Sucrose fed-batch strategy enhanced biomass, polysaccharide, and ganoderic acids production in fermentation of Ganoderma lucidum 5.26.

Ganoderma, as a Chinese traditional medicine, has multiple bioactivities. However, industrial production was limited due to low yield during Ganoderma fermentation. In this work, sucrose was found to greatly enhance intracellular polysaccharide (IPS) content and specific extracellular polysaccharide (EPS) production rate. The mechanism was studied by analyzing the activities of enzymes related to polysaccharide biosynthesis. The results revealed that sucrose regulated the activities of phosphoglucomutase and phosphoglucose isomerase. When glucose and sucrose mixture was used as carbon source, biomass, polysaccharide and ganoderic acids (GAs) production was greatly enhanced. A sucrose fed-batch strategy was developed in 10-L bioreactor, and was scaled up to 300-L bioreactor. The biomass, EPS and IPS production was 25.5, 2.9 and 4.8 g/L, respectively, which was the highest biomass and IPS production in pilot scale. This study provides information for further understanding the regulation mechanism of Ganoderma polysaccharide biosynthesis. It demonstrates that sucrose fed-batch is a useful strategy for enhancing Ganoderma biomass, polysaccharide and GAs production.

Rhizobium tubonense sp. nov., isolated from root nodules of Oxytropis glabra.

Four rhizobial strains, designated CCBAU 85046(T), CCBAU 85051, CCBAU 85048 and CCBAU 85049, isolated from root nodules of Oxytropis glabra grown in Tibet, China, were previously defined, using amplified 16S rRNA gene restriction analysis, as a novel group within the genus Rhizobium. To clarify their taxonomic position, these strains were further analysed and compared with reference strains of related bacteria using a polyphasic approach. The 16S rRNA gene analysis showed that the four isolates formed a distinct phylogenetic lineage in the genus Rhizobium. The isolates showed highest sequence similarity (97.8  %) to Rhizobium indigoferae CCBAU 71042(T). Phenotypic and physiological tests, DNA-DNA hybridization, phylogenetic analyses of housekeeping genes recA, atpD and glnII and fatty acid profiles also indicated that these four strains constitute a novel group distinct from recognized species of the genus Rhizobium. Based on this evidence, strains CCBAU 85046(T), CCBAU 85051, CCBAU 85048 and CCBAU 85049 represent a novel species in the genus Rhizobium, for which the name Rhizobium tubonense sp. nov. is proposed. The type strain is CCBAU 85046(T) (=LMG 25225(T) =HAMBI 3066(T)) and its DNA G+C content is 59.52 % (T(m)). Strain CCBAU 85046(T) could form effective nodules on plant species Vigna unguiculata and Medicago sativa but not on its host of origin Oxytropis glabra.

WITHDRAWN: Ensifer psoraleae sp. nov. and Ensifer sesbaniae sp. nov., isolated from the root nodules of Psoralea corylifolia, Sesbania cannabina and other legumes.

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Proposal of Ensifer psoraleae sp. nov., Ensifer sesbaniae sp. nov., Ensifer morelense comb. nov. and Ensifer americanum comb. nov.

In a survey of rhizobia associated with the native legumes in Yunnan Province, China, seven and nine strains isolated from the root nodules of Psoralea corylifolia, Sesbania cannabina and Medicago lupulina were respectively classified into the novel genomic species groups I and II in the genus Ensifer (former Sinorhizobium) based on the sequence analyses of the 16S rRNA gene. Analyses of concatenated housekeeping genes (atpD, recA and glnII) further revealed that they were distinct lineages in the genus, and group I was most similar to Ensifer terangae and Ensifer garamanticus (both with 94.2% similarity), while group II was most similar to Ensifer adhaerens (94.0%). These groups could be distinguished from closely related species by DNA-DNA relatedness, MALID-TOF MS, cellular fatty acid profiles and a series of phenotypic characters. Therefore, two novel species were proposed: Ensifer psoraleae sp. nov. (seven strains, type strain CCBAU 65732(T)=LMG 26835(T)=HAMBI 3286(T)) and Ensifer sesbaniae sp. nov. (nine strains, type strain CCBAU 65729(T)=LMG 26833(T)=HAMBI 3287(T)). They had a DNA G+C mol% (Tm) of 58.9 and 60.4, respectively. Both of the type strains formed effective nodules on common bean (Phaseolus vulgaris) and their hosts of origin. In addition, the previously described species Sinorhizobium morelense and Sinorhizobium americanum were renamed as Ensifer morelense comb. nov. and Ensifer americanum comb. nov. according to the accumulated data from different studies.

Rhizobium tibeticum sp. nov., a symbiotic bacterium isolated from Trigonella archiducis-nicolai (Sirj.) Vassilcz.

Isolated from root nodules of Trigonella archiducis-nicolai (Sirj.) Vassilcz. grown in Tibet, China, cells of the bacterial strains CCBAU 85039(T) and CCBAU 85027 were Gram-negative, aerobic, motile, non-spore-forming rods that formed colonies that were semi-translucent and opalescent on yeast extract-mannitol agar. In numerical taxonomy, SDS-PAGE analysis of whole-cell proteins and DNA-DNA hybridization, the two strains were very similar and were different from reference strains of defined Rhizobium species. In the phylogeny based on 16S rRNA gene sequences, they were most similar to Rhizobium etli CFN 42(T) (98.2 % similarity) and R. leguminosarum USDA 2370(T) (97.6 %). Sequence analyses of the housekeeping genes recA, atpD and glnII and the 16S-23S rRNA intergenic spacer, phenotypic characteristics and cellular fatty acid profiles strongly suggested that these two strains represented a novel species within Rhizobium. Cross-nodulation tests and sequencing of nifH and nodA genes showed that these two strains were symbiotic bacteria that nodulated Trigonella archiducis-nicolai, Medicago lupulina, Medicago sativa, Melilotus officinalis, Phaseolus vulgaris and Trigonella foenum-graecum. Based on the results, the novel species Rhizobium tibeticum sp. nov. is described to accommodate the two strains. The type strain is CCBAU 85039(T) (=LMG 24453(T) =CGMCC 1.7071(T)). The DNA G+C content of this strain is 59.7 mol% (T(m)).

Rhizobial resource associated with epidemic legumes in Tibet.

A total of 128 bacterial test strains originated from Astragalus, Caragana, Gueldenstaedtia, Medicago, Melilotus, Oxytropis, Trifolium, and Vicia grown in Tibet were characterized phenotypically and genomically. Based upon the consensus of grouping results, they were identified as 16 putative species. Twenty-five test strains belonging to seven putative species of Agrobacterium, Bradyrhizobium, and Rhizobium might be nonsymbiotic bacteria and the remaining 103 test strains were symbiotic bacteria belonging to Mesorhizobium, Rhizobium, and Sinorhizobium meliloti. Although no novel taxon was detected in the symbiotic bacteria, several characters including the alkaliphilic psychrotolerance revealed that the Tibetan rhizobia could be ecotypes adapted to the local conditions. The results also demonstrated that frequent lateral transfer of symbiotic genes might have happened in the Tibetan rhizobia since nodC genes similar to that of S. meliloti were found in several Rhizobium test strains and all the Mesorhizobium species had very similar nodC genes despite their genomic background. All of these findings demonstrated that the Tibetan rhizobia were an important resource for further studies on rhizobial ecology and application.