30 



Prof. Marshall Ward. Experiments on the [Feb. 16, 



Experiments with Spores and Food Material on Separate Plates. 



In order to test still further the accuracy of my previous conclu- 

 sions, that the bactericidal action of the sunlight is direct, and not 

 due to secondary effects, owing to changes in the food material, I 

 carried out the following modifications of the experiments. 



Agar tubes of anthrax, cultivated for several days at 25° C, were 

 selected and examined for spores ; those were selected which exhibited 

 a rich crop of thoroughly mature spores. As is well known, it is 

 possible to remove from such cultures the thick, yellowish, creamy 

 layer of spore material without taking any appreciable quantity of 

 the agar, and great care was employed to accomplish this successfully. 

 Having shaken up these spores in sterile distilled water in a test- 

 tube, I poured a few drops of the infected liquid into each of several 

 Petri's dishes at 70° C, in a hot-air oven at that temperature, and 

 replaced the dishes to allow the water to evaporate into the hot air 

 during the cooling down of the whole. After two or three hours 

 the water had evaporated completely ; or, if not, the oven was heated 

 up again to 70°, and the drying completed, leaving a thin film of dry 

 spores sticking to the bottom of the glass dish. All bacilli and 

 immature spores were of course killed by the high temperature, and 

 only the thoroughly ripe and resistant spores left alive. 



These plates were then wrapped up with suitable stencil plates, &c, 

 exactly as in my previous experiments, and exposed to sunlight, as 

 before described. 



For each plate of spores alone thus prepared, I also made a corre- 

 sponding plate of agar alone, and exposed these side by side with 

 the plates of spores. 



Obviously, if the bactericidal action was direct, it ought to be 

 manifested on the cultures of spores alone, if those spores not killed 

 by the light could be made to develop in a thin layer of fresh agar 

 poured on the plates after exposure ; whereas, if indirect, and due to 

 changes in the agar, the effect ought to be visible by spreading a thin 

 layer of fresh spores over the agar which had been insolated. 



If both effects occur, then both sets of plates should show the 

 positive results. 



Of course this was a case where negative results with both plates 

 could not be used for any useful conclusions; for it is quite con- 

 ceivable that the destructive light-action might not occur in dry 

 spores, or that some volatile body* might be formed in the agar, 

 which was rapidly dissipated in the water used for the sowing of 

 fresh spores after insolation ; or again, the agglutinated layers of dry 

 spores might shelter one another from the sun's rays, and so inter- 



* E.g., it is not inconceivable that hydrogen peroxide might be formed from the 

 moisture in the agar. 



