Jerome T. Syverton

THE PATHOGENESIS OF THE RABBIT PAPILLOMA‐TO‐CARCINOMA SEQUENCE

A naturally-occurring cutaneous papilloma is commonly present on wild hares captured in states bordering the Mississippi River.25* 40, 41 This epithelial tumor became interesting to many investigators following the discovery in 1933 by Shope41 of its virus etiology and the demonstration by Rous and 34 and others’j, 24, 48 that the benign virus-induced growth may terminate in an epidermoid carcinoma. A wide variety of investigations of this tumor and its etiological agent resulted. Our interest in this disease and in the causative virus has been continuous since June, 1934.46-54 Since then, we have kept in our rabbit colony more than 1700 wild cottontail and domestic rabbits with papillomatosis. Our cumulative data and comparative studies51* j2, b4 reflect a wide experience that has confirmed and extended work previously reported. It is the purpose of the present paper to summarize these findings! and the reported experiences of other investigators in an attempt to understand two phenomena of fundamental importance: (a) the role of Shope’s papilloma virus in the papilloma-tocarcinoma sequence and (b) the biological phenomenon that results during the virus papilloma-to-carcinoma sequence in the disappearance, or “masking,” of the causative virus in papillomas of domestic rabbits and in cancers of domestic rabbits and wild hares.

Isolation and Elimination of Pleuropneumonia-Like Organisms From Mammalian Cell Cultures.

Summary One hundred sixty-six sublines representative of 68 culture lines of cells of human, rabbit, monkey, mouse, pig, cow and other species origin were tested repeatedly for contamination with pleuropneumonia-like organisms over a period of 18 months. Cells freshly dispersed with trypsin were inoculated into a medium containing pancreatic digest of beef heart enriched with yeast extract and human ascitic fluid. Of the tested sublines, 57% were contaminated with PPLO. Repeated tests with the heart digest medium on 89 sublines were consistently positive, while 5 sublines yielded variable results. PPLO strains isolated from 2 of the variable cell sublines grew sparsely and slowly, but growth of other PPLO strains was luxuriant in the test medium. Fifteen cell lines carrying PPLO of varying colonial morphology were treated for 3 weeks with 100 μg of kanamycin per ml of medium. Treated sublines yielded no PPLO during subsequent cultivation for nearly a year in the absence of kanamycin. A contaminated strain of human esophageal epithelium has been freed of PPLO by brief exposure to kanamycin. Required time of exposure appeared inversely related to kanamycin concentration employed.

Cortisone and Roentgen Radiation in Combination as Synergistic Agents for Production of Lethal Infections.

Summary A simple method is described for the production of rapidly progressive lethal infections. It consists of the administration of two readily available agents in combination, cortisone and x-radiation, to laboratory animals in preparation for test within the next 24 hours as recipients of a microorganism. Cortisone and x-radiation each over a wide dosage range act synergistically to potentiate the enhancive effect that may result from the employment of either agent singly. The result of this effect is a remarkable alteration in the susceptibility of a test animal to infection by any of a variety of infectious agents, as evidenced by rapid weight loss, death and extensive histopathological lesions. The ready applicability of the method is illustrated in this paper by experiments that utilized four microbiological agents: poliomyelitis virus, a Coxsackie virus, Candida albicans, and Blastomyces dermatitidis. The experimental studies that employed these agents and the findings for other viruses, bacteria and fungi will be reported in detail elsewhere.

Mammalian cell-virus relationship. III. Poliovirus production by non-primate cells exposed to poliovirus ribonucleic acid.

Summary Exposure of non-primate cells in vitro and in vivo to poliovirus RNA resulted in production of infectious poliovirus, without induction of apparent cytopathogenic effect in vitro or disease in vivo.

Mammalian Cell Cultures for Study of Influenza Virus. I. Preparation of Monolayer Cultures with Collagenase.

Summary Primary monolayer cultures of human, rabbit and porcine fetal and adult lung were prepared conveniently by dispersion of tissue with commercially available collagenase in phosphate-free salt solution.

Propagation of poliomyelitis virus in cultures of monkey and human testicular tissues.

