560 



FISHERY BULLETIN OF THE FISH AND WILDLIFE SERVICE 



Table 6. — Growth of sulfonamide-resixtant and sulfonamide-sensitive strains of Aeroiiionas salnioiiicida in buffered, nutrient 



broth with various sulfonamides 



IQuestion mark (?) Indicates reliable reading could not be made because of precipitate in medium] 



broth were determined with laboratory-prepared 

 sodium saUs of sulfadiazine, sulfameraziiie, sul- 

 famethazine, and sulfisoxazole. Ten strains of 

 sulfonamide-sensitive and 10 resistant cultures 

 were used. Serial dilutions of sulfonamides ranged 

 from 1:100 to 1:102,400. Control test tubes 

 contained medium without sulfonamides. After 

 24 hours of incubation the resistant strains grew 

 well in a sulfonamide concentration of 1:100 

 (1 : 200 with sulf amerazine) . Growth of the sensi- 

 tive strains was absent in the following concen- 

 trations and in all lower dilutions: 



Sulfadiazine 1 : 12,800 



Sulfameraziiie 1 :6,400 



Sulfamethazine 1 : 6,400 



Sulfisoxazole 1 : 6,400 



The amount of growth was much reduced up to 

 the highest dilution of the sulfonamides tested 

 (1:102,400, or 10 Mg- per ml.). After 48 hours 

 the results differed very little except in the medium 

 with sulfamethazine which showed abundant 

 growth up to 1:200 and scanty at 1:100 (the 

 highest concentration tested). 



The final series of experiments was conducted 

 for the purpose of selecting a reliable method of 

 determining the drug sensitivity of A. mlmonicida 

 under either laboratorv or field conditions. 



In a preliminary test, the optimum pH range 

 for the appearance of growth inhibition zones on 

 an agar medium was determined. Filter paper 

 disks (6 mm. in diameter) * were impregnated 

 with 1 mg. of filter-sterilized sodium sulfadiazine 

 dissolved in 0.02 ml. of water and air dried at 

 37° C. Standard furunculosis medium (buffered, 

 adjusted to pH 7.0, 7.5, 8.0, 8.5, and 9.0, and 

 sterilized) was distributed in 20-ml. quantities in 

 10 petri plates (10-cm. in diameter), the medium 

 was inoculated by swabbing tlie surface with a 

 24-hour culture of A. salmonicida, and disks 

 were positioned. Results were recorded after 24 

 and 48 hours of incubation at 20° C. Zones 

 were distinctly visible after 24 hours, but the 

 width of the zones could be more conveniently 

 determined after 48 hours. The widths of the 

 zones with sulfonamide-sensitive strains at various 

 pH levels were as follows: 



pH 7.0 II mm. 



pH 7.5-8.0 15 mm. 



pH 8.5 16 mm. 



pH 9.0 17 mm. 



There were no clear zones with sulfa-resistant 

 strains. The best results, based on the width of 



< No. 740-E , C. Schleicher &. Sehuell Co. 



