Abstract
The employment of fertilized egg has proven of value for the cultivation of some bacteria and a number of filterable viruses. The chorio-allantoic membrane has been mostly used as the site of infection. Since the cultivation of anaerobic microorganisms in unfertilized and developing fertilized eggs has not been recorded, the preliminary results of such a study with an obligatory anaerobe, Cl. tetani, is herewith communicated.
Technic. A pure toxigenic strain of Cl. tetani, recently isolated from a patient, was used. Cultures in cooked meat tube after 48 hours' growth were used as the inoculum. Unfertilized and developing fertilized hens' eggs of different age were employed. After the position of the embryo and the air sac was identified by transillumination and marked off and the shell opposite the air sac was sterilized with tincture of iodine, a small opening was made with a carborundum disc attached to a dental drill, leaving the inner shell membrane intact, which was later cut with a small knife after resterilization with 70% alcohol and heat. A loopful of inoculum was then introduced through this opening to the center of the yolk sac, and in the case of fertilized eggs toward a point at some distance from the embryo. The shell aperture was then closed with melted hard paraffin. The eggs were then incubated at 37°C. Different batches of them were taken out and examined grossly and by stained smears microscopically. Aerobic contaminations were excluded in every instance by absence of growth of the eggs' content on blood agar plates.
Results. At first, cultivation was done in unfertilized eggs. Fifty-four eggs were divided into 9 groups of 6 each: One group unheated, others heated in water bath at 90°C for 0.5, 1.0, 1.5, 2.0, 3.0, 5.0, 10.0 minutes respectively, and still others fully cooked by boiling for over 15 minutes.
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