DNA in producer cell lines. The physical state of the Epstein-Barr virus (EBV) DNA in three cell lines which spontaneously produce virus has been characterized. Circular EBV DNA molecules have been found in P3HR-I, B95-8 and M8I cells. The size of the intracellular M8I circular EBV DNA molecules is comparable with the linear virus genome isolated from virus particles but the circular P3HR-I and B95-8 DNA molecules are shorter than the virion DNA . In addition to the circular form, some EBV DNA with physical properties indicative of integrated sequences was found in all three producer cell lines. There was no marked change in the amount of either the circular or integrated forms of EBV DNA when these producer cell lines were grown in the presence of phosphonacetic acid to suppress the spontaneous virus production which occurs in a small percentage of the cells in untreated cultures.


Molecular relationship of the DNA and RNA of intracytoplasmic A particles to mouse and mammary tumour virus genomes. We have directly tested the hypothesis that single-stranded cytoplasmic A particle-associated DNA (ss CAP DNA) is a murine mammary tumour virus (MMTV) proviral intermediate by hybridizing 125I-labelled ss CAP DNA to MMTV RNA or to MMTV complementary DNA (cDNA). 125I-labelled CAP DNA did not form duplexes with either MMTV RNA or MMTV cDNA. In contrast, CAP RNA hybridized readily with MMTV cDNA. CAP RNA contained all the MMTV virus sequences, but at lower concentrations than in MMTV virus particles. Single-stranded CAP DNA hybridized readily with mouse DNA from several sources. A study of the rate of hybridization of CAP DNA to cell DNA at various driver to probe ratios showed that its rate of hybridization is similar to that of tumour cell DNA reassociation. Further, in reassociation studies accelerated by using the phenol emulsion reassociation technique (PERT), CAP DNA originally isolated as single-stranded DNA was shown to reanneal (70%), to protect 125I-cell DNA to the same extent (67%) and to do so with kinetics of reassociation equivalent to that of mouse DNA. Although CAP DNA isolates were slightly enriched for MMTV specific sequences when compared to total cellular DNA, we conclude that the majority of ss CAP-associated DNA is equivalent to a random sample of total tumour cell DNA.


Phosphorylation of a ribosomal protein and of virus-specific proteins in cells infected with herpes simplex virus. In cells infected with herpes simplex virus a protein associated with the small subunit of ribosomes became phosphorylated. It was not detectably labelled with 14C-amino acids added after infection and is therefore probably a cellular protein. The phosphorylated ribosomal proteins from HSV-1- and HSV-2-infected cells were indistinguishable electrophoretically and had an apparent mol. wt. of about 48 000. Phosphorylation of the 48K protein was detected 2 to 3 h after infection and reached a maximum rate at 4 to 5 h. It was prevented by adding cycloheximide at 2 h, or actinomycin at 1.5 h p.i., or azetidine at the beginning of infection. The phosphorylation did not occur on reversal of a cycloheximide block in the presence of actinomycin, confirming that it is not caused by a virus alpha-polypeptide. Virus that had been irradiated with u.v. light, although still able to suppress synthesis of cellular protein and DNA, did not induce phosphorylation of the 48K ribosomal protein. Therefore the phosphorylation is not responsible for the suppression of host synthesis. The alpha polypeptides ICP 4, 0, 22 and 27 are also phosphorylated but, in contrast to that of the ribosomal protein, their phosphorylation does not depend on the synthesis of beta and gamma polypeptides. It is probably mediated by a host enzyme.


Complementation with ts mutants of herpes simplex virus. Tests to detect complementation between ts mutants of herpes simplex virus types 1 and 2 infectious centre and yield of virus assay were investigated. Progeny analysis of both intratypic and intertypic complementation showed a considerable proportion of recombinant or ts+ virus in the progeny; this was more marked in the infectious centre tests. Virus of intermediate temperature-sensitivity was produced in intratypic as well as intertypic complementation. Reduction in the input multiplicity of one of the two mutants in the test to extremely low levels did not prevent complementation, suggesting that non-infectious particles probably contribute to complementation. Demonstration of virus DNA synthesis in mixedly-infected cells at the non-permissive temperature was used to detect complementation between DNA-negative mutants.


The serological relationship of varicella-zoster virus to other primate herpesviruses. The serological relationship between viruses producing varicella-like exanthems in several subhuman primates, varicella-zoster virus (VZV) and herpes simplex viruses (HSV) types 1 and 2, was investigated. All the primate viruses were related to VZV and some to HSV also. These results show that there is a group of varicella viruses, comprising four distinct entities, some of which may be useful for the establishment of an animal model for varicella.


Virus-specific DNA sequences present in cells which carry the herpes simplex virus thymidine kinase gene. Two independently derived cell lines which carry the herpes simplex type 2 thymidine kinase gene have been examined for the presence of HSV-2-specific DNA sequences. Both cell lines contained 1 to 3 copies per cell of a sequence lying within map co-ordinates 0.2 to 0.4 of the HSV-2 genome. Revertant cells, which contained no detectable thymidine kinase, did not contain this DNA sequence. The failure of EcoR1-restricted HSV-2 DNA to act as a donor of the thymidine kinase gene in transformation experiments suggests that the gene lies close to the EcoR1 restriction site within this sequence at a map position of approx. 0.3. The HSV-2 kinase gene is therefore approximately co-linear with the HSV-1 gene.


Purification and identification of the RNA-dependent RNA polymerase of foot-and-mouth disease virus. The RNA-dependent RNA polymerase induced in BHK 21 cells by infection with foot-and-mouth disease virus has been isolated from the replication complex. It contains a major, virus-coded protein with mol. wt. 56 000 which appears from serological studies and tryptic peptide mapping to be the same as the virus infection associated (VIA) antigen and the protein P56 found in cells infected with the virus. Other virus coded proteins and a host cell protein were present in the partially purified replication complex but were removed by digestion with ribonuclease T1, leaving only the major virus coded protein. The tryptic peptide maps of the VIA antigen of the seven serotypes of the virus were similar, suggesting a high level of conservation in that region of the genome coding for the RNA polymerase of each type.


Quantitative electron microscopy of intracytoplasmic type A particles at kinetochores of metaphase chromosomes isolated from Chinese hamster and murine cell lines. We have successfully isolated and spread individual chromosomes of CHO-KI cells for electron microscopic karyotyping. Controlled preparation permitted a quantitative evaluation of the association between endogenous intracytoplasmic type A virus precursor particles and the centromeric region (kinetochores) of isolated chromosomes at prophase and metaphase. Our results suggest the transfer of type A particles from the cytoplasmic to the centromeric regions during early metaphase in conjunction with microtubule assembly at a time when the kinetochores are structurally mature and capable of binding microtubules. Preliminary comparable studies of the endogenous M432 virus propagated in murine cells support these findings. Our results are discussed with respect to mechanisms of intracellular movement of virus precursor particles and the interference with components of both the cytoskeleton and the mitotic apparatus.


Evidence for secondary structure in poliovirus virion RNA demonstrated by antibodies against double-stranded RNA. Poliovirus particle RNA has been considered to have little secondary structure. Specific binding of poliovirion RNA by antibodies against double-stranded RNA (dsRNA) has been confirmed and further characterized by a radioimmunoassay using Staphylococcus aureus protein A to precipitate the nucleic acid-antibody complex. Competitive binding studies between virion single-stranded RNA (ssRNA) and poly(I).poly(C) demonstrated that the dsRNA effectively inhibited binding of radiolabelled poliovirion RNA by the anti-dsRNA antibodies but the virion RNA was a poor competitor of radiolabelled ds RNA. This indicates that both RNAs reacted with the same species of antibodies in the sera, but avidity of the antibodies for dsRNA was greater than for the poliovirion RNA.


Conservation and variation in Orthopoxvirus genome structure. Orthopoxvirus DNA from representative strains of rabbitpox, vaccinia, monkeypox, variola, cowpox and ectromelia viruses was analysed by cleavage with restriction endonucleases HindIII, XhoI or SmaI. Genome mol. wt. vary from about 120 x 10(6) for rabbitpox to about 145 x 10(6) for cowpox. Physical maps of cleavage sites are similar and characteristic for strains of the same Orthopoxvirus type. The distribution of HindIII sites suggests that an internal region of mol. wt. about 30 x 10(6) is highly conserved between Orthopoxvirus genomes although some type-specific differences occur within this region, especially with strains of ectromelia virus. Conservation of internal sequences is less marked following analysis with XhoI although cleavages within this central region of particular genomes appear to represent a subset of preferred sites. Endonuclease SmaI cleaves exceptionally infrequently and distinguishes variola, monkeypox, vaccinia, cowpox or ectromelia viruses. Type specific differences result largely from extensive, near terminal variations in length and sequence. Representative Orthopoxvirus genomes have rapidly renaturing terminal restriction fragments confirming the presence of near terminal, covalent cross-links. Terminal restriction fragments from the same or different genomes generally cross hybridize indicating the presence of near terminal repetitions of mol. wt. up to 6 x 10(6) and which share at least a subset of common sequences. Variola strains however, appear to lack such sequences from one specific terminus which maps shorter than that of related viruses.


