10 M. LEVINE 



The medium consisted of 1.0% peptone, 0.1% glucose and 0.2 to 0.7% phos- 

 phate. Inoculation of the agar plates was made from a 24-hour broth culture. 

 A concentration of 0.4-0.5% of the phosphate gave best results with 6 cultures 

 examined. It is interesting in this connection to note that the titratable acidity 

 with phenolphthalein was in each instance -f 0.7%. The H-ion concentration 

 was much more varied, probably 7.1 with the 0.2% of the phosphate and 7.8 

 with the 0.7% of the buffer salt, as indicated by subsequent experiments. 



The influence of the H-ion concentration on the growth of the dysentery 

 bacilli seems marked on solid medium. It has been my experience that in 

 liquid medium the effect of the H-ion concentration is not so evident. 



Experiments on the effect of the concentration of dipotassium phosphate 

 repeated with 39 strains using 3 concentrations of the salt (0.2, 0.45 *and 0.7%, 

 respectively), showed that: 



1. The phenolphthalein titration is a poor index of the true acidity of the 

 medium. The variation in the titrable acidity was close to the limit of experi- 

 mental error, whereas the difference in H-ion concentration with the different 

 quantities of phosphate was marked and distinct. 



2. The optimum reaction is not the same for all strains of dysentery. Two 

 (5.1%) grew best with the largest quantity of phosphate, four (10.2%) with 

 the least amount of phosphate and twelve (30.7%) show their optimum growth 

 when 0.45% of dipotassium phosphate was used. Seventeen (43.6%) did equally 

 well on all of the 3 mediums. Considering all cultures, we find 33 (84.7%) to 

 have done as well or better with 0.45% phosphate than on either of the other 

 concentrations of this salt. The H-ion concentration with 0.4% of the phos- 

 phate is generally 7.4 or 7.5. This quantity was selected as probably the most 

 reliable and desirable. 



Choice of Indicator. A distinct and noninhibitory indicator is an important 

 adjunct to the successful isolation of dysentery bacilli. It was hoped that the 

 eosin and methylene-blue combination of Holt, Harris and Teague, which was 

 found so valuable in water work, might be successfully employed, particularly 

 as it was reputed to be noninhibitory. Thirty-nine strains of dysentery bacilli 

 were inoculated on agar with and without the indicator from a 24-hour peptone 

 phosphate culture. 



The composition of the medium was: 



Agar 1.5% 



Peptone 1.0% 



Dipotassium phosphate 0.4% 



Glucose 0.1% 



Indicator per 100 c c of above 



Eosin 2% yellowish aq 2.0 c c 



Methylene-blue 0.5% aq .' 2.0 c c 



(The P H of this medium was 7.5). 



B. dys. Shiga was markedly inhibited. A slight growth was observed on 

 prolonged incubation (48-72 hours). Sixteen, or 50%, of the mannite fermenting 

 dysentery strain were partially inhibited. 



Of a number of indicators tried, the china-blue rosolic acid mixture was 

 found to be the least inhibitory when working with pure cultures. Similar 

 results were obtained with artificial suspensions of dysentery organisms in 

 normal stools. 



