The effect of calcium nutrition on the production of diffusible antigens by Rhizobium trifolii

Rhixobium trvolii grown in a defined medium deficient in calcium yielded additional diffusible antigens as compared with organisms re-ceiving calcium or strontium. The two kinds of organisms were indis-tinguishable in respect of their somatic agglutinogens. The same addi-tional precipitinogens were released from Ca-adequate bacteria by mechanical disintegration, freezing, drying, lysozyme or chloroform. Their occurrence in the untreated Ca-deficient bacteria is attributed to autolysis of these more fragile bacteria. The precipitinogen ( ‘ a ’ band) that was common to the Ca-adequate and Ca-deficient bacteria was not shared by a second strain of R. trifolii nor by 2 strains of R. meliloti. This ‘a ’ antigen was heat stable and appeared to be a component of the cell wall, from which it could be separated in diffusible form by formamide extraction of whole bacteria, trypsin digestion of broken bacteria and by mechanical disintegration of prepared cell walls. Neither of the additional antigens (‘b’, ‘c’) was related to agglutination; both were found in Ca-deficient or mechanically broken bacteria of the other strain of R. trifolii; ‘c ’ only was similarly shared with R. meliloti. The ‘b ’ band antigen appeared to be an intracellular component and was heat labile. Like ‘ a ’, ‘ c ’ was heat stable and associated with cell wall, but evidently was not ordinarily exposed, although readily released by lysis or mechanical breakage. The extracellular polysaccharide was neither antigenic nor haptenic, but the preparation obtained from Ca-deficient and mechani-cally broken Ca-adequate organisms was persistently associated with a co-precipitating antigenic component (chiefly ‘ a ’ band)