Effect of Hardening and Fixation on Gram Reaction.

Abstract

In a recent study of sections of the livers and spleens removed from mice, dead of experimentally produced anthrax, we were surprised to find that although the young culture used to induce the disease had been sharply gram-positive, the majority of the organisms with which the tissues teemed, were either completely or partially decolorized by gram. The experiment was repeated with Saccharomyces cerevisiae, a sharply gram-positive organism, more stable in this respect than B. anthracis. A heavy suspension of this organism was injected into the anterior abdominal vein of an etherized frog after the other vessels of the liver had been ligated. The liver thus impregnated with yeasts was immediately removed, and pieces of it were put into Zenker's fluid and into formalin. These were run through in the usual manner and cut in paraffine. The sections were stained by gram. (Burke's Modification). A large majority of the organisms were found to be completely or partially decolorized. This reversal of the gram reaction (similar to that recently described as following exposure of B. anthracis to acriviolet 1 ) might be thought of as occasioned by the contact of the organisms with the animal body, or as resulting from exposure of the organisms to the various chemicals used in hardening and fixation.

An inquiry was therefore instituted as to the effect of these processes on gram reaction. On account of the danger of this kind of experiment if done with B. anthracis, Saccharomyces cerevisiae was used. A heavy suspension of these yeast cells was made and put into two tubes. To one of these Zenker's fluid was added, to the other formalin. At the end of 24 hours, the specirnens were centrifugated.