Oxidation of pyruvate, acetate, succinate, and glutamate was compared in Bacillus thuringiensis, B. alvei, B. lentimorbus, and B. popilliae. Cells of B. thuringiensis and B. alvei in transition from vegetative growth to sporulation oxidized these substrates by tricarboxylic acid (TCA) cycle reactions. No TCA cycle activity was exhibited by B. lentimorbus and B. popilliae cells that do not sporulate. B. popilliae decarboxylated C-1 of pyruvate and glutamate; B. lentimorbus, C-1 of pyruvate only. B. thuringiensis and B. alvei oxidized pyruvate and acetate at a much higher rate in the absence of amino acids and related compounds than when these nutrients were exogenously supplied; in contrast, there was no appreciable increase in C-1 decarboxylation of pyruvate by B. lentimorbus and B. popilliae. No nutrient effect was observed on succinate and glutamate oxidation in any of these four organisms.

The catabolic pathways for use of glucose in proliferating vegetative cells of Bacillus thuringiensis, B. alvei, B. lentimorbus, and B. popilliae were studied by radiorespirometry. These organisms dissimilate glucose predominately via the Embden–Meyerhof–Parnas pathway and to a lesser extent by the pentose phosphate pathway. Extent of participation of concurrent pathways varied with each organism. Tentative evidence suggests that B. popilliae and B. lentimorbus, grown in a yeast extract – glucose medium, lack a fully operational tricarboxylic acid (TCA) cycle. Dilution of this medium slightly enhanced TCA cycle activity in B. popilliae but had no effect with B. lentimorbus. Radiorespirometric data regarding glutamic acid oxidation also were obtained for each bacterium. All organisms studied except B. lentimorbus were capable of oxidizing glutamic acid to carbon dioxide.

Spores of the insect pathogen Bacillus popilliae Dutky have been formed in vitro from vegetative cultures. The procedure results reproducibly in 0.1 to 0.3% spore formation in cells of colonies grown on a solid medium under strictly denned conditions. Sporulation requires a selected strain of the organism, NRRL B-2309S, a relatively large and specific concentration of certain yeast extracts, a specific type of agar, the complete absence of glucose, the presence of acetate, and a pH within the range 7.2 to 7.5. Spore formation occurs slowly during 2- to 4-week incubation periods in surface colonies present in limited numbers on agar plates. Some of the spores formed in this manner survive heating for 15 minutes at 80 °C, and vegetative cultures derived from such spores are pathogenic via injection for larvae of the Japanese beetle, Popillia japonica Newman.

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Growth characteristics of the insect pathogen, Bacillus popilliae Dutky, were studied by propagation in shaken flasks and in 2-liter fermentors. Maximal populations between 5 X 108 and 2 X 109 viable cells per milliliter of culture medium routinely were obtained in incubation periods of 18 to 24 hr at 30 C in a medium composed of 1.5% yeast extract, 0.6% K2HPO4, and 0.2% glucose or trehalose. The carbohydrate required for growth in liquid media was fermented with the formation of 2 meq of acid per mmole of carbohydrate utilized; acid products ordinarily were not subsequently metabolized. B. popilliae is an aerobe, and the amount of growth obtained varied with aeration to an optimum at oxygen absorption rates of about 0.5. Maximal populations persist in a culture for periods of only 1 to 4 hr; cessation of growth was followed immediately by rapid death of cultures, so that less than 1% of the cells remained viable after 48 hr, and viability often was lost entirely by the end of 72 hr of incubation. No cytological evidence for spore formation was observed under any growth condition. Death was not associated with lysis of the cells, although extensive granulation ultimately occurred. Continuous neutralizaiton, augmented buffering, various techniques of dialysis, or slow feeding of the carbohydrate did not markedly alleviate the characteristic death of the cultures.