Cellular basis for susceptibility to mouse cytomegalovirus: evidence from tracheal organ culture. Tracheal organ cultures were prepared from strains of mice with known susceptibility to the lethal effects of mouse cytomegalovirus. When organ cultures from susceptible mice were infected with this virus, characteristic c.p.e. developed and virus titres greater than 10(5) p.f.u./ml were produced 2 to 3 weeks after infection. Identically infected tracheal organ cultures from resistant mice consistently failed to develop significant virus c.p.e. and produced 10- to 100-fold lower titres of virus. When fibroblast cultures were infected with mouse cytomegalovirus, however, no consistent differences between susceptible and resistant mouse strains were observed. Since cytomegalovirus replicates in epithelial cells both in vivo and in tracheal organ culture, these results suggest that resistance may be based in part upon innate genetic susceptibility of epithelial cells to cytomegalovirus infection.


Activation of the endogenous proviral genes in mouse cells is not followed by increased sensitivity to deoxyribonuclease I digestion. The disposition of chromosome proteins about the endogenous proviral DNA of BALB-c mouse has been studied. The sensitivity of the endogenous proviral DNA sequences to deoxyribonuclease I (DNaseI) was analysed in BALB-c mouse tissues (liver and spleen) and in the cell line JLS-V9 which does not produce virus. On all of these preparations the endogenous proviral DNA was as sensitive to DNase I digestion as total chromatin. Since the proviral genes in JLS-V9 cells were silent, it was of interest to study possible changes in the chromatin structure following virus induction by iododeoxyuridine. We could not detect any increase in the sensitivity of the endogenous proviral DNA to DNase I digestion following induction. The induction was very efficient, however, since 60% of the cells responded to produce intracellular virus antigens.


An outbreak of gastroenteritis associated with calicivirus in an infant home. In October 1977 an outbreak of acute infectious diarrhea occurred in an infant home in the city of Sapporo, Japan. Of 34 residents aged two to 20 months, 26 (77%) suffered from diarrhea. In ten of these patients the diarrhea was accompanied by vomiting. Electron microscopic examinations revealed typical calicivirus particles in eight faecal specimens, seven of which were from the group of 26 affected patients (28%) and one of which was from the group of eight infants without symptoms (13%). Immune electron microscopy tests for antibody responses against one of the isolated strains of calicivirus were carried out on 27 paired pre- and post-outbreak sera. Seroconversions were demonstrated in 18 of 19 (95%) affected infants and in six of eight (75%) unaffected infants. One patient with lack of antibody response was the youngest child--two months old. Periodic surveys on enteric viruses circulating in the home revealed that calicivirus was specifically associated with the outbreak of gastroenteritis. These observations provide further evidence for the causative role of calicivirus in acute gastroenteritis in children.


Comparison of four-layer radioimmunoassay and electron microscopy for detection of human rotavirus. A four-layer radioimmunoassay (RIA) using polystyrene beads as the solid phase, anti-rota guinea pig IgG as primary antibody, anti-rota rabbit IgG as secondary antibody, and 125I-labelled sheep anti-rabbit immunoglobulin as indicator antibody has been developed for the detection of human rotavirus in stool specimens. A comparison was made of the developed RIA, routine electron microscopy, and research electron microscopy of 147 unconcentrated stool specimens from patients with infantile gastroenteritis. In routine electron microscopy 17 (11.6%) false-positive or false-negative results were obtained when compared with research electron microscopy. Each specimen positive in research electron microscopy was positive in RIA, and six additional RIA positives were found from 58 electron microscopy negative specimens. A confirmatory test was necessary to find out marginally positive but nonspecific reactions in RIA. The developed radioimmunoassay is slightly more sensitive than research electron microscopy of unconcentrated stool specimens and considerably more sensitive and more specific than routine electron microscopy.


Replication of murine cytomegalovirus in murine Y-1 cells. Murine Y-1 cells, a continuous epithelial cell line derived from an adrenal adenocarcinoma, are highly susceptible to murine cytomegalovirus (MCMV) infection. Inoculation of these cells with MCMV results in production of infectious virus and is associated with development of both cytopathic changes and the expression of viral-specific nuclear and cytoplasmic antigens. Examination of the ultrastructure of infected Y-1 cells reveals MCMV in various stages of replication. Cytomegalovirus replication in this hormone-responsive epithelial cell line should provide a model system in which virus--host cell interactions and their effects on cellular metabolism can be studied.


The effect of sodium thiocyanate on virus structure. A study of two different virus types after elution from immunosorbent columns by sodium thiocyanate has shown that virus degradation occurs. The two virus types studied were hepatitis B and rotavirus. Hepatitis B antigen (HBAg) was only slowly degraded and retained many of its morphological features, although in an altered form; rotavirus was highly sensitive to the chaotropic agent, losing both its viability and its morphological integrity. During the process of disassembly a previously undetected inner component of rotavirus could be visualised, which was termed the "core."


Purification of poliovirus by affinity chromatography. Poliovirus type 1 (Mahoney strain) has been purified by retention on a Sepharose 4B-antibody column and elution with 3M K thiocyanate. The virus was recovered in excellent yield and its purity was as high as that achieved by detergent treatment followed by sucrose gradient centrifugation. The column could be re-used and its capacity was sufficiently high to make it a useful method for the purification of milligram quantities of the virus.


The occurrence of calicivirus in infants with acute gastroenteritis. Calicivirus was detected in 8 (1.2%) of 647 hospitalized patients during a survey of acute gastroenteritis in infants and young children, conducted between December 1974 and September 1977. Morphologically calicivirus was approximately 30 nm in diameter with an easily recognizable staining "star of David" configuration. Its buoyant density in cesium chloride was 1.38-1.40 gm/ml. The serologic response to calicivirus by immune electron microscopy (IEM) was demonstrated only in paired sera from patients who shed the virus in their stools. The results suggest that calicivirus might be a cause of acute gastroenteritis in infants and young children.


Evaluation of bone-marrow scanning with technetium-99m sulfur colloid in pediatric oncology. Eighty-six technetium-99m sulfur colloid (Tc-SC) bone-marrow scans in 56 pediatric oncology patients were reviewed. The distribution of the sulfur colloid was similar to that in adult bone marrow in normal children older than 10 yr, and involved progressively more marrow of the extremities in normal children under 10 years of age. After irradiation or chemotherapy there was an extension of the Tc-SC to peripheral marrow sites. There was also diminished uptake of the tracer in sites corresponding to irradiated areas. In most patients there was recovery of these defects by 6 mo after completion of therapy. Tumor replacement of the marrow was reflected in the scans, and the extent of the scan defect paralleled the course of the disease. In four patients, despite normal bone scans and radiographs, marrow-scan abnormalities due to tumor replacement were present and confirmed by needle aspiration and/or biopsy. In two other patients, the marrow-scan abnormality preceded radiographic and histologic evidence of tumor metastasis. Two patients who responded clinically showed persistent defects; biopsy in one revealed fibrosis. Technetium-99m sulfur colloid bone-marrow scanning appears to be a sensitive monitor of marrow alteration caused by metastases, irradiation damage, or tissue fibrosis in children receiving treatment for cancer.


Comparison of Tc-99m pyrophosphate and Tc-99m methylene diphosphonate in acute myocardial infarction: concise communication. This study compared Tc-99m pyrophosphate (PPi) and Tc-99m methylene diphosphonate (MDP) for myocardial infarct imaging in 24 patients with diagnosed acute myocardial infarction. The radiopharmaceuticals were administered randomly and interpreted without knowledge of the sequence used. Twenty-three patients (96%) had positive Tc-99m PPi scintigrams, but only 17 (71%) had a positive Tc-99m MDP study (P less than 0.05). In addition, a comparison of the relative intensity with each agent revealed greater intensity with Tc-99m in 21 cases, equal intensity in two cases, and less intensity in only one case (p less than 0.001). These findings support the superiority of Tc-99m PPi as the agent of choice for myocardial scintigraphy in acute infarction.


Scintigraphic, electrocardiographic, and enzymatic diagnosis of perioperative myocardial infarction in patients undergoing myocardial revascularization. To assess the incidence of perioperative myocardial infarction, 214 consecutive patients were evaluated 1-5 days after coronary bypass surgery, using Tc-99m pyrophosphate (TcPPi) myocardial imaging, serial electrocardiograms (ECG), and enzyme levels (SGOT, LDH, CPK). On the basis of the clinical course and scintigraphic, enzymatic, and ECG changes, the diagnosis of perioperative infarction was definite in 17 of 241 cases (7.9%) and probable in six of 241 (2.8%). In all of these 23 patients, TcPPi scans were abnormal; one additional patient had a false-positive scintigram. Only 13 of the 23 had ECG evidence of infarction, but there were no false positives. We set the threshold for abnormality of enzyme changes quite high, owing to experience in more than 900 postoperative patients (SGOT greater than 200, LDH greater than 500, CPK greater than 500 on the same day). Using these criteria, 22 of the 23 infarct patients had abnormal enzymes, and six others were falsely positive. These results indicate a relatively low sensitivity for the ECG in diagnosing perioperative infarction, but the lack of false positives suggests high specificity. The sensitivity and specificity of the enzymes and the TcPPi image were both excellent and quite similar; the main difference was a reduction of certainty of infarction with the enzyme criteria, caused by the six patients whose enzyme values were falsely positive. Considering its sensitivity, specificity, and ability to locate and to a certain extent quantitate necrosis. TcPPi imaging is probably the most valuable means of diagnosing perioperative myocardial infarction.


