Isabel M. Morgan

Observations on Acquired Cellular Resistance to Equine Encephalomyelitis Virus

Summary Acquired, non-specific cellular resistance to equine encephalomyelitis virus has been described and differentiated from specific immunity induced by vaccination or passive immunization.

Protective Antibodies against Equine Encephalomyelitis Virus in the Serum of Laboratory Workers

It has been shown recently 1 that a large proportion of mice and guinea pigs develop, with increasing age, physiological or structural barriers that prevent certain viruses from invading the central nervous system. This resistance is demonstrable when virus is given peripherally, as, for example, intraäbdominally or intramuscularly, but not when it is injected directly into the brain. It is not a result of prior infection nor is it associated with the presence of protective substance in the serum. Furthermore, in the recent epidemic of equine encephalomyelitis (E. E.) in man in southeastern Massachusetts, 2 children were predominantly affected. The older animals which resist the E. E. viruses develop systemic infection, as is evidenced by the finding of virus in the circulation and later the presence of protective antibodies in the serum. In view of this suggestive relationship of age of both the experimental animal and man to clinically apparent infection with this virus, it was thought desirable to undertake a study of the protective capacity of the serum in certain individuals in our laboratory, who had been in contact with the E. E. virus for a period extending from 1 to over 6 years. The results would indicate whether a clinically inapparent infection, as determined by the presence of protective antibody, could possibly have occurred during that time. Serum-protection tests were carried out in mice by the intraabdominal method of Olitsky and Harford; 3 that is, by injecting by the intraäbdominal route, 0.03 cc of a mixture of equal parts of test serum and virus-dilutions (tenfold dilutions were used) into 15-day-old mice. Each mixture was given to groups of 3 or 4 mice. The Eastern strain of virus (E. E. E.) was derived from a stock which was frequently passaged in mouse brain and was again passaged through mouse brains immediately before use.

Vaccination with Various Western Equine Encephalomyelitis Viruses; Comparison as Antigens and as Test Inocula.∗:

Conclusions (1) The immunological identity of 4 strains of W.E.E. virus was confirmed by tests in mice. (2) Results of comparative tests for their antigenic potency suggested that the one most recently isolated may be somewhat superior to the older laboratory strains. (3) Mice vaccinated with comparable doses of either of the 4 strains showed equal degrees of resistance to 3 of the strains, while tests with the 4th, the current R.I. strain, indicated that this strain is not a sensitive indicator of the degree of immunity of vaccinated mice. (4) Parallel tests in which vaccinated mice were challenged intracerebrally with the current and with earlier samples of the R.I. strain indicate that after its 9th or 10th year of propagation in laboratory mice, certain changes may have taken place which rendered this strain increasingly less suitable for use in challenge tests for immunity.

Immunizing Capacity of Virus of Eastern Equine Encephalomyelitis Inactivated by ultraviolet Light.

The inactivating effect of ultraviolet light on viruses as well as bacteria has been observed repeatedly. That viruses so inactivated may be effective as immunizing antigens has already been shown for at least 2 viruses capable of causing disease in man. With a vaccine prepared by irradiating mouse-brain infected with rabic virus, Webster and associates 1 successfully immunized mice and dogs against a subsequent injection of active virus. Salk, Lavin, and Francis 2 compared the antigenic potency of epidemic-influenza virus following irradiation with that of active virus. In high concentrations, irradiated virus was nearly as effective an immunizing antigen as active virus; when lower concentrations were tested, a hundredfold loss in immunizing capacity was found to have occurred during irradiation. Ultraviolet light has been applied to the virus of equine encephalomyelitis, Eastern strain (E.E.E.), by Sharp and associates; 3 they studied the molecular stability of ultraviolet-treated virus. The preparation of an immunizing antigen produced by irradiation of E.E.E. virus with ultraviolet light is reported here. Chick embryos 7-days-old were inoculated with 0.1 cc 10-3 suspension of E.E.E. virus-infected embryo in 0.85% saline solution. Embryos removed from the egg 18-20 hours after inoculation were rinsed in saline solution, ground in a mortar and made to a 10% suspension in saline or Tyrodes solution. This suspension was centrifuged at 2000 rpm for 10 minutes; the centrifugation was repeated with the supernate; the supernate then obtained was spun in a Swedish angle-centrifuge at 4000 rpm for 45 minutes. About 30 cc of the final supernate were transferred to a quartz test-tube, with an internal diameter of 2.1 cm, which was placed in the center of a quartz-mercury resonance lamp in the form of a spiral∗ with an internal diameter of 9 cm.

Influence of Age on Rate of Immune Response of Mice to Formolized Equine Encephalomyelitis Virus

Summary The rate of development of neutralizing antibodies in serum of mice immunized with formalin-inactivated virus of Eastern equine encephalomyelitis has been shown to increase progressively with age. The antibodies in serum of mice immunized at a very early age did not reach the maximum titer found in mice immunized when older. The low degree of active immunity to intracerebral injection of active virus induced in mice 14 days old at the beginning of immunization did not increase from 2 to 4 weeks after immunization. During that interval, mice immunized at 3 months of age maintained a high degree of active immunity.

Relation of Age to Immune Response of Mice to Formolized Equine Encephalomyelitic Virus

Summary The ability of mice to be immunized by means of formolized virus of Eastern equine encephalomyelitis increases with age, as shown by the strikingly higher resistance of older immunized mice to the intracerebral injection of active virus, as well as by the amount of neutralizing antibodies developed.