Albert S. Kaplan

The susceptibility of monkey kidney cells to poliovirus in vivo and in vitro

Abstract A comparison of the reaction to poliovirus by kidney cells in vivo and in vitro showed that the kidneys of rhesus or cynomolgus monkeys failed to support the multiplication of Y-SK virus when inoculated in vivo , but cells from these kidneys were highly susceptible in vitro . This is in contrast to the cells of capuchin monkeys which were resistant in vitro as well as in vivo . The inoculation of monkeys with cortisone did not enhance their in vivo susceptibility to virus. There was also no significant difference in the susceptibility of cultures derived from such animals as compared with those from control animals. The inoculation of the kidneys of living animals with virus suspended in trypsin also did not increase the susceptibility of the kidneys in vivo . The in vitro sensitivity of the susceptible kidney cells from animals inoculated in vivo was shown also by the fact that in several instances, when virus could not be detected in the supernatant fluids of kidney homogenates, virus could usually be found in cultures made from the other kidneys of these animals. Cells from the kidneys of a rhesus monkey inoculated 24 hours previously were plated by the plaque technique to determine the number of infectious centers. The number of plaques obtained by titration of a portion of these cells disrupted by homogenization was not significantly different. This was interpreted to mean that virus was not growing in the cells in vivo even to a limited extent but that virus was most likely adsorbed to the cells during the process of trypsinization.

Comparison of susceptible and resistant cells to infection with poliomyelitis virus.

Ever since the discovery of the virus by Landsteiner and Popper, pdiovirus has manifested an extremely narrow host range. Even in vitro strains of the three virus types have been found to multiply and cause cytopathic changes in cultures of tissue derived from primate sources only. A wide variety of tissues of primate origin are susceptible to the action of poliovirus,2 but human kidney tissue: as well as that from rhesus and cynomolgus monkeys: seems to be more susceptible than most of the normal tissues employed for virus growth. The results of studies on the susceptibility of tissues from various species have been obtained either with tissue fragments embedded in a plasma clot, or with suspended tissue fragment cultures. Since monolayer cultures of renal epithelium are more susceptible to poliovirus, we considered it possible that such cultures obtained by trypsinization6 from various species might prove more susceptible than hitherto suspected. The purpose of this report is to compare the reactions to poliovirus of epithelial cells derived from the kidneys of four different species. The animals selected for this study were newborn rabbits, immature hamsters, and capuchin and rhesus monkeys. These animals show a progressively greater susceptibility to poliovirus in the order listed. The kidneys from these animals were trypsinized and cultures prepared in tubes, using the lactalbumin hydrolysate-calf serum medium, as outlined in Doctor Melnicks paper. When the monolayers had formed, each was inoculated with Brunhilde, Y-SK, and Leon strains of poliovirus which had been through 25 passages in cultures of rhesus monkey kidney. Only the rhesus kidney cells supported the growth of these polioviruses. This is illustrated in TABLE 1 for the capuchin monkey. The three representative strains of poliovirus were incubated with cultures of capuchin renal epithelial cells. No cytopathic changes were noted and, after two weeks of incubation, the culture fluids were harvested and titrated in rhesus kidney cultures. The results show that the three types of tissue-culture virus persisted without giving any evidence of having multiplied, since the titer of virus found did not differ significantly from that of the control tubes without tissue. Experiments with capuchin cultures inoculated with human stools proved to contain poliovirus suggest, however, that renal epithelial cells from this species may support the growth of type 1 poliovirus. Capuchin cultures were inoculated with three different specimens of human stools, each stool containing a different virus type. The fluids were harvested a t the end of two weeks and titrated in rhesus kidney cultures. As the results in TABLE 1 show, the fluids from the cultures of capuchin cells inoculated with type 1 fecal virus contained a t least 10,000 times more virus than the control tubes without tissue. Similar results were obtained on second passage of this material in capuchin

Differences in Thermostability of Antigenically Related Strains of Poliomyelitis Virus.

Summary Three rodent-adapted strains of poliomyelitis virus, all belonging to the same immunologic type, were found to differ in their thermostability; these were, in decreasing order of resistance, Y-SK, MEF1, and Lansing. The differences between these strains became evident only when they were suspended in cream or ice cream before being heated. All three strains suspended in water were inactivated by heating at 71.1 °C for 15 seconds. When heated in cream the viruses withstood 79.5°C for 15 seconds and in ice cream they withstood 82.2°C for 25 seconds, with Y-SK resisting inactivation to the greatest extent.

Oral Administration of Coxsackie Viruses to Newborn and Adult Mice.

Summary (1) Newborn mice may be infected by oral administration of at least 5 different strains, representing 4 immunologically distinct Coxsackie, or C, viruses. Adult mice were resistant. (2) Newborn mice surviving oral administration may produce neutralizing antibodies. (3) Adult female mice fed C virus did not transmit neutralizing antibodies to their offspring, in contrast to females inoculated parenterally. (4) In a comparison of routes of inoculation of newborn mice it was found that higher titers may be reached by subcutaneous than by oral administration. The subcutaneous route proved to be about 10,000 times more sensitive than the oral route. (5) Direct contact between inoculated and uninoculated mice did not result in the transmission of infection.

Multiplication of virulent poliovirus in capuchin monkey kidney cultures without microscopically observed cytopathogenicity.

Summary Cultures of trypsinized kidneys of South American capuchin monkeys (Cebus capucina) supported the multiplication of Type 1 polioviruses, even though no cytopathic changes were observed by microscopic examination of the cultures. The failure to observe cytologic changes was found to be due to the small proportion of cells in the culture capable of supporting the growth of virus. After 4 to 5 tissue culture passages the viruses remained virulent for the monkey brain and spinal cord. No evidence was obtained for the propagation of Type 2 and 3 strains in the capuchin cultures.

Dual Antibody Response to Coxsackie and Poliomyelitis Viruses in Patients with Paralytic Poliomyelitis.

Summary Both poliomyelitis virus and Coxsackie virus (C virus) were isolated from the acute phase stools of several paralytic patients during an epidemic of poliomyelitis which occurred in Easton, Pa., in 1949. In 2 such cases, neutralizing antibody tests were carried out with acute and convalescent serum reacting with a suspension of virus obtained directly from acute phase stools. It was found that both patients responded to their illness by simultaneously developing antibodies both to poliomyelitis virus and to C virus.