Biodistribution and pharmacokinetics of S-35-labeled 5-thio-D-glucose in hamsters bearing pancreatic tumors. 5-thio-D-glucose (5-TDG) exerts profound effects on rapidly metabolizing tissues. We have used liquid scintillation counting to study the tissue distribution and pharmacokinetics of S-35-labeled 5-TDG in hamster models of pancreatic tumors. In the normal hamster, initial uptake of S-35 activity into kidney, liver, and blood was high, but rapidly decreased with time. The pancreatic uptake (% dose/g) never exceeded 0.75%. This level occurred only at the earliest times after administration. Uptake in all three tumor models exceeded that in pancreatic tissue within 15 min of injection. The highest tumor-to-pancreas ratio was seen in the duct-tumor model, which also exhibited the most favorable tumor-to-tissue ratio when compared with kidney, liver and muscle. Favorable ratios were most pronounced at 6 and 24 hr after injection. These studies provide impetus for the use of 5-TDG as a model compound for the synthesis of potentially useful agents for clinical detection of pancreatic tumors.


A rapid and precise method for the determination of vitamin D3 in rat skin by high-performance liquid chromatography. In order to develop the investigations into photobiogenesis of vitamin D3, a rapid and precise method for the determination of the vitamin in rat skin was established by using high-performance liquid chromatography (HPLC). The proposed method included saponification of small pieces of rat skin, extraction of the unsaponifiable matter and application to HPLC using "Zorbax SIL" (straight-phase) as an adsorbent and 0.5% isopropanol in n-hexane as a mobile phase. The applicable lower limit of the method was 2ng of vitamin D3/cm2 of subcutaneous tissue-removed skin and it was possible to assay a concentration higher than 2 ng/cm2. The proposed method was applied to determine the content of vitamin D3 in rat skin obtained from in vivo and in vitro irradiation experiments. In the in vitro experiment, the yield of vitamin D3 increased in proportion to the irradiation time. On the other hand, the yield in the in vivo experiment showed a proportional increase similar to the in vitro experiment until 60 min irradiation, while a nearly constant value was obtained by irradiation for longer than 60 min. When the rat skin obtained from the in vitro experiment was irradiated with monochromatic UV rays in the range s60-350 nm, the most effective wavelength for the formation of vitamin D3 was confirmed to be 303 nm, which differs from the result obtained from the experiment in a test tube (295 nm). Moreover, the yield of vitamin D3 by irradiation with UV rays below 288 nm was extremely low, which again differed from the results of a test tube experiment. These differences were thought to be due to the filter effect of the malpighian layer in the epidermis of rat skin.


Dystrophic calcification in calcifying epithelial odontogenic tumor: an X-ray diffraction and electron spin resonance study. X-ray diffraction and electron spin resonance (ESR) analyses were conducted on calcifying epithelial odontogenic tumor (CEOT) associated with abundant calcification in order to compare its mineral structure with that of surrounding cortical bone. The material under investigation was removed from the canine-premolar region of the left maxilla of a 39-year-old woman. The principal findings were that the mineral phase of CEOT consists of apatite crystals and that the crystallinity of the crystals is higher than that of bone apatite. ESR spectra of CEOT are similar to those of bone sample. Quantitative ESR analysis indicates that CEOT shows a higher crystallinity coefficient -- defined by Ostrowski et al. (1972, 1974) as the ratio of spin concentration to the ash content of the sample -- than bone. Based on the data, it can be concluded that the calcification in CEOT has a higher crystalline structure than the cortical bone.


Tryptic peptide analysis of avian oncovirus gag and pol gene products. Radiolabeled tryptic peptides of the gag and pol gene products of avian oncoviruses were examined. This analysis included Rous-associated virus 2 structural proteins and the Pr76gag and P180gag-pol proteins in Rous-associated virus 2-infected chicken embryo cells. The methionine- and cysteine-containing tryptic peptides of virion internal structural proteins were present in both Pr76gag and P180gag-pol, suggesting that there was no loss of gag gene-coding sequences during the generation of P180gag-pol. No overlap of gag and pol gene structural information was detected. Analysis of intermediates in the processing of Pr76gag and translation inhibition mapping with pactamycin yielded the following order of structural proteins within the Rous-associated virus 2 Pr76gag precursor: NH2-p19-p12-p27-p15-COOH. The gag and pol sequences missing in the endogenous gsmp120 protein of uninfected gs+ chicken cells were identified by comparison with those of Rous-associated virus 2 P180gag-pol.


Characterization of the proteins of intracisternal type A and extracellular oncornavirus-like particles produced by MOPC-460 myeloma cells. The mouse plasmacytoma cell line, MOPC-460, produces both intracisternal and intracytoplasmic A-type particles when grown as a solid tumor. When these cells are grown either as an ascites tumor or in tissue culture, a third type of particle is produced extracellularly. This particle, the "myeloma-associated virus," is closely related to, and probably an alternate form of, the intracisternal A-type particle. The proteins present in these two types of particles were compared by tryptic peptide mapping. Both types of particles were found to contain essentially the same major proteins of 76,000 (p76), 68,000 to 70,000 (p68-70), and 45,000 (p45) daltons, in addition to varying amounts of smaller proteins. The relative proportions of all these proteins varied from preparation to preparation in an unpredictable way. The p45, p68, and p70 proteins all contained sequences found in p76, suggesting precursor-product relationships of p76 leads to p70 leads to p45 for solid tumor A-type particles and p76 leads to p68 leads to p45 for extracellular myeloma-associated virus. In addition, immune precipitation experiments have established that p76 contains at least some of the antigenic determinants characteristic of murine leukemia virus p30. This confirms earlier nucleic acid hybridization studies which indicated a moderate degree of relatedness between MOPC-460 A-type particles and several standard murine leukemia and sarcoma viruses. Taken together, our results provide evidence supporting the concept that MOPC-460 A-type particles may represent aberrant forms of C-type murine viruses.


Characterization of infectious oncornaviruses from MOPC-460 plasmacytomas: their relation to A-type particles. MOPC-460 mouse plasmacytoma cells produce intracellular A-type particles and extracellular oncornavirus-like particles ("myeloma-associated virus," abbreviated MAV). The genomes of these two particles are closely related. During attempts to establish infections with MOPC-460 extracellular particles, we isolated ecotropic and xenotropic infectious forms of murine leukemia virus. We have investigated the relation of these isolates to A-type particles and to MAV by nucleic acid hybridization. Using complementary DNA probes prepared from the two isolates, we found that these infectious murine leukemia viruses differ from A-type particles and from MAV. Moreover, we found that MAV is the predominant extracellular component: the ecotropic and xenotropic forms of murine leukemia virus were present at only low levels (less than 5%) in MAV preparations. Neither the SC-1 cells infected with ectropic murine leukemia virus nor the mink cells infected with xenotropic murine leukemia virus showed any A-type particles in their cytoplasm when examined by electron microscopy. Our inability to demonstrate infection by the A-type particle-related component, MAV, suggests that these may be defective.


Mutant of herpes simplex virus type 2 with temperature-sensitive lesions affecting virion thermostability and DNase activity: identification of the lethal mutation and physical mapping of the nuc-lesion. We had previously shown that a temperature-sensitive (ts) mutant of herpes simplex virus type 2 strain HG52, ts13, induced a heat-labile DNase activity in infected cells (B. Francke, H. Moss, M. C. Timbury, and J. Hay, J. Virol. 26:209-213, 1978). Earlier work indicated that the mutant also possessed temperature-sensitive infectivity (I. W. Halliburton and M. C. Timbury, J. Gen. Virol. 30:207-221, 1976). In this study temperature-stable revertants of ts13 have been isolated; examination of them revealed that ts13 is a double mutant, with genetically distinct temperature-sensitive lesions affecting nuclease activity and particle stability. The lethal mutation, in the cell system studied, is the latter. Revertants, which all maintain the nuclease lesion, grew well at a high temperature. Physical mapping of the nuclease lesion placed it between 0.12 and 0.21 (fractional length) on the virus genome, quite distant from the lethal mutation at 0.64 to 0.70.


Cellular proteins associated with simian virus 40 early gene products in newly infected cells. Immunoprecipitates of extracts of simian virus 40-infected permissive monkey kidney cells contained two proteins with molecular weights of 56,000 and 32,000 (52K and 32K) in addition to the known viral early gene products. Immunoprecipitates of cells infected with the 0.54-0.59 deletion mutants that lack the viral 17K gene product did not contain thhe 56K and 32K proteins. The additional proteins appeared in immunoprecipitates of deletion mutant extracts if unlabeled extracts of wild-type-infected cells were added before addition of antiserum. The proteins can also be identified in uninfected cells by co-precipitation with unlabeled viral proteins. Thus, it appears that the 56K and 32K proteins are cellular products that associate with the viral proteins, the 17K in particular, and are indirectly immunoprecipitated by anti-tumor serum.


