Virulent Yersinia pestis was grown on heart infusion blood agar and examined by scanning electron microscopy, exposing the fraction 1 envelope antigen on cell surfaces as a lumpy coating that spilled into the surrounding milieu. The amount of antigen depended on both the duration and temperature of incubation. Comparison of the cultures grown at 37 degrees C for 24, 48, and 72 h with those grown at 22 degrees C for identical periods demonstrated that the antigen increased in amount with the length of incubation, and that the overall production of antigen was much greater of 37 degrees C than at 22 degrees C. These experiments visually confirmed the findings of our previous immunological studies. Moreover, we established that the closely related, virulent organism Y. pseudotuberculosis bears no such antigenic coating at any temperature or incubation period. In addition, the emergence of multiple flagella was noted when Y. pseudotuberculosis was grown at 22 degrees C in a liquid medium, whereas Y. pestis remained without these organelles. These observations preceptibly corroborated the absence of fraction 1 envelope antigen and the presence of flagella, respectively, for distinguishing Y. pseudotuberculosis from Y. pestis.