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Folia microbiologica 62(6), 453-461 (2017). doi:10.1007/s12223-017-0516-9, Published by Springer, [S.l.]

Homopolysaccharides, consisting of one sugar, are described by use of the suffix “an”. Thus a polyglucose (polyglucopyranose) is termed “glucan”; others are heteropolysaccharides that may contain several different constituent residues. s(1,3)-D-Glucans (GLs) are the most commonly used term for homopolysaccharides that has s(1,3)-D-linkages in the backbone, and may possess s-D-glucosidic linkages at position 6 in different, often repeating units (branches). In the nature GLs are widespread and are there found in plants, micro-algae, bacteria, yeast and mushrooms. GLs vary in length and their degree of branching. Curdlan (Alcaligenes faecalis var. myxogenes IFO 13140) is a linear, unbranched GL, whereas scleroglucan and SSG (Sclerotium glucanium and Sclerotinia sclerotiorum IFO 9395, respectively), schizophyllan (Schizophyllum commune) and lentinan (Lentinus edodes) have a main chain composed of s(1,3)-D-glucopyranosyl residues with residues carrying a s(1,6)-D-linked single glucopyranosyl group. Laminarin (Laminaria digitata) and laminaran (Laminaria hyperborea) have more or less extensive branches through s(1,6)-D-linkages from the backbone. Yeast glucan is a particulate carbohydrate that consists of glucose and mannose. GL’s aqueous solubility ranges from insoluble (curdlan at room temperature) via gelling (scleroglucan) to fully soluble (laminarin and partly laminaran). The solubility in water is dependent on temperature [1, 2]. The use of GLs as immunostimulants in aquaculture is discussed.

A genomic library of A. faecalis A1501 was constructed in the broad-host-range cosmid vector pLA2917. A plasmid clone pLAF58 carrying ntrBC-like genes was obtained by colony hybridization, using heterologous probes. The ntrBC genes were localized on a 9.5kb SacI fragment. Establishment of the complete nucleotide sequence of the ntrBC region is in progress.

There are about 20 million hectares of saline land in North and East China, which causes a loss of approximately 30 million tons of crop yield annually. Some studies conducted in China showed that biofertilizer containing salt-tolerant nitrogen-fixing bacteria could promote growth of plants and increase the yield of crops (Ping et al., 1996). Alcaligenes faecalis A1501, which was isolated from paddy soils in South China in 1980, could grow well in solid or liquid medium containing 0.8 mol/L NaCl Table 1 Effect of various concentrations of NaCl on growth of A. faecalis frown in solid medium Na+(mol/L) 0.2 0.4 0.6 0.8 1.0 1.2 Solid medium +++ +++ ++ ++ + - +++, grow well; ++, grow normally; +, grow slowly; -, no growth. . Growth of bacteria was repressed in solid medium containing 1.2 mol/L NaCl. A1501 could grow very slowly even if the NaCl concentration reached 1.6 mol/L in liquid medium. A1501 could produce IAA under salt stress and maintain N-fixing activity until the NaCl concentration was higher than 1.0 mol/L Table 2 Identification of IAA produced by A. faecalis A1501 under salt street (0.4 M NaCl) Time of inclubation (h) A1501 Klebsiella sp. 101 Klebsiella NIAB-I 24 ++ + - 48 ++ + - 72 ++ + - 96 ++ + - 120 +++ ++ - +, IAA production; -, no IAA production. Table 3 Associative nitrogen fixation between host rice and A. faecalis A1501 Concentration of NaCl(%) 0.01 0.5 1.0 3.0 Rice roots inoculated with A1501 1045.3* 1565.4 956.7 799.1 Pure culture of A1501 681.0 755.1 803.6 456.5 *Activity Unit is ‘nmol C2H4 produred/h. g f.w. roots’. . Southern hybridization of total DNA from A1501 grown under condition of different NaCl concentration with Klebsiella pneumoniae nifHDK DNA probe show that homologous hybridization bands under the normal condition were identifical to those of the cells grown under the salt stress.

The strain under study, Alcaligenes faecalis A15, is a diazotroph isolated from the (endo)rhizosphere of paddy rice. To study the interaction of Alcaligenes faecalis A15 with rice we have applied the gus A reporter-gene technology. With a fusion constitutively expressing the gus A gene we have analysed the colonization patterns of rice roots and the environmental factors that affect colonization. With a transcriptional fusion of the Alcaligenes faecalis A15 nifH gene and gusA we were able to visualize the expression of the nitrogen fixation genes, both in planta and in free-living conditions. Finally, we have compared nifH expression in Azospirillum brasilense, Azospirillum irakense, and Alcaligenes faecalis A15 under plant root-associated conditions.

An examination into the effect of bioreactor design on the production of s-l,3-glucan exopolysaccharide (EPS) by selected strains of Alcaligenes faecalis and Agrobacterium radiobacter has revealed that the use of alternative means of agitation and mixing, whereby shear stress is reduced, leads to an increase in both the quantity and quality of the recoverable polymer. Whereas Rushton-type turbine impellers were effective in providing high rates of oxygen transfer for high cell density fermentations, the shear stress resulting from this type of agitation resulted in about a 30–40% reduction in the amount of recoverable polymer after a 5-day batch fermentation. With turbine impellers, the specific rate of polymer production (Qp) decreased significantly after 40–50hrs (stationary-phase). However, the Qp remained constant (approx. 100mg EPS/g cell/hr) over the entire time course of the batch fermentation (90 hrs) when ‘shear’ was reduced; for example, when (i) a marine-type propeller was substituted for the flat-blade turbine impellers, or (ii) mixing and oxygen transfer were accomplished in a non-rotary, vibro-fermentor. Systems with reduced ‘apparent shear’ also proved to be effective in the production of a higher quality polymer product as judged by such indicators as intrinsic viscosity and MW of the alkali-solubilized isolated polymer. The strength of the irreversible thermal gel was also markedly improved for polymer recovered from bioreactors configured with agitation devices designed to reduce shear stress on the cell/polymer complex. Other physicochemical properties were consistent with these high-quality novel polymers being ‘curdlan’-like, s-1,3-glucan exopolysaccharide.