Isolation of a soluble and template-dependent poliovirus RNA polymerase that copies virion RNA in vitro. A soluble RNA-dependent RNA polymerase was isolated from poliovirus-infected HeLa cells and was shown to copy poliovirus RNA in vitro. The enzyme was purified from a 200,000-X-g supernatant of a cytoplasmic extract of infected cells. The activity of the enzyme was measured throughout the purification by using a polyadenylic acid template and oligouridylic acid primer. The enzyme was partially purified by ammonium sulfate precipitation, glycerol gradient centrifugation, and phosphocellulose chromatography. The polymerase precipitated in a 35% saturated solution of ammonium sulfate, sedimented at about 7S on a glycerol gradient, and eluted from phosphocellulose with 0.15 M KC1. The polymerase was purified about 40-fold and was shown to be totally dependent on exogenous RNA for activity and relatively free of contaminating nuclease. The partially purified polymerase was able to use purified polio virion RNA as well as a template. Under the reaction conditions used, the polymerase required an oligouridylic acid primer and all four ribonucleside triphosphates for activity. The optimum ratio of oligouridylic acid molecules to poliovirus RNA molecules for priming activity was about 16:1. A nearest-neighbor analysis of the in vitro RNA product shows it to be heteropolymeric. Annealing the in vitro product with poliovirus RNA product shows it to be heteropolymeric. Annealing the in vitro product with poliovirus RNA rendered it resistant to RNase digestion, thus suggesting that the product RNA was complementary to the virion RNA template.


Terminal sequences of vesicular stomatitis virus RNA are both complementary and conserved. The nucleotide sequences at the 5' and 3' termini of RNA isolated from the New Jersey serotype of vesicular stomatitis virus [vsV(NJ)] and two of its defective interfering (DI) particles have been determined. The sequence differs from that previously demonstrated for the RNA from the Indiana serotype of VSV at only 1 of the first 17 positions from the 3' terminus and at only 2 of the first 17 positions from the 5' terminus. The 5'-terminal sequence of VSV(NJ) RNA is the complement of the 3'-terminal sequence, and duplexes which are 20 bases long and contain the 3' and 5' termini have been isolated from this RNA. The RNAs isolated from DI particles of VSV(NJ) have the same base sequences as do the RNAs from the parental virus. These results are in sharp contrast to those obtained with the Indiana serotype of VSV and its DI particles, in which the 3'-terminal sequences differ in 3 positions within the first 17. However, with both serotypes, the 3'-terminal sequence of the DI RNA is the complement of the 5'-terminal sequence of the RNA from the infectious virus. These findings suggest that the 3' and 5' RNA termini are highly conserved in both serotypes and that the 3' terminus of DI RNA is ultimately derived by copying the 5' end of the VSV genome, as recently proposed (D. Kolakofsky, M. Leppert, and L. Kort, in B. W. J. Mahy and R. D. Barry, ed., Negative-Strand Virus and the Host Cell, 1977; M. Leppert, L. Kort, and D. Kolakofsky, Cell 12:539-552, 1977; A. S. Huang, Bacteriol. Rev. 41:811-8218 1977).


Cell fusion for genetic analysis of two nonconditional Rous sarcoma virus replication mutants. Procedures for characterizing replication-defective viruses in nonpermissive mammalian cells were developed and applied to three nonvirogenic Rous sarcoma virus (RSV)-transformed mammalian cell lines--B4, a line of Bryan virus-transformed hamster cells, and two SRD-RSV transformed rat cell lines, LR3/1 and LR3/2. Cell fusion was used to study virus complementation. The three cell lines (i) fused with helper virus-infected chicken cells and the host range of the rescued virus examined, (ii) tested for complementation by fusion with chicken cells exhibiting various patterns of endogenous virus expression, (iii) fused with chicken cells infected with the temperature-sensitive replication mutant LA334 and assayed for complementation at permissive and nonpermissive temperatures, and (iv) tested for complementation of defective viruses in other RSV-transformed mammalian cell lines by fusing pairs of nonvirogenic cell lines and permissive chicken cells. Based upon these complementation studies, we concluded that B4 virus is defective only in the env gene, LR3/) virus is an absolute mutant in the gag and/or pol genes, and LR3/2 virus is a leaky env mutant. Clones of LR3/1 and LR3/2 virus-infected chicken cells were established, and the results obtained from the characterization of these viruses in permissive avian cells substantiates the conclusions reached in the fusion-rescue studies.


Conserved polynucleotide sequences among the genomes of papillomaviruses. The DNAs of different members of the Papillomavirus genus of papovaviruses were analyzed for nucleotide sequence homology. Under standard hybridization conditions (Tm - 28 degrees C), no homology was detectable among the genomes of human papillomavirus type 1 (HPV-1), bovine papillomavirus type 2 (BPV-2), or cottontail rabbit (Shope) papillomavirus (CRPV). However, under less stringent conditions (i.e., Tm - 43 degrees C), stable hybrids were formed between radiolabeled DNAs of CRPV, BPV-1, or BPV-2 and the HindIII-HpaI A, B, and C fragments of HPV-1. Under these same conditions, radiolabeled CRPV and HPV-1 DNAs formed stable hybrids with HincII B and C fragments of BPV-2 DNA. These results indicate that there are regions of homology with as much as 70% base match among all these papillomavirus genomes. Furthermore, unlabeled HPV-1 DNA competitively inhibited the specific hybridization of radiolabeled CRPV DNA to bpv-2 DNA fragments, indicating that the homologous DNA segments are common among these remotely related papillomavirus genomes. These conserved sequences are specific for the Papillomavirus genus of papovaviruses as evidenced by the lack of hybridization between HPV-1 DNA and either simian virus 40 or human papovavirus BK DNA under identical conditions. These results indicate a close evolutionary relationship among the papillomaviruses and further establish the papillomaviruses and polyoma viruses as distinct genera.


Oligoribonucleotide map and protein of CMII: detection of conserved and nonconserved genetic elements in avian acute leukemia viruses CMII, MC29, and MH2. RNA and protein of the defective avian acute leukemia virus CMII, which causes myelocytomas in chickens, and of CMII-associated helper virus (CMIIAV) were investigated. The RNA of CMII measured 6 kilobases (kb) and that of CMIIAV measured 8.5 kb. By comparing more than 20 mapped oligonucleotides of CMII RNA with mapped and nonmapped oligonucleotides of acute leukemia viruses MC29 and MH2 and with mapped oligonucleotides of CMIIAV and other nondefective avian tumor viruses, three segments were distinguished in the oligonucleotide map of CMII RNA: (i) a 5' group-specific segment of 1.5 kb which was conserved among CMII, MC29, and MH2 and also homologous with gag-related oligonucleotides of CMIIAV and other helper viruses (hence, group specific); (ii) an internal segment of 2 kb which was conserved specifically among CMII, MC29, and MH2 and whose presence in CMII lends new support to the view that this class of genetic elements is essential for oncogenicity, because it was absent from an otherwise isogenic, nontransforming helper, CMIIAV; and (iii) a 3' group-specific segment of 2.5 kb which shared 13 of 14 oligonucleotides with CMIIAV and included env oligonucleotides of other nondefective viruses of the avian tumor virus group (hence, group specific). This segment and analogous map segments of MC29 and MH2 were not conserved at the level of shared oligonucleotides. CMII-transformed cells contained a nonstructural, gag gene-related protein of 90,000 daltons, distinguished by its size from 110,000-daltom MC29 and 100,000-dalton MH2 counterparts. The gag relatedness and similarity to the 110,000-dalton MC29 counterpart indicated that the 90,000-dalton CMII protein is translated from the 5' and internal segments of CMII RNA. The existence of conserved 5' and internal RNA segments and conserved nonstructural protein products in CMII, MC29, and MH2 indicates that these viruses belong to a related group, termed here the MC29 group. Viruses of the MC29 group differ from one another mainly in their 3' RNA segments and in minor variations of their conserved RNA segments as well as by strain-specific size markers of their gag-related proteins. Because (i) the conserved 5' gag-related and internal RNA segments and their gag-related, nonvirion protein products correlate with the conserved oncogenic spectra of the MC29 group of viruses and because (ii) the internal RNA sequences and nonvirion proteins are not found in nondefective viruses, we propose that the conserved RNA and protein elements are necessary for oncogenicity and probably are the onc gene products of the MC29 group of viruses.


Comparative analysis of GS and BK virus genomes. Analysis of heteroduplexing between the genomes of GS virus, a BK-group virus, and the prototype BK virus revealed one region of nonhomology. Further analysis by cleavage of viral DNA with the restriction endonucleases EcoRI, HindIII, and HaeIII revealed that base changes in the GS virus genome spanned 0.6 to 0.7 map unit from the EcoRI site. Large T and small t antigens of GS virus appear to be similar in size to the BK virus antigens.


Apparent posttranscriptional block to anaerobic induction of endogenous leukemia virus. Uninfected Fischer rat cells were induced by anaerobic stress to transcribe high levels of endogenous type C leukemia virus RNA. Complete 35S virus RNA with attached polyadenylic acid sequences was found associated with polysomes, indicating functional mRNA. Since no mature virus was released under these conditions, the presence of a posttranscriptional block to complete virus synthesis is strongly indicated.


Structure and function of adenovirus type 12 defective virions. Purified human adenovirus type 12 preparations contain defective virions with a lighter density. These defective virions were isolated, and their biological functions and DNA were characterized. They can induce early and late antigens in infected cells and tumors in newborn hamsters with similar efficiency as complete virions. The majority of the DNA molecules from light virions contain deletions mapping near 16% from the left-hand end of the genome. Mechanisms for the generation of these molecules are discussed.


