36 LABOE.VTOKY BACTEEIOLOGY. 



EXEECISE XXVI. 



ISOLATION OF DENITKIFIERS. 



1. By means of a sterile platinum loop make transfers 

 from tubes 3, i, 5 and 6 of the previous experiment, into 

 tubes of sterile water. 



2. Then inoculate from these dilutions into three tubes 

 of liquefied and cooled (40° C.) "Giltay" agar in series. 



3. Pour Petri dishes as usual. 

 ■±. Incubate 4-5 days at 20° C. 



5. Inoculate from several well isolated colonies appearing 

 on these plates into tubes of sterile "Giltay" solution. 



6. Incubate 4—5 da}'s at 20° C, and if cultures are not 

 pure replate as before until pure cultures are obtained. 



7. Test qualitatively for the disappearance of nitrates as 

 in the preceding experiment. 



8. If nitrates are absent inoculations are made from each 

 culture into 100 cc. portions of "Giltay" solution in 500 cc. 

 Jena flasks, and when a qualitative test shows that all 

 nitrates have disappeared the solutions are digested by the 

 regular Kjeldahl method and the total nitrogen determined. 

 The difference between the amount of nitrogen found and 

 that supplied as nitrate will give the loss of nitrogen as gas. 



9. Examine the pure cultures of denitrifiers obtained and 

 identify by the usual cultural and morphological descrip- 

 tions. 



