Scott, H. W. (Ohio Agricultural Experiment Station, Wooster), and B. A. Dehority. Vitamin requirements of several cellulolytic rumen bacteria. J. Bacteriol. 89:1169-1175. 1965.-Four strains of cellulolytic bacteria recently isolated from in vitro rumen fermentations were used in this study. Nine water-soluble vitamins were tested in single-deletion and single-addition plus biotin experiments, each with and without charcoal-extracted casein hydrolysate. Bacteroides succinogenes A3C and B21a required only biotin under the above experimental conditions. Ruminococcus flavefaciens B34b showed an absolute requirement for biotin and was stimulated by p-aminobenzoic acid (PABA) in the single-deletion experiments. In the single-addition plus biotin experiments, PABA and, to a lesser extent, vitamin B(12) appeared to be required for maximal growth. The presence or absence of casein hydrolysate did not affect the vitamin requirements for the aforementioned three strains. In the single-deletion experiments, R. flavefaciens Cla showed an absolute requirement for biotin and, when casein hydrolysate was omitted, for B(12). When casein hydrolysate was present, no requirement for B(12) could be observed. In the single-addition experiments where the basal medium contained biotin and casein hydrolysate or B(12), PABA was required for maximal growth; however, the single deletion of PABA caused only slight retardation of growth. Investigation of the B(12) or casein hydrolysate requirement of Cla revealed that a mixture of purified amino acids simulating casein hydrolysate satisfied this requirement. Subsequent work indicated that this requirement could be satisfied by the amino acid methionine.