Impaired maturation of mouse mammary tumor virus precursor polypeptides in lymphoid leukemia cells, producing intracytoplasmic A particles and no extracellular B-type virions. Processing of polypeptides of the mouse mammary tumor virus, a type B retrovirus, was investigated in a transplanted thymic lymphoma cell line of the GR strain (GRSL). This cell line was maintained in vivo in ascites form and in vitro as a suspension culture. GRSL cells produce clusters of intracytoplasmic A particles and are virtually deficient in the production of mature extracellular B-type particles. As control, a mammary tumor cell line of the same mouse strain capable of complete virion synthesis was used. The kinetics of viral polypeptide synthesis were studied by pulse labeling with various isotopes (including (35)S and (32)P), followed by immunoprecipitation of cell lysates with monospecific antisera to the major mouse mammary tumor virus gag and env proteins, p27 and gp52, respectively. Both the primary gag and env precursor polypeptides were synthesized in the GRSL cells, but their conversion into viral proteins was impaired. The major gag precursor, Pr73(gag), was stable over a period of 8 h, and mature viral core polypeptides could not be detected. Also, the highly phosphorylated intermediates in the proteolytic processing of Pr73(gag) in virus-producing cells were absent in GRSL cells. By immunoprecipitation, Pr73(gag) was detected in a GRSL particle fraction with the density of intracytoplasmic A particles. The precursor for envelope proteins, Pr73(env), was turned over without the generation of mature viral envelope components gp52 and gp36. The in vivo-transplanted ascites GRSL cells, however, were shown to express gp52 on the cell surface together with a 73,000-dalton polypeptide, as indicated by cell surface iodination and immunoprecipitation.


Human cytomegalovirus-induced immediate early antigens: analysis in sodium dodecyl sulfate-polyacrylamide gel electrophoresis after immunoprecipitation. Immediate early antigen (IEA) induced in human lung fibroblasts by human cytomegalovirus was characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis after immunoprecipitation with IEA-positive human sera. Two polypeptides of 76,000 daltons (76K) and 82K were detectable within 90 min after infection. Polypeptides of similar molecular weight were also found in immunoprecipitates of human cytomegalovirus-infected cells nonpermissive for virus replication. IEA is located within the nucleus, although some of the 76K material appears to be located on the outer nuclear membrane. Raising salt concentrations in the extraction buffer increased antigen extraction. The contribution of these IEA polypeptides to IEA nuclear fluorescent staining is discussed.


Incomplete genomes of the parvovirus minute virus of mice: selective conservation of genome termini, including the origin for DNA replication. Deletion mutants of minute virus of mice arising during a single high-multiplicity passage and after serial undiluted passage have been isolated, and the incomplete viral genomes contained therein have been analyzed. The DNA isolated from incomplete virions derived from a single high-multiplicity passage was heterogeneous, ranging in size from 15 to 70% of the intact viral genome, with an average molecular length of approximately, 2,000 nucleotides. Two distinct types of molecules, designated as type I D-DNA and type II D-DNA, could be distinguished on the basis of their degree of secondary structure, and these were present in roughly equal amounts. Type I D-DNAs were predominantly single-stranded, recombinant molecules in which the self-complementary sequences derived from both genomic termini were conserved. The 5' terminus was modified relative to the analogous wild-type structure. Although virtually all of the wild-type genome sequence was seen in the total type I D-DNA population, sequences which map between coordinates 47.3 and 87.1 were clearly underrepresented. However, the extent and position of the deletions in individual molecules varied significantly. The shortest molecules in the population lacked between 90 and 95% of the internal wild-type genome sequence and consisted of sequences derived almost exclusively from within 5.0 map units (250 nucleotides) at both ends of the viral genome. Moreover, these miniature recombinant molecules were selectively amplified during serial undiluted passage and were therefore believed to contain all of the critical recognition sites necessary for the replication of minute virus of mice viral DNA. Type II D-DNAs were virus-specific, double-stranded hairpin molecules whose complementary strands were covalently continuous at variable sites distal to the 5' end of the viral minus strand. In sharp contrast to the type I genomes, these hairpin molecules consisted of sequences which mapped entirely at the 5' end of the viral genome between positions 85.0 and 100. Furthermore, type II molecules were gradually lost from the total D-DNA population during serial undiluted passage, suggesting that these molecules are not competent for DNA Replication but arise as the result of fatal replication errors. Deletion mutants of the type described here for minute virus of mice should be valuable generally as aids to future studies on parvovirus DNA replication, transcription, and cell-virus interactions.


Electron microscopic mapping of RNA transcribed from the late region of polyoma virus DNA. The polyoma virus (Py) RNA species transcribed from the L DNA strand of the "late" region of the Py genome in Py-infected mouse cells have been mapped by hybridization with specific fragments of Py DNA followed by electron microscopic visualization of the hybrids. Total cellular polyadenylated Py-specific RNA molecules having an S value in the range of 16S to 20S were purified by oligodeoxythymidylic acidcellulose column chromatography, preparative hybridization with Py DNA, and sucrose gradient centrifugation. Cytoplasmic Py-specific RNA was similarily purified, except that it was not fractionated by sucrose gradient centrifugation. Hybrids of these RNA molecules and Py DNA fragments were spread for electron microscopy by either the cytochrome c technique or the bacteriophage T4 gene 32 protein method. The polyadenylic acid at the 3'-end of the RNA in the hybrids was identified by labeling with simian virus 40 DNA circles to which polybromodeoxyuridylic acid tails had been covalently attached. These experiments revealed the presence of three L DNA strand transcripts in both RNA preparations. Two of these RNA molecules were found to be spliced from chains transcribed from two noncontiguous parts of the late region. The third molecule either is a continuous transcript of the entire late region or contains a splicing feature which is too small to be reliably observed by the electron microscope methods used. The 5'-ends of the three RNA species map within a region extending from 68 to 70 map units on the Py restriction endonuclease map. Each of the two spliced molecules contains a 5'-terminal leader sequence transcribed from a DNA segment with an estimated length of 60 to 110 nuvleotides. The 3'-ends of the leaders map at 66.7 +/- 1.0 and 66.4 +/- 0.50 map units. In these molecules the 5'-ends of the other part (the main body) map at 59.4 +/- 0.90 and 49.4 +/- 2.0 map units, respectively. The 3'-termini of all three RNA species map at 24 to 25 map units.


Physical map of biologically active Harvey sarcoma virus unintegrated linear DNA. BALB/c JLS V9 cells recently infected with Harvey sarcoma virus-murine leukemia virus (HSV-MuLV) complex contained unintegrated HSV linear DNA of 6.0-kilobase pair mass. The cells also contained two HSV closed circular DNA species along with MuLV-encoded linear and closed circular DNA species. HSV 6.0-kilobase pair linear DNA induced focal transformation upon transfection of NIH 3T3 mouse fibroblasts, and the biological activity of HSV DNA did not require helper MuLV functions. A physical map of restriction endonuclease cleavage sites along HSV 6.0-kilobase pair linear DNA was derived. Comparison of this map with one for Moloney MuLV DNA showed that the HSV and Moloney MuLV genomes are identical near their viral RNA 3' ends.


Assembly of vesicular stomatitis virus nucleocapsids in vivo: a kinetic analysis. Pulse-chase labeling and cell fractionation were used to examine the pathways taken by the three nucleocapsid polypeptide species of vesicular stomatitis virus into nucleocapsids and then into virions. An improved method of polyacrylamide gel electrophoresis resolved nucleocapsid polypeptides N and NS from cellular actin, facilitating accurate quantitation of the viral polypeptides. Contrary to previous belief, the rate of NS synthesis was found to be a constant fraction of total virus protein synthesis throughout infection, indicating a consistent mechanism of virus protein synthesis regulation. In the kinetic studies, each polypeptide species displayed the following characteristic behavior. (i) Structural polypeptide N was the only species that entered a metabolically active soluble pool before assembly into nucleocapsids. The size of this pool increased with time after infection, causing an increasing delay in the appearance of pulse-labeled N molecules in nucleocapsids. (ii) Throughout infection, the entire complement of L molecules entered nucleocapsids immediately after their synthesis, without diversion through a soluble pool. (iii) Although 75% of newly synthesized molecules of the transcriptase-associated protein NS entered a soluble pool, they never emerged from the compartment. At all times after infection, about 25% of the NS molecules bypassed the soluble pool and entered nucleocapsids directly after their synthesis, as if in concert with L. These results indicate that VSV nucleocapsid assembly in vivo is a stepwise process, comprising an initial condensation of N with the viral RNA, followed by attachment of L and NS, analogous to the stepwise assembly of Sendai virus nucleocapsids. (D. W. Kingsbury, C.-H. Hsu, and K. G. Murti. Virology 91:86-94, 1978). About half of the intracellular nucleocapsids were recovered in a form that sedimented at anomalously low centrifugal forces, reflecting an association with large cellular organelles. This attachment was mediated mainly by electrostatic forces, since these "bound" nucleocapsids were released by elevated salt concentrations. The kinetic behavior of nucleocapsid polypeptides was the same in both fractions, providing no evidence for a division of nucleocapsid functions between cellular compartments.


