Production of Bacterial Cellulose by Cocultivation of Komagataeibacter sucrofermentans with Producers of Dextran Leuconostoc mesenteroides and Xanthan Xanthomonas campestris

Abstract

Bacterial cellulose (BC) is an extracellular product of bacterial metabolism. BC has the same molecular formula as plant cellulose, but their structures are significantly different. Due to its unique properties (high degree of crystallinity, purity, good water-holding capacity), bacterial cellulose is widely used in many areas of human life. However, despite all the advantages of BC over plant polymers, its production is a relatively expensive process. Thus, one of the ways to increase the polymer yield can be the joint cultivation of a bacterial cellulose producer strain with other polysaccharide producers. From literature data, it is known that there is a positive effect of some water-soluble polysaccharides on the yield of BC. In addition, many biosynthetic genes remain silent and not expressed in vitro, thereby severely limiting the chemical diversity of microbial compounds that can be obtained by fermentation. In contrast, the cocultivation of two or more different microorganisms mimics a real “situation” where microorganisms coexist in complex microbial communities. It has been proven that competition or antagonism that occurs during cocultivation leads to a significant increase in existing compounds and/or to the accumulation of new compounds that are not found in axial cultures of the producer strain. The purpose of this work was to study cocultivation as a way to increase the yield of bacterial cellulose during the cultivation of BC producers with other polysaccharide-forming strains. The strain of Komagataeibacter sucrofermentans B-11267 was used as a BC producer, Xanthomonas campestris was used as a xanthan producer, and Leuconostoc mesenteroides was used as a dextran producer. The cultivation was carried out under dynamic conditions on a medium with molasses. The polysaccharide yield was expressed as the absolute dry weight of the polymers per unit volume of the culture medium. We have studied the morphology of bacterial cellulose using atomic force microscopy (AFM), FTIR spectroscopy. Crystallinity was checked by X-ray diffraction analysis. The interest in bacterial cellulose makes it necessary to synthesize it in large quantities on an industrial scale. The problem of increasing productivity was solved by cocultivating the bacterial cellulose producer Komagataeibacter sucrofermentans with the dextran producer Leuconostoc mesenteroides and xanthan producer Xanthomonas campestris, since the addition of water-soluble polysaccharides is known to increase viscosity of the medium and facilitate the dispersion of bacterial cellulose granules, thereby increasing the number of free cells, which can accelerate sugar consumption and polymer formation. At the first stage of the study, the selection of the most optimal conditions for cocultivation of the BC producer with the producers of xanthan and dextran was carried out, namely, the optimal pH value of the medium. Monoculture of bacteria X. campestris, L. mesenteroides, and K. sucrofermentans was carried out at different pH values (see Figs. 1–3). Based on the data obtained, it can be stated that the most optimal pH value for cocultivation of microorganisms is pH 5.0. In this regard, at the second stage of the work, joint cultivation of the BC producer strain K. sucrofermentans with the xanthan and dextran producers X. campestris and L. mesenteroides, respectively, on molasses medium was carried out. From the data presented (see Fig. 4), it can be seen that the largest amount of polysaccharide is formed on day 3 of cocultivation of the bacterial cellulose producer and the dextran producer. The amount of bacterial cellulose was 5.99 ± 0.02 g/L, i.e., two and a half times higher than the amount of polymer formed during monocultivation of the bacterial cellulose producer: 2.25 ± 0.05 g/L. Cocultivation of the bacterial cellulose producer strain with the xanthan producer did not lead to an increase in the polysaccharide yield. Therefore, no further study of cocultivation of these microorganisms was carried out. The success of joint cultivation of bacterial cellulose and dextran producer strains was assessed and the properties of the obtained polysaccharide were studied using AFM, FTIR spectroscopy, and X-ray structural analysis. The surface relief of the bacterial cellulose was studied by AFM (see Fig. 7). The analysis of AFM images showed the presence of an association of K. sucrofermentans and L. mesenteroides cells in the BC. Also, the obtained bacterial cellulose was investigated using the method of FTIR spectroscopy (see Fig. 8). The IR spectra show the similarity of the detected peaks with the literature data on the peaks corresponding to bacterial cellulose. To determine the degree of crystallinity, the structure of cellulose was studied by X-ray structural analysis (see Fig. 9). The degree of crystallinity of cellulose samples is 64 and 32% for monocultivation of K. sucrofermentans and cocultivation of K. sucrofermentans and L. mesenteroides, respectively.