A simplified method for cultivating spirochaetes on liquid media

Abstract

The method I wish to describe is a modification of the original method of Noguchi 1 for cultivating spirochaetes, which I think might be of interest to those who are called upon to cultivate these organisms. Noguchi's method, although very efficient, offers one great disadvantage, namely that the rubber stopper connection between the upper and lower parts of the apparatus often becomes a source of contamination of the culture. The method I propose consists in utilizing the principles of anaerobic cultivation in general as well as those special features which were worked out by Noguchi. This new method can be used in two different ways. Firstly, one can use the tube as shown on Fig. 2 which instead of having the rubber connection of Noguchi joining its two parts as shown in Fig. 1, is made entirely out of one glass tube; but otherwise can be used exactly as Noguchi's apparatus, namely the lower part in which the piece of rabbit kidney is put before the tube is drawn out, is filled with the ascitic broth or sheep serum water up to the point where the tube broadens out again; another piece of tissue is placed in the upper portion of the tube and this tube is filled with the ascitic agar into which the spirochaetæ culture is placed. 2 Sterile paraffine oil in a thin layer is placed above the agar and the tube is incubated. Spirochaetes during their growth filter through into the lower portion of the tube exactly as in Noguchi's method. This method is especially convenient when one intends to open the tube many times to examine its contents. The other and better way however of cultivating spirochaetes which does away entirely with the upper part of the tube: is the following: I put a piece of tissue at the bottom of the tube; draw it out as before; introduce by means of a capillary pipette the spirochaetae culture and ascitic broth in the lower tube; connect the tube with the vacuum pump, as shown in Fig. 6, warming the lower part of the tube in a water bath at 37″ to facilitate the exhaustion of the air; cover the ascitic broth, after exhaustion, with sterile paraffine oil by means of a special arrangement taking advantage of the negative pressure in the tube, and finally seal the lower part of the tube at the point of strangulation as shown in Fig. 7.