Further morphological characterization and structural proteins of infectious bursal disease virus. An outer layer surrounding the capsid of infectious bursal disease virus was evident from electron micrographs of intact virus particles having diameters of 62 to 63 nm. The capsid was found to be composed of large morphological units or capsomeres, measuring about 12 nm in diameter. The architecture of the capsid appears to be that of T = 3 symmetry, with a probable 32 morphological units by rotational enhancement of image detail. Structural proteins of infectious bursal disease virus consist of seven species, two major and five minor polypeptides. These are P1 to P7, with molecular weights of 133 x 10(3), 124 x 10(3), 98 x 10(3), 51 x 10(3), 33 x 10(3), 26 x 10(3), and 23 x 10(3), respectively.


Secondary structures in polyoma DNA.

Secondary structures in polyoma DNA. Three reproducible secondary-structure features were observed on single strands of polyoma virus DNA mounted for electron microscopy by the T4 gene 32 protein technique: (i) a hairpin fold-back extending from 92.9 +/- 0.8 to 95.0 +/- 0.7 map units; (ii) a small loop extending from 63.2 +/- 3.1 to 68.5 +/- 2.8 map units; and (iii) a big loop extending from 51.9 +/- 2.3 to 68.9 +/- 2.1 map units. Both loops are bounded by inverted repeat stems of length 40 +/- 20 base pairs. The stem sequences around 68.5 and 68.9 of the large and small loops overlap, either partially or completely. Several lines of evidence indicate that the inverted repeat stems of the two secondary-structure loops lie in the regions of polyoma virus DNA flanking and probably very close to the sequences that are spliced out in the formation of the late 16S and 18S messages, whereas the hairpin fold-back appears to map at a splicing point of an early message. These structures may therefore be important for the processing of the primary transcripts to form the early and late messages.


Polymorphism of B-tropic leukemia viruses from BALB/c mice: association of a p30 antigen with N- versus B-tropism. Comparison of a number of murine leukemia virus clones by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed extensive protein polymorphism among B-tropic, but not N-tropic, isolates from BALB/c mice, particularly in migration of p30 proteins. A type-specific radioimmunoassay for p30 was developed which uniformly discriminated all B-tropic viruses from N-tropic viruses of BALB/c origin. N- and B-tropic viruses of C57BL/6 and AKR Fv-1b/b origin could also be distinguished by this assay.


Immune response to the src gene product in mice bearing tumors induced by injection of avian sarcoma virus-transformed mouse cells. A single subcutaneous injection of 10(7) live cells of the highly tumorigenic avian sarcoma virus (Schmidt-Ruppin strain, subgroup D)-transformed BALB/c line into BALB/c mice resulted in the production of an antiserum specific for the avian sarcoma virus gene product pp60src. All sera taken from mice 3 weeks after injection of tumor cells contained antibodies to pp60src. Immunoprecipitation experiments showed that all sera precipitated pp60src from Schmidt-Ruppin-infected chicken cells, but only a portion of these sera precipitated pp60src from chicken cells infected with other strains of avian sarcoma virus, i.e., Prague and Bratislava-77. Analysis of the cross-reactivity patterns of these antisera demonstrated a minimum of three to four antigenic determinants on pp60src. The findings reported here should facilitate the production of monoclonal antibodies to pp60src, which in turn will provide highly specific probes for further investigations into the structure and function of this protein.


Expression of simian virus 40 gene A affects tubulin stability. The stability of tubulins present in crude extracts of untransformed BALB/c-3T3  mouse fibroblasts, Chinese hamster lung cells, and various of their simian virus 40 transformants was assessed by measurement of their individual colchicine-binding decay rates. In all cases studied the decays followed the kinetics of first-order reactions, and rates were reduced at low temperatures and by vinblastine sulfate. Under all assay conditions, including different temperatures and protein concentrations, tubulins of normal cells decayed considerably faster than those of simian virus 40-transformed cells. Experiments performed with a number of Chinese hamster lung cell clones transformed with temperature-sensitive simian virus 40 gene A mutants showed a clear correlation between increased tubulin stability and the expression of gene A function. These results suggest that it is T-antigen, the viral gene A product, that affects tubulin.


Methylation of nuclear simian virus 40 RNAs. Nuclear RNAs, pulse-labeled with [methyl-3H]methionine and [3H]uridine under optimal conditions, were prepared from cells infected with simian virus 40 at a late time after infection. The labeled RNAs were hybridized with restriction fragments of simian virus 40 DNA. Levels of methyl-3H-labeled RNA annealing with the early (E) or late (L) strands of a particular restriction fragment were compared with [3H]uridine-labeled RNA annealing with the same fragment. The methyl-3H-labeled RNA hybridizing to the various restriction fragments was eluted and analyzed for caps and internal 6-methyladenosine residues. Caps of transcripts of both the L and E strands were primarily located in a fragment containing the origin for viral DNA replication. The highest level of the internal 6-methyladenosine residues in simian virus 40 nuclear RNA was located on the L strand within a fragment from 83 to 0.0 map units and in RNA transcripts from the E strand within a fragment from 37 to 67 map units. Portions of both these regions represent intervening sequences not represented in cyotplasmic mRNA's. Lower levels of internal methylation were found in other locations as well. The potential role of internal 6-methyladenosine residues in RNA processing and transport from the nucleus to the cytoplasm is discussed.


Adenovirus early function required for protection of viral and cellular DNA. Studies were done to characterize a DNA-negative temperature-sensitive (ts) mutant of human adenovirus type 2, H2 ts111. The temperature-sensitive defect, which was reversible on shift-down in the absence of protein synthesis, was expressed as early as 2 h postinfection, and the results of density-labeling experiments are in agreement with at least a DNA replication initiation block. On shift-up, after allowing viral DNA synthesis at permissive temperatures, the newly synthesized viral DNA and the mature viral DNA were cleaved into fragments which sedimented as a broad peak with a mean coefficient of 10-12S. This cleavage was more marked in the presence of hydroxyurea as the DNA synthesis inhibitor. Parental DNA in infected cells was degraded to a much lesser extent regardless of the incubation temperature. In contrast, the parental DNA was strongly degraded when early gene expression was permitted at 33 degrees C before shift-up to 39.5 degrees C. Furthermore, cellular DNA was also degraded at 39.5 degrees C in ts111-infected cells, the rate of cleavage being related to the multiplicity of infection. This cleavage effect, which did not seem to be related to penton base-associated endonuclease activity, was also enhanced when early gene expression was allowed at 33 degrees C before shift-up. The ts111 defect, which was related to an initiation block and endonucleolytic cleavage of viral and cellular DNA, seemed to correspond to a single mutation. The implication of the ts111 gene product in protection of viral and cellular DNA by way of a DNase-inhibitory function is discussed.


Inhibition of herpes simplex virus-induced DNA polymerase activity and viral DNA replication by 9-(2-hydroxyethoxymethyl)guanine and its triphosphate. The effect of the nucleoside analog 9-(2-hydroxyethoxymethyl)guanine (acycloguanosine) on herpes simplex virus type 1 DNA synthesis was examined. Acycloguanosine inhibited herpesvirus DNA synthesis in virus-infected cells. The synthesis of host cell DNA was only partially inhibited in actively growing cells at acycloguanosine concentrations several hundred-fold greater than the 50% effective dose for herpes simplex virus type 1. Studies using partially purified enzymes revealed that the triphosphate of this compound inhibited the virus-induced DNA polymerases (DNA nucleotidyltransferases) to a greater degree than the DNA polymerase of the host cell, that the inhibition was dependent upon the base composition of the template, and that the triphosphate was a better substrate for the virus-induced polymerases than for the alpha cellular DNA polymerases.


Identification of transcribed regions of Epstein-Barr virus DNA in Burkitt lymphoma-derived cells. RNA was extracted from the Burkitt lymphoma-derived cell line Raji and from Burkitt lymphoma tumor biopsies, isotope labeled in vitro by iodination with 125I, and hybridized to electrophoretically separated restriction endonuclease fragments of Epstein-Barr virus DNA on nitrocellulose membranes. The results indicated that only certain parts of the Epstein-Barr virus genome are represented as polyribosomal RNA in Raji cells, with a pronounced dominance of RNA sequences complementary to a 2.0 x 10(6)-dalton segment of Epstein-Barr virus DNA located close to the left end of the viral genome. A map of virus-specific polyribosomal RNA sequences was constructed, which indicated that a minimum of three regions of the Epstein-Barr virus genome are expressed in Raji cells. Total-cell RNA preparations from five Burkitt lymphoma biopsies contained RNA sequences homologous to the same regions of Epstein-Barr virus DNA as polyribosomal RNA from Raji cells, albeit at different relative proportions.


Adenovirus DNA template for late transcription is not a replicative intermediate. The relationship between adenovirus replication and late transcription has been investigated using viral replication and transcription complexes isolated from infected HeLa cell nuclei. These two types of complexes extracted from adenovirus type 2-infected cell nuclei did not sediment at the same rate on sucrose gradients. Viral replicative intermediates were quantitatively precipitated by immunoglobulins raised against purified 72,000-dalton DNA-binding protein, whereas viral transcription complexes remained in the supernatant. These results show that late transcription does not occur on active replication complexes or on 72,000-dalton DNA-binding protein-containing replicative intermediates inactive in DNA synthesis. Additional evidence is presented indicating that it is very unlikely that replicative intermediates lacking the 72,000-dalton DNA-binding protein could be the template for late transcription.