Summary In experiments in which monkey or human testicular cells in tissue culture were infected with poliomyelitis virus, Lansing and Yale-SK strains, proof was obtained that the virus in 3 passage series had been propagated. At the time this report was written it had been established for these 3 passage series that the minimal dilution factor based on tissue replacements ranged from 1014.6 to 1018 and, when assessed by fluid replacement, from 1029 to 1036. The LD50 of each strain of virus was determined on successive transfers, and the identity of each strain of virus was established by neutralization tests and histopathological findings in monkeys dead from the injection of tissue culture virus. Control experiments and other tests made known that propagation of poliomyelitis virus did not occur in the absence of viable testicular cells and that an extraneous virus was not inadvertently acquired during the course of these studies. It is plain that the results of the present investigation substantiate the observations of other workers that poliomyelitis virus can propagate in extraneural tissues. From these findings it is apparent that the assumption of an obligate neurotropism for poliomyelitis virus no longer is tenable. This observation and the demonstration regularly of the natural occurrence in poliomyelitis of virus in the oro-intestinal tract emphasize the need for a renewal of investigative studies of the factors that make possible invasion of the central nervous system to result in the overt and residual manifestations of poliomyelitis.

Absorption and translocation of mammalian viruses by plants. I. Survival of mouse encephalomyelitis and poliomyelitis viruses in soil and plant root environment.

Abstract The rates of degradation at 30° of strains FA and GD VII mouse encephalomyelitis viruses and of types 1 (Mahoney) and 2 (YSK) polioviruses in liquid and sand suspensions and in soil were determined. Mouse encephalomyelitis virus, strain FA, survived 3–4 weeks in sterile or nonsterile soil; virus in sterile or nonsterile liquid suspension survived for from 7 to 12 days. Soil pH influenced survival of GD VII virus which persisted for 3–4 weeks in alkaline soil, at least 5 weeks in neutral soil, but could not be recovered from acid soil after incubation for 24 hours. Type 1 poliovirus was recoverable from sterlle soil for 6–7 weeks, and from nonsterile soil for 2–3 weeks. Soil and soil clays adsorbed and held firmly large quantities of poliovirus. In the environment of roots of tomato plants cultivated hydroponically: (a) types 1 and 2 polioviruses were inactivated at similar rates, and (b) type 1 virus from cell culture was inactivated more rapidly than virus in monkey brain suspension. Monkey brain type 1 virus was inactivated more rapidly in the environment of pea roots than in that of tomato roots.

Replication of poliovirus in primate cell cultures maintained under anaerobic conditions.

Abstract Poliovirus, Type 1 (Mahoney strain), was shown to replicate in HeLa cells maintained anaerobically. While the latent period of virus development under anaerobic conditions compared to aerobic conditions was increased, the yield of virus per destroyed cell was similar. Infection by poliovirus did not significantly alter either oxygen consumption by HeLa cells maintained aerobically, or anaerobic acid production until cytopathology was evident. Absence of oxygen did not affect absorption of poliovirus by cells.

Propagation in vitro of Poliomyelitis Viruses VII. pH Change of HeLa Cell Cultures for Assay.

Summary A method is described for use of suspended HeLa cells and phenol red pH indicator for assay of poliomyelitis virus or antibody. A simple medium containing yeast extract and 4% chicken or monkey serum is used to maintain the cultures during incubation with virus or virus-serum mixtures. Infectious virus is revealed by unchanged color of the medium over a 6 day incubation period; neutralization of virus by antibody permits normal cellular metabolism during this period to produce acid with a change in color of the phenol red indicator in the medium to yellow. Evidence is presented for the agreement of results obtained by this method with those of the conventional established cell technic.

Absorption and translocation of mammalian viruses by plants. II. Recovery and distribution of viruses in plants.

Abstract Pea, tomato, potato, and lettuce plants in modified hydroponic culture were exposed to strain FA mouse encephalomyelitis virus or to type 1 (Mahoney) or 2 (YSK or MEF-1) polioviruses, under conditions permitting independent tests for absorption and translocation. FA virus regularly entered plant roots and attained significant concentration; acropetal translocation occurred infrequently. Type 1 poliovirus as mixed suspension of monkey brain tissue extract and cell culture virus was absorbed by tomato plant roots but not translocated to aerial parts; Mahoney and YSK polioviruses in tissue culture fluid, and MEF-1 poliovirus in mouse brain suspension, were neither absorbed nor translocated by plants. In summary of positive experiments, virus was recovered from 47 of 52 plant roots and from the aerial portions of 6 of the 52 plants. Evidence is presented that instances of virus absorption and translocation by plants did not in fact originate from (a) accidental contamination, (b) activation of latent virus in test mice, or (c) capillary ascension of virus along root exteriors.