037 - Relation of Vaccinal Immunity to the Persistence of the Virus in Rabbits

I once heard a claim from an anti-vaccine proponent that immunity from vaccines, instead of clearing the pathogen from the body the way the immune system does in the course of a “natural” infection, instead forces pathogens such as the measles virus to remain latent in cells, hiding in our bodies in an invisible state, until such time as our immunity might wane and they can be released to cause havoc. While I don’t think this claim was well-supported by any evidence, my encountering it did cause the current study to pique my interest.¹ It may not be very satisfying to my curiosity about the above claim though.

Some background: what was apparently a new technology, important at the time and foundational for this study, was electrophoresis (called cataphoresis by the authors Olitsky and Long): this technique, still used today mostly for separating all the proteins or nucleic acids in a sample based on their size, involves putting the sample into a gel immersed in a buffer full of electrolytes, and then running an electric charge from one side of the gel to the other. Since proteins and nucleic acids have an ionic charge, the current will pull them to one electrode or the other, depending on their charge. The larger ones get caught and stuck more as they’re pulled through the gel, so they move more slowly, while smaller ones can fit through smaller spaces, so they move more quickly.

In this study, though, their samples were whole viruses, rather than individual proteins or nucleic acids. People were trying electrophoresis on all kinds of different pathogens (viruses, but also protozoan parasites and virus-like bacteria^2–5) to see whether they moved toward the anode or the cathode. The technique herein was used to isolate and concentrate virus from animal tissues, virus that was too scarce to be detected by other methods (at the time). I should mention that in any case, “detection” of the virus involved inoculating other rabbits with the sample and seeing if they produced a characteristic infection reaction. It wasn’t good to be a rabbit in that lab in those days.

The hypothesis Olitsky and Long were testing was based on previous observations by others, that viruses could be detected in animal’s (and sometimes human’s) tissues longer after that organism had recovered from the viral illness. For example, polio virus had been detected from an infected monkey and a person several months after they had recovered. The authors speculated that immunity to a virus depends on the persistent presence of that virus in an animal’s tissues; after the virus is completely gone, the animal is no longer immune. (This observation and hypothesis didn’t distinguish between a “natural” infection or a live attenuated immunization, such as vaccinia inoculation as a vaccine against smallpox, so it wouldn’t support the anti-vaccine claim above in any case.)

Their model for this study was vaccinia (cowpox) virus infecting the testicles of rabbits. They took rabbits that had been infected at various times past, from 12 to 133 days previous to the study, and recovered. Samples of these animals’ tissues were taken under anesthesia, keeping the animals alive so they could be tested for immunity later, and electrophoresis was used to detect virus in the tissue samples. Olitsky and Long detected virus in samples from each of these rabbits, even the one infected more than 4 months before. (Note of contention: I don’t think they had any negative controls to make sure they weren’t just detecting contamination or something.)

Then the authors took five more rabbits that had recovered from viral infection 119 to 183 days before and tested them for virus. They took samples, seemingly randomly, from spleen, testicles, and skin, and checked for virus using electrophoresis. In the rabbits infected earlier, no virus was isolated, and they appeared to have lost their immunity; in those infected later, the authors did detect virus, and the rabbits seemed immune. So they concluded that the persistent presence of virus was necessary for immunity.

The next step was based on another previous finding that immunity seemed to be transmissible from mother to offspring. This was based on some observation of a pregnant mother and her newborn, and some preliminary animal study. So Olitsky and Long tested this hypothesis in two pregnant rabbits, one infected soon before giving birth, and the other infected but given time to recover before birth. The former’s offspring seemed immune, and electrophoresis recovered virus from the testicles of a male baby rabbit; but the latter’s offspring seemed susceptible to infection as normal. So the conclusion was that immunity (and persistent infection) is heritable only when the mother is infected at the time of birth.

These are interesting results, but the quality of evidence seems pretty low. There weren’t very good controls, and the sample sizes were quite small. Also it is in rabbits, which isn’t necessarily a good model animal. This poor quality seems borne out by later studies, some by the same researchers, that contradicted or found better explanations for these results:

"By the means employed in this investigation, vaccinal immunity has been shown to endure beyond the persistence of recoverable virus."⁶ (Similar results in other studies^7,8)

"Whatever the detailed mechanism, it is evident in our study that newborn rabbits have been made tolerant immunologically toward the antigens of vaccinia virus for at least a relatively brief period. Perhaps tolerance also explains the early observations of Olitsky and Long [037, this study] made on a single pregnant rabbit and its 3 offspring."⁹

References:

1.  Olitsky, P. K. & Long, P. H. Relation of Vaccinal Immunity to the Persistence of the Virus in Rabbits. J. Exp. Med. 50, 263–272 (1929).

2.  Olitsky, P. K., Rhoads, C. P. & Long, P. H. The Effect of Cataphoresis on Poliomyelitis Virus. J. Exp. Med. 50, 273–277 (1929).

3.  Kligler, I. J. Recovery of Fowl-pox Virus from Vaccines by Cataphoresis. Br. J. Exp. Pathol. 12, 42 (1931).

4.  Kligler, I. J. & Olitzki, L. Cataphoresis Experiments with Typhus Virus. Br. J. Exp. Pathol. 12, 69 (1931).

5.  Salle, A. J. The Electrical Behavior of Leishmania donovani. J. Infect. Dis. 49, 450–454 (1931).

6.  Morgan, I. M. & Olitsky, P. K. A Study of Immunity in Rabbits from Two to Three Years After Infection with Vaccine Virus with Attempts to Recover Active Virus. J. Immunol. 39, 1–15 (1940).

7.  Pearce, J. M. The Persistence of Vaccine Virus in Rabbits Immunized to Vaccinia by a Previous Infection and the Relationship of Immunity to Latent Virus. J. Infect. Dis. 66, 130–137 (1940).

8.  Olitsky, P. K. & Casals, J. Concepts of the Immunology of Certain Virus Infections. Bull. N. Y. Acad. Med. 21, 356 (1945).

9.  Flick, J. A. & Pincus, W. B. Inhibition of the Lesions of Primary Vaccinia and of Delayed Hypersensitivity Through Immunological Tolerance in Rabbits. J. Exp. Med. 117, 633–646 (1963).