T lymphocyte-dependent difference in susceptibility between DDD and C3H mice to mouse hepatitis virus, MHV-3. After intraperitoneal inculation with a virulent mouse hepatitis virus, MHV-3, 50% lethal dose in DDD mice was about 7 log10 higher than that in C3H mice. Histopathologically, splenic lymphocytes especially of the thymus-dependent area were more severely affected in susceptible C3H mice than in DDD mice. In the liver of C3H mice, virus multiplied exponentially after inoculation, attaining 10(6) PFU at moribund stage, while virus multiplication in DDD mice was much less prominent decreasing remarkably at day 5 or later. The virus could multiply in the primary culture of spleen cells from C3H mice but not in those from DDD mice, and cells supporting virus growth seemed to be a theta-positive population of lymphocytes. No difference was observed between the two strains of mice in the ability of peritoneal macrophages to support virus growth in vitro as well as serum interferon levels after infection.


Correlation of cyclic AMP and cyclic GMP levels with changes in contractile force of dog ventricular myocardium during cholinergic antagonism of positive inotropic actions of histamine, glucagon, theophylline and papaverine. On the dog isolated right ventricular muscle, experiments were carried out in order to elucidate the characteristics of the cholinergic antagonism against the positive inotropic action (PIA) induced via different subcellular mechanisms. The relationship between cyclic AMP and cyclic GMP levels, and contractile force during cholinergic antagonism was assessed. Carbachol (10 microM) by itself decreased only slightly the tension developed, but inhibited prominently the PIAs of isoprenaline, histamine, glucagon, theophylline and papaverine. The action of dibutyryl cyclic AMP was inhibited less than PIAs of other agents mentioned above. In contrast, carbachol did not affect the PIAs of calcium and g-strophanthin. The antagonism by carbachol of PIAs of isoprenaline, histamine, glucagon, theophylline and papaverine was accompanied by a reduction of the intracellular cyclic AMP level elevated previously by these agents, and by an elevation of the intracellular cyclic GMP level. A good correlation was found between changes in the tension developed, and cyclic AMP and cyclic GMP levels during the cholinergic antagonism of PIAs induced by these agents in the dog ventricular myocardium.


Effects of phenylephrine and isoproterenol on the activity of cyclic AMP-dependent protein kinase of hypothyroid rat tissues. The present study was designed to examine the properties of cyclic AMP-dependent protein kinase from normal and hypothyroid rat tissues. The activity of the liver enzyme decreased in hypothyroid rats and this reduced activity may be due to the decrease in synthesis of enzyme protein. The enzyme activity from the kidney of hypothyroid rats increased by 35% in the absence of cyclic AMP. Further, the enzyme from the kidney of hypothyroid animals responded to phenylephrine and isoproterenol. In the heart, the basal enzyme activity was the same in both groups, but the enzyme from the heart of hypothyroid animals did not respond to isoproterenol. When hypothyroid rats were treated with 3,3',5-triiodothyronine, the enzyme activities from those rat tissues showed essentially the same level as those of the corresponding normal ones. These results suggest that thyroid hormone regulates adrenergic agonists-mediated responses both at the sites of adrenoceptors and that of protein phosphorylation. Furthermore, the dependence on thyroid hormone varies according to the tissue.


[Influence of hyperthermia on toxic side effects of cytostatic agents (author's transl)]. 14 patients with tumors in generalised stages (Hodgkin's disease, oat-cell-carcinoma, Fibrosarcoma, acute leukemia) were treated altogether 54 times with cytostatics in combination with hyperthermia. The temperature was induced by microwaves with a frequency of 27 MHz. A temperature of 40 degrees C was reached after 40 min. At this point cytostatics were applicated; afterwards the temperature was maintained for one hour at 40-40,5 degrees C. Cardia, pulmonary and circulatory complications did not occur. Controls of the laboratory parameters could exclude effects on electrolytes, muscles, blood, liver and kidney. The laboratory controls were made before, during, immediately after and 24 h after hyperthermia. There were no signs for an enhancement of toxicity typical for cytostatics. The observations are compared to the results of other investigators. To date the therapeutic effect of this treatment can not be stated.


Plants used as abortifacients and emmenagogues by Spanish New Mexicans. Individuals of Spanish and Mexican descent in New Mexico have used a number of plants as emmenagogues and abortifacients. Of the plants used, cotton root bark (Gossypium sp.), inmortal ((Asclepias capricornu Woodson), poleo chino (Hedeoma oblongifolia (Gray) Heller), rue Ruta graveolens L.), wormseed (Chenopodium ambrosioides L.), and three species of Artemesia seem to be used most widely. Of these, the cotton root bark, when used as an abortifacient, seems to exhibit the lowest toxicity. Rue is notable because of its use independently within different cultures, but may exhibit toxic side effects when used as an abortifacient. Seven other plants are outlined on the basis of anecdotal and folkloric reports. Investigations are underway to look at use effectiveness, side effects, impact on fertility, and acceptance among cultures of the Southwestern United States.


Transposon 10 promoted deletions and inversions in the transfer genes of R100-1. Spontaneous tetracycline-sensitive, transfer-deficient mutants of R100-1 were selected and analysed by genetic complementation tests and with the restriction endonuclease EcoR1. While some of the Tets Tra- mutants were caused by a single deletion event which removed the Tetr genes and extended into the neighbouring transfer genes, other mutants were the result of the deletion of the Tetr genes within Tn10 which was accompanied by an inversion of adjacent DNA sequences. A clustering of deletion and inversion endpoints occurred in the traA gene. Some of the transfer genes of R100-1 were assigned to EcoR1 fragments.


A kinetic analysis of cell division, and induction and stability of recA protein in U.V. Irradiated ion+ and ion-strains of Escherichia coli K12. Kinetic analysis of induction of recA protein synthesis after U.V. irradiation does not show correspondence with the kinetics of division inhibition in ion+ and ion- strains. When the induction of recA protein after U.V. is drastically reduced by rifampicin treatment, no effect on the kinetics of division inhibition is observed.


Chromosomal mobilization from a recA mutant of Pseudomonas putida. A methyl methane sulfonate sensitive mutant of P. putida strain PpG1 is also extremely sensitive to UV-rays, compared to parent wild type cells. This mutant behaves typically as recombination less (recA) mutants of Escherichia coli and Pseudomonas aeruginosa, since as a recipient, it exhibits extremely low frequency of recombination following conjugational, transductional, and transformational gene transfer. Sex factor plasmids such as K-XYL or TOL can mobilize chromosomal genes equally well both from recA+ and recA801 donor cells, suggesting that host recombination functions are not necessary for mobilization of chromosomal genes by such plasmids.


Altered linkage values in phage P22--mediated transduction caused by distant deletions or insertions in donor chromosomes. The effects of distant deletions or insertions in the Salmonella typhimurium donor strains on P22--mediated cotransducibility of genetic markers was studied. We found that deletions of histidine operon, unit 44 of the chromosome map, changed the linkage of markers purF and aroC (unit 49) and pyrF and trpA (unit 34). They did not change the linkage of more distant markers pyrE and cysE. The effect of three types of insertions was examined. The donor strains carried F factor, Tn10 transposon or pi-his duplication inserted close to histidine operon. These insertions caused alteration of purF-aroC linkage while pyrF-trpA cotransduction values were not affected. These data show that the effect of the chromosome rearrangements extends to at least 5% of S. typhimurium chromosome length and may reach as much as 10% of it. Our results are in agreement with the model of Chelala and Margolin (1974) concerning formation of transduction particles. They indicate that the cotransducibility changes caused by deletions or insertions extent further than it might have been expected from previous reports.


Sensitivity of chemically treated spores of Clostridium perfringens type A to an initiation protein. Extraction of Clostridium perfringens type A spores with dithiothreitol (DTT), DTT plus sodium dodecyl sulphate (DTT-SDS), urea-mercaptoethanol (UME), or alkali, solubilized from 18.6 to 46.5 of the total dry weight of spores. The initiation of germination and lysis of such treated spores with lysozyme and an initiation protein (IP) from the culture supernatant fluid of sporulating cells of C. perfringens was studied under various conditions. The ability of lysozyme and the crude IP to induce germination and lysis of extracted spores was concentration dependent up to 0.5 microgram/ml and 5.6 mg/ml respectively. IP showed an optimum of activity between pH 7 and 8 for DTT-SDS and DTT extracted spores, and between pH 6 and 9 for UME extracted spores. The optimum temperature of activity for IP was 55 degrees C. Dissimilarities in the extent to which lysozyme and the IP initiated germination and lysis of spores extracted by various methods may have been a reflection of the differences in amounts of protein solubilized by each treatment.


Measurement of pyrimidine dimers in spheroplasts of Bacillus subtilis. A method is described for making spheroplasts of Bacillus subtilis which are permeable to exogenous enzymes. Conditions are described for measuring small numbers of pyrimidine dimers in the DNA of UV-irradiated cells by use of a partially purified Micrococcus luteus extract containing an enzyme specific for pyrimidine dimers. The system will detect as few as 10-12 pyrimidine dimers per genome.


