LACTOBACILLUS: YEAST INTERRELATIONSHIPS

Abstract

Progress has been made during the past several decades in the isolation and identification of the flora and fauna from the rumen, soil, and other natural environments, and in the determination of the functions of and products formed by these isolates in pure culture. On the other hand, actual microbial interrelationships have been little studied because of their complexity. Mixed cultures are especially difficult to study in vitro. Whether various microorganisms behave similarly in pure and mixed culture is, to a large extent, unknown. The experiments reported here describe a comparative study of certain microorganisms in pure and mixed culture. The experiments were conducted using a relatively simple system as a model, or basis, from which attempts to gain fundamental information on mixed culture systems might depart. MATERIALS AND METHODS Lactobacillus plantarum, and a strain of Sac-charomyces carlsbergensis (Culture YB-5050, A.R.S., U.S.D.A., Peoria, Illinois), were isolated from a sour dough starter culture and were used in the first series of experiments. I actobacillus casei strain ISU-2K3 and the sour dough yeast were used in the second series of experiments. The combined effects of temperature, initial pH, and medium on mixed cultures of bacterium and yeast and on pure cultures of the organisms were determined. Through observations made in preliminary experiments, it was decided that temperatures of 22, 27, 32, and 37 C would be used, together with initial pH values of 4, 5, 6, and 7 and an incubation period of 48 hr. Two different media were used: sucrose-flour broth (medium I) and sucrose broth (medium II). Medium I had the following composition: beef extract (Difco), 3 g; tryptose (Difco), 10 g; sucrose, 5 g; sodium chloride, 5 g; distilled water, 1 liter; sterile flour, 35 g per 100 ml of broth. The flour was sterilized in ethylene oxide and added after heat sterilization of the medium. Medium II was identical to medium I, except the flour was omitted. Concentrated lactic acid or 6 N sodium hydroxide were used to adjust the pH of the media, which were subsequently auto-claved for 12 min at 121 C. Since the addition of flour and sterilization caused changes in the pH of the media, pH adjustments were made to compensate for the effects of these two factors. Inocula were prepared by culturing the organisms in medium II for 18 hr at 32 C. Two milliliters of inoculum were used per 200 ml of medium in a 500-ml Erlenmeyer flask. The lactobacilli were enumerated using tomato juice agar (Difco) and the yeasts were enumerated on Littman's oxgall agar (Difco). To observe the real effects of each of the factors, both series of experiments consisted of a factorial whereby each series of treatments was tested in all combinations; then the data were subjected to analyses of variance (Snedecor, 1956). RESULTS As is shown in the tables and figures, there were many similarities in the maximal population densities of the yeast and lactobacilli in pure and dual cultures under most of the experimental conditions used. To avoid confusion and for the sake of brevity, the similarities will, in large part, be ignored so that certain differences which might be attributed to microbial interactions can be emphasized. L. plantarum:S. carlsbergensis interrelationships. (1) Effects of temperature:-A significant (P = 0.01) lactobacillus:yeast interaction was noted in medium I (Table 1), although this same interaction did not approach significance (P = 0.05) in medium II. Yeast counts were markedly reduced when the yeast was grown in association with L. plantarum at 37 C (Fig. 1); however, the lactobacillus counts were unaffected. (2) Effects of pH:-Microbial interactions 519