The physical state of herpes simplex virus DNA in infected human cells. Treatment of herpes simplex virus type 1 (HSV-1)-infected human embryo lung (HEL) cells with phosphonoacetic acid (PAA) resulted in complete inhibition of HSV DNA replication. DNA was extracted from PAA-treated HEL cells infected with HSV-1 and centrifuged in a neutral CsCl density gradient. The HSV DNA sequences in the nuclei of PAA treated cells at 24 hr post infection banded at the same density as free HSV DNA (1.725 g/cm3), but a significant amount of viral DNA sequences were detected in the regions of cell DNA (1.700 g/cm3) as well as in the intermediate fractions as determined by hybridization with 3H HSV complementary RNA. The viral DNA sequences of lower deisntiy did not change in density by recentrifugation in a CsCl density gradient, but did change to the density of free viral DNA after treatment with EcoR1 restriction endonuclease. When the DNA from the nuclei of PAA treated cells was analyzed in an alkaline glycerol gradient, more than 95% of the viral DNA sequences were found in the free viral DNA fractions. Since the viral and cellular hybrid DNA represented approximately 33% of the total viral DNA sequences, it is concluded that some of the HSV DNA sequences in PAA treated, infected cells are associated with cell DNA by alkali-labile bonds.


Heterogeneous RNA of the Prague strain of Rous sarcoma virus caused by undiluted passages. The size of genomic RNA in PR-RSV A passaged in chick embryo fibroblasts (CEF) or quail embryo fibroblasts (QEF) was determined by gel electrophoresis. The results showed that 3 undiluted passages resulted in heterogeneity of RNA. The heterogeneity of the smaller incomplete RNAs in the virus stock was decreased by diluted passage or cloning, but RNA of the b subunit size and a subunit RNA of complete genome size were relatively stable. These heterogeneous RNAs were characterized by hybridization analysis. The RNAs from 4 peaks hybridized with both cDNAtotal and cDNAsrc to appreciable extents, indicating that they were derived from viral RNA and that at least some of them contained the src sequence. This finding and the failure to isolate a td mutant from the undiluted-passaged virus stock or from some subclones that had a and b subunits of RNA indicate that the td virus was only a minor constituent of the incomplete virus population caused by undiluted passages. Some viruses with incomplete RNA in the virus stock could produce foci with the aid of td B77 or RAV-60. The emergence of rd viruses by undiluted passages was indicated.


Combination radiotherapy and chemotherapy for small cell carcinoma of the lung. Twenty previously untreated patients with small cell carcinoma of the lung were treated with cyclophosphamide, 400 mg/m2 and Adriamycin, 40 mg/m2 IV on day 1, followed by cytosine arabinoside, 20 mg/m2, every 12 hours subcutaneously on days 5--9; this regimen was repeated every 28 days. On days 14--28 of the first cycle, each patient received 3,000 rads to the primary tumor and whole brain. Following eleven courses, Adriamycin was discontinued and patients received cyclophosphamide, 800 mg/m2 IV on day 1 and methotrexate, 15 mg/m2 IV on days 5--7. This regimen was repeated every 28 days. Toxicity included nausea, vomiting, alopecia, leukopenia, thrombocytopenia, and esophagitis. Overall response rate was 65%. Media survival in limited disease was 14.5 months, and in extended disease it was 4.5 months. This combination is active in localized small cell carcinoma but provides no superiority over other regimens.


Inflammatory fibrous histiocytoma: response to non-surgical therapy: a case report. A case of inflammatory fibrous histiocytoma arising in soft tissue near the sacrum is presented. The patient's mode of presentation, clinical course, and tumor histology were typical of this disease. The tumor was inoperable, and radiotherapy combined with doxorubicin, cyclophosphamide, and intermediate-dose oral methotrexate produced a dramatic complete response lasting over 27 months. Favorable results with nonsurgical therapy have not previously been reported for this disease. The gratifying result obtained, although of relatively short duration to date, indicates that combined modality therapy may provide significant back-up to aggressive surgery. The use of these agents in an adjuvant setting merits study.


[Worldwide eradication of smallpox or from Sarah Nelmes to Ali Maow Maalin]. It cannot be disputed that the WHO plan for the eradication of smallpox has had considerable success. There is no endemic smallpox any longer in the world. Nevertheless caution is advised and continued supervision must not be neglected. There are still countries where supervision is not satisfactory, there is uncertainty about animal reservoirs and it should be assumed from the beginning that up to the present no epidemic disease could be completely eradicated. For this reason it is inevitable that supplies of vaccine should be available. A vaccine from heat sensitive strains is offered which is obtained from cell cultures.


Heterogeneity of neoplastic cells derived from a hamster tumor induced by avian sarcoma virus B77. Cells derived from a hamster tumor induced by avian sarcoma virus Bratislava 77 (B77) in vivo, were cultivated in vitro. After few passages two morphologically different cell lines were isolated from the parental culture (B77/H). One cell line consisted of fibroblastic cells (B77/H/fi and the other from epithelioid cells (B77/H/ep). Cells of both lines were highly tumorigenic in neonatal syngeneic hamsters, B77/H/ep cells were able to form colonies in soft agar and contained complete integrated B77 viral genome. In contrast, the B77/H/fi cells grew poorly in soft agar and did not contained B77 virus genome.


Calf serum- and cell surface proteins released from cultured avian sarcoma virus-transformed and untransformed rat fibroblasts. Cell surface and calf serum proteins were released in vitro from cultured virus-transformed, chemically transformed and normal rat embryo fibroblasts in 0.2 M urea containing serum-free culture medium. Fibronectin (LETS protein) was found in considerably higher amount in medium from normal rat embryo fibroblasts than in that from transformed cells. Major calf serum protein released from cultured normal and transformed rat cells corresponded by its electrophoretic mobility to calf serum albumin. No substantial differences in electrophoretic patterns of calf serum proteins released from normal and transformed cells into the medium were found. Time course of release of serum proteins from cell surface was measured quantitatively with the aid of radioiodinated Staphylococcus aureus protein A radioimmune assay. More than 50% of cell bound calf serum proteins remained on cell surface after 24 hours of incubation in serum-free medium.


Study of the avian sarcoma virus infection of chemically transformed cells. BHK-21/13 cells transformed with various chemical carcinogens and mutagens were tested for susceptibility to avian sarcoma virus infection. The chemically transformed BHK cells were highly tumorigenic. The treatment of these cells in vitro with avian sarcoma virus strain Schmidt-Ruppin D resulted in chemically transformed and viral infected cells with different rescuability of the integrated virus genome. The different rescuability is not due to the difference in virus penetration as was shown by vesicular stomatitis virus Schmidt-Ruppin virus pseudotype VSV(SR-D) technique. The sarcoma virus genome in the doubly transformed cells was not inducible by various virus inducers.


Sero-epidemiologic study of the Epstein--Barr virus infectivity in a healthy Cuban population. The indirect immunofluorescence test was used to study the level of humoral antibodies against viral capsid antigen of the Epstein--Barr virus (VCA-EBV), and early antigen (EA) in the sera of 322 healthy donors of the Havana City province. The age of donors ranged from several months up to 95 years. The results obtained indicate that the healthy population of Cuba is infected by EBV as well as in other countries, even though the spread and the course of infection have its specific characteristics. In the early childhood population group (3--4 years old) the infection is relatively more frequent (73%) than in adults (62--77%), despite the fact that the country is located in the tropical zone. In all the studied age groups the geometric mean titers (GMT) of antibodies were found to be lower in comparison with the other populations studied. The increase of EBV-antibody levels in the group of donors above 55 years of age, together with a large number of seropositive individuals, increase in the number of persons with high anti-VCA-EBV antibody titers (1 : 160), agrees with the results obtained by other investigators. This may be associated with some form of depression of cell-mediated immune response. Infectious mononucleosis-morbidity in the studied age groups was found to be low in 3--4-year- and 15--19-year-old age groups, where higher levels of antibodies against EA of EBV were found.


Endogenous opiates: through 1978.

Endogenous opiates: through 1978. The discovery of receptors in the brain for opiates and the structure of the endogenous peptides for these receptors has led to an explosion of interest in this field. The present review is the first of an annual series. It summarizes many of the highlights of research with opiate peptides published with a date of 1978 or earlier.


[Peripheral nerve injuries following implantation of total endoprosthesis in the hip joint and their treatment]. The complication rate after 844 implantations of total endoprosthesis in the hip-joint, in the form of postoperative nerve injuries was found to be about 1 per cent. A number of causes may be responsible for nerve lesions, and some of these may be eliminated with a sparing operative technique. Although there are good prospects for the disappearance of paralytic symptoms, meaningful pysiotherapeutic after-care is very important. In the paralytic stage consistent exponential electrotherapy is recommended, and for recurring voluntary mobility active exercises against resistance. This will greatly improve the functional result of an operation with regard to nerve lesions.


[A study through factor analysis of the connection between 7 variables concerning blood lipides, with a particular reference to the interpretation of the lipidograms (author's transl)]. The AA. examined 7 variables concerning blood lipides and studied their internal relationship by means of a method, the Factor Analysis, which aims at determining the least amount of Common Factors which can sum up the information contained in the same variables. In this case three Common Factors (F) could be obtained, two of which were connected to the interpretation of the data of the lipidogram. The AA. discuss the signification of the determined Factors; they have even calculated the scores of the same Factors for each single individual and derived from the frequence distributions of the scores of F1 and F2 the normal values of the same Factors.